Photoinduced inverse Sonogashira coupling reaction

Alkynes are widely used in chemistry, medicine and materials science. Here we demonstrate a transition-metal and photocatalyst-free inverse Sonogashira coupling reaction between iodoalkynes and (hetero)arenes or alkenes under visible-light irradiation. Mechanistic and computational studies suggest that iodoalkynes can be directly activated by visible light irradiation, and an excited state iodoalkyne acted as an “alkynyl radical synthetic equivalent”, reacting with a series of C(sp²)–H bonds for coupling products. This work should open new windows in radical chemistry and alkynylation method.

A transition-metal and photocatalyst-free, photoinduced inverse Sonogashira coupling reaction was developed. Under visible-light irradiation, the excited state iodoalkyne acted as an “alkynyl radical synthetic equivalent”.

Alkynes are among the most important class of compounds in organic chemistry. Because of their structural rigidity, special electronic properties and numerous methods available for the functionalization of the triple bond, alkynes are important tools and structural elements both in medicinal chemistry and materials sciences.¹ Therefore, the development of a new methodology to introduce carbon–carbon triple bonds is of great importance in organic chemistry. The Sonogashira coupling reaction is typically used for the formation of C(sp)–C(sp²) bonds starting from hetero(aryl) halides and terminal alkynes.² Recently, “inverse Sonogashira coupling” involving the direct alkynylation of unreactive C(sp²)–H bonds with readily available alkynyl halides has received growing interest in the development of a complementary strategy (Fig. 1a). Various main-group and transition metals have been developed to promote this transformation.³ In addition, a photomediated Sonogashira reaction without a photocatalyst was also developed by several groups (Fig. 1b).⁴Open in a separate windowFig. 1Models of alkynylation. (a) Conventional inverse Sonogashira reaction. (b) Photomediated Sonogashira reaction. (c) SOMOphilic alkynylation. (d) Photoinduced inverse Sonogashira reaction.In recent years, SOMOphilic alkylnylation (SOMO = singly occupied molecular orbital) has become an excellent method of introducing alkynyl groups (Fig. 1c).⁵ Based on photoredox and transition metal catalysis, numerous in situ generated radicals undergo α-addition and β-elimination to alkynyl reagents, like the broadly applicable ethynylbenziodoxolone (EBX) reagent. Various radical alkynylations were thus discovered by Li,⁶ Chen,⁷ Waser,⁸ and many other groups.⁹ However, extending the scope of radical precursors, more atom–economic reactions, and a deeper understanding of the mechanism in these transformations are still highly desirable.After the discovering of trityl radicals by Gomberg in 1900, the “rational” era of radical chemistry has since begun.¹⁰ Now, the development of radical reactions, especially those involving C(sp³) and C(sp²) radicals, enables rapid access to drug discovery, agrochemistry, materials science, and other disciplines.¹¹ However, the C(sp) radical remains a baffling species. Due to their very high energy, short life time, and limited and harsh preparation methods, alkynyl radicals remain an elusive species, which just exists in some extreme environments, like outer-space and the petrochemical industry.¹² Even though alkynyl radicals have been proposed as intermediates for some alkynylation methods, they were regarded as mysterious species and ignored by organic chemists for a long time.¹³ Recently, two approaches have been developed to aid the alkynyl radical generation step. In 2015, Hashmi and collaborators reported a [Au₂(μ-dppm)₂]²⁺ catalyzed free radical–radical C(sp)–C(sp³) bond coupling reaction between iodoalkynes and aliphatic amines.¹⁴ Under irradiation of sunlight, the dimeric gold complex was proposed to reduce the iodine acetylide to an alkynyl radical. In 2017, Li developed a transition-metal-free alkynylation reaction between iodoalkyne and 2-indolinone.¹⁵ Iodoalkynes could release alkynyl radicals under high temperature conditions. In 2019, we reported an Au(i) and Ir(iii) catalyzed alkynylative cyclization of o-alkylnylphenols with iodoalkynes, wherein the photosensitized energy transfer promoted the oxidative addition of a gold(i) complex with iodoalkynes.¹⁶ Based on our continuous interest in haloalkyne and photo-chemistry, we proposed that an iodoalkyne could be a potential “alkynyl radical precursor” under light irradiation. In this work, we uncovered a novel mode of transition-metal and photocatalyst-free, direct photoexcitation of iodoalkynes for the inverse Sonogashira coupling reaction with arenes, heteroarenes, and alkenes via an “alkynyl-radical type” transfer (Fig. 1d).     

0D–1D hybrid nanoarchitectonics: tailored design of FeCo@N–C yolk–shell nanoreactors with dual sites for excellent Fenton-like catalysis

Heterogeneous Fenton-like processes are very promising methods of treating organic pollutants through the generation of reactive oxygen containing radicals. Herein, we report novel 0D–1D hybrid nanoarchitectonics (necklace-like structures) consisting of FeCo@N–C yolk–shell nanoreactors as advanced catalysts for Fenton-like reactions. Each FeCo@N–C unit possesses a yolk–shell structure like a nanoreactor, which can accelerate the diffusion of reactive oxygen species and guard the active sites of FeCo. Furthermore, all the nanoreactors are threaded along carbon fibers, providing a highway for electron transport. FeCo@N–C nano-necklaces thereby exhibit excellent performance for pollutant removal via activation of peroxymonosulfate, achieving 100% bisphenol A (k = 0.8308 min⁻¹) degradation in 10 min with good cycling stability. The experiments and density-functional theory calculations reveal that FeCo dual sites are beneficial for activation of O–O, which is crucial for enhancing Fenton-like processes.

Novel 0D–1D hybrid nanoarchitectonics consisting of FeCo@N–C yolk–shell nanoreactors are developed for Fenton-like reaction. With the multilevel advantages of this design, FeCo@N–C nano-necklaces exhibit excellent performance for BPA removal.

Advanced oxidation processes (AOPs) are one of the most promising strategies to eliminate organic contaminants, sustainably generating reactive oxygen species (ROS) to ideally destroy all non-biodegradable, recalcitrant, toxic, or membrane-permeable organic impurities.^1–4 Among these AOPs, sulfate radical (SO₄˙⁻)-based Fenton-like processes have gained increasing attention as a water treatment strategy because of the strong oxidation potential of SO₄˙⁻ (3.1 V vs. normal hydrogen electrode) at wider pH ranges. SO₄˙⁻ is mainly produced by physical or chemical methods for activation of persulfate salts, such as peroxymonosulfate (PMS) and persulfate.^5–9 Over the past two decades, heterogeneous catalysis has emerged as the most effective approach to water treatment, with much effort dedicated to developing better catalysts, including transition metal-based and carbonaceous materials.^10,11 Unfortunately, most metal-based catalysts suffer from leaching of toxic metal ions, which can thwart their practical application,^12,13 and although carbonaceous catalysts produce no secondary pollution, their cycle performance is always depressed.¹⁴ There is therefore an urgent need to find robust catalysts with adequate activity and stability for Fenton-like processes.To achieve superior performance, an ideal Fenton-like catalyst should contain oxidants with favorably reactive centers for cleavage of peroxyl bonds (O–O), have structure optimized for target pollutant attraction, and have chainmail to protect the vulnerable active sites for long periods.^15–17 Recent studies have demonstrated Co–N–C active sites prefer to activate the O–O of PMS.¹⁸ Furthermore, introducing Fe-doping into the Co–N–C system not only suppresses Co²⁺ leaching, but also modulates the pyrrolic-N content, which is the adsorption site for capture of bisphenol A (BPA).¹⁹ We previously discovered that Co@C yolk–shell nanoreactors could enhance the catalytic activity because of the confinement effect in the nano-spaces between the core and shell, while the carbon shell acted like a chainmail protecting the Co active sites, keeping them highly reactive after five cycles.^20,21Combining different kinds of materials to generate novel hybrid material interfaces can enable the creation of new kinds of chemical and physical functionalities that do not currently exist. However, one cannot simply mix these materials in an uncontrolled manner, because the ensemble of interfaces created by random mixing tends to favour thermodynamically stable interfaces that are functionally less active. Therefore, to prepare new materials with high functionality, it is necessary to carefully control the hybridization of components in interfacial regions with nanometric or atomic precision. By further hybridization of different components e.g., zero to one dimension (0D–1D) hybrid structures, we can prepare the structure to increase not only the specific surface area but also the interfacial region between different materials.In this work, we report novel 0D–1D hybrid nanoarchitectonics (necklace-like structures) consisting of FeCo@N–C yolk–shell nanoreactors as a PMS activator for Fenton-like processes. This catalyst has multilevel advantages: (i) each FeCo@N–C unit is a well-formed yolk–shell nanoreactor, which can guarantee sufficient contact of reactants and active sites, as well as defend them for good durability; (ii) all single nanoreactors are threaded along the carbon fibers, providing a highway for electron transport; and (iii) all the carbon fibers constructed into a thin film with macroscopic structure, which overcomes the complex recyclability of powder catalysts. Benefiting from favorable composition and unique structure, the FeCo@N–C catalyst delivers excellent performance for BPA removal via activation of PMS accompanied with good stability.The synthesis processes of necklace-like nanoarchitecture containing FeCo@N–C yolk–shell nanoreactors are illustrated in Fig. 1a. First, uniform Fe–Co Prussian blue analogue (Fe–Co PBA) nanocubes with an average size of 800–900 nm (Fig. 1b) are encapsulated in polyacrylonitrile (PAN) nanofibers by electrospinning. The obtained necklace-like FeCo PBA–PAN fibers (Fig. 1c) are then pyrolyzed at 800 °C in N₂ atmosphere to produce FeCo@N–C nano-necklaces. The scanning electron microscopy (SEM) image (Fig. 1d) of the FeCo@N–C shows this necklace-like morphology with its large aspect ratio, with the FeCo@N–C particles strung along the PAN-derived carbon fibers. A broken particle (Fig. 1e) shows that the FeCo@N–C has a yolk–shell architecture, which is also identified by transmission electron microscopy (TEM). Fig. 1f and g show the well-defined space between the inner yolk and outer shell, which is attributed to the volume shrinkage of the original Fe–Co PBAs. During pyrolysis, Fe–Co PBA is reduced to FeCo (inner yolk) and PAN is carbonized (outer carbon shell), resulting in the unique necklace-like nanoarchitecture.^22–24 The high-resolution TEM in Fig. 1h shows a lattice fringe of 0.20 nm, which matches well with the (110) plane of FeCo alloy.²⁵ The scanning transmission electron microscopy (STEM) image (Fig. 1i) and corresponding elemental map (Fig. 1j) indicate that FeCo nanocrystals are well distributed in the inner core with some small FeCo nanocrystals located on external carbon shells. Furthermore, the control samples of Fe@N–C and Co@N–C nano-necklaces, prepared by only replacing the Fe–Co PBA nanocubes with Fe–Fe PB and Co–Co PBA (Fig. S1^†), also demonstrate the versatility of this synthetic strategy. The formation of hierarchical porous structure, beneficial to the PMS transportation on the surface of catalysts, could be determined by N₂ adsorption–desorption isotherms and corresponding pore volume analysis (Fig. S2 and Table S1^†).Open in a separate windowFig. 1(a) Preparation of FeCo@N–C necklace-like nanoarchitecture. SEM images of (b) Fe–Co PBA cubic particles and (c) the electrospun FeCo PBA–PAN fibers. (d and e) SEM, (f and g) TEM, and (h) high-resolution TEM images of FeCo@N–C nano-necklaces. (i) STEM and (j) the corresponding elemental mappings of C, N, Fe, and Co.The X-ray diffraction patterns of the as-prepared products are depicted in Fig. S3,^† with one prominent diffraction peak centered at 44.8° corresponding to the (110) lattice plane of FeCo alloy. All the products also have a characteristic signal at 26°, implying that graphite carbon is formed during pyrolysis. Raman spectroscopy further analyzed the crystal structures and defects of the FeCo@N–C nano-necklaces (Fig. S4^†), where peaks found at 1349 cm⁻¹ and 1585 cm⁻¹ index the disordered (D band) and graphitic carbon (G band), respectively.²⁶ X-ray photoelectron spectroscopy investigated the composition and valence band spectra of FeCo@N–C nano-necklaces. The survey spectrum (Fig. S5a^†) reveals the presence of Fe (1.4%), Co (1.2%), C (86.4%), N (4.5%), and O (6.5%) in the composite. The high-resolution N 1s spectrum (Fig. S5b^†) exhibits broad peaks at 398.1, 401.1, and 407.4 eV, corresponding to the pyridinic-N, graphitic-N, and σ* excitation of C–N, respectively.²⁷ The high-resolution Fe 2p spectrum (Fig. S5c^†) shows a broad peak at 707.4 eV, attributed to Fe⁰. Similarly, the 777.5 eV peak observed in the Co 2p spectrum (Fig. S5d^†) corresponds to Co⁰, implying that FeCo dual sites have formed.²⁸ The oxidation state of these sites was investigated by ⁵⁷Fe Mössbauer spectroscopy, which found a sextet in the Mössbauer spectrum of the FeCo@N–C nano-necklaces attributed to FeCo dual sites (Fig. 2a and Table S2^†).²⁹ The coordination environment of the FeCo dual sites was also verified by X-ray absorption fine structure (XAFS) spectroscopy. Fig. 2b shows that the X-ray absorption near-edge structure (XANES) spectra of the Fe K-edge, which demonstrates a similar near-edge structure to that of Fe foil, illustrating that the main valence state of Fe in FeCo@N–C nano-necklaces is Fe⁰. Furthermore, the extended-XAFS (EXAFS) spectra (Fig. 2c) displays a peak at 1.7 Å, which is ascribed to the Fe–N bond, and a remarkable peak at approximately 2.25 Å corresponding to the metal–metal band.^10,30 The Co K-edge and EXAFS spectra (Fig. S6^†) also confirm the presence of Co–N and the metal–metal band. These results provide a potential structure of the FeCo dual sites in the FeCo@N–C nano-necklaces, as illustrated in Fig. 2d.Open in a separate windowFig. 2(a) ⁵⁷Fe Mössbauer spectra of FeCo@N–C nano-necklaces at 298 K. (b) Fe K-edge XANES spectra of FeCo@N–C nano-necklaces and Fe foil. (c) Corresponding Fourier transformed k³-weighted of the EXAFS spectra for Fe K-edge. (d) Possible structure of the FeCo dual sites.This dual-metal center and necklace-like structure may be beneficial to enhance catalytic performance. Fig. 3a shows the Fenton-like performance for BPA degradation compared to Fe@N–C nano-necklaces, Co@N–C nano-necklaces, and FeCo@N–C particles (Fe–Co PBA directly carbonized without electrospinning). Here, the FeCo@N–C nano-necklaces display a higher catalytic performance, with BPA completely removed in 7 min. To clearly compare their catalytic behavior, the kinetics of BPA degradation was fitted by the first-order reaction. As shown in Fig. 3b, FeCo@N–C nano-necklaces exhibit the highest apparent rate constant (k = 0.83 min⁻¹), which is approximately 6.4, 2.6, and 1.2 times that of FeCo@N–C particles, Fe@N–C nano-necklaces, and Co@N–C nano-necklaces, respectively. The significantly enhanced performance of FeCo@N–C nano-necklaces suggests that the FeCo dual sites and necklace-like nanoarchitecture are crucial. Furthermore, the concentration of BPA and PMS in the solution is higher than that in yolk–shell nanoreactor, resulting a concentration gradient which helps to accelerate the diffusion rates of reactants (Fig. 3c).^31,32 For these nano-necklaces, the carbon shell acts like a chainmail protecting the FeCo active sites from attack by molecules and ions, and all the nanoreactors are threaded along the carbon fibers, providing a highway for electron transport, which is important for SO₄˙⁻ generation (SO₄˙⁻ production as eqn, HSO₅⁻ + e⁻ → SO₄˙⁻ + OH⁻). Electrochemical impedance spectroscopy further confirms the good conductivity of the FeCo@N–C nano-necklaces (Fig. 3d). In addition, the concentration of metal-ion leaching and cycling performance (Fig. 3e and f) reveal the high reusability of FeCo@N–C nano-necklaces, with 95% BPA removal in 20 min after five cycles, which is also proved by the SEM and TEM characterization (Fig. S7^†). The effect of other reaction parameters on the BPA degradation, such as pH, reaction temperature, PMS or catalysts dosage, and common anions, were investigated in detail (Fig. S8–S11^†). All the results demonstrate that FeCo@N–C nano-necklaces deliver a better performance for PMS catalysis. In addition, the turnover frequency (TOF) value of FeCo@N–C nano-necklaces is 5.5 min⁻¹ for BPA degradation, which is higher than many previously reported catalysts (detailed catalytic performance comparison as shown in Table S3^†).Open in a separate windowFig. 3(a) BPA degradation efficiency in different reaction systems and (b) the corresponding reaction rate constants. (c) Schematic illustration of PMS activation in FeCo@N–C nano-necklaces. (d) Nyquist plots of the catalysts. (e) The metal leaching in different reaction systems. (f) Cycling performance of FeCo@N–C nano-necklaces for BPA removal. Reaction conditions: [catalyst] = 0.15 g L⁻¹, [BPA] = 20 mg L⁻¹, [PMS] = 0.5 g L⁻¹, T = 298 K, and initial pH = 7.0.To examine the enhanced catalytic activity, radical quenching experiments were conducted. As shown in Fig. 4a, when NaN₃ is added to the reaction solution as a scavenger for ¹O₂, there is no significant reduction of BPA decomposition, implying that non-radicals are not the dominant reactive species. By comparison, when tert-butanol (TBA) (radical scavenger for ˙OH) is added, there is a slight (2.8%) decrease in BPA removal. However, if methanol (radical scavenger for SO₄˙⁻ and ˙OH) is added, the efficiency of BPA degradation declines by up to 59.2%, indicating that the major radicals generated from the PMS activation are SO₄˙⁻;³³ the presence of these radicals is also verified by electron paramagnetic resonance (EPR) (Fig. 4b). Furthermore, the significant inhibition ratio can be observed when KI (quencher for the surface) is added, demonstrating that BPA degradation is mainly attributed to reactions with SO₄˙⁻, which is produced by a surface catalytic process.³⁴Open in a separate windowFig. 4(a) Effects of the radical scavengers on BPA degradation. (b) EPR spectra of SO₄˙⁻ and ˙OH. (c) The energy profiles of PMS on FeCo@N–C nano-necklaces surface. (d) Optimized configurations of PMS adsorbed on FeCo@N–C nano-necklaces.Density-functional theory was applied to calculate the surface energy of PMS activation at FeCo dual sites (Fig. 4c, d and S12^†). The dissociation barrier of PMS into SO₄˙⁻ and OH⁻ is −2.25 eV, which is much lower than that on an Fe or Co single site, suggesting that cleavage of O–O bonds of PMS occurs more easily on FeCo dual sites. This is because FeCo dual sites provide two anchoring sites for the dissociated O atoms, leading to more efficient activation of O–O. The FeCo@N–C nano-necklaces can reduce the energy barrier of O–O bond breaking, which results in high activity for PMS activation and thus high productivity of SO₄˙⁻.     

One-electron bonds in copper–aluminum and copper–gallium complexes

Odd-electron bonds have unique electronic structures and are often encountered as transiently stable, homonuclear species. In this study, a pair of copper complexes supported by Group 13 metalloligands, M[N((o-C₆H₄)NCH₂PⁱPr₂)₃] (M = Al or Ga), featuring two-center/one-electron (2c/1e) σ-bonds were synthesized by one-electron reduction of the corresponding Cu(i) ⇢ M(III) counterparts. The copper bimetallic complexes were investigated by X-ray diffraction, cyclic voltammetry, electron paramagnetic spectroscopy, and density functional theory calculations. The combined experimental and theoretical data corroborate that the unpaired spin is delocalized across Cu, M, and ancillary atoms, and the singly occupied molecular orbital (SOMO) corresponds to a σ-(Cu–M) bond involving the Cu 4p_z and M ns/np_z atomic orbitals. Collectively, the data suggest the covalent nature of these interactions, which represent the first examples of odd-electron σ-bonds for the heavier Group 13 elements Al and Ga.

Hanging on by a thread. Formally zerovalent copper complexes with an Al(iii) or Ga(iii) support were investigated. The combined experimental and theoretical data corroborate the presence of an odd-electron σ-bond between Cu and the Group 13 center.

Odd-electron σ-bonds, where the electrons are delocalized between two atoms, can occur as two-center/one-electron (2c/1e) or two-center/three-electron (2c/3e) interactions. Proposed by Pauling in 1931,¹ odd-electron σ-bonds have garnered attention because of their fundamental importance to chemical bonding and their relationship to radical species generated during oxidative stress in biological systems.^2–14 Examples of compounds exhibiting odd-electron bonding are typically homonuclear (like H₂⁺, He₂⁺, and alkali metal dimers) and transiently stable, limiting them to spectroscopic characterization.^1,11,15–18The first solid-state structure of a formally one-electron σ-bond was a tetraphosphabenzene species (Fig. 1a) which was formed by the coupling of two diphosphirenyl radicals.¹⁹ Following this discovery, the formation of discrete 2c/1e σ-bonds, where the odd-electron is delocalized between two homonuclear main group centers, was reported for B·B and then extended to P·P.^8,17,20 Of note, the first solid-state structure of a B·B compound was reported in only 2014 (Fig. 1b).²¹ Examples of 2c/1e σ-bonds between the heavier Group 13 congeners are even more lacking because of the greater propensity for their unpaired spins to couple, forming larger more stable clusters.⁸ To our knowledge, there are only three structurally characterized examples of odd-electron bonds for the heavy Group 13 atoms,²² and these examples are all homonuclear π-radicals (Fig. 1c).^23–26Open in a separate windowFig. 1Select examples of structurally characterized molecules (a–d) featuring odd-electron bonds.Heteronuclear odd-electron σ-bonds are also rare. The Cu(TPB) complex, where TPB is a trisphosphinoborane, is the single structural example of a 2c/1e bond between heteroatoms (Fig. 1d).²⁷ The authors described the bonding as Cu·B, where the unpaired electron is heavily polarized toward B. A theoretical study predicted that such a bond would also exist between Cu and Al, but no heavier analogues of Cu(TPB) have been synthesized to date.²⁸ Furthermore, the heavier Group 13 elements by virtue of their lower electronegativity compared to B should facilitate greater covalent interactions with the Cu center.Hence, we sought to target formally zerovalent Cu complexes supported by Al(III) or Ga(III) as an extension of the previously reported isoelectronic nickelate species and Cu(TPB).²⁹ Herein, we describe the synthesis, structure, spectroscopic characterization, and DFT calculations of cationic [CuML]⁺ complexes (L = [N((o-C₆H₄)NCH₂PⁱPr₂)₃]³⁻; M = Al and Ga) as well as their one-electron reduced metalloradical counterparts that feature discrete 2c/1e bonds.     

Monitoring single Au38 nanocluster reactions via electrochemiluminescence

Herein, we report for the first time single Au₃₈ nanocluster reaction events of highly efficient electrochemiluminescence (ECL) with tri-n-propylamine radicals as a reductive co-reactant at the surface of an ultramicroelectrode (UME). The statistical analyses of individual reactions confirm stochastic single ones influenced by the applied potential.

Herein, we report for the first time single Au₃₈ nanocluster reaction events of highly efficient electrochemiluminescence (ECL) with tri-n-propylamine radicals as a reductive co-reactant at the surface of a Pt ultramicroelectrode (UME).

Single entity measurements have been introduced by Bard and Wightman based on the collisions/reactions of individual nanoparticles and molecules at an ultramicroelectrode (UME).^1–9 Since then, the field of single entity electrochemistry has gradually attracted several research groups and has become a frontier field of nanoelectrochemistry and electroanalytical chemistry.^8,10–14 For instance, it has been shown that the chemistry of the electrode surface plays an important role in the collision/reaction events and the kinetics of reaction processes.^15–21 Dasari et al. reported that hydrazine oxidation and proton reduction can be detected using single Pt nanoparticles on the surface of a mercury or bismuth modified Pt UME, and the material of the electrode was found to affect the shape of current–time transients.^22,23 Fast scan cyclic voltammetry provides better chemical information about transient electrode–nanoparticle interactions, which is otherwise difficult to obtain with constant-potential techniques.²⁴ There are only a few reports on photoelectrochemical systems including semiconductor nanoparticles designed to detect single nanoparticles in the course of photocatalysis processes.^25–28 More importantly, owing to the nature of stochastic processes of single entity reactions, statistical analyses have shown substantial influences on the understanding of the underlying processes.Electrochemiluminescence or electrogenerated chemiluminescence (ECL),²⁹ as a background-free technique,^30–32 was also utilized to detect individual chemical reactions and single Pt nanoparticle collisions based on the reaction between the Ru(bpy)₃²⁺ complex and tri-n-propylamine (TPrA) radicals on the surface of an ITO electrode.^2,33,34 It was found that the size of the nanoparticles, the origin of the interaction between particles and the electrode surface, the concentration of species generation, and the lifetime of individual electrogenerated nanocluster species (i.e., Au₃₈²⁺, Au₃₈³⁺, and Au₃₈⁴⁺) in conjunction with the reactivity of those oxidized species with co-reactant radical intermediates (i.e., TPrA radical) play crucial roles in the frequency of the ECL reaction events leading to individual ECL responses. More strikingly, a higher ECL reaction frequency is directly proportional to the amount of collected ECL light.²¹ Chen and co-workers also employed ECL to study stationary single gold-platinum nanoparticle reactivity on the surface of an ITO electrode.³⁵ Lin and co-workers monitored the hydrogen evolution reaction in the course of “ON” and “OFF” ECL signals.³⁶ Recently, we performed a systematic and mechanistic ECL study of a series of gold nanoclusters, with the general formula of Au_n(SC₂H₄Ph)_m^z (n = 25, 38, 144, m = 18, 24, 60 and z = −1, 0, +1), where near-infrared (NIR) ECL emission was observed.³⁷ There are several enhancement factors, such as catalytic loops^38,39 that improve the signal to noise ratio. The Wightman group was able to report single ECL reactions based on the capability of ECL.⁷ Furthermore, thus far, we have explored ECL mechanisms and reported the ECL efficiency of five different gold nanoclusters i.e., Au₂₅(SR)₁₈¹⁻, Au₂₅(SR)₁₈⁰, Au₂₅(SR)₁₈¹⁺, Au₃₈(SR)₂₄⁰ and Au₁₄₄(SR)₆₀⁰, among which the Au₃₈(SR)₂₄⁰/TPrA system revealed outstanding ECL efficiency, ca. 3.5 times higher than that of Ru(bpy)₃²⁺/TPrA as a gold standard. Therefore, we decided to focus on the Au₃₈ (SR)₂₄⁰/TPrA system. It was discovered that the ECL emission of these nanomaterials can be tuned through varying the applied potential and local concentration of the desired co-reactant.Herein, for the first time we report on ECL via a single Au₃₈(SC₂H₄Ph)₂₄ nanocluster (hereafter denoted as Au₃₈ NC) reaction (eq. (1)) in the vicinity of an UME in the presence of TPrA radicals as a reductive co-reactant.1where x is the oxidation number that can be either 0, 1, 2, 3 or 4. Single ECL spikes (Fig. 1A) along with ECL spectroscopy were used for elucidating individual reaction events. Indeed, each single ECL spike demonstrates a single Au₃₈^(x−1)* reaction product. Au₃₈ NCs were synthesized according to procedures reported by us and others, and fully characterized using UV-Visible-NIR, photoluminescence, ¹HNMR spectroscopy and MALDI mass spectrometry to confirm the Au₃₈ nanocluster synthesis (details are provided in ESI, Sections 1–3, Fig. S1–S4^†).^38,40,41Fig. 2 (left) shows a differential pulse voltammogram (DPV) in an anodic scan of a 2 mm Pt disc electrode immersed in 0.1 mM Au₃₈ acetonitrile/benzene solution containing 0.1 M TBAPF₆ as the supporting electrolyte. There are five discrete electrochemical peaks at which Au₃₈⁰ was oxidized to Au₃₈⁺ (E°′ = 0.39 V), Au₃₈²⁺ (E°′ = 0.60 V), and Au₃₈^3+/4+ (E°′ = 0.99 V) and reduced to Au₃₈⁻ (E°′ = −0.76 V) and Au₃₈²⁻ (E°′ = −1.01 V).^38,40,41Open in a separate windowFig. 1(A) An example of the reaction event transient of 10 μM Au₃₈ in benzene/acetonitrile (1 : 1) containing 0.1 M TBAPF₆ in the presence of 20 mM TPrA at 0.9 V vs. SCE, acquired at 15 ms time intervals using a 10 μm Pt UME. The white dashed-line indicates the threshold to identify single ECL spikes. (B) Illustration of a single nanocluster ECL spike. (C) ECL instrumentation with an inset showing ECL spike generation in the vicinity of the Pt UME.Open in a separate windowFig. 2Anodic DPV for Au₃₈ (left), reaction energy diagram of Au₃₈²⁺ and TPrA· (middle) along with the ECL–voltage curve (right) in an anodic potential scan at a 2 mm Pt disk electrode immersed in a solution of 10 μM Au₃₈ with 20 mM TPrA.The rich electrochemistry of Au₃₈ NCs is well-matched with that of co-reactants such as TPrA to generate near infrared-ECL (NIR-ECL), and the ECL emission efficiency of the Au₃₈/TPrA system is 3.5 times larger than that of the Ru(bpy)₃²⁺/TPrA co-reactant ECL system.²⁷Thus, it is of utmost interest to investigate the ECL generation of the above co-reactant system in single reactions, which improves the ECL signal detection sensitivity. To perform the ECL experiment a solution of 10 μM Au₃₈ NC with 20 mM TPrA was prepared. We first confirmed the ECL light generation of such solution along with its blank solution containing only TPrA using a typical 2 mm diameter Pt disk electrode (Fig. 2, S5 and S6^†).A 10 μm Pt UME electrode, which is electrochemically inert (Fig. S7^†), was utilized to investigate the ECL of single NC reactions under potentiostatic conditions, at which a specific positive bias potential was applied to oxidize both Au₃₈ and TPrA. Fig. 1A shows a typical ECL–time transient current curve (ECL intensity versus time) at 0.90 V vs. SCE, which was acquired using a photomultiplier tube (PMT, R928) for a duration of 1800 s at data acquisition time intervals of 15 ms (Fig. 1C and ESI, Section 3^†). Fig. 1B represents an exemplary event of a single ECL spike with a sharp increase followed by a decay in the ECL intensity. It is observed from the many spikes in Fig. 1B that this process can reoccur with a high probability in the vicinity of the UME, probably due to an electrocatalytic reaction loop (Fig. 1C). Indeed, ECL intensity was enhanced in this way as an already relaxed species, i.e., Au₃₈^z+1*, participates in an oxidation step to regenerate Au₃₈^z+1 to react with the TPrA radical (TPrA˙).Once photons resulting from the excited state relaxation in the vicinity of the UME are captured by the PMT, individual reaction events can be observed (Fig. 1A with the instrumentation schematic shown in Fig. 1C). As shown in Fig. 3A, there are many ECL spikes during 1800 s of measurement, each of which represents an individual ECL generation reaction in the vicinity of the UME surface. It is worth noting that there are several spikes with various intensities. This is most likely due to the Brownian motion which is random movement due to the diffusion of individual nanocluster species such as Au₃₈⁰, Au₃₈¹⁺, Au₃₈²⁺, etc., electrogenerated at the local applied potentials. Long and co-workers⁴² proposed that silver nanoparticle collision on the surface of a gold electrode follows Brownian motion, leading to several types of surface-nanoparticle response peak shapes. In fact, the observed ECL spikes, shown in Fig. 1C, with a rise and an exponential decay suggested that Au₃₈ nanocluster species diffuse directly through the electrode double-layer, move towards the tunneling region of the electrode surface, collide⁴² and become oxidized, react with TPrA radicals thereafter to produce excited states, and emit ECL. It is worth emphasizing that this path could be partially different for each individual nanocluster owing to the angle and direction relative to the electrode surface. The single Au₃₈ NC reaction behaviour at various bias potentials was investigated following the electrochemical energy diagram shown in Fig. 2, middle. For example, at a bias potential of 0.70 V (the green spot on the DPV in Fig. 2), Au₃₈⁰ undergoes two successive oxidation reactions to Au₃₈²⁺ and TPrA oxidation and deprotonation start to generate TPrA·. In fact, at a very close oxidation potential to Au₃₈²⁺, TPrA is also oxidized to its corresponding cation radical (ca. 0.80 V vs. SCE) Fig. S6,^† followed by deprotonation to form the TPrA radical.³⁸ The TPrA· with a very high reduction power (E°′ = −1.7 eV)⁴³ injects one electron to the LUMO orbital of the nanocluster and forms excited state Au₃₈⁺*, as illustrated in the reaction energy diagram in Fig. 2, middle.³⁸ Then, Au₃₈⁺* emits ECL light while relaxing to the ground state. For another instance, at 1.10 V vs. SCE (the red spot on the DPV in Fig. 2), Au₃₈⁰ is oxidized to Au₃₈^3/4+ feasibly. At this potential, the TPrA radical is generated massively in the vicinity of the electrode. The efficient electron transfer between the TPrA radical and Au₃₈^3/4+ generates both Au₃₈²⁺* and Au₃₈³⁺* that emit light at the same wavelength of 930 nm.³⁸ The results of such interactions produced a transient composed of many ECL events (Fig. 3A), which is an indication of bias potential enforcement on the nanocluster light emission.Open in a separate windowFig. 3Single-nanocluster ECL photoelectron spectroscopy of Au₃₈. ECL–time transients (A), statistics of the number of photons (B), histogram of the single reaction time between sequential spikes (C) and accumulated ECL spectrum (D) for a 10 μm Pt UME at 1.1 V immersed in a 10 μM Au₃₈ nanocluster solution in benzene/acetonitrile (1 : 1) containing 0.1 M TBAPF₆ in the presence of 20 mM TPrA. (E)–(H) The counterpart plots to (A)–(D) for the UME biased at 0.7 V. # represents the number.We further tried to collect the current–time traces of such events; however, owing to the high background current originating from the high concentration of TPrA relative to that of the nanocluster, no noticeable spikes in the current were observed.In order to study the photochemistry and understand deeply the single nanocluster reactions, ECL–time transients were collected at different applied potentials (i.e., 0.7, 0.8, 0.9 and 1.1 V vs. SCE) as labelled in green, brown, purple, and red on the DPV in Fig. 2, respectively. The transients were further analysed using our home-written MATLAB algorithm adapted from that for nanopore electrochemistry.⁴⁴ The population of individual events was identified by applying an appropriate threshold to discriminate ECL spikes from the noise as demonstrated in Fig. 1A. In fact, the applied algorithm also assisted us to learn about the raising time and intensity of each spike, as well as photons of individual spikes. For instance, Fig. 3A shows another typical transit for 1800 s at an UME potential bias of 1.1 V for the ECL events. Indeed, the integrated area of each peak, the charge of the photoelectrons at the PMT, is directly proportional to the number of photons emitted from individual reactions (see ESI, Section 5^†). Basically, the PMT amplifies the collected single photon emitted in the course of light-to-photoelectron conversion (see ESI, Section 6 and Fig. S8^†) and translates a single photon into photoelectrons. The extracted charge of each ECL reaction, Q_ECL, was then converted to the corresponding number of photons by dividing by the gain factor, g, which is 1.55 × 10⁶ (Fig. S8^†), following eqn (2):2The histograms of the number of photons show a Gaussian distribution (Fig. 3B) with a reaction frequency of 53.5 ± 2.9 at E = 1.1 V, whereas at a lower potential of 0.7 V the reaction frequency drops to 18.5 ± 1.7 (Fig. 3F). This indicates that there is a three-fold lower reaction occurrence at the lower potential. The integration of the Gaussian fitting at 1.1 V and 0.7 V also reveals a three-fold drop from 3.3 × 10⁵ to 1.2 × 10⁵ photons over 1800 s.To further explore the effect of electrode potential bias on the single Au₃₈ NCs ECL reaction, potentials lower than 1.1 and higher than 0.7 V, ca. 0.8 and 0.9 V (brown and purple labels in Fig. 2), were applied. In fact, the resulting ECL–time transients show a lower population of single spikes (Fig. S12A and ESI,^†). The integrated Gaussian curve values support the ECL–time transient observations with ∼4.1 × 10⁴ and ∼6.5 × 10⁴ photons, respectively. In fact, it is unlikely that the PMT would get more than two events in the duration, owing to the following reasons: (i) it has been shown that only 5.5% of incoming photons can be effectively converted to photoelectron signals by our R928 PMT during our absolute efficiency calibration, ESI Section 6 and Fig. S8–S19^†;⁴⁵ (ii) spherical ECL emission is proven to be detected for a substantial small part upon examination of our detection system for the absolute ECL quantum efficiency;⁴⁵ (iii) Au₃₈ nanocluster ECL emissions occur at 930 nm, which is almost at the wavelength detection limit of our PMT response curve.^38,45In addition, we evaluated the stochastic (a series of random events at various probability distributions) nature of the observed events and extracted the reaction time interval (τ) at various potentials. The resulting graph shows an exponential decay (Fig. 3C) as expressed in eqn (3):3where frequency (λ) gives the mean rate of the event and A represents the fitting amplitude. One can expect to obtain the distribution of the number of emitted photons and spatial brightness function. In fact, the exponential decay is a clear indication of random single reaction events as Whiteman and co-workers described for a 9,10-diphenylanthracene (DPA) ECL system in the annihilation pathway.^7,46 At a potential of 1.1 V, λ and A are found to be 4.98 ± 0.02 ms⁻¹ and 80.4 ± 3.2, whereas at 0.7 V, λ and A turned out to be 32.9 ± 1.6 ms⁻¹ and 9.5 ± 0.1 (Fig. 3C and G). Indeed, the lower potential of 0.70 V vs. SCE is high enough to generate the TPrA radical along with Au₃₈²⁺, thereby leading to excited Au₃₈⁺*, Fig. 3E. One can conclude that at the applied potentials of 0.7 V and 1.1 V, Au₃₈⁰ is oxidized to Au₃₈²⁺ and Au₃₈⁴⁺, resulting in the generation of Au₃₈⁺* and Au₃₈³⁺* under static conditions. Thus, there are higher populations of ECL spikes with no discrepancy in the number of collected photon distributions. However, at two intermediate potentials, i.e., 0.8 and 0.9 V, a dynamic behaviour which is due to the mixed oxidation of Au₃₈ species, in the vicinity of the UME, is observed. In fact, at these two applied potentials, the local concentration of the corresponding gold nanoclusters (i.e., Au₃₈³⁺ and Au₃₈⁴⁺) is not sufficient to produce significant ECL spikes. We also attempted to collect the ECL spectrum using a charge-coupled device (CCD) camera, which is relatively more sensitive in the NIR region (e.g., λ > 900 nm, Fig. S16^†). Fig. 3D and H display an accumulated spectrum at 1.1 and 0.7 V vs. SCE, which is collected for 30 minutes. The fitted accumulated ECL spectrum indicates an ECL peak emission at 930 nm and supports higher reactivity at 1.1 V than that at 0.7 V.³⁸ To confirm that the observed ECL spikes and accumulated spectra are generated based on the oxidation of Au₃₈ nanoclusters in the presence of TPrA radicals, ECL–time transients were recorded upon holding an applied potential at which no faradaic process occurs. Fig. S11^† represents ECL–time curves and accumulated ECL spectra at 0.0 V and 0.4 V. One can notice that no appreciable ECL signal can be observed.In addition, we investigate the Pearson cross-correlation (ρ) between the intensities of ECL spikes with τ as shown in Fig. S14^† in which there is a positive correlation at 0.7 and 1.0 V and a negative correlation at 0.8 and 0.9 V. In fact, ρ evaluates whether there is a stationary random process between the two defined parameters (see ESI, Section 6^†). Interestingly, the frequency of the reaction at different applied potentials revealed decay from 0.7 to 0.8 V, followed by an upward trend to 0.9 and 1.1 V vs. SCE (Fig. S15^†). This could be additional support for the transition stage at 0.8 and 0.9 V, where the applied potential as the major driving force to generate oxidized forms (e.g., Au₃₈³⁺ and Au₃₈⁴⁺) governs the flux of the nanocluster species that reach the vicinity of the electrode. Furthermore, the effectiveness of electron transfer reaction kinetics between the radical species, i.e., Au₃₈^z+1 and TPrA radical, competes with the flow of the incoming nanoclusters. It is worth mentioning that each of the ECL single event experiments was repeated three times, and very similar results were obtained. Moreover, lower (5 μM) and higher (20 μM) concentrations of Au₃₈ in the presence of 20 mM were tested. In fact, the former shows a smaller number of single reactions; however the later revealed a larger number of multiple reactions (Fig. S13^†).In summary, in this communication we demonstrated that Au₃₈ NC ECL at the single reaction level can be monitored using a simple photoelectrochemical setup following a straightforward protocol. Indeed, we have rich basic knowledge about the ECL mechanisms of various gold nanoclusters with different charge states (Au₂₅(SR)₁₈¹⁺, Au₂₅(SR)₁₈⁰, Au₂₅(SR)₁₈¹⁻) and various sizes (Au₂₅(SR)₁₈⁰, Au₃₈(SR)₂₄⁰, Au₁₄₄(SR)₆₀⁰) in fine detail. Thus, the ECL emission mechanisms of gold clusters, including the contribution of each charge state and influence of various concentrations of co-reactants, are well known. For instance, in our previous studies^{38,39,47–49} we clearly identified three charge states of an Au₂₅(SR)₁₈¹⁻/TPrA system and we discovered that at a high concentration of TPrA the reduction in the bulk solution of gold nanoclusters influences the ECL emission wavelength. We also have learnt that the Au₃₈/TPrA system is a co-reactant independent of co-reactant concentration. Furthermore, an extensively higher concentration of TPrA provides a dominant reaction over any unknown decomposition reaction at higher oxidation states of Au₃₈. It was discovered that the population of ECL reactions is directly governed by the applied bias potential on a Pt UME. This work is a strong indication of the high sensitivity of the ECL technique in detecting single ECL reactions in a simple solution, which complements those reported by the Bard group using rubrene, for instance, embedded in an organic emulsion in the presence of TPrA or oxalate as a co-reactant.^50,51 These systems needed a substantial ECL enhancement in the presence of an ionic liquid as the supporting electrolyte and emulsifier. The current approach can be further extended to investigate other molecules and nanomaterials'' electrocatalytic processes at the single reaction level.     

A Paal–Knorr agent for chemoproteomic profiling of targets of isoketals in cells

Natural systems produce various γ-dicarbonyl-bearing compounds that can covalently modify lysine in protein targets via the classic Paal–Knorr reaction. Among them is a unique class of lipid-derived electrophiles – isoketals that exhibit high chemical reactivity and critical biological functions. However, their target selectivity and profiles in complex proteomes remain unknown. Here we report a Paal–Knorr agent, 4-oxonon-8-ynal (herein termed ONAyne), for surveying the reactivity and selectivity of the γ-dicarbonyl warhead in biological systems. Using an unbiased open-search strategy, we demonstrated the lysine specificity of ONAyne on a proteome-wide scale and characterized six probe-derived modifications, including the initial pyrrole adduct and its oxidative products (i.e., lactam and hydroxylactam adducts), an enlactam adduct from dehydration of hydroxylactam, and two chemotypes formed in the presence of endogenous formaldehyde (i.e., fulvene and aldehyde adducts). Furthermore, combined with quantitative chemoproteomics in a competitive format, ONAyne permitted global, in situ, and site-specific profiling of targeted lysine residues of two specific isomers of isoketals, levuglandin (LG) D2 and E2. The functional analyses reveal that LG-derived adduction drives inhibition of malate dehydrogenase MDH2 and exhibits a crosstalk with two epigenetic marks on histone H2B in macrophages. Our approach should be broadly useful for target profiling of bioactive γ-dicarbonyls in diverse biological contexts.

Natural systems produce various γ-dicarbonyl-bearing compounds that can covalently modify lysine in protein targets via the classic Paal–Knorr reaction.

Synthetic chemistry methods have been increasingly underscored by their potential to be repurposed as biocompatible methods for both chemical biology and drug discovery. The most-known examples of such a repurposing approach include the Staudinger ligation¹ and the Huisgen-based click chemistry.² Moreover, bioconjugation of cysteine and lysine can be built upon facile chemical processes,³ while chemoselective labelling of other polar residues (e.g., histidine,⁴ methionine,⁵ tyrosine,⁶ aspartic and glutamic acids^7,8) requires more elaborate chemistry, thereby offering a powerful means to study the structure and function of proteins, even at a proteome-wide scale.The classical Paal–Knorr reaction has been reported for a single-step pyrrole synthesis in 1884.^9,10 The reaction involves the condensation of γ-dicarbonyl with a primary amine under mild conditions (e.g., room temperature, mild acid) to give pyrrole through the intermediary hemiaminals followed by rapid dehydration of highly unstable pyrrolidine adducts (Fig. S1^†).Interestingly, we and others have recently demonstrated that the Paal–Knorr reaction can also readily take place in native biological systems.^11–13 More importantly, the Paal–Knorr precursor γ-dicarbonyl resides on many endogenous metabolites and bioactive natural products.¹⁴ Among them of particular interest are isoketals¹⁵ (IsoKs, also known as γ-ketoaldehydes) which are a unique class of lipid derived electrophiles (LDEs) formed from lipid peroxidation (Fig. S2^†)¹⁶ that has emerged as an important mechanism for cells to regulate redox signalling and inflammatory responses,¹⁷ and drive ferroptosis,¹⁸ and this field has exponentially grown over the past few years. It has been well documented that the γ-dicarbonyl group of IsoKs can rapidly and predominantly react with lysine via the Paal–Knorr reaction to form a pyrrole adduct in vitro (Fig. 1).¹⁵ Further, the pyrrole formed by IsoKs can be easily oxidized to yield lactam and hydroxylactam products in the presence of molecular oxygen (Fig. 1). These rapid reactions are essentially irreversible. Hence, IsoKs react with protein approximately two orders of magnitude faster than the most-studied LDE 4-hydoxynonenal (4-HNE) that contains α,β-unsaturated carbonyl to generally adduct protein cysteines by Michael addition (Fig. S3^†).¹⁵ Due to this unique adduction chemistry and rapid reactivity, IsoKs exhibit intriguing biological activities, including inhibition of the nucleosome complex formation,¹⁹ high-density lipoprotein function,²⁰ mitochondrial respiration and calcium homeostasis,²¹ as well as activation of hepatic stellate cells.²² Furthermore, increases in IsoK-protein adducts have been identified in many major diseases,²³ such as atherosclerosis, Alzheimer''s disease, hypertension and so on.Open in a separate windowFig. 1The Paal–Knorr precursor γ-dicarbonyl reacts with the lysine residue on proteins to form diverse chemotypes via two pathways. The red arrow shows the oxidation pathway, while the blue one shows the formaldehyde pathway.Despite the chemical uniqueness, biological significance, and pathophysiological relevance of IsoKs, their residue selectivity and target profiles in complex proteomes remain unknown, hampering the studies of their mechanisms of action (MoAs). Pioneered by the Cravatt group, the competitive ABPP (activity-based protein profiling) has been the method of choice to analyse the molecular interactions between electrophiles (e.g., LDEs,²⁴ oncometabolites,²⁵ natural products,^26,27 covalent ligands and drugs^28–30) and nucleophilic amino acids across complex proteomes. In this regard, many residue-specific chemistry methods and probes have been developed for such studies. For example, several lysine-specific probes based on the activated ester warheads (e.g., sulfotetrafluorophenyl, STP;³¹N-hydroxysuccinimide, NHS³²) have recently been developed to analyse electrophile–lysine interactions at a proteome-wide scale in human tumour cells, which provides rich resources of ligandable sites for covalent probes and potential therapeutics. Although these approaches can also be presumably leveraged to globally and site-specifically profile lysine-specific targets IsoKs, the reaction kinetics and target preference of activated ester-based probes likely differ from those of γ-dicarbonyls, possibly resulting in misinterpretation of ABPP competition results. Ideally, a lysine profiling probe used for a competitive ABPP analysis of IsoKs should therefore possess the same, or at least a similar, warhead moiety. Furthermore, due to the lack of reactive carbonyl groups on IsoK-derived protein adducts, several recently developed carbonyl-directed ligation probes for studying LDE-adductions are also not suitable for target profiling of IsoKs.^33,34Towards this end, we sought to design a “clickable” γ-dicarbonyl probe for profiling lysine residues and, in combination with the competitive ABPP strategy, for analysing IsoK adductions in native proteomes. Considering that the diversity of various regio- and stereo- IsoK isomers¹⁵ (a total of 64, Fig. S2^†) in chemical reactivity and bioactivities is likely attributed to the substitution of γ-dicarbonyls at positions 2 and 3, the “clickable” alkyne handle needs to be rationally implemented onto the 4-methyl group in order to minimize the biases when competing with IsoKs in target engagement. Interestingly, we reasoned that 4-oxonon-8-ynal, a previously reported Paal–Knorr agent used as an intermediate for synthesizing fatty acid probes³⁵ or oxa-tricyclic compounds,³⁶ could be repurposed for the γ-dicarbonyl-directed ABPP application. With this chemical in hand (herein termed ONAyne, Fig. 2A), we first used western blotting to detect its utility in labelling proteins, allowing visualization of a dose-dependent labelling of the proteome in situ (Fig. S4^†). Next, we set up to incorporate this probe into a well-established chemoproteomic workflow for site-specific lysine profiling in situ (Fig. 2A). Specifically, intact cells were labelled with ONAyne in situ (200 μM, 2 h, 37 °C, a condition showing little cytotoxicity, Fig. S5^†), and the probe-labelled proteome was harvested and processed into tryptic peptides. The resulting probe-labelled peptides were conjugated with both light and heavy azido-UV-cleavable-biotin reagents (1 : 1) via Cu^I-catalyzed azide–alkyne cycloaddition reaction (CuAAC, also known as click chemistry). The biotinylated peptides were enriched with streptavidin beads and photoreleased for LC-MS/MS-based proteomics. The ONAyne-labelled peptides covalently conjugated with light and heavy tags would yield an isotopic signature. We considered only those modified peptide assignments whose MS1 data reflected a light/heavy ratio close to 1.0, thereby increasing the accuracy of these peptide identifications. Using this criterium, we applied a targeted database search to profile three expected probe-derived modifications (PDMs), including 13 pyrrole peptide adducts (Δ273.15), 77 lactam peptide adducts (Δ289.14), and 557 hydroxylactam peptide adducts (Δ305.14), comprising 585 lysine residues on 299 proteins (Fig. S6 and S7^†). Among them, the hydroxylactam adducts were present predominately, since the pyrrole formed by this probe, the same as IsoKs, can be easily oxidized when being exposed to O₂. This finding was in accordance with a previous report where the pyrrole adducts formed by the reaction between IsoK and free lysine could not be detected, but rather their oxidized forms.³⁷ Regardless, all three types of adducts were found in one lysine site of EF1A1 (K387, Fig. S8^†), further confirming the intrinsic relationship among those adductions in situ.Open in a separate windowFig. 2Adduct profile and proteome-wide selectivity of the γ-dicarbonyl probe ONAyne. (A) Chemical structure of ONAyne and schematic workflow for identifying ONAyne-adducted sites across the proteome. (B) Bar chart showing the distribution of six types of ONAyne-derived modifications formed in situ and in vitro (note: before probe labelling, small molecules in cell lysates were filtered out through desalting columns).State-of-the-art blind search can offer an opportunity to explore unexpected chemotypes (i.e., modifications) derived from a chemical probe and to unbiasedly assess its proteome-wide residue selectivity.^38,39 We therefore sought to use one of such tools termed pChem³⁸ to re-analyse the MS data (see Methods, ESI^†). Surprisingly, the pChem search identified three new and abundant PDMs (Fig. 1 and Table S1^†), which dramatically expand the ONAyne-profiled lysinome (2305 sites versus 585 sites). Overall, these newly identified PDMs accounted for 74.6% of all identifications (Fig. 2B and Table S2^†). Among them, the PDM of Δ287.13 (Fig. 1 and S7^†) might be an enlactam product via dehydration of the probe-derived hydroxylactam adduct. The other two might be explained by the plausible mechanism as follows (Fig. 1). The endogenous formaldehyde (FA, produced in substantial quantities in biological systems) reacts with the probe-derived pyrrole adduct via nucleophilic addition to form a carbinol intermediate, followed by rapid dehydration to a fulvene (Δ285.15, Fig. S7^†) and immediate oxidation to an aldehyde (Δ301.14, Fig. S7^†). In line with this mechanism, the amount of FA-derived PDMs was largely eliminated when the in vitro ONAyne labelling was performed in the FA-less cell lysates (Fig. 2B and Table S3^†). Undoubtedly, the detailed mechanisms underlying the formation of these unexpected PDMs require further investigation, and so does the reaction kinetics. Regardless, all main PDMs from ONAyne predominantly target the lysine residue with an average localization probability of 0.77, demonstrating their proteome-wide selectivity (Fig. S9^†).Next, we adapted an ABPP approach to globally and site-specifically quantify the reactivity of lysine towards the γ-dicarbonyl warhead through a dose-dependent labelling strategy (Fig. 3A) that has been proved to be successful for other lysine-specific probes (e.g., STP alkyne).³¹ Specifically, MDA-MB-231 cell lysates were treated with low versus high concentrations of ONAyne (1 mM versus 0.1 mM) for 1 h. Probe-labelled proteomes were digested into tryptic peptides that were then conjugated to isotopically labelled biotin tags via CuAAC for enrichment, identification and quantification. In principle, hyperreactive lysine would saturate labelling at the low probe concentration, whereas less reactive ones would show concentration-dependent increases in labelling. For fair comparison, the STP alkyne-based lysine profiling data were generated by using the same chemoproteomic workflow. Although 77.5% (3207) ONAyne-adducted lysine sites can also be profiled by STP alkyne-based analysis, the former indeed has its distinct target-profile with 930 lysine sites newly identified (Fig. S10 and Table S4^†). Interestingly, sequence motif analysis with pLogo⁴⁰ revealed a significant difference in consensus motifs between ONAyne- and STP alkyne-targeting lysines (Fig. S11^†).Open in a separate windowFig. 3ONAyne-based quantitative reactivity profiling of proteomic lysines. (A) Schematic workflow for quantitative profiling of ONAyne–lysine reactions using the dose-dependent ABPP strategy (B) Box plots showing the distribution of R_10:1 values quantified in ONAyne- and STP alkyne-based ABPP analyses, respectively. Red lines showing the median values. ***p ≤ 0.001 two-tailed Student''s t-test. (C) Representative extracted ion chromatograms (XICs) showing changes in the EF1A1 peptide bearing K273 that is adducted as indicated, with the profiles for light and heavy-labelled peptides in blue and red, respectively.Moreover, we quantified the ratio (R_{1 mM:0.1 mM}) for a total of 2439 ONAyne-tagged lysines (on 922 proteins) and 17904 STP alkyne-tagged lysines (on 4447 proteins) across three biological replicates (Fig. S12 and Table S5^†). Strikingly, only 26.7% (651) of quantified sites exhibited nearly dose-dependent increases (R_{1 mM:0.1 mM} > 5.0) in reactivity with ONAyne, an indicative of dose saturation (Fig. 3B and C). In contrast, such dose-dependent labelling events accounted for >69.1% of all quantified lysine sites in the STP alkyne-based ABPP analysis.³¹ This finding is in accordance with the extremely fast kinetics of reaction between lysine and γ-dicarbonyls (prone to saturation). Nonetheless, by applying 10-fold lower probe concentrations, overall 1628 (80.2%) detected lysines could be labelled in a fully concentration-dependent manner with the median R_10:1 value of 8.1 (Fig. 3B, C, S12 and Table S5^†). Next, we asked whether the dose-depending quantitation data (100 μM versus 10 μM) can be harnessed to predict functionality. By retrieving the functional information for all quantified lysines from the UniProt Knowledgebase, we found that those hyper-reactive lysines could not be significantly over-represented with annotation (Fig. S12^†). Nonetheless, among all quantified lysines, 509 (25.1%) possess functional annotations, while merely 2.5% of the human lysinome can be annotated. Moreover, 381 (74.8%) ONAyne-labelled sites are known targets of various enzymatic post-translational modifications (PTMs), such as acetylation, succinylation, methylation and so on (Fig. S13^†). In contrast, all known PTM sites accounted for only 59.6% of the annotated human lysinome. These findings therefore highlight the intrinsic reactivity of ONAyne towards the ‘hot spots’ of endogenous lysine PTMs.The aforementioned results validate ONAyne as a fit-for-purpose lysine-specific chemoproteomic probe for competitive isoTOP-ABPP application of γ-dicarbonyl target profiling. Inspired by this, we next applied ONAyne-based chemoproteomics in an in situ competitive format (Fig. 4A) to globally profile lysine sites targeted by a mixture of levuglandin (LG) D₂ and E₂, two specific isomers of IsoKs that can be synthesized conveniently from prostaglandin H₂ (ref. 41) (Fig. S2^†). Specifically, mouse macrophage RAW264.7 cells (a well-established model cell line to study LDE-induced inflammatory effects) were treated with 2 μM LGs or vehicle (DMSO) for 2 h, followed by ONAyne labelling for an additional 2 h. The probe-labelled proteomes were processed as mentioned above. For each lysine detected in this analysis, we calculated a control/treatment ratio (R_C/T). Adduction of a lysine site by LGs would reduce its accessibility to the ONAyne probe, and thus a higher R_C/T indicates increased adduction. In total, we quantified 2000 lysine sites on 834 proteins across five biological replicates. Among them, 102 (5.1%) sites exhibited decreases of reactivity towards LGs treatment (P < 0.05, Table S6^†), thereby being considered as potential targets of LGs. Notably, we found that different lysines on the same proteins showed varying sensitivity towards LGs (e.g., LGs targeted K3 of thioredoxin but not K8, K85 and K94, Table S6^†), an indicative of changes in reactivity, though we could not formally exclude the effects of changes in protein expression on the quantified competition ratios. Regardless, to the best of our knowledge, the proteome-wide identification of potential protein targets by IsoKs/LGs has not been possible until this work.Open in a separate windowFig. 4ONAyne-based in situ competitive ABPP uncovers functional targets of LGs in macrophages. (A) Schematic workflow for profiling LGs–lysine interactions using ONAyne-based in situ competitive ABPP. (B) Volcano plot showing the log₂ values of the ratio between the control (heavy) and LGs-treated (light) channels and the −log₁₀(P) of the statistical significance in a two-sample t-test for all quantified lysines. Potential targets of LGs are shown in blue (R_C/T>1.2, P < 0.05), with the validated ones in red. (C) Bar chart showing the inhibitory effect of 2 μM LGs on the cellular enzymatic activity of MDH2. Data represent means ± standard deviation (n = 3). Statistical significance was calculated with two-tailed Student''s t-tests. (D) Pretreatment of LGs dose-dependently blocked ONAyne-labelling of MDH2 in RAW264.7 cells, as measured by western blotting-based ABPP. (E and F) LGs dose-dependently decreased the H2BK5 acetylation level in RAW 264.7 cells, as measured either by western blotting (E) or by immunofluorescence imaging (F). n = 3. For G, nuclei were visualized using DAPI (blue).We initially evaluated MDH2 (malate dehydrogenase, mitochondrial, also known as MDHM), an important metabolic enzyme that possesses four previously uncharacterized liganded lysine sites (K157, K239, K301 and K329, Fig. 4B) that are far from the active site (Fig. S14^†). We found that LGs dramatically reduced the catalytic activity of MDH2 in RAW264.7 cells (Fig. 4C), suggesting a potentially allosteric effect. We next turned our attention to the targeted sites residing on histone proteins, which happen to be modified by functionally important acetylation, including H2BK5ac (Fig. 4B) that can regulate both stemness and epithelial–mesenchymal transition of trophoblast stem cells.⁴² We therefore hypothesized that rapid adduction by LGs competes with the enzymatic formation of this epigenetic mark. Immunoblotting-based competitive ABPP confirmed that LGs dose-dependently blocked probe labelling of H2B (Fig. 4D). Further, both western blots and immunofluorescence assays revealed that LG treatment decreased the level of acetylation of H2BK5 (average R_C/T = 1.3, P = 0.007) in a concentration-dependent manner (Fig. 4E and F). Likewise, a similar competitive crosstalk was observed between acetylation and LG-adduction on H2BK20 (average R_C/T = 1.2, P = 0.01) that is required for chromatin assembly⁴³ and/or gene regulation⁴⁴ (Fig. 4B and S15^†). Notably, these findings, together with several previous reports by us and others about histone lysine ketoamide adduction by another important LDE, 4-oxo-2-noenal,^11,45,46 highlight again the potentially important link between lipid peroxidation and epigenetic regulation. In addition to the targets validated as above, many other leads also merit functional studies considering diverse biological or physiologic effects of LGs in macrophages.     

A therapeutic keypad lock decoded in drug resistant cancer cells

A molecular keypad lock that displays photodynamic activity when exposed to glutathione (GSH), esterase and light in the given order, is fabricated and its efficacy in drug resistant MCF7 cancer cells is investigated. The first two inputs are common drug resistant tumor markers. GSH reacts with the agent and shifts the absorption wavelength. Esterase separates the quencher from the structure, further activating the agent. After these sequential exposures, the molecular keypad lock is exposed to light and produces cytotoxic singlet oxygen. Among many possible combinations, only one ‘key’ can activate the agent, and initiate a photodynamic response. Paclitaxel resistant MCF7 cells are selectively killed. This work presents the first ever biological application of small molecular keypad locks.

Information processing therapeutics with an implemented keypad lock logic gate selects input order for activation in drug resistant cancer cells.

The complex nature of diseases such as cancer necessitates smarter drugs that can discriminate each disease state or regulate drug efficacy spatially and/or temporally. With this intention, activatable drugs, drugs with on demand release properties are developed with promising selectivity.^1–4 Information processing therapeutics which are based on molecular logic gate operations are another approach to solve this problem.^5–7 Molecular logic gates are small compounds using Boolean logic operations to process inputs (i.e. the analyte concentration), and give an output as a result (fluorescence, and therapeutic activity etc.).⁸ Selective drug activation, release, multiple-analyte sensing and theranostic applications of these devices have been explored by us and others.^5,9–19Among the operations that can be carried out using small molecules, keypad locks provide an alternative application in information security.²⁰ This logic operation can give a specific output when the inputs are given in the correct form and correct sequence. For the device, each input is considered as an AND logic operation where the history of the process is also considered. A pioneering example was reported by Margulies and Shanzer in 2007 where energy transfer is modulated by chelation of Fe³⁺ in a pH dependent manner.²¹ Later, various other devices were introduced with advanced properties such as more than 2 input responsiveness and error detection capability.^22–24 All-photonic logic gates to address chemical waste production is extensively studied by Gust, Andréasson and Pischel.^25,26 Beside small molecule keypad locks, enzymes, antibodies, and DNA hybrids are used to achieve the same goal.^27–30 Although their potential use in molecular cryptology is highlighted, so far, there is no solid biological application of small molecule keypad locks.In the research presented here, a molecular keypad lock is developed which displays a photodynamic therapeutic output when a molecule is exposed to analytes in the correct order and type (PS3, Fig. 1). Two inputs of the system are chosen to be the common markers of drug resistant tumours: glutathione (GSH) and esterase enzyme (E). Cancer cells develop resistance to traditional chemotherapy in time by changing the protein expression or metabolite content of the cell. This adaptation of cancer cells is an obstacle for their treatment and needs to be addressed. Glutathione is a tripeptide used in reductive biochemical synthesis and it is known to be present in elevated levels in rapidly dividing cells such as cancer cells.³¹ A high GSH level is reported to contribute to drug resistance, since GSH adducts of the drugs are exported out of the cell much more rapidly.^32,33 Likewise, esterase enzyme activity is known to be associated with drug detoxification as this enzyme contributes to the chemical conversion of the drug.^34,35 Glutathione and esterase enzyme are chosen to be the first two inputs of the molecular keypad lock, the first two digits of the password. In the research, light is used as the final input. Although trivial, light is essential for photodynamic activity and spatiotemporal control of irradiation, further improving selectivity of the therapy.Open in a separate windowFig. 1Chemical structures of model photosensitizers (PS1 and PS2) and a molecular keypad lock (PS3). Ester bonds (red) are prone to hydrolysis by the esterase enzyme. Distyryl sites of the photosensitizers (blue) can react with thiol nucleophile provided that it is bound to an electron deficient group (i.e. pyri-dinium).Keypad lock PS3 is a photodynamic therapy (PDT) agent. PDT is a non-invasive method used for the treatment of surface cancers and certain other diseases ranging from atherosclerosis to macular degeneration.^36–39 In this therapy, a photosensitizer is excited with light, and produces cytotoxic singlet oxygen (¹O₂) thereby triggering apoptosis or necrosis of the cell, initiating an immune response and blocking microvasculature.⁴⁰ In the research, a boradiazaindecene (BODIPY) photosensitizer is used to benefit from versatile chemistry and spectroscopic properties.^41–45Near-IR absorbing PS3 shown in Fig. 1 is the molecular keypad lock and it is synthesized in 13 steps (Scheme S1^†). PS3 and model compound PS2 have heavy atoms on the structure to favour intersystem crossing required for transition to the triplet state and hence ¹O₂ generation occurs.⁴³ Ester bonds on the structure of PS3 are prone to cleavage by esterase enzyme. Distryryl bonds on the PS3 (blue) tend to reduce or form an adduct with thiol nucleophiles when it is activated by the pyridinium electron withdrawing group.⁴⁶ This property lies at the heart of sequential operation of esterase and GSH. When GSH reacts with electron poor double bonds, the extended conjugated structure is broken and PS3-a is generated (Fig. 2). This structure has absorption below 550 nm, like brominated core BODIPY molecules (compound 8, Scheme S1^†), and therefore can be excited with a green light. A quencher (green) is attached to ensure that photodynamic activity is OFF until esterase cleaves the ester bond. This is because of the energy transfer from the photosensitizer to this module, until esterase separates the photosensitizer. Since PS3 lacks absorption around the 500–550 nm region, it is inactive until GSH reacts with the compound. However, the GSH reacted photosensitizer does show absorption in this region; so, in order to avoid full activation just by GSH, a quencher module is attached. Spectral overlap between the BODIPY core (see the structure of compound 8 in the ESI,^† similar to that of PS3-a in terms of conjugation) and quencher (Q) can be seen from UV-Vis absorption and fluorescence spectra (Fig. 3 and S1^†). By this way, the photosensitizer is chemically modulated by GSH to ensure excitation, and then esterase enzyme inhibits energy transfer by removing the quencher. Lastly a green light is used to excite the photosensitizer leading to generation of photodynamic action. Since light is necessary for the final excitation of the molecule, it should always be the last input. If the order of esterase and GSH changes, as shown in Fig. 2, activation is not expected to take place since cleavage of the ester bonds generates 4-hydroxybenzyl derivative on PS3, which spontaneously faces 1,4-elimination to generate pyridine (Fig. S2^†).⁴⁷ Pyridine on its own is not sufficiently electron withdrawing to favour nucleophilic attack of double bonds by GSH and to activate it as demonstrated below. Therefore, the photosensitizer preserves extended conjugation and essentially lacks absorption at the wavelength of excitation.Open in a separate windowFig. 2Sequential operation of GSH and esterase. GSH can only react with BODIPY distyryl units when the structure has electron withdrawing pyridinium, either reducing it or forming an adduct. Esterase enzyme cleaves ester bonds, liberating the photosensitizer from the quencher module (green). Initial esterase activity converts the pyridinium unit to pyridine, thereby decreasing the reactivity of double bonds with GSH.Open in a separate windowFig. 3Normalized UV-Vis absorption and fluorescence spectra of PS1–3 in 2% water in THF (a and b). Samples are excited at 600 nm. Spectral changes of PS3 (10 μM) alone (black) or PS3 upon exposure to 0.5 mM GSH (c) and 10U esterase (d) for 90 min and 60 min at 37 °C, in 2% water in THF, respectively. A new peak at 544 nm appears upon incubation with GSH which is attributed to reduced PS3 and/orthe GSH-adduct. Esterase treatment increases the relative intensity of the shoulder peak around 600 nm.In order to understand the response of the PS3 to GSH, a molecule is incubated with 0.5 mM of GSH at 37 °C for 90 min. A new peak at 544 nm appears in UV-Vis absorption spectra consistent with the hypothesis (Fig. 3c, S1 and S9^†). The formation of the GSH adduct (PS3-a) is demonstrated by Liquid Chromatography Mass spectrometry analysis (Fig. S3^†). When control module PS1 is exposed to the same conditions, this new peak is not detected indicating that the pyridine bearing structure is neither activated enough for the nucleophilic substitution by GSH nor did it display PDT activity (Fig. S4 and S5^†). On the other hand, GSH treated pyridinium bearing PS2 immediately displayed a colour change indicative of broken conjugation (Fig. S6^†). When PS3 is incubated with esterase for 1 h, a small hypsochromic shift in the absorption peak is detected as a shoulder to the parent peak which is attributed to the conversion of pyridinium to pyridine (PS3-c, Fig. 3d). The control PS3 sample which is incubated under the same conditions but lacks esterase does not show an enhancement of this peak (Fig. 3d, black). High Resolution Mass Spectrometry analysis of the esterase treated PS2 samples confirm the hydrolysis of the ester and subsequent formation of the pyridine compound (Fig. S7^†). Esterase treated samples display an increase in the emission intensity when excited at 620 nm (Fig. S8^†). This is attributed to the initial quenching of the quencher module by the pyridinium photosensitizer. Analysis of the absorption and emission spectra suggest that the quencher module of PS3 can induce energy transfer to the pyridinium photosensitizer (Fig. 3). Once separated by esterase, fluorescence of the quencher module increases. In the case of GSH treated sample, a small enhancement in emission upon excitation at 500 nm is observed (Fig. S9^†). Note that the GSH adduct (or PS3 with reduced double bonds) has higher absorption at this wavelength, which would be the reason for the increase in emission intensity. In the spectral analysis organic solvents with a low water content are used to monitor the formation of water-insoluble, neutral, pyridine-bearing intermediate species.In the project, the molecular keypad lock is aimed to unlock in the presence of drug resistant tumour markers and get activated. Activation cannot take place when the input order differs. To demonstrate this, photodynamic action in the presence of all three inputs in a different order is investigated. ¹O₂ production can be followed by using trap molecule, 1,3-diphenylisobenzofuran (DPBF).⁴⁸ This molecule reacts with ¹O₂ and loses its absorption at 418 nm. The effect of different input combinations on the PDT action are given in Fig. 4. In the first 15 min, all samples are kept in the dark. Under such conditions no ¹O₂ generation is detected, which indicates lack of dark activity. DPBF is exposed to light from a LED source (peak 505 nm) under the same experimental conditions and no decrease in the absorption is detected. This control experiment eliminates the photodegradation of DPBF in the absence of a photosensitizer. Upon irradiation before the activation of the photosensitizer by GSH and esterase, no ¹O₂ generation is observed as expected. The results show that ¹O₂ generation, and the subsequent decrease in DPBF absorption, are significantly more in the input order of glutathione, esterase enzyme and light, consistent with the proposed mode of activation.Open in a separate windowFig. 4 ¹O₂ generation ability of PS3 (0.1 μM) when three inputs are given in a different order. All samples contain 50 μM of ¹O₂ trap molecule DPBF. In the first 15 minutes samples are kept in the dark. GSH is added in 0.5 mM concentration and incubated for 90 min at 37 °C. Samples are incubated with 10U esterase for 1 h at 37 °C. An LED light is irradiated from a 30 cm distance for 45 min.To analyse the effect of PDT action in the cell, a drug resistant cell line is generated. MCF7 cells are exposed to an increased dose of traditional cancer therapeutic agent paclitaxel as described in the literature.⁴⁹ When the spindle-shaped morphology is obtained following maximum drug dose application, cells are reported to have drug resistance. At this stage, PS3 is applied to both normal and drug resistant cells. When cell viabilities at various concentrations are analysed, it has been found that the light toxicity of PS3 is significantly enhanced in drug resistant cells (Fig. 5). The IC50 values of irradiated samples are calculated to be 124.8 μM for MCF7 cells. This value is reduced to 52.5 μM in paclitaxel resistant MCF7 (Pac-MCF7) indicating improved cytotoxicity in these cells. Efficient induction of apoptosis is also proved by Annexin V and PI staining (Fig. 6). Under dark conditions, cells do not have significant loss of viability. Upon irradiation, resistant cells are more prone to apoptosis by the photosensitizer. Relative singlet oxygen generation abilities and results of cell culture experiments altogether confirm selective activation in drug resistant cells.Open in a separate windowFig. 5Change in the cell viability of normal and paclitaxel resistant MCF7 cells (Pac-MCF7) in the presence of PS3 at various concentrations. For each group, cell viability is analysed both after incubation in the dark or after irradiation with a 505 nm LED light source from a distance of 10 cm. Average values of three independent experiments are used.Open in a separate windowFig. 6Apoptosis induction by PS3 (25 μM) in normal and paclitaxel resistant MCF7 cancer cells under dark conditions and upon irradiation with a 505 nm LED light from 10 cm distance. Scale bars: 50 μm.     

Modular assembly of MOF-derived carbon nanofibers into macroarchitectures for water treatment

The organized assembly of nanoparticles into complex macroarchitectures opens up a promising pathway to create functional materials. Here, we demonstrate a scalable strategy to fabricate macroarchitectures with high compressibility and elasticity from hollow particle-based carbon nanofibers. This strategy causes zeolitic imidazolate framework (ZIF-8)-polyacrylonitrile nanofibers to assemble into centimetre-sized aerogels (ZIF-8/NFAs) with expected shapes and tunable functions on a large scale. On further carbonization of ZIF-8/NFAs, ZIF-8 nanoparticles are transformed into a hollow structure to form the carbon nanofiber aerogels (CNFAs). The resulting CNFAs integrate the properties of zero-dimensional hollow structures, one-dimensional nanofibers, and three-dimensional carbon aerogels, and exhibit a low density of 7.32 mg cm⁻³, high mechanical strength (rapid recovery from 80% strain), outstanding adsorption capacity, and excellent photo-thermal conversion potential. These results provide a platform for the future development of macroarchitectured assemblies from nanometres to centimetres and facilitate the design of multifunctional materials.

A scalable strategy is established to generate macroarchitectures based on MOF-related nanofibers. The modular assembly of macroarchitectures with luffa-like structures exhibits high mechanical strength and low densities.

The assembly of simple nanoparticles (such as silica, polystyrene and metal–organic frameworks) into macroarchitectures has a unique attraction for engineering materials due to their variable sizes, shapes, and chemical and physical properties.^1–3 As a novel nanomaterial, the formed macroarchitecture with three-dimensional (3D) porous interconnected network structures has broad application prospects in various fields, including environment treatment, chemical sensing, energy storage, catalysis, and advanced electronic devices.^4–7 Moreover, the functions of macroarchitectures are mainly determined by the fundamental building blocks. On account of large surface area, high porosity and more exposure to active sites, the complex macroarchitectures, which are assembled by building blocks with hollow structures, possess greater advantages.^8,9In the past few decades, in order to seek high-performance hollow building blocks for macroarchitectures, much effort has been put into it. In particular, the emergence of one-dimensional (1D) carbon hollow nanostructures, including hollow porous carbon nanofibers (HPCNs)¹⁰ and carbon nanotubes (CNTs)¹¹ promotes the rapid development of this field. The HPCN- and CNT-based macroarchitectures realize the transformation from 1D nanomaterials to three-dimensional (3D) macroscopic materials with excellent properties (e.g., electrochemical energy storage and antimicrobial air filtration).^12,13 Furthermore, these macroarchitectures can not only retain the characteristics of the 1D material, but also generate many new kinds of features (e.g., high specific surface area, high mechanical strength, and low density) that the components do not possess. Unfortunately, though the HPCN- and CNT-based macroarchitectures exhibit improved conductivity and stability properties, the synthesis of their building blocks is usually expensive and complex.¹⁴ Besides, the assembly of building blocks into 3D macroarchitectures usually exhibits relatively poor mechanical properties and requires some adhesives or templates, which have to be eliminated by extra strategies.¹⁵ These complex synthetic procedures and less favourable structural stability largely hinder the scale-up production of carbon aerogels and their practical applications.To address these issues, we explore a novel and scalable method to synthesize functional macroarchitectures with robust mechanical properties fabricated from MOF-derived carbon nanofibers through manipulating nano-sized particles (MOFs) and micron-sized fibers. First, zeolitic imidazolate framework (ZIF-8) nanoparticles, which are spectacular for their large nitrogen content and surface area,^16–18 are embedded into polyacrylonitrile (PAN)/polyvinylpyrrolidone (PVP) nanofibers to form ZIF-8-PAN/PVP composite nanofibers. Subsequently, the ZIF-8-PAN/PVP nanofibers are assembled into centimetre-sized nanofiber aerogels (ZIF-8/NFAs) by a freeze-drying technique. After preoxidation and carbonization of ZIF-8/NFAs, the carbon nanofiber aerogels with hollow and porous interlayer structures are fabricated (named C-ZIF-8-CNFAs). The interlayer structure of C-ZIF-8-CNFAs is very similar to natural luffa consisting of a network of elastic frameworks. As with the interconnected nanofibers in the interlayer structure, cellulose skeletons are in the interior of luffa interconnected in a highly uniform manner to maximize strength, a porous structure. Therefore, we refer to our porous structure as luffa-like. The prepared C-ZIF-8-CNFAs exhibit a low density of 7.32 mg cm⁻³, high specific surface area (288 m² g⁻¹), large hierarchical pore volume (0.22 cm⁻³ g⁻¹), high mechanical strength (rapid recovery from 80% strain), outstanding adsorption capacity, and excellent photo-thermal conversion potential. Fig. 1a depicts the assembly strategy of macroarchitectures (C-ZIF-8-CNFAs) schematically. The preparation process begins with the fabrication of ZIF-8/nanofibers (Fig. S1b^†) using the method that ZIF-8 nanoparticles (Fig. S1a^†) are incorporated into nanofibers by electrospinning (specific preparation methods in the ESI^†). Upon homogenization in a mixed solution of ultrapure water and tert-butanol, the ZIF-8/nanofibers become wrapped around each other and dispersed uniformly. Subsequently, the homogenized nanofiber dispersion is frozen in a mold followed by freeze-drying into uncrosslinked ZIF-8/NFAs. To build further robust bonding among nanofibers, the obtained uncrosslinked ZIF-8/NFAs are preoxidized at 250 °C to form crosslinked ZIF-8/NFAs with a welding structure under the action of polyvinylpyrrolidone (PVP), providing elastic resilience to the resultant NFAs. Ultimately, the resulting preoxidized ZIF-8/NFAs are carbonized at 900 °C to form hollow C-ZIF-8-CNFAs under a N₂ atmosphere. As a result of the carbonization, the organic ligands of ZIF-8 and molecules of PAN and PVP are decomposed and transformed into N-doped carbon materials. In addition, the Zn²⁺ in ZIF-8 nanoparticles is reduced to metallic Zn and then evaporated at high temperature.^19–21 Nanoscale 0D MOFs are assembled into micron-scale 1D fibers, which are then assembled into centimeter-scale 3D carbon aerogels (Fig. 1b). This novel approach enables the super assembly on a multi-dimensional scale, which realizes the macroscopic application of nanoparticles and the functionalization of CAs. In order to verify that the performance of C-ZIF-8-CNFAs is improved after the introduction of ZIF-8, pure CNFAs (Fig. S2^†) without ZIF-8 (the synthetic process is shown in the ESI^†) are also prepared.Open in a separate windowFig. 1Preparation steps for C-ZIF-8-CNFAs. (a) More fabrication details for C-ZIF-8-CNFAs. (b) Schematic illustration of the fabrication of CNFAs.As illustrated in Fig. 2a, when the ZIF-8/nanofiber dispersion solution is frozen, the uniformly dispersed ZIF-8/nanofibers are extruded by the growth of ice crystals and assembled among ice crystals. After the sample is frozen completely, the nanofibers become lapped and locked into a 3D nanofibrous network. Subsequently, the ZIF-8/NFAs with a luffa-like structure are obtained after the sublimation of ice crystals through the freeze-drying process.²² Moreover, ZIF-8/NFAs can be made into diverse desired shapes such as cylinders, cubes, moon-like shapes, star-like shapes, heart-like shapes and intricate shapes of the letters (Fig. 2b). Fig. 2c shows the obvious reduction of the intensity of ZIF-8/NFA XRD patterns compared to the original ZIF-8, but the site hardly changes, which confirms that the introduced ZIF-8 nanoparticles are not destroyed during the electrospinning and the preparation process of aerogels. After the preoxidation and carbonization steps, the typical C-ZIF-8-CNFAs with an ultra-low density of 7.32 mg cm⁻³ can freely stand on the tip of a red maple leaf (Fig. 2b). The scanning electron microscopy (SEM) images in Fig. 2d–f show that C-ZIF-8-CNFAs have the hierarchical porous luffa-like structure with three kinds of pores (the picture of an actual luffa shown in the inset of Fig. 2d). The porous structure exhibits obvious rectangular pores of ∼25 μm, and the wall of these pores is made of interconnecting nanofibers (Fig. 2e). Meanwhile, the secondary pores of ∼1.5 μm are formed by the welded nanofibers that are interconnected with each other. The nanoscale pores of ∼200 nm also exist in these nanofibers, which come from the carbonization of ZIF-8 nanoparticles (Fig. 2f). As observed from the magnified SEM image (Fig. S3^†), the welded structure resulting from preoxdiation is still preserved through carbonization. Remarkably, ZIF-8 nanoparticles encapsulated by PAN and PVP are transformed into a hollow structure after carbonization. As can be seen from the TEM image (Fig. 2g), the ZIF-8-derived hollow structure is evenly dispersed in PAN/PVP-derived carbon nanofibers. Because Zn²⁺ ions coordinate with –C N groups existing on the surface of PAN/PVP nanofibers, the ZIF-8 particles become tightly encapsulated by the PAN/PVP layers. During carbonization, the PAN/PVP layers make ZIF-8 shrink from inside to outside, thereby leading to the generation of a hollow structure. Moreover, the confined carbonization process within the PAN/PVP matrix prevents the irreversible fusion and aggregation of carbonized ZIF-8 nanoparticles.¹⁹Open in a separate windowFig. 2(a) Schematical illustration of the formation principles for the hierarchical cellular structure. (b) Photographs of ZIF-8/NFAs with diverse shapes and the lightweight C-ZIF-8-CNFAs standing on the tip of a red maple leaf. (c) Wide-angle XRD patterns. (d–g) SEM (d–f) and TEM (g) images showing the microstructure of C-ZIF-8-CNFAs at various magnifications. (h and i) N₂ adsorption–desorption isotherm and pore-size distribution curve of CNFAs and C-ZIF-8-CNFAs.After ZIF-8 nanoparticles were introduced into CNFAs, the properties have been improved significantly. To comprehend the variation of the porous characteristics in C-ZIF-8-CNFAs, nitrogen (N₂) adsorption–desorption measurements were carried out. As observed from Fig. 2h and i, C-ZIF-8-CNFAs have a larger specific surface area of 288.3 m² g⁻¹ and pore volume of 0.22 cm⁻³ g⁻¹, while the specific surface area and pore volume of CNFAs are only 12.1 m² g⁻¹ and 0.01 cm⁻³ g⁻¹, respectively. Meanwhile, C-ZIF-8-CNFAs also have a hierarchical porous structure with micropores, mesopores, and macropores (Fig. 2i). Because of the existence of the hierarchical porous structure in C-ZIF-8-CNFAs, they have a lower density relative to CNFAs (20.73 mg cm⁻³) (Table S1^†). The chemical compositions and graphitic structure of C-ZIF-8-CNFAs are investigated by X-ray powder diffraction (XRD) and X-ray photoelectron spectroscopy (XPS). The XRD pattern (Fig. S4a^†) of C-ZIF-8-CNFAs only exhibits two broad peaks at about 25° and 44°, corresponding to the (002) and (101) diffraction facets of the graphitic structure, respectively.²³ The correlative XPS spectrum shows that C-ZIF-8-CNFAs consist of C, N and O (Fig. S4b^†). The high-resolution N 1s spectra (Fig. S4c^†) can be deconvoluted into four peaks: pyridinic N (398.6 eV), pyrrolic N (399.4 eV), graphitic N (400.9 eV), and oxidized N (403.7 eV), respectively. The percentage of nitrogen and its types are listed in Table S2.^† There is no Zn content thus indicating that it had evaporated during the carbonization process at high temperature (at 900 °C for 3 hours).²⁴ Furthermore the N content of C-ZIF-8-CNFAs increases with the introduction of ZIF-8 compared to CNFAs clearly (Table S3^†).In stark contrast to the hard and brittle characteristics of conventional carbon aerogels, C-ZIF-8-CNFAs show robust mechanical properties, sustaining large compressive strain without fracture (insets in Fig. 3a and movie S1^†). The compressive stress–strain (σ–ε) curves (Fig. 3a) show the compressive process of C-ZIF-8-CNFAs and two typical deformation regimes could be recognized: a Hookean or linear elastic regime of ε < 50% with a stable tangent modulus, and a densification regime of ε > 50% with σ and dσ/dε increasing sharply. When the maximum compressive strain increases from 30% to 80%, the maximum compressive stress steeply increases from 1.2 to 25.9 kPa, indicating that C-ZIF-8-CNFAs can bear over 25 000 times their own weight without cracking. Moreover, the cyclic compression test of the C-ZIF-8-CNFAs is performed to validate their durable cycling performance by applying 50 loading–unloading fatigue cycles at a large ε of 50% (Fig. 3b). C-ZIF-8-CNFAs hardly undergo plastic deformation at all after the 50th cycle, which is a huge improvement over traditional CAs with a crisp character. As shown in Fig. 3c, C-ZIF-8-CNFAs retain nearly 100% of the initial value of the Young''s modulus, maximum stress and energy loss coefficient, indicating that their strength or stiffness has no significant decline highlighting their exceptional structural robustness. Two reasons could account for the excellent mechanical elasticity of C-ZIF-8-CNFAs. On the one hand, the compressive strain is absorbed by the bending of the connected ZIF-8/nanofibers between layers; on the other hand, the strain that continues to grow (beyond 50%) is absorbed by the densification of minor pores, which is formed by the welding of PVP (Fig. 3d).²⁵ The two reasons can also be confirmed by the sharp increase of the compressive stress after ε > 50% (Fig. 3a).Open in a separate windowFig. 3(a) Compressive stress–strain curve of C-ZIF-8-CNFAs at different strains. The insets are photographs of C-ZIF-8-CNFAs under a compressing and releasing cycle (ε = 80%). (b) A 50-cycle compressive fatigue test with ε of 50%. (c) The Young^''s modulus, energy loss coefficient, and maximum stress versus compressive cycles. (d) Sketch of the changes in the hierarchical porous structure with compressive deformation.MOF-based derived CNFAs with hollow structures, ultra-low density, extraordinary mechanical elasticity, and large surface area are conducive to applications in various fields, including environment governance, energy absorption, and energy storage.^26–28 As a proof of concept, we evaluated the organic solvent absorption and photo-thermal conversion performance of C-ZIF-8-CNFAs. Excellent hydrophobicity is one of the important factors to ensure the absorption efficiency of organic solvents. As demonstrated in Fig. 4a, C-ZIF-8-CNFAs exhibit high hydrophobicity with a water contact angle of 142° and the water contact angle has no significant change after 120 s. The water droplet and the absorbed oil droplet are on the surface of C-ZIF-8-CNFAs, indicating the effective oil/water selectivity of C-ZIF-8-CNFAs (Fig. S5^†). As shown in Fig. 4b, C-ZIF-8-CNFAs demonstrate extraordinary absorption capacities for common oils and various organic solvents, 90–200 times their own weight, principally depending on the density and viscosity of the solvents. For example, the adsorption capacity for carbon tetrachloride (ρ = 1.595 g cm⁻³) is much higher than the adsorption capacity for n-hexane (ρ = 0.66 g cm⁻³). We also compared the maximum adsorption capacity of C-ZIF-8-CNFAs and CNFAs for several common organic solvents and oils. The adsorption performance of C-ZIF-8-CNFAs is also demonstrated to be better than that of CNFAs (Fig. S6^†). According to the cross-sectional diagram of the adsorption of organic solvents (Fig. 4c), CNFAs only absorb organic solvents through the capillary phenomenon of the channel, which is formed by nanofibers, while the absorbed organic solvent can also enter into the hollow structure formed by carbonization of ZIF-8 inside the nanofiber of C-ZIF-8-CNFAs. Therefore, the performance of C-ZIF-8-CNFAs is significantly improved after the addition of ZIF-8 nanoparticles. Moreover, the adsorption capacity of C-ZIF-8-CNFAs for organic solvents is greater than that of previously reported aerogels (Fig. S7^†). To verify the excellent adsorption performance of C-ZIF-8-CNFAs in practical application, the oil/water separation and cyclic adsorption experiments were carried out. As illustrated in Fig. S8,^† C-ZIF-8-CNFAs can quickly absorb heavy organic solvents such as carbon tetrachloride (dyed with oil red) sunk at the bottom of water, thus indicating their potential application for selectively removing oils from water. Recyclability and reusability are also crucial to evaluate the practical application possibility of adsorption materials. In view of the outstanding elasticity and structural robustness of C-ZIF-8-CNFAs, we chose ethanol as the absorption solvent for recycling tests. Through simple heating, ethanol absorbed by C-ZIF-8-CNFAs can be readily removed. As demonstrated in Fig. S9a and b,^† even after 10 cycles, C-ZIF-8-CNFAs still retained over 90% adsorption capacity for ethanol and their mass was reduced by less than 10%.Open in a separate windowFig. 4(a) Dynamic behaviors of a water droplet on the surface of C-ZIF-8-CNFAs. (b) Adsorption efficiency of C-ZIF-8-CNFAs towards commonly used organic solvents and oils. (c) Schematic illustration of the organic solvent adsorption process inside the CNFAs and C-ZIF-8-CNFAs. (d) Mass changes of evaporated water versus time under 1 sun illumination. (e) The temperature of C-ZIF-8-CNFAs and seawater under 1 sun illumination as a function of irradiation time. (f) The infrared images (IR) show the temperature distribution of C-ZIF-8-CNFAs and seawater under 1 sun illumination with an irradiation time of 0, 30, and 60 min.Given the abundant porous structure and blackbody characteristic, C-ZIF-8-CNFAs are also promising materials for interfacial solar steam generation (ISSG). To investigate the ISSG performance, the evaporation mass change of seawater and C-ZIF-8-CNFAs is measured under one sun illumination. As shown in Fig. 4d, the seawater in C-ZIF-8-CNFAs achieves the maximum evaporation rate of 3.74 kg m⁻² h⁻¹, which is 5.12 times the evaporation rate of bulk seawater (0.73 kg m⁻² h⁻¹) and 2.34 times faster than the evaporation rate of general 2D ISSG (∼1.6 kg m⁻² h⁻¹). An infrared camera is used to trace the surface temperatures of C-ZIF-8-CNFAs and bulk seawater under one-sun illumination to evaluate the photothermal behavior of ISSG (Fig. 4e). The surface temperature of C-ZIF-8-CNFAs presents a quick increase in 5 min and eventually reaches a stable state (∼48 °C) after 10 min, while the bulk seawater temperature stays unchanged (∼25 °C). The consecutive infrared images in Fig. 4f show the equilibrium temperature distribution and heat localization effect of C-ZIF-8-CNFAs and bulk seawater in 60 min under one-sun illumination.     

Single-atom cobalt-hydroxyl modification of polymeric carbon nitride for highly enhanced photocatalytic water oxidation: ball milling increased single atom loading

Expediting the oxygen evolution reaction (OER) is the key to achieving efficient photocatalytic overall water splitting. Herein, single-atom Co–OH modified polymeric carbon nitride (Co-PCN) was synthesized with single-atom loading increased by ∼37 times with the assistance of ball milling that formed ultrathin nanosheets. The single-atom Co-N₄OH structure was confirmed experimentally and theoretically and was verified to enhance optical absorption and charge separation and work as the active site for the OER. Co-PCN exhibits the highest OER rate of 37.3 μmol h⁻¹ under visible light irradiation, ∼28-fold higher than that of common PCN/CoO_x, with the highest apparent quantum yields reaching 4.69, 2.06, and 0.46% at 400, 420, and 500 nm, respectively, and is among the best OER photocatalysts reported so far. This work provides an effective way to synthesize efficient OER photocatalysts.

Single-atom Co^II-OH modified polymeric carbon nitride synthesized with increased single-atom loading under the assistance of ball milling exhibits high photocatalytic water oxidation activity with Co-N₄OH as the highly active site.

Massive fuel energy consumption induced environmental and ecological problems, especially the greenhouse effect, and the resultant extreme climates and rise in sea level are threatening human life.¹ As a potential substitution for fuel energy, hydrogen energy conversion from solar energy via photocatalytic water splitting attracts great attention from scientists.^2–5 However, the photocatalytic hydrogen evolution efficiency from overall water splitting is still restricted by the sluggish oxygen evolution reaction (OER) that involves energy absorption, four-electron transfer, breakage of O–H bonds, and formation of O–O bonds,^6,7 and thus efficient OER photocatalysts become the key to achieving efficient overall water splitting. Though numerous hydrogen evolution photocatalysts have been reported, research on OER photocatalysts is mainly around a few semiconductors including BiVO₄, WO₃, Ag₃PO₄, α-Fe₂O₃, etc.8–11 and their activity is not high enough yet for practical applications. Therefore, exploring high-efficiency OER photocatalysts is still necessary.Polymeric carbon nitride (PCN) was first reported in 2009 (ref. 12) as a photocatalyst with a layered melon-type orthorhombic structure,¹³ and thereafter quickly became a “star” photocatalyst thanks to its advantages of being visible-light responsive and metal-free, non-toxic, and low cost, and its relatively high chemical stability.¹⁴ Because of several self-deficiencies including fast photogenerated charge recombination and a narrow optical absorption spectrum, PCN exhibits relatively low photocatalytic activity.¹⁵ Then, a series of strategies were put forward successively to enhance the photoactivity of PCN, such as enhancement of crystallinity,¹⁶ morphological control,¹⁷ structural modification¹⁸ (including extensively researched single atom modification in recent years^19,20), exfoliation,²¹ construction of hetero-(homo-)junctions,²² and loading of noble metals.²³ Though photocatalytic water splitting on PCN was extensively researched in the past, the research was mainly around the hydrogen evolution half-reaction used for exploring properties and the catalytic mechanism of photocatalysts, and little research was focused on the industrially useable overall water splitting process owing to the sluggish OER.¹⁵ Therefore, enhancing the photocatalytic OER activity of PCN becomes the key to practical applications.To increase OER rates of PCN, several kinds of methods were proposed, such as rational design of compound cocatalysts (e.g., CoO_x, IrO₂, CoP, CoPi, RhO_x, RuO_x, PtO_x, MnO_x, Co(OH)₂, Ni(OH)₂, and CoAl₂O₄ (ref. 24–30)), modification of carbon dots and carbon rings,^31,32 fabrication of special architectures of PCN (e.g., PCN quantum dot stacked nanowires³³), and single-atom (e.g., B, Co, and Mn^34–36) modification. For instance, Zhao and coauthors prepared B and N-vacancy comodified PCN that exhibits the highest OER rate of ∼28 μmol h⁻¹ (ref. 36) and recently their group further used these B doped PCN ultrathin nanosheets to fabricate a Z-scheme heterojunction for overall water splitting with a solar-to-hydrogen efficiency reaching ∼1.2%.³⁷ Comparatively, PCN loaded with compound cocatalysts can only enhance OER activity to a limited degree and there are finite methods for carbon modification and special architecture fabrication. Single-atom modification shows a bright prospect, on account of metal atoms capable of being inserted into the framework of PCN and effectively increasing the OER activity. However, reported single metal atom modification routes are all based on direct ion adsorption on PCN or calcination of mixtures of metal salts and PCN feedstocks.^34,35,38 New routes need be explored to increase effective loading of single atoms in PCN. Besides, the metal-OH structure is considered efficient for the OER,^30,39,40 and a single metal atom-OH structure has never been reported for modification of PCN, though Mn–OH was thought to play a key role in the OER process.³⁴Ball milling is an extensively used versatile and scalable way for preparation of heterogeneous catalysts and even single-atom catalysts,^41,42 but was rarely used in synthesis of PCN-based single-atom photocatalysts. In this work, we synthesized single-atom Co–OH modified PCN (Co-PCN) with the single-atom content in PCN highly increased with the assistance of ball milling. The simple synthetic route is shown in Fig. 1a. PCN was ball-milled to obtain BM-PCN that then adsorbed Co²⁺ till saturation to form BM-PCN/Co which was calcined to obtain BM-PCN/Co-c (Co-PCN). For comparison, PCN was directly used to adsorb Co²⁺ till saturation to form PCN/Co which was calcined to obtain PCN/Co-c. PCN mainly comprises large blocks with the size of several micrometers (Fig. S1^†), while BM-PCN contains massive irregular particles with the size reduced to several hundreds of nanometers (Fig. S2^†), indicative of high efficacy of ball milling. BM-PCN/Co-c exhibits a similar morphology as BM-PCN (Fig. 1b and S3^†) and PCN/Co-c exhibits a similar morphology to PCN (Fig. S4^†), but the surface area and mesopore volume of BM-PCN and BM-PCN/Co-c are not higher than those of PCN and PCN/Co-c (Fig. S5^†), manifesting that ball-milling and subsequent calcination did not form massive mesopores, which accords well with the particle size variation from several micrometers (before ball milling) to several hundreds of nanometers (after ball milling). However, the Co content in BM-PCN/Co-c, BM-PCN/Co, PCN/Co-c, and PCN/Co was measured to be 0.75, 0.50, 0.02, and ∼0.02 wt%, respectively, by inductively coupled plasma mass spectrometry (ICP-MS). The ∼37 times higher Co content in BM-PCN/Co-c than in PCN/Co-c suggests the ball-milling enhanced adsorption of Co²⁺ on surfaces of BM-PCN, which should arise mainly from the ball-milling induced increase of surface energy and adsorption sites.⁴³Open in a separate windowFig. 1(a) Schematic illustration for synthesis of single-atom Co^II-OH modified PCN (BM-PCN/Co-c); and (b) SEM, (inset in b) TEM, (c) AFM, (d) EDS elemental mapping, and (e) HAADF-STEM images of BM-PCN/Co-c.The TEM image shows the existence of small and ultrathin nanosheets in BM-PCN/Co-c (inset in Fig. 1b) which can also be observed in the atomic force microscopy (AFM) image with a thickness of ∼7–10 nm and lateral size of <70 nm (Fig. 1c), and formation of these ultrathin nanosheets results from the ball milling of PCN.⁴⁴ It should be noted that most formed ultrathin nanosheets with high surface energy may stack into compact particles upon ball milling, leading to no increase of the total surface area. Energy dispersive X-ray spectroscopy (EDS) elemental mapping images of BM-PCN/Co-c indicate homogeneous distribution of C, N, O, and Co elements in the sample (Fig. 1d). The high-angle annular dark-field scanning transmission electron microscopy (HAADF-STEM) image of BM-PCN/Co-c shows massive white spots (marked by circles) with a mean size of <1 Å dispersed in the sample (Fig. 1e and S6^†), which should correspond to single-atom Co.To further verify the single-atom Co structure in BM-PCN/Co-c, Co K-edge X-ray absorption near-edge structure spectroscopy (XANES) and extended X-ray absorption fine structure (EXAFS) analysis were performed. As shown in Fig. 2a, the absorption-edge position of BM-PCN/Co-c is quite close to that of CoO and their peak positions are similar and far from those of other reference samples, indicating that the valence of Co in BM-PCN/Co-c is about +2. The bonding structure around Co was determined by Fourier transformed (FT) k³-weighted EXAFS analysis. As shown in Fig. 2b, a distinct single Co-ligand peak at ∼1.6 Å for BM-PCN/Co-c is observed, which prominently differs from the Co–Co coordination peak at ∼2.2 Å for Co foil and the Co^II–O coordination peak at ∼1.7 Å for CoO. The wavelet transform (WT) contour plot of BM-PCN/Co-c shows only one intensity maximum (Fig. S7^†), and the Cl 2p core-level XPS spectrum of BM-PCN/Co-c reveals no residue of Cl (Fig. S8^†). These further indicate the single-atom dispersion of Co species. Apparently, the Co-ligand peak is almost consistent with the Co^II–N peak for Co porphyrin, suggesting that the single-atom Co in BM-PCN/Co-c mainly coordinates with N. Least-square EXAFS curve fitting was performed to confirm quantitative structural parameters of Co^II in BM-PCN/Co-c, as shown in Fig. 2c, S9, and S10 and Table S1.^† Simple Co–N single-shell fitting of BM-PCN/Co-c (Fig. S10^†) gave a coordination number of 5.6 ± 0.4 (Table S1^†), that is, Co^II coordinates with five atoms. Considering that the PCN monolayer provides four appropriate N coordination sites at most,⁴⁵ Co^II likely coordinates with four N atoms and one OH atom. Thus, we further performed Co–N₄/Co–O double-shell fitting (Fig. 2c) and the obtained R-factor (0.0011) remarkably reduces relative to that from Co–N single-shell fitting (0.0035), indicative of rationality of the proposed Co^II–N₄OH structure. Confirmed Co–N and Co–O bond lengths are 2.04 and 2.15 Å, respectively (Table S1^†).Open in a separate windowFig. 2(a) Co K-edge XANES and (b) EXAFS spectra of Co foil, Co porphyrin (Copr), CoO, Co₃O₄, Co₂O₃, and BM-PCN/Co-c; EXAFS (c) R space-fitting and (inset in c) K space-fitting curves of BM-PCN/Co-c; (d) optimized structure of PCN and Co-doped PCN with different doping configurations and calculated formation energies (e) of Co doped PCN; and (e) Co 2p and (f) O 1s core-level XPS spectra of samples.To further confirm rationality of the Co–N₄OH coordination structure, density functional theory (DFT) calculations were conducted. As shown in Fig. 2d, three possible Co^II coordination structures in the PCN monolayer were explored. The Co–N₄OH structure without removal of H from PCN exhibits a much lower formation energy (∼0.15 eV) than Co–N₄ and Co–N₃ structures with removal of two H atoms from PCN (∼2.51 and 3.55 eV), demonstrating a high probability of existence of the Co–N₄OH structure in BM-PCN/Co-c. This structure can also be evidenced by X-ray photoelectron spectroscopy (XPS). As shown in Fig. 2e, the Co 2p core-level XPS spectrum of BM-PCN/Co-c shows two distinct peaks at binding energies of 796.8 and 781.4 eV beside satellite peaks, corresponding to Co 2p_1/2 and 2p_3/2 of Co^II ions.⁴⁶ The spectrum of BM-PCN/Co also shows two Co 2p peaks but at a binding energy ∼1.1 eV higher, suggesting variation of the Co coordination structure from BM-PCN/Co to BM-PCN/Co-c. PCN/Co-c exhibits no peaks because of its low Co content. Fig. 2f shows O 1s core-level spectra of PCN, BM-PCN, BM-PCN/Co-c, and PCN/Co-c. All the samples exhibit one peak at a binding energy of ∼532.0 eV, ascribed to surface hydroxyl species,⁴⁷ but an additional peak could be obtained for BM-PCN or BM-PCN/Co-c after deconvolution. The peak at a binding energy of ∼531.3 eV for BM-PCN should be assigned to adsorbed H₂O at new active adsorption sites generated by ball milling. This peak can also be observed in the spectrum of BM-PCN/Co, but with a ∼0.1 eV shift to a higher binding energy (Fig. S11^†) owing to the influence of adsorbed Co^II ions. The peak at ∼531.2 eV for BM-PCN/Co-c should be assigned to Co–OH,⁴⁸ given that there is only one O 1s peak for BM-PCN-c (synthesized by direct calcination of BM-PCN) (Fig. S11^†). The calculated Co/O(–Co) molar ratio, based on the XPS data, is ∼1.07 (Table S2^†), close to 1, consistent with the Co–N₄OH coordination structure.In C 1s and N 1s core-level XPS spectra, BM-PCN, BM-PCN/Co-c, PCN/Co-c, and BM-PCN/Co exhibit similar peaks to PCN (Fig. S12a–d^†), indicative of their similar framework structure which can also be evidenced by their similar N/C molar ratios, 1.53 (Table S3^†), but the N–H peak of BM-PCN shifts ∼0.2 eV to a lower binding energy relative to that of PCN, likely arising from the ball-milling induced destruction of intralayer hydrogen bonds (Fig. S13^†). The Co content in BM-PCN/Co, BM-PCN/Co-c, and PCN/Co-c is too low to cause detectable variation of C 1s and N 1s peaks. Similar FT-IR absorption bands of the samples (Fig. S14a and b^†) also indicate their basic frame structure, but in enlarged spectra (Fig. S14c^†), ν(C–N) and ν(C N) absorption bands of BM-PCN shift 16 cm⁻¹ to a higher wavenumber and 19 cm⁻¹ to a lower wavenumber, respectively, relative to those of PCN at 1242 and 1640 cm⁻¹,⁴⁹ likely resulting from the ball-milling induced hydrogen bond destruction, and the shift of these two absorption bands turns smaller for BM-PCN/Co-c, suggesting calcination-induced reforming of the destroyed hydrogen bonds, which is consistent with the XPS results (Fig. S12c^†). Besides, BM-PCN exhibits a wider and relatively stronger ν(N–H)/ν(O–H) absorption band than PCN (Fig. S14a^†), probably owing to the hydrogen bond destruction and new adsorbed H₂O, while this absorption band for BM-PCN/Co-c becomes much weaker, suggesting hydrogen bond reforming and loss of new adsorbed H₂O (Fig. 2f). Zeta potentials of the samples dispersed in water reflect variation of surface adsorbed hydroxyl species. As shown in Fig. S15a,^† all the samples exhibit negative zeta potentials because of dissociation of surface hydroxyl species. The zeta potentials, following the order PCN (−24 mV) > BM-PCN (−41 mV) < BM-PCN/Co-c (−30 mV) ≈ PCN/Co-c (−28 mV), suggest the ball milling-induced increase of surface hydroxyls in BM-PCN and calcination-induced decrease in BM-PCN/Co-c, consistent with the FT-IR results.Solid-state ¹³C magic-angle-spinning nuclear magnetic resonance (NMR) spectra of PCN, BM-PCN, BM-PCN/Co-c, and PCN/Co-c show two similar peaks at chemical shifts of ∼164 and 156 ppm (Fig. S15b^†), ascribed to C−NH_x and N C–N, respectively,⁵⁰ indicating their similar molecular framework, but in enlarged spectra, BM-PCN exhibits ∼0.3° movement of the N C–N peak to a lower chemical shift compared with PCN, because of the ball-milling induced hydrogen bond destruction, and the C−NH_x peak of BM-PCN/Co-c moves ∼0.2° to a lower chemical shift, likely owing to formation of the C–N–Co structure whose peak lies close to the C−NH_x peak.⁵¹ The XRD patterns of the samples are shown in Fig. S15c.^† PCN and PCN/Co-c exhibit typical diffraction peaks of melon-type carbon nitride with a layered orthorhombic structure and peaks at 13.1° and 27.6° correspond to (210) and (002) facets, respectively,^13,52 but BM-PCN reveals remarkably decreased peak intensity and ∼0.2° shift of the (002) peak to a lower 2θ (indicative of the increased interlayer distance) relative to PCN, demonstrating the ball-milling induced hydrogen bond destruction and substantial decrease of crystallinity. The remarkable decrease of crystallinity and almost no change of the surface area of BM-PCN, compared with those of PCN, further suggest that ball milling may form massive thin nanosheets (Fig. 1c) most of which stack into compact particles (Fig. 1b) owing to their high surface energy. In comparison with BM-PCN, BM-PCN/Co-c exhibits a narrower (002) peak, suggesting enhanced crystallinity owing to the calcination-induced hydrogen bond reforming, consistent with the FT-IR results. On the whole, it is likely the ball-milling induced destruction of hydrogen bonds that contributes largely to the increase of surface energy and new active adsorption centers and thus Co²⁺ adsorption on BM-PCN.Optical absorption capability of samples was investigated by UV-vis diffuse reflectance spectroscopy (DRS). As shown in Fig. 3a, BM-PCN/Co-c, BM-PCN, and PCN/Co-c exhibit considerably higher, lower, and similar optical absorption than/to PCN, respectively. For BM-PCN/Co-c, the optical absorption enhancement at a wavelength of <400 nm may benefit from the electron-rich Co that enhances π–π* transitions in heptazine rings,⁵³ and the Urbach tail absorption should arise from the Co–OH doping.^54,55 Bandgaps (E_g) of PCN, BM-PCN, BM-PCN/Co-c, and PCN/Co-c were roughly confirmed as 2.70, 2.81, 2.56, and 2.73 eV, respectively, via the formula E_g/eV = 1240/(λ_ed/nm)⁵⁶ where λ_ed is the absorption edge determined by solid lines in the spectra. The wider E_g of BM-PCN probably results from the quantum size effect of massive ultrathin crystal nanosheets (Fig. 1c) formed by ball milling, and the narrower E_g of BM-PCN/Co-c arises from the Co–OH doping that was then verified by DFT calculations. As shown in Fig. S16,^† the calculated E_g of BM-PCN/Co-c, ∼1.90 eV, is much smaller than that of PCN (2.57 eV), in accordance with the experimental results. For PCN, the conduction band (CB) is contributed by C 2p and N 2p orbitals and the valence band (VB) mainly by N 2p orbitals, while for BM-PCN/Co-c, the CB is contributed by Co 3d, C 2p, and N 2p orbitals and the VB mainly by Co 3d and N 2p orbitals (Fig. S16c and d^†), effectively manifesting that the narrowing of E_g of BM-PCN/Co-c results from the Co–OH doping. In addition, there are prominent doping levels (E_d) in the bandgap of BM-PCN/Co-c, mainly contributed by Co 3d and O 2p orbitals (Fig. S16d^†), effectively proving the Co–OH doping effect in BM-PCN/Co-c. Similar calculation results have been reported for Pt–OH modified carbon nitride.⁵⁷ Given that the experimental Co content (0.75 wt%) is much lower than the theoretical (6.71 wt%), practical doping levels in the bandgap may approach more to the VB. CB edges of the samples (E_CB) could be roughly determined by using Mott-Schottky plots (Fig. S17^†) and their Fermi levels (E_f) were subsequently confirmed based on VB-XPS spectra (Fig. S18^†). Energy band levels of the samples are shown in Fig. 3b, and it seems that ball milling causes a slight downshift of the VB edge (E_VB) of BM-PCN, favorable for photocatalytic water splitting, but the Co–OH doping causes a slight downshift of E_CB and upshift of E_VB of BM-PCN/Co-c. It is noteworthy that the E_d close to the VB edge (E_VB) can capture photogenerated holes⁵⁸ and thus the single-atom Co–OH works as the active site for the OER (Fig. 3b).Open in a separate windowFig. 3(a) UV-vis diffuse reflectance spectra of PCN, BM-PCN, BM-PCN/Co-c, and PCN/Co-c; (b) energy band levels of the samples and schematic illustration for water oxidation on BM-PCN/Co-c; (c) photoluminescence spectra, (d) time-resolved fluorescence spectra, and (e) anodic photocurrent (J_a) response of the samples; and (f) EPR spectra of the samples in the dark and under visible light irradiation. Data in (d) are the results of fitting decay curves to a tri-exponential model. Dark J_a in (e) was set as zero for distinct comparison.Spectroscopy and photoelectrochemical tests were conducted to evaluate photogenerated charge separation and transfer performance. As shown in Fig. 3c, photoluminescence (PL) spectra of all the samples show one emission peak, basically corresponding to their bandgap emission. BM-PCN exhibits weaker PL intensity than PCN, revealing a decreased photogenerated charge recombination efficiency, which originates from faster charge transfer from the inside to the surface of ultrathin nanosheets (Fig. S19^†) and trapped by surface states.⁵⁹ BM-PCN/Co-c exhibits the lowest PL intensity and the PL intensity of PCN/Co-c is lower than that of PCN, which arises from the E_d capturing photogenerated holes to reduce their direct recombination with electrons beside the ultrathin nanosheet effect in BM-PCN/Co-c. Fig. 3d shows time-resolved fluorescence spectra of the samples. Decay curves were well fitted to a tri-exponential model (S3) and the obtained results are shown in Fig. 3d. Three lifetimes (τ₁–τ₃) and their mean lifetime (τ_m, 89.2 ns) of BM-PCN are all much longer than those of PCN (τ_m = 17.9 ns), further suggesting the faster charge transfer from the inside to the surface of ultrathin nanosheets in BM-PCN, decreasing the direct charge recombination efficiency, but with subsequent surface radiative recombination.⁶⁰ Interestingly, the τ₁–τ₃ and τ_m (10.8 ns) of BM-PCN/Co-c are much shorter than those of PCN, which should result from faster transfer of holes to E_d that effectively decreases the charge recombination efficiency, with subsequent nonradiative energy transformation.⁶¹ The Co–OH doping effect also makes PCN/Co-c exhibit shorter τ₁–τ₃ and τ_m (16.5 ns) than PCN. Fig. 3e shows the photocurrent response of the samples. Their anodic photocurrent density follows the order PCN < PCN/Co-c < BM-PCN < BM-PCN/Co-c, indicating gradually increased photogenerated charge separation efficiencies,⁶² basically consistent with the PL results. The relatively high photocurrent response of BM-PCN benefits from the applied bias that effectively inhibits surface recombination of photogenerated charge carriers.To assess charge mobility of the samples, their electrochemical impedance spectroscopy (EIS) spectra were tested with high-frequency data simply fitted to an equivalent circuit (Fig. S20^†). The obtained charge transfer resistance (R_ct) follows the order PCN (26 Ω) > BM-PCN (18 Ω) ≈ PCN/Co-c (19 Ω) > BM-PCN/Co-c (13 Ω). Apparently, BM-PCN/Co-c exhibits smaller R_ct than BM-PCN and PCN/Co-c, and PCN/Co-c exhibits smaller R_ct than PCN, indicating the highest charge transfer performance of BM-PCN/Co-c⁶³ which originates from the single-atom Co modification⁶⁴ that may increase the electron density to facilitate charge transport. The smaller R_ct of BM-PCN than that of PCN indicates the additional favorable effect of ultrathin nanosheets.⁶⁵Fig. 3f shows electron paramagnetic resonance (EPR) spectra of the samples. All reveal one single Lorentzian line centered at a g of 2.0039, attributed to unpaired electrons in heptazine rings.⁶⁶ In the dark, the EPR signal intensity follows the order PCN < BM-PCN < PCN/Co-c < BM-PCN/Co-c, and the stronger signal of BM-PCN than that of PCN results from formation of ultrathin nanosheets that enhances delocalization of unpaired electrons, while the stronger signal of BM-PCN/Co-c and PCN/Co-c mainly benefits from the Co doping that increases the delocalized electron density.⁶⁷ Under visible light irradiation, the samples exhibit remarkable signal enhancement, following the sequence PCN < BM-PCN < PCN/Co-c < BM-PCN/Co-c, similar to that of the signal intensity in the dark, suggesting that the increase in the delocalized electron density facilitates charge photoexcitation. The high delocalized electron density favors charge transport, consistent with the EIS results, and the high photoexcited charge density benefits enhancement of photocatalytic activity.Photocatalytic OER activity of various samples was well evaluated using Ag⁺ as the sacrificial agent (Fig. S21^†). The Co content in BM-PCN/Co-c was optimized according to the photocatalytic OER rates and BM-PCN-c exhibits no detectable OER activity (Fig. S22^†), indicating indispensability of the Co–OH structure for the OER. The influence of the calcination temperature (T_c °C) of BM-PCN/Co on OER rates of BM-PCN/Co-c (T_c = 460) and BM-PCN/Co-cT_c was investigated and BM-PCN/Co-c exhibits the highest photoactivity (Fig. 4a), manifesting that the optimal calcination temperature is 460 °C. Under both simulated solar light and visible light irradiation (λ ≥ 420 nm), BM-PCN/Co-c exhibits substantially higher OER activity than PCN/Co-c (Fig. 4b), further suggesting the significance of the single-atom Co loading amount, and remarkably higher activity than common PCN/CoO_x (with 0.75 wt% Co, obtained via photodeposition) and BM-PCN-c/Co(OH)₂ (with 0.75 wt% Co), demonstrating the high efficacy of the single-atom distribution of Co–OH in BM-PCN/Co-c. Besides, urea was used as the feedstock to synthesize carbon nitride (marked as PCN-urea) with a larger surface area (76 m² g⁻¹ (ref. 68)) than PCN, and PCN-urea was further used to synthesize PCN-urea/Co-c similar to the synthesis of BM-PCN/Co-c. The OER activity of BM-PCN/Co-c is prominently higher than that of PCN-urea/Co-c (with the optimized Co content and Co single atom distribution, Fig. S23^†), suggesting the significant role of ball milling in fabricating the single-atom Co–N₄OH structure. To quantitively compare photoactivity of the samples, their mean OER rates under visible light illumination for 2 h are shown in Fig. 4c. The OER rate of BM-PCN/Co-c can reach ∼37.3 μmol h⁻¹, about 13.8, 28.7, 2.6, and 2.0 times those of PCN/Co-c, PCN/CoO_x, BM-PCN-c/Co(OH)₂, and PCN-urea/Co-c, respectively. Comparatively, less N₂ was generated for BM-PCN/Co-c (Fig. S24^†), further demonstrating the significance of single-atom Co–OH modification.Open in a separate windowFig. 4(a) The influence of the calcination temperature (T_c °C) of BM-PCN/Co on photocatalytic OER activity of BM-PCN/Co-c (T_c = 460) and BM-PCN/Co-cT_c, under Xe-lamp illumination, with AgNo₃ as the sacrificial agent; (b) photocatalytic oxygen evolution on various samples under Xe-lamp illumination with or without using a 420-nm filter; (c) corresponding OER rates of the samples in 2 h; (d) photocatalytic OER rates of BM-PCN/Co-c under irradiation with various monochromatic light sources for 12 h; (e) apparent quantum yields (AQYs) of BM-PCN/Co-c at different wavelengths and reaction times and the highest AQY at every wavelength, along with the UV-DRS spectrum; and (f) proposed mechanism for photocatalytic water oxidation on the single-atom Co^II-OH structure.Photocatalytic oxygen evolution on BM-PCN/Co-c was also tested under monochromatic light irradiation (Fig. S25^†). Apparently, BM-PCN/Co-c can exhibit OER activity even at a wavelength of 500 nm. The mean OER rate in 12 h decreases from 1.85 to 0.54 μmol h⁻¹ with increasing wavelengths from 400 to 500 nm (Fig. 4d), independent of light intensity of the Xe lamp and is mainly dependent on optical absorption capability of BM-PCN/Co-c at various wavelengths (Fig. 3a). Fig. 4e shows apparent quantum yields (AQYs) of BM-PCN/Co-c at different reaction times and wavelengths. Basically, there are maxima of AQYs with increasing reaction time at every wavelength, suggesting the adverse effect of excessive photodeposited Ag on surfaces of samples. These maxima are shown in Fig. 4e and accord well with the UV-vis DRS spectrum with increasing wavelengths. The maxima of AQYs at 400, 420, 450, and 500 nm can reach 4.69, 2.06, 1.07, and 0.46%, respectively. Compared with the reported photocatalytic OER results for PCN (Table S4^†), BM-PCN/Co-c exhibits the top-class performance.To investigate chemical stability of BM-PCN/Co-c, the cyclic OER experiment was conducted. After five consecutive runs, OER rates of BM-PCN/Co-c decrease less (Fig. S26a^†), with the morphology similar to the original (Fig. S26b^†). Co single atoms in the sample could still be distinctly observed by HAADF-STEM (Fig. S26c and d^†). In addition, N 1s core-level XPS spectra of BM-PCN/Co-c are almost similar before and after the cyclic experiment (Fig. S26e^†). These indicate the high stability of the basic framework structure of the sample. However, Co 2p core-level spectra show remarkable differences before and after the experiment, not only the Co^II peak shift, probably owing to ion (e.g., IO₄⁻) adsorption, but also formation of a large amount of Co^III (Fig. S26f^†). Coexistence of Co^II/Co^III may suggest the photocatalytic OER mechanism.The proposed OER mechanism based on the Co–OH structure is shown in Fig. 4f, according to the reported results in Mn doped PCN.³⁴ Four holes are needed to complete four oxidation steps and obtain one O₂ molecule. The first step starting with one hole may involve formation of the Co^III O bond. The Co–N₄OH structure should facilitate the water oxidation more compared with that of Co–N₄ without OH coordination, by leaving out the initial adsorption process of H₂O molecules.³⁴ On the whole, the high photocatalytic OER activity of Co-PCN benefits from the Co–N₄OH structure that not only effectively enhances optical absorption, and charge separation and transport, but also works as the highly active site for the OER.     

Hydroxy-directed fluorination of remote unactivated C(sp3)–H bonds: a new age of diastereoselective radical fluorination

We report a photochemically induced, hydroxy-directed fluorination that addresses the prevailing challenge of high diastereoselectivity in this burgeoning field. Numerous simple and complex motifs showcase a spectrum of regio- and stereochemical outcomes based on the configuration of the hydroxy group. Notable examples include a long-sought switch in the selectivity of the refractory sclareolide core, an override of benzylic fluorination, and a rare case of 3,3′-difluorination. Furthermore, calculations illuminate a low barrier transition state for fluorination, supporting our notion that alcohols are engaged in coordinated reagent direction. A hydrogen bonding interaction between the innate hydroxy directing group and fluorine is also highlighted for several substrates with ¹⁹F–¹H HOESY experiments, calculations, and more.

We report a photochemical, hydroxy-directed fluorination that addresses the prevailing challenge of high diastereoselectivity. Numerous motifs showcase a range of regio- and stereochemical outcomes based on the configuration of the hydroxy group.

The hydroxy (OH) group is treasured and versatile in chemistry and biology.¹ Its ubiquity in nature and broad spectrum of chemical properties make it an attractive source as a potential directing group.² The exploitation of the mild Lewis basicity exhibited by alcohols has afforded several elegant pathways for selective functionalization (e.g., Sharpless epoxidation,³ homogeneous hydrogenation,⁴ cross-coupling reactions,⁵ among others⁶). Recently, we reported a photochemically promoted carbonyl-directed aliphatic fluorination, and most notably, established the key role that C–H⋯O hydrogen bonds play in the success of the reaction.⁷ Our detailed mechanistic investigations prompt us to postulate that other Lewis basic functional groups (such as –OH) can direct fluorination in highly complementary ways.⁸ In this communication, we report a hydroxy-directed aliphatic fluorination method that exhibits unique directing properties and greatly expands the domain of radical fluorination into the less established realm governing high diastereoselectivity.⁹Our first inclination that functional groups other than carbonyls may influence fluorination regiochemical outcomes was obtained while screening substrates for our published ketone-directed radical-based method (Scheme 1).^8a In this example, we surmised that oxidation of the tertiary hydroxy group on substrate 1 cannot occur and would demonstrate functional group tolerance (directing to C11, compound 2). Surprisingly, the two major regioisomers (products 3 and 4) are derivatized by Selectfluor (SF) on C12 and C16 – indicative of the freely rotating hydroxyl directing fluorination. Without an obvious explanation of how these groups could be involved in dictating regiochemistry, we continued the mechanistic study of carbonyl-directed fluorination (Scheme 2A). We established that the regioselective coordinated hydrogen atom abstraction occurs by hydrogen bonding between a strategically placed carbonyl and Selectfluor radical dication (SRD).⁷ However, we noted that the subsequent radical fluorination is not diastereoselective due to the locally planar nature of carbonyl groups. Thus, we posed the question: are there other directing groups that can provide both regio- and diastereoselectivity? Such a group would optimally be attached to a sp³ hybridized carbon; thus the “three dimensional” hydroxy carbon logically comes to mind as an attractive choice, and Scheme 1 illustrates the first positive hint.Open in a separate windowScheme 1Observed products for the fluorination of compound 1.Open in a separate windowScheme 2(A) Proposed mechanism, (B) β-caryophyllene alcohol hypochlorite derivative synthetic probe, (C) isodesmic relation of transition states showing the general importance of the hydroxy group to reactivity (ωB97xd/6-31+G*), and (D) ¹H NMR experiment with Selectfluor and various additives at different concentrations.We began our detailed study with a simple substrate that contains a tertiary hydroxyl group. Alcohol 5 was synthesized stereoselectively by the reaction of 3-methylcyclohexanone, FeCl₃, and 4-chlorophenylmagnesium bromide;¹⁰ the 4-chlorophenyl substituent allows for an uncomplicated product identification and isolation (aromatic chromophore). We sought to determine optimal reaction conditions by examination of numerous photosensitizers, bases, solvents, and light sources (7 Although we utilize cool blue LEDs (sharp cutoff ca. 400 nm), CFLs (small amount of UVB (280–315 nm) and UVA (315–400 nm)) are useable as well.¹¹ A mild base additive was also found to neutralize adventitious HF and improve yields in the substrates indicated (

Reversing electron transfer in a covalent triazine framework for efficient photocatalytic hydrogen evolution

Covalent triazine-based frameworks (CTFs) have emerged as some of the most important materials for photocatalytic water splitting. However, development of CTF-based photocatalytic systems with non-platinum cocatalysts for highly efficient hydrogen evolution still remains a challenge. Herein, we demonstrated, for the first time, a one-step phosphidation strategy for simultaneously achieving phosphorus atom bonding with the benzene rings of CTFs and the anchoring of well-defined dicobalt phosphide (Co₂P) nanocrystals (∼7 nm). The hydrogen evolution activities of CTFs were significantly enhanced under simulated solar-light (7.6 mmol h⁻¹ g⁻¹), more than 20 times higher than that of the CTF/Co₂P composite. Both comparative experiments and in situ X-ray photoelectron spectroscopy reveal that the strong interfacial P–C bonding and the anchoring of the Co₂P cocatalyst reverse the charge transfer direction from triazine to benzene rings, promote charge separation, and accelerate hydrogen evolution. Thus, the rational anchoring of transition-metal phosphides on conjugated polymers should be a promising approach for developing highly efficient photocatalysts for hydrogen evolution.

Reversing the electron transfer in a covalent triazine-based framework by Co₂P anchoring achieved highly efficient photocatalytic hydrogen evolution from water splitting.

Photocatalytic water splitting into hydrogen fuels has been considered as a promising technique for converting solar energy into chemical energy.^1–3 To achieve this target, it is necessary to design and construct photocatalysts with high solar-to-hydrogen (STH) conversion efficiency.^4,5 Recently, covalent triazine-based frameworks (CTFs),^6,7 as a new class of conjugated polymer materials, have attracted significant attention in the photocatalytic water splitting field owing to their visible-light response, organized architecture, adjustable pore-size, and controllable functionalization.^8–12 However, owing to the high charge recombination and fewer active sites for the hydrogen evolution reaction (HER), the photocatalytic activities of pristine CTFs are very low. To overcome this drawback, noble-metal platinum (Pt) is generally required in CTF-based photocatalytic systems as the HER cocatalyst for promoting charge separation and catalyzing hydrogen generation.^13–15 However, the high cost and scarcity of metallic Pt greatly limit the large-scale commercial applications. Thus, it is highly desirable to explore economical materials as noble-metal substitutes for achieving comparable or even superior hydrogen evolution activities.Recently, transition-metal phosphides (TMPs) have attracted intensive attention as HER cocatalysts for photocatalytic water splitting, owing to their unique structural and electronic properties.^16–20 Up to now, a variety of semiconductor materials, including metal oxides,^21,22 metal sulfides,^23,24 g-C₃N₄,^25,26 MOFs,^27,28etc., decorated with TMPs have been extensively investigated, and the hydrogen generation activity in some reports is even higher than that of Pt cocatalysts. However, related studies about the decoration of TMP cocatalysts on CTF photocatalysts for highly efficient hydrogen generation have not been reported so far. Taking into account the molecular structure of CTFs, the nitrogen sites in the triazine frameworks could easily coordinate with transition metal ions to form nitrogen–metal interactions.^29–31 In contrast, the coordination of TMPs with CTFs is relatively difficult owing to the high electronegativity of P sites which could draw electrons from metal atoms.¹⁷ Moreover, most reported TMPs were fabricated directly from precursor metal salts, oxides, etc.,^32–35 and the resultant large-dimensions usually lead to very limited contact-interface with CTFs, or even a physical mixture form. Accordingly, the charge transfer between TMPs and CTFs is significantly restrained. Thus, effective anchoring of TMPs on CTFs for achieving highly efficient photocatalytic hydrogen evolution still remains a great challenge.Herein, a one-step phosphidation strategy has been developed to achieve phosphorus bonding with the benzene rings of CTFs and the anchoring of well-defined dicobalt phosphide (Co₂P) nanocrystals (∼7 nm). The photocatalytic results clearly reveal that an excellent hydrogen evolution activity (7.6 mmol h⁻¹ g⁻¹) is achieved, which is much higher than that of the CTF/Co₂P composite (0.37 mmol h⁻¹ g⁻¹). More detailed studies confirm that the phosphidation strategy could effectively facilitate the interfacial bonding between CTFs and Co₂P nanocrystals. More importantly, the SI-XPS results clearly suggest that the charge transfer direction in CTFs is completely reversed, and the photo-generated electrons efficiently transferred from the triazine rings to P-bonded benzene rings, where the Co₂P cocatalyst attracted electrons through the interfacial P–C bonds for efficient hydrogen evolution. To our knowledge, this is the first report on incorporating TMPs on a CTF photocatalyst for enhancing the hydrogen evolution activity. Fig. 1A shows the basic procedures for the preparation of CTF polymer anchored Co₂P nanocrystals. Briefly, the CTFs with adsorbed cobalt ions were directly phosphatized by thermal decomposition of NaH₂PO₂ under an Ar atmosphere, which could simultaneously achieve P atom bonding with benzene rings and the anchoring of the Co₂P cocatalyst (marked as P-CTF-Co₂P). Fig. 1B shows the typical transmission electron microscopy (TEM) image of the obtained sample, clearly revealing that well-defined Co₂P nanocrystals with an average diameter of 5∼9 nm were uniformly dispersed on the CTF surface. Furthermore, the high-resolution TEM (HR-TEM) image (Fig. 1C) of the formed nanocrystals exhibited two lattice fringes with d-spacing values of 0.20 and 0.21 nm, respectively, which could be well indexed to the (211) and (121) planes of orthorhombic Co₂P crystals.^36,37Fig. 1D shows the X-ray diffraction (XRD) pattern of the obtained P-CTF-Co₂P sample. For comparison, the pristine CTF, and Co₂P nanocrystals have also been studied. It can be clearly seen that for the P-CTF-Co₂P sample, except for the two broad diffraction peaks at 7.3° and 26.1° attributed to the in-plane (100) facets and interlayer (001) stacking of CTFs,^38,39 the other XRD peaks well matched with those of the Co₂P crystals,^40,41 confirming the successful incorporation of the Co₂P cocatalyst in the CTFs. Furthermore, energy-dispersive X-ray (EDX) elemental mapping (Fig. 1E) clearly reveals the uniform distribution of C, N, Co and P elements in the whole detection region, further confirming the uniform dispersion of Co₂P on CTF polymers.Open in a separate windowFig. 1(A) Basic procedures and the ideal structure scheme for fabricating P-CTF-Co₂P catalysts; (B) TEM image and (C) HR-TEM image of the P-CTF-Co₂P catalyst; (D) XRD patterns of CTFs, Co₂P and P-CTF-Co₂P catalysts; (E) EDX elemental mapping images of the P-CTF-Co₂P catalyst.To further explore the P-bonding sites, high-resolution X-ray photoelectron spectroscopy (XPS) was performed on both P-CTF-Co₂P and pristine CTFs. As shown in Fig. 2A, for pristine CTF samples, the C 1s peak could be well fitted into two peaks located at 284.8 and 286.8 eV, which could be assigned to C–C C and N–C N bonds,^38,39,42 respectively. Notably, after the phosphidation treatment, a new peak at 285.3 eV attributed to P–C bonds was detected in the P-CTF-Co₂P sample,^43–45 and the C–C C to N–C N ratio was significantly decreased from 10.2 (pristine CTFs) to 7.1 (P-CTF-Co₂P). In contrast, compared with the N 1s spectrum of the pristine CTF sample, no evident change of peak shape and intensity could be detected in the P-CTF-Co₂P sample (Fig. 2B). These results clearly reveal that after the phosphidation treatment, P atoms should mainly bond with the carbon sites of the benzene rings in CTFs instead of the triazine rings.^46,47 To further confirm this inference, XPS studies on phosphatized CTFs without Co₂P nanocrystals (marked as P-CTFs) were also performed. It can be clearly seen from Fig. S9^† that similar changes for the C 1s and N 1s peaks of P-CTF-Co₂P (Fig. 2A and B) are observed in the P-CTF sample. Moreover, in both P-CTF-Co₂P and P-CTF samples, an evident P 2p peak corresponding to P–C bonds could be observed (Fig. 2C),⁴⁸ further indicating the bonding of P atoms with C atoms on the benzene rings in CTFs after the phosphidation treatment. However, note that the binding energy (BE) position of P–C peaks in P-CTF-Co₂P (133.6 eV) is slightly higher than that of P-CTF (133.2 eV), which should be attributed to the anchoring with Co₂P nanocrystals. In addition to the XPS investigations, solid-state cross-polarization magic angle spinning carbon-13 and phosphorus-31 nuclear magnetic resonance (¹³C-NMR and ³¹P-NMR) spectroscopy studies were further performed to explore the bonding sites of phosphorus atoms in the P-CTF (Fig. S10^†). The Fourier-transformed infrared (FTIR) spectrum was also employed (Fig. 2D). It can be clearly observed that the typical FTIR peaks of the triazine frameworks and the stretching vibrations of carbon–nitrogen (C–N) (1521 cm⁻¹ and 1354 cm⁻¹) in P-CTF-Co₂P are consistent with those of pristine CTFs.^38,39 However, the peaks at 1672 cm⁻¹ and 1014 cm⁻¹ corresponding to the stretching vibrations of C C and C–H in benzene rings were evidently decreased in P-CTF-Co₂P compared with those of pristine CTFs.^49,50 Combining the results of XPS, NMR, and FTIR, it can be concluded that in the obtained P-CTF-Co₂P sample, the carbon sites of the benzene rings in the CTFs were partially bonded with P-atoms, which should anchor Co₂P through P–Co bonding.Open in a separate windowFig. 2High-resolution XPS spectra of (A) C 1, (B) N 1s and (C) P 2p in CTF, P-CTF and P-CTF-Co₂P samples (the insets show the molecular structures); (D) Fourier transform infrared spectroscopy (FT-IR) of CTF and P-CTF-Co₂P samples.Furthermore, the hydrogen evolution activities of the P-CTF-Co₂P (2 wt% Co₂P) sample were evaluated under simulated solar light irradiation. For comparison, the CTF/Co₂P composite and pristine CTFs were also measured under the same conditions. As shown in Fig. 3A, the P-CTF-Co₂P sample exhibits a much higher H₂ evolution activity (15.2 mmol g⁻¹) than the CTF/Co₂P composite (0.7 mmol g⁻¹) at 2 h, while no evident hydrogen generation could be detected in the pristine CTF sample. To further explore the crucial roles of interfacial P-bonding in CTFs and the anchoring of the Co₂P cocatalyst, the hydrogen evolution rates of various samples were calculated and their comparison is shown in Fig. 3B. Obviously, compared with the excellent activity of P-CTF-Co₂P (7.6 mmol h⁻¹ g⁻¹), CTF/Co₂P and P-CTF/Co₂P only exhibit very low hydrogen evolution rates of 0.37 and 0.38 mmol h⁻¹ g⁻¹, respectively, while no evident activity could be detected for CTFs, Co₂P, and P-CTF samples. The above results clearly confirm that the significant enhancement of the hydrogen evolution activity of the P-CTF-Co₂P sample should be mainly attributed to the P-bonding and the anchoring of the Co₂P cocatalyst. Furthermore, the photocatalytic performances of CTFs decorated with Pt nanoparticles were also studied (the inset of Fig. 3B, Fig. S15^†), as Pt is generally recognized as the most active cocatalyst for hydrogen generation. Amazingly, the hydrogen production rates of the Pt cocatalyst at different amounts (1∼5 wt%) were all lower than that of the P-CTF-Co₂P sample, indicating that the rational bonding of the Co₂P cocatalyst on CTFs should be a promising strategy for achieving highly efficient photocatalytic hydrogen evolution.Open in a separate windowFig. 3(A) Photocatalytic H₂ evolution tests of the as-prepared photocatalysts; (B) comparative presentation of the hydrogen evolution rates; (C) ultraviolet-visible diffuse reflectance spectrum of the P-CTF-Co₂P photocatalyst (red solid line) and wavelength-dependent hydrogen production activities of the P-CTF-Co₂P photocatalyst within 2 h; light source: a 300 W Xe lamp equipped with various cut-off filters. (D) The cycling photocatalytic tests of P-CTF-Co₂P and the CTF/Co₂P composite; (E) I–t curves of various catalysts at −0.3 V (vs. SCE); (F) electrochemical impedance spectroscopy under light of the various catalysts. Measurements were conducted in 0.2 mol L⁻¹ Na₂SO₄ electrolyte solution under AM 1.5 G illumination.Furthermore, the relationship between wavelength and the hydrogen evolution activity of the P-CTF-Co₂P photocatalyst was studied and is shown in Fig. 3C. With increasing the wavelength from full-spectrum light irradiation (λ > 300) to 500 nm, the hydrogen evolution rate significantly decreased from 7.6 to 0.06 mmol h⁻¹ g⁻¹, which is generally consistent with their absorption spectrum. The highest value of the apparent quantum yields (AQYs) attained is 31.8% at 365 nm. Moreover, the photocatalytic stability and durability of P-CTF-Co₂P and CTF/Co₂P were examined by cycling experiments. As shown in Fig. 3D, the CTF/Co₂P composite demonstrated relatively poor stability, and there is no evident activity after only one cycling experiment, resulting from the incompact combination between CTFs and Co₂P (Fig. S5^†). In contrast, the P-CTF-Co₂P sample exhibits relatively high stability, and no evident inactivation was detected during the whole test. These results further confirm that in addition to enhancing activities, the interfacial P–C bonds and the anchoring of the Co₂P cocatalyst could also effectively promote the hydrogen evolution stability. To further confirm the efficient charge separation and electron transfer in P-CTF-Co₂P, photoelectrochemical (PEC) tests were performed. As shown in Fig. 3E, the amperometric I–t curves clearly reveal that P-CTF-Co₂P exhibits higher photocurrent density than pristine CTFs and CTF/Co₂P samples, confirming its more efficient charge separation capability. Moreover, electrochemical impedance spectroscopy (EIS) was also performed to explore the interfacial charge transfer process.^51,52 As shown in Fig. 3F, the P-CTF-Co₂P sample with the smallest arc radius revealed remarkably increased interfacial charge transport efficiency. These PEC results clearly demonstrate the highly efficient photogenerated charge separation and transfer in the P-CTF-Co₂P photocatalyst, which are highly consistent with the above photocatalytic hydrogen evolution results.The photo-induced charge separation and transfer in the excited state are crucial processes for determining the photocatalytic activity. Herein, we have demonstrated an in situ irradiation X-ray photoelectron spectroscopy (SI-XPS) technique for exploring the intrinsic charge separation and transfer mechanisms between Co₂P and CTFs. As shown in Fig. S20 and S21,^† no evident binding energy (BE) shift could be observed in both pristine CTF and Co₂P samples, confirming their relatively poor charge separation capability. Amazingly, for the P-CTF-Co₂P sample (Fig. 4), distinct variations for C 1s, N 1s, P 2p, and Co 2p peaks were detected in the excited state. More specifically, the C 1s and N 1s peaks in the CTFs shifted towards the high BE region by 0.3 eV accompanied by the broadening of peak shape (Fig. 4B and C), while the P 2p and Co 2p peaks of the Co₂P cocatalyst shifted towards the low BE direction by 0.3 and 0.4 eV (Fig. 4D and E), respectively.Interestingly, as shown in Fig. 4D, the P 2p peaks attributed to interfacial P–C bonds exhibited no evident BE change under dark and light irradiation. On the basis of the above SI-XPS results, it can be concluded that under light irradiation, the photo-excited electrons effectively transferred from CTFs to Co₂P through the P-bonding sites as well as the interfacial P–C bonds, leading to electron enrichment in Co₂P nanocrystals and hole enrichment in CTFs. To further confirm the crucial roles of interfacial P-bonding in CTFs and the anchoring of the Co₂P cocatalyst in promoting charge separation, SI-XPS studies for the CTF/Co₂P composite were also conducted. As shown in Fig. S22,^† no new peak or shape change of C 1s, N 1s, P 2p, and Co 2p peaks could be detected in CTF/Co₂P compared with pristine CTFs and Co₂P in the ground state, indicating no interfacial bonding in the CTF/Co₂P composite. Furthermore, in the excited state, no evident BE shifts and shape change could be observed, indicating the poor charge-separation capability of CTF/Co₂P, which is highly consistent with its fairly low hydrogen evolution activity. These SI-XPS results further confirm the crucial roles of interfacial P-bonding in CTFs and the anchoring of the Co₂P cocatalyst in efficiently promoting charge separation and enhancing the photocatalytic activity.Open in a separate windowFig. 4(A) Schematic illustration of the SI-XPS technique for direct observation of electron transfer in the excited state. The (B) C 1s, (C) N 1s, (D) P 2p and (E) Co 2p of SI-XPS spectra in the P-CTF-Co₂P sample tested under dark and light illumination.On the basis of the above results, it can be concluded that the P-bonding and Co₂P-cocatalyst anchoring should reverse the photo-induced electron transfer direction from the triazine to benzene rings in CTFs. To further confirm this speculation, the photocatalytic behavior of various CTF-based photocatalysts was studied and their electron transfer directions in excitation states have been proposed (Fig. 5A and B). First, when Co²⁺ ions were further bonded with the N sites of the triazine rings in the P-CTF-Co₂P sample (marked as P-CTF-Co₂P-Co), the hydrogen evolution activity remarkably increased from 7.6 up to 8.4 mmol h⁻¹ g⁻¹. This result clearly reveals that the Co–N bonding in P-CTF-Co₂P could further promote charge separation due to the effective hole trapping on Co sites. In contrast, when the Pt cocatalyst was decorated on the P-CTF-Co₂P sample (marked as P-CTF-Co₂P–Pt) to form Pt–N coordination, the photocatalytic activity significantly decreased to 4.6 mmol h⁻¹ g⁻¹, mainly resulting from the electron transfer competition between Pt and Co₂P. Furthermore, the decoration of the single Pt cocatalyst on CTFs promoted the photo-induced electron transfer from the benzene rings to triazine rings, and the photocatalytic activity achieved was up to 3.8 mmol h⁻¹ g⁻¹. Furthermore, in situ FTIR spectroscopy was performed for exploring the intermediate products in the photocatalytic process of the P-CTF-Co₂P sample with co-adsorption of H₂O vapor (Fig. S27^†), which also demonstrated the direction of electron transfer combined with the in situ XPS results. Accordingly, a possible mechanism has been proposed for clarifying the hydrogen evolution activities of the P-CTF-Co₂P photocatalyst (Fig. 5B). Owing to the P-bonding and the anchoring of Co₂P nanocrystals, the photo-generated electrons could effectively transfer from the triazine rings to benzene rings, where the electrons are trapped by the Co₂P cocatalyst through the interfacial P–C bonds for the hydrogen evolution reaction. Simultaneously, the holes left behind in the CTFs participated in oxidation reactions. More importantly, this mechanism may provide a new insight for understanding the crucial roles of interfacial P-bonding in CTFs and their bonding with Co₂P in promoting the charge separation for efficient hydrogen evolution.Open in a separate windowFig. 5(A) Photocatalytic hydrogen evolution activities of various samples and the scheme of electron-transfer direction in the excited state. (B) The ideal structural illustration of the interfacial bonding and charge transfer of the P-CTF-Co₂P photocatalyst for hydrogen evolution.     

HydroFlipper membrane tension probes: imaging membrane hydration and mechanical compression simultaneously in living cells

HydroFlippers are introduced as the first fluorescent membrane tension probes that report simultaneously on membrane compression and hydration. The probe design is centered around a sensing cycle that couples the mechanical planarization of twisted push–pull fluorophores with the dynamic covalent hydration of their exocyclic acceptor. In FLIM images of living cells, tension-induced deplanarization is reported as a decrease in fluorescence lifetime of the dehydrated mechanophore. Membrane hydration is reported as the ratio of the photon counts associated to the hydrated and dehydrated mechanophores in reconvoluted lifetime frequency histograms. Trends for tension-induced decompression and hydration of cellular membranes of interest (MOIs) covering plasma membrane, lysosomes, mitochondria, ER, and Golgi are found not to be the same. Tension-induced changes in mechanical compression are rather independent of the nature of the MOI, while the responsiveness to changes in hydration are highly dependent on the intrinsic order of the MOI. These results confirm the mechanical planarization of push–pull probes in the ground state as most robust mechanism to routinely image membrane tension in living cells, while the availability of simultaneous information on membrane hydration will open new perspectives in mechanobiology.

HydroFlippers respond to membrane compression and hydration in the same fluorescence lifetime imaging microscopy histogram: the responses do not correlate.

The detection and study of membrane mechanics in living cells is a topic of current concern.^1–14 To enable this research, appropriate chemistry tools, that is small-molecule fluorescent probes that allow imaging of membrane tension, are needed.¹⁵ With the direct imaging of physical forces being intrinsically impossible, design strategies toward such probes have to focus on the suprastructural changes caused by changes in membrane tension.¹⁵ These suprastructural changes are divers, often interconnected, and vary with the composition of the membrane.^15–25 Beyond the fundamental lipid compression and decompression, they include changes in membrane curvature, from rippling, buckling and budding to tubules extending from the membrane and excess lipid being ejected. Of similar importance are changes in membrane organization, particularly tension-induced phase separation and mixing, i.e. assembly and disassembly of microdomains. Consequences of these suprastructural changes include microdomain strengthening and softening and changes in membrane hydration and viscosity.^16–25The currently most developed fluorescent flipper probes have been introduced^26,27 to image membrane tension by responding to a combination of mechanical compression and microdomain assembly in equilibrium in the ground state.¹⁵ Extensive studies, including computational simulations,²⁸ have shown that flipper probes align non-invasively along the lipid tails of one leaflet and report changes in membrane order and tension as changes in fluorescent lifetimes and shifts of excitation maxima.¹⁵ Among other candidates, solvatochromic probes respond off-equilibrium in the excited state to changes in membrane hydration and have very recently been considered for the imaging of membrane tension in living cells.^29–36 So far not considered to image tension, ESIPT probes also report off equilibrium in the excited state on membrane hydration, but for different reasons.^37,38 Mechanosensitive molecular rotors respond off equilibrium in the excited state to changes in microviscosity.^{17,30,32,39–53} The same principle holds for the planarization of bent, papillon or flapping fluorophores.^54–57 The response of all possible probes to tension can further include less desired changes in positioning and partitioning between different domains, not to speak of more catastrophic probe aggregation, precipitation, disturbance of the surrounding membrane structure, and so on. Although the imaging of membrane tension is conceivable in principle with most of above approaches, the complex combination of parameters that has to be in place can thus far only be identified empirically, followed by much optimization.¹⁵The force-induced suprastructural changes are accompanied by the alteration in several unrelated physical properties of membranes. It is, for instance, well documented that membrane hydration increases with membrane disorder, from solid-ordered (S_o) to liquid-disordered (L_d) phases.^29,58 Increasing cholesterol content decreases membrane hydration in solid- and liquid-ordered membranes.⁵⁹ However, studies in model membranes also indicate that membrane hydration and membrane fluidity do not necessarily correlate.⁵⁹ The dissection of the individual parameters contributing to the response of fluorescent membrane tension probes would be important for probe design and understanding of their responses, but it remains a daunting challenge. In this study, we introduce fluorescent flipper probes that simultaneously report on mechanical membrane compression and membrane hydration at equilibrium in the ground state. Changes of both in response to changes in membrane tension and membrane composition are determined in various organelles in living cells.The dual hydration and membrane tension probes are referred to as HydroFlippers to highlight the newly added responsiveness to membrane hydration. The mechanosensing of lipid compression in bilayer membranes by flipper probes has been explored extensively.¹⁵ Fluorescent flippers²⁷ like 1 are designed as bioinspired⁶⁰ planarizable push–pull probes²⁶ (Fig. 1). They are constructed from two dithienothiophene fluorophores that are twisted out of co-planarity by repulsion of methyls and σ holes on sulfurs^61,62 next to the twistable bond. The push–pull system is constructed first from formal sulfide and sulfone redox bridges in the two twisted dithienothiophenes. These endocyclic donors and acceptors are supported by exocyclic ones, here a trifluoroketone acceptor and a triazole donor.⁶³ To assure stability, these endo- and exocyclic donors are turned off in the twisted ground state because of chalcogen bonding and repulsion, respectively.⁶²Open in a separate windowFig. 1The dual sensing cycle of HydroFlippers 1–5, made to target the indicated MOIs in living cells and responding to membrane compression by planarization and to membrane hydration by dynamic covalent ketone hydration. With indication of excitation maxima (ref. 63) and fluorescence lifetimes (this study).Mechanical planarization of the flipper probe establishes conjugation along the push–pull systems, electrons flow from endocyclic donors to acceptors, which turns on the exocyclic donors and acceptors to finalize the push–pull system.⁶² This elaborate, chalcogen-bonding cascade switch has been described elsewhere in detail, including high-level computational simulations.⁶² The planar high-energy conformer 1dp excels with red shifted excitation and increased quantum yield and lifetime compared to the twisted conformer 1dt because the less twisted Franck-Condon state favors emission through planar intramolecular charge transfer (PICT) over non-radiative decay through twisted ICT, or conical intersections.¹⁵Flipper probe 1 was considered for dual responsiveness to membrane tension and hydration because of the trifluoroketone acceptor.⁶³ Dynamic covalent hydration of 1dt yields hydrate 1ht.^64–76 Blue-shifted excitation and short lifetime of 1ht are not expected to improve much upon planarization because the hydrate is a poor acceptor and thus, the push–pull system in 1hp is weak. The dynamic covalent chemistry of the trifluoroketone acceptor has been characterized in detail in solution and in lipid bilayer membranes.⁶³To explore dual responsiveness to membrane tension in any membrane of interest (MOI) in living cells, HydroFlippers 2–5 were synthesized. While HydroFlipper 1 targets the plasma membrane (PM), HydroFlippers 2–4 were equipped with empirical targeting motifs.⁷⁷ HydroFlipper 5 terminates with a chloroalkane to react with the self-labeling HaloTag protein, which can be expressed in essentially any MOI.⁷⁸ Their substantial multistep synthesis was realized by adapting reported procedures (Schemes S1–S4^†).The MOIs labeling selectivity of HydroFlippers was determined in HeLa Kyoto (HK) cells by confocal laser scanning microscopy. Co-localization experiments of flippers 1–4 with the corresponding trackers gave Pearson correlation coefficients (PCCs) >0.80 for the targeting of mitochondria, lysosomes and the endoplasmic reticulum (ER, Fig. S4–S6^†). HydroFlipper 5 was first tested with stable HGM cells, which express both HaloTag and GFP on mitochondria (referred to as 5_M).^78,79 The well-established chloroalkane penetration assay demonstrated the efficient labeling of HaloTag protein by 5 as previously reported HaloFlippers (Fig. S3^†).⁷⁸ By transient transfection, HydroFlippers 5 were also directed to lysosomes (5_L), Golgi apparatus (GA, 5_G)⁸⁰ and peroxisomes (5_P) with HaloTag and GFP expressed on their surface.⁷⁸ PCCs >0.80 for co-localization of flipper and GFP emission confirmed that MOI labeling with genetically engineered cells was as efficient as with empirical trackers (Fig. S7–S11^†).Dual imaging of membrane compression and hydration was envisioned by analysis of fluorescence lifetime imaging microscopy (FLIM) images using a triexponential model (Fig. 2).⁸¹ FLIM images of ER HydroFlipper 4 in iso-osmotic HK cells were selected to illustrate the concept (Fig. 3a). Contrary to classical flipper probes, the fluorescence decay curve of the total FLIM image (Fig. 2a, grey) showed a poor fit to a biexponential model (Fig. 2a, cyan, b). Consistent with their expected dual sensing mode, a triexponential fit was excellent (Fig. 2a, dark blue, c). Lifetimes τ_1i = 4.3 ns (†) were obtained besides background. This three-component model was then applied to every pixel of FLIM images (Fig. 3c). The resulting reconvoluted FLIM histogram revealed three clearly separated populations for τ₁ (red), τ₂ (green), and background (τ₃, blue, Fig. 2d). Maxima of these three clear peaks were at the lifetimes estimated by triexponential fit of the global decay curve, thus demonstrating the validity of the methodology at necessarily small photon counts. Irreproducible fitting would give randomly scattered data without separated peaks.Open in a separate windowFig. 2(a) Fluorescence decay curve (grey, corresponding to the total image, not to a single pixel) with biexponential (cyan) and triexponential fit (dark blue). (b, c) Residual plots for bi- (b) and triexponential fit (c). (d) Histogram with the intensities associated with the τ₁ (red), τ₂ (green), and τ₃ (blue, background) components obtained by triexponential fit of the fluorescence decay curve of each pixel of the FLIM image, fit to Gaussian function (black solid curves).Open in a separate windowFig. 3FLIM images of HK cells labelled with ER flipper 4 before (a, c) and after (b, d) hyper-osmotic shock, showing average lifetimes τ_av (a, b) and τ₁ (c, d) from triexponential reconvolution; scale bars = 10 μm. (e) Distribution of the photon counts associated with the τ₁ component of 4 in HK cells after triexponential reconvolution of FLIM images before (c, τ_1i) and after (d, τ_1h) hyper-osmotic shock, showing decreasing lifetimes for τ₁ (4d). (f) The dehydration factor dh_i defined as total integrated photon counts for τ₁ (Στ₁) divided by Στ₂ (i.e., dh_i = area Στ_1i/area Στ_2i) for 4 in strongly hydrated ER (dh_i < 2, turquoise) and 1 in weakly hydrated plasma membrane (dh_i > 6, purple) of HK Kyoto cells under iso-osmotic conditions.Dual response of HydroFlippers to changes in membrane tension^a

Simplifying and expanding the scope of boron imidazolate framework (BIF) synthesis using mechanochemistry

Mechanochemistry enables rapid access to boron imidazolate frameworks (BIFs), including ultralight materials based on Li and Cu(i) nodes, as well as new, previously unexplored systems based on Ag(i) nodes. Compared to solution methods, mechanochemistry is faster, provides materials with improved porosity, and replaces harsh reactants (e.g. n-butylithium) with simpler and safer oxides, carbonates or hydroxides. Periodic density-functional theory (DFT) calculations on polymorphic pairs of BIFs based on Li⁺, Cu⁺ and Ag⁺ nodes reveals that heavy-atom nodes increase the stability of the open SOD-framework relative to the non-porous dia-polymorph.

Mechanochemistry enables rapid access to boron imidazolate frameworks (BIFs), including ultralight materials based on Li and Cu(i) nodes, as well as new, previously unexplored systems based on Ag(i) nodes.

Mechanochemistry^1–7 has emerged as a versatile methodology for the synthesis and discovery of advanced materials, including nanoparticle systems^8–10 and metal–organic frameworks (MOFs),^11–15 giving rise to materials that are challenging to obtain using conventional solution-based techniques.^16–18 Mechanochemical techniques such as ball milling, twin screw extrusion¹⁹ and acoustic mixing^20,21 have simplified and advanced the synthesis of a wide range of MOFs, permitting the use of simple starting materials such as metal oxides, hydroxides or carbonates,^22,23 at room temperature and without bulk solvents, yielding products of comparable stability and, after activation, higher surface areas than solution-generated counterparts.^24–29 The efficiency of mechanochemistry in MOF synthesis was recently highlighted by accessing zeolitic imidazolate frameworks (ZIFs)^30,31 that were theoretically predicted, but not accessible under conventional solution-based conditions.¹⁷The advantages of mechanochemistry in MOF chemistry led us to address the possibility of synthesizing boron imidazolate frameworks (BIFs),^32–34 an intriguing but poorly developed class of microporous materials analogous to ZIFs, comprising equimolar combinations of tetrahedrally coordinated boron(iii) and monovalent Li⁺ or Cu⁺ cations as nodes (Fig. 1A–C). Although BIFs offer an attractive opportunity to access microporous MOFs with lower molecular weights, particularly in the case of “ultralight” systems based on Li⁺ and B(iii) centers, this family of materials has remained largely unexplored – potentially due to the need for harsh synthetic conditions, including the use of n-butyllithium in a solvothermal environment.^32–34Open in a separate windowFig. 1Structures of previously reported BIFs with: (A) zni-, (B) dia-, or (C) SOD-topology (M = Li, Cu); (D) tetrakis(imidazolyl)boric acids used herein for mechanochemical BIF synthesis; and (E) schematic representation of the herein developed mechanosynthesis of dia- and SOD BIF polymorphs based on Li, Cu or Ag metal nodes.We now show how switching to the mechanochemical environment enables lithium- and copper(i)-based BIFs to be prepared rapidly (i.e., within 60–90 minutes), without elevated temperatures or bulk solvents, and from readily accessible solid reactants, such as hydroxides and oxides (Fig. 1D and E). While the mechanochemically-prepared BIFs exhibit significantly higher surface areas than the solvothermally-prepared counterparts, mechanochemistry allows for expanding this class of materials towards previously not reported Ag⁺ nodes. The introduction of BIFs isostructural with those based on Li⁺ or Cu⁺ but comprising of Ag⁺ ions, enables a periodic density-functional theory (DFT) evaluation of their stability. This reveals that switching to heavier elements as tetrahedral nodes improves the stability of sodalite topology (SOD) open BIFs with respect to close-packed diamondoid (dia) topology polymorphs.As a first attempt at mechanochemically synthesis of BIFs, we targeted the synthesis of previously reported zni-topology LiB(Im)₄ and CuB(Im)₄ frameworks (Li-BIF-1 and Cu-BIF-1, respectively, Fig. 1A) using a salt exchange reaction between LiCl or CuCl with commercially available sodium tetrakis(imidazolyl)borate (Na[B(Im)₄]) (Fig. 2A). Milling of LiCl and Na[B(Im)₄] in a 1 : 1 stoichiometric ratio for up to 60 minutes led to the appearance of Bragg reflections consistent with the target Li-BIF-1 (CSD MOXJEP) and the anticipated NaCl byproduct. The reaction was, however, incomplete, as seen by X-ray reflections of Na[B(Im)₄] starting material. In order to improve reactant conversion, we explored liquid-assisted grinding (LAG), i.e. milling in the presence of a small amount of a liquid phase (measured by the liquid-to-solid ratio η³⁵ in the range of ca. 0–2 μL mg⁻¹). Using LAG conditions with acetonitrile (MeCN, 120 μL, η = 0.5 μL mg⁻¹) led to the complete disappearance of reactant X-ray reflections, concomitant with the formation of Li-BIF-1 alongside NaCl within 60 minutes.Open in a separate windowFig. 2(A) Reaction scheme for the mechanochemical synthesis of Li-BIF-1 by a salt metathesis strategy. Selected PXRD patterns for: (B) Na[B(Im)₄] (C) LiCl, (D) simulated Li-BIF-1 (CSD MOXJPEP) and (E) synthesized BIF-1-Li by LAG for 60 minutes with MeCN (η = 0.5 μL mg⁻¹), (F) CuCl, (G) simulated Cu-BIF-1 (CSD MOXJIT), and (H) synthesized BIF-1-Cu by LAG for 60 minutes with MeOH (η = 0.50 μL mg⁻¹). Asterisks denote NaCl, a byproduct of the metathesis reaction. (Fig. 2B–E, also see ESI^†). The copper-based zni-CuB(Im)₄ (Cu-BIF-1) was readily obtained from CuCl within 60 minutes using similar LAG conditions. We also explored LAG with methanol (MeOH), revealing that the exchange reaction to form NaCl took place with both LiCl and CuCl starting materials. With LiCl, however, the PXRD pattern of the product could not be matched to known phases involving Li⁺ and B(Im)₄⁻ (see ESI^†). With CuCl as a reactant, LAG with MeOH (η = 0.5 μL mg⁻¹) cleanly produced Cu-BIF-1 alongside NaCl (see ESI^†).Next, we explored an alternative synthesis approach, analogous to that previously used to form ZIFs and other MOFs: an acid–base reaction between a metal oxide or hydroxide and the acid form of the linker: tetrakis(imidazolato)boric acid, HB(Im)₄ (Fig. 3A).^36–40 Neat milling LiOH with one equivalent of HB(Im)₄ in a stainless steel milling assembly led to the partial formation of Li-BIF-1, as evidenced by PXRD analysis (see ESI^†). Complete conversion of reactants into Li-BIF-1 was achieved in 60 minutes by LAG with MeCN (η = 0.25 μL mg⁻¹), as indicated by PXRD analysis (Fig. 3B–E), Fourier transform infrared attenuated total reflectance spectroscopy (FTIR-ATR), thermogravimetric analysis (TGA) in air, and analysis of metal content by inductively-coupled plasma mass spectrometry (ICP-MS) (see ESI^†).Open in a separate windowFig. 3(A) Reaction scheme for the mechanochemical synthesis of Li-BIF-1 using the acid–base strategy. Selected PXRD patterns for: (B) H[B(Im)₄] (C) LiOH, (D) simulated Li-BIF-1 (CSD MOXJPEP), (E) synthesized BIF-1-Li by LAG for 60 minutes with MeCN (η = 0.25 μL mg⁻¹), (F) Cu₂O, (G) simulated Cu-BIF-1 (CSD MOXJIT), and (H) synthesized Cu-BIF-1 by ILAG for 60 minutes with MeOH (η = 0.50 μL mg⁻¹) and NH₄NO₃ additive (5% by weight).Neat milling of HB(Im)₄ with Cu₂O under similar conditions gave a largely non-crystalline material, as evidenced by PXRD (see ESI^†). Switching to the ion- and liquid-assisted grinding (ILAG) methodology, in which the reactivity of a metal oxide is enhanced by a small amount of a weakly acidic ammonium salt, and which was introduced to prepare zinc and cadmium ZIFs from respective oxides,^37–40 enabled the synthesis of Cu-BIF-1 from Cu₂O. Specifically, PXRD analysis revealed complete disappearance of the oxide in samples obtained by ILAG with either MeOH or MeCN (η = 0.5 μL mg⁻¹) in the presence of NH₄NO₃ additive (5% by weight, see ESI^†). Notably, achieving complete disappearance of Cu₂O reactant signals also required switching from stainless steel to a zirconia-based milling assembly, presumably due to more efficient energy delivery.⁴¹ After washing with MeOH, the material was characterized by FTIR-ATR, TGA in air, and analysis of metal content by ICP-MS (see ESI^†).Whereas both the metathesis and acid–base approaches can be used to mechanochemically generate Li- and Cu-BIF-1, the latter approach has a clear advantage of circumventing the formation of the NaCl byproduct. Consequently, in order to further the development of mechanochemical routes to other BIFs, we focused on the acid–base strategy. As next targets, we turned to MOFs based on tetrakis(2-methylimidazole)boric acid H[B(Meim)₄],³⁶ previously reported³² to adopt either a non-porous diamondoid (dia) topology (BIF-2) or a microporous sodalite (SOD) topology (BIF-3) with either Li⁺ or Cu⁺ as nodes (Fig. 4). Attempts to selectively synthesize either Li-BIF-2 or Li-BIF-3 by neat milling or LAG (using MeOH or MeCN as liquid additives) with LiOH and a stoichiometric amount of HB(Meim)₄ were not successful. Exploration of different milling times and η-values produced only mixtures of residual reactants with Li-BIF-2, Li-BIF-3, and/or not yet identified phases (see ESI^†). Consequently, we explored milling in the presence of 2-aminobutanol (amb), which is a ubiquitous component of solvent systems used in the solvothermal syntheses of BIFs.^32,33 Gratifyingly, using a mixture of amb and MeCN in a 1 : 3 ratio by volume as the milling liquid led to an effective strategy for the selective synthesis of both the dia-topology Li-BIF-2 (CSD code MOXKUG), and the SOD-topology Li-BIF-3 (CSD code MUCLOM).^‡ The selective formation of phase-pure samples of Li-BIF-2 and Li-BIF-3 was confirmed by PXRD analysis, which revealed an excellent match to diffractograms simulated based on the previously reported structures (Fig. 4B–G). Systematic exploration of reaction conditions, including time (between 15 and 60 minutes) and η value (between 0.25 and 1 μL mg⁻¹) revealed that the open framework Li-BIF-3 is readily obtained at η either 0.75 or 1 μL mg⁻¹ after milling for 45 minutes or longer (Fig. 4B–G, also see ESI^†).^§ Lower η-values of 0.25 and 0.5 μL mg⁻¹ preferred the formation of the dia-topology Li-BIF-2, which was obtained as a phase-pure material upon 60 minutes milling at η = 0.5 μL mg⁻¹, following the initial appearance of a yet unidentified intermediate. The preferred formation of Li-BIF-2 at lower η-values is consistent with our previous observations that lower amounts of liquid promote mechanochemical formation of denser MOF polymorphs.³⁷Open in a separate windowFig. 4(A) Reaction scheme for the mechanochemical synthesis of Li-BIF-3. Comparison of selected PXRD patterns for the synthesis of Li-BIF-2 and Li-BIF-3: (B) H[B(Meim)₄] reactant; (C) LiOH reactant; (D) simulated for Li-BIF-3 (CSD MUCLOM); (E) simulated for Li-BIF-2 (CSD MOXKUG); (F) Li-BIF-3 mechanochemically synthesized by LAG for 60 minutes with a 1 : 3 by volume mixture of amb and MeCN (η = 1 μL mg⁻¹); and (G) Li-BIF-2 mechanochemically synthesized by LAG for 60 minutes with a 1 : 3 by volume mixture of amb and MeCN (η = 0.5 μL mg⁻¹). Comparison of selected PXRD patterns for the synthesis of Cu-BIF-2 and Li-BIF-3: (H) Cu₂O; (I) Cu-BIF-3 (CSD MOXJOZ); (J) Cu-BIF-2 (CSD MUCLIG); (K) Cu-BIF-3 mechanochemically synthesised by ILAG for 60 minutes using NH₄NO₃ ionic additive (5% by weight) and MeOH (η = 1 μL mg⁻¹); and (L) mechanochemically synthesised Cu-BIF-2 by ILAG for 90 minutes using NH₄NO₃ ionic additive (5% by weight) and MeOH (η = 0.5 μL mg⁻¹).Samples of both Li-BIF-2 and Li-BIF-3 after washing with MeCN were further characterized by FTIR-ATR, TGA in air, and analysis of metal content by ICP-MS (see ESI^†). Nitrogen sorption measurement on the mechanochemically obtained Li-BIF-3, after washing with MeCN and evacuation at 85 °C, revealed a highly microporous material with a Brunauer–Emmett–Teller (BET) surface area of 1010 m² g⁻¹ (Fig. 5A), which is close to the value expected from the crystal structure of the material (1200 m² g⁻¹, 32 For direct comparison with previous work,³² we also calculated the Langmuir surface area, revealing an almost 40% increase (1060 m² g⁻¹) compared to samples made solvothermally (762.5 m² g⁻¹) (Fig. 5A, inset).Experimental Brunauer–Emmett–Teller (BET) and Langmuir surface area (in m² g⁻¹) of mechanochemically synthesized SOD-topology BIFs, compared to previously measured and theoretically calculated values, along with average particle sizes (in nm) established by SEM and calculated energies (in eV) for all Li-, Cu-, and Ag-BIF polymorphs. The difference between calculated energies for SOD- and dia-polymorphs in each system is given as ΔE (in kJ mol⁻¹)

Separation of pyrrolidine from tetrahydrofuran by using pillar[6]arene-based nonporous adaptive crystals

Pyrrolidine, an important feedstock in the chemical industry, is commonly produced via vapor-phase catalytic ammoniation of tetrahydrofuran (THF). Obtaining pyrrolidine with high purity and low energy cost has extremely high economic and environmental values. Here we offer a rapid and energy-saving method for adsorptive separation of pyrrolidine and THF by using nonporous adaptive crystals of per-ethyl pillar[6]arene (EtP6). EtP6 crystals show a superior preference towards pyrrolidine in 50 : 50 (v/v) pyrrolidine/THF mixture vapor, resulting in rapid separation. The purity of pyrrolidine reaches 95% in 15 min of separation, and after 2 h, the purity is found to be 99.9%. Single-crystal structures demonstrate that the selectivity is based on the stability difference of host–guest structures after uptake of THF or pyrrolidine and non-covalent interactions in the crystals. Besides, EtP6 crystals can be recycled efficiently after the separation process owing to reversible transformations between the guest-free and guest-loaded EtP6.

Here we offer a rapid and energy-saving method for adsorptive separation of pyrrolidine and tetrahydrofuran by using nonporous adaptive crystals of per-ethyl pillar[6]arene.

Pyrrolidine is an important feedstock in the chemical industry that has been widely used in the production of food, pesticides, daily chemicals, coatings, textiles, and other materials.¹ Particularly, pyrrolidine is a raw material for organic synthesis of medicines such as buflomedil, pyrrocaine, and prolintane.² Moreover, pyrrolidine is also used as a solvent in the semi-synthetic process of simvastatin, one of the best-selling cardiovascular drugs.³ In the chemical industry, there are many preparation methods for pyrrolidine. The most common way to obtain pyrrolidine is the gas-phase catalytic method using tetrahydrofuran (THF) and ammonia as raw materials;⁴ this is carried out at high temperature under catalysis by solid acids. However, separating pyrrolidine from the crude product is difficult because of similar molecular weights and structures between pyrrolidine (b.p. 360 K and saturated vapor pressure = 1.8 kPa at 298 K) and THF (b.p. 339 K and saturated vapor pressure = 19.3 kPa at 298 K), which result in complicated processes and large energy consumption.⁵ Therefore, it is worthwhile to find energy-efficient and simple methods to separate pyrrolidine from THF.Many techniques and materials, including porous zeolites, metal–organic frameworks (MOFs), and porous polymers, have facilitated energy-efficient separations of important petrochemicals and feedstocks, including THF and pyrrolidine.^6,7 However, some drawbacks of these materials cannot be ignored.⁸ For example, the relatively low thermal and moisture stabilities of MOFs limit their practical applications. Therefore, the development of new materials with satisfactory chemical and thermal stabilities for pyrrolidine/THF separation is of high significance.In the past decade, pillararenes have been widely studied in supramolecular chemistry.⁹ Owing to their unique pillar structures and diverse host–guest recognitions, pillararenes have been used in the construction of numerous supramolecular systems.¹⁰ Recently, nonporous adaptive crystals (NACs) of macrocycles, which have shown extraordinary performance in adsorption and separation, have been developed by our group as a new type of adsorption and separation materials.¹¹ Unlike MOFs, covalent-organic frameworks (COFs), and other materials with pre-existing pores, NACs do not have “pores“ in the guest-free form, whereas they adsorb guest vapors through cavities of macrocycles and spaces between macrocycles. NACs have been applied in separations of many significant chemicals such as alkane isomers, aromatics, and halohydrocarbon isomers.¹² However, such materials have never been used to separate pyrrolidine and THF. Herein, we utilized pillararene crystals as a separation material and realized the selective separation of pyrrolidine from a mixture of pyrrolidine and THF. We found that nonporous crystals of per-ethyl pillar[6]arene (EtP6) exhibited a shape-sorting ability at the molecular level towards pyrrolidine with an excellent preference, while crystals of per-ethyl pillar[5]arene (EtP5) did not (Scheme 1). In-depth investigations revealed that the separation was driven by the host–guest complexation between pyrrolidine and EtP6, which resulted in the formation of a more stable structure upon adsorption of pyrrolidine vapor in the crystalline state. EtP6 crystals can also adsorb THF. However, when these two chemicals simultaneously exist as the vapor of a 50 : 50 (v/v) mixture, EtP6 prefers pyrrolidine as an adsorption target. Compared with previously reported NAC-based separation, this separation took place rapidly. 95% purity was achieved in 15 min, and the purity increased to 99.9% after 2 h of separation. Moreover, pyrrolidine was removed upon heating, along with the structural transformation of EtP6 back to its original state, endowing EtP6 with excellent recyclability.Open in a separate windowScheme 1Chemical structures and cartoon representations: (a) EtP5 and EtP6; (b) THF and pyrrolidine.EtP5 and EtP6 were prepared as previously described and then a pretreatment process was carried out to obtain guest-free EtP5 and EtP6 (Fig. S1–S4†).¹³ According to powder X-ray diffraction (PXRD) patterns, activated EtP5 and EtP6 (denoted as EtP5α and EtP6β, respectively) were crystalline, and the patterns matched previous reports (Fig. S5 and S6^†).¹⁴ Studies from our group indicated that EtP5α and EtP6β crystals were nonporous, presumably due to their dense packing modes.We first investigated the adsorption capabilities of EtP5α and EtP6β towards pyrrolidine and THF vapors. Based on time-dependent solid–vapor adsorption procedures, both EtP5α and EtP6β showed good ability to adsorb pyrrolidine and THF vapors. As shown in Fig. 1a, the adsorption amount of THF in EtP5α was higher than that of pyrrolidine. It took 6 hours for EtP5α to reach saturation points for adsorption of both pyrrolidine and THF vapors. The final storage of THF in EtP5α was 2 : 1 (molar ratio to the host), whereas the storage of pyrrolidine was 1 : 1. It seemed that the THF vapor was favored to occupy EtP5α, which was ascribed to the relatively lower boiling point of THF. A similar phenomenon was found for EtP6β. Time-dependent solid–vapor adsorption experiments for pyrrolidine demonstrated that it took just 1 hour to reach the saturation point, while it took 4 hours for the THF vapor (Fig. 1b). The adsorption amount of THF vapor was twice that of pyrrolidine. ¹H NMR spectra and thermogravimetric analyses (TGA) further confirmed the adsorption and storage of THF and pyrrolidine in both hosts (Fig. S7–S16†). Meanwhile, in the desorption process, adsorbed pyrrolidine and THF in EtP6β were easily released under reduced pressure and heating. Based on these data, it was clear that pyrrolidine could be adsorbed rapidly by both EtP5α and EtP6β in molar ratios = 1 : 1, while THF could be captured in a relatively slow process. Structural changes after adsorption of these two vapors were analyzed via PXRD experiments, in which varying degrees of changes before and after adsorption were observed, evidencing the appearance of new crystal structures (Fig. 1c and d). Nevertheless, only slight differences were observed in the PXRD patterns after the adsorption of THF or pyrrolidine, which might be ascribed to the structural similarity of the two molecules.Open in a separate windowFig. 1Time-dependent solid–vapor adsorption plots of (a) EtP5α and (b) EtP6β for single-component pyrrolidine and THF vapors. PXRD patterns of (c) EtP5α and (d) EtP6β: (I) original activated crystals; (II) after adsorption of THF vapor; (III) after adsorption of pyrrolidine vapor.To study the mechanism of adsorption, guest-loaded single crystals were obtained by slowly evaporating either THF or pyrrolidine solutions of pillararenes (Tables S2 and S3^†). In the crystal structure of THF-loaded EtP5 (2THF@EtP5, Fig. 2a and S17^†),^11a two THF molecules are in the cavity of one EtP5 molecule driven by multiple C–H⋯O hydrogen bonds and C–H⋯π bonds. EtP5 assembles into honeycomb-like infinite edge-to-edge 1D channels. In the crystal structure of pyrrolidine-loaded EtP5 (pyrrolidine@EtP5, Fig. 2b and S19^†), one pyrrolidine molecule, stabilized by C–H⋯π interactions and C–H⋯O hydrogen bonds between hydrogen atoms on pyrrolidine and oxygen atoms on EtP5, is found in the cavity of EtP5. It''s worth mentioning that a hydrogen atom which is linked with the N atom of pyrrolidine also forms a strong hydrogen bond with an oxygen atom on the ethoxy group of EtP5. EtP5 forms imperfect 1D channels because of partial distortion of orientation. The PXRD patterns simulated from these crystal structures matched well with the experimental results (Fig. S18 and S20^†), which verified that the uptake of vapors transformed EtP5α into pyrrolidine-loaded EtP5.Open in a separate windowFig. 2Single crystal structures: (a) 2THF@EtP5; (b) pyrrolidine@EtP5.In the crystal structure of THF-loaded EtP6 (2THF@EtP6, Fig. 3a and S21^†), one EtP6 molecule encapsulated two THF molecules in its cavity with C–H⋯O interactions, forming a 1 : 2 host–guest complex. Although 1D channels are observed, EtP6 adopts a slightly different conformation, caused by the presence of THF. Moreover, the PXRD pattern of EtP6β after adsorption of THF vapor matches well with that simulated from 2THF@EtP6, which is evidence for the structural transformation upon adsorption. In the crystal structure of pyrrolidine-loaded EtP6 (pyrrolidine@EtP6, Fig. 3b and S23^†), a 1 : 1 host–guest complex with pyrrolidine is found. Driven by C–H⋯π interactions and C–H⋯O hydrogen bonds formed by hydrogen atoms on pyrrolidine and oxygen atoms on EtP6, one pyrrolidine molecule is in the cavity of EtP6 with the nitrogen atom inside the cavity. The window-to-window packing mode of hexagonal EtP6 molecules in pyrrolidine@EtP6 contributes to the formation of honeycomb-like infinite edge-to-edge 1D channels, favorable for guest adsorption. Likewise, the PXRD result of EtP6β after adsorption of pyrrolidine is in line with the simulated pattern of pyrrolidine@EtP6, indicating that EtP6β transformed into pyrrolidine@EtP6 in the presence of pyrrolidine (Fig. S22 and S24^†).Open in a separate windowFig. 3Single crystal structures: (a) 2THF@EtP6; (b) pyrrolidine@EtP6.According to the adsorption ability and different crystal structures after adsorption of guest vapors, we wondered whether EtP5α or EtP6β could separate mixtures of THF and pyrrolidine. We first evaluated separation by EtP5α. GC analysis indicated that the adsorption ratios of THF and pyrrolidine were 65.7% and 34.3%, respectively, when EtP5α was exposed to 50 : 50 (v/v) pyrrolidine/THF mixture vapor (Fig. 4a and S25^†). Such adsorption was also illustrated by ¹H NMR (Fig. S26^†). Although EtP5α showed a preference for THF, the selectivity is not satisfactory and cannot be applied to industrial separation. The less satisfactory selectivity may be ascribed to the similar crystal structures of EtP5 after adsorption of THF or pyrrolidine and insufficient strong stabilizing interactions. The PXRD pattern of EtP5α after adsorption of the 50 : 50 (v/v) pyrrolidine/THF mixture vapor exhibited minor differences compared with that simulated from either 2THF@EtP5 or pyrrolidine@EtP5, due to poor selectivity (Fig. 4b).Open in a separate windowFig. 4(a)Time-dependent solid–vapor adsorption plot for EtP5α in the presence of 50 : 50 (v/v) pyrrolidine/THF mixture vapor. (b) PXRD patterns of EtP5α: (I) original EtP5α; (II) after adsorption of THF vapor; (III) after adsorption of pyrrolidine vapor; (IV) after adsorption of pyrrolidine/THF mixture vapor; (V) simulated from the single crystal structure of pyrrolidine@EtP5α; (VI) simulated from the single crystal structure of 2THF@EtP5α. (c) Time-dependent solid–vapor adsorption plot for EtP6β in the presence of 50 : 50 (v/v) pyrrolidine/THF mixture vapor. (d) PXRD patterns of EtP6β: (I) original EtP6β; (II) after adsorption of THF vapor; (III) after adsorption of pyrrolidine vapor; (IV) after adsorption of pyrrolidine/THF mixture vapor; (V) simulated from the single crystal structure of pyrrolidine@EtP6β; (VI) simulated from the single crystal structure of 2THF@EtP6β.Nevertheless, selective separation of THF and pyrrolidine was achieved with EtP6β. As shown in Fig. 4c, time-dependent solid–vapor adsorption experiments for a 50 : 50 (v/v) pyrrolidine/THF mixture were conducted. Unlike the phenomenon in single-component adsorption experiments, uptake of pyrrolidine by EtP6β increased and reached the saturation point rapidly (less than 2 hours), while capture of THF was negligible. According to the NMR and GC results (Fig. S27 and S28^†), the purity of pyrrolidine was determined to be 99.9% after 2 hours of adsorption, which indicates the remarkable selectivity of EtP6β for pyrrolidine. The PXRD pattern of EtP6β after adsorption of the mixture was consistent with that from single-component adsorption, indicating the structural transformation in the crystalline state upon selective capture of pyrrolidine from the mixture. Although THF and pyrrolidine have similar molecular structures, their non-covalent interactions with EtP6 are different. We assume that the hydrogen bond between N–H and the oxygen atom on EtP6 stabilizes pyrrolidine and leads to such selectivity. More importantly, compared with previous adsorption processes using NACs reported by our group, the selective separation of pyrrolidine was completed rapidly. According to the GC results, the purity of pyrrolidine reached around 95% in the initial 15 min, while it usually takes hours for selective separations of other substrates using NACs. Increasing the adsorption time to 2 h improves the purity to over 99%. The rapid separation of pyrrolidine with high purity using EtP6β shows great potential in industrial applications.Apart from selectivity, recyclability is also an important parameter for an adsorbent. Consequently, recycling experiments were carried out by heating pyrrolidine@EtP6 under vacuum at 100 °C to remove adsorbed pyrrolidine. According to TGA and PXRD analysis, the recycled EtP6 solid maintained crystallinity and structural integrity that were the same as those of activated EtP6 crystals (Fig. S29 and S30^†). Besides, it is worth mentioning that the recycled EtP6 solids were still capable of separating mixtures of pyrrolidine and THF without loss of performance after being recycled five times (Fig. S31^†).In conclusion, we explored the separation of pyrrolidine/THF mixtures using NACs of EtP5 and EtP6. Pyrrolidine was purified using EtP6 from a 50 : 50 (v/v) pyrrolidine/THF mixture with a purity of 99.9%, but EtP5 exhibited selectivity towards THF. Moreover, the separation of pyrrolidine by EtP6 was extremely fast so that over 95% purity was determined within 15 min of adsorption. The rapid separation is unique among NAC-based separations. Single-crystal structures revealed that the selectivity depended on the stability of the new structures after adsorption of the guests and the non-covalent interactions in the host–guest complexes. PXRD patterns indicated that the structures of the host crystals changed into the host–guest complexes after adsorption. Additionally, the NACs of EtP6 exhibited excellent recyclability over at least five runs; this endows EtP6 with great potential as an alternative adsorbent for rapid purification of pyrrolidine that can be applied in practical industry. The fast separation with such simple NACs in this work also reveals that minor structural differences can cause significant changes in properties, which should provide perspectives on designs of adsorbents or substrates with specifically tailored binding sites.     

Electrooxidative o-carborane chalcogenations without directing groups: cage activation by copper catalysis at room temperature

Copper-catalyzed electrochemical direct chalcogenations of o-carboranes was established at room temperature. Thereby, a series of cage C-sulfenylated and C-selenylated o-carboranes anchored with valuable functional groups was accessed with high levels of position- and chemo-selectivity control. The cupraelectrocatalysis provided efficient means to activate otherwise inert cage C–H bonds for the late-stage diversification of o-carboranes.

Copper-catalyzed electrochemical cage C–H chalcogenation of o-carboranes has been realized to enable the synthesis of various cage C-sulfenylated and C-selenylated o-carboranes.

Carboranes are polyhedral molecular boron–carbon clusters, which display unique properties, such as a boron enriched content, icosahedron geometry and three-dimensional electronic delocalization.¹ These features render carboranes as valuable building blocks for applications to optoelectronics,² as nanomaterials, in supramolecular design,³ organometallic coordination chemistry,⁴ and boron neutron capture therapy (BNCT) agents.⁵ As a consequence, considerable progress has been witnessed in transition metal-catalyzed regioselective cage B–H functionalization of o-carboranes⁶ and different functional motifs have been incorporated into the cage boron vertices.^7–10 However, progress in this research arena continues to be considerably limited by the shortage of robust and efficient methods to access carborane-functionalized molecules. While C–S bonds are important structural motifs in various biologically active molecules and functional materials,¹¹ strategies for the assembly of chalcogen-substituted carboranes continue to be scarce. A major challenge is hence represented by the strong coordination abilities of thiols to most transition metals, which often lead to catalyst deactivation.¹² While copper-catalyzed B(4,5)–H disulfenylation of o-carboranes was achieved,^7e elevated reaction temperature was required, and 8-aminoquinoline was necessary as bidentate directing group. The bidentate directing group¹³ needs to be installed and removed, which jeopardizes the overall efficacy. Likewise, an organometallic strategy was recently devised for cysteine borylation with a stoichiometric platinum(ii)-based carboranes.¹⁴ Meanwhile, oxidative cage B/C–H functionalizations largely call for noble transition metal catalysts¹⁵ and stoichiometric amounts of chemical oxidants, such as expensive silver(i) salts.¹⁶In recent years, electricity has been identified as an increasingly viable, sustainable redox equivalent for environmentally-benign molecular synthesis.^17,18 While significant advances have been realized by the merger of electrocatalysis with organometallic bond activation,¹⁹ electrochemical carborane functionalizations continue unfortunately to be underdevelopment. In sharp contrast, we have now devised a strategy for unprecedented copper-catalyzed electrochemical cage C–H chalcogenations of o-carboranes in a dehydrogenative manner, assembling a variety of C-sulfenylated and C-selenylated o-carboranes (Fig. 1a). It is noteworthy that our electrochemical cage C–S/Se modification approach is devoid of chemical oxidants, and does not need any directing groups, operative at room temperature.Open in a separate windowFig. 1Electrochemical diversification of o-carboranes and optimization of reaction conditions. ^aReaction conditions: procedure A: 1a (0.10 mmol), 2a (0.3 mmol), CuOAc (15 mol%), 2-PhPy (15 mol%), LiOtBu (0.2 mmol), TBAI (2.0 equiv.), solvent (3 mL), platinum cathode (10 mm × 15 mm × 0.25 mm), graphite felt (GF) anode (10 mm × 15 mm × 6 mm), 2 mA, under air, r.t., 16 h. ^bYield was determined by ¹H NMR with CH₂Br₂ as the internal standard. ^cIsolated yields in parenthesis. ^dKI (1.0 equiv.) as additive. ^eProcedure B: 2 (0.3 mmol), LiOtBu (0.2 mmol), TBAI (2.0 equiv.), solvent (3.0 mL), 2 mA, r.t., 3 h, then adding 1a (0.10 mmol), 2-PhPy (15 mol%), CuOAc (15 mol%), 2 mA, rt, 16 h. ^f2b (0.3 mmol), LiOtBu (0.2 mmol), KI (1.0 equiv.), TBAI (2.0 equiv.), solvent (3.0 mL), 2 mA, r.t., 3 h, then adding 1a (0.10 mmol), 2-PhPy (15 mol%), CuOAc (15 mol%), r.t., 16 h. TBAI = tetrabutylammonium iodide, TBAPF₆ = tetrabutylammonium hexafluorophosphate. DCE = 1,2-dichloroethane, THF = tetrahydrofuran.We commenced our studies by probing various reaction conditions for the envisioned copper-catalyzed cage C–H thiolation of o-carborane in an operationally simple undivided cell setup equipped with a GF (graphite felt) anode and a Pt cathode (Fig. 1b and Table S1^†). After extensive experimentation, we observed that the thiolation of substrate 1 proceeded efficiently with catalytic amounts of CuOAc and 2-phenylpyridine, albeit in the presence of 2 equivalents LiOtBu as the base, and 2 equivalents n-Bu₄NI as the electrolyte at room temperature under a constant current of 2 mA (entry 1). The yield was reduced when other copper sources or additives were used (entries 2–5). Surprisingly, n-Bu₄NPF₆ as the electrolyte failed to facilitate the carborane modification, indicating that n-Bu₄NI operates not only as electrolyte, but also as a redox mediator (entry 6). Altering the stoichiometry of the electrolyte or using KI did not improve the performance (entries 7–8). Product formation was not observed, when the reaction was conducted with DCE as the solvent, while CH₃CN resulted in a drop of the catalytic performance (entries 9–10). Control experiments confirmed the essential role of the electricity and the catalyst (entries 11–12), while a sequential procedure was found to be beneficial (entries 13–15).With the optimized reaction conditions in hand, we explored the versatility of the cage C–H thiolation of o-carborane 1a with different thiols 2 (Scheme 1). Electron-rich as well as electron-deficient substituents on the arenes were found to be amenable to the electrocatalyzed C–H activation, providing the corresponding thiolation products 3aa–3ao in good to excellent yields. Thereby, a variety of synthetically useful functional groups, such as fluoro (3ae, 3am), chloro (3af, 3ak, 3an) and bromo (3ag, 3al), were fully tolerated, which should prove instrumental for further late-stage manipulations. Various disubstituted aromatic and heterocyclic thiols afforded the corresponding cage C–S modified products 3ap–3as. Notably, aliphatic thiols efficiently underwent the electrochemical transformation to provide the corresponding cage alkylthiolated products 3at–3au. Notably, the halogen-containing thiols (2e–2f, 2k–2n and 2q) reacted selectively with o-carboranes to deliver the desired products without halide coupling byproducts being observed. The connectivity of the products 3aa, 3am and 3ao was unambiguously verified by X-ray single crystal diffraction analysis.²²Open in a separate windowScheme 1Electrochemical C–H thiolation of o-carborane 1a. (a) Procedure B. (b) KI (1 equiv.). (c) Cul as the catalyst.Encouraged by the efficiency of the cupraelectro-oxidative cage C–H thiolation, we became intrigued to explore the chalcogenantion of differently-decorated o-carboranes 1 (Scheme 2). Electronically diverse carboranes 1 served as competent coupling partners, giving the corresponding thiolation products 4bo–4do with high levels of efficacy in position-selective manner. The strategy was not restricted to phenyl-substituted o-carboranes. Indeed, substrates bearing benzyl and even alkyl groups also performed well to deliver the desired products 4eo–4ga. It is noteworthy that the C–H activation approach was also compatible with selenols to give the o-carboranes 4av–4fv. The molecular structures of the carborane 4br and 4av were unambiguously verified by single-crystal X-ray diffraction.²²Open in a separate windowScheme 2Electrochemical cage C–H chalcogenation of o-carboranes. (a) Procedure B. (b) KI (1 equiv.).Scaffold functionalization of the thus obtained carborane 3ag provided the alkynylated derivative 5a and amine 5b (Scheme 3), giving access to carborane-based host materials of relevant to phosphorescent organic light-emitting diodes.²⁰Open in a separate windowScheme 3Late-stage diversification.Next, we became attracted to delineating the mode of the cupraelectro-catalyzed cage C–H chalcogenation. To this end, control experiments were performed (Scheme 4a). First, electrocatalysis in the presence of TEMPO or Ph₂C CH₂ gave the desired product 3aa. EPR studies of thiol 2a, LiOtBu and THF under the electrochemical conditions showed a small radical signal, which might be attributed to a thiol radical.²¹ Second, the cupraelectrocatalysis occurred efficiently in the dark. Third, detailed cyclovoltammetric analysis of the thiol and iodide mediator (Scheme 4b and ESI^†)²¹ revealed an irreversible oxidation of the thiol anion at E_p = −0.62 V vs. Ag/Ag⁺ and two oxidation events for the iodide, including an irreversible oxidation at E_p = 0.12 V vs. Ag/Ag⁺ and a reversible oxidation at E_p = 0.44 V vs. Ag/Ag⁺, which is in good agreement with the literature reported iodide oxidation potentials,^18c,d and is suggestive of the preferential oxidation of the iodide as a redox mediator. In this context, the use of n-Bu₄NI as a redox mediator to achieve copper-catalyzed electrochemical arene C–H aminations had been documented.^18d Furthermore, we calculated the redox potential of complex C by means of DFT calculations at the PW6B95-D4/def2-TZVP + SMD(MeCN)//TPSS-D3BJ/def2-SVP level of theory.²¹ These studies revealed a calculated oxidation half-wave potential for complex C is E^o,calc_1/2 = −0.08 V vs. SCE. Hence, iodide is a competent redox mediator to achieve the transformation from complex C to complex D. Analysis of non-covalent interactions²¹ in complex C (Fig. 2) show the presence of a weak stabilization interaction between the chalcogen''s anisole group and the 2-phenylpyridine. In contrast, in complex D these interactions were found more relevant between the o-carborane phenyl group and the chalcogen aromatic motif.Open in a separate windowFig. 2Non-covalent interaction plots for the complexes C and D. Strong attractive interactions are shown in blue, weak attractive interactions are given in green, while red corresponds to repulsive interactions. Ar = 4-MeOC₆H₄.Open in a separate windowScheme 4Control experiments and cyclic voltammograms.On the basis of the aforementioned findings,¹⁸ a plausible reaction mechanism is proposed in Scheme 5, which commences with an anodic single electron-transfer (SET) oxidation of the thiol anion E to form the sulfur-centered radical F. Subsequently, the copper(i) species A reacts with the sulfur radical F to deliver copper(ii) complex B, which next reacts with o-carborane 1 in the presence of LiOtBu to generate a copper(ii)-o-carborane complex C. Thereafter, the complex C is oxidized by the anodically generated redox mediator I₂ to furnish the copper(iii) species D,^18d which subsequently undergoes reductive elimination, affording the final product and regenerating the catalytically active complex A. Alternatively, the direct oxidation of copper(ii) complex C by electricity to generate copper(iii) species D can not be excluded at this stage.^18a,bOpen in a separate windowScheme 5Proposed reaction mechanism.In conclusion, a sustainable electrocatalytic C–H chalcogenation of o-carboranes with thiols and selenols was realized at room temperature by earth abundant copper catalysis. The C–H activation was characterized by mild reaction conditions and high functional group tolerance, leading to the facile assembly of various o-carboranes. Thereby, a transformative platform for the design of cage C–S and C–Se o-carboranes was established that avoids chemical oxidants by environmentally-sound electricity in the absence of directing groups. A plausible mechanism of paired electrolysis was established by detailed mechanistic studies.     

Direct synthesis of pentasubstituted pyrroles and hexasubstituted pyrrolines from propargyl sulfonylamides and allenamides

Multisubstituted pyrroles are important fragments that appear in many bioactive small molecule scaffolds. Efficient synthesis of multisubstituted pyrroles with different substituents from easily accessible starting materials is challenging. Herein, we describe a metal-free method for the preparation of pentasubstituted pyrroles and hexasubstituted pyrrolines with different substituents and a free amino group by a base-promoted cascade addition–cyclization of propargylamides or allenamides with trimethylsilyl cyanide. This method would complement previous methods and support expansion of the toolbox for the synthesis of valuable, but previously inaccessible, highly substituted pyrroles and pyrrolines. Mechanistic studies to elucidate the reaction pathway have been conducted.

This method is a toolbox for the synthesis of valuable, but previously inaccessible, highly substituted pyrroles and pyrrolines.

Pyrroles are molecules of great interest in a variety of compounds including pharmaceuticals, natural products and other materials. Pyrrole fragments for example are key motifs in bioactive natural molecules, forming the subunit of heme, chlorophyll and bile pigments, and are also found in many clinical drugs, including those in Fig. 1a.¹ Although many classical methods of pyrrole synthesis, including the Paal–Knorr condensation,² the Knorr reaction,³ the Hantzsch reaction,⁴ transition metal-catalyzed reactions,⁵ and multicomponent coupling reactions,⁶ have been developed over many years, the efficient synthesis of multisubstituted pyrroles is still challenging. In condensation syntheses of pyrroles, the major problems lie in the extended syntheses of complex precursors and limited substitution patterns that are allowed. Multicomponent reactions are superior when building pyrrole core structures with more substituents. Among these, the [2+2+1] cycloaddition reaction of alkynes and primary amines is attractive because of the readily available alkyne and amine substrates and the ability to construct fully substituted pyrroles.⁷ However, with the exception of some rare examples,⁸ most [2+2+1] cycloaddition reactions afford pyrroles with two or more identical substituents. The synthesis of multisubstituted pyrroles with all different substituents from simple starting materials therefore remains a major challenge.Open in a separate windowFig. 1Previous reports and this work on propargylamides transformation.Easily accessible propargylamides are classical, privileged building blocks broadly utilized for the synthesis of a large variety of heterocyclic molecules such as pyrroles, pyridines, thiazoles, oxazoles and other relevant organic frameworks.⁹ For example, Looper¹⁰et al. reported the synthesis of 2-aminoimidazoles from propargyl cyanamides and Eycken¹¹ reported a method starting from propargyl guanidines which undergo a 5-exo-dig heterocyclization as shown in Fig. 1b. Subsequently, Wan¹²et al. revealed the cyclization of N-alkenyl propargyl sulfonamides into pyrroles via sulfonyl migration. Inspired by these transformations and multi-substituted pyrrole synthesis, we report herein a direct synthesis of pentasubstituted pyrroles and hexasubstituted pyrrolines with all different substituents from propargyl sulfonylamides and allenamides.Previously, Zhu,¹³ Ji¹⁴ and Qiu^13b,15 reported efficient syntheses of 2-aminopyrroles from isocyanides. Ye¹⁶ and Huang¹⁷ independently developed gold-catalyzed syntheses of 2-amino-pentasubstituted pyrroles with ynamides. Despite the many advantages of these methods, they all afford protected amines, rather than free amines. The deprotection of these amines may cause problems in further transformations of the products. Our method delivers pyrroles with an unprotected free amino group and are often complementary to the previously well-developed classical methods.Initially, the cyclization reaction of N-(1,3-diphenylprop-2-yn-1-yl)-N-ethylbenzenesulfonamide (1a) with trimethylsilyl cyanide (TMSCN) was carried out with Ni(PPh₃)₂Cl₂ as a catalyst, a base (Cs₂CO₃) and DMF as a solvent. Different metal catalysts, such as Ni(PPh₃)₂Cl₂, Pd(OAc)₂, Cu(OAc)₂, and Co(OAc)₂ provided the desired product with similar yields ( EntryCat.BaseSolventYield1Ni(PPh₃)₂Cl₂Cs₂CO₃DMF67%2Pd(OAc)₂Cs₂CO₃DMF65%3Cu(OAc)₂Cs₂CO₃DMF65%4Co(OAc)₂Cs₂CO₃DMF63%5Cs₂CO₃DMF65%6KFDMFTrace7K₃PO₄DMFTrace8K₂CO₃DMF48%9KOHDMF52%10KO^tBuDMF46%11Et₃NDMFTrace12Cs₂CO₃CH₃CN18%13Cs₂CO₃DME23%14Cs₂CO₃TolueneTrace15Cs₂CO₃DCETrace16Cs₂CO₃DioxaneTraceOpen in a separate window^aReaction conditions: 1a (0.1 mmol, 1 equiv.), TMSCN (0.3 mmol, 3 equiv.), cat. (0 or 10 mol%), base (0.3 mmol, 3 equiv.) and solvent (1 mL), at 80 °C for 10 h; isolated yield.With the optimal reaction conditions in hand, we investigated the scope of this reaction. As shown in Fig. 2, the transformation tolerates a broad variety of substituted propargylamides (1). The R¹ group could be an aryl group containing either electron-donating groups or electron-withdrawing groups, and the corresponding products (2b–2h) were obtained in yields of 62–80%. The substituent R¹ could also be an alkyl group such as 1-hexyl in which case the reaction provided the corresponding pyrrole (2i) in 53% yield. Exploration of the R² substituent was also conducted. Electron-rich and electron-deficient substituents in the aromatic ring of R² gave the desired products (2j–2o) with yields of 70–81%. The product bearing a furyl group (2p) can be produced in 61% yield. However, when R² group is an aliphatic group, the reaction failed to provide the desired product. Substituent groups R³, such as benzyl (2q) or 3,4-dimethoxyphenylethyl (2r) were also compatible in the reaction, providing the corresponding products in moderate yields. Significantly, this method has the potential to produce core structures (for example 2s) similar to that in Atorvastatin. Interestingly, when alkynyl substituted isoquinolines (1t–1v) were used as the substrates, the reactions smoothly afforded fused pyrrolo[2,1-α]isoquinoline derivatives (2t–2v), members of a class of compounds that are found widely in marine alkaloids and exhibit anticancer and antiviral activity.¹⁸Open in a separate windowFig. 2Substrate scope of propargylamides. Reaction conditions: 1 (0.20 mmol, 1 equiv.), TMSCN (0.60 mmol, 3 equiv.), Cs₂CO₃ (0.60 mmol, 3 equiv.) and DMF (2 mL), at 80 °C for 10 h; isolated yield.Allenes are key intermediates in the synthesis of many complex molecules.¹⁹ As a subtype of allenes, allenamines are also useful as reaction intermediates.²⁰ Although the transformation of allenamides to multisubstituted pyrroles has not been previously recorded, this reaction probably goes through the allenamide intermediates which can be derived from propargyl sulfonamides under basic conditions. To verify this hypothesis, the trisubstituted allenamide (3) was synthesized and subjected to the standard reaction conditions. A pyrrole (2a) was isolated in 82% yield from this reaction (Fig. 3). This result confirmed our assumption and raised a new question: is it possible to build hexasubstituted pyrrolines from tetrasubstituted allenamides? A range of tetrasubstituted allenamides²¹ was tested under the standard reaction conditions, and the hexasubstituted pyrrolines were obtained as is shown in Fig. 4. The R¹ group could be an aryl substituent or an alkyl chain, and the corresponding products (5a–5e) were obtained with good yields. Various aryl groups with either electron-donating groups or electron-withdrawing groups in the aromatic ring of R² provided the desired products (5f–5k) in 62–83% yields. In addition, the difluoromethyl group can also be replaced by a phenyl group, and the reaction provided the corresponding product 5l in 82% yield. It is worth noting that these pyrroline products are not easily accessible from other methods.Open in a separate windowFig. 3Synthesis of substituted pyrroles from allenes.Open in a separate windowFig. 4Substrate scope of tetrasubstituted allenamides. Reaction conditions: 4 (0.10 mmol, 1 equiv.), TMSCN (0.30 mmol, 3 equiv.), K₂CO₃ (0.30 mmol, 3 equiv.) and DMF (1 mL), at 80 °C for 10 h, isolated yield.Some synthetic applications of this method are shown in Fig. 5. The amide is a naturally occurring and ubiquitous functional group. When using benzoyl chloride to protect the free amino group of the fully-substituted pyrrole (2a), a bis-dibenzoyl amide (6) was obtained in the presence of a base, triethylamine while the monobenzoyl protected amide (7) was obtained in the presence of pyridine as the base (Fig. 5a). This method also provides a straightforward approach to pyrrole fused lactam structures (Fig. 5b). For examples, a five-membered lactam and a six-membered lactam were generated separately in a one pot reaction, directly from, (8 and 10), respectively. Taking advantage of this method, an analogue of the drug Atorvastatin was synthesized in 5 steps (Fig. 5c), demonstrating the synthetic value of the reaction.Open in a separate windowFig. 5Synthetic applications.Mechanistic experiments were performed (Fig. 6) to explore the mechanism of the reaction. When 3 equivalents of TEMPO were added, the reaction was not inhibited and the desired product (2a) was formed in 62% yield (Fig. 6a). This result suggested that the reaction might not involve a radical process. To probe the reaction further, a kinetic study was conducted (Fig. 6b). According to this study, the propargylamide (1a) was completely converted to an allenamide (3a) in 10 min under the standard conditions. The multi-substituted pyrrole (2a) was then gradually produced from the intermediate allenamide and no other reaction intermediates were observed or identified. On the other hand, DFT calculations of substrates 3b and 4a were carried out at the B3LYP-D3(SMD)/Def2-TZVP//B3LYP-D3/Def2-SVP level of theory to identify the natural bond orbital (NBO) charges on the carbons of the allene moieties. NBO charges on the internal carbon in both 3b and 4a are 0.11 and 0.18, respectively (Fig. 6c) indicating that the nucleophilic addition of cyanide anion onto the internal carbon should be reasonable as opposed to its addition onto the terminal carbon. Pathways of the cyano addition to 3b were also calculated (Fig. 6d). The transition state of cyano addition on the internal carbon (TS1), is indeed much lower than addition on the terminal carbon (TS2). The intermediate of internal carbon addition int1, is more stable than int2, implying that the internal carbon addition pathway is not only kinetically but also thermodynamically favoured.Open in a separate windowFig. 6Mechanistic studies and proposed mechanism.Based on the results of these mechanistic studies, a plausible reaction mechanism for the synthesis of pentasubstituted pyrroles and hexasubstituted pyrrolines is proposed and is shown in Fig. 6e. First, under basic conditions, the propargylamide isomerizes to an intermediate allenamide (A), which can be attacked nucleophilically by the cyanide anion to afford an intermediate imine (B) with release of the sulfonyl group. Then, the second cyanide anion attacks the imine to form an intermediate (C), which can undergo cyclization and protonation to afford the fully substituted pyrrole (2). Similarly, the hexasubstituted pyrroline product (5) can be obtained from double nucleophilic attack of the intermediate (A) by the cyanide ion.     

Strong non-Arrhenius behavior at low temperatures in the OH + HCl → H2O + Cl reaction due to resonance induced quantum tunneling

The OH + HCl → H₂O + Cl reaction releases Cl atoms, which can catalyze the ozone destruction reaction in the stratosphere. The measured rate coefficients for the reaction deviate substantially from the Arrhenius limit at low temperatures and become essentially independent of temperature when T < 250 K, apparently due to quantum tunneling; however, the nature of the quantum tunneling is unknown. Here, we report a time-dependent wave packet study of the reactions on two newly constructed potential energy surfaces. It is found that the OH + HCl reaction possesses many Feshbach resonances trapped in a bending/torsion excited vibrational adiabatic potential well in the entrance channel due to hydrogen bond interaction. These resonance states greatly induce quantum tunneling of a hydrogen atom through the reaction barrier, causing the reaction rates to deviate substantially from Arrhenius behavior at low temperature, as observed experimentally.

The OH + HCl reaction possesses many Feshbach resonances trapped in the hydrogen bond well in the entrance channel, which substantially enhance the reaction rates at low temperatures.

In the classical picture, a chemical reaction with an energetic barrier can only occur at collision energies higher than the barrier, which leads to the well-known Arrhenius formula for chemical reaction rates. However, chemical reactions can happen at energies below the reaction barrier through quantum tunneling,^1–3 resulting in the deviation of the reaction rates from the Arrhenius behavior at low temperatures. The effect of quantum tunneling on the reaction rates increases with decreasing reaction temperature, and hence becomes especially important in low-temperature environments such as the interstellar medium and atmospheric processes.^4,5 Reaction resonances are quasi-trapped quantum states in the transition state region with some lifetime, and can substantially promote quantum tunneling through the reaction barrier. Over the past decades, great efforts have been devoted to detecting resonances in chemical reactions and to studying their structures and dynamics.^6–14 Theoretical studies on the O(³P) + HCl → OH + H reaction using the accurate ³A′′ potential energy surface revealed that the tunneling induced by the resonances trapped in the van der Waals well in the reactant channel can substantially enhance the thermal rate constants at low temperatures.^15–17 A combined experimental and theoretical investigation discovered that resonance-induced quantum tunneling dramatically enhances the reactivity of the F + p-H₂ → HF + H reaction,¹⁴ and is fully responsible for the unusually high chemical reactivity of the reaction in the low temperature interstellar medium. Recently, a quantum dynamics calculation showed that the presence of two resonance peaks strongly influence the rotational quenching of HF (j = 1, 2) with H, leading to an up to two-fold increase in the thermal rate coefficients at the low temperatures characteristic of the interstellar medium.¹⁸ Thus, understanding quantum tunneling, and in particular, resonance-induced quantum tunneling, in chemical reactions is of general interest and fundamental importance to low-temperature chemistry.The OH + HCl → H₂O + Cl reaction is of great importance in atmospheric chemistry because it releases Cl atoms from one of the principal chlorine-containing species in the stratosphere, HCl. The Cl atoms generated from the reaction can catalyze the ozone destruction reaction in the stratosphere, which was responsible for the formation of the ozone hole over Antarctica.¹⁹ Since the chlorine-catalyzed ozone destruction is proportional to the steady-state Cl atom concentration, which is directly controlled by the rate of the reaction, extensive studies have been carried out to measure the rate coefficient for the reaction with high accuracy over a wide range of temperatures.^20–29 The measured rate coefficients exhibit a small activation energy of a few hundred K, deviate substantially from the Arrhenius limit at low temperatures and become essentially independent of temperature when T < 250 K.^25,28,29 In addition, a large H/D kinetic isotope effect has also been found.^21,22,26,28 All these observations suggest the presence of an important quantum tunneling effect in the reaction.The dynamics of this reaction and its reverse have also attracted great attention in the past decades. In particular, the endothermic Cl + H₂O → HCl + OH reaction with a late barrier has been extensively investigated as a benchmark system for mode specificity and bond selectivity chemistry.^30–35 Recently, the construction of two high-quality potential energy surfaces (PESs) in its ground electronic state using the PIP-NN method have substantially advanced the theoretical study of the dynamics and kinetics of the system. The first PES was based on a large number of ab initio data points calculated at the multi-reference configuration interaction (MRCI) level of theory by Li, Dawes, and Guo (LDG),³³ and the second one was fitted to ab initio energy points obtained using an explicitly correlated unrestricted coupled-cluster method with single, double, and perturbative triple excitations (UCCSD(T)-F12b) and the augmented correlation-consistent polarized valence triple-zeta (aug-cc-pVTZ, or AVTZ) basis set by Zuo, Zhao, Guo, and Xie (ZZGX) with a fitting error of 6.9 meV.³⁶ The static barrier height is 2.86 and 2.23 kcal mol⁻¹, respectively, for these two PESs. On both PESs, there exists a well of about 3.5 kcal mol⁻¹ in the OH + HCl entrance channel due to the hydrogen bond (HB) interaction between OH and HCl. Extensive quantum dynamics studies on the PESs have revealed many interesting features of the reaction in both directions. In particular, time-dependent wave packet calculations for the title reaction with OH in the ground and vibrational excited states found one or two broad peaks in the total reaction probabilities, which are presumed to be the signature of the resonances supported by the reactant complex well.^36,37 However, the impact of these peaks on the reaction rates has not been investigated. Ring-polymer molecular dynamics (RPMD) calculations^38,39 were also carried out on both PESs to compute the thermal rate coefficients for the reaction.^40–42 It was found that the RPMD rates on the LDG PES underestimate the experimental data, while the RPMD rates on the latest ZZGX PES agree with the experimental results much better, and do not decrease further when the temperature falls below 300 K, apparently due to quantum tunneling. Unfortunately, RPMD calculations cannot provide any clue regarding the nature of the quantum tunneling.Therefore, despite the significant progress that has been made in theoretical studies of the system, some key issues still remain to be addressed: Are the broad peaks in the total reaction probabilities obtained from the time-dependent wave packet calculations indeed the signature of the resonances in the reaction? If not, do there exist resonances in the reaction? How do resonances affect the reaction rates at low temperature? Here, we report a quantum dynamics study of the reaction on two new and more accurate PESs. Good agreement is achieved between the rate coefficients calculated on these two PESs and the experimental data. Our calculations reveal that the HB well in the entrance channel of OH + HCl supports many low energy resonance states. These resonance states substantially enhance the quantum tunneling effect and have an important impact on the reaction rates at low temperatures.In order to improve the fitting accuracy of the ZZGX PES, we constructed two new PESs using the fundamental invariant neural network (FI-NN) method, fitted to ∼70 000 ab initio energy points calculated at the UCCSD(T)-F12a and UCCSD(T)-F12b levels of theory, respectively, both with the AVTZ basis set. The fitting RMSE is 3.07 and 3.12 meV, respectively, for the F12a and F12b PES, which is about half that for the ZZGX PES.³⁶ The spin–orbit coupling of the channels of both the reagent OH and the product Cl have been included using FI-NN fitting to about 38 000 points calculated at the MRCI/aug-cc-pVTZ level of theory with a fitting error of 0.19 meV. The CASSCF wave-function with an active space of (5e, 3o) was used as a reference for MRCI. Details of the new PESs are provided in the ESI^†. The static barrier height for PESa is 0.088 eV (0.095 eV with SO correction included), and that for PESb is 0.097 eV (0.104 eV). As can be seen from Tables S1 and S2,^† the geometries and energies of all the stationary points for the F12b PES without SO correction are in good agreement with the ZZGX PES. Table S2^† also shows the corresponding complete basis set (CBS) energies for these stationary points based on AVTZ, AVQZ and AV5Z calculations. Because the F12b energies are slightly closer to the CBS results, we will present the dynamical results obtained on the F12b PES in the main text and provide those for the F12a PES in the SI.On the new PESs, we carried out potential-averaged five-dimensional (PA5D) time-dependent wave packet^43,44 calculations to obtain the total reaction probabilities for the reaction by freezing the non-reacting OH bond in its ground vibrational state. Tests revealed that the PA5D treatment is capable of providing reaction probabilities for the ground rovibrational initial state that are essentially identical to those obtained using the full six-dimensional approach, as shown in Fig. S2.^†Fig. 1(A) shows the total reaction probabilities for the HCl + OH reaction as a function of collision energy calculated on the F12b PES with both reagents in the ground rovibrational state at propagation times of 60 000, 120 000, 360 000, and 2 400 000 a.u. At high collision energies, the reaction probabilities converge quickly with respect to the propagation time, and one can barely see any difference among the four reaction probability curves, which exhibit smooth increases with collision energy. However, in the low collision energy region, large differences appear for different propagation times. At t = 60 000 a.u., the reaction probability presents a smooth curve with some small and broad oscillations, as was observed in the wave packet calculations of Guo and coworkers.³⁶Open in a separate windowFig. 1(A) Total reaction probabilities for the ground initial state of the OH + HCl → Cl + H₂O reaction on the F12b PES at wave packet propagation times of T = 60 000, 120 000, 360 000, and 2 400 000 a.u. (B) Same as (A), except showing the collision energy between 0.0 and 0.04 eV. The crosses mark the points for which the wavefunctions are shown in Fig. 2. (C) Total reaction probabilities for some partial waves J = 0, 30, 60, and 90 as a function of the collision energy. (D) Same as (C) except showing the collision energy between 0.0 and 0.04 eV.When the propagation time is increased to t = 120 000, the reaction probabilities at collision energies below 0.05 eV increase substantially. With further increasing the propagation time to 360 000 a.u., many oscillatory structures emerge at collision energies below the barrier height of 0.104 eV, in particular in the very low collision energy region as shown in Fig. 1(B). These sharp structures become fully converged after around 2 400 000 a. u. of wave packet propagation (∼58 ps). The reaction probability even at a collision energy close to zero reaches 3–4%. The convergence of the total reaction probabilities on the F12a PES is very similar to that on the F12b PES (Fig. S3^†), except that the final converged reaction probabilities for these two PESs exhibit a small shift, apparently due to slightly different barrier heights. Therefore, it is clear that reaction resonances exist in the title reaction in the very low collision energy region, and that these resonance states substantially induce quantum tunneling and enhance the reactivity. The lifetimes for these resonance states are quite long, with many being longer than 6.5 ps and having corresponding widths smaller than 0.1 meV. Fig. 1(C) presents converged (t = 2 400 000 a.u. ≈ 58 ps) total reaction probabilities for the total angular momentum J = 0, 30, 60, and 90. With increasing J, the reaction probability curve shifts to higher energy. In the low collision energy region (<0.05 eV), the total reaction probabilities for J = 30 exhibit rich oscillatory structures as in the J = 0 case (Fig. 1(D)), which are expected to have a great influence on the rate constant at low temperature. With further increase of the total angular momentum, the influence of these resonances on the total reaction fades due to the centrifugal barrier, which prevents the low-energy wave function from entering the well. They only leave a small trace in the total reaction probabilities at low energies for J = 60, and do not have any effect for J = 90.To understand the nature of these resonances, we calculated scattering wave functions at two collision energies (1.08 and 4.26 meV) with the peak reaction probabilities indicated by x in Fig. 1(B). Fig. 2(A) shows the two dimensional (2D) contour at the collision energy of 1.08 meV in the Jacobi coordinates HCl bond length (r_H–Cl) and center of mass distance between OH and HCl (R_HCl–OH), with the other coordinates integrated. As can be seen, the wave function is localized in the HB well region in the entrance channel with a few nodes in the R coordinate and no node in the r_H–Cl coordinate. Inspection of the scattering wave function for the bending and torsion coordinates reveals nodes exist in these coordinates (Fig. S4 and S5^†). The 2D contour in the coordinates of R_HCl–OH and r_H–Cl at the collision energy of 4.26 meV shown in Fig. 2(B) looks similar to that shown in Fig. 2(A), except with more nodes in the R direction. Therefore, the observed resonance states in the reaction are Feshbach resonances trapped in a bending/torsion excited vibrationally adiabatic potential (VAP) well in the reactant complex region due to the HB interaction.Open in a separate windowFig. 2Reactive scattering wave functions for the OH + HCl → Cl + H₂O reaction on the F12b PES in the two Jacobi coordinates R(HCl–OH) and r(H–Cl) with other coordinates integrated at the collision energies of 1.08 (A) and 4.26 meV (B). The contour lines are the corresponding 2D PESs along the two reactive bonds R(HCl–OH) and r(H–Cl) with other coordinates optimized. The geometries for the saddle point and the HB minimum are displayed in (A). The coordinate units in the figures are a₀. Fig. 3(A) shows the accurate rate constants for the initial ground rovibrational state, k_g, based on the probabilities for J = 0, 30, 60, 90 using the uniform J-shifting approach with a temperature-dependent shifting constant.^45–47 A test shows that the J-shifting scheme based on these four individual J values only introduces a few percent error to the rate constants in the temperature region considered here (Fig. S6^†). As can be seen from the figure, with decreasing temperature, k_g first decreases rapidly from 1000 K to 700 K, then decreases slowly. It reaches a minimum at T ≈ 260 K, and increases slowly with further decrease of the temperature. At temperatures lower than 300 K, the rate constants for the ground rovibrational initial state are larger than the measured thermal rate coefficients, with k_g being larger than k_exp by ∼70% at T = 200 K.Open in a separate windowFig. 3(A) Accurate rate constants, k_g, for the initial ground rovibrational state of the HCl + OH → H₂O + Cl reaction calculated on the F12b PES, in comparison with k^JS_g (obtained using the J-shifting approximation) and k^NR_g (based on the background reaction probabilities for J = 0 with the resonance contribution removed shown below); (B) the background reaction probabilities up to E = 0.1 eV by connecting some valleys of the reaction probabilities marked by x.Also shown in Fig. 3(A) are the rate constants for the ground rovibrational initial state, k^JS_g, obtained from the J = 0 reaction probabilities using the J-shifting approximation (see ESI^† for details). As can be seen, the J-shifting approximation works very well at high temperatures around 1000 K, but begins to overestimate the rates with decreasing temperature. At T = 500 K, k^JS_g is about 10% higher than the true rate. At temperatures below 300 K, the J-shifting approximation underestimates the reaction rate, with k^JS_g being smaller than k_g by about 16% at T = 200 K. Overall, the J-shifting approximation works fairly well for the ground rovibrational initial state, although there are numerous resonance peaks in the reaction probabilities in the low collision energy region.Now we consider the issue of the effect of the resonance structures found in the reaction on the rate constant. For a reaction system with isolated resonances, it is rather straightforward to remove the resonance contributions from the reaction probabilities by fitting the resonance peaks to some Lorentzian functions and to obtain smooth background scattering probabilities, as demonstrated in the F + HD reaction and recently in the inelastic scattering of H + HF (ref. 6 and 18). However, the OH + HCl reaction possesses numerous highly overlapped resonances in the low energy region, as shown in Fig. 1(A), and it is impractical to fit these resonance peaks as accurately as the isolated resonances. Instead, we obtained an approximate background reaction probability curve, which is shown in Fig. 3(B), by smoothly connecting some resonance valleys as shown in the figure. The rate constants k^NR_g calculated using the background curve shown in Fig. 3(B) with the J-shifting approximation were compared with the original k_g in Fig. 3(A). Given the fact that the J-shifting approximation works fairly well for k^JS_g, as shown in Fig. 3(A), it is very reasonable to expect that will work even better for the k^NR_g values obtained from the background reaction probabilities with the resonance contribution removed. As can be seen, k^NR_g exhibits rate behavior typical for systems with a low barrier with some quantum tunneling effects. At T = 1000 K, k^NR_g is essentially identical to k_g, but decreases much faster than k_g as the temperature drops. At T = 200 K, k_g is larger than k^NR_g by a factor of about 5.6 (9.8 × 10⁻¹³vs. 1.74 × 10⁻¹³ cm³ s⁻¹), indicating that the reaction probabilities in the low collision energy region due to resonances substantially enhance the rate constants for the reaction at low temperatures. It is worthwhile to note that for systems with overlapped resonances like that shown in Fig. 3(B), the reaction probabilities at the valleys must be considerably higher than the true background reaction probabilities; therefore, the background curve shown in Fig. 3(B) is the upper limit of the background reaction probabilities and the rate based on the curve shown in Fig. 3(A) is also the upper limit of the rate for the ground rovibrational initial state without the resonance contributions. Therefore, the true enhancement due to the resonances must be larger than that shown in Fig. 3(A).For reliable comparison with the measured thermal rate coefficients, we must take into account the contributions from all the thermally populated initial states of the reagents. Due to the very large number of thermally populated rotational states for this reaction even at 200 K, we opted to calculate the cumulative reaction probabilities, NE(E), (the sum of the reaction probabilities for all the initial states with a fixed total energy) for the reaction from which the thermal rate constants can be reliably evaluated.^48–52 The transition state wave packet calculations were carried out to obtain NE(E) using the details given in the ESI^†. Due to huge computational efforts required to obtain the cumulative reaction probabilities for J > 0, we only calculated NE(E) for J = 0 and employed the J–K-shifting approximation^45,49,53 to obtain the thermal rate constant. NE(J = 0, E) as a function of total energy measured with respect to the ground rovibrational energy of OH and HCl is presented in Fig. S7.^†In Fig. 4, we present thermal rate constants for the reaction calculated on both the F12a and F12b PES, together with the rate constants for the ground rovibrational state (k_g) and the previous experimental measurements.^{21,22,24–29} As can be seen, the thermal rate coefficients are smaller than k_g over the entire temperature region, indicating that reagent rotation excitations diminish the reaction rates. Overall, the thermal rate coefficients calculated on both PESs agree with the experimental results rather well, with the F12a PES slightly overestimating and the other PES slightly underestimating compared to the experimental measurements. As observed in the experiments, the thermal rate coefficients decrease quickly with decreasing temperature in the high-temperature region, but decrease much more slowly at low temperatures, in particular at T < 300 K, substantially deviating from Arrhenius behavior.Open in a separate windowFig. 4Thermal rate constants of the HCl + OH → H₂O + Cl reaction calculated on both the F12a and F12b PES, compared with rate constants for the ground rovibrational state, the RPMD rates on ZZGX PES and the previous experimental measurements.^{21,22,24–29} Experimental data are taken from Husain et al. (in circles),^21,22 Molina et al. (in diamonds),²⁴ Ravishankara et al. (in upward-pointing triangles),²⁵ Smith et al. (in squares),²⁶ Sharkey et al. (in triangle left),²⁷ Battin-Leclerc et al. (in downward-pointing triangles),²⁸ and Bryukov et al. (in leftward-pointing triangles).²⁹Also shown in Fig. 4 are the RPMD rates calculated (see ESI^† for details) on the F12a, F12b and ZZGX PESs.⁴² As shown, the RPMD rates on the F12b PES agree with those on the ZZGX PES extremely well, although the barrier heights for the F12b PES are higher by 9 meV. The present RPMD rates agree with the J–K-shifting quantum rates rather well, except at 200 K, at which the RPMD rates are overestimated by about 40%. As a result, the RPMD rate is higher than the experimental value at T = 200 K, in particular for the F12a PES. Previous studies have shown that RPMD tends to underestimate reaction rates in the strong quantum tunneling region, even for systems with resonances such as the O(³P) + HCl → OH + H reaction;⁵⁴ however, it overestimates the rates for the F + H₂ reaction with pronounced post-barrier Feshbach resonances⁵⁵ and some insertion reactions.⁵⁶ The discrepancy between the RPMD and J–K-shifting quantum rate at T = 200 K could be caused by a possible underestimation of the rate by the J–K-shifting approximation, as in the J-shifting approximation of the ground rovibrational initial state shown in Fig. 3(A). On the other hand, the rapid increase in the rate with decreasing temperature (from 250 K to 200 K) could also be a feature of bimolecular reaction rates in the high-pressure limit for reactions with a pre-reactive minimum.⁵⁷ RPMD rate theory in its original form employs a free energy calculation,^38,39 which allows thermalization in the pre-reactive minimum, sampling tunneling pathways with energies not accessible in the low-pressure limit. Therefore, RPMD models a rate process that resembles rates in the high-pressure limit, which could lead to an overestimation of the rate at low temperature. A recent study also reported a similar issue with RPMD.⁵⁸ The authors attributed it to spurious resonances in RPMD favoring energy transfer in the pre-reactive minimum, which is similar to our argument. Certainly, more quantum dynamics calculations to provide rigorous quantum rates at low temperatures to assess the accuracy of the J–K-shifting approximation and the RPMD method on this reaction system with strong HB interaction in the entrance channel would be highly desirable.Therefore, the OH + HCl reaction possesses many long-lifetime Feshbach resonances trapped in a bending excited VAP well in the entrance channel due to HB interaction. These resonance states substantially induce quantum tunneling of the hydrogen atom through the reaction barrier and enhance the reactivity in the low-collision region. Consequently, the reaction rate for the reaction becomes essentially independent of temperature in the low-temperature region and deviates substantially from Arrhenius behavior, as observed experimentally. The resonance states in the reaction are very different in location from those in the F + H₂/H₂O reactions,^{8,11,12,14,59–63} which are trapped in the VAP well in the product channel. Furthermore, they are trapped in the bending/torsion excited VAP well with HCl in the ground vibrational state, unlike those in the F + H₂/H₂O reactions with HF in vibrationally excited states.^8,63,64 In nature, these resonance states in the reaction arise from HB interaction as in the F + H₂O (v = 0) reaction,^11,62,63 but are different from those in the F + H₂/HD/HOD (v = 1) reactions^{8,10,12,14,64–66} due to chemical bond softening. Because the resonance states are trapped in the bending/torsion excited VAP well, their lifetimes are much longer than those observed in the F + H₂/H₂O reactions. As a result, they should have an important impact on the differential cross sections. More efforts, and in particular more joint efforts involving theory and experiment, should be devoted to studying these Feshbach resonances in this reaction of great atmospheric importance in detail.     

Photostable polymorphic organic cages for targeted live cell imaging

Fluorescent microscopy is a powerful tool for studying the cellular dynamics of biological systems. Small-molecule organic fluorophores are the most commonly used for live cell imaging; however, they often suffer from low solubility, limited photostability and variable targetability. Herein, we demonstrate that a tautomeric organic cage, OC1, has high cell permeability, photostability and selectivity towards the mitochondria. We further performed a structure–activity study to investigate the role of the keto–enol tautomerization, which affords strong and consistent fluorescence in dilute solutions through supramolecular self-assembly. Significantly, OC1 can passively diffuse through the cell membrane directly targeting the mitochondria without going through the endosomes or the lysosomes. We envisage that designing highly stable and biocompatible self-assembled fluorophores that can passively diffuse through the cell membrane while selectively targeting specific organelles will push the boundaries of fluorescent microscopy to visualize intricate cellular processes at the single molecule level in live samples.

In this article, we demonstrate the relatively unexplored potential of organic cages for use in targeted live cell imaging and highlight the importance of inter- and intramolecular interactions to stabilize and improve the performance of fluorophores.

Organic fluorophores are of great interest in many research fields especially live cell imaging, which allows for noninvasive observation and monitoring of biological processes. Small molecule fluorophores can be functionalized and tuned with high precision to tag a diverse variety of biological targets; however, limited water solubility, cell-permeability, photostability and targetability have halted the biomedical translation of many promising fluorescent molecules.¹ Ultimately, new generations of fluorescent tags were designed to include new functionalities that can improve their physical properties and cell penetration such as long aliphatic side chains and cell penetrating peptides.^2–6 In addition, various strategies have been employed to improve the photostability of these compounds by the addition of electron withdrawing groups, conformational changes, encapsulation or incorporation within host macrocycles.^7–13Major efforts in the organic fluorophores development have been directed towards designing stable probes targeting the mitochondria as it plays a central role in the generation of adenosine triphosphate (ATP), central metabolism, and apoptosis.^14,15 Successful examples of mitochondria targeting fluorophores include triphenyl phosphonium cations, heterocyclic aromatic cations, macrocyclic amphiphiles, BODIPY derivatives and mitochondria targeted peptides that have limited solubility.^16–21 Commercially available dyes for mitochondrial imaging such as Mito Tracker Red have a high photobleaching tendency and can form covalent bonds with the mitochondrial respiratory chain complex I resulting in increased cytotoxicity. Consequently, various organic, inorganic and hybrid platforms were developed ranging from small nanoparticles to self-assembled frameworks on the quest for improved mitochondrial imaging agents.^22–27Porous organic cages have received major attention over the last decade as they are easily prepared, soluble in a range of solvents and are intrinsically porous.^28–31 They have shown excellent applicability in molecular recognition, sensing and hydrocarbon separation but their biomedical imaging applications have never been investigated.^32–35 Recently, self-assembled organic imine stacks showed impressive electroluminescence and live cell imaging.³⁶ Moreover, an intracellular targeted macrocyclic nanohoop showed fast cell uptake and two-photon live cell fluorescence imaging.³⁷ In this work, a tautomeric organic cage (OC1) that shows unprecedented photostability for live-cell imaging with high biocompatibility, cell permeability and mitochondrial targetability is reported (Fig. 1). We hypothesize that the keto–enol tautomerism is playing a major role in promoting the stability and sustaining the fluorescence of this platform. Such effect has been previously reported in other organic dyes such as oxyluciferin and HDFL.^38,39Open in a separate windowFig. 1Tautomeric forms of OC1 and chemical structures of OC2 and OC3.Investigating other organic cages with less hydroxyl groups (OC2) or replacing the hydroxyl groups with methoxy substituents (OC3) resulted in the loss of cell stability, retention and targetability (Fig. 1). The presence of the –OH groups in OC1 not only enables keto–enol tautomerization but also promotes H-bonding interactions, which ultimately improves solubility and cell permeability. We envision that understanding the nature of the intra- and intermolecular interactions in organic probes can lead to a profound enhancement in the photobleaching efficacy in addition to cell permeability. OC1 can effectively image the mitochondria over 3 h with high selectivity and without any loss of efficacy (no photo bleaching), which is comparable to the commercially used MitoTracker RedFM.OC1, a [2 + 3] cage, was synthesized following a reported procedure and its structure was confirmed by ¹H Nuclear Magnetic Resonance (NMR) and TOF-Mass analysis (Fig. S1–S4^†).³² Dynamic light scattering (DLS) of OC1 revealed an average size of 30–50 nm in DMSO–H₂O (2–5%) with a negative zeta potential of −5 mV (Fig. S5^†). Single crystal X-ray analysis revealed that OC1 crystallizes in a triclinic crystal system with chiral P1 space group and the asymmetric unit consists of two molecules of [2 + 3] cages (Fig. S6^†). Crystal structure analysis suggests that the imine cage crystallizes as the keto tautomeric form (Fig. 2a). This results in the formation of intramolecular N–H⋯O hydrogen bonding between the imine (N–H) hydrogen and the keto (C O) oxygen (d_{C–OH actual} = 1.36 Å; d_{C O actual} = 1.25–1.26 Å vs. d_{C O for OC1} = 1.260 Å and d_{C N actual} = 1.25 Å vs. d_{C–NH for OC1} = 1.30–1.35 Å) (Fig. S6^†). Three different windows of each cage are connected with three different cages in the same plane through weak C–H⋯O hydrogen bonding (d = 2.414 Å, and θ = 60°), which creates isolated pores at the center of the cage (Fig. 2b). Furthermore, the packing diagram suggests the formation of one-dimensional infinite extrinsic pores along the b-axis (Fig. 2c and S6^†). OC1 is stable in aqueous solutions even at low pH as well as in cell media (Fig. S7–S10^†).Open in a separate windowFig. 2(a) SCXRD and crystal image of OC1. (b) OC1 self-assembled through hydrogen bonding interactions. (c) Packing of OC1 along b-axis.To test our original proposal that organic cages can act as effective imaging agents, we first investigated the UV-visible spectrum of OC1 in DMSO, which exhibited two characteristic peaks at λ_max = 286 and 330 nm that can be assigned to π–π* and n–π* transitions, respectively (Fig. S11^†). Fluorescence emission spectra showed that OC1 exhibits two major peaks at around 411 and 497 nm upon excitation at 330–350 nm in DMSO, while excitation at 400 and 480 nm showed characteristic emission peaks at 503 and 538 nm, respectively (Fig. S12^†). This behavior can be explained by the presence of two emissive forms due to the keto–enol tautomerization. The quantum yield (QY) of OC1 in DMSO (0.25) and chloroform (0.4) was also tested. Molar extinction coefficient of OC1 was estimated as 46 806 M⁻¹ cm⁻¹ in DMSO (Fig. S13–S14^†). The quantum yield of OC1 in DMSO are about half that of in chloroform that is significantly matched with the reported intracellular targeted macrocyclic nanohoop structure.³⁷ The fluorescent properties of OC1 were maintained at different pH values (4, 7 and 10) (Fig. S15^†). We hypothesize that this mode of assembly, as supported by SCXRD, promotes stability and prevents aggregation-induced quenching that is very common in small molecule dye fluorophores. The keto–enol tautomerism is boosting intra- and intermolecular hydrogen bonding interactions that can be contributing to the emissive characteristics of this system. In fact, the presence of the phenolic OH groups have been reported to have critical effects on the structural and spectroscopic properties of organic cages.^31,40We then proceeded to test OC1 for cell imaging by first testing the cytotoxicity of OC1 in MCF-7 cells (Fig. S16a^†). The cells were treated with a solution of concentrations ranging from 3.9–500 μg ml⁻¹ for 24 h. Cell death was measured using MTT assay demonstrating the excellent biocompatibility of OC1 with an IC50 of > 500 μg ml⁻¹. MCF-7 cells were then either co-incubated with OC1 and LysoTracker Red or MitoTracker RedFM, the cells are then fixed with 4% formaldehyde (w/v) and imaged on a confocal laser-scanning microscope (CLSM). Interestingly, OC1 showed strong colocalization in the mitochondria with a Pearson''s correlation coefficient of 0.9751 (Fig. S16b and c^†). Real time imaging of MCF-7 cells at different time intervals after co-internalization of OC1 and MitoTracker showed a clear signal overlap on a subcellular level (Fig. 3 and MOV1), which ultimately supports that OC1 can selectively target the mitochondria.Open in a separate windowFig. 3Live cell imaging of MCF-7 cells incubated with 250 μg ml⁻¹ of OC1 for 3 h showing the uptake and colocalization of OC1 (abs/em ∼495/519 nm) with the commercial probe MitoTracker RedFM (abs/em ∼581/644 nm) in the mitochondria (scale bar at 50 μm).These encouraging results piqued our interest to investigate other organic cages in order to have a better understanding of the mechanistic factors in play. As the tautomeric activity is well reported to affect the photophysical properties of organic compounds, we compared OC1 to an organic cage with less phenolic OHs (OC2) and an organic cage with only methoxy substituents (OC3). OC2 and OC3 were synthesized and characterized according to reported procedures (Fig. S17–S25^†).^32,41 Absorbance, emission properties and stabilities of OC2 and OC3 in DMSO were also verified. OC2 showed one major characteristic emission peak at around 547 nm upon excitation at 350 and 440 nm. The quantum yield (QY) of OC2 in DMSO (6.7%) and chloroform (11.1%) was also tested (Fig. S13 and S14^†). OC3 showed emission at 494 and 537 nm upon excitation at 320 nm and 480 nm upon excitation at 420 nm (Fig. S26^†). The quantum yield (QY) and molar extinction coefficient of OC3 in DMSO is negligible. Cytotoxicity and biocompatibility studies of OC2 and OC3 showed a relatively higher cytotoxicity compared to OC1 (Fig. S27^†). As to targetability and photostability, OC2 and OC3 showed less selectivity with colocalization in both mitochondria and lysosome where the fluorescent signal slowly disappeared, which implies the OC2 and OC3 are less stable than OC1 (Fig. S28^†). Moreover, FACS data revealed that OC1 showed the highest uptake percentage in comparison to OC2, and OC3 with an uptake percentage of 86.4%, 78%, and 69%, respectively (Fig. S29^†). Based on this data, we can conclude that indeed the hydroxyl groups, and their prompted keto–enol tautomerism, are playing a major role in stabilizing the overall structure through hydrogen bonding while simultaneously improving photostability and targetability. As a control, we tested the aldehyde starting material, 1,3,5-triformylphloroglucinol, to verify that OC1 is being uptaken as a whole and not as the aldehyde building block or a possible decomposition product. The experiment clearly demonstrates that 1,3,5-triformylphloroglucinol as a fluorophore had no selectivity towards the mitochondria and is initially distributed in the cytoplasm and finally localizes in the lysosomes for degradation (Fig. S30^†).As for the uptake mechanism, we initially hypothesized that OC1 will be taken up by cells through endocytosis, which is the typical biological pathway of nanoparticles and nanoassemblies.^42,43 Surprisingly, we discovered that OC1 can passively diffuse rather than being actively transported through the cell membrane. Passive transport is a type of membrane transport that does not require energy to move substances across cell membranes as it mainly relies on the second law of thermodynamics. We tested OC1 uptake in the presence of different endocytic inhibitors including chlorpromazine (CPZ), filipin (FIL), and amiloride to inhibit clathrin-mediated, caveolae mediated, and macropinocytotic endocytosis pathways, respectively.⁴⁴ All cells were treated with the inhibitors for 30 min, followed by 3 h incubation with OC1. The cellular uptake data showed that there was no significant reduction in the uptake using the active transport inhibitors (Fig. 4a). Moreover, the passive transport was tested by incubating OC1 (125 μg ml⁻¹) with MCF-7 for 3 h at different temperatures (4, 27, or 37 °C). The uptake was monitored by confocal microscopy (Fig. 4b) and quantified by FACS (Fig. 4c). Interestingly, no obvious reduction of uptake was observed in all the cases.Open in a separate windowFig. 4Cellular uptake mechanism of OC1. (a) Cell uptake of OC1 in the presence of endocytosis inhibitors to investigate endocytic uptake mechanism. (b) MCF-7 cell uptake CLSM of OC1 at 37 °C, 25 °C, and 4 °C, respectively (scale bar at 30 μm). (c) Quantification of cell fluorescence relative to 37 °C.After verifying the targetability and the uptake mechanism of OC1, we focused our attention on photostability. Since the commercially available mitochondrion tracker dyes normally have moderate photostability, it is critical to evaluate the photostability of OC1 in live cells. We first compared the photostability of OC1 with the commercially available MitoTracker RedFM. Since the excitation/emission wavelengths of OC1 (abs/em ∼495/519 nm) and MitoTracker RedFM (abs/em ∼581/644 nm) are distant from each other, we performed simultaneous scans for both dyes for 100 scans at 1.5 s per scan by incubating OC1 and MitoTracker RedFM with MCF-7 cells at 37 °C for 48 h. The results showed that at a laser power density of P ∼ 0.4132 W cm⁻²OC1 (ε = 46 806 M⁻¹ cm⁻¹) had higher retention and photostability than that of the MitoTracker RedFM where OC1 retained 54% of its initial fluorescence intensity while the fluorescence of MitoTracker RedFM dropped below 15% (Fig. 5a). Moreover, OC1 showed high photostability even at 72 h post-staining (Fig. 5b). Finally, it was noted that throughout the 72 h of incubation of OC1 with MCF-7, over three generations occurred in the cell culture and showed fluorescent labeling of the mitochondria suggesting that OC1 can be passed through to daughter cells upon cell division.Open in a separate windowFig. 5Intracellular retention and photostability of OC1. (a) Photostability of OC1 in MCF-7 cells co incubated with MitoTracker RedFM for 48 h. Plot of the sum intensity of each frame normalized to time 0 (frame 1) and plotted against time (seconds). Intensity data analysis was done using Python and plotted on Excel. (b) CLSM images (72 h post incubation) showing OC1 distribution while most of the MitoTracker RedFM signal was lost (Scale bar at 30 μm).     

Supramolecular encapsulation of redox-active monomers to enable free-radical polymerisation

Extended polymeric structures based on redox-active species are of great interest in emerging technologies related to energy conversion and storage. However, redox-active monomers tend to inhibit radical polymerisation processes and hence, increase polydispersity and reduce the average molecular weight of the resultant polymers. Here, we demonstrate that styrenic viologens, which do not undergo radical polymerisation effectively on their own, can be readily copolymerised in the presence of cucurbit[n]uril (CB[n]) macrocycles. The presented strategy relies on pre-encapsulation of the viologen monomers within the molecular cavities of the CB[n] macrocycle. Upon polymerisation, the molecular weight of the resultant polymer was found to be an order of magnitude higher and the polydispersity reduced 5-fold. The mechanism responsible for this enhancement was unveiled through comprehensive spectroscopic and electrochemical studies. A combination of solubilisation/stabilisation of reduced viologen species as well as protection of the parent viologens against reduction gives rise to the higher molar masses and reduced polydispersities. The presented study highlights the potential of CB[n]-based host–guest chemistry to control both the redox behavior of monomers as well as the kinetics of their radical polymerisation, which will open up new opportunities across myriad fields.

Extended polymeric structures based on redox-active species are of great interest in emerging technologies related to energy conversion and storage.

Polyviologens are redox-active polymers based on N-substituted bipyridinium derivatives which have emerged as promising materials for energy conversion and storage.^1–5 Their physicochemical properties can be adjusted through copolymerisation of the redox-active viologen monomers.^6–8 The resultant materials are stable, water soluble and exhibit fast electron transfer kinetics. Polyviologens have been commonly fabricated through step-growth polymerisation in linear and dendritic architectures,^9–13 as supramolecular polymers,^14–16 networks,^6,17,18 and covalent organic frameworks.^19,20 Alternatively, anionic/cationic or metathesis-based polymerisations are used to avoid interference of radical-stabilising monomers with the radical initiators, however, these techniques are highly water- and/or oxygen-sensitive.^21,22 When free-radical polymerisation (FRP) is conducted in the presence of viologen species, its reduction can cause a depletion of active radicals and thus disruption of the polymerisation process. Despite varying solvents, comonomers and initiator loadings, the direct FRP of viologen-containing monomers remains therefore limited to molar masses of 30 kDa.^23–25 Accessing higher molar masses has been possible via post-polymerisation modification,^26–28 which has impacted the electrochemical properties of the resultant materials.^29,30 Alternative strategies to access higher molar masses of redox-active polymers and control their polymerisation are highly desirable.Incorporation of cucurbit[n]uril (CB[n]) macrocycles have lead to a variety of functional materials through host–guest chemistry.^31–34 Moreover, the redox chemistry of viologens can be modulated through complexation with CB[n].^35–38 Specifically, CB[n] (n = 7, 8) can tune the redox potential of pristine viologens and efficiently sequester monoreduced viologen radical cations, avoiding precipitation in aqueous environments. Further to this, we recently demonstrated that the viologen radical cation is stabilised by −20 kcal mol⁻¹ when encapsulated in CB[7].³⁹Consequently, we envisioned that incorporating CB[n]s as additives prior to polymerisation could (i) overcome current limits in accessible molar masses, (ii) increase control over FRP of viologen-based monomers through encapsulation and (iii) enable separation of radical species avoiding aggregation.Here, we demonstrate a new approach to control FRP of redox-active monomers leading to high molar masses and decreased dispersity of the resultant polymers. In absence of CB[n], co-polymerisation of the N-styryl-N′-phenyl viologen monomer 1²⁺ and N,N-dimethylacrylamide (DMAAm) only occurs at high initiator loadings (>0.5 mol%, Fig. 1a), leading to low molecular weights and high polydispersity. Using our synthetic approach, 1²⁺ is efficiently copolymerised with DMAAm in the presence of CB[n] (n = 7, 8) macrocycles resulting in control of the polymer molar mass across a broad range, 4–500 kDa (Fig. 1b). Finally, CB[n] are successfully removed from the polymer via competitive host–guest binding and dialysis. Spectroscopic and electrochemical studies revealed that solubilisation/stabilisation of the reduced species and/or shielding of the redox-active monomers from electron transfer processes was responsible for this enhancement.Open in a separate windowFig. 1Schematic representation of the investigated polymerisation. (a) Conventional free radical polymerisation either completely fails to copolymerise redox-active monomers (low initiator loading) or delivers copolymers with limited molar masses and high dispersities (high initiator loading). (b) CB[n]-mediated protection suppresses interference of viologen monomers with radicals formed through the initiation process facilitating copolymerisation. The molar mass of the resulting copolymers is readily tunable via the amount of present CB[n] macrocycles and the CB[n] is post-synthetically removed via competitive binding to yield the final copolymer with desired molar mass. Cl⁻ counter-ions are omitted for clarity.Recent studies on symmetric aryl viologens demonstrated 2 : 2 binding modes with CB[8] and high binding constants (up to K_a ∼ 10¹¹ M⁻²).^40,41 Incorporation of polymerisable vinyl moieties, in combination with the relatively static structure of their CB[n] host–guest complexes, was postulated to allow polymerisation without unfavorable side reactions. The asymmetric N-styryl-N′-phenyl viologen monomer 1²⁺ prepared for this study (Fig. S1a and S2–S13^†) displays a linear geometry and was predicted to bind CB[n] (n = 7, 8) in a 2 : 1 and 2 : 2 binding fashion (Fig. S1b^†).^40,42 Binding modes between CB[n] (n = 7, 8) and 1²⁺ were investigated through titration experiments (¹H NMR and ITC) which confirmed the formation of 1·(CB[7])₂ and (1)₂·(CB[8])₂ (see Fig. S25 and S26^†). ¹H NMR titration of CB[7] with 1²⁺ demonstrates encapsulation of both aryl moieties (including the vinyl group) through upfield chemical shifts of the respective signals (Fig. 2a). Similar upfield shifts were observed for CB[8] (Fig. 2c). Different para-aryl substituents (vinyl vs. hydrogen) resulted in either head-to-tail or head-to-head (1)₂·(CB[8])₂ dimers (Fig. S1b and S26^†), a previously reported phenomenon.⁴³ Nonetheless, the reversible nature of the complex renders the vinyl group temporarily available for copolymerisation. In the presence of CB[8], 1²⁺ yields polymer molar masses of up to 500 kDa as its complexation is more robust. ITC data confirmed binding stoichiometry, with binding constants of K_a = 2.64 × 10⁶ M⁻¹ for 1·(CB[7])₂ and K_a = 9.02 × 10¹⁰ M⁻² for (1)₂·(CB[8])₂ (Table S2, Fig. S29a and b^†).Open in a separate windowFig. 2Supramolecular complexation of 1²⁺ and CB[n]. ¹H NMR spectra of 1²⁺ at (a) χ_CB[7] = 2, (b) χ_CB[n] = 0 and (c) χ_CB[8] = 1 in D₂O. Cl⁻ counter-ions are omitted for clarity.The free radical copolymerisation of 1²⁺ and DMAAm ([M] = 2 M), in the absence of CB[n], was based on optimised DMAAm homopolymerisations (Fig. S14 and S15^†) and full conversion was confirmed by ¹H NMR spectroscopy (Table S1 and Fig. S16^†). 1²⁺ was maintained at 1 mol% relative to DMAAm and by varying the radical initiator concentration molar masses of up to 30 kDa with broad dispersities (Đ = 11.4) were obtained (Fig. S17^†). Lower initiator concentrations (<0.25 mol%) limited polymerisation (M_n = 3.7 kDa) and size exclusion chromatography elution peaks exhibited extensive tailing, suggesting that 1²⁺ engages in radical transfer processes.To verify our hypothesis that CB[n] macrocycles can modulate the redox behavior of 1²⁺, FRP of 1²⁺ and DMAAm was conducted with varying amounts of CB[n] (n = 7, 8) (Fig. 3, S18 and S20^†). Full conversion of all monomers including their successful incorporation into the polymer was verified via¹H NMR spectroscopy and SEC (Fig. S18 and S21–S23^†). Using CB[7], the molar mass of the copolymers was tunable between M_n = 3.7–160 kDa (Fig. 3b and S21a^†). Importantly, in the presence of CB[8], a broad range of molar masses M_n = 3.7–500 kDa were accessible for 0 < χ_CB[8] < 1.2 (Fig. S20 and S21b^†). Increasing the CB[n] (n = 7, 8) concentration caused dispersity values to converge to Đ = 2.2 (χ_CB[8] = 1.2, χ is the ratio of CB[n] to the redox-active monomer, Fig. S20^†). The copolymers were purified by addition of adamantylamine (competitive binder) prior to dialysis to deliver CB[n]-free redox-active copolymers (Fig. S23^†).Open in a separate windowFig. 3(a) In situ copolymerisation of DMAAm with 1²⁺ and CB[7]. (b) Molar mass and dispersity vs. amount of CB[7] in the system. Fitted curve is drawn to guide the eye. Cl⁻ counter-ions are omitted for clarity.The range of molar masses obtainable through addition of CB[n] (n = 7, 8) correlated with the measured K_a (Fig. 3b and S20^†). Binding of 1²⁺ to CB[8] was stronger and therefore lower concentrations of CB[8] were required to shift the binding equilibrium and mitigate disruption of the polymerisation. Dispersity values reached a maximum at χ_CB[7] = 0.6 or χ_CB[8] = 0.3, suggesting 1⁺˙ is only partially encapsulated. Consequently, higher CB[n] concentrations can enable FRP with lower initiator concentrations (0.10 mol%, Fig. S19^†), which demonstrates the major role of complexation to modulate electron accepting properties of 1²⁺.The redox-active monomer 1²⁺ can engage with propagating primary radicals (P_m˙) to either be incorporated into the growing polymer chain (P_m–1²⁺˙) or to abstract an electron deactivating it (P_m). This deactivation likely occurs through oxidative termination producing 1⁺˙ (energetic sink), inactive oligo- and/or polymer chains (P_m) and a proton H⁺, causing retardation of the overall polymerisation. Oxidative terminations have been previously observed in aqueous polymerisations of methyl methacrylate, styrenes and acrylonitriles that make use of redox initiator systems.^44–47 Another example by Das et al. investigated the use of methylene blue as a retarder, with the primary radical being transferred to a methylene blue electron acceptor via oxidative termination, altogether supporting the outlined mechanism of our system (extended discussion see ESI, Section 1.4^†).⁴⁸The process of retardation can, however, be successfully suppressed, when monomer 1²⁺ is encapsulated within CB[n] macrocycles. Herein the formation of 1·(CB[7])₂ or (1)₂·(CB[8])₂ results in shielding of the redox-active component of 1²⁺ from other radicals within the system, hampering other electron transfer reactions. This inhibits termination and results in extended polymerisation processes leading to higher molar mass polymers through mitigation of radical transfer reactions. Moreover, suppressing the formation of 1⁺˙ through supramolecular encapsulation minimises both π and σ dimerisation of the emerging viologen radical species,³⁹ preventing any further reactions that could impact the molar mass or polydispersity of the resulting polymers.Cyclic voltammetry (CV) and UV-Vis titration experiments were conducted to provide insight into the impact of CB[n] on the redox behavior and control over FRP of 1²⁺. Excess of CB[n] (n = 7, 8) towards 1²⁺ resulted in a complete suppression of electron transfer processes (Fig. S31 and S32^†). Initially, 1²⁺ shows a quasi-reversible reduction wave at −0.44 V forming 1⁺˙ (Fig. 4a). Increasing χ_CB[7], this reduction peak decreases and shifts towards more negative potentials (−0.51 V, χ_CB[7] = 1) accompanied by the formation of 1²⁺·(CB[7])₁. A second cathodic peak emerges at −0.75 V due to the increased formation of 1²⁺·(CB[7])₂. At χ_CB[7] = 2, this peak shifts to −0.80 V, where it reaches maximum intensity, once 1²⁺·(CB[7])₂ is the dominating species in solution. When 2 < χ_CB[7] < 4, the intensity of the reduction peak decreases and the complexation equilibrium is shifted towards the bound state, complete suppression of the reduction peak occurs at χ_CB[7] = 4. Similarly, the oxidation wave intensity is reduced by 95% at χ_CB[7] = 4 causing suppression of potential oxidative radical transfer processes (Fig. 4c).Open in a separate windowFig. 4Mechanism of the CB[n]-mediated (n = 7, 8) strategy for the controlled copolymerisation of redox-active monomer 1²⁺. (a) Cyclic voltammogram with varying amounts of CB[7]. (b) UV-Vis titration of 1²⁺ with varying amounts of CB[7]. (c) Intensity decay of the oxidation peak at −0.27 V and change in absorption maximum of 1⁺˙ at 590 nm vs. χ_CB[7]. (d) Electron transfer processes of 1²⁺ to generate 1⁺˙ and 1⁰. (e) Reduction of 1²⁺ resulting in precipitation of 1⁰. (f) Stabilisation of 1⁺˙ through encapsulation with CB[7]. (g) Protection of 1²⁺ from redox processes through CB[7]-mediated encapsulation.The concentration of 1⁺˙ can be monitored using UV-Vis (Fig. 4b and S34^†).⁴⁹ Absorbance at 590 nm (λ_max) vs. χ_CB[7] was plotted and the concentration of 1⁺˙ increases, reaching a maximum at χ_CB[7] = 4 (Fig. 4c). When χ_CB[7] > 4, a decrease in concentration of 1⁺˙ was observed. We postulate the following mechanism: at χ_CB[7] = 0, 1²⁺ is reduced to produce high concentrations of 1⁺˙ that partially disproportionates to form 1⁰, which precipitates (Fig. 4e and S34^†). When 0 < χ_CB[7] < 4, increasing amounts of green 1⁺˙ are stabilised through encapsulation within CB[7] suppressing disproportionation (Fig. 4c (cuvette pictures), Fig. 4f). For χ_CB[7] > 4, 1²⁺ is protected from reduction through encapsulation (Fig. 4g).To further demonstrate applicability of this strategy, we chose another viologen-based monomer 2²⁺ for copolymerisation (Fig. 5a). As opposed to 1²⁺, CB binds predominantly to the styryl moiety of 2²⁺ (Fig. S27 and S28^†).⁵⁰ ITC data showed that 2²⁺ binds CB[7] in a 1 : 1 fashion with a binding affinity of K_a = 2.32 × 10⁶ M⁻¹ (Fig. S30 and Table S2^†). Monomer 2²⁺ was also analysed via CV and showed three reversible reduction waves at −0.91 V, −0.61 V (viologen) and 0.40 V (styrene). Similar to 1²⁺, excess CB[7] selectively protects the molecule from redox processes, while the vinyl moiety remains accessible (Fig. 5a, S33c and d^†). For CB[8], only partial suppression of electron transfer processes was observed (Fig. S33e and f^†). We therefore chose CB[7] as an additive to increase control over FRP of 2²⁺ (Fig. 5b). Copolymerisation of 2²⁺ (1 mol%) and DMAAm ([M] = 2 M) at χ_CB[7] = 0 resulted in M_n = 28 kDa. When χ_CB[7] = 0.1, 0.2 or 0.3, M_n increased gradually from 124 to 230 and 313 kDa, respectively, demonstrating the potential of this strategy for FRP of redox-active monomers. Higher percentages of CB[7] led to copolymers with presumably higher molar masses causing a drastic decrease in solubility that prevented further analysis. Investigations on a broader spectrum of such copolymers, including those with higher contents of 2²⁺ are currently ongoing.Open in a separate windowFig. 5(a) Cyclic voltammogram of viologen-containing monomer 2²⁺ and its complexation with CB[n] (n = 7, 8) at a concentration of 1 mM using a scan rate of 10 mV s⁻¹ in 0.1 mM NaCl solution. (b) Molar mass and dispersity of 2²⁺-containing copolymers vs. equivalents of CB[7]. Cl⁻ counter-ions are omitted for clarity.In conclusion, we report a supramolecular strategy to induce control over the free radical polymerisation of redox-active building blocks, unlocking high molar masses and reducing polydispersity of the resulting polymers. Through the use of CB[n] macrocycles (n = 7, 8) for the copolymerisation of styrenic viologen 1²⁺, a broad range of molar masses between 3.7–500 kDa becomes accessible. Our mechanistic investigations elucidated that the redox behavior of monomer 1²⁺ is dominated by either CB[n]-mediated stabilisation of monoradical cationic species or protection of the encapsulated pyridinium species from reduction. In the stabilisation regime (χ_CB[7] < 4), 1²⁺ is reduced to form the radical cation 1⁺˙, which is subsequently stabilised through CB[7] encapsulation. Upon reaching a critical concentration of CB[7] (χ_CB[7] > 4), the system enters a protection-dominated regime, where reduction of 1²⁺ is suppressed and the concentration of 1⁺˙ diminishes. The resulting copolymers can be purified by use of a competitive binder to remove CB[n] macrocycles from the product. This strategy was successfully translated to a structurally different redox-active monomer that suffered similar limitations. We believe that the reported strategy of copolymerisation of redox-active monomers will open new avenues in the synthesis of functional materials for energy conversion and storage as well as for applications in electrochromic devices and (nano)electronics.