Mapping the Complete Photocycle that Powers a Large Stokes Shift Red Fluorescent Protein

Large Stokes shift (LSS) red fluorescent proteins (RFPs) are highly desirable for bioimaging advances. The RFP mKeima, with coexisting cis- and trans-isomers, holds significance as an archetypal system for LSS emission due to excited-state proton transfer (ESPT), yet the mechanisms remain elusive. We implemented femtosecond stimulated Raman spectroscopy (FSRS) and various time-resolved electronic spectroscopies, aided by quantum calculations, to dissect the cis- and trans-mKeima photocycle from ESPT, isomerization, to ground-state proton transfer in solution. This work manifests the power of FSRS with global analysis to resolve Raman fingerprints of intermediate states. Importantly, the deprotonated trans-isomer governs LSS emission at 620 nm, while the deprotonated cis-isomer's 520 nm emission is weak due to an ultrafast cis-to-trans isomerization. Complementary spectroscopic techniques as a table-top toolset are thus essential to study photochemistry in physiological environments.     

Molecular-Orbital Delocalization Enhances Charge Transfer in π-Conjugated Organic Semiconductors

π-Conjugated organic semiconductors are promising materials for surface-enhanced Raman scattering (SERS)-active substrates based on the tunability of electronic structures and molecular orbitals. Herein, we investigate the effect of the temperature-mediated resonance-structure transitions of poly(3,4-ethylenedioxythiophene) (PEDOT) in poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate) (PEDOT : PSS) films on the interactions between substrate and probe molecules, thereby affecting the SERS activity. Absorption spectroscopy and density functional theory calculations show that this effect occurs mainly due to delocalization of the electron distribution in molecular orbitals, effectively promoting the charge transfer between the semiconductor and probe molecules. In this work, we investigate for the first time the effect of electron delocalization in molecular orbitals on SERS activity, which will provide new design ideas for the development of highly sensitive SERS substrates.     

Achieving Molecular Recognition of Structural Analogues in Surface-Enhanced Raman Spectroscopy: Inducing Charge and Geometry Complementarity to Mimic Molecular Docking

Molecular recognition of complex isomeric biomolecules remains challenging in surface-enhanced Raman scattering (SERS) spectroscopy due to their small Raman cross-sections and/or poor surface affinities. To date, the use of molecular probes has achieved excellent molecular sensitivities but still suffers from poor spectral specificity. Here, we induce “charge and geometry complementarity” between probe and analyte as a key strategy to achieve high spectral specificity for effective SERS molecular recognition of structural analogues. We employ 4-mercaptopyridine (MPY) as the probe, and chondroitin sulfate (CS) disaccharides with isomeric sulfation patterns as our proof-of-concept study. Our experimental and in silico studies reveal that “charge and geometry complementarity” between MPY's binding pocket and the CS sulfation patterns drives the formation of site-specific, multidentate interactions at the respective CS isomerism sites, which “locks” each CS in its analogue-specific complex geometry, akin to molecular docking events. Leveraging the resultant spectral fingerprints, we achieve > 97 % classification accuracy for 4 CSs and 5 potential structural interferences, as well as attain multiplex CS quantification with < 3 % prediction error. These insights could enable practical SERS differentiation of biologically important isomers to meet the burgeoning demand for fast-responding applications across various fields such as biodiagnostics, food and environmental surveillance.     

Semiconductor SERS on Colourful Substrates with Fabry-Pérot Cavities

Non-metallic materials have emerged as a new family of active substrates for surface-enhanced Raman scattering (SERS), with unique advantages over their metal counterparts. However, owing to their inefficient interaction with the incident wavelength, the Raman enhancement achieved with non-metallic materials is considerably lower with respect to the metallic ones. Herein, we propose colourful semiconductor-based SERS substrates for the first time by utilizing a Fabry-Pérot cavity, which realize a large freedom in manipulating light. Owing to the delicate adjustment of the absorption in terms of both frequency and intensity, resonant absorption can be achieved with a variety of non-metal SERS substrates, with the sensitivity further enhanced by ≈100 times. As a typical example, by introducing a Fabry-Pérot-type substrate fabricated with SiO₂/Si, a rather low detection limit of 10⁻¹⁶ M for the SARS-CoV-2S protein is achieved on SnS₂. This study provides a realistic strategy for increasing SERS sensitivity when semiconductors are employed as SERS substrates.     

Expanding the Range of Bioorthogonal Tags for Multiplex Stimulated Raman Scattering Microscopy

Multiplex optical detection in live cells is challenging due to overlapping signals and poor signal-to-noise associated with some chemical reporters. To address this, the application of spectral phasor analysis to stimulated Raman scattering (SRS) microscopy for unmixing three bioorthogonal Raman probes within cells is reported. Triplex detection of a metallacarborane using the B−H stretch at 2480–2650 cm⁻¹, together with a bis-alkyne and deuterated fatty acid can be achieved within the cell-silent region of the Raman spectrum. When coupled to imaging in the high-wavenumber region of the cellular Raman spectrum, nine discrete regions of interest can be spectrally unmixed from the hyperspectral SRS dataset, demonstrating a new capability in the toolkit of multiplexed Raman imaging of live cells.     

Vibrational Properties of LaNb0.8M0.2O4-δ (M=As,Sb, V,and Ta)

LaNb_0.8M_0.2O_4-δ (where M=As, Sb, V, and Ta) oxides with pentavalent elements of different ionic sizes were synthesized by a solid-state reaction method. The vibrational properties of these oxides have been investigated. These studies revealed that the substituent element influences both Debye temperature value as well as the Raman active vibrational modes. Additionally, the low-temperature vibrational properties of LaNb_0.8Sb_0.2O_4-δ have been determined to show the phase transition occurrence at 260 K which is lower than previously reported.     

Separating Vibrational Coherences in Ground/Excited Electronic States of Solvent/Solute Following Non-Resonant/Resonant Impulsive Excitation

In a quest to track down the origin of coherent vibrational motions observed in femtosecond pump-probe transients, whether they arise from ground/excited electronic state of solute or are contributed by the solvent, we demonstrate a method for extricating vibrations under resonant and non-resonant impulsive excitations using a diatomic solute in condensed phase (iodine in carbon tetrachloride) with aid of spectral dispersion of the chirped broadband probe. Most importantly, we show how a sum over intensities for a select region of detection wavelengths and Fourier transform of data over select temporal window untwine contributions from vibrational modes of different origins. Thus, in a single pump-probe experiment, vibrational features specific to solute as well as solvent are disentangled that are otherwise spectrally overlapping and are non-separable in conventional (spontaneous/stimulated) Raman spectroscopy employing narrowband excitation. We envision wide-ranging applications of this method to unveil vibrational features in complex molecular systems.     

Activation-induced layered structure in NiCoAl by atomic modulation for energy storage application

The surface activation of alloys favors their electrochemical interactions, ion diffusivity, and the rapid kinetics of ions and electrons, leading to the formation of self-supported layered double hydroxides (LDHs) in them. However, the formation of LDHs at different depths in the alloy upon activation, their electronic/atomic structures, and their electrochemical charge storage mechanism, have not been thoroughly explored. Herein, Ni ion-substituted CoAl alloys are prepared by arc melting and activated by KOH electrolyte, which is responsible for the modulation of the atomic configuration as confirmed by XRD. Raman depth mapping demonstrates how the LDHs vary with depth upon activation and that the octahedral and tetrahedral symmetry sites of CoO and Co₃O₄ are responsible for the formation of the layered structures of CoOOH and Co(OH)₂, respectively. The activated Ni₁₀Co₈₅Al₅ has a superior volumetric capacitance of 4.15 F/cm³ at 0.5 mA/g, which is 38.6 times that of an unactivated one, and excellent cyclic stability up to 5000 cycles, and a voltage of 0.54 V generated from a fabricated supercapacitor cell. X-ray Absorption Spectroscopy (XAS) analysis indicates greater charge transfer by Co than by Ni and the modulation of the local atomic structures facilitates electrochemical charge storage in Ni₁₀Co₈₅Al₅. This work presents an easy route for the development of advanced LDHs, and the mechanism of electrochemical charge storage in them.     

Effect of non-covalent interactions in 2,5-di-Me-pyrazine-di-N-oxide - methanol – Carbon nanotube electrocatalytic system

Catalytic oxidation of methanol (MeOH) in the absence of noble metals and noble metal oxides as catalysts, and the use of metal-free materials are inexpensive and attractive process for practical use in electrocatalysis, sensors, and in direct methanol fuel cells. In previous works, it was found that the use of single-walled (SWCNT) or multi-walled (MWCNT) carbon nanotube paper electrodes instead of GC increases the catalytic efficiency of organic compounds oxidation in the presence of aromatic di-N-oxides by several times. In this work, the effect of non-covalent interactions on the catalytic efficiency of MeOH oxidation in the presence of 2,5-di-Me-pyrazine-di-N-oxide (Pyr₁) in 0.1 M Bu₄NClO₄ solution in acetonitrile at SWCNT and MWСNT paper electrodes was studied by the methods of quantum chemical modeling, Raman spectroscopy, and using electrochemical data. New factors determined the features of mechanism of MeOH oxidation on CNT electrodes and lead to an increase in the catalytic efficiency of the electrode process in comparison with the GC electrode were established.     

Development of bioorthogonal SERS imaging probe in biological and biomedical applications

Living-cell imaging demands high specificity,sensitivity,and minimal background interference to the targets of interest.However,developing a desirable imaging probe that can possess all the above features is still challenging.The bioorthogonal surface-enhanced Raman scattering(SERS) imaging has been recently emerged through utilizing Raman reporters with characteristic peaks in Raman-silent region of cells(1800-2800 cm~(-1)),which opens a revolutionary avenue for living-cell imaging with multiplexing capability.In this review,we focus on the recent advances in the technology development and the biological and biomedical applications of the living-cell bioorthogonal SERS imaging technique.After introduction of fundamental principles for bioorthogonal tag or label,we present applications for visualization of various intracellular components and environment including proteins,nucleic acids,lipids,pH and hypoxia,even for cancer diagnosis in tissue samples.Then,various bioorthogonal SERS imaging-guided thera py strategies have been discussed such as photothera py and surge ry.In conclusion,this strategy has great potential to be a flexible and robust tool for visualization detection and diseases diagnosis.