Investigation of the stability of thiosialosides toward hydrolysis by sialidases using NMR spectroscopy

[formula: see text] 1H NMR spectroscopy has been used to investigate whether the alpha(2-->6)-linked thiosialoside 3 and the alpha(2-->3)-linked thiosialoside 9 are hydrolyzed in the presence of Vibrio cholerae sialidase. Similarly, the hydrolysis of the O-ketosides Neu5Ac-2-O-alpha-(2-->3)-Gal beta Me (4) and the alpha-(2-->6)-sialyllactoside 7, representing natural alpha(2-->3)- and alpha(2-->6)-linked sialosides, respectively, was investigated. The results of the 1H NMR experiments clearly demonstrate that the thiosialosides are not hydrolyzed by Vibrio cholerae sialidase. As expected, the O-sialosides are hydrolyzed to give N-acetyl-alpha-D-neuraminic acid as the first product of substrate cleavage.     

Molecular Modification of Single Cobalt Sites Boosts the Catalytic Activity of CO2 Electroreduction into CO

Electroreduction of CO₂ into carbonaceous fuels or industrial chemicals using renewable energy sources is an ideal way to promote global carbon recycling. Thus, it is of great importance to develop highly selective, efficient, and stable catalysts. Herein, we prepared cobalt single atoms (Co SAs) coordinated with phthalocyanine (Co SAs-Pc). The anchoring of phthalocyanine with Co sites enabled electron transfer from Co sites to CO₂ effectively via the π-conjugated system, resulting in high catalytic performance of CO₂ electroreduction into CO. During the process of CO₂ electroreduction, the Faradaic efficiency (FE) of Co SAs-Pc for CO was as high as 94.8 %. Meanwhile, the partial current density of Co SAs-Pc for CO was −11.3 mA cm⁻² at −0.8 V versus the reversible hydrogen electrode (vs RHE), 18.83 and 2.86 times greater than those of Co SAs (−0.60 mA cm⁻²) and commercial Co phthalocyanine (−3.95 mA cm⁻²), respectively. In an H-cell system operating at −0.8 V vs RHE over 10 h, the current density and FE for CO of Co SAs-Pc dropped by 3.2 % and 2.5 %. A mechanistic study revealed that the promoted catalytic performance of Co SAs-Pc could be attributed to the accelerated reaction kinetics and facilitated CO₂ activation.     

A self-assembly pipette tip graphene solid-phase extraction coupled with liquid chromatography for the determination of three sulfonamides in environmental water

A sensitive, economical, and miniaturized self-assembly pipette tip graphene solid-phase extraction (PT-G-SPE) coupled with liquid chromatography fluorescence detection (LC-FD) was developed for rapid extraction and determination of three sulfonamide antibiotics (SAs) in environmental water samples. The PT-G-SPE cartridge, assembled by packing 1.0 mg of graphene as sorbent into a 100 μL pipette tip, showed high adsorption capacity for the SAs owing to the large surface area and unique structure of graphene. The factors that affected the extraction efficiency of PT-G-SPE, including sample volume, pH, sorbent amount, washing solvent and eluent solvent were optimized. Good linearity for SAs was obtained in a range of 2–4000 pg mL⁻¹ with correlation coefficients (r²) ≥ 0.9993. The recoveries of the SAs at three spiked levels ranged from 90.4% to 108.2% with relative standard deviations (RSD) ≤ 6.3%. In comparison with other sorbents such as C₁₈, HLB, SCX, PCX, and multiwalled carbon nanotubes, one advantage of using graphene as sorbent of pipette tip solid-phase extraction (PT-SPE) was that PT-G-SPE could adsorb larger sample volume (10 mL) at a small amount of sorbent (1 mg) and low solvent consumption with good extraction efficiency, which not only increased the fraction of analytes to LC and the sensitivity of SAs determination, but also reduced the cost and pollution.     

Convenient Synthesis and Purification of N-Trifluoroacetylated Neuraminic Acid Tetraol: A Key Precursor for the Synthesis of New Sialyl Donors Containing Labile O-Acyl Protecting Groups

A convenient synthesis and separation of α- and β-anomers of methyl (phenyl 3,5-dideoxy-2-thio-5-trifluoroacetamido-D-glycero-D-galacto-nonulopyranosid)onate (6a and 6b) on a multigram scale was developed. Both α- and β-isomers of 6 were obtained as crystals suitable for safe storage. The β-isomer forms a crystalline solvate with methanol. Fully O-trichloroacetylated and O-trifluoroacetylated N-trifluoroacetyl thiosialosides were synthesized in an efficient manner from the β-tetraol 6b.     

A Bimetallic Zn/Fe Polyphthalocyanine‐Derived Single‐Atom Fe‐N4 Catalytic Site:A Superior Trifunctional Catalyst for Overall Water Splitting and Zn–Air Batteries

Developing an efficient single‐atom material (SAM) synthesis and exploring the energy‐related catalytic reaction are important but still challenging. A polymerization–pyrolysis–evaporation (PPE) strategy was developed to synthesize N‐doped porous carbon (NPC) with anchored atomically dispersed Fe‐N₄ catalytic sites. This material was derived from predesigned bimetallic Zn/Fe polyphthalocyanine. Experiments and calculations demonstrate the formed Fe‐N₄ site exhibits superior trifunctional electrocatalytic performance for oxygen reduction, oxygen evolution, and hydrogen evolution reactions. In overall water splitting and rechargeable Zn–air battery devices containing the Fe‐N₄ SAs/NPC catalyst, it exhibits high efficiency and extraordinary stability. This current PPE method is a general strategy for preparing M SAs/NPC (M=Co, Ni, Mn), bringing new perspectives for designing various SAMs for catalytic application.     

The differential modulation of USP activity by internal regulatory domains, interactors and eight ubiquitin chain types

Ubiquitin-specific proteases (USPs) are papain-like isopeptidases with variable inter- and intramolecular regulatory domains. To understand the effect of these domains on USP activity, we have analyzed the enzyme kinetics of 12 USPs in the presence and absence of modulators using synthetic reagents. This revealed variations of several orders of magnitude in both the catalytic turnover (k_cat) and ubiquitin (Ub) binding (K_M) between USPs. Further activity modulation by intramolecular domains affects both the k_cat and K_M, whereas the intermolecular activators UAF1 and GMPS mainly increase the k_cat. Also, we provide the first comprehensive analysis comparing Ub chain preference. USPs can hydrolyze all linkages and show modest Ub-chain preferences, although some show a lack of activity toward linear di-Ub. This comprehensive kinetic analysis highlights the variability within the USP family.     

Surfactant Hydrogels for the Dispersion of Carbon‐Nanotube‐Based Catalysts

Novel hydrogel phases based on positively charged and zwitterionic surfactants, namely, N‐[p‐(n‐dodecyloxybenzyl)]‐N,N,N‐trimethylammonium bromide (pDOTABr) and p‐dodecyloxybenzyldimethylamine oxide (pDOAO), which combine pristine carbon nanotubes (CNTs), were obtained, thus leading to stable dispersions and enhanced cross‐linked networks. The composite hydrogel featuring a well‐defined nanostructured morphology and an overall positively charged surface was shown to efficiently immobilise a polyanionic and redox‐active tetraruthenium‐substituted polyoxometalate (Ru₄POM) by complementary charge interactions. The resulting hybrid gel has been characterised by electron microscopy techniques, whereas the electrostatic‐directed assembly has been monitored by means of fluorescence spectroscopy and ζ‐potential tests. This protocol offers a straightforward supramolecular strategy for the design of novel aqueous‐based electrocatalytic soft materials, thereby improving the processability of CNTs while tuning their interfacial decoration with multiple catalytic domains. Electrochemical evidence confirms that the activity of the catalyst is preserved within the gel media.     

Desymmetrization of 1,4‐Pentadien‐3‐ol by the Asymmetric 1,3‐Dipolar Cycloaddition of Azomethine Imines

Desymmetrization of the divinyl carbinol 1,4‐pentadien‐3‐ol was accomplished by the asymmetric 1,3‐dipolar cycloaddition of azomethine imines based on a magnesium‐mediated, multinucleating chiral reaction system utilizing diisopropyl (R,R)‐tartrate as the chiral auxiliary. The corresponding optically active trans‐pyrazolidines, each with three contiguous stereogenic centers, were obtained with excellent regio‐, diastereo‐, and enantioselectivity, with results as high as 99 % ee. This reaction was shown to be applicable to both aryl‐ and alkyl‐substituted azomethine imines. The use of a catalytic amount of diisopropyl (R,R)‐tartrate was also effective when accompanied by the addition of MgBr₂.     

Monitoring of sulfonamide antibacterial residues in milk and egg by polymer monolith microextraction coupled to hydrophilic interaction chromatography/mass spectrometry

A simple, rapid, and sensitive method for the determination of traces of thirteen sulfonamide antibacterials in milk and eggs is presented. This method is based on the combination of polymer monolith microextraction (PMME) technique with hydrophilic interaction chromatography/mass spectrometry (HILIC/MS). The extraction was performed with a poly(methacrylic acid-ethylene glycol dimethacrylate) monolithic capillary column while the subsequent separation was carried out on a Luna NH₂ column by HILIC. To obtain optimum results, several parameters relating to HILIC and PMME were investigated. After optimization, acetonitrile (contain 0.05% formic acid, v/v) was used as the elution solution, which was well compatible with the mobile phase in HILIC. Good linearities were obtained for thirteen SAs with the correlation coefficients (R²) above 0.997. The limits of detection (S/N = 3) of the method were found to be 0.4–5.7 ng mL⁻¹ of SAs in whole milk and 0.9–9.8 ng g⁻¹ of SAs in eggs. The recoveries of thirteen SAs in two matrices ranged from 80.4 to 119.8%, with relative standard deviations less than 11.8%.     

Development of a New Class of Inhibitors for the Malarial Aspartic Protease Plasmepsin II Based on a Central 7‐Azabicyclo[2.2.1]heptane Scaffold

Plasmepsin II (PMII), a malarial aspartic protease involved in the catabolism of hemoglobin in parasites of the genus Plasmodium, and renin, a human aspartic protease, share 35% sequence identity in their mature chains. Structures of 4‐arylpiperidine inhibitors complexed to human renin were reported by Roche recently. The major conformational changes, compared to a structure of renin, with a peptidomimetic inhibitor were identified and subsequently modeled in a structure of PMII (Fig. 1). This distorted structure of PMII served as active‐site model for a novel class of PMII inhibitors, according to a structure‐based de novo design approach (Fig. 2). These newly designed inhibitors feature a rigid 7‐azabicyclo[2.2.1]heptane scaffold, which, in its protonated form, is assumed to undergo ionic H‐bonding with the two catalytic Asp residues at the active site of PMII. Two substituents depart from the scaffold for occupancy of either the S1/S3 or S2′‐pocket and the hydrophobic flap pocket, newly created by the conformational changes in PMII. The inhibitors synthesized starting from N‐Boc‐protected 7‐azabicyclo[2.2.1]hept‐2‐ene ( 6 ; Schemes 1–5) displayed up to single‐digit micromolar activity (IC₅₀ values) toward PMII and good selectivity towards renin. The clear structure? activity relationship (SAR; Table) provides strong validation of the proposed conformational changes in PMII and the occupancy of the resulting hydrophobic flap pocket by our new inhibitors.     

Spatially Heterogeneous Nature of Self‐Catalytic Reaction in Hetero‐Double Helix Formation of Helicene Oligomers

A 1:1 mixture of pseudoenantiomeric aminomethylenehelicene oligomers, (P)‐tetramer and (M)‐pentamer, in fluorobenzene show a self‐catalytic phenomenon in the formation of hetero‐double helices from random coils. This study visualizes the spatially heterogeneous nature of the self‐catalytic reaction in dilute solution. UV/Vis imaging analysis of the mixture at 70 °C, containing random coils, exhibits a homogeneous bright area. When the solution is cooled from 70 to 30 °C and held at that temperature, dark domains of approximately 1 mm in size appear, which move approximately at a rate of 1 mm min^?1. The dark domains indicate that weaker UV/Vis absorption results from the formation of hetero‐double helices, which is supported by circular dichroism (CD) imaging experiments. Then a homogeneous mixture is regenerated upon heating to 55 °C, as shown by CD imaging. Under self‐catalytic conditions, a homogeneous solution spontaneously changed to a heterogeneous solution in the process of hetero‐double‐helix formation.     

Cytochrome c embraced in sodium dodecyl sulfate nano-micelle as a homogeneous nanostructured peroxidase

A homogeneous nanostructured enzyme (artificial peroxidase, AP) with suitable catalytic efficiency was generated using bovine heart cytochrome c (Cyt c) and sodium dodecyl sulfate nano-micelles in 50?mM phosphate buffer pH 10.5 at 25?°C. The Michaelis?CMenten (K _m) and catalytic rate (k _cat) of the AP were determined to be 21.6?±?1.2???M and 0.474?±?0.013?s^?1, respectively. The catalytic efficiency of the AP was 0.0219?±?0.002???M^?1s^?1, which was 30?±?1.5?% as efficient as the native horseradish peroxidase (HRP). The mean diameter of AP was measured to be 6.4?nm using dynamic light scattering technique. The UV?CVis spectrometry, circular dichroism, surface tension, isothermal titration calorimetric and electrochemistry methods were utilized for additional characterization of the AP. Together our results suggest that the AP generated here can be used in place of HRP in industrial and commercial fields under some extreme conditions.     

Development of Bio-inspired Composite Materials for the Detection of Traces of Silver Present in Water: Use of Taguchi Methodology to Design Low-cost Carbon Paste Electrodes

Carbon Paste Electrodes (CPE) modified with Grapefruit-Peels (GP) functionalized with Urea (GPU) and, Melamine (GPM) were designed for the detection of Ag⁺ in water. Taguchi L₉ methodology was used to determine the optimal graphite-Active material ratio. The best electrochemical response was for CPE-GPU with an 80 : 20 ratio. The results obtained showed a linear detection range between 0.5 to 28 μg L⁻¹, with a detection limit of 0.73 μg L⁻¹ and a limit of quantification of 1.04 μg L⁻¹. Attributable to CPE-GPU characteristics: electroactive surface area 0.175 cm², roughness factor 3.87, resistance 0.09 Ω and, mostly −NH₂ groups on its surface. The composite material offers a viable option to be used for the determination of silver traces in situ in industrial processes.     

Structure‐Based Design of Nonpeptidic Thrombin Inhibitors: Exploring the D‐Pocket and the Oxyanion Hole

Structure‐activity relationships for new members of a class of nonpeptidic, low‐molecular‐weight inhibitors of thrombin, a key serine protease in the blood coagulation cascade, are described. These compounds, which originate from X‐ray‐structure‐based design, feature a conformationally rigid, bi‐ or tricyclic core from which side chains diverge into the four major binding pockets (distal D, proximal P, recognition or specificity S1, and oxyanion hole O) at the thrombin active site (Fig. 1). Phenylamidinium is the side chain of choice for the S1 pocket, while the most active inhibitors orient an i‐Pr group into the P‐pocket (Table 1). The key step in the synthesis of the inhibitors is the construction of the central bi‐ or tricyclic scaffold by 1,3‐dipolar cycloaddition of an in situ prepared azomethine ylide and an N‐substituted maleimide (Schemes 1–3, and 8–10). One series of compounds was designed to explore the binding features of the large hydrophobic D pocket. This pocket provides space for lipophilic residues as bulky as benzhydryl groups. A new strategy was developed, allowing introduction of these sterically demanding substituents very late in the synthesis (Schemes 5 and 6). Benzhydryl derivative (±)‐ 2 was found to be the most selective member (K_i (trypsin)/K_i (thrombin)=1200) of this class of nonpeptidic thrombin inhibitors, while the ‘dipiperonyl' analog (±)‐ 3 (K_i=9 nM , 7.60‐fold selectivity) displays the highest potency of all compounds prepared so far (Table 1). A second series of inhibitors features side chains designed to orient into the oxyanion hole and to undergo H‐bonding with the backbone NH groups lining the catalytic site of the enzyme. Unfortunately, neither activity nor selectivity could be substantially improved by introduction of these substituents (Table 2). Presumably, the high degree of pre‐organization and the rigidity of the tightly bound scaffolds prevents the new substituents from assuming a position that would allow favorable interactions in the oxyanion hole. However, the oxyanion hole and the S1′ pocket next to it were found to be capable of accommodating quite large groups, which leaves much room for further exploration.     

On-line coupling of dynamic microwave-assisted extraction to solid-phase extraction for the determination of sulfonamide antibiotics in soil

A rapid technique based on dynamic microwave-assisted extraction (DMAE) coupled on-line with solid-phase extraction (SPE) was developed for the determination of sulfonamides (SAs) including sulfadiazine, sulfameter, sulfamonomethoxine and sulfaquinoxaline in soil. The SAs were first extracted with acetonitrile under the action of microwave energy, and then directly introduced into the SPE column which was packed with neutral alumina for preconcentration of analytes and clean-up of sample matrix. Subsequently, the SAs trapped on the alumina were eluted with 0.3% acetic acid aqueous solution and determined by liquid chromatography-tandem mass spectrometry. The DMAE parameters were optimized by the Box-Behnken design. Maximum extraction efficiency was achieved using 320 W of microwave power; 12 mL of extraction solvent and 0.8 mL min⁻¹ of extraction solvent flow rate. The limits of detection and quantification obtained are in the range of 1.4-4.8 ng g⁻¹ and 4.6-16.0 ng g⁻¹ for the SAs, respectively. The mean values of relative standard deviation of intra- and inter-day ranging from 2.7% to 5.3% and from 5.6% to 6.7% are obtained, respectively. The recoveries of SAs obtained by analyzing four spiked soil samples at three fortified levels (20 ng g⁻¹, 100 ng g⁻¹ and 500 ng g⁻¹) were from 82.6 ± 6.0% to 93.7 ± 5.5%. The effect of standing time of spiked soil sample on the SAs recoveries was examined. The recoveries of SAs decreased from (86.3-101.9)% to (37.6-47.5)% when the standing time changed from one day to four weeks.     

A SAR-based mechanistic study on the combined toxicities of sulfonamides and quorum sensing inhibitors on Escherichia coli

Quorum sensing inhibitors (QSIs) are promising alternatives to antibiotics, but they are discharged into the environment after their use cycle. This poses joint effects on the organisms in the environment. Therefore, it is of great importance to study the combined toxicities of QSIs and antibiotics. In this study, we investigated the single and combined toxicities of four potential QSIs and 11 sulfonamides (SAs) on Escherichia coli. The results revealed that the single toxicities of SAs were greater than those of QSIs, and the toxicities were found positively related to the binding energies (E_bind) with their target proteins, for both antibiotics and QSIs. The combined toxicities of the binary mixtures were observed to be either antagonism or addition. The antagonism could be explained by the phenomenon that QSIs changed SAs molecules into ionic forms, preventing the SA molecules entering the bacteria. Furthermore, it was found that the ratios of the effective concentration (the actual concentration involved in the interaction with the proteins) in the antagonistic cases were higher than those in the additive cases. This study would benefit both rational use of the drug combination and ecological risk assessment of antibiotics and QSIs in the real environment.     

DPAMPP in catalytic asymmetric reactions: enantioselective synthesis of l-homophenylalanine

l-Homophenylalanine 1, a key intermediate of most commercially important ACE inhibitors, was prepared via catalytic asymmetric hydrogenation of (Z)-2-acetamido-4-phenylcrotonate 3a with a cationic rhodium complex of (1R,2S)-DPAMPP, followed by acid-hydrolysis of the protecting groups in good yield with >99.9% enantioselectivity.     

Variation dynamics of palladium(II) and tin(II) concentrations in activating solution

A study of the possibility of optimizing the efficiency of a catalytic palladium layer demonstrated that the minimum palladium adsorption providing a sufficient catalytic activity of the surface is Γ_min = 3.178 × 10^–6 mol m^–2, which can be reached at a palladium dichloride concentration of 0.035 g L^–1. It was shown that palladium(II) activates the surface of the insulator (is sorbed) from a true, rather than a colloid solution.     

Synthesis of novel 1,3-oxazino[6,5-b]indole-2,4-(3H,9H)-dione and 2H-1,3,5-oxadiazino[3,2-a]indole-2,4-(3H)-dione from oxindoles

Two novel tricyclic ring systems were designed as serine protease inhibitors and synthesized from oxindoles; the first series, 1,3-oxazino[6,5-b]indole-2,4-(3H,9H)-diones, were prepared from 2,3-dihydro-2-oxo-(1H)-indole-3-carboxamides and phosgene in tetrahydrofuran with triethylamine. The second, a 2H-1,3,5-oxadiazino[3,2-a]indole-2,4-(3H)-dione was made using a similar cyclization on 2,3-dihydro-2-oxo-N-phenyl-(1H)-indole-1-carboxamide.     

Control Mechanism for cis Double‐Bond Formation by Polyunsaturated Fatty‐Acid Synthases

Polyunsaturated fatty acids (PUFAs) such as docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and arachidonic acid (ARA) are essential fatty acids for humans. Some microorganisms biosynthesize these PUFAs through PUFA synthases composed of four subunits with multiple catalytic domains. These PUFA synthases each create a specific PUFA without undesirable byproducts, even though the multiple catalytic domains in each large subunit are very similar. However, the detailed biosynthetic pathways and mechanisms for controlling final‐product profiles are still obscure. In this study, the FabA‐type dehydratase domain (DH_FabA) in the C‐subunit and the polyketide synthase‐type dehydratase domain (DH_PKS) in the B‐subunit of ARA synthase were revealed to be essential for ARA biosynthesis by in vivo gene exchange assays. Furthermore, in vitro analysis with truncated recombinant enzymes and C₄‐ to C₈‐acyl ACP substrates showed that ARA and EPA synthases utilized two types of DH domains, DH_PKS and DH_FabA, depending on the carbon‐chain length, to introduce either saturation or cis double bonds to growing acyl chains.