Divergent Regioselective Synthesis of Functionalized 1,2,3-1H-Triazoles from Nitriles and Arylazides under Metal- and/or Solvent-free Conditions.

Highly selective and divergent syntheses, which are crucial in both organic synthesis and medicinal chemistry, involve significant advancements in compound accessibility. By modifying α-cyano esters into α-cyano ketones, the synthesis pathway broadens to include a diverse range of 4-CN, 5-amino, and 5-arylamino derivatives of 1,2,3-triazoles, which are achieved notably through the Dimroth rearrangement. This versatility extends further with the potential for a triple cascade reaction, leading to the production of carboximidamide compounds, which are facilitated by the Cornforth rearrangement. Advancements in compound accessibility not only expand the repertoire of synthesized molecules but also open new avenues for potential pharmacological agents. Building on these findings, we have developed an innovative and efficient method for the divergent synthesis of functionalized 1,2,3-triazoles. This method strategically utilizes α-cyanocarbonyls and arylazides by harnessing their reactivity and compatibility to orchestrate a variety of molecular transformations. By optimizing these substrates, our goal is to simplify synthetic routes, improve product yields, and accelerate the discovery and development of new chemical entities with promising biological activities.

Universal, Time-Cost-Effective, and Inside-Out Strategy for Synthesis of Plant-Based Flow Catalysis Microreactors.

The utilization of plant-based flow catalytic microreactors has been increasingly gaining traction in the fields of water treatment, energy generation, and biotechnological science due to their inherent channel structures, renewable properties, and environmentally friendly nature. The conventional outside-in strategy for synthesizing plant-based monolithic microreactors typically entails prolonged hydrothermal modification, extensive chemical usage, or energy-intensive equipment. The present study presents a universal inside-out strategy for the rapid synthesis of monolithic catalytic microreactors derived from plant materials. This approach enables the direct formation of catalytic metal nanoparticles within specific plant microchannels through regioselective deposition, resulting in reduced chemical usage and an accelerated process. Moreover, this method effectively minimizes the required catalyst dosage. In this process, the plant monolith's aligned, narrow, and accessible channels provided a higher contact area, shorter diffusion path, and abundant oxygen-containing functional groups for rapid transformation of metal salt precursors into catalytic metal nanoparticles with excellent dispersion. The inside-out strategy can be extended to various plant-based monoliths and diverse metal/metal oxide/MOF materials within the plant monolith, thereby offering a facile, time- and cost-effective universal approach for skillfully designing plant-based flow microreactors for a wide range of applications.

Improving regioselectivity of phenylalanine aminomutase from Taxus chinensis by semi-rational mutagenesis for the biocatalytic amination of cinnamates.

The occupancy of the binding pocket by the substrate ultimately determines the outcome of enzyme catalysis. Previous engineering and substrate scope of phenylalanine aminomutase from Taxus chinensis (TcPAM) has generated valuable knowledge about the regioselectivity with biocatalytic potentials for the preparation of α- and ß-phenylalanine and their derivatives. However, the significantly different regioselectivity during the amination of cinnamates by TcPAM is not fully understood. In this study, we take a reconstruction approach to change the whole binding pocket of TcPAM for probing the factors affecting the regioselectivity, resulting in variant C107S/Q319M/I431V reaching a 25.5-fold enhancement of the ß/α product ratio toward trans-cinnamate acid. Furthermore, when substituted cinnamates were used as substrates, the regioselectivity was strongly correlated with various changes in the binding pocket, and value-added 2-Cl-α-Phe (100% α-selectivity) and 4-CH3-ß-Phe (98% ß-selectivity) were individually verified by the mutants L104A and Q319M at a preparative scale, exemplifying the application feasibility of our engineering strategy. The present study uncovered the cooperative connection between aromatic binding and carboxylate binding to affect the regioselectivity, which provides new insights into the determinants of the regioselectivity possessed by TcPAM and paves the way for its biocatalytic applications on phenylalanine derivatives.

An Exploratory Study of the Enzymatic Hydroxycinnamoylation of Sucrose and Its Derivatives.

Phenylpropanoid sucrose esters are a large and important group of natural substances with significant therapeutic potential. This work describes a pilot study of the enzymatic hydroxycinnamoylation of sucrose and its derivatives which was carried out with the aim of obtaining precursors of natural phenylpropanoid sucrose esters, e.g., vanicoside B. In addition to sucrose, some chemically prepared sucrose acetonides and substituted 3'-O-cinnamates were subjected to enzymatic transesterification with vinyl esters of coumaric, ferulic and 3,4,5-trimethoxycinnamic acid. Commercial enzyme preparations of Lipozyme TL IM lipase and Pentopan 500 BG exhibiting feruloyl esterase activity were tested as biocatalysts in these reactions. The substrate specificity of the used biocatalysts for the donor and acceptor as well as the regioselectivity of the reactions were evaluated and discussed. Surprisingly, Lipozyme TL IM catalyzed the cinnamoylation of sucrose derivatives more to the 1'-OH and 4'-OH positions than to the 6'-OH when the 3'-OH was free and the 6-OH was blocked by isopropylidene. In this case, Pentopan reacted comparably to 1'-OH and 6'-OH positions. If sucrose 3'-O-coumarate was used as an acceptor, in the case of feruloylation with Lipozyme in CH3CN, 6-O-ferulate was the main product (63%). Pentopan feruloylated sucrose 3'-O-coumarate comparably well at the 6-OH and 6'-OH positions (77%). When a proton-donor solvent was used, migration of the 3'-O-cinnamoyl group from fructose to the 2-OH position of glucose was observed. The enzyme hydroxycinnamoylations studied can shorten the targeted syntheses of various phenylpropanoid sucrose esters.

New Bis(pyrazolo[5,1-b]quinazolines) and Bis(9H-xanthenediones) Linked to Alkane Cores: One-Pot Synthesis, Antibacterial Screening, and SAR Study.

Effective one-pot methods were used to synthesize some new alkane-linked bis(pyrazolo[5,1-b]quinazolines) and bis(9H-xanthenediones). The first series was produced, in 80-88% yields, via the reaction of one equivalent of the appropriate bis(aldehydes) with two equivalents of 1H-pyrazole-3,5-diamine and dimedone in DMF at 150 ºC for 5-6 h. The second series was prepared, in 82-89% yields, via the reaction one equivalent of the appropriate bis(aldehydes) with four equivalents of dimedone in acetic acid at 120 ºC for 4-5 h. The new products displayed a broad range of antibacterial activity against different bacterial strains. Generally, the antibacterial activity of the alkane-linked bis(pyrazolo[5,1-b]quinazoline) units is more than 2-fold their bis(9H-xanthenedione) analogues. The (p-tolylthio)methyl)-linked bis(pyrazolo[5,1-b]quinazolines) demonstrate the best antibacterial activity with MIC/MBC values up to 3.3/6.6 µM.

Functional characterization, structural basis, and protein engineering of a rare flavonoid 2'-O-glycosyltransferase from Scutellaria baicalensis.

Glycosylation is an important post-modification reaction in plant secondary metabolism, and contributes to structural diversity of bioactive natural products. In plants, glycosylation is usually catalyzed by UDP-glycosyltransferases. Flavonoid 2'-O-glycosides are rare glycosides. However, no UGTs have been reported, thus far, to specifically catalyze 2'-O-glycosylation of flavonoids. In this work, UGT71AP2 was identified from the medicinal plant Scutellaria baicalensis as the first flavonoid 2'-O-glycosyltransferase. It could preferentially transfer a glycosyl moiety to 2'-hydroxy of at least nine flavonoids to yield six new compounds. Some of the 2'-O-glycosides showed noticeable inhibitory activities against cyclooxygenase 2. The crystal structure of UGT71AP2 (2.15 Å) was solved, and mechanisms of its regio-selectivity was interpreted by pK a calculations, molecular docking, MD simulation, MM/GBSA binding free energy, QM/MM, and hydrogen‒deuterium exchange mass spectrometry analysis. Through structure-guided rational design, we obtained the L138T/V179D/M180T mutant with remarkably enhanced regio-selectivity (the ratio of 7-O-glycosylation byproducts decreased from 48% to 4%) and catalytic efficiency of 2'-O-glycosylation (k cat/K m, 0.23 L/(s·µmol), 12-fold higher than the native). Moreover, UGT71AP2 also possesses moderate UDP-dependent de-glycosylation activity, and is a dual function glycosyltransferase. This work provides an efficient biocatalyst and sets a good example for protein engineering to optimize enzyme catalytic features through rational design.

Regioselective Electrochemical Construction of Csp2-Csp2 Linkage at C5-C5' Position of 2-Oxindoles via an Intermolecular Anodic Dehydrogenative Coupling.

Applying electricity as a reagent in synthetic organic chemistry has attracted particular attention from synthetic chemists worldwide as an environmentally benign and cost-effective technique. Herein, we report the construction of the Csp2-Csp2 linkage at the C5-C5' position of 2-oxindole utilizing electricity as the traceless oxidant in an anodic dehydrogenative homo-coupling process. A variety of 3,3-disubstituted-2-oxindoles were subjected to dimerization, achieving yields of up to 70% through controlled potential electrolysis at an applied potential of 1.5 V versus Ag/Ag+ nonaqueous reference electrode. This electro-synthetic approach facilitates the specific assembly of C5-C5' (para-para coupled) dimer of 3,3-disubstituted-2-oxindole without the necessity of any external oxidants or additives and DFT (Density Functional Theory) calculations provided confirmation of this pronounced regioselectivity. Furthermore, validation through control experiments and voltammetric analyses substantiated the manifestation of radical-radical coupling (or biradical pathway) for the dimerization process.

Kinetic and in silico structural characterization of norbelladine O-methyltransferase of Amaryllidaceae alkaloids biosynthesis.

Amaryllidaceae alkaloids are a diverse group of alkaloids exclusively reported from the Amaryllidaceae plant family. In planta, their biosynthesis is still not fully characterized; however, a labeling study established 4'-O-methylnorbelladine as the key intermediate compound of the pathway. Previous reports have characterized O-methyltransferases from several Amaryllidaceae species. Nevertheless, the formation of the different O-methylnorbelladine derivatives (3'-O-methylnorbelladine, 4'-O-methylnorbelladine, and 3'4'-O-dimethylnorbelladine), the role, and the preferred substrates of O-methyltransferases are not clearly understood. In this study, we performed the biochemical characterization of an O-methyltransferase candidate from Narcissus papyraceus (NpOMT) in vitro and in vivo, following biotransformation of norbelladine in Nicotiana benthamiana having transient expression of NpOMT. Docking analysis was further used to investigate substrate preferences, as well as key interacting residues of NpOMT. Our study shows that NpOMT methylates norbelladine preferentially at the 4'-OH position in vitro and in planta. Interestingly, NpOMT also catalyzed the synthesis of 3',4'-O-dimethylnorbelladine from norbelladine and 4'-O-methylnorbelladine during in vitro enzymatic assay. Furthermore, we show that NpOMT methylates 3,4-dihydroxybenzylaldehyde and caffeic acid in a nonregiospecific manner to produce meta/para monomethylated products. This study reveals a novel catalytic potential of an Amaryllidaceae O-methyltransferase and its ability to regioselectively methylate norbelladine in the heterologous host N. benthamiana.

Traceless Nucleophile Strategy for C5-Selective C-H Sulfonylation of Pyridines.

The functionalization of pyridines is crucial for the rapid construction and derivatization of agrochemicals, pharmaceuticals, and materials. Conventional functionalization approaches have primarily focused on the ortho- and para-positions, while achieving precise meta-selective functionalization, particularly at the C5 position in substituted pyridines, remains a formidable challenge due to the intrinsic electronic properties of pyridines. Herein, we present a new strategy for meta- and C5-selective C-H sulfonylation of N-amidopyridinium salts, which employs a transient enamine-type intermediate generated through a nucleophilic addition to N-amidopyridinium salts. This process harnesses the power of electron donor-acceptor complexes, enabling high selectivity and broad applicability, including the construction of complex pyridines bearing valuable sulfonyl functionalities under mild conditions without the need for an external photocatalyst. The remarkable C5 selectivity, combined with the broad applicability to late-stage functionalization, significantly expands the toolbox for pyridine functionalization, unlocking access to previously unattainable meta-sulfonylated pyridines.

Insight into C4 Selectivity in the Light-Driven C-H Fluoroalkylation of Pyridines and Quinolines.

Given the prevalence of pyridine motifs in FDA-approved drugs, selective fluoroalkylation of pyridines and quinolines is essential for preparing diverse bioisosteres. However, challenges are often faced with conventional Minisci reactions in achieving precise regioselectivity owing to competing reaction sites of pyridine and the limited availability of fluoroalkyl radical sources. Herein, we present a light-driven, C4-selective fluoroalkylation of azines utilizing N-aminopyridinium salts and readily available sulfinates. Our approach employs electron donor-acceptor complexes, achieving highly C4-selective fluoroalkylation under mild conditions without an external photocatalyst. This practical method not only enables the installation of CF2H groups but also allows for the incorporation of CF2-alkyl groups with diverse functional entities, surpassing the limitations of previous methods. The versatility of the radical pathway is further demonstrated through straightforward three-component reactions involving alkenes and [1.1.1]propellane. Detailed experimental and computational studies have elucidated the origins of regioselectivity, providing profound insights into the mechanistic aspects.

Regioselective photocyclodimerization of 2-anthracenecarboxylic acid through ATP hydrolysis-driven conformational change using simulation prediction-designed GroEL mutant.

GroEL, a chaperone protein responsible for peptide and denatured protein folding, undergoes substantial conformational changes driven by ATP binding and hydrolysis during folding. Utilizing these conformational changes, we demonstrated the GroEL-mediated regioselective photocyclodimerization of 2-anthracenecarboxylic acid (AC) using ATP hydrolysis as an external stimulus. We designed and prepared an optimal GroEL mutant to employ in a docking simulation that has been actively used in recent years. Based on the large difference in the motif of hydrogen bonds between AC and GroEL mutant compared with the wild-type, we predicted that GroELMEL, in which the 307‒309th amino acid residues were mutated to Ala, could alter the orientation of bound AC in GroEL. The GroELMEL-mediated photocyclodimerization of AC can be used for regioselective inversion upon ATP addition to a moderate extent.

Nickel-Catalyzed α-selective Hydroalkylation of Vinylarenes.

C(sp3) centers adjacent to (hetero) aryl groups are widely present in physiologically active molecules. Metal-hydride-catalyzed hydroalkylation of alkenes represents an efficient means of forging C(sp3)-C(sp3) bonds, boasting advantages as a wide source of substrates, mild reaction conditions, and facile selectivity manipulation. Nevertheless, the hydroalkylation of vinylarenes encounters constraints in terms of substrate scope, necessitating the employment of activated alkyl halides or alkenes containing chelating groups, remains a challenge. In this context, we report a general nickel-hydride-catalyzed hydroalkylation protocol for vinylarenes. Remarkably, this system enables α-selective hydroalkylation of both aryl and heteroaryl alkenes under an extra ligand-free condition, demonstrating excellent coupling efficiency and selectivity. Furthermore, through the incorporation of chiral bisoxazoline ligands, we have achieved regio- and enantioselective hydroalkylation of vinylpyrroles, thereby facilitating the synthesis of α-branched alkylated pyrrole derivatives.

Regioselective thexyldimethylsilylation of (1➔3)-glucans - Does the linkage type matter.

α- and ß-(1➔3)-linked polysaccharides dissolved in N,N-dimethyl acetamide (DMA) were subjected to conversion with thexyldimethylchlorosilane (TDMS-Cl) in presence of pyridine as base. A degree of substitution of TDMS groups (DSSi) between 0.7 and 1.0 was achieved indicating that the ß-(1➔3)-linked curdlan (DSSi 0.7) is less reactive than α-(1➔3)-linked glucans (DSSi ca. 1). The synthesis sequence of permethylation, desilylation, and acetylation afforded the corresponding acetyl-methyl derivatives, where unaffected OH groups were methylated and TDMS groups were replaced by acetyl moieties. NMR spectroscopic investigations revealed a highly selective silylation of the primary OH group at position 6 while leaving the secondary OH moieties unaffected. This pronounced selectivity was found to be distinctly higher compared to cellulose and starch. Conversion of (1➔4)-linked polysaccharides dissolved in DMA/LiCl with TDMS-Cl leads to products consisting of both 6-mono-O- and 2,6-di-O- silylated repeating units. Regioselective 6-mono-O-silylation requires the hazardous use of liquid ammonia.

Synthesis of regioselectively protected building blocks of benzyl ß-d-glucopyranoside.

Reported herein is the synthesis of benzyl ß-d-glucopyranoside and its derivatives that provide straightforward access to 3,4-branched glycans. Modes to diversify the synthetic intermediates via introduction of various temporary protecting groups have been demonstrated.

Engineering regioselectivity of glycosyltransferase for efficient polydatin synthesis.

Resveratrol is a promising functional ingredient applied in food products. However, low bioavailability and poor water solubility, which can be improved by glycosylation, hinder its application. A uridine diphosphate-dependent glycosyltransferase (UGT) from Bacillus subtilis 168 (named UGTBS) presents potential application for resveratrol glycosylation; nonetheless, imprecise regioselectivity renders the synthesis of resveratrol-3-O-ß-D-glucoside (polydatin) difficult. Therefore, molecular evolution was applied to UGTBS. A triple mutant Y14I/I62G/M315W was developed for 3-OH glycosylation of resveratrol and polydatin accounted for 91% of the total product. Kinetic determination and molecular docking indicated that the enhancement of hydrogen bond interaction and altered conformation of the binding pocket increases the enzyme's affinity for the 3-OH group, stabilizing the enzyme-substrate intermediate and promoting polydatin formation. Furthermore, a fed-batch cascade reaction by periodic addition of resveratrol was conducted and nearly 20 mM polydatin was obtained. The mutant Y14I/I62G/M315W can be used for polydatin manufacture.

Identify Regioselective Residues of Ginsenoside Hydrolases by Graph-Based Active Learning from Molecular Dynamics.

Identifying the catalytic regioselectivity of enzymes remains a challenge. Compared to experimental trial-and-error approaches, computational methods like molecular dynamics simulations provide valuable insights into enzyme characteristics. However, the massive data generated by these simulations hinder the extraction of knowledge about enzyme catalytic mechanisms without adequate modeling techniques. Here, we propose a computational framework utilizing graph-based active learning from molecular dynamics to identify the regioselectivity of ginsenoside hydrolases (GHs), which selectively catalyze C6 or C20 positions to obtain rare deglycosylated bioactive compounds from Panax plants. Experimental results reveal that the dynamic-aware graph model can excellently distinguish GH regioselectivity with accuracy as high as 96-98% even when different enzyme-substrate systems exhibit similar dynamic behaviors. The active learning strategy equips our model to work robustly while reducing the reliance on dynamic data, indicating its capacity to mine sufficient knowledge from short multi-replica simulations. Moreover, the model's interpretability identified crucial residues and features associated with regioselectivity. Our findings contribute to the understanding of GH catalytic mechanisms and provide direct assistance for rational design to improve regioselectivity. We presented a general computational framework for modeling enzyme catalytic specificity from simulation data, paving the way for further integration of experimental and computational approaches in enzyme optimization and design.

Regioselective alkylation of a versatile indazole: Electrophile scope and mechanistic insights from density functional theory calculations.

Herein, we report a pair of regioselective N 1- and N 2 -alkylations of a versatile indazole, methyl 5-bromo-1H-indazole-3-carboxylate (6) and the use of density functional theory (DFT) to evaluate their mechanisms. Over thirty N 1- and N 2-alkylated products were isolated in over 90% yield regardless of the conditions. DFT calculations suggest a chelation mechanism produces the N 1-substituted products when cesium is present and other non-covalent interactions (NCIs) drive the N 2-product formation. Methyl 1H-indazole-7-carboxylate (18) and 1H-indazole-3-carbonitrile (21) were also subjected to the reaction conditions and their mechanisms were evaluated. The N 1- and N 2-partial charges and Fukui indices were calculated for compounds 6, 18, and 21 via natural bond orbital (NBO) analyses which further support the suggested reaction pathways.

Br-to-Cl Transformation Guided the Formation of Polyhalogenated Dibenzo-p-dioxins/Dibenzofurans.

Polyhalogenated dibenzo-p-dioxins/dibenzofurans (PXDD/Fs) are commonly released into the environment as byproducts of combustion processes, accompanied by flue gases. Chlorinated (Cl) and brominated (Br) precursors play crucial roles in forming PXDD/Fs. However, the specific contributions of Cl-precursors and Br-precursors to PXDD/Fs formation have not been fully elucidated. Herein, we demonstrate that the formation of Br-precursors can increase the fraction of polychlorinated dibenzo-p-dioxins/dibenzofurans (PCDD/Fs) congeners substituted at specific positions, such as 1,2,3,4,6,7,8-HpCDD, OCDD, 2,3,4,7,8-PeCDF, and 2,3,4,6,7,8-HxCDF. This is attributed to the electrophilic chlorination reaction of the Br-precursors, which includes the Br-to-Cl transformation pathway, following the principle of regioselectivity. The observed formation of polybrominated/chlorinated dibenzo-p-dioxins/benzofurans (PBCDD/Fs) from 1,2-dibromobenzene (1,2-DiBBz) as a Br precursor provides direct evidence supporting the proposed Br-to-Cl transformation. Quantum chemical calculations are employed to discuss the principle of regioselectivity in the Br-to-Cl transformation, clarifying the priority of the position for electrophilic chlorination. Additionally, the concentration of PCDD/Fs formed from 1,2-DiBBz is 1.6 µg/kg, comparable to that of polybrominated dibenzo-p-dioxins/dibenzofurans (PBDD/Fs) (2.4 µg/kg), highlighting the potential of brominated organic pollutants as precursors for PCDD/Fs formation. This study provides three potential pathways for PCDD/Fs formation from Br-precursors, establishing a theoretical foundation for elucidating the formation mechanism of PXDD/Fs in the coexistence of Cl and Br.

Regioselectivity in Pyrene-Templated Polymerization Using MOFs as 1D Porous Scaffolds.

Physicochemical properties of polymers strongly depend on the arrangement and distribution of attached monomers. Templated polymerization using porous crystalline materials appears as a promising route to gain control on the process. Thus, we demonstrate here the potential of metal-organic frameworks as scaffolds with a versatile and very regular porosity, well adapted for the regioselective oxidative polymerization of pyrene. This photoresponsive monomer was first encapsulated within the one-dimensional (1D) microporosity of the robust zirconium(IV) carboxylate metal-organic framework (MOF) (MIL-140D) to, later, undergo in situ oxidative polymerization, enabling the growth of a highly selective polypyrene (PPyr) regioisomer over other potential polymer configurations. To confirm the polymerization and the geometry control of pyrene, the resulting composites were exhaustively characterized by powder X-ray diffraction (PXRD), thermogravimetric analysis (TGA), N2 sorption measurements, scanning transmission electron microscopy coupled with energy-dispersive X-ray (STEM-EDX) spectroscopy, and fluorescence spectroscopy. Among others, photoluminescence quenching and emission shift in the solid state demonstrated the presence of PPyr inside the MOF porosity. Furthermore, an in-depth joint analysis combining solid-state, magic-angle spinning (MAS) 1H and 13C NMR spectroscopy, Fourier transform infrared (FTIR) spectroscopy, matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy (MALDI-TOF MS), and molecular simulations (grand canonical Monte Carlo (GCMC) and density functional theory (DFT)) allowed the elucidation of the spatial, host-guest interactions driving the polymerization reaction.

REACTION KINETICS OF THE BENZYLATION OF ADENINE IN DMSO: REGIO-SELECTIVITY GUIDED BY ENTROPY.

The factors governing the regio-selectivity of the alkylation of adenine have been of interest for many years due to the biological importance of adenine derivatives, however, no reaction kinetic studies have been conducted. Herein, we report the rate constants and activation parameters of the benzylation of adenine under basic conditions in DMSO in the absence and presence of 15-crown-5 ether using real-time 1H NMR spectroscopy. The reaction is second-order for the formation of the N9- and N3-benzyladenine products, with a regio-selectivity factor 2.3 in favour of the N9-adduct. The Gibbs free energy of activation amounts to 87±2 kJ mol-1 for both reactions. The formation of the N9-adduct is more activated by 7 kJ mol-1, but its effect is offset by a less negative activation entropy, demonstrating that the long-contested reason for the regioselectivity in the benzylation of adenine is dominated by compensation of entropy and enthalpy in the transition state. The kinetic parameters obtained in the presence of the 15-crown-5 ether indicate that the crown ether forms a complex with an adenine-sodium ion-pair, increasing the activation barrier. However, the Gibbs free energy in the absence and presence of the crown ether remains constant.