Asymmetric Reduction Of Prochiral Ketones Research Articles

Vigna radiata (mung) as superior natural biobased catalyst for asymmetric reduction of prochiral acetophenone

Asymmetric reduction of prochiral ketone via biocatalysis is one of the most promising and straight forward routes to prepare optically active alcohols. In this work, whole cells of Vigna radiata (also called mung) catalyzed enantioselective reduction of prochiral aromatic ketones was investigated. Effects of various parameters such as co-substrate, temperature, cell loading, substrate concentration on conversion and enantioselectivity were studied in a systematic manner by using acetophenone as the model compound. Glucose was found to be the most effective co-substrate for co-factor regeneration among different co-substrates employed. Whole cells of mung have shown excellent thermostability and high substrate tolerance (up to 200 mM). Under optimum conditions, maximum conversion of 93.44% and enantioselectivity of 98.92% were obtained in 9 h. Good conversion and enantioselectivity were observed with various substituted aromatic ketones. This is a cheap, clean, and eco-friendly process for preparation of chiral secondary alcohols compared to chemical processes.

Enantioselective Regulation of Short‐Chain Dehydrogenases via Key Sites in its Loop and Adjacent Regions for the Enantiodivergent Reduction of Difficult‐To‐Reduce Ketones

AbstractShort‐chain dehydrogenases (SDRs) are powerful catalysts for the asymmetric reduction of prochiral ketones in pharmaceutical products. Herein, through gene mining and evolutionary analysis, we obtained two major types of SDRs (SDR‐1 and SDR‐2) for the enantioselective complementary reduction of N‐Boc‐piperidone, which gives the corresponding product (R)‐ or (S)‐N‐Boc‐piperidol, an intermediate of the interleukin inhibitor and lymphoma treatment drug (imbruvica), respectively. By integrating multiple sequence alignment, site‐directed mutagenesis and computational modeling, we proposed a “loop regulation” mechanism for the enantioselective control of SDRs, through which residues in the loop region could potentially fine‐tune their enantioselectivity. Further, site‐directed mutagenesis assays showed that two key residues (L201 and F205 for SDR‐1, F92 and H93 for SDR‐2) in the loop and its adjacent region played critical roles in fine‐tuning the enantioselectivity of SDRs. Understanding this mechanism of SDR stereo preference in catalyzing asymmetric reduction, we further switched the enantioselectivity of the homologous enzymes. The obtained enzymes catalyzed the enantiodivergent synthesis of chiral heterocyclic alcohols with different ring sizes and substituents (25–99% conversion and 25–99% ee (R/S)), including piperidols, 4‐hydroxy azepanes, 3‐hydroxy azepanes and pyrrolidinols. These findings could potentially guide future attempts at protein engineering of stereoselective SDRs.

Development of a novel magnetic metal-organic framework for the immobilization of short-chain dehydrogenase for the asymmetric reduction of pro-chiral ketone

Short-chain dehydrogenase/reductase (SDR) acts as a biocatalyst in the synthesis of chiral alcohols with high optical purity. Herein, we achieved immobilization via crosslinking on novel magnetic metal–organic framework nanoparticles with a three-layer shell structure (Fe3O4@PDA@Cu (PABA)). The results of scanning electron microscopy, Fourier-transform infrared spectroscopy, X-ray diffraction, X-ray photoelectron spectroscopy, and energy dispersive X-ray spectroscopy confirmed the morphology and cross-linking property of immobilized SDR, which was more durable, stable, and reusable and exhibited better kinetic performance than free enzyme. The SDR and glucose dehydrogenase (GDH) were co-immobilized and then used for the asymmetric reduction of COBE and ethyl 2-oxo-4-phenylbutanoate (OPBE). These finding suggest that enzymes immobilized on novel MOF nanoparticles can serve as promising biocatalysts for asymmetric reduction prochiral ketones into chiral alcohols.

Modified distance optimization method for the asymmetric bioreduction conditions of phenyl(thiophen-2-yl)methanone by Weissella paramesenteroides N7

Chirality plays a significant part in life since it is closely linked to carrying out the many metabolic processes that make up a living being. Chiral secondary alcohols such as diaryl-, aryl heteroaryl-, and diheteroaryl are employed in pharmaceuticals as drug intermediates. Although biocatalytic asymmetric reduction of prochiral ketones containing aromatic and heteroaromatic groups is widely known, biocatalytic reduction of heteroaromatic prochiral ketones containing sulfur heteroatoms is rarely known. Very few studies exist in the literature, including the biocatalytic reduction of phenyl(thiophen-2-yl)methanone (1). Moreover, the biocatalytic reduction of 1 with a mathematical modelling and optimization technique is unknown until now. In this study, Weissella paramesenteroides N7 biocatalyst for the asymmetric bioreduction of 1 using a novel modified distance optimization method. Optimization conditions were found as pH = 6.46, temperature = 26 °C, incubation period = 71 h, agitation speed = 200 rpm by the modified distance optimization method, and it was determined that the conversion and enantiomeric excess (ee) under these conditions could be 98.7% and 98%, respectively. Under these proposed optimization conditions, (S)-phenyl(thiophen-2-yl)methanol ((S)-2) was obtained with >99% ee, >99% conversion, and 97% yield. In addition, 11.29 g of 1 was completely converted into (S)-2 (11.07 g, 97% isolated yield) under optimized conditions. This is the first report about the fabrication of enantiopure (S)-2 in high gram scale using a biocatalyst and a novel modified distance optimization technique. In this study, the successful applicability of the new modified distance optimization method in biocatalytic asymmetric reduction reactions has been successfully demonstrated.

Optimization of asymmetric bioreduction conditions of 1-indanone by Leuconostoc mesenteroides N6 using a face-centered design-based multi-objective optimization model

There has been an increasing interest in biocatalysts over the past few decades in order to obtain high efficiency, high yield, and environmentally benign procedures aiming at the manufacture of pharmacologically relevant chemicals. Lactic Acid Bacteria (LAB), a microbial group, can be employed as biocatalysts while performing asymmetric reduction of prochiral ketones. In this study, Leuconostoc mesenteroides N6 was used for the asymmetric bioreduction 1-indanone. And then, a novel and innovative face-centered design-based multi-objective optimization model was used to optimize experimental conditions. Also, the experimental design factors were defined as agitation speed, incubation period, pH, and temperature for optimization to acquire the maximum enantiomeric excess (ee) and conversion rate (cr) values. When using the face-centered design-based multi-objective optimization model, the optimum culture conditions corresponded to 96.34 and 99.42%, ee and cr responses, respectively, were pH = 5.87, incubation temperature = 35 °C, incubation period = 50.88 h, and agitation speed = 152.60 rpm. Notably, the validation experiment under the optimum model conditions confirmed the model results. This study demonstrated the importance of the optimization and the efficiency of the face-centered design-based multi-objective model.

New Anti-Prelog Stereospecific Whole-Cell Biocatalyst for Asymmetric Reduction of Prochiral Ketones.

The biocatalytic asymmetric reduction of prochiral ketones for the production of enantiopure alcohols is highly desirable due to its inherent advantages over chemical methods. In this study, a new bacterial strain capable of transforming ketones to corresponding alcohols with high activity and excellent enantioselectivity was discovered in a soil sample. The strain was subsequently identified as Bacillus cereus TQ-2 based on its physiological characteristics and 16S rDNA sequence analysis. Under optimized reaction conditions, the resting cells of B. cereus TQ-2 converted acetophenone to enantioenriched (R)-1-phenylethanol with 99% enantiometric excess following anti-Prelog's rule, which is scarce in biocatalytic ketone reduction. The optimum temperature for the cells was 30 °C, and considerable catalytic activity was observed over a broad pH range from 5.0 to 9.0. The cells showed enhanced catalytic activity in the presence of 15% (v/v) glycerol as a co-substrate. The catalytic activity can also be substantially improved by adding Ca2+ or K+ ions. Moreover, the B. cereus TQ-2 cell was highly active in reducing several structurally diverse ketones and aldehydes to form corresponding alcohols with good to excellent conversion. Our study provides a versatile whole-cell biocatalyst that can be used in the asymmetric reduction of ketones for the production of chiral alcohol, thereby expanding the biocatalytic toolbox for potential practical applications.

Structural analysis of an anthrol reductase inspires enantioselective synthesis of enantiopure hydroxycycloketones and β-halohydrins

Asymmetric reduction of prochiral ketones, particularly, reductive desymmetrization of 2,2-disubstituted prochiral 1,3-cyclodiketones to produce enantiopure chiral alcohols is challenging. Herein, an anthrol reductase CbAR with the ability to accommodate diverse bulky substrates, like emodin, for asymmetric reduction is identified. We firstly solve crystal structures of CbAR and CbAR-Emodin complex. It reveals that Tyr210 is critical for emodin recognition and binding, as it forms a hydrogen-bond interaction with His162 and π-π stacking interactions with emodin. This ensures the correct orientation for the stereoselectivity. Then, through structure-guided engineering, variant CbAR-H162F can convert various 2,2-disubstituted 1,3-cyclodiketones and α-haloacetophenones to optically pure (2S, 3S)-ketols and (R)-β-halohydrins, respectively. More importantly, their stereoselectivity mechanisms are also well explained by the respective crystal structures of CbAR-H162F-substrate complex. Therefore, this study demonstrates that an in-depth understanding of catalytic mechanism is valuable for exploiting the promiscuity of anthrol reductases to prepare diverse enantiopure chiral alcohols.

Recent Advances in Organocatalyzed Asymmetric Reduction of Prochiral Ketones: An Update

AbstractChiral alcohols are important synthetic intermediates and building blocks for the synthesis of drugs, agrochemicals, and natural products. Asymmetric reduction of prochiral ketones has been the most investigated method for accessing chiral alcohols. In this regard, organocatalyzed asymmetric reduction, as a complementary method to transition-metal- and enzyme-catalyzed reactions, has attracted tremendous interest in the past decades due to the reactions with such catalysts being metal-free and easy to operate, and principally, the ease of recovery and the ability to reuse the catalysts. Following up on a comprehensive overview on organocatalyzed asymmetric reductions of prochiral ketones in early 2018, this short review is intended to summarize the recent progress in this area from the beginning of 2018 until the end of August 2021.1 Introduction2 Boron-Based Chiral Organocatalysts2.1 Boron-Containing Chiral Schiff Base Catalysts2.2 Chiral Alpine-Borane Catalysts2.3 Boron-Containing Chiral Frustrated Lewis Pair Catalysts2.4 Chiral Borate Ester–Amine Complex Catalysts3 Phosphorus-Based Chiral Organocatalysts3.1 Chiral Phosphoric Acid Organocatalysts3.2 Chiral Phosphinamide and Phosphoramide Organocatalysts4 Chiral Ionic Liquid Organocatalysts5 Chiral-Oxazoline-Based Organocatalysts6 Conclusion and Outlook

Organosilanes in Metal-Catalyzed, Enantioselective Reductions

The growth and development of an extensive range of metals complexed with chiral ligands for the purpose of catalyzing a variety of reactions in an enantioselective manner has been impressive for its scope, chemical yields and high ee values. Chief among the classes of synthetic transformations has been that of the asymmetric reduction of prochiral substrates. These include the asymmetric reduction of prochiral ketones, imines, unsaturated aldehydes, ketones, esters, nitriles and olefins, as well as a number of asymmetric coupling reactions. In concert with these efficient catalyst systems, organosilanes have the ability to carry out any number of organic reductions under a variety of conditions. In these reactions, which in reality are hydrosilylations (hydrosilylation and reduction are considered interchangeable herein) in which the initially resulting silylated product is hydrolyzed to the ultimate desired functionality, the ability to sterically and electronically alter the organosilane reductant can contribute to the overall success of the transformation. In this review, we present a thorough compilation of the literature covering the use of organosilanes in metal-catalyzed asymmetric reductions (hydrosilylations) and coupling reactions.

The Development of Organocatalytic Asymmetric Reduction of Carbonyls and Imines Using Silicon Hydrides

AbstractThe asymmetric reduction of prochiral ketones and ketimines represents one of the most important and practical chemical transformations toward the synthesis of valuable chiral alcohols and amines. Among the existing strategies, organocatalyzed asymmetric reduction of carbonyls and imines using silicon hydrides is attractive due to low‐cost, chemical stability, and easy handling in experiments. In this review, we wish to highlight the recent progress made in the past fifteen years in this field. Four catalytic systems of different activation modes are presented and discussed in detail. We aim to help shed light on common features that enable highly enantioselective silicon hydride reductions through organocatalysis and provide the design principles for the development of more effective catalytic systems.

Optimization of asymmetric reduction conditions of 1-(benzo [d] [1,3] dioxol-5-yl) ethanone by Lactobacillus fermentum P1 using D-optimal experimental design-based model

The biocatalytic asymmetric reduction of prochiral ketones is a significant transformation in organic chemistry as chiral carbinols are biologically active molecules and may be used as precursors of many drugs. In this study, the bioreduction of 1-(benzo [d] [1,3] dioxol-5-yl) ethanone for the production of enantiomerically pure (S)-1-(1,3-benzodioxal-5-yl) ethanol was investigated using freeze-dried whole-cell of Lactobacillus fermentum P1 and the reduction conditions was optimized with a D-optimal experimental design-based optimization methodology. This is the first study using this optimization methodology in a biocatalytic asymmetric reduction. Using D-optimal experimental design-based optimization, optimum reaction conditions were predicted as pH 6.20, temperature 30 °C, incubation time 30 h, and agitation speed 193 rpm. For these operating conditions, it was estimated that the product could be obtained with 94% enantiomeric excess (ee) and 95% conversion rate (cr). Besides, the actual ee and cr were found to be 99% tested under optimized reaction conditions. These findings demonstrated that L. fermentum P1 as an effective biocatalyst to obtain (S)-1-(1,3-benzodioxal-5-yl) ethanol and with the D-optimal experimental design-based optimization, this product could be obtained with the 99% ee and 99% cr. Finally, the proposed mathematical optimization technique showed the applicability of the obtained results for asymmetric reduction reactions.

Enantioselective reduction of heterocyclic ketones with low level of asymmetry using carrots

A whole spectrum of biocatalysts for asymmetric reduction of prochiral ketones is well known including the Daucus carota root. However, this type of reaction is still challenging when pro-chiral ketones present low level of asymmetry, like heterocyclic ketones. In this work, 4,5-dihydro-3(2H)-thiophenone (1), 2-methyltetrahydrofuran-3-one (2), N-Boc-3-pyrrolidinone (3), 1-Z-3-pyrrolidinone (4) and 1-benzyl-3-pyrrolidinone (5) were studied in order to obtain the respective enantioselective heterocyclic secondary alcohols. Except for 5, the corresponding alcohols were obtained in high values of conversion and with high selectivity. In order to circumvent the low isolated yield of the corresponding chiral alcohol from 2, we observed that the use of carrots in the absence of water is feasible. Addition of co-solvents was needed to the water-insoluble ketones 3 and 4. Comparatively, baker’s yeast was used for bio reductions of 1, 3 and 4. And in terms of conversion, selectivity and work-up, the use of carrots were a more efficient biocatalyst, as well as a viable method for obtaining 5-member heterocyclic secondary alcohols.

Promotion of the Asymmetric Reduction of Prochiral Ketone with Recombinant E. coli Through Strengthening Intracellular NADPH Supply by Modifying EMP and Introducing NAD Kinase

The intracellular NADPH insufficient supply is the main bottleneck to the synthesis of chiral alcohols by asymmetric reduction with whole-cell catalysis. Herein, we provide a novel strategy to strengthen intracellular NADPH supply through introducing an NADP+-dependent glyceraldehyde 3-phosphate dehydrogenase (gapB from Bacillus subtilis 168) into the Embden-Meyerhof pathway and a NAD kinase (yfjB from E. coli MG1655) to further enhance the NADP(H) pool. A recombinant E. coli (E. coli BL21 (DE3)/pETDuet-1-gapB-yueD&pET28a-yfjB) was constructed to co-express gapB and yfjB with a carbonyl reductase gene yueD together. The result showed that the intracellular NADPH amount increased by 134.4% with the strategy. To the model reaction (asymmetric reduction of acetophenone to S-phenyl ethanol), the yield was 3.7-fold with this strategy compared to the control. This provides a technological route for strengthening the intracellular NADPH supply in E. coli for biocatalysis and biosynthesis.

Asymmetric reduction of aromatic heterocyclic ketones with bio-based catalyst Lactobacillus kefiri P2

Chiral heterocyclic secondary alcohols have received much attention due to their widespread use in pharmaceutical intermediates. In this study, Lactobacillus kefiri P2 biocatalysts isolated from traditional dairy products, were used to catalyze the asymmetric reduction of prochiral ketones to chiral secondary alcohols. Secondary chiral carbinols were obtained by asymmetric bioreduction of different prochiral substrates with results up to > 99% enantiomeric excess (ee). (R)-1-(benzofuran-2-yl)ethanol 5a, which can be used in the synthesis of pharmaceuticals such as bufuralols potent nonselective β-blockers antagonists, Amiodarone (cardiac anti-arrhythmic), and Benziodarone (coronary vasodilator), was produced in gram-scale, high yield and enantiomerically pure form using L. kefiri P2 biocatalysts. The gram-scale production was carried out, and 9.70 g of (R)-5a in enantiomerically pure form was obtained in 96% yield. Also, production of (R)-5a in terms of yield and gram scale through catalytic asymmetric reduction using the biocatalyst was the highest report so far. This is a cost-effective, clean and eco-friendly process for the preparation of chiral secondary alcohols compared to chemical processes. From an environmental and economic perspective, this biocatalytic method has great application potential, making it a green and sustainable way of synthesis.

Characterization of a novel aldo-keto reductase with anti-Prelog stereospecificity from Corallococcus sp. EGB

The asymmetric reduction of prochiral ketones is a promising process for synthesis of optically active alcohols. The aldo-keto reductase (AKR) is an attractive candidate of biocatalyst, due to its high enantioselectivity and environmentally friendly reaction conditions. In this work, nine putative AKR encoding genes from Corallococcus sp. EGB were cloned and expressed in Escherichia coli. Of these produced enzymes (CoAKRs), CoAKR7 exhibited reductive activity to various ketones and ketoesters, especially very high activity toward ethyl 4-chloro-3-oxobutanoate (COBE) with NADPH as the coenzyme. The CoAKR7 was optimally active at pH 7.0 and 50 °C. The apparent Km and Vmax for COBE was 14.18 U/mg and 0.269 mM, respectively. Moreover, CoAKR7 catalyzed an anti-Prelog reduction of COBE to (S)-ethyl-4-chloro-3-hydroxybutanoate (CHBE) with e.e. >99%. Enzyme-substrate-cofactor docking analysis elucidated the molecular mechanism of the substrate stereospecificity, providing basis for protein engineering of these enzymes for applications in the synthesis of valuable chemicals.

Conformational Dynamics‐Guided Loop Engineering of an Alcohol Dehydrogenase: Capture, Turnover and Enantioselective Transformation of Difficult‐to‐Reduce Ketones

AbstractDirected evolution of enzymes for the asymmetric reduction of prochiral ketones to produce enantio‐pure secondary alcohols is particularly attractive in organic synthesis. Loops located at the active pocket of enzymes often participate in conformational changes required to fine‐tune residues for substrate binding and catalysis. It is therefore of great interest to control the substrate specificity and stereochemistry of enzymatic reactions by manipulating the conformational dynamics. Herein, a secondary alcohol dehydrogenase was chosen to enantioselectively catalyze the transformation of difficult‐to‐reduce bulky ketones, which are not accepted by the wildtype enzyme. Guided by previous work and particularly by structural analysis and molecular dynamics (MD) simulations, two key residues alanine 85 (A85) and isoleucine 86 (I86) situated at the binding pocket were thought to increase the fluctuation of a loop region, thereby yielding a larger volume of the binding pocket to accommodate bulky substrates. Subsequently, site‐directed saturation mutagenesis was performed at the two sites. The best mutant, where residue alanine 85 was mutated to glycine and isoleucine 86 to leucine (A85G/I86L), can efficiently reduce bulky ketones to the corresponding pharmaceutically interesting alcohols with high enantioselectivities (∼99% ee). Taken together, this study demonstrates that introducing appropriate mutations at key residues can induce a higher flexibility of the active site loop, resulting in the improvement of substrate specificity and enantioselectivity.magnified image

Recent trends in organocatalyzed asymmetric reduction of prochiral ketones

This review depicts the recent practices followed in organocatalyzed asymmetric reduction of prochiral ketones, highlighting the main organocatalysts used for the past seven years.

Asymmetric Reduction of Prochiral Ketones by Using Self-Sufficient Heterogeneous Biocatalysts Based on NADPH-Dependent Ketoreductases.

The development of cell-free and self-sufficient biocatalytic systems represents an emerging approach to address more complex synthetic schemes under nonphysiological conditions. Herein, we report the development of a self-sufficient heterogeneous biocatalyst for the synthesis of chiral alcohols without the need to add an exogenous cofactor. In this work, an NADPH-dependent ketoreductase was primarily stabilized and further co-immobilized with NADPH to catalyze asymmetric reductions without the addition of an exogenous cofactor. As a result, the immobilized cofactor is accessible, and thus, it is recycled inside the porous structure without diffusing out into the bulk, as demonstrated by single-particle in operando studies. This self-sufficient heterogeneous biocatalyst was used and recycled for the asymmetric reduction of eleven carbonyl compounds in a batch reactor without the addition of exogenous NADPH to achieve the corresponding alcohols in 100 % yield and >99 % ee; this high performance was maintained over five consecutive reaction cycles. Likewise, the self-sufficient heterogeneous biocatalyst was integrated into a plug flow reactor for the continuous synthesis of one model secondary alcohol, which gave rise to a space-time yield of 97-112 g L-1 day-1 ; additionally, the immobilized cofactor accumulated a total turnover number of 1076 for 120 h. This is one of the few examples of the successful implementation of continuous reactions in aqueous media catalyzed by cell-free and immobilized systems that integrate both enzymes and cofactors into the solid phase.

Co‐immobilized Phosphorylated Cofactors and Enzymes as Self‐Sufficient Heterogeneous Biocatalysts for Chemical Processes

AbstractEnzyme cofactors play a major role in biocatalysis, as many enzymes require them to catalyze highly valuable reactions in organic synthesis. However, the cofactor recycling is often a hurdle to implement enzymes at the industrial level. The fabrication of heterogeneous biocatalysts co‐immobilizing phosphorylated cofactors (PLP, FAD+, and NAD+) and enzymes onto the same solid material is reported to perform chemical reactions without exogeneous addition of cofactors in aqueous media. In these self‐sufficient heterogeneous biocatalysts, the immobilized enzymes are catalytically active and the immobilized cofactors catalytically available and retained into the solid phase for several reaction cycles. Finally, we have applied a NAD+‐dependent heterogeneous biocatalyst to continuous flow asymmetric reduction of prochiral ketones, thus demonstrating the robustness of this approach for large scale biotransformations.

Co-immobilized Phosphorylated Cofactors and Enzymes as Self-Sufficient Heterogeneous Biocatalysts for Chemical Processes.

Enzyme cofactors play a major role in biocatalysis, as many enzymes require them to catalyze highly valuable reactions in organic synthesis. However, the cofactor recycling is often a hurdle to implement enzymes at the industrial level. The fabrication of heterogeneous biocatalysts co‐immobilizing phosphorylated cofactors (PLP, FAD+, and NAD+) and enzymes onto the same solid material is reported to perform chemical reactions without exogeneous addition of cofactors in aqueous media. In these self‐sufficient heterogeneous biocatalysts, the immobilized enzymes are catalytically active and the immobilized cofactors catalytically available and retained into the solid phase for several reaction cycles. Finally, we have applied a NAD+‐dependent heterogeneous biocatalyst to continuous flow asymmetric reduction of prochiral ketones, thus demonstrating the robustness of this approach for large scale biotransformations.