Debaryomyces hansenii as a new biocatalyst in the asymmetric reduction of substituted acetophenones

Chiral secondary alcohols are convenient mediator for the synthesis of biologically active compounds and natural products. In this study fifteen yeast strains belonging to three food originated yeast species Debaryomyces hansenii, Saccharomyces cerevisiae and Hanseniaspora guilliermondii were tested for their capability for the asymmetric reduction of acetophenone to 1-phenylethanol as biocatalyst microorganisms. Of these strains, Debaryomyces hansenii P1 strain showed an effective asymmetric reduction ability. Under optimized conditions, substituted acetophenones were converted to the corresponding optically active secondary alcohols in up to 99% enantiomeric excess and at high conversion rates. This is the first report on the enantioselective reduction of acetophenone by D. hansenii P1 from past?rma, a fermented Turkish meat product. The preparative scale asymmetric bio reduction of 3-methoxy acetophenone 1g by D. hansenii P1 gave (R)-1-(3-methoxyphenyl) ethanol 2g 82% yield, and >99% enantiomeric excess. Compound 2g can be used for the synthesis of (+)-NPS-R-568 [3-(2-chlorophenyl)-N-[(1R)-1-(3-methoxyphenly) ethyl] propan-1-amine] which have a great potential for the treatment of primary and secondary hyper-parathyroidism. In addition, D. hansenii P1 successfully reduced acetophenone derivatives. This study showed that this yeast can be used industrially to produce enantiomerically pure chiral secondary alcohols, which can be easily converted to different functional groups.     

Production of enantiomerically pure (S)-phenyl(pyridin-2-yl)methanol with Lactobacillus paracasei BD101

Abstract

Asymmetric reduction studies of heteroaryl ketones, including phenyl(pyridin-2-yl)methanone in enantioselective form with biocatalysts are very few, and chiral heteroaryl alcohols have been synthesized generally in the small scale. In this study, seven bacterial strains have been used to produce the (S)-phenyl(pyridin-2-yl)methanol in high enantiomeric excess and yield. Among the tested strains, Lactobacillus paracasei BD101, was found to be the best biocatalyst for the reducing phenyl(pyridin-2-yl)methanone to the (S)-phenyl(pyridin-2-yl)methanol at gram scale. The asymmetric bioreduction conditions were systematically optimized using L. paracasei BD101, which demonstrated excellent enantioselectivity and high level of conversion for the bioreduction reaction. (S)-phenyl(pyridin-2-yl)methanol, which is an analgesic, was produced enantiomerically pure form in the first time on gram scale using a biocatalyst. In total, 5.857?g of (S)-phenyl(pyridin-2-yl)methanol in enantiomerically pure form (>99% enantiomeric excess) was obtained in 52?h with 93% yield using whole cells of L. paracasei BD101. Enantiomerically pure (S)-phenyl (pyridin-2-yl)methanol, which is an analgesic, was first produced in the gram scale using a biocatalyst with excellent ee (>99%) and yield (93%).     

Whole cell application of Lactobacillus paracasei BD101 to produce enantiomerically pure (S)‐cyclohexyl(phenyl)methanol

In this study, a total of 10 bacterial strains were screened for their ability to reduce cyclohexyl(phenyl)methanone 1 to its corresponding alcohol. Among these strains, Lactobacillus paracasei BD101 was found to be the most successful biocatalyst to reduce the ketones to the corresponding alcohols. The reaction conditions were systematically optimized for the reducing agent L paracasei BD101, which showed high enantioselectivity and conversion for the bioreduction. The preparative scale asymmetric reduction of cyclohexyl(phenyl)methanone ( 1 ) by L paracasei BD101 gave (S)‐cyclohexyl(phenyl)methanol ( 2 ) with 92% yield and >99% enantiomeric excess. The preparative scale study was carried out, and a total of 5.602 g of (S)‐cyclohexyl(phenyl)methanol in high enantiomerically pure form (>99% enantiomeric excess) was produced. L paracasei BD101 has been shown to be an important biocatalyst in asymmetric reduction of bulky substrates. This study demonstrates the first example of the effective synthesis of (S)‐cyclohexyl(phenyl)methanol by the L paracasei BD101 as a biocatalyst in preparative scale.     

Synthesis of Enantiomerically Enriched Drug Precursors by Lactobacillus paracasei BD87E6 as a Biocatalyst

Global sales of single enantiomeric drug products are growing at an alarming rate every year. A total of 7 bacterial strains were screened for their ability to reduce acetophenones to its corresponding alcohol. Among these strains Lactobacillus paracasei BD87E6 was found to be the most successful biocatalyst to reduce the ketones to the corresponding alcohols. The reaction conditions were systematically optimized for the reducing agent Lactobacillus paracasei BD87E6, which showed high enantioselectivity and conversion for the bioreduction. The preparative scale asymmetric reduction of 3‐methoxyacetophenone ( 1h ) by Lactobacillus paracasei BD87E6 gave (R)‐1‐(3‐methoxyphenyl)ethanol ( 2h ) with 92% yield and 99% enantiomeric excess. Compound 2h could be used for the synthesis of (S)‐rivastigmine which has a great potential for the treatment of Alzheimer's disease. This study demonstrates that Lactobacillus paracasei BD87E6 can be used as a biocatalyst to obtain chiral carbinol with excellent yield and selectivity. The whole cell catalyzed the reductions of ketone substrates on the preparative scale, demonstrating that Lactobacillus paracasei BD87E6 would be a valuable biocatalyst for the preparation of chiral aromatic alcohols of pharmaceutical interest.     

Bio-catalytic asymmetric synthesis of β-adrenergic receptor blocker precursor: (R)-2-bromo-1-(naphthalen-2-yl)ethanol

Abstract

Aromatic α-halohydrins, particularly 2-haloethanols as significant precursor of drugs, can easily be converted to chiral β-adrenergic receptor blockers. Eight strains of Lactobacillus curvatus were tested as biocatalysts for asymmetric reduction of 2-bromo-1-(naphthalen-2-yl)ethanone 1 to 2-bromo-1- (naphthalen-2-yl) ethanol 2. The parameters of the bioreduction were optimized using L. curvatus N4, the best biocatalyst found. As a result, (R)-2-bromo-1-(naphthalen-2-yl)ethanol 2, which can be β-adrenergic receptor blocker precursor, was produced for the first time in high yield and enantiomerically pure form using biocatalysts. Moreover, the gram scale synthesis was performed and 7.54?g of (R)-2 was synthesized as enantiopure form (enantiomeric excess >99%) in 48?h. The important advantages of this process are that it produces of (R)-2 for the first time in enantiopure form, in excellent yield and under environmentally friendly and moderate reaction conditions. This system is of the potential to be applied at a commercial scale.     

Synthesis of enantiopure (S)-6-chlorochroman-4-ol using whole-cell Lactobacillus paracasei biotransformation

Chromane, which has a fused cyclic structure, is a significant molecule that can be found in the structure of many important compounds. Lactobacillus paracasei BD101 was demonstrated as whole-cell biocatalyst for the synthesis of (S)-6-chlorochroman-4-ol with immense enantioselectivity. The conditions of asymmetric reduction were optimized one factor by one factor using L paracasei BD101 to achieve enantiomerically pure product and complete conversion. Using these obtained optimization conditions, asymmetric reduction of 6-chlorochroman-4-one was performed under environmentally friendly conditions; 6-chlorochroman-4-one, having a fused cyclic structure as previously noted to be difficult to asymmetric reduction with biocatalysts, was enantiomerically reduced to (S)-6-chlorochroman-4-ol with an enantiomeric excess >99% on a high gram scale. This study is the first example in the literature for the enantiopure synthesis of (S)-6-chlorochroman-4-ol using a biocatalyst. Also notably, the optical purity of (S)-6-chlorochroman-4-ol obtained in this study through asymmetric bioreduction using whole-cell biocatalyst is the highest value in the literature. In this study, (S)-6-chlorochroman-4-ol was produced on a gram scale by an easy, inexpensive, and environmentally friendly method, which has shown the production of valuable chiral precursors for drug synthesis and other industrial applications. This study provides a convenient method for the production of (S)-6-chlorochroman-4-ol, which can meet the industrial green production demand of this chiral secondary alcohol.     

Enzymatic resolution of homoallyllic alcohols using various Rhizopus species

The asymmetric resolution of various 1-aryl-3-buten-1-ols via microbial hydrolysis of the corresponding acetates has been investigated using different Rhizopus species. The chosen species, R. arrhizus (wild type), efficiently hydrolyzed 1-phenyl- and 1-para-substituted phenyl-3-buten-1-ol acetates, producing the enantiomerically pure (R)-alcohols with 53–65% yields. Although the antipode acetates were obtained with 9–52% enantiomeric excess, the (S)-alcohols were amenable in > 99% enantiomeric excess via a R. arrhizus mediated asymmetric reduction of the corresponding ketones.     

Isolation of a Bacillus strain producing ketone reductase with high substrate tolerance

Target reaction-oriented screening from soil samples yielded a ketone reductase-producing Bacillus sp., strain ECU0013, which exhibits excellent stereoselectivity, high substrate tolerance and is capable of regenerating the required cofactor with glucose as a co-substrate. Whole-cells catalyzed the asymmetric reduction of 2-chloro-1-phenylethanone (50 mM) to (R)-2-chloro-1-phenylethanol with a 93.3% conversion rate and 99% e.e. (enantiomeric excess). A variety of ketones were enantioselectively reduced by resting cells, giving corresponding chiral alcohols with good to excellent e.e. values. These results suggest the potential of this strain for the industrial production of chiral halogenated aromatic alcohols.     

Resolution,determination of enantiomeric purity and chiral recognition mechanism of new xanthone derivatives on (S,S)‐whelk‐O1 stationary phase

The enantioresolution and determination of the enantiomeric purity of 32 new xanthone derivatives, synthesized in enantiomerically pure form, were investigated on (S ,S )‐Whelk‐O1 chiral stationary phase (CSP). Enantioselectivity and resolution (α and R_S) with values ranging from 1.41–6.25 and from 1.29–17.20, respectively, were achieved. The elution was in polar organic mode with acetonitrile/methanol (50:50 v/v ) as mobile phase and, generally, the (R )‐enantiomer was the first to elute. The enantiomeric excess (ee ) for all synthesized xanthone derivatives was higher than 99%. All the enantiomeric pairs were enantioseparated, even those without an aromatic moiety linked to the stereogenic center. Computational studies for molecular docking were carried out to perform a qualitative analysis of the enantioresolution and to explore the chiral recognition mechanisms. The in silico results were consistent with the chromatographic parameters and elution orders. The interactions between the CSP and the xanthone derivatives involved in the chromatographic enantioseparation were elucidated.     

β‐Hydroxyamide‐Based Ligands and Their Use in the Enantioselective Borane Reduction of Prochiral Ketones

Hydroxyamide‐based ligands have occupied a considerable place in asymmetric synthesis. Here we report the synthesis of seven β‐hydroxyamide‐based ligands from the reaction of 2‐hydroxynicotinic acid with chiral amino alcohols and test their effect on the enantioselective reduction of aromatic prochiral ketones with borane in tetrahydofuran (THF). They produce the corresponding secondary alcohols with up to 76% enantiomeric excess (ee) and good to excellent yields (86‐99%). Chirality 26:21–26, 2013. © 2013 Wiley Periodicals, Inc.     

Green synthesis of enantiopure quinoxaline alcohols using Daucus carota

Green chemistry comprises a new approach in the synthesis of biologically active compounds using biocatalysts, thus diminishing the hazards for human health and environmental pollution. Asymmetric bioreduction is one of the most widely employed strategies in chemoenzymatic synthesis to produce enantiomerically pure chiral alcohols. The present study highlights the use biocatalyst Daucus carota for selective bioreduction of quinoxaline ketones 1a‐6a to their corresponding optically pure alcohols 1b‐6b in high yields (up to 84%) and good enantioselectivity (up to 98%). The absolute configuration of the chiral product (R)‐1‐(3‐methyl 7‐nitroquinoxalin‐2‐yl) ethan‐1‐ol 2b was confirmed by X‐ray crystallography studies. The chiral R‐configuration of the products obtained was confirmed by absolute configuration studies and was assigned following anti‐Prelogs rule. Quinoxaline pharmacophores form a part of well‐known potent drug molecules; hence, the chiral products were studied for determination of their molecular properties using SwissADME property analyser. All the chiral products show no Lipinski rule violations and are expected to have good oral bioavailability. As per the molecular properties prediction studies, the compound 6b (R)‐1‐(6,7‐dichloro‐3‐ methylquinoxalin‐2‐yl) ethanol is observed to show the best physicochemical properties to be a good lead molecule. Thus, the sustainable methodology was developed, and it confirms the synthesis of novel quinoxaline chiral alcohols in a simple, inexpensive, and eco‐friendly condition using D carota.     

Synthesis of optically pure 1,2-epoxypropane by microbial asymmetric reduction of chloroacetone

A number of bacteria and yeast was screened for asymmetric reduction of prochiral chloroacetone into chiral 1-chloro-2-propanol, which is chemically convertible into chiral 1,2-epoxypropane. In this way Rhodotorula glutinis produced optically pure S-1,2-epoxypropane with 98% enantiomeric excess and in a relatively high final concentration. The enzyme that catalysed the asymmetric reduction was an NAD(P)H-dependent alcohol dehydrogenase. Reduction of racemic 3-chloro-2-butanone resulted in mixtures of cis and trans-2,3-epoxybutane, indicating that no enantioselective reduction of this haloketone occurred. Correspondence to: C. A. G. M. Weijers     

Highly Enantioselective Production of Chiral Secondary Alcohols Using Lactobacillus paracasei BD101 as a New Whole Cell Biocatalyst and Evaluation of Their Antimicrobial Effects

Chiral secondary alcohols are valuable intermediates for many important enantiopure pharmaceuticals and biologically active molecules. In this work, we studied asymmetric reduction of aromatic ketones to produce the corresponding chiral secondary alcohols using lactic acid bacteria (LAB) as new biocatalysts. Seven LAB strains were screened for their ability to reduce acetophenones to their corresponding alcohols. Among these strains, Lactobacillus paracasei BD101 was found to be the most successful at reducing the ketones to the corresponding alcohols. The reaction conditions were further systematically optimized for this strain and high enantioselectivity (99%) and very good yields were obtained. These secondary alcohols were further tested for their antimicrobial activities against important pathogens and significant levels of antimicrobial activities were observed although these activities were altered depending on the secondary alcohols as well as their enantiomeric properties. The current methodology demonstrates a promising and alternative green approach for the synthesis of chiral secondary alcohols of biological importance in a cheap, mild, and environmentally useful process.     

Highly enantioselective bioreduction of prochiral ketones by stem and germinated plant of Brassica oleracea variety italica

Abstract

An eco-friendly and environmentally benign asymmetric reduction of a broad range of prochiral ketones employing Brassica oleracea variety italica (stems and germinated plant) as a novel biocatalyst was developed. It was found that B. oleracea variety italica could be used effectively for enantioselective bioreduction in aqueous medium with moderate to excellent chemical yield and enantiomeric excess (ee). This process is more efficient and generates less waste than conventional chemical reagents or microorganisms. Both R- and S-configurations were obtained by these asymmetric reactions. The best ee were achieved for pyridine derivatives (92–99%). The ee in germinated plant reactions were significantly higher than those of stem reactions. The low cost and the easy availability of these biocatalysts suggest their possible use for large scale preparations of important chiral alcohols.     

Chemical Composition,Enantiomeric Analysis,AEDA Sensorial Evaluation and Antifungal Activity of the Essential Oil from the Ecuadorian Plant Lepechinia mutica Benth (Lamiaceae)

This study describes the GC‐FID, GC/MS, GC‐O, and enantioselective GC analysis of the essential oil hydrodistilled from leaves of Lepechinica mutica (Lamiaceae), collected in Ecuador. GC‐FID and GC/MS analyses allowed the characterization and quantification of 79 components, representing 97.3% of the total sample. Sesquiterpene hydrocarbons (38.50%) and monoterpene hydrocarbons (30.59%) were found to be the most abundant volatiles, while oxygenated sesquiterpenes (16.20%) and oxygenated monoterpenes (2.10%) were the minor components. In order to better characterize the oil aroma, the most important odorants, from the sensorial point of view, were identified by Aroma Extract Dilution Analysis (AEDA) GC‐O. They were α‐Pinene, β‐Phellandrene, and Dauca‐5,8‐diene, exhibiting the characteristic woody, herbaceus, and earthy odors, respectively. Enantioselective GC analysis of L. mutica essential oil revealed the presence of twelve couples and two enantiomerically pure chiral monoterpenoids. Their enantiomeric excesses were from a few percent units to 100%. Moreover, the essential oil exhibited moderate in vitro activity against five fungal strains, being especially effective against M. canis, which is a severe zoophilic dermatophyte causal agent of pet and human infections.     

Optimizing the attractiveness of pheromone baits used for trapping the four‐eyed spruce bark beetle Polygraphus poligraphus

Bark beetles have caused extensive damage to forests in central Sweden during the past decade, and the four‐eyed spruce bark beetle, Polygraphus poligraphus, seems to be involved. However, its role in these bark beetle outbreaks is still not clear. The purpose of this study was to develop an efficient pheromone bait for P. poligraphus, which would make it possible to study the species more carefully and thereby contribute to protect exposed forests in an environmentally friendly way. Three field studies were conducted in 2015, 2016 and 2018 in Medelpad, county of Västernorrland, Sweden. The pheromone of P. poligraphus, (?)‐terpinen‐4‐ol, was tested at different release rates and in different enantiomeric purities, to find the most attractive formulation for the beetles. It was also tested in combination with racemic frontalin, a compound which has previously been shown to produce a synergistic effect together with (?)‐terpinen‐4‐ol of low enantiomeric purity; 52% ee. Other compounds, chosen based on responses from electroantennographic studies, were also tested in an attempt to find additional attractants and repellents for P. poligraphus. The most attractive treatment tested was enantiomerically pure (?)‐terpinen‐4‐ol (99% ee). When the enantiomeric purity was lower (50% ee), the trap catches was lowered to levels comparable to the catches for unbaited control traps. A strong synergistic effect with frontalin was observed for (?)‐terpinen‐4‐ol of low enantiomeric purity (50% ee) but not for the enantiomerically pure compound (99% ee). The release rate of (?)‐terpinen‐4‐ol (99% ee) was shown to be an important factor. For the combination of frontalin and (?)‐terpinen‐4‐ol (50% ee), the attraction seemed strongest when (?)‐terpinen‐4‐ol was released at a higher rate than frontalin. An interesting and novel result was that a repellent compound, α‐terpineol, was identified in our studies. Our results from field studies and electroantennography recordings also indicate that (+)‐terpinen‐4‐ol is a repellent for P. poligraphus.     

Comparative Study of Cyanobacteria as Biocatalysts for the Asymmetric Synthesis of Chiral Building Blocks

The three representative cyanobacteria, Synechococcus PCC7942, Anabaena variabilis, and Nostoc muscorum, were studied for their ability to asymmetrically reduce the prochiral ketones 2′‐3′‐4′‐5′‐6′‐pentafluoroacetophenone, ethyl 4‐chloroacetate, 4‐chloroacetophenone, and ethylbenzoylacetate to the corresponding chiral alcohols. Photosynthesis as well as respiration was applied for intracellular regeneration of the NAD(P)H cofactor. It was shown for the first time that all cyanobacteria were able to reduce the prochiral ketones asymmetrically without light for cofactor regeneration. By comparison of the cell specific product formation capacities of cyanobacteria with typical heterotrophic whole cell biocatalysts in batch processes, it is shown that comparable or, in some cases, better performances at high enantiomeric excess (ee > 99.8 %) are obtained. As a consequence of a generally strong product inhibition, in situ product removal must be applied in order to restore process efficiency when using cyanobacteria as biocatalysts.     

Green and scalable synthesis of chiral aromatic alcohols through an efficient biocatalytic system

Chiral aromatic alcohols have received much attention due to their widespread use in pharmaceutical industries. In the asymmetric synthesis processes, the excellent performance of alcohol dehydrogenase makes it a good choice for biocatalysts. In this study, a novel and robust medium-chain alcohol dehydrogenase RhADH from Rhodococcus R6 was discovered and used to catalyse the asymmetric reduction of aromatic ketones to chiral aromatic alcohols. The reduction of 2-hydroxyacetophenone (2-HAP) to (R)-(-)-1-phenyl-1,2-ethanediol ((R)-PED) was chosen as a template to evaluate its catalytic activity. A specific activity of 110 U mg⁻¹ and a 99% purity of e.e. was achieved in the presence of NADH. An efficient bienzyme-coupled catalytic system (RhADH and formate dehydrogenase, CpFDH) was established using a two-phase strategy (dibutyl phthalate and buffer), which highly raised the tolerated substrate concentration (60 g l⁻¹). Besides, a broad range of aromatic ketones were enantioselectively reduced to the corresponding chiral alcohols by this enzyme system with highly enantioselectivity. This system is of the potential to be applied at a commercial scale.     

The resolution of acyclic P‐stereogenic phosphine oxides via the formation of diastereomeric complexes: A case study on ethyl‐(2‐methylphenyl)‐phenylphosphine oxide

As an example of acyclic P‐chiral phosphine oxides, the resolution of ethyl‐(2‐methylphenyl)‐phenylphosphine oxide was elaborated with TADDOL derivatives, or with calcium salts of the tartaric acid derivatives. Besides the study on the resolving agents, several purification methods were developed in order to prepare enantiopure ethyl‐(2‐methylphenyl)‐phenylphosphine oxide. It was found that the title phosphine oxide is a racemic crystal‐forming compound, and the recrystallization of the enantiomeric mixtures could be used for the preparation of pure enantiomers. According to our best method, the (R)‐ethyl‐(2‐methylphenyl)‐phenylphosphine oxide could be obtained with an enantiomeric excess of 99% and in a yield of 47%. Complete racemization of the enantiomerically enriched phosphine oxide could be accomplished via the formation of a chlorophosphonium salt. Characterization of the crystal structures of the enantiopure phosphine oxide was complemented with that of the diastereomeric intermediate. X‐ray analysis revealed the main nonbonding interactions responsible for enantiomeric recognition.     

Functional characterization of salt‐tolerant microbial esterase WDEst17 and its use in the generation of optically pure ethyl (R)‐3‐hydroxybutyrate

The two enantiomers of ethyl 3‐hydroxybutyrate are important intermediates for the synthesis of a great variety of valuable chiral drugs. The preparation of chiral drug intermediates through kinetic resolution reactions catalyzed by esterases/lipases has been demonstrated to be an efficient and environmentally friendly method. We previously functionally characterized microbial esterase PHE21 and used PHE21 as a biocatalyst to generate optically pure ethyl (S)‐3‐hydroxybutyrate. Herein, we also functionally characterized one novel salt‐tolerant microbial esterase WDEst17 from the genome of Dactylosporangium aurantiacum subsp. Hamdenensis NRRL 18085. Esterase WDEst17 was further developed as an efficient biocatalyst to generate (R)‐3‐hydroxybutyrate, an important chiral drug intermediate, with the enantiomeric excess being 99% and the conversion rate being 65.05%, respectively, after process optimization. Notably, the enantio‐selectivity of esterase WDEst17 was opposite than that of esterase PHE21. The identification of esterases WDEst17 and PHE21 through genome mining of microorganisms provides useful biocatalysts for the preparation of valuable chiral drug intermediates.