Electrochemical Enantioselective Nickel-Catalyzed Cross-Coupling of Aldehydes with Aryl Iodides.

The preparation of enantioenriched diarylmethanol derivatives is described using nickel-catalyzed electrochemical cross-couplings between various alkyl/aryl aldehydes and aryl iodides. Performed in an electrochemical cell equipped with an iron anode and a nickel cathode, this electrocatalytic variant led to the scalemic targeted products in the presence of 2,2-bis((4R,5S)-4,5-diphenyl-4,5-dihydrooxazol-2-yl)acetonitrile (L2), as enantiopure cyano-bis(oxazoline) ligand. X-ray structure analysis of a pre-catalyst, for instance the [Ni(II)(L2)2] complex, with L2 as an anionic bisoxazolinate ligand, confirms the chemical formulation of one nickel surrounded by two ligands. The redox behavior of the new Ni complexes generated in situ was first assessed by cyclic voltammetry showing a redox wave at ca. -1.5 V that can be assigned to the two-electron reduction of the Ni(II) center to the Ni(0) state. Oxidative addition between the electrogenerated Ni(0) complex and aryl iodide was evidenced. An intense current was observed in presence of a mixture of the two substrates pertaining an electrocatalytic process. Interestingly, we found that the sacrificial iron anode plays a crucial role in the catalytic mechanism.

Biocatalytic approach for the synthesis of chiral alcohols for the development of pharmaceutical intermediates and other industrial applications: A review.

Biocatalysis has emerged as a strong tool for the synthesis of active pharmaceutical ingredients (APIs). In the early twentieth century, whole cell biocatalysis was used to develop the first industrial biocatalytic processes, and the precise work of enzymes was unknown. Biocatalysis has evolved over the years into an essential tool for modern, cost-effective, and sustainable pharmaceutical manufacturing. Meanwhile, advances in directed evolution enable the rapid production of process-stable enzymes with broad substrate scope and high selectivity. Large-scale synthetic pathways incorporating biocatalytic critical steps towards >130 APIs of authorized pharmaceuticals and drug prospects are compared in terms of steps, reaction conditions, and scale with the corresponding chemical procedures. This review is designed on the functional group developed during the reaction forming alcohol functional groups. Some important biocatalyst sources, techniques, and challenges are described. A few APIs and their utilization in pharmaceutical drugs are explained here in this review. Biocatalysis has provided shorter, more efficient, and more sustainable alternative pathways toward existing small molecule APIs. Furthermore, non-pharmaceutical applications of biocatalysts are also mentioned and discussed. Finally, this review includes the future outlook and challenges of biocatalysis. In conclusion, Further research and development of promising enzymes are required before they can be used in industry.

Rational redesign of the loop dynamics of carbonyl reductase LfSDR1 to improve the stereoselectivity for asymmetric synthesis of bulky chiral alcohols.

Engineering biocatalysts with enhanced stereoselectivity is highly desirable, and active-site loop dynamics play an important role in its regulation. However, knowledge of their precise roles in catalysis and evolution is limited. Here, we used the strategy of Rosetta enzyme design combined molecular dynamic simulations (MDs) to reprogram the landscapes of the key active-site loop dynamics of the carbonyl reductase LfSDR1 to improve stereoselectivity. The key flexible loop in the active site showed the potential to regulate the catalytic properties. A library of virtual variants was produced using the Rosetta design and assessed dynamic effect of the loop with the aid of MDs. A potential candidate was obtained with significant stereoselectivity (ee > 99 %) compared to the wild-type (ee = 42 %) without loss of catalytic activity or thermostability. The molecular basis of the catalytic property enhancement was flanked by MDs, which revealed the role of the G92L mutation in regulating loop dynamics to stabilize the environment of the active site. Finally, a series of the challenge bulky substrate derivatives were assessed using the G92L variant, and all showed improved stereoselectivity ee > 99 %. This study provides novel insights for improving stereoselectivity through rational engineering of the loop dynamics of biocatalysts.

Optical Relay Sensing of Cryptochiral Alcohols Displaying α-, ß-, γ- and δ-Stereocenters or Chirality by Virtue of Isotopic Substitution.

A reaction-based optical relay sensing strategy that enables accurate determination of the concentration and enantiomeric ratio (er) of challenging chiral alcohols exhibiting stereocenters at the α-, ß-, γ- or even δ-position or hard-to-detect cryptochirality arising from H/D substitution is described. This unmatched application scope is achieved with a conceptually new sensing approach by which the alcohol moiety is replaced with an optimized achiral sulfonamide chromophore to minimize the distance between the covalently attached chiroptical reporter unit and the stereogenic center in the substrate. The result is a remarkably strong, red-shifted CD induction that increases linearly with the sample er. The CD sensing part of the tandem assay is seamlessly coupled to a redox reaction with a quinone molecule to generate a characteristic UV response that is independent of the enantiopurity of the alcohol and thus allows determination of the total analyte concentration. The robustness and utility of the CD/UV relay are further verified by chromatography-free asymmetric reaction analysis with small aliquots of crude product mixtures, paving the way toward high-throughput chiral compound screening workflows which is a highly sought-after goal in the pharmaceutical industry.

Merging High-Pressure Syngas Metal Catalysis and Biocatalysis in Tandem One-Pot Processes for the Synthesis of Nonchiral and Chiral Alcohols from Alkenes in Water.

While simultaneously proceeding reactions are among the most fascinating features of biosynthesis, this concept of tandem processes also offers high potential in the chemical industry in terms of less waste production and improved process efficiency and sustainability. Although examples of one-pot chemoenzymatic syntheses exist, the combination of completely different reaction types is rare. Herein, we demonstrate that extreme "antipodes" of the "worlds of catalysis", such as syngas-based high-pressure hydroformylation and biocatalyzed reduction, can be combined within a tandem-type one-pot process in water. No significant deactivation was found for either the biocatalyst or the chemocatalyst. A proof-of-concept for the one-pot process starting from 1-octene was established with >99 % conversion and 80 % isolated yield of the desired alcohol isomers. All necessary components for hydroformylation and biocatalysis were added to the reactor from the beginning. This concept has been extended to the enantioselective synthesis of chiral products by conducting the hydroformylation of styrene and an enzymatic dynamic kinetic resolution in a tandem mode, leading to an excellent conversion of >99 % and an enantiomeric ratio of 91 : 9 for (S)-2-phenylpropanol. The overall process runs in water under mild and energy-saving conditions, without any need for intermediate isolation.

Co-Immobilization of ADH and GDH on Metal-Organic-Framework: An Effective Biocatalyst for Asymmetric Reduction of Ketones.

Chiral alcohols are not only important building blocks of various bioactive natural compounds and pharmaceuticals, but can serve as synthetic precursors for other valuable organic chemicals, thus the synthesis of these products is of great importance. Bio-catalysis represents one effective way to obtain these molecules, however, the weak stability and high cost of enzymes often hinder its broad application. In this work, we designed a biological nanoreactor by embedding alcohol dehydrogenase (ADH) and glucose dehydrogenase (GDH) in metal-organic-framework ZIF-8. The biocatalyst ADH&GDH@ZIF-8 could be applied to the asymmetric reduction of a series of ketones to give chiral alcohols in high yields (up to 99 %) and with excellent enantioselectivities (>99 %). In addition, the heterogeneous biocatalyst could be recycled and reused at least four times with slight activity decline. Moreover, E. coli containing ADH and GDH was immobilized by ZIF-8 to form biocatalyst E. coli@ZIF-8, which also exhibits good catalytic behaviours. Finally, the chiral alcohols are further converted to marketed drugs (R)-Fendiline, (S)-Rivastigmine and NPS R-568 respectively.

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.

Biocatalytic hydrogen-transfer to access enantiomerically pure proxyphylline, xanthinol, and diprophylline.

Alcohol dehydrogenases (ADHs; EC 1.1.1.1) have been widely used for the reversible redox reactions of carbonyl compounds (i.e., aldehydes and ketones) and primary or secondary alcohols, often resulting in optically pure hydroxyl products with high added value. In this work, we report a concise chemoenzymatic route toward xanthine-based enantiomerically pure active pharmaceutical ingredients (API) - proxyphylline, xanthinol, and diprophylline employing various recombinant short-chain ADHs with (R)- or (S)-selectivity as key biocatalysts. By choosing the appropriate ADH, the (R)- as well as the (S)-enantiomer of proxyphylline was prepared in excellent enantiomeric excess (99-99.9% ee), >99% conversion, and the isolated yield ranging from 65% to 74%, depending on the used biocatalyst (ADH-A from Rhodococcus ruber or a variant derived from Lactobacillus kefir, Lk-ADH-Lica). In turn, E. coli/ADH-catalyzed bioreduction of the carbonylic precursor of xanthinol and diprophylline furnished the corresponding (S)-chlorohydrin in >99% ee, >99% conversion, and 80% yield (in the case of Lk-ADH-Lica); while the (R)-counterpart was afforded in 94% ee, 64% conversion, and 41% yield (in the case of SyADH from Sphingobium yanoikuyae). After further chemical functionalization of the key (S)-chlorohydrin intermediate, the desired homochiral (R)-xanthinol (>99% ee) was obtained in 97% yield and (S)-diprophylline (>99% ee) in 90% yield. The devised biocatalytic method is straightforward and thus might be considered practical in the manufacturing of title pharmaceuticals.

Ru-Catalyzed Asymmetric Addition of Arylboronic Acids to Aliphatic Aldehydes via P-Chiral Monophosphorous Ligands.

Chiral alcohols are among the most widely applied in fine chemicals, pharmaceuticals and agrochemicals. Herein, the Ru-monophosphine catalyst formed in situ was found to promote an enantioselective addition of aliphatic aldehydes with arylboronic acids, delivering the chiral alcohols in excellent yields and enantioselectivities and exhibiting a broad scope of aliphatic aldehydes and arylboronic acids. The enantioselectivities are highly dependent on the monophosphorous ligands. The utility of this asymmetric synthetic method was showcased by a large-scale transformation.

Nickel-Catalyzed Intermolecular Asymmetric Addition of Aryl Iodides across Aldehydes.

Enantioenriched alcohols comprise much of the framework of organic molecules. Here, we first report that chiral nickel complexes can catalyze the intermolecular enantioselective addition of aryl iodides across aldehydes to provide diverse optically active secondary alcohols using zinc metal as the reducing agent. This method shows a broad substrate scope under mild reaction conditions and precludes the traditional strategy through the pre-generation of organometallic reagents. Mechanistic studies indicate that an in situ formed arylnickel, instead of an arylzinc, adds efficiently to aldehydes, forming a new C-C bond and a chiral nickel alkoxide that may be turned over by zinc powder.

Evolutionary coupling-inspired engineering of alcohol dehydrogenase reveals the influence of distant sites on its catalytic efficiency for stereospecific synthesis of chiral alcohols.

Alcohol dehydrogenase (ADH) has attracted much attention due to its ability to catalyze the synthesis of important chiral alcohol pharmaceutical intermediates with high stereoselectivity. ADH protein engineering efforts have generally focused on reshaping the substrate-binding pocket. However, distant sites outside the pocket may also affect its activity, although the underlying molecular mechanism remains unclear. The current study aimed to apply evolutionary coupling-inspired engineering to the ADH CpRCR and to identify potential mutation sites. Through conservative analysis, phylogenic analysis and residues distribution analysis, the co-evolution hotspots Leu34 and Leu137 were confirmed to be highly evolved under the pressure of natural selection and to be possibly related to the catalytic function of the protein. Hence, Leu34 and Leu137, far away from the active center, were selected for mutation. The generated CpRCR-L34A and CpRCR-L137V variants showed high stereoselectivity and 1.24-7.81 fold increase in k cat /K m value compared with that of the wild type, when reacted with 8 aromatic ketones or ß-ketoesters. Corresponding computational study implied that L34 and L137 may extend allosteric fluctuation in the protein structure from the distal mutational site to the active site. Moreover, the L34 and L137 mutations modified the pre-reaction state in multiple ways, in terms of position of the hydride with respect to the target carbonyl. These findings provide insights into the catalytic mechanism of the enzyme and facilitate its regulation from the perspective of the site interaction network.

Visible-Light-Driven Catalytic Deracemization of Secondary Alcohols.

Deracemization of racemic chiral compounds is an attractive approach in asymmetric synthesis, but its development has been hindered by energetic and kinetic challenges. Here we describe a catalytic deracemization method for secondary benzylic alcohols which are important synthetic intermediates and end products for many industries. Driven by visible light only, this method is based on sequential photochemical dehydrogenation followed by enantioselective thermal hydrogenation. The combination of a heterogeneous dehydrogenation photocatalyst and a chiral molecular hydrogenation catalyst is essential to ensure two distinct pathways for the forward and reverse reactions. These reactions convert a large number of racemic aryl alkyl alcohols into their enantiomerically enriched forms in good yields and enantioselectivities.

The revenge of Zygosaccharomyces yeasts in food biotechnology and applied microbiology.

Non-conventional yeasts refer to a huge and still poorly explored group of species alternative to the well-known model organism Saccharomyces cerevisiae. Among them, Zygosaccharomyces rouxii and the sister species Zygosaccharomyces bailii are infamous for spoiling food and beverages even in presence of several food preservatives. On the other hand, their capability to cope with a wide range of process conditions makes these yeasts very attractive factories (the so-called "ZygoFactories") for bio-converting substrates poorly permissive for the growth of other species. In balsamic vinegar Z. rouxii is the main yeast responsible for converting highly concentrated sugars into ethanol, with a preference for fructose over glucose (a trait called fructophily). Z. rouxii has also attracted much attention for the ability to release important flavor compounds, such as fusel alcohols and the derivatives of 4-hydroxyfuranone, which markedly contribute to fragrant and smoky aroma in soy sauce. While Z. rouxii was successfully proposed in brewing for producing low ethanol beer, Z. bailii is promising for lactic acid and bioethanol production. Recently, several research efforts exploited omics tools to pinpoint the genetic bases of distinctive traits in "ZygoFactories", like fructophily, tolerance to high concentrations of sugars, lactic acid and salt. Here, I provided an overview of Zygosaccharomyces industrially relevant phenotypes and summarized the most recent findings in disclosing their genetic bases. I suggest that the increasing number of genomes available for Z. rouxii and other Zygosaccharomyces relatives, combined with recently developed genetic engineering toolkits, will boost the applications of these yeasts in biotechnology and applied microbiology.

Highly Active Cooperative Lewis Acid-Ammonium Salt Catalyst for the Enantioselective Hydroboration of Ketones.

Enantiopure secondary alcohols are fundamental high-value synthetic building blocks. One of the most attractive ways to get access to this compound class is the catalytic hydroboration. We describe a new concept for this reaction type that allowed for exceptional catalytic turnover numbers (up to 15 400), which were increased by around 1.5-3 orders of magnitude compared to the most active catalysts previously reported. In our concept an aprotic ammonium halide moiety cooperates with an oxophilic Lewis acid within the same catalyst molecule. Control experiments reveal that both catalytic centers are essential for the observed activity. Kinetic, spectroscopic and computational studies show that the hydride transfer is rate limiting and proceeds via a concerted mechanism, in which hydride at Boron is continuously displaced by iodide, reminiscent to an SN 2 reaction. The catalyst, which is accessible in high yields in few steps, was found to be stable during catalysis, readily recyclable and could be reused 10 times still efficiently working.

Computational design of highly stable and soluble alcohol dehydrogenase for NADPH regeneration.

Nicotinamide adenine dinucleotide phosphate (NADPH), as a well-known cofactor, is widely used in the most of enzymatic redox reactions, playing an important role in industrial catalysis. However, the absence of a comparable method for efficient NADP+ to NADPH cofactor regeneration radically impairs efficient green chemical synthesis. Alcohol dehydrogenase (ADH) enzymes, allowing the in situ regeneration of the redox cofactor NADPH with high specific activity and easy by-product separation process, are provided with great industrial application potential and research attention. Accordingly, herein a NADP+-specific ADH from Clostridium beijerinckii was selected to be engineered for cofactor recycle, using an automated algorithm named Protein Repair One-stop Shop (PROSS). The mutant CbADH-6M (S24P/G182A/G196A/H222D/S250E/S254R) exhibited a favorable soluble and highly active expression with an activity of 46.3 U/mL, which was 16 times higher than the wild type (2.9 U/mL), and a more stable protein conformation with an enhanced thermal stability: Δ T 1 / 2 60 min = + 3.6 °C (temperature of 50% inactivation after incubation for 60 min). Furthermore, the activity of CbADH-6M was up-graded to 2401.8 U/mL by high cell density fermentation strategy using recombinant Escherichia coli, demonstrating its industrial potential. Finally, the superb efficiency for NADPH regeneration of the mutant enzyme was testified in the synthesis of some fine chiral aromatic alcohols coupling with another ADH from Lactobacillus kefir (LkADH).

Combined Photoredox/Enzymatic C-H Benzylic Hydroxylations.

Chemical transformations that install heteroatoms into C-H bonds are of significant interest because they streamline the construction of value-added small molecules. Direct C-H oxyfunctionalization, or the one step conversion of a C-H bond to a C-O bond, could be a highly enabling transformation due to the prevalence of the resulting enantioenriched alcohols in pharmaceuticals and natural products,. Here we report a single-flask photoredox/enzymatic process for direct C-H hydroxylation that proceeds with broad reactivity, chemoselectivity and enantioselectivity. This unified strategy advances general photoredox and enzymatic catalysis synergy and enables chemoenzymatic processes for powerful and selective oxidative transformations.

A facile lipase-catalyzed KR approach toward enantiomerically enriched homopropargyl alcohols.

Compounds possessing propargylic (prop-2-ynylic) system are very important building blocks for organic chemistry. Among them, preparation of enantiomeric homopropargyl alcohols (but-3-yn-1-ols) constitutes a key-challenge for asymmetric synthesis and thus drawn tremendous attention from the synthetic community in the last few decades. In this work, the catalytic performance of a set of commercial lipases has been investigated for enantioselective transesterification of 1-phenylhomopropargylic alcohols under kinetically-controlled conditions. Lipase from Burckholderia cepacia (BCL) immobilized either on ceramic (Amano PS-C II) or diatomaceous earth (Amano PS-IM) turned out to be the most active and enantioselective enzyme preparations (E ≫ 500) furnishing both resolution products of the racemic 1-phenylbut-3-yn-1-ol in highly enantiomerically enriched form (up > 99% ee). Variable reaction parameters, such as the acyl-group donor reagent as well as solvent, were additionally screened to establish their impact on the stereochemical outcome. For optimal biocatalytic systems established with model substrate, the enzymatic transformations were extended toward preparative-scale KR of 8 other differently para-phenyl-substituted homopropargylic sec-alcohols, which resulted in the synthesis of (S)-alcohols (96-100% ee) and the respective (R)-acetates (92-100% ee) in 19-44% yield, accordingly. Additionally, the crystal structure of (1R)-1-(4-nitrophenyl)but-3-yn-1-yl acetate has been evaluated for the first time and helped to assess stereopreference of the studied BCL.

Enantioselective synthesis of enantiopure chiral alcohols using carbonyl reductases screened from Yarrowia lipolytica.

AIMS: We aimed to explore Yarrowia lipolytica carbonyl reductases as effective biocatalysts and to develop efficient asymmetric reduction systems for chiral alcohol synthesis. METHODS AND RESULTS: Yarrowia lipolytica carbonyl reductase genes were obtained via homologous sequence amplification strategy. Two carbonyl reductases, YaCRI and YaCRII, were identified and characterized, and used to catalyse the conversion of 2-hydroxyacetophenone (2-HAP) to optically pure (S)-1-phenyl-1,2-ethanediol. Enzymatic assays revealed that YaCRI and YaCRII exhibited specific activities of 6·96 U mg-1 (99·8% e.e.) and 7·85 U mg-1 (99·9% e.e.), respectively, and showed moderate heat resistance at 40-50°C and acid tolerance at pH 5·0-6·0. An efficient whole-cell two-phase system was established using reductase-expressing recombinant Escherichia coli. The conversion of 2-HAP (20·0 g l-1 ) conversion with the solvent of dibutyl phthalate was approximately 70-fold higher than in water. Furthermore, the two recombinant E. coli displayed biocatalyst activity and enantioselectivity towards several different carbonyl compounds, and E. coli BL21 (DE3)/pET-28a-yacrII showed a broad substrate spectrum. CONCLUSIONS: A new whole-cell recombinant E. coli-based bioreduction system for enantiopure alcohol synthesis with high enantioselectivity at high substrate concentrations was developed. SIGNIFICANCE AND IMPACT OF THE STUDY: We proposed a promising approach for the efficient preparation of enantiopure chiral alcohols.

Generation of novel family of reductases from PCR based library for the synthesis of chiral alcohols and amines.

Biocatalysis has shown tremendous potential in the synthesis of drugs and drug intermediates in the last decade. Screening of novel biocatalysts from the natural genome space is the growing trend to replenish the harsh chemical synthetic routes, commonly used in the pharmaceutical and chemical industry. Here, we report a novel ketoreductase (KERD) and a nitrile reductase isolated from the PCR based library generated from the genome of Rhodococcus ruber and Bacillus subtilis, respectively. Both the proteins are hypothetical in nature as there is no putative homology found in the database, although both the enzymes have significant activity towards the synthesis of chiral alcohols and amines. Enzyme activity over a wide range of substrates (aromatic and aliphatic) for both the novel catalysts was observed. From the unique gene sequence to activity over a broad range of substrate and >99% conversion at higher concentrations (100 mM and above) entitles both the hypothetical enzymes as novel. The novel KERD has shown >99% selectivity for the synthesis of (S)-phenylethanol which makes it a potential candidate for industrial catalysis. The novel nitrile reductase has also shown promising activity for the synthesis of (R)-2-phenylethanolamine, which is a difficult moiety to synthesize chemically. In this report, starting from a homology based library, two highly potent whole cell biocatalysts are obtained.

A Comparative Study on Asymmetric Reduction of Ketones Using the Growing and Resting Cells of Marine-Derived Fungi.

Whole-cell biocatalysts offer a highly enantioselective, minimally polluting route to optically active alcohols. Currently, most of the whole-cell catalytic performance involves resting cells rather than growing cell biotransformation, which is one-step process that benefits from the simultaneous growth and biotransformation, eliminating the need for catalysts preparation. In this paper, asymmetric reduction of 14 aromatic ketones to the corresponding enantiomerically pure alcohols was successfully conducted using the growing and resting cells of marine-derived fungi under optimized conditions. Good yields and excellent enantioselectivities were achieved with both methods. Although substrate inhibition might be a limiting factor for growing cell biotransformation, the selected strain can still completely convert 10-mM substrates into the desired products. The resting cell biotransformation showed a capacity to be recycled nine times without a significant decrease in the activity. This is the first study to perform asymmetric reduction of ketones by one-step growing cell biotransformation.