- Efficient synthesis of a chiral precursor for angiotensin-converting enzyme (ace) inhibitors in high space-time yield by a new reductase without external cofactors
-
A new reductase, CgKR2, with the ability to reduce ethyl 2-oxo-4-phenylbutyrate (OPBE) to ethyl (R)-2-hydroxy-4-phenylbutyrate ((R)-HPBE), an important chiral precursor for angiotensin-converting enzyme (ACE) inhibitors, was discovered. For the first time, (R)-HPBE with >99% ee was produced via bioreduction of OPBE at 1 M without external addition of cofactors. The space-time yield (700 g·L-1·d -1) was 27 times higher than the highest record.
- Shen, Nai-Dong,Ni, Yan,Ma, Hong-Min,Wang, Li-Juan,Li, Chun-Xiu,Zheng, Gao-Wei,Zhang, Jie,Xu, Jian-He
-
-
Read Online
- A practical asymmetric synthesis of a 1,7-enyne A-ring synthon en route toward the total synthesis of vitamin D3 analogues
-
A concise synthesis of a key A-ring synthon of 1α,25-dihydroxyvitamin D3, 1, has been achieved in 8 steps starting from readily available, inexpensive ethyl-4-chloroacetoacetate. This synthon serves as one of two main coupling partners in our previously developed Pd-catalyzed alkylative enyne cyclization leading toward the total synthesis of la,25-dihydroxyvitamin D3 and potentially useful analogues.
- Trost, Barry M.,Hanson, Paul R.
-
-
Read Online
- Stereoselective reduction of α- and β-keto esters with aerobic thermophiles, Bacillus strains
-
The first example of stereoselective reduction with aerobic thermophiles is reported. Various α- and β-keto esters were reduced stereoselectively to the corresponding alcohols by the aerobic thermophiles, Bacillus strains. In particular, the reduction of ethyl 3-methyl-2-oxobutanoate with B. stearothermophilus DSM 297 gave the corresponding (R)-alcohol with high yield in excellent enantioselectively (> 99% e.e.). The conversions of keto esters to the corresponding hydroxy esters with Bacillus strains were increased by introduction of glycerol in the reaction mixture as an additive.
- Ishihara, Kohji,Iwai, Keisuke,Yamaguchi, Hitomi,Nakajima, Nobuyoshi,Nakamura, Kaoru,Ohshima, Toshihisa
-
-
Read Online
- A NEW METHOD FOR STEREOCHEMICAL CONTROL OF MICROBIAL REDUCTION. REDUCTION OF β-KETO ESTERS WITH BAKERS' YEAST IMMOBILIZED BY MAGNESIUM ALGINATE
-
Yeast reduction of methyl 3-oxopentanoate gives the L-hydroxy ester when bakers' yeast is immobilized by magnesium alginate and the reaction is run under a high concentration of magnesium ion.The D-hydroxy ester is obtained under normal reaction conditions.
- Nakamura, Kaoru,Kawai, Yasushi,Oka, Shinzaburo,Ohno, Atsuyoshi
-
-
Read Online
- Preparation of structurally diverse chiral alcohols by engineering ketoreductase CgKR1
-
Ketoreductases are tools for the synthesis of chiral alcohols in industry. However, the low activity of natural enzymes often restricts their use in industrial applications. On the basis of computational analysis and previous reports, two residues (F92 and F94) probably affecting the activity of ketoreductase CgKR1 were identified. By tuning these two residues, the CgKR1-F92C/F94W variant was obtained that exhibited higher activity toward all 28 structurally diverse substrates examined than the wild-type enzyme. Among them, 13 substrates have a specific activity over 50 U mg-1 (54-775 U mg-1). Using CgKR1-F92C/F94W as a catalyst, five substrates at high loading (>100 g-1 L-1) were reduced completely in gramscale preparative reactions. This approach provides accesses to pharmaceutically relevant chiral alcohols with high enantioselectivity (up to 99.0% ee) and high space-time yield (up to 583 g-1 L-1 day-1). Molecular dynamics simulations highlighted the crucial role of residues 92 and 94 in activity improvement. Our findings provide useful guidance for engineering other ketoreductases, especially those possessing a similar active pocket to that in CgKR1.
- Zheng, Gaowei,Liu, Yuan-Yang,Chen, Qi,Huang, Lei,Yu, Hui-Lei,Lou, Wen-Yong,Li, Chun-Xiu,Bai, Yun-Peng,Li, Ai-Tao,Xu, Jian-He
-
-
Read Online
- Practical Enzymatic Route to Optically Active 3-Hydroxyamides. Synthesis of 1,3-Aminoalcohols
-
Candida antarctica lipase (CAL) is a very efficient catalyst for the enantioselective aminolysis of different racemic 3-hydroxyesters with aliphatic amines.The degree of enantioselectivity exhibited by the lipase depends on the substrate and nucleophile, but in the most cases, the E values obtained are very satisfactory.CAL also catalyzes the aminolysis of ethyl (+/-)-3,4-epoxybutyrate and the epoxyamide was achieved with high e. e.The chemical reduction of the 3-hydroxyamides obtained by enzymatic aminolysis yields the corresponding 1,3-aminoalcohols.
- Garcia, Maria Jesus,Rebolledo, Francisca,Gotor, Vicente
-
-
Read Online
- Efficient synthesis of an ε-hydroxy ester in a space-time yield of 1580 g L-1 d-1 by a newly identified reductase RhCR
-
A new NADH-dependent carbonyl reductase RhCR capable of efficiently reducing the ε-ketoester ethyl 8-chloro-6-oxooctanoate (ECOO) to give ethyl (S)-8-chloro-6-hydroxyoctanoate [(S)-ECHO], an important chiral precursor for the synthesis of (R)-α-lipoic acid, was identified from Rhodococcus sp. ECU1014. Using recombinant Escherichia coli cells expressing RhCR and glucose dehydrogenase used for the regeneration of cofactor, 440 g L-1 (2 M) of ECOO were stoichiometrically converted to (S)-ECHO in a space-time yield of 1580 g L-1 d-1 without the external addition of any expensive cofactor.
- Chen, Rui-Jie,Zheng, Gao-Wei,Ni, Yan,Zeng, Bu-Bing,Xu, Jian-He
-
-
Read Online
- Lipase-catalyzed asymmetric synthesis of 6-(3-chloro-2-hydroxypropyl)-1,3-dioxin-4-ones and their conversion to chiral 5,6-epoxyhexanoates
-
Highly enantioselective syntheses of (R)- and (S)-6-(3-chloro-2-hydroxypropyl)-1,3-dioxin-4-ones by means of lipase-catalyzed kinetic resolutions are described. Chiral dioxinones thus obtained have been converted to optically active 5,6-epoxyhexanoates, which are important precursors for a series of biologically active compounds.
- Sakaki,Sakoda,Sugita,Sato,Kaneko
-
-
Read Online
- Baker's yeast: Improving the D-stereoselectivity in reduction of 3-oxo esters
-
The stereoselectivity of baker's yeast in the reduction of ethyl 3- oxopentanoate was shifted towards the corresponding (R)-hydroxy ester by sugar, heat treatment and allyl alcohol. The highest enantiomeric excesses obtained with baker's yeast with a good reduction capacity, 92-97%, were achieved by combining allyl alcohol and sugar; heat treatment did not increase the stereoselectivity further. With the use of this technique, ethyl (R)-3-hydroxyhexanoate, >99% ee, and ethyl (S)-4-chloro-3-hydroxybutanoate, 82-90% ee, were produced from the corresponding esters, and for the first time an excess of the (R)-enantiomer of ethyl 3-hydroxybutanoate was obtained with ordinary baker's yeast.
- Dahl, Allan C.,Fjeldberg, Morten,Madsen, Jorgen gaard
-
-
Read Online
- Stereochemical Control in Microbial Reduction. 8. Stereochemical Control in Microbial Reduction of β-Keto Esters
-
Stereochemistry of the reduction of β-keto esters with bakers' yeast is controlled by the addition of a certain α,β-unsaturated carbonyl compound (or its reduced form).Glucose also exerts the same effect.The additives tend to shift the stereochemistry of the reduction toward the production of D-hydroxy ester.Namely, methyl 3-oxopentanoate and ethyl 3-oxo-6-heptenoate were reduced to the corresponding D-hydroxy esters with excellent stereoselectivities and chemical yields.The enones are supposed to inhibit the enzymes that produce the L-hydroxy ester, whereas glucose plays a role to activate the enzymes that produce the D-hydroxy ester.
- Nakamura, Kaoru,Kawai, Yasushi,Oka, Shinzaburo,Ohno, Atsuyoshi
-
-
Read Online
- Stereoselective reduction of alkyl 3-oxobutanoate by carbonyl reductase from Candida magnoliae
-
The enantioselective reduction of alkyl 3-oxobutanoates by carbonyl reductase (S1) from Candida magnoliae was investigated. S1 reduced alkyl 4-halo-3-oxobutanoates to the corresponding enantiomerically pure (S)-3-hydroxy esters. Escherichia coli HB101 transformant co-overproducing the S1 and glucose dehydrogenase from Bacillus megaterium, produced optically pure alkyl 4-substituted-3-hydroxybutanoates in a two-phase water/organic solvent system.
- Yasohara, Yoshihiko,Kizaki, Noriyuki,Hasegawa, Junzo,Wada, Masaru,Kataoka, Michihiko,Shimizu, Sakayu
-
-
Read Online
- Synthesis of a chiral alcohol using a rationally designed Saccharomyces cerevisiae reductase and a NADH cofactor regeneration system
-
Ethyl (S)-4-chloro-3-hydroxy butanoate (ECHB) is a building block for the synthesis of hypercholesterolemia drugs that function as 3-hydroxy-3- methylglutaryl coenzyme A reductase inhibitors. Yeast reductase YDL124W has been shown to convert ethyl 4-chloro-3-oxo butanoate (ECOB) into (S)-ECHB in an enantioselective manner. In that reduction, YDL124W absolutely prefers NADPH as a cofactor to NADH. NADPH is, however, much more expensive and unstable than NADH. In this study, four amino acid residues that directly interact with the 2′-phosphate group of adenosine ribose of NADPH were identified based on the homology model and docking model of YDL124W with ECOB and NADPH. Among various single mutants altered in these target residues, R264H evidenced noticeably increased reductase activity with NADH. The double mutant, R264H/R27Y, had more profoundly enhanced reductase activity with NADH. Kinetic analysis results demonstrated that these mutants had reduced Km values toward NADH and increased kcat/Km values in comparison with the wild-type. Computer modeling showed that no hydrogen bond could be formed between the wild-type and 2′-hydroxyl group of adenosine ribose of the NADH cofactor, whereas three hydrogen bonds and one π-π stacking interaction could be formed between R264H or R264H/R27Y mutants and the NADH cofactor. These two mutants could efficiently produce (S)-ECHB using NADH as a cofactor in conjunction with formate dehydrogenase as a coupling enzyme.
- Jung, Jihye,Park, Seongsoon,Kim, Hyung Kwoun
-
-
Read Online
- A novel method to synthesize (L)-β-hydroxyl esters by the reduction with bakers' yeast
-
β-Keto esters are reduced stereoselectively into the corresponding (L)-β-hydroxyl esters in the presence of ethyl chloroacetate.
- Nakamura, Kaoru,Kawai, Yasushi,Ohno, Atsuyoshi
-
-
Read Online
- Asymmetric synthesis of (S)-ethyl-4-chloro-3-hydroxy butanoate using a Saccharomyces cerevisiae reductase: Enantioselectivity and enzyme-substrate docking studies
-
Ethyl (S)-4-chloro-3-hydroxy butanoate (ECHB) is a building block for the synthesis of hypercholesterolemia drugs. In this study, various microbial reductases have been cloned and expressed in Escherichia coli. Their reductase activities toward ethyl-4-chloro oxobutanoate (ECOB) have been assayed. Amidst them, Baker's yeast YDL124W, YOR120W, and YOL151W reductases showed high activities. YDL124W produced (S)-ECHB exclusively, whereas YOR120W and YOL151W made (R)-form alcohol. The homology models and docking models with ECOB and NADPH elucidated their substrate specificities and enantioselectivities. A glucose dehydrogenase-coupling reaction was used as NADPH recycling system to perform continuously the reduction reaction. Recombinant E. coli cell co-expressing YDL124W and Bacillus subtilis glucose dehydrogenase produced (S)-ECHB exclusively.
- Jung, Jihye,Park, Hyun Joo,Uhm, Ki-Nam,Kim, Dooil,Kim, Hyung-Kwoun
-
-
Read Online
- Practical application of recombinant whole-cell biocatalysts for the manufacturing of pharmaceutical intermediates such as chiral alcohols
-
We have developed efficient biocatalytic processes for the preparation of chiral alcohols, such as (R)-1,3-butanediol, ethyl (S)-4-chloro-3-hyroxybutanoate, ethyl (R)-4-chloro-3-hyroxybutanoate, (S)-5-chloro-2-pentanol, (R)-5-chloro-2-pentanol, and (S)-cyclopropylethanol by stereospecific enzymatic oxidoreduction on a practical level. These chiral alcohols are very important synthons for the synthesis of various pharmaceutical intermediates that lead to antibiotics and inhibitors of HMG-CoA reductase. Here, we present practical applications on biocatalysis using novel recombinant whole-cell biocatalysts that catalyzed enantioselective oxidation and asymmetric reduction with a coenzyme regeneration system.
- Matsuyama, Akinobu,Yamamoto, Hiroaki,Kobayashi, Yoshinori
-
-
Read Online
- A novel highly stereoselective synthesis of chiral 5- and 4,5-substituted 2-oxazolidinones
-
A novel highly stereoselective synthesis of chiral mono- and bicyclic 4- and 4,5-substituted 2-oxazolidinones starting from β-keto esters was developed. After bioreduction with S. cerevisiae the resulting homochiral β-hydroxy esters are transformed into their hydrazides. Treatment with NaNO2/H+ then furnishes 2-oxazolidinones in high e.e. (~99%) and d.e. (>99%). The ring formation proceeds via a highly concerted sextet rearrangement with full retention of configuration at the stereocentres. Enantiopure 1,2-amino alcohols can subsequently be obtained by saponification of the 2-oxazolidinone products.
- Bertau,Buerli,Hungerbuehler,Wagner
-
-
Read Online
- Synthesis of both enantiomers of ethyl-4-chloro-3-hydroxbutanoate from a prochiral ketone using Candida parapsilosis ATCC 7330
-
Candida parapsilosis ATCC 7330 when grown in a medium containing glycerol reduced ethyl-4-chloro-3-oxobutanoate to (R)-ethyl-4-chloro-3-hydroxybutanote (ee >99%, yield: 94%) while glucose and sucrose grown cells yielded (S)-ethyl-4-chloro-3-hydroxybutanote (ee >99%, yield: 96%). The activity of ethyl-4-chloro-3-oxobutanoate reductase was higher in glucose-grown cells (160 U/g protein) when compared to sucrose (158 U/g protein) and glycerol (22 U/g protein). Both the enantiomers of ethyl-4-chloro-3-hydroxybutanoate (ee >99%) can thus be obtained using Candida parapsilosis ATCC 7330 by altering the carbon source in the growth medium.
- Kaliaperumal, Tarjan,Gummadi, Sathyanarayana N.,Chadha, Anju
-
-
Read Online
- Molecular Cloning and Overexpression of the Gene Encoding an NADPH-Dependent Carbonyl Reductase from Candida magnoliae, Involved in Stereoselective Reduction of Ethyl 4-Chloro-3-oxobutanoate
-
An NADPH-dependent carbonyl reductase (S1) isolated from Candida magnoliae catalyzed the reduction of ethyl 4-chloro-3-oxobutanoate (COBE) to ethyl (S)-4-chloro-3-hydroxybutanoate (CHBE), with a 100% enantiomeric excess, which is a useful chiral building block for the synthesis of pharmaceuticals. The gene encoding the enzyme was cloned and sequenced. The S1 gene comprises 849 bp and encodes a polypeptide of 30,420 Da. The deduced amino acid sequence showed a high degree of similarity to those of the other members of the short-chain alcohol dehydrogenase superfamily. The S1 gene was overexpressed in Escherichia coli under the control of the lac promoter. The enzyme expressed in E. coli was purified to homogeneity and had the same catalytic properties as the enzyme from C. magnoliae did. An E. coli transformant reduced COBE to 125 g/1 of (S)-CHBE, with an optical purity of 100% enantiomeric excess, in an organic solvent two-phase system.
- Yasohara, Yoshihiko,Kizaki, Noriyuki,Hasegawa, Junzo,Wada, Masaru,Kataoka, Michihiko,Shimizu, Sakayu
-
-
Read Online
- A PRACTICAL ASYMMETRIC SYNTHESIS OF CARNITINE
-
The first efficient chemical synthesis of (R)-carnitine has been accomplished on the basis of homogenous enantioselective hydrogenation of ethyl 4-chloro-3-oxobutanoate.
- Kitamura, M.,Ohkuma, T.,Takaya, H.,Noyori, R.
-
-
Read Online
- Cloning, expression and enzymatic characterization of an aldo-keto reductase from Candida albicans XP1463
-
An aldo-keto reductase encoding gene caakr was cloned from Candida albicans XP1463 (CCTCC M 2014382), and heterologously expressed in Escherichia coli. The aldo-keto reductase CaAKR is NADH-dependent with a molecular weight of approximately 38.6 kDal including a His6-Tag. It is active and stable at 30°C and pH 7.0. The maximal reaction rate (Vmax), apparent Michaelis-Menten constant (Km) and catalytic constant (kcat) for t-butyl 6-cyano-(5R)-hydroxy-3-oxohexanoate ((R)-1a) were 11.50 mmol/L min, 1.91 mmol/L and 218.50 min-1. Besides atorvastatin's chiral synthon t-butyl 6-cyano-(3R,5R)- dihydroxy hexanoate ((R,R)-1b), it can synthesize N,N-2-dimethyl-(3S)-hydroxy-3-(2-thienyl)-1-propanine ((S)-9b) and methyl 1-[E]-2-[3-[3-[2-(7-chloro-2-quinoliny) ethenyl] phenyl]-(3S)-hydroxy propy] benzoate ((S)-10b), the chiral intermediates of duloxetine and montelukast, displaying potential applications in pharmaceutical industry. 2015 Elsevier B.V. All rights reserved.
- Wang, Ya-Jun,Liu, Xiao-Qing,Luo, Xi,Liu, Zhi-Qiang,Zheng, Yu-Guo
-
-
Read Online
- Asymmetric synthesis of the chiral synthon ethyl (S)-4-chloro-3- hydroxybutanoate using Lactobacillus kefir
-
Lactobacillus kefir was used as the whole cell biocatalyst for the asymmetric reduction of ethyl 4-chloro acetoacetate 1 to the chiral synthon ethyl (S)-4-chloro-3-hydroxybutanoate 2. Ketoester 1 was obtained as micro-droplets, without the use of an organic solvent as substrate reservoir. 2 (1.2 M) was produced using 2-propanol as co-substrate with a final yield of 97% within 14 h. A high space-time yield and a high specific product capacity of 85.7 mmol/L h and of 24 mmol/gDCW were measured. The enantiomeric excess of the (S)-alcohol 2 was 99.5%.
- Amidjojo, Maya,Weuster-Botz, Dirk
-
-
Read Online
- Immobilized whole cells as effective catalysts for chiral alcohol production
-
Recombinant Escherichia coli overexpressing the gene LbADH, which encodes for an alcohol dehydrogenase from Lactobacillus brevis, was successfully transformed and cultured. The cells are able to catalyze the reduction of pro-chiral ketones, e.g. ethyl acetoacetate into R-()ethyl hydroxybutyrate (EHB) with high conversion and enantiomeric excess 99%. Immobilizing the whole cells in alginate beads leads to a catalyst with improved stability and ease of handling while maintaining the high activity of the free cells. The whole-cell catalyst was tested in a stirred batch reactor (CSTR) and in a continuously operated packed-bed reactor. An Mg2+ concentration of 2 mM was crucial for maintaining the activity of the biocatalyst. After a partial optimization of the process conditions, a productivity of 1.4 gEHB gwcw-1 h-1 could be maintained in a continuous flow reactor over a prolonged period of time. CSIRO 2009.
- Ng, Jeck Fei,Jaenicke, Stephan
-
-
Read Online
- Synthesis of a new class of chiral 1,5-diphosphanylferrocene ligands and their use in enantioselective hydrogenation
-
A new family of ferrocenylphosphane ligands has been prepared. Their flexible synthesis allows many structural modifications. The asymmetric induction of these ligands was examined in the hydrogenation of functionalized C=C, C=O, and C=N bonds. The enantioselectivity of the reaction was strongly dependent on the substituent R at the position α to the ferrocene moiety. In many cases, both enantiomeric β-hydroxyesters of the reduction product can be obtained by simply replacing a dimethylamino group in the ligand with a methyl group.
- Ireland, Tania,Tappe, Katja,Grossheimann, Gabi,Knochel, Paul
-
-
Read Online
- Immobilized microorganisms in the reduction of ethyl 4-chloro acetoacetate
-
Microorganisms were used to reduce ethyl 4-chloroacetoacetate (CAAE) to ethyl (S)-4-chloro-3-hydroxybutanoate [(S)-CHBE]. Mucor ramannianus provided 98% conversion with 84% ee. Free cells of Kluyveromyces marxianus led to 95% conversion with 81% ee. After a fractionary factorial design to study the reaction conditions, calcium alginate immobilized cells of K. marxianus furnished the product with 99% conversion with 91% ee.
- Ribeiro, Joyce Benzaquem,de Souza Ramos, Aline,Fiaux, Sorele Batista,Leite, Selma Gomes Ferreira,Ramos, Maria da Conceicao Klaus V.,de Aquino Neto, Francisco Radler,Antunes, Octavio A.C.
-
-
Read Online
- A novel reductase from Candida albicans for the production of ethyl (S)-4-chloro-3-hydroxybutanoate
-
A novel NADPH-dependent reductase (CaCR) from Candida albicans was cloned for the first time. It catalyzed asymmetric reduction to produce ethyl (S)-4-chloro-3-hydroxybutanoate ((S)-CHBE). It contained an open reading frame of 843 bp encoding 281 amino acids. When co-expressed with a glucose dehydrogenase in Escherichia coli, recombinant CaCR exhibited an activity of 5.7 U/mg with ethyl 4-chloro-3-oxobutanoate (COBE) as substrate. In the biocatalysis of COBE to (S)-CHBE, 1320mM (S)-CHBE was obtained without extra NADP+/NADPH in a water/butyl acetate system, and the optical purity of the (S)-isomer was higher than 99% enantiomeric excess.
- An, Mingdong,Cai, Ping,Yan, Ming,Hao, Ning,Wang, Shanshan,Liu, Huan,Li, Yan,Xu, Lin
-
-
Read Online
- Deep Eutectic Solvents as Media in Alcohol Dehydrogenase-Catalyzed Reductions of Halogenated Ketones
-
The application of deep eutectic solvents (DESs) in biotechnological processes has gained an outstanding relevance, as they can be used as greener media to obtain higher productivities and selectivities. In the present contribution, an eutectic mixture composed of choline chloride (ChCl): glycerol (1 : 2 mol/mol) has been used as a reaction medium in combination with Tris?SO4 50 mM buffer pH 7.5, applied to the alcohol dehydrogenase (ADH)-catalyzed reduction of various carbonyl precursors of chiral halohydrins. These alcohols are key intermediates of biologically active compounds, and hence they are of industrial interest. In the presence of up to 50 % v/v of DES, these biotransformations were achieved up to 300–400 mM of the α-halogenated ketone substrate, getting access to the final compounds with excellent conversions (usually >90 %) and enantiomeric excess (ee >99 %). Among the different ADHs tested, two stereocomplementary enzymes (Lactobacillus brevis ADH and Rhodococcus ruber ADH) afforded the best results, so both alcohol enantiomers could be obtained in all the studied examples. Selected bioreductions were scaled up to 250 mg and 1 g, demonstrating the potential that DESs can offer as media in redox processes for substrates with low solubility in water.
- Ibn Majdoub Hassani, Fatima Zohra,Amzazi, Saaid,Kreit, Joseph,Lavandera, Iván
-
-
Read Online
- Effect of Allyl Alcohol on Reduction of β-Keto Esters by Bakers' Yeast
-
Stereochemical course of the reduction of β-keto esters by bakers' yeast was controlled by treating the reduction systems with allyl alcohol.
- Nakamura, Kaoru,Inoue, Kiyoko,Ushio, Kazutoshi,Oka, Shinzaburo,Ohno, Atsuyoshi
-
-
Read Online
- Altering the substrate specificity of reductase CgKR1 from Candida glabrata by protein engineering for bioreduction of aromatic α-keto esters
-
A versatile keto ester reductase CgKR1, exhibiting a broad substrate spectrum, was obtained from Candida glabrata by genome data mining. It showed the highest activity toward an aliphatic β-keto ester, ethyl 4-chloro-3-oxobutanoate (COBE), but much lower activity toward bulkier α-keto esters with an aromatic group, such as methyl ortho- chlorobenzoylformate (CBFM) and ethyl 2-oxo-4-phenylbutyrate (OPBE). By rational design of the active pocket, the substrate specificity of the reductase was significantly altered and this tailor-made reductase showed a much higher activity toward aromatic α-keto esters (~7-fold increase in k cat/Km toward CBFM) and lower activity toward aliphatic keto esters (~12-fold decrease in kcat/Km toward COBE). Meanwhile, the thermostability of the reductase was enhanced by a consensus approach. Such improvements may yield practical catalysts for the asymmetric bioreduction of these aromatic α-keto esters
- Huang, Lei,Ma, Hong-Min,Yu, Hui-Lei,Xu, Jian-He
-
-
Read Online
- A class of readily available optically pure 7,7′-disubstituted BINAPs for asymmetric catalysis
-
A class of optically pure 7,7′-disubstituted BINAPs have been prepared starting with a catalytic asymmetric oxidative coupling reaction. A general, concise, and straightforward synthetic procedure has been established, and is suitable for all optically pure 7,7′-disubstituted BINAPs 1a-h, regardless of the substituents' structure in the 7,7′-positions. The catalytic potential of this class of ligands has been investigated in the highly enantioselective Rh-catalyzed 1,4-addition of aryl boronic acids to enones (up to 99.8% ee), and Ru-catalyzed asymmetric hydrogenation of simple aromatic ketones (up to S/C=100,000, up to 98% ee) and β-ketoesters (up to S/C=10,000, up to 99.8% ee), respectively.
- Yuan, Wei-Cheng,Cun, Lin-Feng,Mi, Ai-Qiao,Jiang, Yao-Zhong,Gong, Liu-Zhu
-
-
Read Online
- Purification and Characterization of NADPH-Dependent Carbonyl Reductase, Involved in Stereoselective Reduction of Ethyl 4-Chloro-3-oxobutanoate, from Candida magnoliae
-
A NADPH-dependent carbonyl reductase was purified to homogeneity from Candida magnoliae AKU4643 through four steps, including Blue Sepharose affinity chromatography. The enzyme catalyzed the stereoselective reduction of ethyl 4-chloro-3-oxobutanoate to the corresponding (S)-alcohol with a 100% enantiomeric excess, which is a useful chiral building block for the chemical synthesis of pharmaceuticals. The relative molecular mass of the enzyme was estimated to be 76,000 on high performance gel filtration chromatography and 32,000 on SDS polyacrylamide gel electrophoresis. The enzyme reduced α, β-keto esters and conjugated diketones in addition to ethyl 4-chloro-3-oxobutanoate. The enzyme activity was inhibited by quercetin and HgCl2, but not by EDTA. The N-terminal amino acid sequence of the enzyme showed no apparent similarity with those of other oxidoreductases.
- Wada, Masaru,Kataoka, Michihiko,Kawabata, Hiroshi,Yasohara, Yoshihiko,Kizaki, Noriyuki,Hasegawa, Junzo,Shimizu, Sakayu
-
-
Read Online
- Optimization and scale-up of a bioreduction process for preparation of ethyl (S)-4-chloro-3-hydroxybutanoate
-
Ethyl 4-chloro-3-oxobutanoate (COBE) was asymmetrically reduced with Escherichia coli cells expressing a reductase (ScCR) from Streptomyces coelicolor to afford enantiopure ethyl (S)-4-chloro-3-hydroxybutanoate [(S)-CHBE], which is an important precursor for preparing the drug atorvastatin. The substrate load was fixed at 100 g/L, and the concentration of coenzyme NAD+ was limited to 0.1 mM based on cost considerations. Under these conditions, the other reaction parameters were optimized as 25 °C and pH 6.5, with a biocatalyst dose of 10 kU/L in the presence of isopropanol (1.5 equiv of COBE), which acted as a cosubstrate for regenerating NADH. The reaction was performed in a toluene-aqueous biphasic system (1:1, v/v), with agitation at the maximal linear rate of 0.88 m/s. Finally, the bioreaction was performed on a pilot scale using a 50 L thermostated stirred-tank-reactor, affording (S)-CHBE in 85.4% yield and 99.9% ee, and a total turnover number (TTN) of 6060 for the cofactor NAD+. The specific production was calculated to be 36.8 gproduct/gdcw, which is the highest value reported to date among the whole-cell-mediated processes for producing (S)-CHBE.
- Pan, Jiang,Zheng, Gao-Wei,Ye, Qin,Xu, Jian-He
-
-
Read Online
- Asymmetric reduction of substituted α- and β-ketoesters by Bacillus pumilus Phe-C3
-
The enantioselective reduction of substituted α- and β-ketoesters using resting cells of Bacillus pumilus Phe-C3 was investigated. Effects of substrate concentration on the catalytic efficiency of the microorganism were studied. Preparative scale productions were carried out under the optimized conditions with 62.4-91.0% yields and 90.2-97.1% ee. The cells retained 80% of initial activity after recycling for six times.
- He, Chunmao,Chang, Dongliang,Zhang, Jie
-
-
Read Online
- Preparation of ethyl (R)-3-hydroxy-4-chlorobutyrate by selective reduction of (R)-4-(trichloromethyl)-oxetan-2-one: Key intermediate to (R)-carnitine and (R)-4-amino-3-hydroxybutyric acid
-
Selective reduction of (R)-4-(trichloromethyl)-oxetan-2-one in ethanol by catalytic hydrogenation on Pd-C in the presence of KOAc gave directly ethyl (R)-3-hydroxy-4-chlorobutyrate, which can be used as a key intermediate for the synthesis of some biologically active γ-amino-β-hydroxy amino acids, (R)-carnitine and γ-amino-β-hydroxy amino acid (R)-GABOB).
- Song, Choong Eui,Lee, Jae Kyun,Kim, In O.,Choi, Jung Hoon
-
-
Read Online
- Enantioselective hydrogenation of α- and β-functionalized ketones by Ru(II){AMPP} catalysts
-
Neutral ruthenium complexes bearing aminophosphine-phosphinite ligands have been applied in the enantioselective hydrogenation of α- (5-7) and β- (8-10) functionalized ketones. The corresponding hydroxycompounds are produced with enantiomeric excesses up to 63 and 85% respectively for the two types of substrates.
- Hapiot, Frederic,Agbossou, Francine,Mortreux, Andre
-
-
Read Online
- 2-(1S)-Camphanoyloxy-2′-phosphanylbiphenyl Ligands – Synthesis, Structure, and Preliminary Tests in Transition-Metal Catalysis
-
Diastereoisomer separation of the (1S)-camphanic acid 2-isopropylphenylphosphanyl-phenyl ester 1 exemplifies the potential of (1S)-camphanoyl chloride for enantiomer separation of hydroxyl-functional asymmetric phosphanes. Esterification of lithium 2′-phosphanylbiphenyl-2-olates, generated from the respective 2-OH or 2-OSiMe3 precursors 2aOH and 2b–fSi, furnished the 2-(1S)-camphanoyloxy-biphenylphosphanes 3a–c as 1:1 mixtures of diastereomers with low barriers for interconversion by rotation around the C–C axis (ΔG# = 70–73 kJ mol–1 for 3a and 3c by 31P VT NMR spectroscopy). The P-asymmetric compounds 3d–f form 1:1 mixtures of stereoisomers. There is a tendency to cocrystallization of two preferred diastereoisomers, as shown by the crystal structure analyses of 3dD and 3fD, and in solution, there is a tendency toward partial isomerization to the sterically less-favored atropisomers. The [RhCl(cod)(3dD)] complex 4dD, however, seems stable in solution. Excess 2dLi reacted with (1S)-camphanoyl chloride preferentially to form the (SP,Rax,1S) isomer, which was separated by crystallization as enantiopure 3dE, characterized by single-crystal XRD. Preliminary screening tests of this ligand in Rh-catalyzed asymmetric hydrogenations of N-(1-phenylvinyl)acetamide allowed high conversion and up to 59 % ee. Hydrosilylation of acetophenone proceeded with 78 % conversion and 48 % ee; Suzuki–Miyaura couplings of 1-bromo-2-naphthol with PhB(OH)2, in the presence of 3b/[Pd(OAc)2], gave yields up to 98 %.
- Wawrzyniak, Piotr,Kindermann, Markus K.,Thede, Gabriele,Thede, Richard,Jones, Peter G.,Enthaler, Stephan,Junge, Kathrin,Beller, Matthias,Heinicke, Joachim W.
-
-
Read Online
- Asymmetric whole cell biotransformations in biphasic ionic liquid/water-systems by use of recombinant Escherichia coli with intracellular cofactor regeneration
-
Ionic liquids such as [BMIM][PF6] and [BMIM][NTF] are already known as good alternatives to organic solvents in biphasic biotransformation. Herein, we report about a systematic procedure based on physical properties to identify more commercially available ionic liquids exhibiting the potential to improve the efficiency of whole cell biocatalyses. This approach resulted in the identification of seven other water immiscible ionic liquids. These ionic liquids were rated by their biocompatibility, their substrate- and product-specific distribution coefficients and by for example performed asymmetric reductions of several prochiral ketones. With the use of a recombinant Escherichia coli as biocatalyst, overproducing a Lactobacillus brevis alcohol dehydrogenase and a Mycobacterium vaccae N10 formate dehydrogenase for cofactor regeneration, the great potential of asymmetric whole cell biotransformations in biphasic ionic liquid/water-systems were demonstrated in simple batch processes.
- Braeutigam, Stefan,Bringer-Meyer, Stephanie,Weuster-Botz, Dirk
-
-
Read Online
- A genomic search approach to identify carbonyl reductases in Gluconobacter oxydans for enantioselective reduction of ketones
-
The versatile carbonyl reductases from Gluconobacter oxydans in the enantioselective reduction of ketones to the corresponding alcohols were exploited by genome search approach. All purified enzymes showed activities toward the tested ketoesters with different activities. In the reduction of 4-phenyl-2-butanone with in situ NAD(P)H regeneration system, (S)-alcohol was obtained with an e.e. of up to 100% catalyzed by Gox0644. Under the same experimental condition, all enzymes catalyzed ethyl 4-chloroacetoacetate to give chiral products with an excellent e.e. of up to 99%, except Gox0644. Gox2036 had a strict requirement for NADH as the cofactor and showed excellent enantiospecificity in the synthesis of ethyl (R)-4-chloro-3-hydroxybutanoate. For the reduction of ethyl 2-oxo-4-phenylbutyrate, excellent e.e. (>99%) and high conversion (93.1%) were obtained by Gox0525, whereas the other enzymes showed relatively lower e.e. and conversions. Among them, Gox2036 and Gox0525 showed potentials in the synthesis of chiral alcohols as useful biocatalysts.
- Chen, Rong,Liu, Xu,Lin, Jinping,Wei, Dongzhi
-
-
Read Online
- Identification of a Robust Carbonyl Reductase for Diastereoselectively Building syn-3,5-Dihydroxy Hexanoate: A Bulky Side Chain of Atorvastatin
-
t-Butyl-6-cyano-(3R,5R)-dihydroxyhexanoate is an advanced chiral precursor for the synthesis of the side chain pharmacophore of cholesterol-lowering drug atorvastatin. Herein, a robust carbonyl reductase (LbCR) was newly identified from Lactobacillus brevis, which displays high activity and excellent diastereoselectivity toward bulky t-butyl 6-cyano-(5R)-hydroxy-3-oxo-hexanoate (7). The engineered Escherichia coli cells harboring LbCR and glucose dehydrogenase (for cofactor regeneration) were employed as biocatalysts for the asymmetric reduction of substrate 7. As a result, as much as 300 g L-1 of water-insoluble substrate was completely converted to the corresponding chiral diol with >99.5% de in a space-time yield of 351 g L-1 d-1, indicating a great potential of LbCR for practical synthesis of the very bulky and bi-chiral 3,5-dihydroxy carboxylate side chain of best-selling statin drugs.
- Gong, Xu-Min,Zheng, Gao-Wei,Liu, You-Yan,Xu, Jian-He
-
-
Read Online
- Genomic mining-based identification of novel stereospecific aldo-keto reductases toolbox from Candida parapsilosis for highly enantioselective reduction of carbonyl compounds
-
Biocatalytic reduction of prochiral ketones offers significant potential in synthesis of optically active alcohols. However, so far the application of aldo-keto reductases (AKRs) in asymmetric reduction has been hampered due to limited availability of AKRs with high enantioselectivity and catalytic efficiency. Based on the genome sequence of Candida parapsilosis, a versatile bioresource for asymmetric reduction, eight open reading frames encoding putative AKRs were discovered and expressed, and the resulted enzymes (CPARs), comprising an AKR toolbox, were evaluated toward various carbonyl substrates. The CPARs were active to the selected substrates, especially 2-hydroxyacetophenone and ethyl 4-chloro-3-oxobutyrate. Additionally, most of them were obviously enantioselective to the substrates and gave alcohol products with optical purity up to 99%e.e. Of the enzymes, CPAR4 was outstanding with excellent enantioselectivity and broad substrate spectrum. All these positive features demonstrate that genomic mining is powerful in searching for novel and efficient biocatalysts of desired reactions for pharmaceuticals and fine chemicals synthesis.
- Guo, Rongyun,Nie, Yao,Mu, Xiao Qing,Xu, Yan,Xiao, Rong
-
-
Read Online
- New method for the preparation of (R)-carnitine
-
A new method for the preparation of (R)-carnitine (1) has been developed from enantiomerically pure (R)-4-(trichloromethyl)-oxetan-2-one [(R)-2] which was easily obtained from the [2 + 2]-cycloaddition of ketene and chloral in the presence of catalytic amounts of poly(acryloyl quinidine). The key intermediate, ethyl (R)-3-hydroxy-4-chlorobutyrate [(R)-5], was prepared by ethanolysis of (R)-2 followed by selective bis-dechlorination of ethyl (R)-3-hydroxy-4,4,4-trichlorobutyrate [(R)-3].
- Song,Lee,Lee,Lee
-
-
Read Online
- Synthesis of ethyl (R)-4-chloro-3-hydroxybutyrate by immobilized cells using amino acid-modified magnetic nanoparticles
-
Fe3O4-Arg was selected as the optimal carrier due to its high activity recovery of immobilized cells in the preparation of Fe3O4-Arg-Cells. The optimal immobilization conditions for the preparation of Fe3O4-Arg-Cells were 30 °C, 4 h, pH 7, and 3 g dry yeast. The activity recovery of immobilized cells reached 76.8 percent. For a batch reduction in a shaker in an alternating magnetic field, Fe3O4-Arg-Cells were used as a catalyst to gain ethyl (R)-4-chloro-3-hydroxybutyrate ((R)-CHBE). For further improvement in reduction productivity, a continuous reduction in the magnetic fluidized bed reactor system (MFBRS) was completed. Under their optimal transformation conditions, it took 24 h for Fe3O4-Arg-Cells to complete the conversion of ethyl 4-chloro-3-oxobutanoate (COBE) (0.8553 mol/L) in the shaker and only 8 h for the batch reduction in an alternating magnetic field. Continuous reduction in MFBRS provided new ideas for the efficient production of (R)-CHBE; 1.5882 mol/L (10 mL) of COBE can be completely converted in 6 h. The conversion and enantiomeric excess (e.e.) of (R)-CHBE were 100 percent and above 99.9 percent respectively, in the three reaction systems mentioned above.
- Dai, Hongqian,Lu, Yuan,Ou, Zhimin,Shi, Hanbing,Sun, Xingyuan,Tang, Lan
-
-
Read Online
- A novel generation of optically active ethyl 4-chloro-3-hydroxybutyrate as a C4 chiral building unit using microbial dechlorination
-
A novel procedure for the generation of optically active ethyl 4-chloro-3-hydroxybutyrate using bacterial cells was developed. Ethyl (S)-4-chloro-3-hydroxybutyrate was prepared by Pseudomonas sp. OS-K-29, which stereoselectively assimilates 2,3-dichloro-1-propanol. The reaction was based on its kinetic dehalogenation for both enantiomers using the resting cells. The obtained 4-chloro-3-hydroxybutyrate and high enantiomeric excess of >98% with a yield of 33% at the microbial resolution step. Moreover, several C4 compounds having the 4-chloro-3-hydroxyl function were also resolved and gave good enantiomeric purities (>95% ee). Ethyl (R)-4-chloro-3-hydroxybutyrate was also obtained with high enantiomeric purity (>98% ee) using the cells of Pseudomonas sp. DS-K-NR818. Copyright (C) Elsevier Science Ltd.
- Suzuki, Toshio,Idogaki, Hideaki,Kasai, Naoya
-
-
Read Online
- Gene cloning and expression of Leifsonia alcohol dehydrogenase (LSADH) involved in asymmetric hydrogen-transfer bioreduction to produce (R)-form chiral alcohols
-
The gene encoding Leifsonia alcohol dehydrogenase (LSADH), a useful biocatalyst for producing (R)-chiral alcohols, was cloned from the genomic DNA of Leifsonia sp. S749. The gene contained an opening reading frame consisting of 756 nucleotides corresponding to 251 amino acid residues. The subunit molecular weight was calculated to be 24,999, which was consistent with that determined by polyacrylamide gel electrophoresis. The enzyme was expressed in recombinant Escherichia coli cells and purified to homogeneity by three column chromatographies. The predicted amino acid sequence displayed 30-50% homology to known short chain alcohol dehydrogenase/reductases (SDRs); moreover, the NADH-binding site and the three catalytic residues in SDRs were conserved. The recombinant E. coli cells which overexpressed lsadh produced (R)-form chiral alcohols from ketones using 2-propanol as a hydrogen donor with the highest level of productivity ever reported and enantiomeric excess (e.e.).
- Inoue, Kousuke,Makino, Yoshihide,Dairi, Tohru,Itoh, Nobuya
-
-
Read Online
- Synthetic applications of the carbonyl reductases isolated from Candida parapsilosis and Rhodococcus erythropolis
-
Synthetic applications of the novel carbonyl reductases isolated from Candida parapsilosis and Rhodococcus erythropolis are reported. A number of different carbonyl compounds such as 3-, 4- and 5-ketoesters, ethyl 4-chloro 3-oxobutanoate, pyruvic aldehyde dimethyl acetal, ethyl α-methyl 3-oxobutanoate and acetophenone are reduced on the preparative scale to the corresponding (S)-hydroxy compounds with high enantiomeric excess (>>94%) and yield (>>83%). Product concentrations in the range of 30 to 200 mM were obtained. The synthesized chiral compounds are valuable building blocks for the synthesis of pharmaceuticals, agrochemicals and natural products.
- Peters,Zelinski,Minuth,Kula
-
-
Read Online
- Enantioselective bioreductive preparation of chiral halohydrins employing two newly identified stereocomplementary reductases
-
Two robust stereocomplementary carbonyl reductases (DhCR and CgCR) were identified through rescreening the carbonyl reductase toolbox. Five reductases were returned through the activity and enantioselectivity assay for α-chloro-1-acetophenone and ethyl 4-chloro-3-oxo-butanate (COBE). Three reductases were stable at elevated substrate loading. Enzymatic characterization revealed that DhCR and CgCR were more thermostable. As much as 330 g COBE in 1 L biphasic reaction mixture was reduced to (S)- and (R)-3-hydroxy-4-chlorobutyrate by DhCR and CgCR (coexpressed with glucose dehydrogenase), with 92.5% and 93.0% yields, >99% ee, and total turnover numbers of 53800 and 108000, respectively. Six other α-halohydrins were asymmetrically reduced to optically pure forms at a substrate loading of 100 g L-1. Our results indicate the potential of these two stereocomplementary reductases in the synthesis of valuable α-halohydrins for pharmaceuticals. This journal is
- Xu, Guo-Chao,Yu, Hui-Lei,Shang, Yue-Peng,Xu, Jian-He
-
-
Read Online
- Highly active mutants of carbonyl reductase S1 with inverted coenzyme specificity and production of optically active alcohols
-
A wild type NADPH-dependent carbonyl reductase from Candida magnoliae (reductase S1) has been found not to utilize NADH as a coenzyme. A mutation to exchange the coenzyme specificity in reductase S1 has been designed by computer-aided methods, including three-dimensional structure modeling and in silico screening of enzyme mutants. Site-directed mutagenesis has been used to introduce systematic substitutions of seven or eight amino acid residues onto the adenosine-binding pocket of the enzyme according to rational computational design. The resulting S1 mutants show NADH-dependency and have lost their ability to utilize NADPH as a coenzyme, but retain those catalytic activities. Kinetic parameter Vmax and Km, values of those mutants for NADH are 1/3- to 1/10-fold those of the wild type enzyme for NADPH. As a model system for industrial production of optically active alcohols, the S1 mutants can be applied to an asymmetric reduction of ketones, cooperating with a coenzyme-regeneration system that uses an NAD-dependent formate dehydrogenase.
- Morikawa, Souichi,Nakai, Takahisa,Yasohara, Yoshihiko,Nanba, Hirokazu,Kizaki, Noriyuki,Hasegawa, Junzo
-
-
Read Online
- A new chiral dipyridylphosphine ligand Xyl-P-Phos and its application in the Ru-catalyzed asymmetric hydrogenation of β-ketoesters
-
A new chiral dipyridylphosphine ligand Xyl-P-Phos has been synthesized and the structure of (S)-Xyl-P-Phos oxide has been characterized by single crystal X-ray diffraction. The ruthenium complex of this ligand, Ru(R-Xyl-P-Phos)(C6H6)Cl2, has been found to be a highly active, enantioselective and air-stable catalyst for the asymmetric hydrogenation of β-ketoesters and shows good potential for industrial applications.
- Wu, Jing,Chen, Hua,Kwok, Wai Him,Lam, Kim Hung,Zhou, Zhong Yuan,Yeung, Chi Hung,Chan, Albert S.C
-
-
Read Online
- Method for continuously preparing (R)-4-halo-3-hydroxy-butyrate by using micro-reaction system
-
The invention belongs to the technical field of chemical engineering, and particularly relates to a method for continuously preparing (R)-4-halo-3-hydroxy-butyrate by using a micro-reaction system. Asubstrate solution containing halogenated acetoacetate and a biological catalytic solution are continuously subjected to an enzyme-catalyzed asymmetric reduction reaction in the micro-reaction systemcomposed of a micro-mixer, a micro-channel reactor and a pH regulator so as to obtain the (R)-4-halo-3-hydroxy-butyrate. Compared with the prior art, the method has the advantages that: the reaction time is only a few minutes, the yield of the product (R)-4-halo-3-hydroxy-butyrate is more than 95 percent, the process is continuous, the automation degree is high, the efficiency is high, the technological process is simple and convenient to operate, and the industrial production is easy.
- -
-
Paragraph 0071-0093
(2021/02/10)
-
- Single-Point Mutant Inverts the Stereoselectivity of a Carbonyl Reductase toward β-Ketoesters with Enhanced Activity
-
Enzyme stereoselectivity control is still a major challenge. To gain insight into the molecular basis of enzyme stereo-recognition and expand the source of antiPrelog carbonyl reductase toward β-ketoesters, rational enzyme design aiming at stereoselectivity inversion was performed. The designed variant Q139G switched the enzyme stereoselectivity toward β-ketoesters from Prelog to antiPrelog, providing corresponding alcohols in high enantiomeric purity (89.1–99.1 % ee). More importantly, the well-known trade-off between stereoselectivity and activity was not found. Q139G exhibited higher catalytic activity than the wildtype enzyme, the enhancement of the catalytic efficiency (kcat/Km) varied from 1.1- to 27.1-fold. Interestingly, the mutant Q139G did not lead to reversed stereoselectivity toward aromatic ketones. Analysis of enzyme–substrate complexes showed that the structural flexibility of β-ketoesters and a newly formed cave together facilitated the formation of the antiPrelog-preferred conformation. In contrast, the relatively large and rigid structure of the aromatic ketones prevents them from forming the antiPrelog-preferred conformation.
- Li, Aipeng,Wang, Ting,Tian, Qing,Yang, Xiaohong,Yin, Dongming,Qin, Yong,Zhang, Lianbing
-
p. 6283 - 6294
(2021/03/16)
-
- Method for preparing 3 -hydroxy -4 -chlorobutyric acid ethyl ester
-
The invention discloses a method for preparing 3 -hydroxy -4 -chlorobutyric acid ethyl ester. The method comprises 4 - chloroacetoacetate as a raw material, methanol as a solvent and sodium borohydride as a reducing agent. The methanol is concentrated to recycle after the reaction is finished. The residue is added with methanol, hydrogen chloride gas is introduced to pH=2 - 3 filtration desalination, and the hydrogen chloride - methanol system is recycled. The residue was distilled under reduced pressure to give ethyl 3 -hydroxy -4 -chlorobutyric acid ethyl ester. The sodium borohydride is low in use amount, no by-product peak occurs, and the yield reaches 83 - 87%. The methanol and hydrogen chloride - methanol two solvent systems are separately recycled, so that the use of a low-boiling-point solvent is avoided. The synthetic process is low in cost, high in yield and environmentally friendly, and can be used for industrial large-scale production.
- -
-
Paragraph 0010-0012
(2021/08/25)
-
- Efficient Nicotinamide Adenine Dinucleotide Phosphate [NADP(H)] Recycling in Closed-Loop Continuous Flow Biocatalysis
-
Biocatalytic redox reactions regularly depend on expensive cofactors that require recycling. For continuous conversions in flow chemistry, this is often an obstacle since the cofactor is washed away. Here, we present a quasi-stationary recycling system for nicotinamide adenine dinucleotide phosphate utilizing an immobilized alcohol dehydrogenase. Four model substrates were reduced with high enantioselectivity as a proof of concept. The two-phase system enables continuous production as well as quick substrate changes. This setup may serve as a general cofactor regeneration module for continuous biocatalytic devices employing (co-)substrates being miscible in organic solvent. The system resulted in space-time yields up to 117 g L?1 h?1 and total turnover numbers for nicotinamide adenine dinucleotide phosphate higher than 12,000 mol/mol are possible. (Figure presented.).
- Baumer, Benedikt,Classen, Thomas,Pohl, Martina,Pietruszka, J?rg
-
p. 2894 - 2901
(2020/04/15)
-
- Efficient Asymmetric Synthesis of Ethyl (S)-4-Chloro-3-hydroxybutyrate Using Alcohol Dehydrogenase SmADH31 with High Tolerance of Substrate and Product in a Monophasic Aqueous System
-
Bioreductions catalyzed by alcohol dehydrogenases (ADHs) play an important role in the synthesis of chiral alcohols. However, the synthesis of ethyl (S)-4-chloro-3-hydroxybutyrate [(S)-CHBE], an important drug intermediate, has significant challenges concerning high substrate or product inhibition toward ADHs, which complicates its production. Herein, we evaluated a novel ADH, SmADH31, obtained from the Stenotrophomonas maltophilia genome, which can tolerate extremely high concentrations (6 M) of both substrate and product. The coexpression of SmADH31 and glucose dehydrogenase from Bacillus subtilis in Escherichia coli meant that as much as 660 g L-1 (4.0 M) ethyl 4-chloroacetoacetate was completely converted into (S)-CHBE in a monophasic aqueous system with a >99.9% ee value and a high space-time yield (2664 g L-1 d-1). Molecular dynamics simulation shed light on the high activity and stereoselectivity of SmADH31. Moreover, five other optically pure chiral alcohols were synthesized at high concentrations (100-462 g L-1) as a result of the broad substrate spectrum of SmADH31. All these compounds act as important drug intermediates, demonstrating the industrial potential of SmADH31-mediated bioreductions.
- Chen, Rong,Liu, Qinghai,Wang, Hualei,Wei, Dongzhi,Xie, Youyu,Yang, Zeyu,Ye, Wenjie
-
p. 1068 - 1076
(2020/07/06)
-
- Efficient asymmetric synthesis of chiral alcohols using high 2-propanol tolerance alcohol dehydrogenase: Sm ADH2 via an environmentally friendly TBCR system
-
Alcohol dehydrogenases (ADHs) together with the economical substrate-coupled cofactor regeneration system play a pivotal role in the asymmetric synthesis of chiral alcohols; however, severe challenges concerning the poor tolerance of enzymes to 2-propanol and the adverse effects of the by-product, acetone, limit its applications, causing this strategy to lapse. Herein, a novel ADH gene smadh2 was identified from Stenotrophomonas maltophilia by traditional genome mining technology. The gene was cloned into Escherichia coli cells and then expressed to yield SmADH2. SmADH2 has a broad substrate spectrum and exhibits excellent tolerance and superb activity to 2-propanol even at 10.5 M (80%, v/v) concentration. Moreover, a new thermostatic bubble column reactor (TBCR) system is successfully designed to alleviate the inhibition of the by-product acetone by gas flow and continuously supplement 2-propanol. The organic waste can be simultaneously recovered for the purpose of green synthesis. In the sustainable system, structurally diverse chiral alcohols are synthesised at a high substrate loading (>150 g L-1) without adding external coenzymes. Among these, about 780 g L-1 (6 M) ethyl acetoacetate is completely converted into ethyl (R)-3-hydroxybutyrate in only 2.5 h with 99.9% ee and 7488 g L-1 d-1 space-time yield. Molecular dynamics simulation results shed light on the high catalytic activity toward the substrate. Therefore, the high 2-propanol tolerance SmADH2 with the TBCR system proves to be a potent biocatalytic strategy for the synthesis of chiral alcohols on an industrial scale.
- Yang, Zeyu,Fu, Hengwei,Ye, Wenjie,Xie, Youyu,Liu, Qinghai,Wang, Hualei,Wei, Dongzhi
-
-
- Method for preparing chiral beta-hydroxycarboxylate compound
-
The invention provides a method for preparing a chiral beta-hydroxycarboxylate compound, comprising the following steps: dissolving [Ir(COD)Cl]2, a ligand and an alkaline additive in a solvent, stirring at room temperature, carrying out in-situ synthesis of a catalyst, dissolving a substrate beta-hydroxycarboxylate compound in a solvent, adding the prepared catalyst, and introducing hydrogen to carry out an asymmetric catalytic hydrogenation reaction on the substrate beta-hydroxycarboxylate compound. The reaction conditions are as follows: pressure is 10-100 atmospheres, the reaction temperature is 0-200 DEG C, and the reaction time is 12-48 hours. The reaction activity and selectivity are high, and the hydrogenation reaction conditions are mild. The method is suitable for various beta-hydroxycarboxylate compounds; the substrate application range is wide; and the reaction process causes little environmental pollution.
- -
-
Paragraph 0075-0081
(2019/08/20)
-
- Cobalt-Catalyzed Alkoxycarbonylation of Epoxides to β-Hydroxyesters
-
Herein, we developed a new and practical catalytic system for the carbonylative synthesis of β-hydroxyesters. By using simple, cheap, and air-stable cobalt(II) bromide as the catalyst, combined with pyrazole and catalytic amount of manganese, active cobalt complex can be generated in situ and can catalyze various epoxides to give the corresponding β-hydroxyesters in moderate to excellent yields. Mechanism studies indicate that pyrazole plays a crucial role in this reaction. Moreover, with the addition of the catalytic amount of manganese, the active cobalt catalyst can be regenerated, which provides a possibility for reusing the cobalt catalyst.
- Xu, Jian-Xing,Wu, Xiao-Feng
-
p. 9907 - 9912
(2019/08/26)
-
- Fine tuning the enantioselectivity and substrate specificity of alcohol dehydrogenase from Kluyveromyces polysporus by single residue at 237
-
Here, S237 was identified to be important in fine tuning the substrate specificity and enantioselectivity of alcohol dehydrogenase from Kluyveromyces polysporus (KpADH). In the reduction of a diaryl ketone, (4-chlorophenyl)-(pyridin-2-yl)-methanone (1a), the highest and lowest enantioselectivity of 96.1% and 27.0% e.e. (R) were obtained with S237A and S237C. Kinetic parameters analysis revealed that S237G, S237A, S237H and S237D displayed improved kcat/Km toward 1a. Various prochiral ketones, including acetophenone, 4-chloroacetophenone and ethyl 2-oxo-4-phenylbutyrate could be asymmetrically reduced by S237C, S237G and S237E with > 99% e.e. This study provides guidance for the application of KpADH in the preparation of chiral secondary alcohols.
- Wang, Yue,Dai, Wei,Liu, Yongmei,Zhang, Zhongwei,Zhou, Jieyu,Xu, Guochao,Ni, Ye
-
-
- Synthesis method of quaternary amine inner salt
-
The invention discloses a synthesis method of quaternary amine inner salt. The synthesis method comprises the following steps: (a) reduction reaction: taking a compound having a structure shown as theformula I as a raw material, carrying out a reduction reaction among the compound, an apoenzyme, a dehydrogenase and a coenzyme in monosaccharide within a certain pH range, removing enzymes with active carbon and performing rectification, so as to obtain a reduced product shown in the original specification, wherein X represents one of chlorine, bromine and iodine in halogens, and R represents one of a saturated alkyl or an unsaturated alkyl; (b) synthesis of the quaternary amine inner salt: carrying out a reaction between an obtained product and trimethylamine under a strong base condition to obtain quaternary amine hydrochloride, exchanging the quaternary amine hydrochloride in ion exchange resin to remove halide ions, performing concentration and refining a concentrated product with alcohol and acetone, so as to obtain the quaternary amine inner salt. The synthesis method has the advantages of being high in yield in each step, simple to operate and mild in reaction conditions, effectively removing enzyme residues by introducing a chiral structure with a high-selectivity enzymatic method, avoiding a reagent with high toxicity and high pollution by utilizing renewable resin for desalting, obtaining the high-purity product, being suitable for industrial production and the like.
- -
-
Paragraph 0066; 0067; 0087
(2019/01/14)
-
- Preparation method of high-purity ethyl 4-chloro-3-hydroxybutyrate
-
The invention belongs to the field of drug preparation and particularly relates to a preparation method of high-purity ethyl 4-chloro-3-hydroxybutyrate. Ethyl 4-chloroacetoacetate is used as a raw material to carry out reacting with ethanol as a solvent and sodium borohydride as a reducing agent; the reacted liquid is dewatered, neutralizing with glacial acetic acid is performed until neutrality,filtering is performed to remove salts, vacuum concentration is performed, a solvent is added to carry out dissolving, washing with water is carried out to remove salts, concentrating is performed, and vacuum distillation is performed to obtain the high-purity ethyl 4-chloro-3-hydroxybutyrate. The ethyl 4-chloro-3-hydroxybutyrate is an intermediate of oxiracetam, a drug to treat senile dementia; as registration of new medicines in China requires research on impurities of pharmaceutical intermediates, namely research on impurity whereabouts and influence of impurities on products, the existingpreparation method of ethyl 4-chloro-3-hydroxybutyrate causes many impurities and low purity, only 84.9%; the preparation method has a reasonable ratio of materials, has good simplicity and low cost,and enables the purity to reach 99.5% and above and the yield to reach 90% and above, and the prepared samples may meet the requirements for medicine registration.
- -
-
Paragraph 0028-0038
(2018/09/11)
-
- A atorvastatin calcium intermediate preparation method
-
The present invention provides a kind of atorvastatin calcium intermediate preparation method, will S - 4 - chloro - 3 - hydroxy nitrile-butadiene and cyano hydrolase, buffer solution dropped into the reactor, controlling the reaction temperature at 20 - 40 °C, dropwise lye control PH in 6 - 8 between, the reaction is complete add acid to adjust the PH to 3 - 4, reaction solution with the solvent, pressure reducing and concentrated to obtain S - 4 - chloro - 3 - hydroxy butyric acid, S - 4 - chloro - 3 - hydroxy butyric acid with ethanol, in a small amount of catalyst catalyzes the esterification, distilling off the excess ethyl alcohol, washing the obtained S - 4 - chloro - 3 - hydroxy butyric acid ethyl ester. The obtained S - 4 - chloro - 3 - hydroxy butyric acid ethyl ester is the Atorvastatin calcium is important intermediates, the invention avoids the use of the great amount of acid ethanol, thereby greatly reducing the workshop the production amount of the tail gas recovery and acid water, avoids a great deal of hydrogen chloride gas to the equipment corrosion of the pipeline, the reaction of the invention high yield, high purity product, the preparation process is simple, and is suitable for industrial production.
- -
-
Paragraph 0019; 0021; 0024; 0027
(2018/09/21)
-
- Production technology of 3-hydroxytetrahydrofuran with high optical purity
-
The invention discloses a production technology of3-hydroxytetrahydrofuran with high optical purity. The production technology comprises the following steps: (1) taking chloroacetoacetic acid ethyl ester as a starting raw material, adding appropriate amount of solvents, chiral catalysts and reducing agents, and reacting at an appropriate temperature to obtain chiral ethyl 4-chloro-3-hydroxybutyrate; (2) taking the chiral ethyl 4-chloro-3-hydroxybutyrate obtained in step (1) as a raw material, adding the appropriate amount of solvents and metal borohydride reducing agents, and reacting at the appropriate temperature to obtain chiral 4-chloro-3-hydroxy-1-butanol; (3) taking the chiral 4-chloro-3-hydroxy-1-butanol obtained in step (2) as the raw material, adding appropriate amount of catalysts and solvents, and reacting at the appropriate temperature to obtain chiral 3-hydroxytetrahydrofuran. According to the production technology of the 3-hydroxytetrahydrofuran with the high optical purity, the chiral 3-hydroxytetrahydrofuran can be produced through a three-step reaction, the shortcomings of complicated production operation and high production cost are solved, and products with high optical purity can be produced.
- -
-
Paragraph 0028; 0032; 0033; 0034; 0041; 0046
(2017/10/06)
-
- Cobalt carbonyl ionic liquids based on the 1,1,3,3-tetra-alkylguanidine cation: Novel, highly efficient, and reusable catalysts for the carbonylation of epoxides
-
A series of novel cobalt carbonyl ionic liquids based on 1,1,3,3-tetra-alkyl-guanidine, such as [1,1-dimethyl-3,3-diethylguanidinium][Co(CO)4] (3a), [1,1-dimethyl-3,3-dibutylguanidinium][Co(CO)4] (3b), [1,1-dimethyl-3,3-tetramethyleneguanidinium][Co(CO)4] (3c), and [1,1-dimethyl-3,3-pentamethyleneguanidinium] [Co(CO)4] (3d), were synthesized in good yields and were also characterized using infrared spectroscopy, ultraviolet-visible spectroscopy, 1H nuclear magnetic resonance (NMR) spectroscopy, 13C NMR spectroscopy, high–resolution mass spectrometry, differential scanning calorimetry, and thermogravimetric analysis. The four compounds exhibited high thermal and chemical stability. In addition, the catalytic performance of these compounds was investigated in the carbonylation of epoxides, with 3a exhibiting the best catalytic activity without the aid of a base as the additive. The catalyst could be reused at least six times without significant decreases of the selectivity or conversion rate. Moreover, the catalyst system exhibited good tolerance with terminal epoxides bearing alkyl, alkenyl, aryl, alkoxy, and chloromethyl functional groups.
- Zhang, Wei,Han, Feng,Tong, Jin,Xia, Chungu,Liu, Jianhua
-
p. 805 - 812
(2017/05/29)
-
- 1,1,3,3-tetra alkyl guanidine carbonyl cobalt metal organic ion liquid and preparation method and application thereof
-
The invention discloses a 1,1,3,3-tetra alkyl guanidine carbonyl cobalt metal organic ion liquid. A structure formula of the ion liquid is shown in the description. The invention also discloses a preparation method of the ion liquid and application of the ion liquid in synthesizing of 3-hydroxy carboxylate by epoxy compound hydroesterification. The 1,1,3,3-tetra alkyl guanidine carbonyl cobalt metal organic ion liquid has the advantages that guanidine cation has the double functions of stabilizing tetra carbonyl cobalt cation and using as a Lewis acid to activate epoxy compound primer; compared with the traditional Co2(CO)8/azacycle catalytic system, the use of a nitrogen-containing organic matter is avoided; the ion liquid can be recycled and repeatedly used, the use rate of carbonyl cobalt is improved, and the production cost is reduced; the stability of the 1,1,3,3-tetra alkyl guanidine carbonyl cobalt metal organic ion liquid is high, the ion liquid can be repeatedly applied to catalysis of epoxy compound hydroesterification for multiple times, the catalytic reaction effect is good, and the separation and recycling are convenient.
- -
-
Paragraph 0061
(2017/08/31)
-
- A preparation method of the compound-carnitine
-
The invention discloses a preparation method of a levocarnitine compound, and the preparation method is applied to the technical field of pharmaceutical chemical synthesis. The preparation method comprises the following steps that (1), 4-chloroacetoacetic acid ethyl ester serves as the initial raw material, under the condition that a solvent exists, an asymmetric catalyst phosphine ligand ruthenium complex is applied for hydrogenation reduction, and (R)-4-chlorine-3-hydroxybutyrate ethyl ester is obtained by vacuum concentration and high vacuum distillation; (2), a trimethylamine squeous solution and the (R)-4-chlorine-3-hydroxybutyrate ethyl ester are added dropwise and slowly in the solvent comprising inorganic base, the dropping speed is controlled, low temperature reaction is carried out, then indoor temperature reaction is carried out, the pH of the mixture is adjusted to be 6 through concentrated hydrochloric acid after reaction is finished, and the levocarnitine compound is obtained by resin column purification. According to the method, the asymmetric catalytic reaction is applied, the two-step reaction that an optical active intermediate with high optical purity and high yield can be obtained and the optical active intermediate is converted to be the levocarnitine compound is the one-pot reaction, the water serves as the reaction solvent, the inorganic base is used for catalysis, the unique process of indoor temperature reaction is adopted, the product quality is good, the purity is high, the yield can reach up to 80%, the reaction steps of the preparation method are short, operation is easy, pollution to environment is small, and green resources are protected.
- -
-
Paragraph 0034; 0035
(2017/04/03)
-
- Asymmetric synthesis of lipitor chiral intermediate using a robust carbonyl reductase at high substrate to catalyst ratio
-
An NADPH-dependent carbonyl reductase (RpCR) from Rhodococcus pyridinivorans was discovered by genome mining for the asymmetric reduction of ethyl 4-chloro-3-oxo-butanoate (COBE). RpCR has been soluble expressed in Escherichia coli BL21(DE3). The highest activity is determined at pH 5.0 and 50 °C toward COBE. The apparent Km and kcat/Km are 0.39 mM and 1747 s-1 mM-1, endowing RpCR with high catalytic efficiency in reduction of COBE. Employing merely 0.1 g recombinant RpCR-GDH in a toluene-aqueous biphasic system, as much as 7.0 g COBE could be asymmetrically reduced into ethyl (S)-4-chloro-3-hydroxybutanoate [(S)-CHBE] (>99% ee) without addition of external cofactor, achieving molar isolation yield of 91%, substrate to biocatalyst ratio of 70 and space-time yield of 1480 g L-1 d-1. Our results indicate the robust RpCR could be potentially applied in the preparation of optically pure (S)-CHBE.
- Xu, Guo-Chao,Tang, Ming-Hui,Ni, Ye
-
supporting information
p. 67 - 72
(2015/11/28)
-
- Preparing method for linezolid and intermediate thereof
-
The invention relates to a preparing method for linezolid and an intermediate thereof. The preparing method includes the step that 4-chloracetyl acetate compound (I) serves as a starting raw material and is subjected to asymmetric chiral reduction, acetylation, condensation, ammonolysis, Hoffman degradation, acetylation and cyclization to obtain (S)-N-[[3-[3-fluoro-4-(4-morpholinyl) phenyl]-2-oxo-5-oxazolidinyl] methyl] acetamide (linezolid). Compared with other linezolid synthesis methods, the preparing method is high in total yield and product purity, raw materials are cheap and easy to obtain, flammable, combustible and poisonous reagents are avoided, and the production technology is safe and environmentally friendly.
- -
-
Paragraph 0046; 0047; 0048
(2016/11/02)
-
- Identification of an ε-keto ester reductase for the efficient synthesis of an (R)-α-lipoic acid precursor
-
Abstract A novel reductase (CpAR2) with unusually high activity toward an ε-keto ester, ethyl 8-chloro-6-oxooctanoate, was isolated from Candida parapsilosis. The asymmetric reduction of ethyl 8-chloro-6-oxooctanoate using Escherichia coli cells coexpressing CpAR2 and glucose dehydrogenase genes gave ethyl (R)-8-chloro-6-hydroxyoctanoate, a key precursor for the synthesis of (R)-α-lipoic acid, in high space-time yield (530 gL-1d-1) and with excellent enantiomeric excess (>99%). This bioprocess was shown to be viable on a 10-L scale. This method provides a greener and more cost-effective method for the industrial production of (R)-α-lipoic acid.
- Zhang, Yu-Jun,Zhang, Wen-Xia,Zheng, Gao-Wei,Xu, Jian-He
-
supporting information
p. 1697 - 1702
(2015/06/02)
-
- A highly efficient designer cell for enantioselective reduction of ketones
-
A designer cell, surf-crs-gdh, coexpressing carbonyl reductase (crs) and glucose dehydrogenase (gdh) on the cell surface has been constructed and its enzyme activities are compared with those of the corresponding cell, cyto-crs-gdh, coexpressing crs and gdh in cytosol. For various ketones, surf-crs-gdh exhibited 48- to 265-fold higher crs activity per unit protein compared to cyto-crs-gdh.
- Srivastava, Gautam,Pal, Mohan,Kaur, Suneet,Jolly, Ravinder S.
-
p. 105 - 108
(2015/02/19)
-
- Asymmetric synthesis of optically active methyl-2-benzamido-methyl-3-hydroxy-butyrate by robust short-chain alcohol dehydrogenases from Burkholderia gladioli
-
Three short-chain alcohol dehydrogenases from Burkholderia gladioli were discovered for their great potential in the dynamic kinetic asymmetric transformation of methyl 2-benzamido-methyl-3-oxobutanoate, and their screening against varied organic solvents and substrates. This is the first report of recombinant enzymes capable of achieving this reaction with the highest enantio- and diastereo-selectivity.
- Chen, Xiang,Liu, Zhi-Qiang,Huang, Jian-Feng,Lin, Chao-Ping,Zheng, Yu-Guo
-
p. 12328 - 12331
(2015/07/27)
-
- Asymmetric synthesis of (S)-4-chloro-3-hydroxybutanoate by sorbose reductase from Candida albicans with two co-existing recombinant Escherichia coli strains
-
An NADPH-dependent sorbose reductase from Candida albicans was identified to catalyze the asymmetric reduction of ethyl 4-chloro-3-oxobutanoate (COBE). The activity of the recombinant enzyme toward COBE was 6.2 U/mg. The asymmetric reduction of COBE was performed with two coexisting recombinant Escherichia coli strains, in which the recombinant E. coli expressing glucose dehydrogenase was used as an NADPH regenerator. An optical purity of 99% (e.e.) and a maximum yield of 1240 mM (S)-4-chloro-3-hydroxybutanoate were obtained under an optimal biomass ratio of 1:2. A highest turnover number of 53,900 was achieved without adding extra NADP+/NADPH compared with those known COBE-catalytic systems.
- Cai, Ping,An, Mingdong,Xu, Sheng,Yan, Ming,Hao, Ning,Li, Yan,Xu, Lin
-
p. 1090 - 1093
(2015/10/05)
-
- Identification of key residues in Debaryomyces hansenii carbonyl reductase for highly productive preparation of (S)-aryl halohydrins
-
Key residues of Debaryomyces hansenii carbonyl reductase in the determination of the reducing activity towards aryl haloketones were identified through combinatorial mutation of conserved residues. This study provides a green and efficient biocatalyst for the synthesis of (S)-aryl halohydrins.
- Xu, Guo-Chao,Shang, Yue-Peng,Yu, Hui-Lei,Xu, Jian-He
-
supporting information
p. 15728 - 15731
(2015/11/02)
-
- Characterization and identification of three novel aldo-keto reductases from Lodderomyces elongisporus for reducing ethyl 4-chloroacetoacetate
-
Lodderomyces elongisporus LH703 isolated from soil samples contained three novel aldo-keto reductases (AKRs) (LEAKR 48, LEAKR 49, and LEAKR 50). The three enzymes were cloned, expressed, and purified to homogeneity for characterization. These three AKRs shared 40% amino acid identity with each other. LEAKR 50 was identified as a member of AKR3 family, whereas the other two LEAKRs were identified as members of two novel AKR families, respectively. All the three AKRs required nicotinamide adenine dinucleotide phosphate as a cofactor. However, they showed diverse characteristics, including optimum catalyzing conditions, resistance to adverse reaction conditions, and substrate specificity. LEAKR 50 was estimated to be a promising biocatalyst that could reduce ethyl 4-chloroacetoacetate with high enantiomeric excess (98% e. e.) and high activity residue under adverse conditions.
- Ning, Chenxi,Su, Erzheng,Wei, Dongzhi
-
p. 219 - 228
(2015/02/05)
-
- Asymmetric bioreduction of ethyl (S)-4-chloro-3-hydroxy butanoate using dried baker's yeast in aqueous/ionic liquid biphasic system
-
A process of highly stereoselective reduction of ethyl 4-chloro-3- oxobutanoate to ethyl (S)-4-chloro-3-hydroxy butanoate with Baker's yeast was established. Compared with the aqueous/organic solvent biphasic system, the aqueous/ionic liquid biphasic system could eliminate high toxicity of organic solvent and make the convenience of product separation. Baker's yeast showed the best catalytic activity and enantioselectivity in the aqueous/[Bmim]PF6 biphasic system among the biphasic systems. Under the optimum conditions: [Bmim]PF6 180 g/L, dried baker's yeast 60 g/L, glucose 0.8 mol/L, ethyl 4-chloro-3-oxobutanoate 0.08 mol/L, temperature 30 °C, pH 7.0, 24 h and shaking speed 180 rpm, the yield and the e.e. value of (S)-4-chloro-3-hydroxy butanoate reached 92.6 % and 95.4 %, respectively.
- Tan, Zhongqin,Du, Huan,Han, Xiaoxiang,Yang, Zheyao,Wu, Xiaodan
-
p. 2695 - 2698
(2014/06/09)
-
- More efficient redox biocatalysis by utilising 1,4-butanediol as a 'smart cosubstrate'
-
1,4-Butanediol is shown to be an efficient cosubstrate to promote NAD(P)H-dependent redox biocatalysis. The thermodynamically and kinetically inert lactone coproduct makes the regeneration reaction irreversible. Thereby not only the molar surplus of cosubstrate is dramatically reduced but also faster reaction rates are obtained.
- Kara, Selin,Spickermann, Dominik,Schrittwieser, Joerg H.,Leggewie, Christian,Van Berkel, Willem J. H.,Arends, Isabel W. C. E.,Hollmann, Frank
-
supporting information
p. 330 - 335
(2013/03/29)
-
- Highly enantioselective double reduction of phenylglyoxal to (R)-1-phenyl-1,2-ethanediol by one NADPH-dependent yeast carbonyl reductase with a broad substrate profile
-
The activity and enantioselectivity of a carbonyl reductase from Pichia pastoris GS115 were evaluated with a series of carbonyl compounds including aryl aldehydes, ketones, α- and β-ketoesters. This recombinant enzyme possessed a broad substrate profile with the ability of reducing both aldehydes and ketones. Especially, the enzyme catalyzed the double reduction of phenylglyoxal to (R)-1-phenyl-1,2-ethanediol with 99% yield and 99% ee by coupling with d-glucose dehydrogenase for the regeneration of cofactor NADPH, representing the first example of effective reduction of both aldehyde and ketone functional groups in one molecule by using only one enzyme. Furthermore, this study provides valuable information for guiding the future application of this versatile biocatalyst.
- Li, Zhe,Liu, Weidong,Chen, Xi,Jia, Shiru,Wu, Qiaqiang,Zhu, Dunming,Ma, Yanhe
-
p. 3561 - 3564
(2013/04/24)
-
- Multi-enzymatic biosynthesis of chiral β-hydroxy nitriles through co-expression of oxidoreductase and halohydrin dehalogenase
-
To establish a system for the efficient one bacterial multi-enzymatic biosynthesis of both (R)- and (S)-β-hydroxy nitriles, we co-expressed alcohol dehydrogenases with opposite stereoselectivities, cofactor regeneration enzymes, and a halohydrin dehalogenase in Escherichia coli. By researching cofactor recycling and various co-expression strategies and by selecting and engineering the halohydrin dehalogenase, we engineered two E. coli strains, which were subsequently used in a cascade of reactions to produce chiral β-hydroxy nitriles with high enantiomeric excess directly from prochiral α-halo ketones. Three valuable pharmaceutical intermediates were prepared by means of this catalytic system, and substrate conversion reached about >99%. More importantly, the system is of low cost because there is no need for expensive cofactors or for expression and purification of the component enzymes. Copyright
- Chen, Shao-Yun,Yang, Chen-Xi,Wu, Jian-Ping,Xu, Gang,Yang, Li-Rong
-
p. 3179 - 3190
(2013/12/04)
-
- Formal total synthesis of the algal toxin (-)-polycavernoside A
-
A concise and largely catalysis-based approach to the potent algal toxin polycavernoside A (1) is described that intercepts a late-stage intermediate of a previous total synthesis; from there on, this challenging target can be reached in a small number of steps. Key to success was a sequence of a molybdenum-catalyzed ring-closing alkyne metathesis (RCAM) reaction to forge the macrocyclic frame, followed by a gold-catalyzed and strictly regioselective transannular hydroalkoxylation of the resulting cycloalkyne that allows the intricate oxygenation pattern of the macrolactone ring of 1 to be properly set. The required cyclization precursor 5 was assembled by the arguably most advanced fragment coupling process based on an Evans-Tishchenko redox esterification known to date, which was optimized to the extent that the precious coupling partners could be used in an almost equimolar ratio (6/7 1:1.3). The preparation of these building blocks features, inter alia, the power of the Sc(OTf) 3-catalyzed Leighton crotylation as well as the superb selectivities of alkene cross metathesis, asymmetric keto-ester hydrogenation, and the Jacobsen epoxidation/epoxide resolution technologies. Copyright
- Brewitz, Lennart,Llaveria, Josep,Yada, Akira,Fürstner, Alois
-
supporting information
p. 4532 - 4537
(2013/04/23)
-
- Facile access to chiral alcohols with pharmaceutical relevance using a ketoreductase newly mined from Pichia guilliermondii
-
Chiral secondary alcohols with additional functional groups are frequently required as important and valuable synthons for pharmaceuticals, agricultural and other fine chemicals. With the advantages of environmentally benign reaction conditions, broad reaction scope, and high stereoselectivity, biocatalytic reduction of prochiral ketones offers significant potential in the synthesis of optically active alcohols. A CmCR homologous carbonyl reductase from Pichia guilliermondii NRRL Y-324 was successfully overexpressed. Substrate profile characterization revealed its broad substrate specificity, covering aryl ketones, aliphatic ketones and ketoesters. Furthermore, a variety of ketone substrates were asymmetrically reduced by the purified enzyme with an additionally NADPH regeneration system. The reduction system exhibited excellent enantioselectivity (>99% ee) in the reduction of all the aromatic ketones and ketoesters, except for 2-bromoacetophenone (93.5% ee). Semi-preparative reduction of six ketones was achieved with high enantioselectivity (>99% ee) and isolation yields (>80%) within 12 h. This study provides a useful guidance for further application of this enzyme in the asymmetric synthesis of chiral alcohol enantiomers. Copyright
- Xu, Guochao,Yu, Huilei,Xu, Jianhe
-
p. 349 - 354
(2013/08/22)
-
- Experimental and computation studies on Candida antarctica lipase B-catalyzed enantioselective alcoholysis of 4-bromomethyl-β-lactone leading to enantiopure 4-bromo-3-hydroxybutanoate
-
Both enantiomers of optically pure 4-bromo-3-hydroxybutanoate, which is an important chiral building block in the syntheses of various biologically active compounds including statins, were synthesized from rac-4-bromomethyl-β- lactone through kinetic resolution. Candida antarctica lipase B (CAL-B) enantioselectively catalyzes the ring opening of the β-lactone with ethanol to yield ethyl (R)-4-bromo-3-hydroxybutanoate with high enantioselectivity (E>200). The unreacted (S)-4-bromomethyl-β-lactone was converted to ethyl (S)-4-bromo-3-hydroxybutanoate (>99% ee), which can be further transformed to ethyl (R)-4-cyano-3-hydroxybutanoate, through an acid-catalyzed ring opening in ethanol. Molecular modeling revealed that the stereocenter of the fast-reacting enantiomer, (R)-bromomethyl-β-lactone, is ~2 A from the reacting carbonyl carbon. In addition, the slow-reacting enantiomer, (S)-4-bromomethyl-β-lactone, encounters steric hindrance between the bromo substituent and the side chain of the Leu278 residue, while the fast-reacting enantiomer does not have any steric clash. Copyright
- Lim, Jung Yun,Jeon, Nan Young,Park, A-Reum,Min, Bora,Kim, Bum Tae,Park, Seongsoon,Lee, Hyuk
-
p. 1808 - 1816
(2013/07/19)
-
- Stereospecific reduction of methyl o-chlorobenzoylformate at 300 g·L-1 without additional cofactor using a carbonyl reductase mined from Candida glabrata
-
In order to search for oxidoreductases suitable for the preparation of methyl (R)-o-chloromandelate [(R)-CMM], the key intermediate for clopidogrel, the homologous proteins of Gre2p were expressed in Escherichia coli, among which CgKR1 showed the most satisfactory activity and stereoselectivity towards methyl o-chlorobenzoylformate (CBFM). Using the crude enzyme of CgKR1 and glucose dehydrogenase (GDH), as much as 300 g·L-1 of CBFM was almost stoichiometrically converted to (R)-CMM with excellent enantiomeric excess (98.7% ee). More importantly, the reaction could be performed without external addition of an expensive cofactor. The substrate profile indicates that keto esters serve as the most suitable substrate, which was confirmed by gram-scale preparations. Homology modeling and docking analysis revealed the molecular basis for the high stereoselectivity of CgKR1. These demonstrate not only the feasibility of in silico mining of novel enzymes based on sequence homology but also the applicability of this new reductase for the practical production of optically active (R)-CMM. Copyright
- Ma, Hongmin,Yang, Linlin,Ni, Yan,Zhang, Jie,Li, Chun-Xiu,Zheng, Gao-Wei,Yang, Huaiyu,Xu, Jian-He
-
experimental part
p. 1765 - 1772
(2012/08/08)
-
- Asymmetric reduction of ethyl (S)-4-chloro-3-hydroxy butanoate in an aqueous-organic solvent biphasic system using dried Baker's yeast
-
A process of highly stereoselective reduction of ethyl 4-chloro-3- oxobutanoate (ECOB) to ethyl (S)-4-chloro-3-hydroxy butanoate [(S)- ECHB] with Baker's yeast was established. The influence of volumetric phase ratio, substrate concentration, the amount of dried yeast cells and glucose on the yield of ethyl (S)-4-chloro-3-hydroxybutanoate and the enantiomeric excess (e.e.) (S) value were also investigated. A high yield and e.e. of ethyl (S)-4-chloro-3-hydroxybutanoate were observed in a biphasic system composed of phosphate buffer (0.05 mol/L, pH 7.0) and isooctane. Under the optimum conditions (in the water/isooctane biphasic system, Vaq/Vorg 3/1, dried yeast cells 50 g/L, glucose 0.6 mol/L, 30 °C, pH 7.0, substrate concentration 0.08 mol/L), the yield of ethyl (S)-4-chloro-3-hydroxybutanoate and the e.e. value of (S)-ECHB reached 93.9 % and 91.4 %, respectively, after 24 h reaction.
- Wu, Xiaodan,Han, Xiaoxiang,Zhou, Lingxiao,Li, Ang
-
scheme or table
p. 5151 - 5154
(2012/10/07)
-
- Ru-BINAP-catalyzed asymmetric hydrogenation of keto esters in high pressure carbon dioxide
-
Asymmetric hydrogenation of ethyl 4-chloro-3-oxobutyrate and dimethyl acetylsuccinate in high pressure CO2 using [RuCl2(C 6H6)]2-(R)-BINAP as the catalyst provides high ee values for the products.
- Turova, Olga V.,Kuchurov, Ilya V.,Starodubtseva, Eugenia V.,Ferapontov, Vladimir A.,Ikonnikov, Nikolai S.,Zlotin, Sergei G.,Vinogradov, Maxim G.
-
experimental part
p. 184 - 186
(2012/10/08)
-
- A new practical synthesis of ethyl (R)-(-)-4-Cyano-3-hydroxybutyrate from (S)-3-chloro-1,2-propanediol
-
A practical chemical synthesis of ethyl (R)-(-)-4-Cyano-3- hydroxybutyrate((R)-CNHB) has been accomplished from (S)-3-chloro-1,2- propanediol, which is a main by-product originating from (S,S)-Salen Co(III) catalyzed by hydrolytic kinetic resolution (HKR) of epichlorohydrin. The new synthetic approach demonstrated an efficient utilization of organic by-product for the asymmetric synthesis of the intermediate of atorvastatin.
- Jiang, Chengjun,Hong, Huabin
-
p. 520 - 521
(2012/11/06)
-