卤醇脱卤酶催化合成(R)-1-氯-3-苯氧基-2-丙醇的工艺优化
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摘要
以卤醇脱卤酶重组湿菌体E. coli BL21(pET28a-HHDH)为催化剂,催化外消旋的1-氯-3-苯氧基-2-丙醇的动力学拆分可以获得光学纯的(R)-1-氯-3-苯氧基-2-丙醇。本文系统地研究了卤醇脱卤酶催化合成光学纯(R)-1-氯-3-苯氧基-2-丙醇的影响因素,对反应pH、反应温度、菌体浓度、亲核试剂N3-浓度和底物浓度进行了探究。结果表明,卤醇脱卤酶催化合成(R)-1-氯-3-苯氧基-2-丙醇的最佳工艺条件为:pH为7.0,反应温度为28℃,菌体浓度为22.5g/L,亲核试剂NaN3的浓度为50mmol/L,底物外消旋1-氯-3-苯氧基-2-丙醇的浓度为10mmol/L。在此工艺条件下,(R)-1-氯-3-苯氧基-2-丙醇的ee值和收率分别为100%和16.97%。
Halohydrin dehalogenase(HHDH) was produced by recombinant strain E. coli BL21(pET28 a-HHDH). This recombinant E. coli was used as whole-cell catalysis to synthesize enantiopure(R)-1-chloro-3-phenoxy-2-propanol. The influencing factors of HHDH catalyzed synthesis of(R)-1-chloro-3-phenoxy-2-propanol were studied in the experiment. The reaction pH, reaction temperature,catalyst concentration, nucleophilic concentration(NaN3) and substrate concentration were investigated.The results showed that the optimal conditions were as follows: pH was 7.0, reaction temperature was28℃, the concentration of catalyst was 22.5 g/L, the concentration of sodium azide was 50 mmol/L, the concentration of racemic 1-chloro-3-phenoxy-2-propanol was 10 mmol/L. Under these conditions, the ee and yield of(R)-1-chloro-3-phenoxy-2-propanol was 100% and 16.97%, respectively.
Halohydrin dehalogenase(HHDH) was produced by recombinant strain E. coli BL21(pET28 a-HHDH). This recombinant E. coli was used as whole-cell catalysis to synthesize enantiopure(R)-1-chloro-3-phenoxy-2-propanol. The influencing factors of HHDH catalyzed synthesis of(R)-1-chloro-3-phenoxy-2-propanol were studied in the experiment. The reaction pH, reaction temperature,catalyst concentration, nucleophilic concentration(NaN3) and substrate concentration were investigated.The results showed that the optimal conditions were as follows: pH was 7.0, reaction temperature was28℃, the concentration of catalyst was 22.5 g/L, the concentration of sodium azide was 50 mmol/L, the concentration of racemic 1-chloro-3-phenoxy-2-propanol was 10 mmol/L. Under these conditions, the ee and yield of(R)-1-chloro-3-phenoxy-2-propanol was 100% and 16.97%, respectively.
引文
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[20] JANSSEN D B, MAJERI?ELENKOV M, HASNAOUI G, et al.Enantioselective formation and ring-opening of epoxides catalysed by halohydrin dehalogenases[J]. Biochemical Society Transactions, 2006,34:291-295.
[21] JIN H X, HU Z C, LIU Z Q, et al. Nitrite-mediated synthesis of chiral epichlorohydrin using halohydrin dehalogenase from Agrobacterium radiobacter AD1[J]. Biotechnology and Applied Biochemistry, 2012, 59(3):170-177.
[22] XUE F, YA X J, TONG Q, et al. Heterologous overexpression of Pseudomonas umsongensis halohydrin dehalogenase in Escherichia coli and its application in epoxide asymmetric ring opening reactions[J].Process Biochemistry, 2018, 75:1359-5113.
[23] ADER U, MUSCHALEK V, SCHNEIDER M P. Chromatographic resolution of chiral intermediates in beta-adrenergic blocker synthesis on chiral stationary phases[J]. Chirality, 1993, 5(7):554.
[24] CHEN C S, FUJIMOTO Y, GIRDAUKAS G, et al. Quantitative analyses of biochemical kinetic resolutions of enantiomers[J]. Journal of the American Chemical Society, 1982, 104(25):7294-7299.
[25] ZHENG G W, YU H L, ZHANG J D, et al. Enzymatic production of lmenthol by a high substrate concentration tolerable esterase from newly isolated Bacillus subtilis ECU0554[J]. Advanced Synthesis&Catalysis, 2010, 351(3):405-414.
[26] LIU Z Q, WU L, ZHENG L, et al. Biosynthesis of tert-butyl(3R,5S)-6-chloro-3,5-dihydroxyhexanoate by carbonyl reductase from Rhodosporidium toruloides in mono and biphasic media[J]. Bioresource Technology, 2017, 249:161-167.
[2] EDYTA?, PLENKIEWICZ J. Asymmetric reduction ofα-thiocyanatoketones by Saccharomyces cerevisiae and Mortierella isabellina—A new route to optically active thiiranes[J]. Tetrahedron Asymmetry, 2007, 18(10):1202-1209.
[3]汪海平,陈正国. 1-氯-3-苯氧基-2-丙醇的合成[J].广东化工,2005, 32(6):22-23.WANG H P, CHEN Z G. Synthesis of 1-chloro-3-phenoxy-2-propanol[J]. Guangdong Chemical Industry, 2005, 32(6):22-23.
[4] GOLUBOK Y O, V'YUNOV K A, PODZOLKOVA O A. Halogenated ethers. XIV. kinetics of the reaction of substituted phenols with1-chloro-2,3-epoxypropane[J]. Zhurnal Organicheskoi Khimii, 1979,15(10):2106-2112.
[5] SABITHA G, SATHEESH BABU R, RAJKUMAR M. Highly regioselective ring opening of epoxides and aziridines using cerium(Ⅲ)chloride[J]. Tetrahedron Letters, 2001, 42:3955-3958.
[6] TR?FF A, BOGáR K, WARNER M, et al. Highly efficient route for enantioselective preparation of chlorohydrins via dynamic kinetic resolution[J]. ChemInform, 2009, 40(12):4807-4810.
[7] CHUNGJYL,CVETOVICHR,AMATOJ,etal.Enantioselectivenitrile anion cyclization to substituted pyrrolidines. a highly efficientsynthesis of(3S, 4R)-N-tert-butyl-4-arylpyrrolidine-3-carboxylic acid[J].ChemInform,2005,36(39):3592-3601.
[8] OHKUMA T, TSUTSUMI K, UTSUMI N, et al. Asymmetric hydrogenation of alpha-chloro aromatic ketones catalyzed by eta6-arene/TsDPEN-ruthenium(Ⅱ)complexes[J]. Organic Letters, 2007, 9(2):255-257.
[9] MCCUBBIN J A, MADDESS M L, LAUTENS M. Enzymatic resolution of chlorohydrins for the synthesis of enantiomerically enriched2-vinyloxiranes[J]. ChemInform, 2007, 2008(2):289-293.
[10]王瑞,许耀辉,王克伟,等.环氧化物水解酶PvEH3的表达及手性邻二醇的合成[J].化工进展, 2018, 37(5):1933-1939.WANG R, XU Y H, WANG K W, et al. Expression of Pv EH3, a Phaseolus vulgarise poxide hydrolase, and synthesis of chiral vicinal diols[J]. Chemical Industry and Engineering Progress, 2018, 37(5):1933-1939.
[11]齐雁斌,马伟超,陈可泉.利用大肠杆菌全细胞催化赖氨酸发酵液生产1,5-戊二胺[J].化工进展, 2017, 36(5):1843-1847.QI Y B, MA W C, CHEN K Q. 1,5-pentanediamine production by using Escherichia coli whole-cell biocatalysis lysine fermentation liquid[J]. Chemical Industry and Engineering Progress, 2017, 36(5):1843-1847.
[12]陈佩,颜家保,武文丽,等.邻二甲苯高效降解菌的分离及其降解特性[J].化工进展, 2016, 35(2):565-569.CHEN P, YAN J B, WU W L, et al. Separation and biodegradation characteristics of a o-xylene degrading strain[J]. Chemical Industry and Engineering Progress, 2016, 35(2):565-569.
[13]邓杨敏,李冰麟,董文博,等.磷脂酰乙醇胺的酶促合成及底物抑制动力学[J].化工进展, 2017, 36(7):2601-2606.DENG Y M, LI B L, DONG W B, et al. Synthesis of phosphatidylethanolamine by enzymatic catalysis and its substrate inhibition kinetics[J]. Chemical Industry and Engineering Progress,2017, 36(7):2601-2606.
[14] YOU Z Y, LIU Z Q, ZHENG Y G. Properties and biotechnological applications of halohydrin dehalogenases:current state and future perspectives[J]. Applied Microbiology&Biotechnology, 2013, 97(1):9-21.
[15]陈少云.氧化还原酶-卤代醇脱卤酶多酶法合成手性β-羟基腈[D].杭州:浙江大学, 2013.CHEN S Y. Multi-enzymatic synthesis of chirolβ-hydroxyniles with oxidoreductase and halohydrin dehalogense[D]. Hangzhou:Zhejiang University, 2013.
[16]王雷,袁京,姚培圆,等.一种来源于运动替斯崔纳菌KA081020-065的新兴卤醇脱卤酶的表达纯化及性质分析[J].生物工程学报,2015, 31(5):659-669.WANG L, YUAN J, YAO P Y, et al. Expression and characterization of a novel halohydrin dehalogenase from Tistrella mobilis KA081020-065[J]. Chinese Journal of Biotechnology, 2015, 31(5):659-669.
[17]郑楷,汤丽霞.一种新型微生物卤醇脱卤酶的研究进展[J].生物技术通讯, 2010, 21(1):121-125.ZHENG K,TANG L X. Current research advances in bacterial halohydrin dehalogenases[J]. Lettersin Biotechnology, 2010, 21(1):121-125.
[18] SCHALLMEY A, SCHALLMEY M. Recent advances on halohydrin dehalogenases—From enzyme identification to novel biocatalytic applications[J]. Applied Microbiology and Biotechnology, 2016, 100(18):7827-7839.
[19] SPELBERG J H, JE V H V, TANG L, et al. Highly enantioselective and regioselective biocatalytic azidolysis of aromatic epoxides[J].Organic Letters, 2001, 3(1):41-43.
[20] JANSSEN D B, MAJERI?ELENKOV M, HASNAOUI G, et al.Enantioselective formation and ring-opening of epoxides catalysed by halohydrin dehalogenases[J]. Biochemical Society Transactions, 2006,34:291-295.
[21] JIN H X, HU Z C, LIU Z Q, et al. Nitrite-mediated synthesis of chiral epichlorohydrin using halohydrin dehalogenase from Agrobacterium radiobacter AD1[J]. Biotechnology and Applied Biochemistry, 2012, 59(3):170-177.
[22] XUE F, YA X J, TONG Q, et al. Heterologous overexpression of Pseudomonas umsongensis halohydrin dehalogenase in Escherichia coli and its application in epoxide asymmetric ring opening reactions[J].Process Biochemistry, 2018, 75:1359-5113.
[23] ADER U, MUSCHALEK V, SCHNEIDER M P. Chromatographic resolution of chiral intermediates in beta-adrenergic blocker synthesis on chiral stationary phases[J]. Chirality, 1993, 5(7):554.
[24] CHEN C S, FUJIMOTO Y, GIRDAUKAS G, et al. Quantitative analyses of biochemical kinetic resolutions of enantiomers[J]. Journal of the American Chemical Society, 1982, 104(25):7294-7299.
[25] ZHENG G W, YU H L, ZHANG J D, et al. Enzymatic production of lmenthol by a high substrate concentration tolerable esterase from newly isolated Bacillus subtilis ECU0554[J]. Advanced Synthesis&Catalysis, 2010, 351(3):405-414.
[26] LIU Z Q, WU L, ZHENG L, et al. Biosynthesis of tert-butyl(3R,5S)-6-chloro-3,5-dihydroxyhexanoate by carbonyl reductase from Rhodosporidium toruloides in mono and biphasic media[J]. Bioresource Technology, 2017, 249:161-167.