固相PGA酶催化一步法合成氨苄西林的工艺研究
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摘要
采用吸附法将青霉素酰化酶(PGA)固定于AB-8树脂上,通过固相酶催化D-苯甘氨酸甲酯与6-氨基青霉烷酸(6-APA)反应制备氨苄西林,对酰化反应中的最适pH条件、溶剂体系、反应温度、反应时间、底物浓度进行实验研究。以氨苄西林收率和水解比为评价条件,固相酶最适催化pH值为6.5,通过正交实验分析表明,在异丙醇-PBS缓冲体系中,350mg PGA/AB-8(固定化酶活性113U·g~(-1))催化下,反应温度10℃,底物浓度(D-PGME∶6-APA)为3.5∶1,反应时间8h,氨苄西林收率达到71.3%。固相PGA酶重复回用6次,氨苄西林收率由71.3%降至63.6%。
Penicillin acylase(PGA)was immobilized on AB-8 resin by adsorption method,and ampicillin was prepared by solid phase enzyme catalyzed reaction of D-phenylglycine methyl ester with 6-aminopenicillanic acid(6-APA).The optimum pH conditions,solvent system,reaction temperature,reaction time,and substrate concentration in the acylation reaction were analyzed.Based on the yield and hydrolysis ratio of ampicillin,the optimum pH of the solid phase enzyme was 6.5.The orthogonal experimental analysis showed that 350 mg PGA/AB-8(immobilized enzyme)in the isopropanol-PBS buffer system.Under the activity of 113 U·g~(-1)),the reaction temperature was 10℃,the substrate concentration(D-PGME∶6-APA)was 3.5∶1,the reaction time was 8 h,and the ampicillin yield was 71.3%.The solid phase PGA enzyme was reused 6 times,and the ampicillin yield was reduced from 71.3%to 63.6%.
Penicillin acylase(PGA)was immobilized on AB-8 resin by adsorption method,and ampicillin was prepared by solid phase enzyme catalyzed reaction of D-phenylglycine methyl ester with 6-aminopenicillanic acid(6-APA).The optimum pH conditions,solvent system,reaction temperature,reaction time,and substrate concentration in the acylation reaction were analyzed.Based on the yield and hydrolysis ratio of ampicillin,the optimum pH of the solid phase enzyme was 6.5.The orthogonal experimental analysis showed that 350 mg PGA/AB-8(immobilized enzyme)in the isopropanol-PBS buffer system.Under the activity of 113 U·g~(-1)),the reaction temperature was 10℃,the substrate concentration(D-PGME∶6-APA)was 3.5∶1,the reaction time was 8 h,and the ampicillin yield was 71.3%.The solid phase PGA enzyme was reused 6 times,and the ampicillin yield was reduced from 71.3%to 63.6%.
引文
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[3]国家药典委员会.中华人民共和国药典第二部[M].北京:化学工业出版社,2015,1 147.
[4]刘芳.青霉素G酰化酶反应机理的说明[J].现代商贸工业,2017,(34):196-198.
[5]张琳,曹晓云.氨苄西林、氨苄西林钠及注射用氨苄西林钠中二聚体及其他相关物质的HPLC检查法[J].天津药学,2008(3):21-23.
[6]Giordano R C.Improving selectivity and productivity of the enzymatic synthesis of ampicillin with immobilized penicillin G acylase.Braz[J].Chem Eng & Technol,2004,21(4):519-529.
[7]Wei D Z,Zhu J H,Cao X J.Enzymatic synthesis of cephalexin in aqueous two-phase systems[J].Bioprocess Eng,2002,11(2-3):95-99.
[8]冯立峰.氨苄西林钠制备方法的研究[J].河北化工,2007,(6):40-41.