生物工程學(xué)報(bào) 25 June 2009, 25(6):863～868
響應(yīng)面法優(yōu)化熱帶假絲酵母104菌株產(chǎn)羰基還原酶發(fā)酵培養(yǎng)基
王普, 孫立明, 何軍邀
浙江工業(yè)大學(xué)藥學(xué)院, 杭州 310032
摘 要: 通過菌種優(yōu)選得到產(chǎn)高選擇性羰基還原酶的熱帶假絲酵母(Candida tropicalis) 104菌株, 可不對(duì)稱還原1-(3,5-二三氟甲基)苯基乙酮生成(S)-1-(3,5-二三氟甲基)苯基乙醇, 對(duì)映體過量值>99.9%,。采用部分因子實(shí)驗(yàn)設(shè)計(jì)考察發(fā)酵培養(yǎng)基中各組分對(duì)產(chǎn)酶的影響, 結(jié)果表明, 酵母粉,、葡萄糖和NH4Cl濃度對(duì)產(chǎn)酶影響顯著,。繼而采用最陡爬坡路徑逼近最大響應(yīng)區(qū)域, 并結(jié)合中心組合實(shí)驗(yàn)和響應(yīng)面對(duì)3個(gè)顯著性因素進(jìn)行分析, 得到優(yōu)化的發(fā)酵培養(yǎng)基組成: 葡萄糖47.14 g/L, 酵母粉13.25 g/L, NH4Cl 2.71 g/L, MgSO4·7H2O 0.4 g/L, KH2PO4 1 g/L和K2HPO4 1 g/L,。采用該優(yōu)化培養(yǎng)基, 供試菌株的羰基還原酶活力達(dá)851.13 U/L, 較優(yōu)化前提高了65.2%,。
關(guān)鍵詞: 熱帶假絲酵母, 羰基還原酶, (S)-1-(3,5-二三氟甲基)苯基乙醇, 響應(yīng)面, 培養(yǎng)基優(yōu)化
Medium optimization for enhanced production of carbonyl reductase by Candida tropicalis 104 by response surface methodology
Pu Wang, Liming Sun, and Junyao He
College of Pharmaceutical Science, Zhejiang University of Technology, Hangzhou 310032, China
Abstract: Using response surface method, we optimized the medium for the asymmetric whole cell biotransformation by Candida tropicalis 104. This strain was used for microbial reduction of 1-[3,5-bis(trifluoromethyl)phenyl] ethanone to (S)-1-[3,5-bis(trifluoromethyl)phenyl] alcohol, with enantiomeric excess(e.e.) reached more than 99.9%. Fractional factorial design was used to evaluate the effects of medium components on carbonyl reductase activity of Candida tropicalis 104. Yeast extract, glucose and NH4Cl were the most important factors among six tested variables that influence the enzyme activity for the biotransformation process. Based on the experimental results, the path of steepest ascent was undertaken to approach the optimal region of these factors. Central composite design and response surface analysis were subsequently employed for further optimization. The optimal medium for Candida tropicalis 104 was composed of (in g/L): glucose 47.14, yeast extract 13.25, NH4Cl 2.71, MgSO4·7H2O 0.4, KH2PO4 1, K2HPO4 1. Under the optimum conditions, the maximum enzyme activity of 852.75 U/L in theory and 851.13 U/L in the experiment were obtained, with an increase of 65.2% compared to the original medium components.
Keywords: Candida tropicalis, carbonyl reductase, (S)-1-[3,5-bis(trifluoromethyl)phenyl] alcohol, response surface methodology, medium optimization
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