中国中药杂志

2015, v.40(23) 4616-4622

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利用一种耐热耐糖β-葡萄糖苷酶制备黄芩素的研究
Preparation of baicalein using thermophilic and sugar-tolerant β-glucosidase

李石平;温建辉;赵祎武;黄文哲;裴建军;王振中;赵林果;萧伟;
LI Shi-ping;WEN Jian-hui;ZHAO Yi-wu;HUANG Wen-zhe;PEI Jian-jun;WANG Zhen-zhong;ZHAO Lin-guo;XIAO Wei;Jiangsu Kanion Parmaceutical Co.,Ltd.;State Key Laboratory of New-tech for Chinese Medicine Pharmaceutical Process;The Key Laboratory for the New Technique Research of Traditional Chinese Medicine Extraction and Purification;College of Chemical Engineering,Nanjing Forestry University;

摘要(Abstract):

对利用一种耐热耐糖β-葡萄糖苷酶水解黄芩苷制备黄芩素进行了研究。该酶催化黄芩苷转化为黄芩素的反应能在醋酸-醋酸钠缓冲液中良好地进行,酶的最适p H 5.5,最适反应温度85℃,温度在85℃以下,p H 5~7.5酶的稳定性良好。酶的最大反应速率Vmax为0.41 mmol·L~(-1)·min~(-1),米氏常数Km为3.31 mmol·L~(-1)。不同的金属离子对酶的活性有不同的影响,醋酸-醋酸钠缓冲液中的Na+对酶有活化作用。反应体系中葡萄糖醛酸浓度低于0.6 mol·L-1时,对酶有活化作用,当单糖浓度大于0.6 mol·L~(-1)时,对酶有抑制作用,当单糖浓度达到1.2 mol·L~(-1)时,酶活抑制率为50%,糖耐受性较好。通过实验得出较佳反应条件为黄芩苷-酶液1 g∶0.2 m L,pH 5.5的醋酸-醋酸钠缓冲液,反应时间为10 h时,反应温度为85℃,酶解率可达97%。
The reaction conditions of baicalin hydrolyzed into baicalein by a kind of thermophilic and sugar-tolerant β-glucosidase were studied in this paper. The β-glucosidase could catalyze baicalin into baicalein well in the acetic acid-sodium acetate buffer. The optimal enzyme activity was at 85 ℃ and p H 5. 5. The enzyme was stable at the temperature less than 85 ℃ and p H range of 5-7. 5.The maximum reaction rate Vmaxand michaelis constant Kmwere 0. 41 mmol·L~(-1)·min~(-1) and 3. 31 mmol·L~(-1) respectively. Different metal ions had different effects on the activity of enzyme. Na+existing in acetic acid-sodium acetate buffer had an activation effect on enzyme. The enzyme activity was enhanced by the concentrations of glucose below 0. 6 mol·L~(-1),and was gradually inhibited when monosaccharide concentration was over 0. 6 mol·L~(-1). When the monosaccharide concentration reached 1. 2 mol·L~(-1),the inhibition rate of enzyme activity was about 50%,which showed good glucose tolerance. The good reaction conditions through the experiment have been determined as follows,the substrate∶ enzyme dose was 1 g∶ 0. 2 m L,acetic acid-sodium acetate buffer pH 5. 5,reaction temperature 85 ℃,reaction time 10 h,and the enzymatic hydrolyzation ratio could reach 97%.

关键词(KeyWords): 耐热;耐糖;β-葡萄糖苷酶;黄芩苷;黄芩素
thermophilic;glucose-tolerance;β-glucosidase;baicalin;baicalein

Abstract:

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基金项目(Foundation): 国家“重大新药创制”科技重大专项(2013ZX09402203)

作者(Author): 李石平;温建辉;赵祎武;黄文哲;裴建军;王振中;赵林果;萧伟;
LI Shi-ping;WEN Jian-hui;ZHAO Yi-wu;HUANG Wen-zhe;PEI Jian-jun;WANG Zhen-zhong;ZHAO Lin-guo;XIAO Wei;Jiangsu Kanion Parmaceutical Co.,Ltd.;State Key Laboratory of New-tech for Chinese Medicine Pharmaceutical Process;The Key Laboratory for the New Technique Research of Traditional Chinese Medicine Extraction and Purification;College of Chemical Engineering,Nanjing Forestry University;

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