藏灵菇源克鲁维酵母M3菌株胆盐水解酶的分离纯化研究  

Purification of bile salt hydrolase from Kluyveromyces marxianus M3 isolated from Tibetan kefir

作　　者：潘昌莉[1] 张红星[1] 耿新[1] 贾慧[1] 刘慧[1] 

机构地区：[1]北京农学院食品科学学院,北京102206

出　　处：《中国酿造》2011年第3期58-62,共5页China Brewing

基　　金：北京市自然科学基金资助项目(No.5092008);北京市重点建设学科资助项目(PXM2009_014207_078172)

摘　　要：为获得藏灵菇马克斯克鲁维酵母M3菌株胆盐水解酶(BSH)的纯品,探讨了BSH粗品的分离纯化方法。粗酶液采用硫酸铵分级沉淀,再以DEAE-Sepharose Fast Flow离子交换介质,分别考察缓冲液pH值、流速和洗脱方式等对BSH分离纯化的影响。结果表明,在40%～70%饱和度的硫酸铵条件下BSH提取效率最高。最佳层析条件为采用pH值为6.5、50mmol/L磷酸钠缓冲体系、1.5mL/min的流速,进行分步洗脱(100mmol/L、350mmol/L、500mmol/L～600mmol/L NaCl及50mmol/L磷酸盐缓冲液3步洗脱)。在优化纯化条件下BSH的比活力可达479.55AU/mg,是原粗酶液的23.66倍。In order to obtain the pure bile salt hydrolase （BSH） produced by Kluyveromyces marxianus M3 isolated from Tibetan kefir, the purification methods of crude BSH were studied. The crude BSH was precipitated by ammonium sulfate, and then was purified by DEAE-Sepharose Fast Flow ion exchange chromatography. The effects of buffer pH, flow rate and elution modes of ion exchange chromatography on the purification of BSH were investigated respectively. The results showed that the highest BSH extraction efficiency appeared when the saturation of ammonium sulfate ranged between 40% and 70%. The optimal chromatography conditions were： 50mmol/L sodium phosphate at pH value 6.5 was used as buffer, the flow rate was 1.5ml/min, and three-step elution of 100mmol/L, 350mmol/L and 500mmol/L-600mmol/L NaCl with 50mmol/L sodium phosphate was adopted. The activity of BSH reached 479.55AU/mg after purified under optimal conditions, which was 20 times higher than that of the crude BSH solution.

关 键 词：藏灵菇 马克斯克鲁维酵母 胆盐水解酶 分离纯化 

分 类 号：TS201.3[轻工技术与工程—食品科学]