木糖利用融合子D2的制备与乙醇发酵特性  

Preparation of Xylose Metabolic Fusant D2 and Its Ethanol Fermentation Characteristics

作　　者：周林[1] 邱瑞娇 林上龙 许少萍[2] 张英[2] ZHOU Lin QIU Ruijiao LIN Shanglong XU Shaoping ZHANG Ying(School of Biosciences and Biopharmaceutics, Guangdong Pharmaceutical University, Guangzhou 510006, China School of Chinese Herbal medicine, Guangzhou University of Chinese Medicine, Guangzhou 510006, China)

机构地区：[1]广东药科大学生命科学与生物制药学院,广东广州510006 [2]广州中医药大学中药学院,广东广州510006

出　　处：《生物质化学工程》2017年第4期17-24,共8页Biomass Chemical Engineering

基　　金：国家自然科学基金资助项目(8140140277);广州中医药大学"青年英才培养工程"资助(QNYC20140112);广东药科大学大学生创新创业训练计划项目(201510573024)

摘　　要：以树干毕赤酵母(Pichia stipitis)1960(Ps1960)与酿酒酵母(Saccharomyces cerevisiae)AADY(ScA ADY)为亲本菌株,采用双亲灭活原生质体技术制备木糖利用融合子,并对其制备条件进行了优化。优化后的原生质体制备条件为Ps1960采用2%蜗牛酶和1%纤维素酶在28℃酶解45 min,20W紫外灯距离10 cm照射3 min灭活;ScA ADY采用1.5%蜗牛酶和1%纤维素酶28℃酶解50 min,55℃水浴50 min灭活;均采用0.6 mol/L山梨醇为渗透压稳定剂。在该条件下,共得到22株融合子。通过测定各融合子在不同培养基条件下的生物量来评价其木糖代谢和乙醇耐受能力,最终获得能利用木糖高效发酵产乙醇、遗传性状稳定的融合子D2,并进行乙醇发酵条件优化。结果表明,在混合糖质量分数8%、木糖和葡萄糖质量比6∶1、5%接种量、30℃、160 r/min、培养72 h条件下,融合子D2发酵产乙醇的产量为40.58 g/L。The xylose metabolic fusants were prepared by fusion of inactivated parental protoplasts with Pichia stipitis 1960（ Ps1960） and Saccharomyces cerevisiae AADY（ ScA ADY） as parental strains. And the preparation conditions were optimized.The results showed that the Ps1960 protoplast was prepared by enzymolysis with 2 % snailase and 1 % cellulase for 45 min at 28 ℃,and then inactivation with 20 W ultroviolet lamp 10 cm away from the culture for 3 min. While ScA ADY protoplast was prepared by enzymolysis with 1. 5 % snailase and 1 % cellulase for 50 min at 28 ℃,and inactivation in water bath for 50 min at 55 ℃. And0. 6 mol/L sorbitol was used as osmotic stabilizer. And then,22 fusants were obtained. The biomasses of the fusants under different culture medium conditions were determined to evaluate the ability of metabolism of xylose and ethanol tolerance. Thus,from 22 fusants,the fusant D2,which could ferment ethanol from xylose,with good hereditary feature was selected for ethanol fermentation. Furtherly,fusant D2 could produce 40. 58 g/L ethanol under the conditions of 8 % mixed sugar,mass ratio of xylose and glucose 6∶1,inoculum size of 5 % and cultured for 72 h with shaking 160 r/min at 30 ℃.

关 键 词：酿酒酵母 树干毕赤酵母 原生质体融合 灭活 生物乙醇 

分 类 号：TQ223.122[化学工程—有机化工] TQ920.6[轻工技术与工程—发酵工程]