优良酵母菌的筛选、鉴定及发酵生产乙醇的研究

Studies on screening,identification and producing ethanol of excellent yeasts

从几种可能富含发酵木糖-葡萄糖酵母菌的材料中分离得到68株酵母菌,分别对其发酵、产乙醇、耐乙醇、木糖利用及木糖-葡萄糖发酵能力进行研究,筛选出发酵木糖-葡萄糖能力较强的酵母菌株。通过镜检观察、耐渗透压、耐受pH值、同化碳源和氮源试验,研究筛选菌株的形态和生理特性,并通过26S rRNA序列测定及系统进化分析对其进行分子鉴定,最后利用木糖-葡萄糖发酵培养基进行产乙醇试验。结果表明:分离菌株中S3和Y4菌株发酵木糖-葡萄糖能力较强,经分子鉴定分别为异常毕赤酵母(Pichia anomala)和热带假丝酵母(Candida tropicalis)。菌株Y4和对照菌株酿酒酵母(Saccharomyces cerevisiae)(N)产乙醇能力显著高于菌株S3(P<0.05),且菌株Y4和N间无显著差异(P>0.05);而S3与N组合其乙醇产量显著高于菌株Y4,N,S3单独及Y4与N组合,但显著低于S3与Y4组合及S3,N,Y4三者组合(P<0.05),其中三者组合的乙醇产量最高,为39.74 g/L。

68 strains of yeasts were isolated from the materials of fresh grass, withered lawn grass, rotted fruit, soil and rice wine and the best strains fermenting xylose-glucose were screened based upon the capabilities of fermenting, producing ethanol, ethanol tolerance, xylose utilization and xylose-glucose fermentation. The morphological and physiological characteristics of selected strains were measured by microscopic observation and the tolerance to osmotic pressure, pH and assimi- lation of carbon and nitrogen, respectively, and they were identified by 26 S rRNA sequencing and the phylogenetic analysis. Finally, the test of producing ethanol from xylose-glucose was carried out.The results showed that strains $3 and Y4 were of stronger ability to ferment xylose-glucose and were identified as Pichia anomala and Candida tropicalis, respectively. Strain Y4 and control strain N had higher ethanol yield than strain $3 （P〈0.05）, and there was no significant difference between strains Y4 and N （P〉0.05）. Ethanol yield by combination of $3 and N was significantly higher than those by S3 ,Y4 and N alone, or the combination of Y4 and N,but significantly lower than those by the combinations of S3 and Y4 or S3, N and Y4 （P〈0.05）.The combination of S3, N and Y4 had the highest ethanol yield（39.74 g/L）.