李子健¹，黄 丹^1，2，张 曼²，郭辉祥³，胡 娟¹，罗 政⁴，罗惠波^1，2*

（1.四川轻化工大学 生物工程学院，四川自贡 643000；

2.四川轻化工大学 酿酒生物技术及应用四川省重点实验室，四川自贡 643000；

3.舍得酒业股份有限公司，四川遂宁 629209；4.四川省古川酒业有限公司，四川邛崃 611530）

摘要：以某制曲车间提供的培养了22 d的浓香型中高温大曲为原料，在不同培养温度下，通过稀释涂布平板法进行多次筛选，并用平板划线法进行分离纯化，得到了两株耐高温真菌。通过形态学及真菌ITS、18 s RNA高通量测序技术鉴定，确定分别为微小根毛酶、布氏横梗霉，两株真菌分别标记为Q1、M1。该菌在45 ℃条件下正常生长繁殖。通过发酵对筛选到的两株耐高温霉菌进行产酶特性比较。以小麦为发酵底物，模拟曲房发酵条件，对温度、湿度进行控制，原料经过润粮、粉碎和灭菌后进行接种、发酵。分别设置发酵温度为25 ℃、45 ℃：发酵湿度为40%、70%、95%。研究发现Q1菌株产糖化酶的能力受湿度影响较大，M1菌株产糖化酶的能力受温度影响较大，Q1、M1菌株所产糖化酶酶活最高分别为126 U/g、122 U/g：Q1菌株在高湿较高温度条件下有很好的产酶能力，蛋白酶活最高达到83.59 U/g，M1菌株产蛋白酶的能力受温度影响较小，受湿度影响大。Q1、M1菌株所产蛋白酶酶活最高分别为83.240 U/g、91.662 U/g。最高糖化酶、蛋白酶酶活均在发酵条件为45 ℃、95%的湿度下获得。

关键词：大曲耐热霉菌：菌种鉴定：糖化酶活力：蛋白酶活力

中图分类号：TS201.3   文献标识码：A   文章编号：1674-506X（2020）03-0001-0007

Isolation and Identification of Two Heat-resistant Molds in Daqu at Medium-high Temperature and Comparison of Their Enzyme Production Characteristics

LI Zi-jian¹, HUANG Dan^1，2, ZHANG Man², GUO Hui-xiang³, HU Juan¹, LUO Zheng⁴, LUO Hui-bo^1，2*

（1.College of Bioengineering, Sichuan University of Science and Engineering, Zigong Sichuan 643000, China;

2.Liquor Making Bio-Technology and Application of Key Laboratory of Sichuan Province, Zigong Sichuan 643000, China;

3.Shede Distillery Co., Ltd., Suining Sichuan 629209, China;

4.Guchuan Distillery Co., Ltd., Qionglai Sichuan 611530, China）

Abstract：Two strains of high temperature resistant fungi were obtained from 22 days of luzhou-flavor medium and high temperature daqu cultured in a daqu making workshop. Through morphological identification an  high-throughput sequencing of fungal ITS and fungal 18 s RNA, the two fungal strains were identified as Rhizomucor pusillus (Q1) and Lichtheimia blakesleeana (M1), which grew and reproduced normally at 45 ℃ . The enzyme production characteristics of two strains of high temperature resistant mold were compared by fermentation. Wheat was used as the fermentation substrate to simulate the conditions of starter fermentation, and the temperature and humidity were controlled. The fermentation temperature was set as 25 ℃ and 45 ℃ respectively. The fermentation humidity is 40% , 70% and 95%. It was found that the ability of strain Q1 to produce saccharinase was greatly affected by humidity, while that of strain M1 was greatly affected by temperature. The highest enzyme activity of saccharinase produced by strains Q1 and M1 was 126 U/g and 122 U/g, respectively. Under high humidity and high temperature, strain Q1 had good enzyme production capacity, and the highest protease activity was 83.59 U/g. The protease production capacity of strain M1 was less affected by temperature and more affected by humidity. The highest enzyme activity of proteases produced by strains Q1 and M1 was 83.240 U/g and 91.662 U/g, respectively. The highest saccharifying enzyme and protease activity were obtained at 45 ℃ and 95% humidity.

Keywords：Daqu heat-resistant mould; identification of strains; activity of saccharifying enzyme; protease activity

doi：10.3969/j.issn.1674-506X.2020.03-001

收稿日期：2020-04-01

基金项目：四川省科技厅2018年重点研发项目（2019YF0518）；四川省古川酒业横向项目（HX2016260）

作者简介：李子健（1987-），男，硕士研究生。研究方向：酿酒生物技术及应用。

*通讯作者

引文格式：李子健,黄丹,张曼,等.中高温大曲中两株耐热霉菌的分离鉴定及产酶特性比较[J].食品与发酵科技,2020,56(3):1-7.