云南传统发酵豆豉中产β-半乳糖苷酶乳酸菌的筛选及其产酶条件的研究

目的从云南豆豉样品中筛选产β-半乳糖苷酶的乳酸菌,并对其产酶条件进行研究。方法从云南省元阳、红河、建水、石屏等地采集豆豉样品,并从中分离得到355株微生物。结果经明胶诱导、脱脂乳平板实验,复筛得到87株蛋白酶产生菌,从中筛选产β-半乳糖苷酶的乳酸菌。通过X-Gal平板实验,共获得34株产β-半乳糖苷酶菌株,通过酶活测定,最终筛选得到1株高产β-半乳糖苷酶菌株GJ-1-3L,经16S rDNA序列分析鉴定为短乳杆菌;GJ-1-3L在以葡萄糖为碳源、多聚蛋白胨为氮源、起始pH 6.5的MRS培养基中,接种量为4%,35℃发酵培养12 h,其β-半乳糖苷酶活性高达6.73 U/mL,Cu2＋、Ba2＋对酶活有抑制作用,而K2HPO4、MgSO4则能促进酶活。结论 GJ-1-3L菌株来源于豆豉,能够产生β-半乳糖苷酶发酵乳糖,同时产生乳酸,其在食品与乳品加工等方面具有很好的应用前景。

Screening of β-galactasidase producing Lactic acid bacteria from Yunnan traditional Douchi and its enzyme producing condition

Objective To screen lactic acid bacteria producing β-galactosidase from Yunnan traditional Douchi（formented soybean） and study the bacteria′s enzyme producing conditions.Method Douchi samples were collected from Yuanyang,Honghe,Jianshui,Shiping regions,Yunnan Province.Result 355 strains were isolated from these samples,among which 87 strains with protease-producing activity were screened by detecting the clear zone on the defatted milk plate,and protease activity was determined by Ninhydrin method.34 strains were selected by X-gal plates.Finally,the high-yield strain GJ-1-3L was rescreened and identified as Lactobacillus brevis according to its 16S rDNA sequence.Its enzyme producing conditions was also studied,and the results revealed that the optimal conditions for producing β-galactosidase were as follows： glucose as carbon source and polypeptone plus yeast powder as nitrogen source,initial pH 6.5,inoculum size 4%,fermentation temperature 35 ℃ and fermentation time 12 h.Under these conditions,the yield of β-galactosidase was 6.73 U/mL.It was also found that Cu2＋ and Ba2＋ could inhibit the enzyme activity,while K2HPO4 and MgSO4 could improve β-galactosidase production.Conclusion GJ-1-3L isolated from Douchi can secrete β-galactosidase and ferment lactose to produce lactic acid,and so has a good prospect in food and dairy processing application.