产β-葡聚糖酶耐盐鲁氏酵母的筛选及鉴定

Identification and Screening of β-glucanase-producing, Salt-tolerant Zygosaccharomyces rouxii

摘要:

从农家自制辣椒酱中分离纯化了一株耐盐酵母，通过β-1,3葡聚糖酶鉴定（刚果红染色法）确定其具有合成、分泌胞外β-1,3葡聚糖酶特性。NaCl耐受性研究确定其具有高耐盐性，能经144 h的适应期后在24% NaCl的培养基B中稳定生长120 h。经26s rRNA基因序列分析鉴定为鲁氏酵母A（Z. Rouxii A）。Z. Rouxii A发酵培养中存在二次生长现象，其生物量在24 h和48 h达到峰值，分别比18 h的6.38 g/L提高了46.55%和87.15%，而21 h后葡萄糖浓度仅维持在1.57 g/L左右，说明：Z. Rouxii A生长过程中合成、分泌的β-葡聚糖酶持续降解发酵培养基中添加的β-1,3-1,6-葡聚糖，为其二次生长提供了碳源和能源。Z. Rouxii A的β-1,3-葡聚糖酶活性曲线与生长曲线基本趋势一致，最大酶活性随着菌体自溶（12 h、24 h和48 h）而迅速降低，48 h达到酶活峰值15.23 U/mL，说明：Z. Rouxii A的β-1,3-葡聚糖酶合成与细胞生长偶联。以上数据确认该菌为产β-葡聚糖酶高耐盐鲁氏酵母。

Abstract:

A salt-tolerant yeast strain was isolated and purified from a homemade chili sauce. Congo red agar plate method confirmed that the strain produced extracellular β-1,3-glucanase. The NaCl-tolerance test showed that the strain had high salt-tolerance, with stable growth observed for 120 h in medium B containing 24% NaCl after an adaptation period of 144 h. The strain was identified as Zygosaccharomyces rouxii A via 26S rRNA sequence analysis. Secondary growth phenomenawere observed during fermentation where the biomass reached peak values at 24 and 48 h, with an increase of 46.55% (9.35 g/L) and 87.15% (11.94 g/L), respectively, compared with that at 18 h (6.38 g/L). However, the glucose concentration remained constant at 1.57 g/L after 21 h. The results indicated that the β-1,3-glucanase produced by Z. rouxii A degraded the 1,3-1,6-glucan present in the substrate, thus providing carbon and energy sources for the secondary growth of cells. Additionally, β-1,3-glucanase activity curve was consistent with the growth curve for Z. rouxii A, which showed that maximum enzyme activity rapidly decreased with cell autolysis (12, 24, and 48 h), and peak enzyme activity level (15.23 U/mL) was reached at 48 h. The results suggest that β-1,3-glucanase synthesis was coupled with cell growth and that the strain could produce this enzyme in the presence of high salt concentration.