产β-葡萄糖醛酸苷酶菌株胃肠道耐受性与酶学特性研究

Gastrointestinal Tolerance and Enzymatic Characteristics of A Strain Producing β-glucuronidase

为发酵转化黄芩苷生产活性产物黄芩素、研发益生菌和中草药黄芩协同联用绿色混合型饲料添加剂产品奠定基础,试验采用酶解底物显色法和薄层层析法,从健康鸡肠道内容物中分离筛选出一株产β-葡萄糖醛酸苷酶的JY24菌株,经菌落形态、生理生化及16SrDNA序列分析,鉴定为枯草芽孢杆菌。结果表明:(1)JY24菌株对人工胃液、人工肠液均有一定耐受能力,胆盐最高耐受浓度为0.3%~0.5%;(2)JY24菌株所产β-葡萄糖醛酸苷酶的最适温度为35℃,40℃、50℃和60℃处理1 h相对酶活分别为72.96%、31.63%和0;(3)最适pH 6.5,在pH 6.0和7.0环境下,酶活相对稳定,处理1 h相对酶活分别为73.54%和73.63%;(4)浓度为2.5 mmol/L的Ca^2+、Fe^3+和Zn^2+可使酶活升高11.48%~21.05%,对该酶有激活作用,Fe^2+和Cu^2+可使酶活性下降15.84%~18.28%,对酶活有抑制作用,K^+、Na^+、Mg^2+和Mn^2+对酶活无明显影响。

To produce baicalein through fermentation transformation of baicalin, develop a green mixed feed additive product with a synergy of probiotics and a Chinese herb medicine Scutellaria baicalensis, a β-glucuronidase-producing strain designated as JY24 was separated from the intestinal contents of healthy chickens by enzymatic substrate chromogenic reaction and thin layer chromatography(TLC). The strain was identified as Bacillus subtilis by bacterial colony morphology, physiological and biochemical analysis and 16 SrDNA sequence analysis. The results indicated 1) JY24 strain showed certain tolerance after being inoculated in artificial gastric juice and intestinal liquid for different time. The highest tolerance concentration of JY24 strain against bile salt was 0.3%~0.5%. 2) The optimum reaction temperature was 35 ℃, and the relative enzyme activity were respectively 72.96%,31.63% and 0 at 40 ℃, 50 ℃ and 60 ℃ for 1 h. 3) The optimum pH was 6.5, under pH6.0 and 7.0 environment, the enzyme activity was relatively stable, and the relative enzyme activity were respectively 73.54% and 73.63% for 1 h. 4) 2.5 mmol/L of Ca^2+,Fe^3+ and Zn^2+ could increase 11.48%~21.05% of the β-glucuronidase activity, whereas Fe^2+ and Cu^2+ decreased 15.84%~18.28% of the enzyme activity,K^+, Na^+, Mg^2+ and Mn^2+ had no significant effect on the enzyme activity.