p-NPG法是目前进行β-葡萄糖苷酶活性测定时最常用的一种方法,但测活条件对不同来源糖苷酶活性有显著影响,不但降低了试验数据的可靠性与真实性,也不利于各文献中菌株酶活力大小的比较。为建立酿酒酵母β-葡萄糖苷酶活性测定条件,以商品酵母红佳酿为研究对象,分别利用单因素及正交试验考察了反应时间、温度、底物浓度及缓冲液pH对酶活性的影响。结果表明最佳测活条件为:采用pH 5.5柠檬酸-磷酸氢二钠缓冲体系,40 mmol/L底物p-NPG,50℃反应10 min;研究发现红佳酿酵母具有较高β-葡萄糖苷酶活性,与其他11株商品酿酒酵母菌株相比其胞外酶活最高,且在培养36 h时达到最大值。

p-NPG(4-nitrophenyl-β-D-glucopyranoside) method is commonly used for β-glucosidase activity determination,however,quantifying conditions significantly influence enzyme activity,not only decrease the reliability and authenticity of results,but also disadvantaged to the comparability among different papers.In order to build β-glucosidase activity determination conditions which suit for wine yeast,a selected yeast Vintage Red was used to test the influence of reaction time,temperature,substrate concentration and pH values on the activity of glucosidase by single factor and orthogonal tests.The results showed that the best conditions for quantifying β-glucosidase activity were as follows:pH 5.5 citrate phosphate buffer,40 mmol /L p-NPG solution,incubation at 50 ℃ for 10 min.Vintage Red yeast also showed relative high β-glucosidase activity,higher in extracellular activity than those of the other11 wine yeasts,and with peak production at 36 h growth.