一种从未经巴氏杀菌的牛奶中纯化碱性磷酸酶的三步方法。
A three step approach for the purification of alkaline phosphatase from non-pasteurized milk.
作者信息
Upadhyay Lata Sheo Bachan, Verma Nishant
机构信息
Department of Biotechnology, National Institute of Technology, Raipur, 492010 India.
出版信息
J Food Sci Technol. 2015 May;52(5):3140-6. doi: 10.1007/s13197-014-1578-1. Epub 2014 Oct 11.
In this study, a three step purification of alkaline phosphatase from non-pasteurized milk has been described. It included cream extraction, n-butanol treatment and acetone precipitation. Different parameters such as buffer concentration, temperature, pH, substrate concentration, acetone and n-butanol treatment were optimized to maximize the enzyme activity. The enzyme was fruitfully purified up to homogeneity from the milk, with percentage recovery and fold purification of 56.17 and 17.67 respectively. The kinetic parameters were determined to be 0.927 mM (Km) and 55.86 μM/min (Vmax), with specific activity of 11.31 U/mg. Other optimized parameters were estimated as a buffer concentration of 0.5 M with pH 9.0, temperature optima at 37 °C, with n-butanol and acetone concentration of 20 % (v/v) and 50 % (v/v) respectively. This approach provides a simple and effective method for the purification of alkaline phosphatase from non-pasteurized milk.
本研究描述了从生牛奶中三步纯化碱性磷酸酶的方法。该方法包括乳脂提取、正丁醇处理和丙酮沉淀。对不同参数如缓冲液浓度、温度、pH值、底物浓度、丙酮和正丁醇处理进行了优化，以最大化酶活性。该酶成功地从牛奶中纯化至均一，回收率和纯化倍数分别为56.17%和17.67倍。动力学参数测定为0.927 mM（Km）和55.86 μM/min（Vmax），比活性为11.31 U/mg。其他优化参数估计为缓冲液浓度0.5 M，pH值9.0，最适温度37°C，正丁醇和丙酮浓度分别为20%（v/v）和50%（v/v）。该方法为从生牛奶中纯化碱性磷酸酶提供了一种简单有效的方法。
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