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核农学报  2015 

水蜜桃多酚氧化酶分离纯化及其酶学特性研究

DOI: 10.11869/j.issn.100-8551.2015.06.1121, PP. 1121-1128

Keywords: 水蜜桃,多酚氧化酶,纯化,酶学特性

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Abstract:

为建立水蜜桃多酚氧化酶(PPO)高效分离纯化方法,并明确该酶的酶学特性,本研究采用柱层析法对该酶进行分离纯化,对其反应特性及稳定性进行研究。结果表明,Phenyl-SepharoseCL-4B疏水柱层析法对水蜜桃PPO的纯化倍数与硫酸铵分级沉淀法相近,但其得率为89%远高于后者;采用Phenyl-SepharoseCL-4B、DEAESepharoseFastFlow和SephaorylHRS-200三级柱层析联用的方法对PPO分离纯化,获得PPO总纯化倍数为12.40倍,总得率为20%;该PPO组分经SDS-聚丙烯酰胺凝胶电泳分析得到单一蛋白条带,其分子量为81kDa。纯化后的水蜜桃PPO对(+)-儿茶素和(-)-表儿茶素的米氏常数分别为0.14mM和0.12mM,最适反应pH值为7.0,最适反应温度为40℃。纯化后的水蜜桃PPO对热以及低pH值都很不稳定。本研究结果为进一步研究PPO酶促褐变机制提供了理论指导,同时也为其他果蔬中PPO的分离纯化提供参考。

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