土豆中酪氨酸酶的提取及植物精油对其激活作用与动力学研究

本实验采取单因素分析法对土豆中酪氨酸粗酶的提取及反应条件进行优化,采用多巴速率氧化法筛选具有酪氨酸酶激活作用的植物精油,并通过酶动力学实验研究其激活机理。结果表明土豆中酪氨酸酶最佳提取条件为:料液比1:10(g/mL)、磷酸盐缓冲液pH6.8;最佳酪氨酸酶反应条件为:温度35℃、时间13 min、底物L-多巴/粗酶加入量比例5:5。在最佳提取条件与反应条件下筛选15种植物精油中发现百里香、茶树、依兰精油对酪氨酸酶激活作用显著(P>0.05),IC₅₀值分别为0.488、0.429、0.480 mg/mL,接近阳性对照补骨脂素IC₅₀值0.328 mg/mL,动力学实验发现百里香、茶树、依兰精油激活类型分别为非竞争型、混合型、混合型。

Extraction of Tyrosinase from Potatoes and Activation of Tyrosinase by Plant Essential Oils and Its Kinetics

The extraction and eaction of tryosinase from potato,and optimization by single factor experiment were been investigated in the study. The plant essential oil which could increase the catalytic activity of tryosinase was been screened by dopa rate oxidation method. Furthermore,enzyme kinetics model was used to study it's catalytic mechanism. Results showed that the optimal extraction conditions of tyrosinase in potatoes were as follows:solid-liquid ratio 1:10 (g/mL),phosphate buffer pH value 6.8. The optimum reaction conditions of tryosinase were as follows:reaction temperature 35℃,reaction time 13 min,and the substrate addition L-Dopa/crude enzyme addition ratio at 5:5. In the screening of 15 plant essential oils under the optimal extraction conditions and reaction conditions,thyme,tea tree and ylang-ylang had significant tryosinase activation(P>0.05),the IC₅₀ values were 0.488,0.429 and 0.480 mg/mL,close to the positive control psoralen IC₅₀ value 0.328 mg/mL. The kinetic experiments found that the activation types of thyme,tea tree and ylang-ylang oil were non-competitive,mixed and mixed.