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从鲫鱼和青虾肝脏中分离谷丙转氨酶(GPT),以比活力为指标,L25(56)正交试验与极差分析法研究了GPT酶活力测定的最适条件,测定了Nic(氯硝柳胺)和CuSO4(硫酸铜)对GPT活力的影响。正交试验分析鲫鱼GPT活力测定最适条件:酶质量浓度2.0g/L、底物浓度2.0μmol/L、反应体系pH值7.0、反应温度30℃、反应时间55min,青虾GPT活力测定最适条件酶质量浓度3.0g/L、底物浓度0.5μmol/L、反应体系pH值7.5、反应温度45℃和反应时间45min。Nic和CuSO4对鲫鱼和青虾GPT活力均显示诱导作用,其0.002g/L时对鲫鱼GPT的诱导率分别为641.01%和61.22%,对青虾的分别为45.99%和67.57%。研究结果表明:2种GPT活力测定条件相差较大,酶质量浓度和底物浓度影响作用最大。鲫鱼和青虾GPT对Nic和CuSO4敏感度高,推定GPT可能是Nic和CuSO4中毒的生化靶点之一。
GPT was isolated from the liver of crucian carp and freshwater shrimp, and the optimum conditions of GPT enzyme activity were determined by L25 (56) orthogonal test and range analysis with specific activity as the indicator, The effect of CuSO4 (copper sulfate) on GPT activity was studied. Orthogonal test analysis GPT activity of crucian carp determination of the optimum conditions: enzyme concentration 2.0g / L, substrate concentration 2.0μmol / L, the reaction system pH 7.0, the reaction temperature 30 ℃, the reaction time 55min, the best determination of freshwater shrimp GPT activity Conditional enzyme concentration 3.0g / L, substrate concentration 0.5μmol / L, the reaction system pH 7.5, the reaction temperature 45 ℃ and the reaction time 45min. Nicotine and CuSO4 showed an induction effect on GPT activity of Carassius auratus and shrimp, and the induction rate of GPT of Carassius auratus at the concentration of 0.002g / L was 641.01% and 61.22% respectively, while they were 45.99% and 67.57% respectively. The results showed that there was a big difference between the two kinds of GPT activity assay conditions, and the effect of enzyme concentration and substrate concentration was the greatest. The carp and freshwater shrimp GPT sensitivity to Nic and CuSO4 high, presumed GPT may be one of the biochemical targets of Nic and CuSO4 poisoning.