福林酚法高通量测定牛奶中蛋白酶活力的优化

Optimizing of High Throughput Determination of Protease Activity in Milk Using Folin-phenol Method

[目的]优化福林酚法测定牛奶中蛋白酶活力的试验条件,解决检测结果重复性差、检测效率不高等问题。[方法]用酶标仪测定牛奶中蛋白酶活力,同步对比各试验条件对测定结果的影响。[结果]酶标仪测定时,显色保温时间15 min以上、取样350μL、福林试剂浓度0.33~0.5 mol/L时,牛奶中蛋白酶活力测定结果重复性好,检测效率高。[结论]酶标仪测定牛奶中蛋白酶活力是一种检测效率高、结果重复性好的方法,更适合乳品企业大批量筛查牛奶中蛋白酶活力。

[Objective]To optimize the conditions for the determination of protease activity in milk using the national standard method-Folin-phenol method.[Method]By measuring the protease activity in milk,the influence of various experimental parameters on the results was compared and analyzed.[Result]When using an enzymelinked immunosorbent assay(ELISA) reader for measurement,the color retention time should be more than 15 minutes,and a sample of 350 μL.When the concentration of Folin reagent is 0.33~0.5 mol/L,the absorbance constant K value of the standard curve meets the requirements.[Conclusion]When detecting protease activity in milk,it is necessary to control the color retention time,sampling volume,and clarity of the sample solution.After the color reaction is fully completed,the system of the standard solution and sample solution has stabilized,and there is no significant change in the absorbance value within 1 hour of further insulation or room temperature storage,which has no significant impact on the results.