果蔬农药残留快速检测固定化酶片的制作研究

Study on the Preparation of Immobilized Enzymes for Rapid Detection of Pesticide Residues in Fruits and Vegetables

[目的]对胆碱酯酶的保藏性进行研究分析,以期得到保障农药残留检测的最佳酶活力。[方法]试验选定高酶活力的大豆酯酶为酶原,通过筛选最适反应温度、反应时间、离子浓度和最适pH,及最佳的反应显色体系来确定反应最适的条件,采用不同的显色体系酶抑制法,研究酶片的制作方法。[结果]酶片载体材料采用硝酸纤维膜,制作每张酶片取20μL大豆酶液,15μL 0.5%的BSA溶液,2μL 0.05%戊二醛于平底试管中。添加0.03 mol/L pH 7.0的磷酸缓冲液至总体积100μL,4℃固定10 h;制作每张显色片,需在1 cm×1 cm大小的滤纸片上加入50μL显色剂,30μL底物,4℃冷风吹干,封膜真空保存。[结论]固兰B盐和α-乙酸奈酯组合,适合作为酶抑制法检测有机磷农药残留的显色剂,且效果明显。

[Objective] The preservation of cholinesterase was studied in order to guarantee the best enzyme activity of pesticide residue detec-tion.[Method] The soybean esterase with high enzyme activity was selected as zymogen, optimum reaction conditions by screening the optimal reaction temperature, reaction time, concentration and pH, reaction and the best color system were determine.The method of making enzyme tablets was studied by using different color system and enzyme inhibition method.[ Result] Nitrocellulose membrane was used for enzyme piece of material, which made each enzyme take 20 μL soybean enzyme liquid, 15 μL 0.5% BSA solution, 2 μL 0.05% glutaraldehyde to the bot-tom of the tube.Add 0.03 mol/L pH 7.0 phosphate buffer and the total volume was 100 μL, 4 ℃ fixed 10 h.Made each color piece, should add 50 μL chromogenic agent in 1 cm ×1 cm filter size, 30 μL substrates, 4 ℃ cold wind blow dry, sealing membrane vacuum preservation. [ Conclusion] The combination of solid blue B salt andα-naphthyl acetate is suitable for the detection of organophosphorus pesticide residue by enzyme inhibition method, and the effect is obvious.