基于对硝基苯酚荧光内滤效应的脂肪酶活性测定

Detection of Lipase Activity Based on Fluorescence Inner Filter Effect of p-Nitrophenol

基于金纳米簇的荧光特性和对硝基苯酚的荧光内滤效应检测脂肪酶活性。利用谷胱甘肽制备金纳米簇,作为反应体系的荧光团,棕榈酸对硝基苯酯在脂肪酶的作用下水解为对硝基苯酚,充当荧光吸收剂,基于荧光内滤机理测定脂肪酶活性,并对反应条件进行优化。结果显示:棕榈酸对硝基苯酯质量浓度1.6 mg/mL、pH 7.5、温度50℃、反应时间20 min时,相对荧光强度(F_(0)-F)与脂肪酶活性在5.6~196 U/L范围内呈正相关,线性方程为y=2.0035+0.9368x,相关系数r2为0.9978,检出限(信噪比为3)1.3 U/L。该方法操作简便,灵敏度高,可应用于脂肪酶活性的测定。

Lipase activity was detected based on the fluorescence properties of gold nanoclusters(AuNCs)and the inner filter effect(IFE)of p-nitrophenol.Glutathione-modified AuNCs was used as fluorophore,and p-nitrophenol produced from the lipase-catalyzed hydrolysis of p-nitrophenol palmitate as fluorescence absorbent.The results showed that under the optimal conditions(p-nitrophenyl palmitate concentration 1.6 mg/mL,pH 7.5,temperature 50℃,and reaction time 20 min),the relative fluorescence intensity(F_(0)-F,y)was positively correlated with lipase activity(x)in the range of 5.6-196 U/L,which was described by the equation y=2.0035+0.9368x,with a correlation coefficient(r2)of 0.9978,and the detection limit was 1.3 U/L(at a signal-to-noise ratio of 3).The developed method is simple and sensitive,and can be applied to the detection of lipase activity.