紫外-荧光分光光度法联用测定熊果酸对胰脂肪酶的抑制作用

在pH 8.0的Tris-HCl缓冲体系中,应用紫外-荧光分光度法作为研究手段,结合酶抑制动力学,研究了熊果酸对胰脂肪酶的抑制作用、抑制类型和抑制动力学常数,并探讨了熊果酸对胰脂肪酶表面疏水性的影响。结果表明,熊果酸是一种的竞争型胰脂肪酶抑制剂(半抑制率IC50为1.87±0.05 mg/mL,抑制常数Ki为0.68±0.03 mg/mL),且失活动力学时间进程能够证明熊果酸能很快的与胰脂肪酶发生作用并迅速的降低酶的活性;荧光光谱分析显示木犀草素能显著增强酪氨酸酶的表面疏水性,使胰脂肪酶活性位点的内部疏水性区域暴露出来,诱发了胰脂肪酶的解折叠,最终导致胰脂肪酶活性的降低。

Measurement of Ursolic Acid on the Inhibitory Activity of Pancreatic Lipase by UV-Fluorescence Spectrophotometer

The inhibitory effect,inhibitory type and inhibitory kinetics on pancreatic lipase by ursolic acid were investigated in pH 8.0 Tris-HC1 buffer system, and the effect of surface hydrophobicity on pancreatic lipase were also probed. The results showed that ursolic acid was a competitive inhibitor （the half inhibition concentration （IC50） and inhibition constant （Ki ） were obtained to be 1.87 ±0.05 mg/mL and 0.68 ±0.03 mg/mL, respectively）, The time-courses of inactivation kinetics time indicated that ursolic acid quickly interacted with pancreatic lipase and inhibited the acticity of enzyme rapidly, Fluorescence spectrum analysis showed that ursolie acid could significantly increase the surface hydrophobicity of pancreatic lipase, and induced the exposed of pancreatic lipase activity sites within the hydrophobic area and the unfolding of pancreatic lipase, finally lead to decrease the pancreatic lipase activity.