| Objective: The person’s health seriously is affected by theresidual pesticides of the fruits and vegetables. In recentyears, people pay attention to the researchs which study howto remove the halogenated aromatic hydrocarbon pesticidetoxicity. Glutathione S-transferase （GST） can catalyze thebinding reaction of halogenated aromatic hydrocarbons poisonand reduced glutathione （GSH）, increases solubility andexcretes detoxification in normal body. The metabolism andelimilation of poisons is promoted by any factor whichincreases the activity of GST. Some fruits and vegetables sachas grapes and tea are rich in gallic acid. It is reported thatgallic acid has detoxification on the organic poisons, however,the detoxification mechanism is not clear and there are a littleof reports about it. In this paper, we stand on the newperspective of the vivo and vitro metabolic enzyme kineticsstudy of the rats to research whether the activity of the body detoxification enzyme （GST） is affected by gallic acid, whichchanges the metabolic rate of halogenated aromatichydrocarbons poison. So, we reveal the molecular mechanism ofdetoxification of the gallic acid, which safeguard the healthydiet, enhance physical fitness, and provide importanttheoretical guid for enhanceing the ability of the bodydetoxification. Methods: At first in vitro, the incubationsystem of the GST in rat liver microsomes was estanblished andthe GST activity was analysed by HPLC. The Vmax, Km andmetabolic clearance rate （CLint） of the GST were determined,which were better for researching the effect of gallic acid onGST and the related molecular mechanism. We evaluated theeffect of gallic acid on the metabolism of1-chloro-2,4-nitrobenzene （CDNB） as a representative of thepoison by IC50, Ki parameters, which preliminary judged twointeraction pattern and lay the foundation for the experimentin vivo. Secondly, in order to establish the evaluation systemof drug metabolism in rats, we estanblished the analysis methodfor determination the plasma concentrations of the DCNB whichwas used as the probe substrate by gas chromatographic.Then,we drew the concentration-time curve of DCNB and calculated thekinetic parameters with DAS3.2.1, which were used to analyze the effect of gallic acid on the metabolism of halogenatedaromatic hydrocarbons poison in vivo. In order to furtherexplore the comperhensive the effects of gallic acid on GST inrats, the mGST-1protein in the liver, kidney, intestine weredetermined by Western Blot, which comprehensively analyze andevaluate the overall impact of gallic acid on GST in vivo.Results: In vitro, the kinetics analysis of the rat livermicrosomal GST showed Vmax85.45nmol min-1 mg protein-1;Km was15.09μ mol L-1; Clint5.66mL min-1 mg protein-1.There was not time dependent inhibition of Gallic acid on GSTenzyme activity in vitro （IC50=14.79μ M）., The enzymeactivity of GST was restored by ultracentrifugation and adding10Km CDNB. The Ki was9.49μ M in inhibition kinetics analysis,which showed gallic acid was a moderate inhibitor for GST andbelonged to the mixed mechanism of reversible inhibition invitro. The CDNB metabolism may be reduced by gallic acid invitro. In vivo, the AUC0-∞of the experimental group decreasedfrom10363.5±1945.6to7075.1±1433.2comparing with thecontrol group. While the experimental group CL/F （14.1±3.4）was significantly higher than the control group （9.6±2.3）.The GST protein expression of liver, kidney, intestinal in theexperimental group was stronger than the control group. Conclusion: Form the dynamics analysis of the phase II enzymes,we inspected the effect of the gallic acid on GST enzymekinetics and exerted the detoxification for the halogenatedaromatic hydrocarbons pesticide. There was inhibitory effectof the gallic acid on GST in vitro, however, the expression ofGST protein in various organs was induced by gallic acid in rats,which increased the absolute content of the GST enzyme in vivoto promote the metabolism and removal of the halogenatedaromatic hydrocarbon pesticides. |
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