| Objiective:Turnning clinical curative effect of ZuoGuiYin anti-aging effect as the research orientation and using modern pharmacology and molecular biology experiment method,from the perspective of science revealled ZuoGuiYin anti-aging effects and its mechanism of action.So it can lay the foundation for further revealing its efficacy material base and guiding the development of new dosage forms.At the same time,it also provide the scientific theory basis for guiding clinical rational use of drugs and improving the clinical curative effect.Methods:(1)Used the high performance liquid chromatography(HPLC)method and Ultraviolet-visible spectroscopy(UV)to establishing the quality evaluation method of ZuoGuiyin,and respectively determined the content of loganin、5-HMF and polysaccharide in ZuoGuiyin extractive,in order to provide a stable and controllable experimental samples for later animal experiment;(2)Using drosophila survival experiment method to exam the influence of ZuoGuiYin on the half the time of deaths average life span and average maximum life span of fruit flies;(3)Using the D-galactose neck back subcutaneous injection method to copy the subacute aging rat model.At the same time,fill the stomach with ZuoGuiYin.Through observing the influence of ZuoGuiyin on the rat’s general status.behavior indicator and the hippocampal tissue pathology and related enzyme activity,in order to discussing the mechanism of ZuoGuiYin of slow the brain aging.On this basis,using the immunohistochemical method to inspect the influence of ZuoGuiYin on the hippocampus tissue apoptosis related proteins of aging rats and discussing the mechanism of action of anti-aging from the angle of molecular biology.Results:(1)The sample quality and peak area of loganin and 5-HMF in ZuoGuiyin extractive had good linear relationship in the range of 0.51～4.08μg(R2＝0.9998)and 0.26～2.04μg(R2= 0.9998).The average sample recovery rate was 101.24%(RSD= 1.40%)and 99.72%(RSD = 1.10%)respectively,the content of logain and 5-HMF were 0.246%and 0.212%;The phenol-sulfuric acid method of standard curve equation is Y= 15.284X—0.1154(R2 =0.9993),and the content of glucose and absorbance has a good linear relationship in the range of 0.0202—0.2018mg/mL,the content of total polysaccharides was 27.00mg/g(RSD=1.23%);(2)Both the high does and the low does of ZuoGuiyin can significantly extend the average life expectancy and the average maximum life span of fruit flies(P<0.05,P<0.01);(3)The rats which aged by the injection of D-galactose,were in the performance of a poor general condition with active oxygen free radicals and lipid peroxide increased.In addition,the aging rats showed up the pathological manifestations of the antioxidant capacity obviously decreased、Cell apoptosis increases and hippocampal tissue morphological structure damage;After the treatment with ZuoGuiyin,General status were improved in each dose group of rats.Furthermore,the vigor of SOD and T-AOC in brain showed varying degrees of increase.the MDA content in Brain tissue showed different degrees of lower、the level of brain hippocampus Bcl-2 protein was expressed and the Bax and Caspase-3 protein were expression.Conclusion:(1)This study established an HPLC method simultaneously determined the content of loganin and 5-HMF and this method was accurate and reliable repeatability.Using the phenol-sulfuric acid method determined the total polysaccharide content in ZuoGuiyin.This method was simple with good accuracy.stable and reliable.So these methods can provide reference for the quality control of ZuoGuiyin;(2)ZuoGuiyin can extend the life expectancy and the highest life span of flies.And the life-span extended along with the augment of ZuoGuiyin.(3)ZuoGuiyin can improve the learning and memory ability of aging rats and improve the shape and structure of brain tissue.Simultaneously,it can improve the antioxidant capacity in the brain tissue and inhibition of cell apoptosis function of normal play.These suggested ZuoGuiyin have anti-aging effect. |
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