Studies On The Antioxidant Effective Component Of Citrus Reticulata’Chachi’Peel Extracts

Topic-based group have already studied the extraction technology of the antioxidantextraction of Citrus reticulata “Chachi” peel in laboratory-scale. We have also investigatedthe antioxidant extraction in vitro experiments and animal experiments， and found that theextraction have significant effects on D-Galactose-induced aging rats. The extraction couldreduce the malondialdehyde(MDA) contents in bloods，liver and brain. It could also increasethe SOD activity in the blood plasma and in the tissue of brain and liver， and the GSH-Pxactivity in the blood.This application of serum medicinal chemistry study of traditionalChinese medicine， extracts in Sera of rats to give drug prototype components for separationand identification;Six Polymethoxylated flavones were isolated and purified from theextracts， in which four of components are checked out into the prototype of bloodcomponents;Model of cellular oxidative stress injury have been constructed， and sixPolymethoxylated flavones and antioxidant activity of extracts tested by cell screening， asantioxidant Citrus Reticulata ‘Chazhi’ peel extracts provide a basis for material basis study ofeffect.Integrated content and the results of this article， summarized as follows:1. To develop a HPLC method for separation and analysis of principle component. Touse a HPLC method and confirm a better chromatographic condition through the mobilephase composition and pH value， eluting gradient， flow rate and the column temperaturefor system inspection. A chromatogram that has well separation， information-rich andmoderate peak time， was also established. 2. Using the HPLC method， to analyze the serum and determine the transitional bloodcomponents in rats after administration of Citrus reticulata “Chachi” peel extracts. Based onthe HPLC chromatogram， and combining references， comparison was carried out amoungthe peaks of blank serum， serum containing drug and Citrus reticulata “Chachi” peelextracts to determine the transitional blood components in rats after administration.Seventheen compounds were obtained after the administration of Citrus reticulate “Chachi”peel.extracts， in which five of them are prototype components， mainly of the bloodprototype components are Polymethoxylated flavones of Nobiletin and Tangeretin and otheringredients.3. Silica gel column chromatography， preparative thin-layer chromatography andrecrystallization were used to isolate the chemical constituents from Petroleum ether extractfraction and ethyl acetate extractsextrac which came from Citrus Reticulata ‘Chazhi’extractsof80%ethanol.And the structures of the compounds were identificated by spectral analysisand physicochemical properties.Then six compounds’ structures were elucidated byspectral(1H-NMR，13C-NMR)and physicochemical methods. They are5，7，8，4’-tetramethoxyflavone，5，6，7，3’，4’-pentamethoxyflavone，Nobiletin，3，5，6，7，8，3’，4’-heptamethoxyflavone， Tangeretin，5-hydroxy-6，7，8，3’，4’-pentamethoxyflavone.Nobiletin，Tangeretin and3，5，6，7，8，3’，4’–heptamethoxyflavonewere detected in the blood;5，7，8，4’-tetramethoxyflavone and5-hydroxy-6，7，8，3’，4’-pentamethoxyflavone were not detected in the blood.4. First， we set up the H2O2-induced cell oxidative stress injury model. And thenscreening the best concentration of H2O2and the best concentration of sample by the way ofdetecting the survival rate of PC12cells with MTT method. Using the best concentration ofsample to culture PC12cells first， and then co-culturing PC12cells with the bestconcentration of H2O2. The antioxidant indexes were the content of MDA and the vitality ofantioxidant enzymes(SOD and GSH-Px).The results showed that an experimental model of oxidative stress was established basedon which， contrast with normal control group， PC12cell injury group induced by hydrogenperoxide in culture can reduce SOD and GSH-Px activity and improve the MDA content.Testing component group on the activity of SOD and GSH-PX reduced inhibition andMDA content reduced.Into the protective effect on cells of the blood components not in bloodcomponents clearly. Polymethoxylated flavones which from Citrus Reticulata ‘Chazhi’ peelextracts play anti-oxidizing effect on neuronal injury by improving SOD and GSH-Px activityand reducing the MDA content.