| Objective:Camellia taliensis(W.W.Smith)Melchior,which belongs to Camellia sect Thea..Camellia taliensis is a closely-related species to Camellia sinensis,a widely cultivated tea plant.The present researches on Camellia taliensis are mainly focused on resources and chemical constituents.The few reports on the biological activity limited on the in vitro antioxidant effects.This has greatly hindered the further development of Camellia taliensis The aim of this study was to analyze the chemical constituents,evaluate the effects of Camellia taliensis on liver injury caused by restraint stress in vivo and explore its mechanism primarily as well.Methods:1.Camellia taliensis leaves(DLC)were harvested in winter and spring.High performance liquid chromatography(HPLC)was utilized to analyze polyphenols and alkaloids qualitatively and quantitatively.2.The in vitro antioxidant capacity index(ORAC)was used to measure the antioxidant activity of DLC.3.The mouse restraint stress model was used to evaluate the effects of DLC.Experimental animals were randomly divided into control group,model group,DLC(200 mg·kg-1?100 mg·kg-1 and 50 mg·kg-1),and vitamin C group(250 mg·kg-1),10 mice in each group Each group were given intragastric administration for 7 days,once a day.The normal group and the model group were given the same volume of normal saline.After the last administration of 30 min,all animals except the control group mice were given restraint stress for 18 hours.Paraffin sections of liver tissue were stained by HE staining,the morphological and structural changes of liver tissue were observed.The activity of glutamic pyruvic transaminase(ALT)in liver tissue was determined by kit to confirm the occurrence of acute liver injury.The level of ORAC in liver tissue was determined by fluorescence enzyme labeling instrument.SOD,GSH,GSH-Px,Vit C and MDA in liver tissue were determined by kits.Griess method was used to determine NO and iNOS was detected by kits.IL-1? and TNF-? were detected by ELISA kit.4.The expression of Nrf-2 mRNA in restraint stress mouse liver tissue was detected by RT-PCR.Western-blot was utilized to determine Nrf2 protein both in nucleus and cytoplasm of hepatocytes.Results:1.Caffeine was the main alkaloid in DLC,with the contents of 3.88%and 4.65%in winter and spring respectively.It also contained a small amount of theobromine,theophylline was undetectable under this chromatographic condition.The concentration of EGCG was the highest in all catechins,the contents in winter and spring were 3.56%and 4.43%.The rank of catechins was:EGCG>ECG>EGC>EC>GCG>C>GA.2.The results of in vitro ORAC test showed that DLC had strong ability to quench free radicals.3.ALT in plasma of restraint stress mice increased significantly;ORAC,SOD,GSH,GSH-Px and Vit C in liver all decreased significantly,along with the remarkable increase of MDA,NO and iNOS;the mRNA of IL-1? and TNF-? also rised significantly;mild morphologic change was observed,the cytoplasm in hepatocytes around the central vein were loose and lightly stained,together with inflammatory cell infiltration in local lobules;With the treatment of DLC,the plasma ALT was lowered,SOD in liver was increased and ORAC was improved.Meanwhile,DLC could inhibited the production of MDA and NO and suppressed iNOS.The mRNA expressions of IL-1? and TNF-? were down-regulated,the ameliorative liver morphology was also observed.4.The mRNA of Nrf 2 in the model group increased significantly.DLC promoted Nrf 2 expression.Conclusion:Caffeine was the main purine alkaloid and the content of EGCG exceeds other catechins in DLC.Most of purine alkaloids and catechins in spring were higher than in winter DLC was a potent antioxidant and effective on acute liver injury induced by restraint stress. |
PDF Full Text Request