Investigation on Hepatoprotective Activity of Some Medicinal Plants

Abstract

Introduction: The present work entitling was aimed to determine hepatoprotective effects of Zizyphus xylopyrus (ZX) leaves and Sphaeranthus indicus(SI) flower head extracts against carbon tetrachloride and alcohol induced hepatotoxicity in in vitro and in vivo as well as to isolate and characterize active compounds responsible for the hepatoprotective activity. Material and Methods: The total flavonoids content (TFC), total phenolic content (TPC) and total tannin content (TTC) were determined using quercetin and tannic acid equivalents, as standards while antioxidant activities of extracts were determined using standard in vitro methods. Extracts were subjected to evaluate in-vitro protective effects against CCl4 induced damage in HepG2 cell line as well as to evaluate in vivo protective, prophylactic and curative effects in alcohol and CCl4 intoxicated rats; respectively using silymarin as standard drug. Results: Quantitative estimation of TFC, TPC and TTC in ZX, showed that among all extracts highest amount of TFC and TPC was found in the ZXEAE where ZXEE extract contained highest amount of TTC. However in case of SI, highest amount of TPC and TTC were found more in SIEE while SIEAE contained highest amount of flavonoids. Among all extracts of ZX, the ZXEAE showed potent antioxidant activity in reducing power, DPPH, OHº, NOº, O2ˉ and inhibition of LPO assay followed by ZXEE. In case of SI, SIEE showed potent antioxidant activity followed by SIEAE. In HepG2 cell line study, treatment with ZXEAE (250 μg/ml) significantly (p<0.001) increased the cell viability by 89.9±2.68% by preventing CCl4 (1% w/v) induced cell damage in in-vitro HepG2 cell line which was comparable to the effects produced by silymarin 95.2±1.66%. Treatment with ZXEE and SIEE moderately increased the cell viability 67.3±2.56% and 63.9±2.56%, respectively. Whereas SIEAE extract showed mild increase in cell viability by 40.6±2.68% as compared to toxicant control. Further, in vitro active extracts were subjected for evaluation protective effects against alcohol (5 gm/kg) in vivo. Study results demonstrated that treatment with ZXEAE and SIEE extracts exhibit significant (p<0.001) dose dependent protective effects against alcohol induced damage; decreasing the elevated level of serum enzymes and MDA as well as increasing the decline level of enzymatic antioxidant; comparable to standard silymarin (50 mg/kg). The ZXEE and SIEAE extracts produce moderate protective effects against alcohol induced damage. Improved histopathological changes supported the serum enzyme levels and antioxidant data results. In case prophylactic study, pretreatment with ZXEAE and SIEE significantly (p<0.001) decreased the CCl4-induced increased serum liver enzymes level which was comparable to the silymarin. Pretreatment with ZXEE and SIEAE produced moderate prophylactic effects. However in curative study, treatment with ZXEAE, significantly (p<0.001) decreased the increased serum liver enzymes levels induced by CCl4 while ZXEE and SIEE produced moderately curative effects; compared to silymarin. Prophylactic and curative potential of the extracts were supported by improved histopathological changes as well as by significant reduction in pentobarbitone sodium induced prolongation of sleeping time. Based on in vitro and in vivo study results, ZXEAE and SIEE extract were subjected to isolation of compounds by coloum chromatography using silica gel (60-120#); which may be responsible for attributing the significant hepatoprotective effects. Compounds-rutin, quercetin and apigenin-7-glycoside, sphaeranthanolide were successfully isolated and characterized from ZXEAE and SIEE; respectively. Conclusion: Based on the phytochemical analysis, in vitro antioxidant activity as well as in vitro and in vivo hepatoprotective screening studies, it is hereby concluded that Zizyphus xylopyrus (leaves) and Sphaeranthus indicus (flower heads) demonstrated potential hepatoprotective activities in the tested models. Results suggested that the activities attributed may be due to antioxidant/immunostimulant property of the selected extracts. It is further concluded that identified flavonoids (rutin, quercetin and apigenin-7-glycoside) and sesquiterpene glycoside (sphaeranthanolide), are responsible for the attributing hepatoprotective effects. However the activity may also be partially by other phytoconstitutents present in these plants extract. Based on present study it is very evident and clear that these plant extracts can be used efficiently as prophylactic and curative measures in case of acute hepatic injury or other hepatic disease and disorders.