Establishment And Application Of The Method To Screen Hepatoprotective Drugs

Objective:(1)To establish a stable and convenient method to screen active sites of hepatoprotective,and active sites in thermal transformation products from swertiamarin also have been detected,which was to find out the effective material bases of mileensis to resist hepatitis.(2)To establish five kinds of models of chemical liver injury, observe hepatoprotective effect of Dai medicine YaJieShaBa,and to preliminarily explore its mechanism,which was to provide prevention and treatment of liver injury in clinic with experimental basis.Methods:(1) To replicate the mice model of acute liver injury by CCl4and detect its proper dose, duration and pathways to cause toxic effect,and by comparing with the positive control group,the stability of this method has been detected by repeating experiments.(2)With the mice model of acute liver injury by CCl4and tracing research including pharmacological screening and chemical separation, active sites in thermal transformation products from swertiamarin have been detected.(3) Hepatoprotective effectiveness of Dai medicine YaJieShaBa and its mechanism have been studied through five different mice models of liver injury caused by CCl4、 D-GLaN、AAP、TAA、ANIT.Results:(1) After a single intraperitoneal injection in mice with solution of Olive oil containing0.2%CCl4(0.1ml/10g.bw), the level of serum ALT and AST in model group is higher than control group (P<0.01),which showed this mice model of acute liver injury was successful. The level of serum ALT in group given the200mg/kg Bifendate was lower than model group(P<0.01),which suggested its effects of protecting liver and reducing transaminase,so the Bifendate has been selected as the positive control medicine.(2)The initial results showed the level of serum ALT in group given soluble fraction (1200mg/kg)in thermal transformation products from swertiamarin was lower than model group (P0.01); Levels of serum ALT in two groups respectively given insoluble fractions (400mg/kg and1200mg/kg) of thermal transformation products from swertiamarinwas were lower than model group (P<0.05、P<0.01);And levels of serum ALT and AST in groups of the soluble fraction and insoluble fraction(both were400mg/kg) of swertiamarin thermal transformation products were lower than model group (P<0.01, P<0.05); Levels of serum ALT and AST in groups of insoluble fractions (200mg/kg and100mg/kg given respectively)were lower than model group (P<0.05);Screening research suggested that the level of serum ALT in group given Sample A (400mg/kg) from insoluble fraction was lower than model group (P<0.05).(3)Injecting D-Galn(800mg/kg)in mice through enterocoelia and AAP(500mg/kg) given orally,after24h, the level of serum ALT and AST in model group were higher than control group (P<0.01),and16h after a single intraperitoneal injection in mice with TAA(100mg/kg), the level of serum ALT and AST in model group was also higher than control group (P<0.01).48h after giving mice100mg/kg ANIT by intragastric administration, the level of serum ALT and Tbil in model group was higher than control group (P<0.01).The results showed experimental methods above can successfully replicate the mice model of acute liver injury.(4) The level of serum ALT in high dose group of YaJieShaBa was lower than the model group by CCl4(P<0.01),and although low dose group of YaJieShaBa could decrease level of serum ALT in mice,comparing with model group,it had no significant difference(P>0.05).The level of serum AST in high dose group of YaJieShaBa was lower than the model group of liver injury by D-Galn(P<0.05),but could not significantly depress the increasing of AST(P>0.05). The level of serum ALT in high dose group of YaJieShaBa was lower than the model group by AAP(P<0.05), but could not significantly depress the increasing of AST(P>0.05). High dose group of YaJieShaBa could not obviously depress the increasing of serum ALT caused by TAA(P>0.05),and high dose group of YaJieShaBa could significantly depress the increasing of serum ALT caused by ANIT(P<0.05),based on above, which showed YaJieShaBa had the protection effect on liver injury caused by above five chemical materials.Conclusions:(1) The stable mice model of acute liver injury can be successfully established20h after a single intraperitoneal injection in mice with solution of Olive oil containing0.2%CC14(0.1ml/10g.bw).(2) Soluble and insoluble fractions of thermal transformation products from swertiamarin could decrease ALT and AST in serum in model group attacked by CCl4,and sample A coming from insoluble fraction could also reduce the ALT activity in serum in mice of liver injury by CCl4, which suggested soluble and insoluble fractions of swertiamarin thermal transformation products,as well as sample A had the effects on protecting liver and reducing transaminase.(3)The different mechanisms of mice models of acute liver injury could be replicated by respectively giving0.2%CC14,800mg/kgD-Galn,500mg/kg AAP,100mg/kg TAA and100mg/kg ANIT,the five different Liver toxicants.(4) ALT and AST activity in the model groups respectively caused by CC14, D-GLaN, TAA,AAP and ANIT could be depressed with YaJieShaBa,which showed YaJieShaBa had the effects on protecting liver and reducing transaminase in these five mice models of acute liver injury.And these effects might work through the depression of lipid peroxidation and enhancing the body functions to eliminate radicals.