Quantitative Analysis And Pharmacokinetics Of Two Components In Evodia Rutaecarpa

Evodia rutaecarpa is the dry and nearly ripe fruit of Euodia rutaecarpa(Juss)Benth.,E.rutaecarpa(Juss)Benth.var.officinalis(Dode)Huang or E.rutaecarpa Benth.(Juss)var.bodinieri(Dode)Huang.Evodia rutaecarpa was first recorded in the "Shennong Herbal Medicine".It is spicy,bitter,hot and belongs to the liver,spleen,stomach and kidney meridian.It can reduce the vomiting and avoid a cold,and it is also the necessary medicine for pain relief in clinical warming.Evodia rutaecarpa can damage the liver blood.It was recorded in the literature of the past dynasties that it is poisonous.The adverse reactions of Evodia rutaecarpa can also be seen in clinical application now.The study of its toxicity in the literature has been documented for a long time and the mainly study is about sliver.The previous experiments of the research group showed that Evodia rutaecarpa(Euodia rutaecarpa(Juss)Benth.)was hepatotoxic.Five potential hepatotoxic material basises were screened.It includes two target components of this experiment.At present,the study on the in vivo process of two components in toxic dose has not been reported.It is very important for us to study pharmacokinetics in vivo for exploring the basis of toxic substances in the liver of Evodia rutaecarpa.In this paper,the potential hepatotoxicity components of Evodia rutaecarpa were studied.A method for the analysis of two components in Evodia rutaecarpa and in plasma was established to study the metabolic kinetics and plasma protein binding rate of evocarpine and dihydrorutaecarpine after hepatotoxicity induced by Evodia rutaecarpa.Study on the in vivo processes of two alkaloids,It also provides the basis for the rational use of Evodia rutaecarpa in clinic.The main research methods and results are as follows:The first part,we established the method for the determination of the contents of evocarpine and dihydroevocarpine in Evodia rutaecarpa.The experimental results showed that the ethanol extract of Evodia rutaecarpa 50%was the toxic part of Evodia rutaecarpa.Therefore,50%ethanol was refluxed to extract Evodia rutaecarpa.The HPLC method was used to simultaneously determine the evocarpine and dihydroevocarpine in the extract of Evodia rutaecarpa.The separation degree of two alkaloids was good.The response was high and the linear relationship was good in the range of 10～50 μg/mL.And the time of the analysis only need 20 mins.The results of the methodological evaluation showed that the method had good specificity and high sensitivity,and the precision,stability and recovery were all in accordance with the requirements of samples.The results showed that the content of dihydrorutaecarpine was(149.83±2.79)μg/g and the content of evocarpine was(462.53±0.89)μg/g.The second part was to establish a method for the determination of evocarpine and dihydroevocarpine in rat plasma by UPLC-MS/MS technique.The treatment method of plasma sample was optimized.The ethyl acetate extraction method with high recovery rate was selected.According to the research method of this paper,two alkaloids and internal standard can be separated well.And the analytical time only need 5 min.There was a good linear relationship between the two components in plasma in the concentration range of 0.1～250 ng/mL.The RSD value of precision and stability was less than 10%.The matrix effects of the two alkaloids were all between 80%～120%,and the recovery rate was between 80%and 120%.The results showed that the method had good specificity and high sensitivity,and the precision,stability,recovery and matrix effect were all in accordance with the requirements of biological samples.In the third part,the metabolic kinetics of evocarpine and dihydroevodiamine were studied.The results showed that evocarpine and dihydroevocarpine were rapidly absorbed in vivo.Pharmacokinetic results indicated that the evocarpine and dihydroevocarpine were rapidly absorbed in vivo.The average peak time in plasma was 0.54±0.10h and 0.58±0.13h,the average peak concentrations were 36.38±9.89 and 22.07±8.34 ng/mL,and the AUC was 119.8±29.8 and 67.25±21.12 ng/mL·h.The blood drug concentration of both decreased fast,and the half-lives were 2.80±0.61 h and 3.49±0.74 h.After about 4 half-lives,The amount of residual in blood was very small,indicating the fast metabolism in vivo.In the fourth part,the plasma protein binding rates of evocarpine and dihydroevocarpine were studied by equilibrium dialysis.The methodology investigation results show that the method of the stability and recovery rate are in line with the requirements of determination of biological sample.The results showed when the concentration of evocarpine and dihydroevocarpine in plasma was 5～60 μg/mL,the plasma protein binding rate of evocarpine was(37.43±2.34)%.The plasma protein binding rate of dihydroevocarpine was(39.53±1.32)%.The above research results would lay foundation for further study on prevention and treatment of hepatotoxicity and drug safety of Evodia rutaecarpa.