To Study The Effective Parts Of Total Lignans In Arctium Tomentosum Mill.seeds And Pharmacokinetics

Objective:To study the main effective components of total lignans in Arctium tomentosum Mill.seeds as the research object,the quality analysis of Arctium tomentosum Mill.seeds from different areas in Xinjiang was analyzed,the best process of extraction and enrichment were investigated and prepared the effective parts of total lignans.The absorption and distribution of arctiin and arctigenin in rats were preliminary studied to provide theoretical data for the further development of the characteristic ethnic medicine of Arctium tomentosum Mill.seeds.Methods:?1?The high performance thin-layer chromatography was used for the qualitative and quantitative analysis of arctiin and arctigenin in Arctium tomentosum Mill.seeds.The determination of the total lignans content in Arctium tomentosum Mill.seeds from the different areas with ultraviolet-visible spectrophotometry in which the arctiin was used as the reference substance was established and the methodological investigation was completed.High performance liquid chromatography was used to determine the content of arctiin in Arctium tomentosum Mill.seeds of different areas and completed the methodological investigation.?2?The single-factor test combined with the method of Box-Behnken design was used to optimize the extraction process of the effective parts of total lignans in Arctium tomentosum Mill.seeds by taking the comprehensive score of total lignans content,arctiin content and extract yield as evaluation indexes.?3?Based on the comprehensive scores of total lignans content and arctiin content as the evaluation index,selected eight kinds of macroporous adsorption resins to adsorb and elute the effective parts of total lignans from Arctium tomentosum Mill.seeds.The effects of sample concentration,sampling velocities,maximum sampling volume,diameter height ratio,water volume of removal impurities,elution solvent and the volume of elution solvent and elution velocity were all investigated on the enrichment of the effective parts of total lignans from Arctium tomentosum Mill.seeds.?4?The effective parts of total lignans was prepared according to the best extraction and enrichment process,and the content of arctiin and arctigenin in the effective parts of total lignans were determined by high performance liquid chromatography and completed the methodological investigation.Ultraviolet-visible spectrophotometry was used to determine the content of total lignans in the effective parts of total lignans and completed the methodological investigation.?5?After intragastric administration the total lignans of Arctium tomentosum Mill.seeds in rats,blood and tissue samples were collected at different time points,and the content of arctiin and arctigenin in plasma and tissue were measured by high performance liquid chromatography.PKSolver 2.0 pharmacokinetic and pharmacodynamics data processing software was used to calculate pharmacokinetic parameters.The distribution of arctiin and arctigenin in tissues also were analyzed.Results:?1?High performance thin-layer chromatography of Arctium tomentosum Mill.seeds:using silica gel G plate,chloroform:methanol?48:5?as the developing solvent,the obtained map had the clear spots and good separation effect,and the content of arctiin and arctigenin is different.The content of arctiin in six batches of Arctium tomentosum Mill.seeds was in the range of 26.15 mg/g⁴2.20 mg/g;the content of arctigenin was in the range of 4.52 mg/g¹8.89 mg/g.The content of total lignans in six batches of Arctium tomentosum Mill.seeds was determined by ultraviolet-visible spectrophotometry,ranging from 73.16 mg/g to 183.97 mg/g.The content of arctiin in six batches of Arctium tomentosum Mill.seeds was determined by HPLC from 17.46 mg/g to 78.34 mg/g.?2?The single factor test showed that the extraction time range was 40 to 80 min,the alcohol addition was 12 to 16 times,the ethanol concentration was 50%to 70%,and the number of extraction times was twice.Combined with the Box-Behnken response surface test,the best extraction process for total lignans in Arctium tomentosum Mill.seeds was finally determined as follows:15 times the amount of 58%ethanol to extract twice under reflux for 62 min each time.?3?The best enrichment process was to use HPD-100 resin,diameter height ratio was 1:10,the concentration of sample was 0.2 g/mL,4 BV at a flow rate of 1.5BV/h.Wash the water-soluble impurities with 7 BV distilled water,4 BV 70%ethanol was eluted at the flow rate of 1 BV/h,the validation experiment showed that the content of total lignans in the effective parts of total lignans obtained by macroporous resin enrichment was more than 90%,and the arctiin content was more than 30%.The results showed that the process was stable and feasible.?4?The content of total lignans in the effective parts of the prepared total lignans was 95.68%,the content of arctiin was 55.71%,and the content of arctigenin was 2.99%.?5?After intragastric administration of effective parts of total lignans from Arctium tomentosum Mill.seeds in rats,the pharmacokinetic parameters of arctiin are:T_max?time for maximum of drug concentration?is 8 h,C_max?maximum of drug concentration?is 39.95?g/mL,AUC_0-t?AUC_0-??the area under concentration-time curve?are 713.12 and 760.43?g·h/mL,MRT?mean residence time?is 19.58 h,ke?elimination rate constant?is 0.06 h^-1,t_1/2?half-life time?is 11.5 h,CL?clearance?is 1.05L/h,V_d?apparent volume of distribution?is 17.46 L.The pharmacokinetic parameters of arctigenin are:T_maxax is 6 h,C_maxax is 1.82?g/mL,(AUC_0-t?AUC_0-?)are 39.36 and 62.15?g·h/mL,MRT is 47.48 h,ke is 0.02 h^-1,t_1/2/2 is 31.61 h,CL is 12.87 L/h,V_d is 586.89L.Arctiin and arctigenin are widely distributed in various tissues of rats,and the drug concentration in the tissues is greater than that in blood.The concentration of arctiin in vivo:small intestine>lung>spleen>heart>kidney>liver>blood,the concentration of arctigenin in vivo:small intestine>spleen>lung>liver>heart>kidney>blood.Conclusion:Based on the preliminary quality analysis of Arctium tomentosum Mill.seeds,the extraction process of the total lignans from the Arctium tomentosum Mill.seeds was optimized,combined with the macroporous resin to enrich the effective parts of total lignans in Arctium tomentosum Mill.seeds,and the effective parts of the total lignans was obtained.After intragastric administration the effective parts of total lignans in rats,blood and tissue samples were measured by the HPLC for quantitation,and plasma concentration-time curves of arctiin and arctigenin in rat plasma were plotted.Arctiin and arctigenin have good pharmacokinetic characteristics,are widely distributed in tissues,and have certain targeting.Arctiin is mainly distributed in the lung,small intestine and other tissues,and arctigenin is mainly distributed in the spleen,small intestine and other tissues.