| Objectives:A isotope labeling technique was used in an nonclinical pharmacokinetics study of[14C]SHR1258 to investigate its pharmacokinetic characteristics and the phenomenon of low recovery in the non-radioactive pharmacokinetics experiment would be explained.What's more,the method involved in pharmacokinetics experiment and related detection of isotope labeling drug would be improved,and it provide a basis for application of isotope labeling technique in drug clinical pharmacokinetic study.Methods:(1)Absorption study:A LC-MS/MS detection method of SHR1258 in plasma was established.The prototype concentration of non-isotope labeling SHR1258,which was administered by single dose oral,in rat plasma was determined to establish the drug-plasma concentration curve and calculate the corresponding pharmacokinetics parameters.Meanwhile,radioactivity of[14C]SHR1258,which was also administered by single dose oral,was determined by LSC to establish the drug-plasma concentration curve and calculate corresponding pharmacokinetics parameters.Finally,the difference of pharmacokinetics between non-isotope labeling SHR1258 and its isotope labeling form was compared.(2)Excretion study:Radioactivity of[14C]SHR1258 was measured by LSC to evaluate its in vivo excretion.Feces and urine of rats who were taken single dose[14C]SHR1258 by oral were collected,and radioactivity of excreta in different period was measured to obtain the excretion of drugs and metabolites based on total radioactivity.The main excretion pathways of SHR1258 was identified and material balance information of SHR1258 was obtained.(3)Metabolism study:LC-MS/MS combined with isotope on line/off line detection was used in feces,bile,urine and plasma sample to confirm the metabolites profile of[14C]SHR-258 in each sample and quantify it.Meantime,the relative molecular mass and primary ion fragment was determined on line,and the structure of major metabolites was identified.Results:(1)Absorption study:Pharmacokinetics parameters of non-radioactive SHR1258(t1/2=3.80 h?V=208 L?CL=38 L?AUC=4478 ng·h/mL)has difference with radioactive SHR1258(t1/2=5.21 h?V=197 L?CL=26 L?AUC=11990 ng-h/mL):Half-life and exposure of radioactive SHR1258 were increased.It is suggested that,except prototype SHR1258,there were other metabolites that eliminated slowly.(2)Excretion study:The radioactive excretion rate of[14C]SHR1258 in 0-48h was higher than 90%(approximately 92%)in feces,and was less than 1%in urine.At the third day(48-72h)and the fourth day(72-96h)after administration,the total radioactivity of feces and urine was less than 1%of the radioactive dose,suggesting that SHR1258 was mainly excreted by feces within 48h.(3)Metabolism study:A total of 22 different metabolites,except prototype MO was identified.15 metabolites in rat feces sample,15 metabolites in bile sample,6 metabolites in plasma sample were detected.6 metabolites were detected in urine sample.Quantifying and structure identification of SHR1258 metabolites were achieved by combinedly using isotope labeling technique and LC-MS/MS.Conclusion:In this study,the understanding of SHR1258 oral absorption by using radioactive isotope labeling technique was improved.Excretion information and characteristic of SHR1258 and its related metabolites was obtained.Fast and efficiently quantifying and structure identifying method was established. |
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