The Clinical Pharmacokinetic Study Of 1, 5-Dicaffeoylquinic Acid

1,5-Dicaffeoylquinic Acid is an active compound isolated from Inula japonica Thunb., which possess a broad spectrum of pharmacological properties, including antioxidant, antibacterial, antihistaminic, hepatoprotectant, and other biological effects. 1,5-Dicaffeoylquinic Acid has drawn more and more attention for their anti-HIV (human immunodeficiency virus) activity. This thesis was designed to finish the clinical pharmacokinetic study and metabolite study, which provide data for the future clinical study and endure the safe and efficacy of clinical medication.Established and validated the LC/MS/MS method for simultaneous determination of 1,5-dicaffeoylquinic acid and its active metabolitesIn this study, a rapid, sensitive and reproducible liquid chromatography-tandem mass spectrometry method for the simultaneous determination of 1,5-Dicaffeoylquinic acid (1,5-DCQA) and its active metabolites, 1-Caffeoyl-5-feruoylquinic acid (1,5-CFQA) and 1,5-O-Diferuoylquinic acid (1,5-DFQA) in human plasma, using Puerarin as internal standard (IS), was developed and validated. Analytes were extracted from plasma samples by liquid-liquid extraction with ethyl acetate, separated on a C18 reversed phase column with 5mM ammonium acetate and acetonitrile as the mobile phase and detected by electrospray ionization mass spectrometry in negative selected reaction monitoring (SRM) mode. The accuracy and precision of the method was acceptable and linearity was good over the range 1-200 ng·mL-1 for clinical pharmacokinetic study in phaseⅠfor each analyte and 0.5-200 ng·mL-1 for clinical pharmacokinetic study in phaseⅡfor 1,5-DCQA. In addition, the specificity, extraction recovery, stability and matrix effect was also satisfactory.The pharmacokinetic study in phaseⅠof 1,5-Dicaffeoylquinic AcidFollowing a single oral administration with different dose of 1,5-DCQA to healthy human (200 mg,400 mg,800 mg), the blood samples were collected at different time. The concentration of 1,5-DCQA acid and its active metabolites in plasma were determined by HPLC-MS/MS method described as above. Pharmacokinetic parameters were calculated by the DAS 2.0 software according the statistical moment theory. It was shown that the pharmacokinetic parameters were fit to linear kinetics characteristics in the range of administration dosage. In the repeated oral administration study there was no significant difference between the the first dose and last dose in AUC (0-t)、Cmax and t1/2 (p>0.05) of 1,5-DCQA, suggesting that 1,5-DCQA was not easily accumulative in vivo and it did not induce or inhibit the metabolic enzyme. The AUC (0-t)and t1/2 of 1,5-DFQA was significant difference between the first dose and last dose, suggesting that 1,5-DFQA was easily accumulative in vivo.The pharmacokinetic study in phaseⅡof 1,5-Dicaffeoylquinic AcidFollowing a repeated oral administration with different dose of 1,5-DCQA to patients (200 mg,300 mg), there was no significant difference between the first dose and last dose in AUC (0-t)、Cmax and t1/2 (p>0.05) of 1,5-DCQA, suggesting that 1,5-DCQA was not easily accumulative and it did not induce or inhibit the metabolic enzyme in patients. The AUC (0-t) of 1,5-DFQA was significant difference between the first dose and last dose, suggesting that 1,5-DFQA was easily accumulative in vivo.The metabolites of 1,5-Dicaffeoylquinic Acid in human with LC-MSn assayTo investigate metabolites of 1,5-DCQA in human urine after oral administration of 1,5-DCQA by using liquid chromatography and tandem electrospray ionization ion trap mass spectrometry (LC-ESI/MSn). A total of 28 metabolites were discovered, including methylated, glucuronidated and methyl-glucuronidated metabolites. Two important metabolites were determined by comparing with synthesized standards. These results suggest that methylation, glucuronidation and isomerization were three important metabolic pathways of 1,5-DCQA in human.