Extensive intestinal first-pass metabolism of arctigenin: Evidenced by simultaneous monitoring of both parent drug and its major metabolites

Elucidation of Arctigenin Pharmacokinetics After Intravenous and Oral Administrations in Rats: Integration of In Vitro and In Vivo Findings via Semi-mechanistic Pharmacokinetic Modeling

“…Besides the parent compound, metabolites formed after absorption could also occur in the GI tract via either efflux from enterocyte or bile secretion. As demonstrated in our previous in situ single-pass rat intestinal perfusion study, over 90% of the AA and AG formed during AR intestinal absorption was found in the perfusate (22), which were in turn susceptible to further biotransformation in the intestinal lumen. To comprehend the potential biotransformation of AR in the GI tract, in vitro incubations of AR, AA, and collected bile (after IV administration of AR) with both simulated GI fluid and rat GI content solution were conducted based on their potential site of appearance in vivo, followed by qualitative and semi-quantitative determinations of relevant metabolites.…”

“…All the above reactions were carried out at 37°C and terminated by the addition of an equal volume of ice-cold methanol containing 20% ascorbic acid and 80 μg/ml diclofenac acid (IS) followed by content analyses of AR and its metabolites using our previously developed ultraperformance liquid chromatography tandem mass spectrometry (UPLC/MS/MS) method (22). The calibration samples were prepared in microsomal matrices similar to those of the reaction samples except for the absence of corresponding enzyme reaction cofactors.…”

“…Briefly, the collected bile samples were diluted with an equal volume of 35% methanol in 25 mM NaH 2 PO 4 buffer (pH 2.5) containing 1% ascorbic acid. After 5-min centrifugation at 16,000×g, 20 μl of supernatant was injected into UPLC/MS/MS for content analyses of AR and its metabolites by our previously developed assay (22). For qualitative detection of AR and its metabolites in bile, multiple reaction mode (MRM) transitions corresponding with DA (m/z 357→83) and potential secondary metabolites were also monitored in addition to the parent compound and the two major metabolites identified in plasma (AR 371→83, AA 389→330, and AG 547→371).…”

“…Analyses of all the obtained plasma samples followed a previously validated UPLC/MS/MS assay method, which demonstrated satisfactory sensitivity, selectivity, accuracy, and precision of quantitative measurement of AR, AA, and AG in rat plasma, as well as recovery, matrix effect, and sample stability (22,30). Briefly, an aliquot of 5 μl IS was added to a 100-μl plasma sample, and 500 μl acetonitrile (ACN) was added into the mixture followed by vortex mixing for 1 min.…”

“…1), were the main compounds found in plasma after oral administration of AR in rats (22,23). Although our previous rat single-pass intestinal perfusion study demonstrated that extensive hydrolysis and glucuronidation of AR occurred when it passed through the intestinal epithelium (22), all its available in vivo pharmacokinetic studies by far monitored only the parent compound (24)(25)(26). Therefore, pharmacokinetic studies providing concrete plasma concentration profiles of both AR and its metabolites are essential for the understanding of its in vivo biotransformation.…”