Sample collection, analytical methods, and pharmacokinetic analysis
Blood samples were collected for PK analysis of carotegrast methyl, carotegrast, and other metabolites including M-I, M-II, and carotegrast glucuronide (carotegrast-gluc) at 0, 0.5, 1, 2, 4, 6, 8, 10, 12, 15, and 24 h post-dose on days 1 and 7. A blood sample was collected on day 14 when subjects visited for follow-up observation. Plasma concentrations of carotegrast methyl, carotegrast, M-I, M-II, and carotegrast-gluc were measured by validated methods using liquid chromatography-tandem mass spectrometry at Toray Research Center, Inc. (Tokyo, Japan). The linear analytical ranges of carotegrast methyl and its metabolites including carotegrast, M-I, and M-II were 0.5 − 500 ng mL-1, and that of carotegrast-gluc was 2.0 − 200 ng mL-1. Plasma concentrations of rifampicin were not measured in this study.
The primary PK parameters analyzed for carotegrast methyl and its metabolites included area under the time concentration curve (AUC) from time of dosing to time of last measurable concentration (AUC0-t), AUC from time of dosing to infinity (AUC0‐inf), maximum drug concentration (Cmax), time to Cmax(Tmax), t1/2, and mean residence time from time of dosing to time of last measurable concentration (MRT0-t).