A novel ultra-sensitive LC-MS/MS method for determination of elobixibat in human plasma; Application to a bioequivalence study on healthy volunteers

Towards a new era for alleviating chronic idiopathic constipation, elobixibat has been recently approved in Japan and completed phase III in clinical trials as a highly potent inhibitor for ileal bile acid transporter. This work provides the field of biomedical analysis and clinical studies with the first bioanalytical method for elobixibat quantitation in real human plasma. A new ultra-sensitive liquid chromatography-tandem mass spectrometric method was developed using elobixibat-d5 as an internal standard. First of all, several preliminary efforts were exerted and directed towards optimizing sample preparation procedures to extract the desired drug at a very low concentration level and to avoid any matrix interference or masking. The trials settled on adopting liquid-liquid extraction using methyl tertiary butyl ether after adding 200 mu L of 10 % formic acid to 500 mu L plasma sample. Chromatographic separation was then conducted using Kinetex (R) EVO C18 column with mobile phase composed of acetonitrile and 20 mM ammonium format acidified with 0.1 % formic acid in ratio of 80:20 (v/v) and pumped at 0.6 mL/min. Positive electrospray ionization was adopted for mass acquisition, operated in multiple reactions monitoring (MRM) mode at m/z 696 -* 593.1 for elobixibat and m/z 701 -* 598.1 for the IS. Full bioanalytical validation as per FDA guidance was done over a range of 20.0-1500.0 pg/mL. The proposed method was successfully exploited for determination of elobixibat in human plasma samples and extended to estimate the pharmacokinetic parameters after administration of a single oral dose of 5 mg elobixibat tablet to thirty healthy volunteers.