Simultaneous determination of imigliptin and its three metabolites in human plasma and urine by liquid chromatography coupled to tandem mass spectrometry

• Imigliptin is a novel anti-diabetic drug.
• Imigliptin and its three metabolites concentration were firstly determined in plasma and urine by LC–MS/MS method.
• The method was proved to be a robust, accurate and specified method.

A specific and sensitive method was firstly developed using high performance liquid chromatography coupled with tandem mass spectrometry (HPLC–MS/MS) to simultaneously quantify imigliptin (KBP-3853) and its three metabolites (KBP-3926, KBP-3902, KBP-5493) in human plasma and urine. Solid-phase extraction (SPE) and direct dilution were used to extract imigliptin and its three metabolites from plasma and urine, respectively. The extracts were injected onto a SymmetryShield RP8 column with a gradient elution of acetonitrile and water containing 5 mM ammonium acetate (pH 7). Ionization of KBP-3853, KBP-3926, KBP-3902, KBP-5493, and XZP-3244 (internal standard, IS) was performed using an electrospray ionization (ESI) source in positive mode and detection was carried out with multiple reaction monitoring (MRM) mode. The lower limits of quantitation (LLOQ) of KBP-3853/KBP-3926/KBP-3902/KBP-5493 in human plasma and urine were 0.500/0.500/0.500/0.500 ng/mL and 20.0/20.0/10.0/10.0 ng/mL, respectively. Inter- and intra-batch precision of imigliptin and its three metabolites were less than 15% and the accuracy was within 85–115% for both plasma and urine. The extraction recoveries of all analytes at three concentration levels were consistent. The specificity, matrix effect, linearity and stabilities under various conditions were validated for imigliptin and its three metabolites in human plasma and urine. In conclusion, the validation results showed that this method was robust, specific, and sensitive and it can successfully fulfill the requirement of clinical pharmacokinetic study of imigliptin hydrochloride in Chinese healthy subjects.