Abstract
5-Amino-1 -methyl quinolinium (5-AMQ) is a potent Nicotinamide N-methyl transferase (NNMT) inhibitor. NNMT is an enzyme that catalyzes the N-methylation of the endogenous substrate nicotinamide, as well as exogenous xenobiotics. NNMT is fundamental to cellular metabolism; NNMT is overexpressed in select tissues (e.g., adipose tissue, skeletal muscle, etc.) in pathophysiological conditions, making it a clinically relevant target for drug development in several chronic diseases including obesity and diabetes. The objective of this study was to develop and validate a simple, sensitive, and reproducible liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantification of 5-AMQin rat plasma and urine samples. 5-AMQwas extracted from plasma and urine by protein precipitation. Chromatographic separation was achieved using an ACE (R) Excel (TM) C-18 column (2 mu m, 50 x 2.1 mm) with a binary gradient solvent system comprising of water (A) and acetonitrile (B) containing 0.1 % formic acid as the mobile phase. Analysis was performed using an API 4000 QTRAP hybrid triple quadruple mass spectrometer and multiple reaction monitoring (MRM) in positive mode at m/z transitions of 159.100 -> 90.00 and 162.200 -> 117.200 for 5-AMQ and the internal standard, respectively. The standard curves of 5-AMQ in rat urine and plasma samples were linear in the concentration range of 10-2500 ng/mL. The intra-day and inter-day precisions and accuracies for 5-AMQ at four concentration levels in rat plasma and urine samples were found to be within the 15 % FDA acceptance range. Similarly, the accuracy and precision of 5-AMQquantification in samples diluted up to 20-fold using blank plasma were within the 15 % acceptable range. Furthermore, the extraction recoveries and matrix effects at three concentration levels of rat plasma samples ranged from 99.5 %-110.6 % and -6.1 %-14.1 %, respectively. 5-AMQ was stable in rat plasma samples subjected to standard storage, preparation, and handling conditions, with less than 15 % variation noted at two concentration levels. The validated, sensitive, and reproducible LC-MS/MS method for 5-AMQ in rat plasma and urine samples was effectively applied to a pharmacokinetic study in rats with IV and oral administration of 5-AMQ, 5-AMQ displayed substantial plasma exposures via IV and oral route, with a mean maximum plasma concentration of 2252 ng/mL after oral administration, mean area under the curve (AUC(0-infinity)) of 3708 h.ng/mL and 14431 h.ng/mL for the IV and oral groups, respectively, mean terminal elimination half-life of 3.80 +/- 1.10 h and 6.90 +/- 1.20 h respectively after intravenous and oral dose, and a good oral bioavailability (F % = 38.4). (C) 2021 Published by Elsevier B.V.