LC–MS/MS Method Assay for Simultaneous Determination of the Apixaban and Metformin in Rat Plasma: Assessment of Pharmacokinetic Drug–Drug Interaction Study

Abstract

A simple, sensitive and accurate LC–MS/MS method was developed and validated for the simultaneous quantification of apixaban (APB) and metformin (MET) in rat plasma using rivaroxaban as internal standard (IS). An Inertsil ODS3 C18 column (150 × 4.6 mm, 5 μm) was used for chromatographic separation with isocratic elution. Multiple reaction monitoring (MRM) using positive-ion ESI mode to monitor ion transitions of m/z 459.8 → 442.8 for APB, m/z 130.2 → 71.2 for MET, m/z 436.8 → 144.9 for IS. The procedure of method validation included selectivity, linearity, precision, accuracy, matrix effect, extraction recovery and stability were conducted according to the guidelines of EMA and FDA. The method was validated over the concentration range of 0.5–250 ng/mL for APB and 8–8000 ng/mL for MET. The intra- and inter-day precision and accuracy of the quality control samples exhibited relative standard deviations (RSD) < 12.5% and the accuracy values ranged from −8.6 to 12.4%. Recovery and matrix effect values variations were all less than 15%. After oral administration APB and MET to rats, the comparison of pharmacokinetic parameters of APB in the single and co-administrated groups showed significant difference in AUC(0-t) from 730.71 ± 121.31 to 573.07 ± 90.13 ng/mL·h, t1/2 from 5.86 ± 3.21 to 4.24 ± 1.15 h and Cmax from113.54 ± 24.04 to 159.42 ± 54.6 ng/mL. The comparison of pharmacokinetic parameters of MET in the single and co-administrated groups showed significant difference in t1/2 from 2.83 ± 1.81 to 3.97 ± 0.57 h and Cmax from 4015.76 ± 873.23 to 3153.6 ± 1012.51 ng/mL. The results indicated that drug–drug interactions (DDI) occurred might be owing to APB affect one or all of OCTs, MATE1, MATE2-K.