Quality-by-design optimized RP-HPLC approach for the therapeutic drug monitoring of glibenclamide and fluoxetine in human plasma

Abstract

Glibenclamide (GLB) and fluoxetine (FLX) were reported to have a potential drug-drug interaction when clinically co-administered. FLX is an inhibitor of CYP2C9 (the enzyme responsible for the metabolism of GLB), thus resulting in increased serum levels of GLB followed by severe hypoglycemia. Therefore, a sensitive, simple, and selective HPLC method is urgently needed for their simultaneous determination in plasma to be utilized for monitoring their therapeutic blood concentration levels. The current study introduces the first RP-HPLC approach for the simultaneous estimation of this binary mixture in less than 7 min. Adopting a quality-by-design (QbD) approach allowed for the designed approach to be optimized. Using a two-level full factorial design (25), the effects of various variables on the chromatographic responses were investigated and optimized. Hypersil BDS C18 column was used for the achievement of the optimum chromatographic separation at 19.9 °C using acetonitrile: sodium dihydrogen phosphate buffer (7.5 mM; pH 4, 60:40 % v/v) as a mobile phase, pumped isocratically at a flow rate of 1.036 mL/min. An injection volume of 30.0 μL was used for the binary mixture with UV detection at 230 nm. FLX and GLB calibration curves revealed an excellent linearity over the ranges of 0.025–10.00 and 0.02–10.00 µg/mL with limits of quantitation of 0.018 and 0.02 µg/mL, respectively. Due to the high sensitivity and selectivity, the method was successfully applied to the assay of the two drugs in spiked human plasma samples with reasonable % recoveries (92.70–108.50 %), indicating good bioanalytical applicability. Therefore, this study could be applied for the therapeutic drug monitoring of GLB and FLX in human plasma to control their potential drug-drug interaction. The suggested approach was also applied to estimate the investigated drugs in their dosage forms with reasonable % recoveries (96.80–102.80 %) and small % RSD values (less than 2 %). The method's excellent greenness was evaluated using three metric tools, including Analytical Eco-Scale, Green Analytical Procedure Index (GAPI), and Analytical Greenness (AGREE). Validation of the proposed method was performed according to ICH and US FDA guidelines.