Design of experiments based reversed-phase high-performance liquid chromatography method for estimation of metformin and glimepiride in combined dosage forms

Background: The present study aimed to systematically develop and validate a robust, rapid, and stability-indicating reversed-phase high-performance liquid chromatography (RP-HPLC) method for the simultaneous estimation of metformin and glimepiride in fixeddose combination tablets. Method development was guided by a design of experiments (DoE) approach using the Box-Behnken design (BBD) to ensure optimal analytical performance. Materials and Methods: Chromatographic separation was achieved on a Cosmosil C18 column using an optimized mobile phase of acetonitrile and water (80:20, v/v) adjusted to pH 3.5. The flow rate was 1 mL/min, and detection was performed at 221 nm. The BBD approach systematically evaluated the effects of wavelength, flow rate, and pH on critical chromatographic parameters, enabling the selection of optimal conditions. Method validation followed ICH Q2 (R2) guidelines for linearity, specificity, precision, accuracy, robustness, limit of detection (LOD), and limit of quantification (LOQ). Results: The optimized method showed excellent linearity (R2 > 0.999) for both drugs, specificity without interference from excipients, and precision with % relative standard deviation Conclusion: The developed RP-HPLC method is simple, rapid, costeffective, and highly reliable for routine quality control of fixed-dose metformin–glimepiride tablets. The DoE approach enhanced method optimization, ensuring robustness and regulatory compliance.