Rapid RP-HPLC Method for Simultaneous Estimation of Metformin, Pioglitazone, and Glimepiride in Human Plasma

An isocratic reversed-phase high-performance liquid chromatography (RP-HPLC) method has been developed for rapid and simultaneous separation and estimation of 3 antidiabetic drugs, namely, metfonnin, pioglitazone, and glimepiride, in human plasma within 3 min. Separation was carried out on a MAGELLEN 5U C-18 (5 mu m, 150 mm x 4.60 mm) using a mobile phase of MeOH-0.025 M KH2PO4 adjusted to pH 3.20 using ortho-phosphoric acid (85:15, v/v) at ambient temperature. The flow rate was 1 mL/min, and the maximum absorption was measured at 235 nm. The retention time of metformin, pioglitazone, and glimepiride was noted to be 1.24, 2.32, and 2.77 min, respectively, indicating a very short analysis time compared to that of other reported methods. Also, limits of detection were reported to be 0.05, 0.26, and 0.10 mu g/mL for metformin, pioglitazone, and glimepiride, respectively, showing a high degree of method sensitivity. The method was then validated according to the FDA guidelines for the determination of the three drugs clinically in human plasma, in particular, regarding pharmacokinetic and bio-equivalence simulation studies.