High-Performance Thin-Layer Chromatographic Assay of Metformin in Urine Using Ion-Pair Solid-Phase Extraction: Application for Bioavailability and Bioequivalence Study of New Microbeads Controlled Release Formulation

A sensitive high-performance thin-layer chromatographic (HPTLC) method was developed for the estimation of metformin in human urine. The drug was extracted from urine using ion-pair solid-phase extraction (IPSPE). Separation was carried out on Merck HPTLC aluminum sheets of silica gel 60 F-254 using a mixture of chloroform-methanol-ammonia 27% w/v (5:5:0.2, v/v) as a mobile phase. This system was found to give dense and compact spots for metformin (retardation factor, R-F, 0.25) following densitometric measurements of the spots at 237 nm. The proposed method was applied to generate urinary excretion data for metformin after administration of two metformin formulations (500 mg extended release, XR, Formulation R, and microbeads controlled release, Formulation T) to six healthy human volunteers in a two-treatment, crossover design. Various pharmacokinetic parameters like peak excretion rate ((dAU/dt)(max)), cumulative amount (Ae(0-24)) of metformin excreted, elimination half-life (t(1/2)), and terminal elimination rate constant (k(el)) were calculated for both the formulations. Statistical comparison of various pharmacokinetic parameters of Formulation T with that of Formulation R revealed that Formulation T is bioequivalent with Formulation R.