Validation of an HPLC-UV Method for the determination of metformin hydrochloride in spiked-Human plasma for the application of therapeutic drug monitoring

Widely used as the mainstay therapy of type-2 diabetes patients, the plasma level of metformin should remain within therapeutic range (0.75–5.00 μg/mL) to gain optimal effectiveness without lactic acidosis. An HPLC-UV method has been developed and validated for the determination of metformin in spiked-human plasma according to the FDA Guidance of Bioanalysis. Optimum determination was achieved by using a mixture of 55% buffer pH 5.2 (6 mM potassium dihydrogen phosphate-6 mM sodium dodecyl sulphate) and 45% acetonitrile with Sunfire® C-18 column, 150x4.6mmx5μm from Waters, and SM7 injector with UV detector at 233 nm of wavelength. The analyte was extracted from plasma using 10% trichloroacetic acid. The assay was linear over the therapeutic steady-state concentration range of 0.50–7.00 μg/mL for metformin HCl with correlation coefficient 0.9999. The limit of detection was 0.14 μg/mL. The obtained results from accuracy, precision, and selectivity tests complied well with the generally accepted criteria for bioanalytical assay. The validated method could be used for determining metformin in human plasma and monitoring the metformin therapy in a clinical setting for dose adjustment, particularly for the elderly and patients with renal impairment as well as for measuring metformin accumulation.