HPLC-DAD determination of Metformin in human plasma using derivatization with p-nitrobenzoyl chloride in a biphasic system

An optimized method for determination of Metformin at ppb level in human plasma is described. Derivatization of Metformin with p-nitrobenzoyl chloride was carried out in a biphasic system. The derivative extracted in the organic layer (dichloromethane) was concentrated and analyzed by HPLC-DAD, using an isocratic elution (water/methanol mixture 65/35), monitored at 280 nm. Good selectivity against co-extracted matrix components was observed. A detection limit of 10 ppb Metformin in human plasma was reached.