OBJECTIVE To develop a HPLC assay for the
determination of metformin in human plasma.METHODS 400 μL
acetonitrile was added
to 200 μL plasma sample with ranitidine as the internal standard for precipitating protein.The supernatant was washed with dichoromethane and 20 μL aqueous was injected to HPLC.The separation was performed on a Zorbax Rx-Sil column(4.6 mm×250 mm,5 μm).The mobile phase consisted of
acetonitrile,0.05 mol·L~(-1)NaH_2PO_4 and 0.05 mol·L~(-1)Na_2HPO_4(75∶25∶5) at the flow rate of 1.0 mL·min~(-1).The detection wavelength was at 236 nm.RESULTS An excellent linearity(r=0.999 5,n=5) was obtained over the range of 0.025～4.0 mg·L~(-1),and the limit of detection was 0.01 mg·L~(-1).The RSDs for 0.05,1.0 and 3.0 mg·L~(-1) check samples were 2.46%～4.96% for intra-run validation(n=5),and 1.85%～3.97% for inter-run validation(n=10),respectively.CONCLUSION The improved method is sensitive,specific,reproducible and easy to process.It is practical and suitable for the pharmacokinetic study of metformin.