DEVELOPMENT AND VALIDATION OF A HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC METHOD FOR THE DETERMINATION OF METHOCARBAMOL IN HUMAN PLASMA

An isocratic HPLC method was developed and validated for the quantitation of methocarbamol in human plasma. Methocarbamol and internal standard in 200 mu l of human plasma were extracted with ethyl acetate, evaporated to dryness and reconstituted in water. Separation was achieved on a reversed-phase C-18 column with a mobile phase of methanol-0.1 M potassium phosphate monobasic-water (35:10:55, v/v/v). The detection was by ultraviolet at 272 nm. Linearity was established at 1-100 mu g/ml (r > 0.999). The limit of quantitation was designed as 1 mu g/ml to suit pharmacokinetic studies. Inter-day precision and accuracy of the calibration standards were 1.0 to 3.6% coefficients of variance (C.V.) and -2.0 to +1.6% relative error (R.E.). Quality controls of 3, 20 and 70 mu g/ml showed inter-day precision and accuracy of 2.5 to 3.6% C.V. and -0.9 to -0.4% R.E. Recovery of methocarbamol was 91.4-100.3% in five different lots of plasma. The method was shown to be applicable on different brands of C-18 columns.