Determination of rifampicin and its main metabolite in plasma and urine inpresence of pyrazinamide and isoniazid by HPLC method

A reversed phase HPLC method is described for the simultaneous estimation o f rifampicin and its major metabolite desacetyl rifampicin, in the presence of isoniazid and pyrazinamide, in human plasma and urine. The assay involv es simple liquid extraction of drug, metabolite and internal standard (rifa pentine) from biological specimens and their subsequent separation on a C-1 8 reversed phase column and single wavelength UV detection. In plasma as we ll as in urine samples, all the three compounds of interest eluted within 1 7 min. Using methanol-sodium phosphate buffer (pH 5.2; 0.01 M) (65.35, v/v) as mobile phase under isocratic conditions, it was established that isonia zid, pyrazinamide and ascorbic acid (added to prevent oxidative degradation of analytes) did not interfere with the analyte peaks. Recoveries (extract ion efficiency) for drug were greater than 90% in both plasma and urine, wh ereas for metabolite the values were found to be 79 and 86% In plasma and u rine, respectively. The plasma and urine methods were precise (total coeffi cient of variation ranged from 5 to 23%) and accurate (-7 to 5% of the nomi nal values) for both the analytes. Individual variance components, their es timates and their contribution to the total variance were also determined. Using the same method, unknown samples supplied by WHO were assayed and goo d correlations were obtained between the found and intended values. The met hod developed proved to be suitable for simultaneous estimation of rifampic in and desacetyl rifampicin in plasma and urine samples. (C) 1999 Elsevier Science B.V. All rights reserved.