BIOANALYTICAL METHOD DEVELOPMENT AND VALIDATION OF PIRFENIDONE BY RPHPLC METHOD AND ITS APPLICATION TO THE DETERMINATION OF DRUG FOOD INTERACTION STUDY IN WISTER RATS

A simple precise, accurate RP-HPLC method has been developed and validated for bioavailability study of pirfenidone in wister rat plasma. The separation and quantization of pirfenidone was achieved on a C18 reversed phase column using the mobile phase in gradient mode constituting of eluant A HPLC grade water (adjusted to pH 3.5) and eluant B 20% acetonitrile and 15% of methanol in the ratio of 60: 40 at a flow rate 1 mlmin-1. Eluted components were detected at 324 nm. The method showed good linearity for Pirfenidone in the range of 50–250ng mL-1, Y=54.97x - 349.5and correlation coefficient R2 is 0.998 respectively. The limit of quantitation (LOQ) and limit of detection (LOD) were found to be 12 and 20 ng mL-1respectively.Thedeveloped method shows good accuracy and precision. . Accuracy ranges from 98.49% to 99.37% with the precision 6.43% to 7.67% in inter-day method. Intra-day method the accuracy ranges from 98.64% to 99.33%with the precision 5.64% to 6.93 %. For bioanalytical study, parameters like Cmax, Tmax, AUC0-t,AUC 0-∞,Keli and T1/2 are compared by statistical analysis. The maximum concentration (Cmax) obtained for pirfenidone before and after food was found to be 1020.76 ng mL-1 and 836.5ng mL-1 respectively. The half life (T1/2) of pirfenidonebefore and after food were calculated and found to be 2.732158 h and 4.009485 h respectively. Area under the curve t AUC.0 of pirfenidone before food was calculated as 3060.95nghr mL-1 and 0 AUC was found to be 3053.07ng mL-1. Area under the curve t AUC.0 of pirfenidone after food was calculated as 2534.16nghr mL-1 and 0 AUC was found to be 2510.64ng mL-1. Elimination rate constant for Pirfenidone was found to be 0.004228 h-1and 0.002881 h-1 respectively. This method was successfully applied to the bioavailability study of pirfenidone.