Optimization of a HPLC-UV bioanalytical method using Box-Behnken design to determine the oral pharmacokinetics of neratinib maleate in Wistar rats.

The current research work reports the development and validation of a sensitive, robust and reproducible bioanalytical method for quantifying neratinib maleate in rat plasma. More than 85% of the drug was extracted from the plasma samples by protein precipitation. The method was optimized using Box-Behnken design, a response surface method. The effect of three critical factors, viz., the pH of the buffer (X1 ), the aqueous phase proportion in the mobile phase (X2 ) and the mobile phase flow rate (X3 ), was studied on two response variables, retention time (Y1 ) and United States Pharmacopoeia (USP) width (Y2 ). With the highest overall desirability function value of 0.943, the obtained optimized method conditions were: X1  = 2.4 ± 0.1; X2  = 66.7 ml, and X3  = 0.85 ml/min. Under the optimized conditions, the values of Y1 and Y2 for a sample containing 1 ppm of the drug were found to be 14.1 min and 0.50 ± 0.003, respectively. Single-dose intravenous bolus (7.5 mg/kg) and oral (15 mg/kg) pharmacokinetic studies were performed to determine the absolute bioavailability of the drug. The optimized bioanalytical method was sensitive enough to capture 95% of the drug eliminated from the body. The absolute oral bioavailability of the drug was 49.30%.