A simultaneous liquid chromatographic analysis of niclosamide and bicalutamide in rat plasma by protein precipitation...

The aim of the study was to develop a simple, sensitive and cost-efficient bioanalytical method for the simultaneous determination of niclosamide and bicalutamide in rat plasma with appropriate validation.

Real-time estimation was carried out using Ultra Flow Liquid Chromatography. The solvent system consisting of 20mM sodium acetate buffer at pH 4.0 and acetonitrile (65: 35% v/v) is used as the mobile phase. The analytes were extracted by protein precipitation with acetonitrile and separated on an Eclipse plus C18 Column (25cm×5cm×4.6μm) with L1 packing in isocratic mode with a flow rate of 1ml/min at 254nm. The developed method was validated on the basis of the bioanalytical guidelines of the US American FDA.

The retention times of niclosamide and bicalutamide were 4.19min and 8.61min, respectively, with a running time of 15minutes. The calibration ranges are 50-600ng/ml for niclosamide and 100-1200ng/ml for bicalutamide. Regression equations for niclosamide and bicalutamide were y=55746x - 1E+06 and y=22051x-576888 with regression coefficient (R2) 0.9952 & 0.9982 using the unweighted and weighted linear regression with a weighting factor of 1/xo, 1/x, 1/√x, and 1/x2. The accuracy of niclosamide and bicalutamide were 46.01ng/ml to 584.10ng/ml and 82.30ng/ml to 1149.13ng/ml, respectively. The percentage recoveries for niclosamide and bicalutamide were 86.51-90.29% & 88.64-93.99%, respectively.

The result of the present study show that the method developed is a simple, fast and precise method and is used in bioavailability and bioequivalence studies as well as during routine therapeutic drug monitoring of niclosamide and bicalutamide.