Application of central composite design and structural analysis to the optimization of HPLC method for simultaneous analysis of hydrochlorothiazide, amlodipine besylate and valsartan

Central composite experimental design was used for fast, simple, and accurate high-performance liquid chromatography (HPLC) determination of hydrochlorothiazide, amlodipine and valsartan in combined dosage forms. This method avoids the disadvantages of the traditional analytical approach, which is time-consuming, involves a large number of runs, and does not allow establishing the multiple interacting parameters. On the basis of preliminary experiments and physicochemical characteristic of analyzed substances, three independent variables (methanol content, pH of the mobile phase, and column temperature) were selected as input, while as dependent variables, six responses (retention time of hydrochlorothiazide, retention time of amlodipine, retention time of valsartan, asymmetry of hydrochlorothiazide peak, asymmetry of amlodipine peak, and asymmetry of valsartan peak) were chosen. Face centered central composite design enables an estimation of factors which have the most importance. After optimizing experimental conditions, a separation was conducted on a Zorbax C-8 (150 mm x 4.6 mm, 5 mu m) column with a mobile phase consisting of methanol-acetonitrile-acetate buffer 40: 20: 40 (v/v/v), pH adjusted to 3.5 with acetic acid, flow rate of 1 mL/min and column temperature of 40 C-deg. The method was successfully applied to the simultaneous separations of these active drug compounds in their commercial dosage forms.