Development and Validation of UV Spectrophotometric, HPLC and RP-HPLC Methods for Simultaneous Estimation of Amlodipline Besylate. Hydrochlorothiazide and Valsartan in their Combined Tablet Dosage Form

Abstract

This paper describes Validated UV Spectrophotometric, high-performance thin-layer chromatography (HPTLC) and Reverse phase high-performance liquid chromatography (RP-HPLC) methods for the Simultaneous estimation of Amlodipine Besylate (AMLO), Hydrochlorothiazide (HCTZ) and Valsartan (VALS) in bulk and combined Tablet Dosage form. The RP-HPLC separation was achieved on a Kromasil C18 column (250×4.6 mm, 5 μm) using mobile phase of 50 mM Potassium dihydrogen o-phosphate buffer phosphate (triethylamine 0.2%), (pH 3.7 adjusted with o-phosphoric acid)-acetonitrile, (56:44, v/v) as the mobile phase at a flow rate of 1.0 mL/min. Quantification was achieved with ultraviolet (UV) detection at 232 nm. The HPTLC separation was achieved on Precoated silica gel 60GF254 using ethyl acetate: methanol: Toluene: Ammonia 25% (7.5:3:2:0.8, v/v/v/v) as the mobile phase. Quantification was achieved with UV detection at 242 nm. UV Spectrophotometric method includes simultaneous equation method that involves measurement of absorbances at three wavelengths i.e. at 237.6 nm (λmax of AMLO), 270.2 nm (λmax of HCTZ), and 249.2 nm (λmax of VALS) in methanol. All the three methods were validated as per ICH guidelines. The comparison of all the three methods was done by applying ANOVA test. The proposed methods for estimation of AMLO, HCTZ and VALS were found to be simple, precise, and accurate are applicable for the simultaneous determination of AMLO, HCTZ and VALS in marketed tablet formulations. Keywords:-Amlodipine Besylate, Hydrochlorothiazide, Valsartan, UV Spectrophotometric, HPTLC, RP-HPLC