MICRO-SCALE METHOD FOR SEPARATION AND QUANTIFICATION OF ATENOLOL AND HYDROCHLOROTHIAZIDE BY SEQUENTIAL INJECTION CHROMATOGRAPHY

A microscale stopped-flow separation manifold, which is termed sequential injection chromatograph (SIC), was exploited for separating and quantifying atenolol and hydrochlorothiazide. A short C18 separation column (4.6 × 25 mm) with a monolithic structure was used to offer a rapid and sufficient separation. Microscale fiber optic spectrometric devices were coupled with the SIC manifold for UV detection at 220 nm. The proposed SIC method was optimized, validated, and applied to both formulations single and binary mixture. The mobile phase composition (MPC) was optimized using the 23 full-factorial design approach while other conditions were optimized using the univariate approach. The optimum MPC was 40 µmol/L phosphate:methanol:acetonitrile (85:7.5:7.5, v/v/v) at pH 3.0. Other optimum conditions were 40 µL/s for flow rate and 40 µL for sample volume. Satisfactory separation with a resolution of 1.6 and numbers of theoretical plates of more than 630 was achieved. The interesting features of the proposed SI...