Method Development and Validation for Quantitative Estimation of Calcium Acetate in Calcium Acetate Capsules by RP-HPLC using Indirect UV Method

Abstract:

Objective: A simple, accurate and economical method has been developed by Reverse Phase High Performance Liquid Chromatography (RP-HPLC) using indirect UV (Ultra Violet) method. The Principle behind the indirect UV method was applied; the copper sulfate pentahydrate with UV absorption in the mobile phase was replaced with calcium ions without UV absorption, resulting in a reduction in UV absorption of the mobile phase. Materials and Methods: Method development was carried out on strong cation exchanger column (Zorbax 300-SCX, 4.6 mm i.d. x 150 mm, particle size 5 μm). The mobile phase consists of 4 mM copper sulfate pentahydrate solution at a flow rate of 2.0 mL per min. Indirect UV detection was performed at 230.0 nm. Results: Retention time for calcium was found about 7.1 min. The developed method was validated in terms of specificity, linearity, accuracy, precision, range, limit of detection, limit of quantification and robustness. Calibration plot was linear over the concentration range of 19 to 1125 µg/mL. System suitability parameters were found 0.8 % RSD, tailing factor 1.1 and more than 8000 theoretical plates. Limit of detection and limit of quantification were found 6.3 µg/mL and 19.0 µg/mL respectively. Conclusion: The high recovery and low relative standard deviation confirm the suitability of the method for determination of calcium acetate in calcium acetate capsules.

Key words: Calcium Acetate, RP-HPLC, Indirect UV method, Chromophore, Validation.