A study had been conducted on the optimization and validation of high performance liquid chromatography (HPLC) method for simultaneous determinationquantitative analysis of whitening agents namely hydroquinone (HQ) and retinoic acid (RA) in a skin bleaching cream cosmetics. The optimization of analytical method uUsing experimental design of response surface method, resulted the optimal condition of mobile phase resulted was of methanol: water: glacial acetic acid (88: 12: 0.4 v/v/v), with flow rate of 1.2 mL/minute. The separation of HQ and RA was achieved column used wasusing column of STAR LP Purospher® stationary phase RP-18 end-capped (250 x 4.6 mm, 5 µm) with column-ovenat temperature of 45°C. Detection was performed using photodiode array (PDA), set at a wavelength of 310 nm. The methodHPLC-PDA was validated by assessing several parameters, namely selectivity, linearity, accuracy, precision, detection limit, quantitation limit, and robustness. The validation results showed that the analytical method was selective, linear with correlation coefficient (r) of ≥ 0.9998, precise with relative standard deviation (RSD) of inter-day precision of ≤ 3.44%, accurate with recovery percentage of HQ and RA of 100.00% and 99.67%, respectively. The detection limits of HQ and RA were of 0.44 and 0.47 mg/g, sample respectively; while the limits of quantitation of HQ and RA were of 5.09 and 1.71 mg/g samples, respectively. The developed method was successfully used for analysis of HQ and RA in commercial cream samples.
Key words: HPLC, hydroquinone, retinoic acid, cosmetics, validation
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