Validation and application of reversed-phase high-performance liquid chromatography for quantitative analysis of acid orange 7 and Sudan II in blusher products

ABSTRACT

The objective of this study was to develop and to validate reversed-phase high-performance liquid chromatography(RP-HPLC) method for the determination of acid orange 7 (AO7) and Sudan II (SII) in blusher products. Separationof AO7 and SII was performed using C18 column (Thermo Synergy Gold 250 mm × 4.6 mm i.d., 5 µm) with mobilephase consisting of acetonitrile-water as a solvent (11 v/v) and delivered in gradient manner. The mobile phase wasdelivered at flow rate of 0.9 ml/minute, using column temperature of 40°C. RP-HPLC was validated by assessingseveral performance characteristics which included selectivity, linearity and range, sensitivity, precision, accuracy, androbustness. The developed RP-HPLC was selective for the separation of AO7 and SII. Over concentration of 10.326–41.304 µg/ml (AO7) and 9.967–39.869 (SII), the method was linear with coefficient of correlation (r) >0.999. Themethod was sensitive as indicated by low limit of detection and limit of quantification. The relative standard deviation(RSD) values during intra-assay and inter-assay were lower than those required by RSD Horwitz. The percentageof recovery was within acceptable ranges as required by Association of Official Analytical Chemists method. Thevalidated RP-HPLC method has been successfully applied for the analysis of AO7 and SII in commercial blusherproducts, and both AO7 and SII were not detected in the tested samples.