Background The U.S. Food and Drug Administration (FDA) batch certifies FD&C Yellow No. 6 (Y6) to ensure that the color additive meets requirements published in the Code of Federal Regulations (CFR), including specifications for seven organic manufacturing impurities consisting of two intermediates, a reaction by-product, an impurity originating from an intermediate, and three subsidiary colors. Objective An ultra-high-performance liquid chromatography (UHPLC) method was developed and validated for determining seven organic impurities in Y6. For comparison, the currently used high-performance liquid chromatography (HPLC) method was also validated. Methods The new UHPLC method uses a 1.7 µm particle size biphenyl column with aqueous ammonium formate and methanol as eluants. Analytes are identified by comparing their retention times and UV-visible spectra to those of reference standards. Calibration is performed in the presence of the dye matrix and analyte levels are determined from their peak areas. Results UHPLC and HPLC validation studies obtained linear calibration curves and excellent values for limits of detection, limits of quantitation, recovery, repeatability, and intermediate precision for all analytes. Survey analyses of 30 samples using the UHPLC and HPLC methods yielded results that were consistent within experimental error; however, the UHPLC method provided more accurate results for two analytes that coelute using the HPLC method. Conclusion The UHPLC method satisfies the accuracy and precision requisites for routine certification of Y6. Highlights The UHPLC method is faster and generates less waste than the HPLC method while delivering better sensitivity, separation, and accuracy.

中文翻译：

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超高效液相色谱法测定 FD&C Yellow No. 6 中的 7 种有机杂质

背景 美国食品和药物管理局 (FDA) 批次认证 FD&C Yellow No. 6 (Y6) 以确保颜色添加剂符合联邦法规 (CFR) 中公布的要求，包括由两种中间体组成的七种有机制造杂质的规格、反应副产物、源自中间体的杂质和三种辅助颜色。目的 开发并验证了一种超高效液相色谱（UHPLC）方法，用于测定 Y6 中的 7 种有机杂质。为了比较，目前使用的高效液相色谱（HPLC）方法也得到了验证。方法 新的 UHPLC 方法使用 1.7 µm 粒径的联苯色谱柱，使用甲酸铵水溶液和甲醇作为洗脱剂。通过将分析物的保留时间和紫外可见光谱与参考标准进行比较来识别分析物。在染料基质存在的情况下进行校准，分析物水平由它们的峰面积确定。结果 UHPLC 和 HPLC 验证研究获得了所有分析物的线性校准曲线和出色的检测限、定量限、回收率、重复性和中间精密度值。使用 UHPLC 和 HPLC 方法对 30 个样品进行调查分析得出的结果在实验误差范围内是一致的；然而，UHPLC 方法为使用 HPLC 方法共洗脱的两种分析物提供了更准确的结果。结论 UHPLC 方法满足 Y6 常规认证的准确度和精密度要求。