Spectrophotometric determination of the concentrations of pyranine dye components in a ternary mixture for dye batch certification

A strategy is proposed to determine the concentrations of components of pyranine, a cosmetic dye, to batch certification by the U.S. Food and Drug Administration. The main component of pyranine is mainly accompanied by two impurities during its synthesis. Harmful impurities were present at significantly lower concentrations than the main component. Despite the high degree of spectral overlap between the two impurities, specific spectral characteristics were exploited to overcome this challenge and determine their concentrations. The proposed method takes advantage of two key spectral peculiarities of the pyranine system: (1) the main component exhibits significant absorption at 404 nm where the impurities have negligible absorbance, and (2) despite being minor components, show intensely strong UV-vis spectra at concentrations comparable to those obtained in pyranine synthesis, facilitating their detection even at low concentrations. These characteristics enabled the development of a two-stage spectrophotometric approach. First, the concentration of the main dye was determined independently of the presence of the other two dyes in the ternary mixture using a direct absorbance method. Subsequently, the concentrations of the two impurities were determined by bivariate spectral calibration at two selected wavelengths. The results obtained by the proposed method were comparable to those obtained using other methods, such as HPLC and TLC. The proposed method is simple and accurate and can be used in quality control for the certification of dye batches. However, it critically depends on the unique spectral characteristics of the pyranine system, in particular the negligible absorption of the impurities at 404 nm and their relatively high absorption intensity comparable to main dye.