Oxidative hair dye products have always been one of the key concerns in cosmetic supervision. The various aromatic amine and phenol dyes used in such products have different degrees of sensitization and toxicities. Therefore, it is necessary to establish a rapid and accurate method for the determination of various dyes. A high performance liquid chromatography (HPLC) method was developed for the determination of 40 dyes in oxidative hair dye products, and the method was optimized in terms of mobile phase type, column temperature, detection wavelength, and extraction solvent concentration. The sample pretreatment method was as follows: weighed 0.5 g sample accurately, added to 10 mL of sodium bisulfite solution containing 70% ethanol, extracted by ultrasonication for 15 min, added to 25 mL of the sodium bisulfite solution. Chromatographic separation was carried out on a Waters Atlantis® T3 MV Kit column (250 mm×4.6 mm, 5 μm) by gradient elution. To protect the column, the sample solution was filtered using a syringe filter with a 0.45-μm membrane before injection. The mobile phase was 0.02 mol/L ammonium acetate aqueous solution (containing 4% acetonitrile) and acetonitrile. The column temperature was varied between 30 ℃ and 35 ℃ during the separation. The dyes were detected using a photodiode array detector. The detection wavelength of 31 dyes was 235 nm, and that of the other nine dyes was 280 nm. The external standard method was used for quantitative analysis. The results showed that the 40 dyes could be separated well. All the dyes had good linearities in their own concentration ranges, with correlation coefficients (r) exceeding 0.999. The limits of detection (LODs) of the 40 dyes ranged from 5 to 168 μg/g, and the limits of quantification (LOQs) ranged from 16 to 504 μg/g. The average spiked recoveries at three levels (2.5, 5.0, and 10 mg/g) of the 40 dyes ranged from 81.4% to 109.6%, with relative standard deviations (RSDs) less than 5%. The stability of each dye in the standard solutions was good within 24 h, with RSDs ranging from 0.2% to 2.2%. To demonstrate the applicability of the method, 12 batches of commercial oxidative hair dye products from different manufacturers were analyzed. In total, 24 kinds of dyes were detected in the 12 batches of samples, and the contents of these dyes ranged from 0.01% to 2.83%. The method greatly increased the types of dyes detected by a single chromatography step, especially the permitted dyes (36 types). Compared with the standard method of Safety and Technical Standards for Cosmetics (2015 edition), this method replenished 13 permitted dyes: 1-hydroxyethyl 4,5-diaminopyrazole sulfate, hydroxyethyl-p-phenylenediaminesulfate, tetraaminopyrimidine sulfate, 2,6-dihydroxyethylaminotoluene, 2-amino-6-chloro-4-nitrophenol, 2-methyl-5-hydroxyethylaminophenol, 3-nitro-p-hydroxyethylaminophenol, 4-hydroxypropylamino-3-nitrophenol, 5-amino-4-chloro-o-cresol, 5-amino-6-chloro-o-cresol, HC yellow No. 2, hydroxybenzomorpholine, and hydroxyethyl-2-nitro-p-toluidine. The sensitivity and accuracy of the results could ensure and the detection efficiency could be improved through validation of the method. The developed method is suitable for the determination of various dyes in oxidative hair dye products and can provide an effective technical means for the supervision of such products.
氧化型染发产品中的多种染发剂具有不同程度的致敏性及其他毒性,建立快速、准确检测多种染发剂的方法,为该类产品监管提供有效的技术手段,十分必要。该研究建立了氧化型染发类产品中40种染发剂的高效液相色谱测定方法。染发产品经含70%乙醇的亚硫酸氢钠水溶液涡旋、超声提取,并经亚硫酸氢钠水溶液稀释后,以0.02 mol/L乙酸铵水溶液(含4%乙腈)和乙腈为流动相,采用Waters Atlantis® T3 MV Kit色谱柱(250 mm×4.6 mm, 5 μm)分离,配合柱温变化进行梯度洗脱,二极管阵列检测器检测,检测波长为235 nm和280 nm,外标法定量。结果表明,40种染发剂在各自范围内线性关系良好,相关系数均大于0.999; 40种染发剂的检出限为5~168 μg/g,定量限为16~504 μg/g;各染发剂在3个添加水平下的平均回收率为81.4%~109.6%, RSD均小于5%;各染发剂标准溶液在24 h内稳定性良好,RSD为0.2%~2.2%。与现行标准检验方法相比,该方法较大程度地增加了单一液相色谱条件下可测定的染发剂,特别是准用染发剂种类(36种),提高了检测效率,并可保证检测结果的灵敏度与准确性,适用于氧化型染发产品中多种染发剂的检测分析。
Keywords: dyes; high performance liquid chromatography (HPLC); oxidative hair dye products.