目的:建立葛根芩连片中可能掺入的染色剂含量测定方法,并采用LC-MS/MS方法进行专属性确证。方法:采用HPLC法测定染色剂含量,使用Kromasil C18(4.6 mm×250 mm,5μm)色谱柱,以乙腈-0.05 mol·L-1乙酸铵溶液(含0.5%冰乙酸)为流动相,梯度洗脱,流速1.0 m L·min-1,检测波长为420 nm和500 nm,柱温35℃。用LC-MS/MS进行确证,以乙腈-0.01 mol·L-1乙酸铵溶液为流动相,梯度洗脱;离子源为电喷雾电离源(ESI),扫描模式为多反应监测(MRM)。结果:建立了HPLC法检测葛根芩连片中可能含有的10种染色剂含量的方法。方法学验证结果表明,柠檬黄、胭脂红、日落黄、亮黄、酸性红73、甲基橙、金橙Ⅱ、金胺O、赤藓红、碱性橙Ⅱ的线性范围分别为1.498 6~49.955 0、0.672 7~22.423 5、1.497 8~49.925 0、0.223 5~7.451 6、0.447 0~14.900 0、0.398 0~13.267 8、0.497 0~16.566 7、0.190 6~6.353 2、0.695 3~23.176 4和0.119 3~3.977 6μg;加样回收率(n=6)分别为97.2%(RSD=0.86%)、99.1%(RSD=0.48%)、98.6%(RSD=0.86%)、101.0%(RSD=1.2%)、98.2%(RSD=0.89%)、98.3%(RSD=1.0%)、97.5%(RSD=0.63%)、96.3%(RSD=0.40%)、97.8%(RSD=1.0%)和96.3%(RSD=0.96%)。在对11个厂家123批样品的检验中,有5个厂家33批样品检出金胺O,检出率为26.8%,含量在0.013~83.51·g·g-1之间,其中1批样品同时检出金胺O和碱性橙Ⅱ。用LC-MS/MS进行专属验证,33批出现HPLC阳性峰样品均检出相应的金胺O和碱性橙Ⅱ。结论:本方法准确、便捷,具有良好的精密度、重现性和稳定性,线性关系良好,可用于葛根芩连片的质量控制。 OBJECTIVE: To establish a method for the determination of the possible incorporation of the dye into the Gegenqinlian contiguous tablet and confirm the specificity by LC-MS / MS. Methods: The content of the dye was determined by HPLC. The chromatographic conditions were as follows: Kromasil C18 (4.6 mm × 250 mm, 5 μm) column, acetonitrile-0.05 mol·L-1 ammonium acetate solution The flow rate was 1.0 m L · min-1, the detection wavelength was 420 nm and 500 nm, and the column temperature was 35 ℃. The sample was confirmed by LC-MS / MS. The mobile phase consisted of acetonitrile-0.01 mol·L-1 ammonium acetate solution with gradient elution. The ion source was electrospray ionization source (ESI) and the scanning mode was multi-reaction monitoring (MRM). Results: The HPLC method was established for the determination of the content of 10 kinds of stains in Gegenqinlian. The validation results showed that the linear range of lemon yellow, carmine, sunset yellow, bright yellow, acidic red 73, methyl orange, golden orange Ⅱ, Auramine O, erythrosine and basic orange Ⅱ were 1.498 6 ~ 49.955 0,0.672 7 ~ 22.423 5,1.497 8 ~ 49.925 0,0.223 5 ~ 7.451 6,0.447 0 ~ 14.900 0,0.398 0 ~ 13.267 8,0.497 0 ~ 16.566 7,0.190 6 ~ 6.353 2,0.695 3 ~ 23.176 4 (RSD = 0.86%), 99.1% (RSD = 0.48%), 98.6% (RSD = 0.86%) and 101.0% (RSD = (RSD = 0.89%), 97.8% (RSD = 1.0%), 96.3% (RSD = 0.89%), 98.3% % (RSD = 0.96%). Among 123 batches of samples from 11 manufacturers, 5 batches of samples of goldmine O were detected in 33 batches of samples from 5 factories, the detection rate was 26.8% and the content was 0.013 ~ 83.51 g · g-1. At the same time detection Auramine O and alkaline orange Ⅱ. By LC-MS / MS for the exclusive validation, 33 batches appear HPLC positive peak samples were detected corresponding Auamine O and basic orange Ⅱ. Conclusion: The method is accurate and convenient, with good precision, reproducibility and stability, good linearity, and can be used for quality control of Gegenqinlian contiguous tablets.