Nishizaki Yuzo, Sato-Masumoto Naoko, Yokota Aki, Mikawa Tsuyoshi, Nakashima Koichi, Yamazaki Taichi, Kuroe Miho, Numata Masahiko, Ihara Toshihide, Ito Yusai, Sugimoto Naoki, Sato Kyoko
a Division of Food Additives , National Institute of Health Sciences , Kawasaki , Japan.
b San-Ei Gen F.F.I., Inc ., Osaka , Japan.
Food Addit Contam Part A Chem Anal Control Expo Risk Assess. 2018 May;35(5):838-847. doi: 10.1080/19440049.2018.1440642. Epub 2018 Feb 22.
To accurately determine carminic acid (CA) and its derivative 4-aminocarminic acid (4-ACA), a novel, high-performance liquid chromatography with photodiode array detector (HPLC/PDA) method using relative molar sensitivity (RMS) was developed. The method requires no analytical standards of CA and 4-ACA; instead it uses the RMS values with respect to caffeine (CAF), which is used as an internal standard. An off-line combination of H-quantitative nuclear magnetic resonance spectroscopy (H-qNMR) and HPLC/PDA was able to precisely determine the RMSs of CA/CAF and 4-ACA/CAF. To confirm the performance of the HPLC/PDA method using RMSs, the CA and 4-ACA contents in test samples were tested using four different HPLC-PDA instruments and one HPLC-UV. The relative standard deviations of the results obtained from five chromatographs and two columns were less than 2.7% for CA/CAF and 1.1% for 4-ACA/CAF. The H-qNMR method was directly employed to analyse the CA and 4-ACA contents in test samples. The differences between the quantitative values obtained from both methods were less than 5% for CA and 3% for 4-ACA. These results demonstrate that the HPLC/PDA method using RMSs to CAF is a simple and reliable quantification method that does not require CA and 4-ACA certified reference materials.
为了准确测定胭脂红酸（CA）及其衍生物4-氨基胭脂红酸（4-ACA），开发了一种使用相对摩尔灵敏度（RMS）的新型高效液相色谱-光电二极管阵列检测器（HPLC/PDA）方法。该方法无需CA和4-ACA的分析标准品；而是使用相对于用作内标的咖啡因（CAF）的RMS值。1H-定量核磁共振波谱（1H-qNMR）与HPLC/PDA的离线组合能够精确测定CA/CAF和4-ACA/CAF的RMS。为了确认使用RMS的HPLC/PDA方法的性能，使用四台不同的HPLC-PDA仪器和一台HPLC-UV对测试样品中的CA和4-ACA含量进行了测试。从五台色谱仪和两根色谱柱获得的结果的相对标准偏差，CA/CAF小于2.7%，4-ACA/CAF小于1.1%。直接采用1H-qNMR方法分析测试样品中的CA和4-ACA含量。两种方法获得的定量值之间的差异，CA小于5%，4-ACA小于3%。这些结果表明，使用相对于CAF的RMS的HPLC/PDA方法是一种简单可靠的定量方法，不需要CA和4-ACA有证标准物质。
