酶解样品-高效液相色谱法测定糯米类制品中10种合成着色剂的含量

Determination of 10 Synthetic Colorants in Glutinous Rice Products by High Performance Liquid Chromatography after Sample Enzymolysis

取糯米类制品5 g,经中温淀粉酶(2 000~7 000 U·g^(-1)) 150 mg于60℃水浴中酶解6 h,得到酶解待测液。在上述溶液中加入调成粥状的聚酰胺粉(0.075~0.15 mm) 1 g,经G3垂溶漏斗抽滤,滤饼依次经60℃水(10 mL×3)淋洗,体积比7∶2∶1的乙醇-氨水-水混合液(15 mL×3)洗脱,所得洗脱液加热浓缩至5 mL左右,浓缩液用水定容至20 mL,离心。所得上清液采用高效液相色谱法测定其中10种合成着色剂的含量,以C_(18)色谱柱为固定相,以不同体积比的甲醇和20 mmol·L^(-1)乙酸铵溶液的混合液为流动相梯度洗脱,二极管阵列检测器在430 nm(检测柠檬黄),500 nm(检测日落黄、新红、胭脂红、苋菜红、诱惑红、偶氮玉红、赤藓红),600 nm(检测靛蓝、亮蓝)等3个波长条件下检测。结果表明:10种合成着色剂的质量浓度均在0.2~20.0 mg·L^(-1)内与其对应的峰面积呈线性关系,检出限(3S/N)为0.19~0.43 mg·kg^(-1)。按标准加入法进行回收试验,回收率为76.9%~94.0%,测定值的相对标准偏差(n=6)均小于4.0%。方法用于分析市售的10批糯米类制品,3批样品检出合成着色剂,分别为柠檬黄、日落黄、亮蓝。本法与国家标准GB 5009.35-2016测定结果相比,有效克服该类食品的假阴性结果。

The samples of glutinous rice products(5 g) were taken, and the enzymatic hydrolysis was carried out in a 60 ℃ water bath for 6 h with 150 mg of medium temperature amylase(2 000-7 000 U·g^(-1)), then the solution of enzymatic hydrolysis to be tested was obtained. 1 g of polyamide powder(0.075-0.15 mm) mixed into porridge was added into the above solution, and filtered through G3 vertical dissolution funnel. The filter cake was successively washed with 60 ℃ water(10 mL×3) and eluted with a mixture of ethanol, ammonia and water at volume ratio of 7∶2∶1(15 mL×3). The obtained eluent was heated and concentrated to about 5 mL. The concentrated solution was made its volume up to 20 mL with water and then centrifuged. 10 synthetic colorants in the supernatant were determined by high performance liquid chromatography(HPLC). C_(18)chromatographic column was used as the stationary phase, the mixture of methanol and 20 mmol·L^(-1)ammonium acetate solution at different volume ratio was used as the mobile phase for gradient elution, and the diode array detector was at 430 nm for detection of citrine, 500 nm for detection of sunset yellow, new red, ponceau 4 R, amaranth, allura red, azorubine and etythrosine B, and 600 nm for detection of indigotin and brilliant blue. As shown by the results, linear relationships between values of peak area and mass concentration of 10 synthetic colorants were found in the same ranges of 0.2-20.0 mg·L^(-1), with detection limits(3 S/N) in the range of 0.19-0.43 mg·kg^(-1). Test for recovery was made by standard addition method, giving results in the range of 76.9%-94.0%, with RSDs(n=6) of the determined values less than 4.0%. This method was applied for analysis of 10 batches of glutinous rice products on the market, and citrine, sunset yellow and brilliant blue were detected respectively in 3 batches of samples. Compared with the determination results of national standard GB 5009.35-2016, this method could effectively overcome the false negative results of this kind of food.