RP-HPLC柱后衍生法检测干酪中6种生物胺  

Simultaneous Determination of Six Biogenic Amines in Cheese by RP-HPLC Coupled to On-Line Post Column Derivatization

作　　者：郭雯[1] 郑冬梅[1] 李晓东[1] 张宏伟[1] 孙爱杰 王建平 

机构地区：[1]东北农业大学乳品科学教育部重点实验室食品学院,哈尔滨150030 [2]黑龙江飞鹤乳业有限公司,黑龙江齐齐哈尔164800

出　　处：《中国食品学报》2015年第5期213-218,共6页Journal of Chinese Institute Of Food Science and Technology

基　　金：国家科技部"十二五"科技支撑计划项目(No.2012BAD12B00-5)

摘　　要：目的：建立干酪中6种生物胺的高效液相色谱-柱后衍生检测方法。方法：采用高氯酸提取干酪中生物胺,以邻苯二甲醛（OPA）为柱后衍生试剂,高效液相色谱-柱后衍生-荧光法检测。采用Agilent Eclipse XDBC18色谱柱;流动相A为乙酸钠-辛烷磺酸钠溶液,流动相B为乙酸钠-辛烷磺酸钠和乙腈混合溶液,流速1.0m L/min,梯度与等度相结合洗脱方式;柱温40℃;荧光探测器激发波长330 nm,发射波长465 nm;衍生剂流量0.3 m L/min,衍生池温度45℃。结果 ：采用本洗脱程序,40 min内6种生物胺完全分离;衍生剂OPA质量浓度0.4 mg/m L,流速0.3 m L/min时衍生效果最佳;6种生物胺标准曲线具有良好的线性关系（R2〉0.9982）,平行样品相对标准偏差（RSD）〈3.8%,方法稳定性良好,6种生物胺样品平均回收率89.25%~103.09%。结论：本法样品预处理简便,灵敏度高,重复性好,准确性高,适合检测干酪中的生物胺。Objective： To develop post-column RP-HPLC method for determination of six biogenic amines in cheese. Methods： Determination of six biogenic amines was applied to on-line post-column derivatization RP-HPLC with fluores- cence detector. Six Biogenie amines were obtained by perchlorie acid from cheese, o-phthaldehyde （OPA） as the post- column derivatization reagent. Biogenie amines were HPLC separated on an Agilent Eclipse XDB-C18 column. The gradi- ent elution and isocratic elution were performed by two mobile phases consisted the mixture of sodium acetate - sodium octanesulphonate - acetonitrile at a flow rate of 1.0 mL/min and 40 ℃. Fluorimetric detection at 330 nm for excitation and 465 nm for emission was applied. The flow rate of the derivatization reagent was 0.3mL/min while the on-line post- column reaction equipment was kept at 45 ℃. Results： The method allows the resolution of a total of six biologically ac- tive amines in a single chromatographic run on a 40 min elution programme. The derivatization was the best when the flow rate of the derivatization reagent was 0.3 mL/min in 0.4 mg/mL OPA solution. The coefficient of determination （Rz） was higher than 0.9982 for all standard curves, relative standard deviation （RSD） among different parallels was 〈 3.8%. The average recoveries were 89.25%-103.09%. Conclusion： The post-column RP-HPLC method described is a reliable procedure to determine six biogenic amines in cheese in less than 40 min of chromatographic elution, showing a satis- factory linearity, sensitivity, precision and stability.

关 键 词：干酪 生物胺 反相高效液相色谱法 柱后衍生法 

分 类 号：O657.72[理学—分析化学] TS252.53[理学—化学]