免疫亲和层析净化-高效液相色谱法检测红曲产品中桔青霉素方法研究

Immuno-affinity chromatographic purification： the study of methods to test citrinin in monascusproducts by high performance liquid chromatography

目的建立红曲产品中桔青霉素的免疫亲和层析净化（IAC）-高效液相色谱（HPLC）检测方法，检测福建相关红曲产品中桔青霉素（CIT）的含量。方法采用IAC—ItPLC的方法对红曲产品中CIT的含量进行测定。HPLC的条件为：色谱柱：C18。反相色谱柱，150．0mm×4．6mm×3μm；流动相：采用乙腈和体积分数为0．1％磷酸溶液，体积比值为65：35，等度洗脱；柱温：28℃；流速：0．8ml／min；荧光检测器波长：激发光波长（λex）为331nm、发射光波长（λem）为500nln。根据峰面积（Y）对CIT浓度（X，ng／m1）进行线性回归，绘制标准曲线，并验证方法的准确度和精密度。采用建立的方法对32种福建红曲产品进行测定和色价比较。结果本研究建立方法的标准曲线为Y=4634．8X-136．42，r=1．000，最低检测限为20肛∥kg，定量限为64μg/kg。在200～800肛∥kg范围内，加标回收率为98．9％～110．0％（n=3），RSD为O．5l％～1．76％。32份样品中，11份红曲米、9份红曲粉和5份红曲色素均检测出CIT，含量为0．212～14．500mg／kg，7份功能性红曲中有4份检mCIT，含量为0．142～0．275mg／kg。结论建立IAC—HPLC法测定红曲产品中CIT的方法。

Objective To establish a method to test citrinin （CIT） in monascus products by immuno-affinity chromatography （IAC）- high performance liquid chromatography （HPLC）, and to detect the content of CIT in monascus products in Fujian province. Methods IAC-HPLC was applied to detect the CIT content in monascus products. The conditions to use HPLC were as follows： Cls reversed-phase chromatographic column, 150. 0 mm×4. 6mm×3 μm; mobile phase：the volume ratio of acetonitrile and 0.1% phosphoIic acid solution at 65： 35; isocratic elution; column temperature： 28℃; flow velocity： 0. 8 ml/min; fluorescence detector, excitation wavelength （λex ） was 331 nm and emission wavelength （λem ） was 500 nm. The standard curved was established by the linear regression of peak area （Y） to CIT content （X, ng/ml）. The accuracy and precision of the method would then be verified. And 32 kinds of monascus products were determined and their color values were compared by this method. Result The standard curve established in this study was Y=4634. 8X - 136. 42, r = 1. 000; whose limits of detection was 20μg/kg and the limits of qualification was 64μg/kg. In the range between 200 and 800 μg/kg, the standard recovery rate was 98.9% - 110. 0% （n = 3）, and the relative standard deviation（RSD） was 0. 51% - 1.76%. Out of the 32 samples, CIT was detected from 11 samples of monascus rice, 9 samples of monascus powder and 5 samples of monascus pigments, the content was around 0. 212 - 14. 500 mg/kg. 4 out of 7 functional monascus samples were detected out CIT, whose content at 0. 142 -0. 275 mg/kg. Conclusion The method to detect CIT in monascus nroducts by IAC-HPLC has been established.