Liquid chromatographic method with immunoaffinity column cleanup for determination of ochratoxin A in barley: collaborative study.
Liquid chromatographic method with immunoaffinity column cleanup for determination of ochratoxin A in barley: collaborative study.
复制标题
免疫亲和柱净化液相色谱法测定大麦中赭曲霉毒素 A：合作研究。
DOI：
10.1093/jaoac/83.6.1377
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发表时间：
2000
影响因子：
1.6
通讯作者：
John Gilbert
中科院分区：
文献类型：
--
作者：
A. C. Entwisle;Alison C. Williams;P. Mann;P. T. Slack;John Gilbert
采用高效液相色谱(LC)和免疫亲和柱层析相结合的方法，对大麦中赭曲霉毒素A的3种浓度水平进行了检测。试验部分用乙腈-水高速混合提取。提取物经过过滤，用磷酸盐缓冲液(PBS)稀释，然后应用于赭曲霉毒素A免疫亲和色谱柱。色谱柱用水洗涤，用甲醇洗脱。然后将溶剂蒸发，残留物重新溶解在注射溶剂中。将该溶液注入反相液相色谱柱后，用荧光检测法测定赭曲霉毒素A。8个低水平天然污染大麦样品和2个空白大麦样品(在信噪比为3：1的情况下，在0.2微克/公斤的检测限值下未发现赭曲霉毒素A)，以及赭曲霉毒素A安瓶、校准剂和添加溶液，被送往13个不同欧洲国家的15个实验室。测试部分以4 ng/g的水平添加赭曲霉毒素A，回收率从65%到113%不等。根据添加样品(盲复本)和自然污染样品(3个水平的盲复本)的结果，重复性的相对标准偏差(RSDR)为4-24%，重复性的相对标准偏差(RSDR)为12-33%。该方法在大麦中赭曲霉毒素A的低水平测定中显示出可接受的实验室内和实验室间精密度，HORRAT值证明了这一点。
A collaborative study was conducted to evaluate a liquid chromatographic (LC) method with immunoaffinity column cleanup for determination of ochratoxin A. The method was tested at 3 concentration levels of ochratoxin A in barley, which represent possible future European regulatory limits. The test portion was extracted with acetonitrile-water by blending at high speed. The extract was filtered, diluted with phosphate-buffered saline (PBS), and applied to an ochratoxin A immunoaffinity column. The column was washed with water and the ochratoxin A eluted with methanol. The solvent was then evaporated and the residue redissolved in injection solvent. After injection of this solution onto reversed-phase LC column, ochratoxin A was measured by fluorescence detection. Eight samples of low level naturally contaminated barley and 2 samples of blank barley (ochratoxin A not found at the limit of detection of 0.2 microg/kg at the signal-to-noise ratio of 3 to 1) were sent, along with ampules of ochratoxin A, calibrant, and spiking solutions, to 15 laboratories in 13 different European countries. Test portions were spiked with ochratoxin A at levels of 4 ng/g, and recoveries ranged from 65 to 113%. Based on results for spiked samples (blind duplicates) and naturally contaminated samples (blind duplicates at 3 levels), the relative standard deviation for repeatability (RSDr) ranged from 4 to 24%, and the relative standard deviation for reproducibility (RSDR) ranged from 12 to 33%. The method showed acceptable within- and between-laboratory precision, as evidenced by HORRAT values, at the low level of determination for ochratoxin A in barley.