摘要
将丁香酚(Eul)与4-溴丁酸叔丁酯进行衍生化反应,合成了半抗原4-(4-烯丙基-2-甲氧基-苯氧基)-丁酸(Eul-4-Tbl)。通过偶联载体蛋白并免疫动物后获取丁香酚多克隆抗体,建立了丁香酚的间接竞争化学发光酶免疫分析方法(ic-CLEIA)。水产样品经乙腈提取,Oasis PRiME HLB固相萃取小柱净化后用于测定。结果表明,ic-CLEIA的最佳反应条件为:包被原质量浓度为0.125μg/mL,抗体稀释40000倍,抗体反应时间为30 min,药物稀释液为PBS。该方法的半抑制浓度(IC_(50))为1.28μg/L,检测线性范围为0.25~6.43μg/L,检出限(LOD,IC_(10))为0.11μg/L。样品中丁香酚的加标回收率为76.6%~108%,相对标准偏差(RSD)为5.9%~12%,测定结果与液相色谱-串联质谱法具有良好的相关性(r=0.9904),可用于水产品中麻醉剂丁香酚残留的快速测定。
Eugenol was derived with tert-butyl 4-bromobutanoate to obtain a hapten 4-(4-allyl-2-methoxy-phenoxy)-butyric acid(Eul-4-Tbl).The hapten was coupled to carrier protein as immunogen for animal immunization,and a polyclonal antibody against eugenol was raised.Thus,an indirect competitive chemiluminescent enzyme-linked immunosorbent assay(ic-CLEIA)was developed for the determination of eugenol.The samples were extracted with acetonitrile,and then cleaned up with an Oasis PRiME HLB solid-phase extraction column.Results showed that the optimized assay conditions were as follows:coating antigen concentration:0.125μg/mL,dilution ratio of antibody:40000,reaction time of antibody:30 min,dilution solution for analyte:PBS.The half inhibition concentration(IC_(50))of the developed ic-CLEIA was 1.28μg/L,the liner range was 0.25-6.43μg/L,and the limit of detection(LOD,IC_(10))was 0.11μg/L.The spiked recoveries of eugenol in aquatic products were in the range of 76.6%-108%,with relative standard deviations of 5.9%-12%.The results obtained by this method exhibited good correlation to those of HPLCMS/MS(r=0.9904).The developed method was suitable for rapid determination of anaesthetic eugenol residues in aquatic product.
作者
王强
王旭峰
赵东豪
张英霞
黄珂
WANG Qiang;WANG Xu-feng;ZHAO Dong-hao;ZHANG Ying-xia;HUANG Ke(South China Sea Fisheries Research Institute,Chinese Academy of Fishery Sciences,Key Laboratory of Aquatic Product Processing,Ministry of Agriculture and Rural Affairs,Guangzhou 510300,China)
出处
《分析测试学报》
CAS
CSCD
北大核心
2022年第3期354-360,共7页
Journal of Instrumental Analysis
基金
“蓝色粮仓科技创新”重点专项(2019YFD0901701)。
关键词
丁香酚
麻醉剂
化学发光酶免疫分析
多克隆抗体
水产品
eugenol
anaesthetic
chemiluminescene enzyme immunoassay
polyclonal antibody
aquatic product
