Rapid analysis of diclofenac in freshwater and wastewater by a monoclonal antibody-based highly sensitive ELISA
Rapid analysis of diclofenac in freshwater and wastewater by a monoclonal antibody-based highly sensitive ELISA
复制标题
DOI：
10.1007/s00216-015-9048-9
复制
发表时间：
2015-11-01
影响因子：
4.3
通讯作者：
Knopp, Dietmar
中科院分区：
文献类型：
--
作者：
Huebner, Maria;Weber, Ekkehard;Knopp, Dietmar
非甾体抗炎药（NSAID）双氯芬酸（DCF）在全球范围内的水环境中发现。因此，它引起了公众对人类健康和野生动物潜在长期影响的日益关注。欧洲联盟最近强调了DCF的重要性，将这种药物列入第一份重点危险物质观察清单，以收集全联盟的监测数据。因此，需要快速和廉价的分析方法来进行高样品量的新鲜和废水监测。在这里，第一次，充分表征的单克隆抗体（mAb）对DCF的产生和高度敏感的ELISA开发。最佳抗体（mAb 12G5）具有高度亲和性（K-D = 1.5 × 10（-10）M），对潜在基质干扰（如pH值（pH范围5.2 - 9.2）、钙离子浓度（高达75 mg/L）和腐殖酸含量（高达20 mg/L））稳定。ELISA校准曲线的检出限（LOD，S/N = 3）和IC 50分别为7.8和44 ng/L。工作范围确定为11 - 180纳克/升。平均而言，发现DCF代谢物5-OH-DCF、4'-OH-DCF和DCF-酰基葡糖苷酸的交叉反应性（CR）约为10%，但与母体化合物相比，其他结构相关的NSAID显示结合<1%。虽然在低ppt范围内的DCF浓度在河流和湖泊的水，较高的值为2.9和2.1微克/升的废水流入和流出，分别被发现。这些结果可以通过固相萃取与LC-MS相结合来证实。
The non-steroidal anti-inflammatory drug (NSAID) diclofenac (DCF) is found worldwide in the aqueous environment. Therefore, it has raised increased public concern on potential long-term impact on human health and wildlife. The importance of DCF has been emphasized by the European Union recently by including this pharmaceutical in the first watch list of priority hazardous substances in order to gather Union-wide monitoring data. Rapid and cheap methods of analysis are therefore required for fresh and wastewater monitoring with high sample load. Here, for the first time, well-characterized monoclonal antibodies (mAbs) against DCF were generated and a highly sensitive ELISA developed. The best antibody (mAb 12G5) is highly affine (K-D = 1.5 x 10(-10) M), stable to potential matrix interferences such as pH value (pH range 5.2-9.2), calcium ion concentration (up to 75 mg/L), and humic acid content (up to 20 mg/L). The limit of detection (LOD, S/N = 3) and IC50 of the ELISA calibration curve were 7.8 and 44 ng/L, respectively. The working range was defined between 11 and 180 ng/L. On average, about 10 % cross-reactivity (CR) was found for DCF metabolites 5-OH-DCF, 4'-OH-DCF, and DCF-acyl glucuronide, but other structurally related NSAIDs showed binding < 1 % compared to the parent compound. While DCF concentrations at the low ppt range were measured in river and lake water, higher values of 2.9 and 2.1 mu g/L were found in wastewater influents and effluents, respectively. These results could be confirmed by solid phase extraction combined with LC-MS.