Jaeger L L, Jones A D, Hammock B D
Department of Entomology, Facility for Advanced Instrumentation, University of California, Davis, California 95616, USA.
Chem Res Toxicol. 1998 Apr;11(4):342-52. doi: 10.1021/tx9701844.
Improved assessments of human exposure to electrophilic chemicals require rapid and inexpensive analytical techniques that can detect specific urinary metabolites at low levels as needed for epidemiological screenings of large populations. The first aim of this study has been to apply rational hapten design strategies to develop a more sensitive and selective enzyme-linked immunosorbent assay for atrazine mercapturic acid. Polyclonal sheep antiserum was generated against an improved hapten, numerous coating antigen chemistries were evaluated, and assay conditions were optimized. An assay was developed with an IC50 of 0.08 +/- 0.02 micrograms/L (K approximately with 10(-)10 M) for atrazine mercapturic acid. The assay exhibited greatest recognition of atrazine mercapturic acid relative to other known urinary metabolites of atrazine as well as other triazine herbicides. The assay was surprisingly selective to atrazine mercapturic acid over the structurally similar simazine mercapturic acid. Urine samples presented matrix effects due in part to the nonspecific effects of urinary salts, but 4-fold dilution of urine achieved an overall method limit of quantitation of 0.3 micrograms/L. Solid-phase extraction strategies were also developed in an attempt to increase the sensitivity of the overall method. However, a weak positive assay response was present in the solid-phase extracts of unspiked urines, resulting in accurate recovery of atrazine mercapturic acid at microgram/L.
要改进对人类接触亲电化学物质的评估，就需要快速且廉价的分析技术，以便能按照对大量人群进行流行病学筛查的要求，检测低水平的特定尿代谢物。本研究的首要目标是应用合理的半抗原设计策略，开发一种针对莠去津巯基尿酸更灵敏且更具选择性的酶联免疫吸附测定法。针对一种改进的半抗原制备了多克隆羊抗血清，评估了多种包被抗原化学性质，并优化了测定条件。所开发的测定法对莠去津巯基尿酸的半数抑制浓度为0.08±0.02微克/升（亲和常数约为10⁻¹⁰摩尔）。相对于莠去津的其他已知尿代谢物以及其他三嗪类除草剂，该测定法对莠去津巯基尿酸的识别能力最强。令人惊讶的是，该测定法对莠去津巯基尿酸的选择性高于结构相似的西玛津巯基尿酸。尿液样本存在基质效应，部分原因是尿盐的非特异性影响，但尿液4倍稀释后实现了整体方法定量限为0.3微克/升。还开发了固相萃取策略，试图提高整体方法的灵敏度。然而，未加标尿液的固相提取物中存在微弱的阳性测定响应，导致微克/升水平的莠去津巯基尿酸回收率准确。
