ID AND DEV OF BIOLOGICAL MARKERS OF HUMAN EXPOSURE TO THE INSECTICIDE PERMETHRI

人类接触杀虫剂氯菊酯的生物标志物的识别和开发

• 批准号: 8362754
• 负责人: BRUCE D HAMMOCK
• 金额: $ 16.38万
• 依托单位: UNIVERSITY OF CALIF-LAWRNC LVRMR NAT LAB
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8362754
• 关键词: Acids; Agricultural Workers; Alcohols; Antibodies; Biological Assay; Biological Markers; Biological Markers; Blood; C14 isotope; Chemicals; Clinical; Collection; Data; Dermal; Detection; Development; Dose; Exposure to; Funding; Glucuronides; Glycine; Grant; Half-Life; Haptens; Health; High Pressure Liquid Chromatography; Hour; Human; Immunoassay; Insect Repellents; Insecticides; Knowledge; Lice; Liquid Chromatography; Liquid substance; Manuscripts; Mass Spectrum Analysis; Measures; Methods; Military Personnel; Monitor; National Center for Research Resources; Pattern; Permethrin; Pesticides; Preparation; Principal Investigator; Publications; Radiolabeled; Research; Research Infrastructure; Resource Development; Resources; Review Literature; Route; Saliva; Sampling; Scintillation Counting; Soaps; Source; System; United States National Institutes of Health; Urine; Validation; accelerator mass spectrometry; analytical method; base; cost; exposed human population; liquid chromatography mass spectrometry; metabolic abnormality assessment; prevent; pyrethroid; radiotracer; tool; ultraviolet; urinary

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Agricultural workers, gardeners and homeowners are routinely exposed to the insecticide permethrin. Also, military personnel are exposed to the permethrin when using the DOD Insect Repellent System and over-the-counter lice soaps use permethrin as the active ingredient. A urinary metabolite of permethrin, that is in high abundance and is relatively stable, may be an ideal biomarker of exposure to this pesticide. In addition, the ratio of one metabolite to another may vary, according to the route of administration. The results of this study would be used to identify candidates for the development of a rapid, sensitive immunochemical based analytical method that can be used to routinely monitor human exposure to permethrin. Objectives: The purpose of this study is to determine the human metabolite(s) of permethrin in urine following dermal exposure that are in greatest abundance and are the most stable. Accelerator mass spectrometry is a method for measuring levels of 14C several orders of magnitude more sensitive than liquid scintillation counting. With this high sensitivity we will conduct human metabolism studies at biologically relevant doses. SPECIFIC AIMS: I. Develop an LC/MS method for separation of permethrin and its putative human metabolites. II. Determine the human metabolite profile of permethrin using accelerator mass spectrometry (AMS). III. Develop an immunoassay to the key metabolite identified in Objective II as a biomarker of human exposure to permethrin. METHODOLOGY: For specific aim I, synthesize metabolite standards; develop an HPLC method to separate the putative pyrethroid metabolites using ultraviolet detection; determine the feasibility of an HPLC/mass spectrometry method for analysis of pyrethroid metabolites. For specific aim II, clinical exposure of humans to radiolabeled permethrin dermally and collection of urine, blood and saliva; separation of samples by methods developed in specific aim I and analysis of separated samples by accelerator mass spectrometry; identification of most prevalent metabolite from resultant data. For specific aim III, synthesis of haptens; development of antibodies; use of the haptens and antibodies in the development of an immunoassay for the most prevalent metabolite; validation of immunoassay. EXPECTED PRODUCTS (MILESTONES): Literature review of putative human metabolites of permethrin; small quantities of synthesized metabolites of permethrin; an HPLC method for separating permethrin metabolites in human urine or saliva; identification of the most abundant human metabolite(s) by accelerator mass spectrometry; an immunoassay to detect the targeted human metabolite of permethrin STATUS/RESULTS TO DATE: Literature review has been completed and putative major metabolites identified. All of the major metabolites have been synthesized or acquired. Using the metabolite standards a high performance liquid chromatography method for their analysis has been developed. This method will later be used to identify metabolites found in human urine samples and the liquid chromatography-mass spectrophotometric method used for validation. Using the chemical knowledge from the preparation of metabolites, synthesis of haptens for immunoassay detection of these molecules is complete. Immunoassays for 3-phenoxybenzoic acid, the glycine conjugate of 3-phenoxybenzoic acid, the glycine conjugate of dichlorovinylchrysanthemic acid (DCCA) and the glucuronide conjugate of 3-phenoxybenzyl alcohol (see publication below) have been developed. An assay for free DCCA is in progress (manuscript in preparation). The assay for 3-phenoxybenzoic acid has been adapted to a sensitive, high throughput method (see publication below). Clinical exposures have been completed. All samples have been measured by AMS for total carbon-14. An estimate of the total dose absorbed (for 4 subjects) ranged from 0.06 to 0.27%. The permethrin is eliminated from the blood with a half life of about 12-24 hours hours. The urinary half life averaged 24 hours. Saliva was sampled, but permethrin does not appear to be excreted by that route. Liquid chromatography analysis of the metabolite pattern in urine is underway. Conclusion: The results of this study will be used to identify candidates for the development of a rapid, sensitive immunochemical based analytical method that can be used to routinely monitor human exposure to permethrin. The ability to carefully monitor the presence of absorbed doses of permethrin will be a useful tool to prevent the possibility of human health effects due to permethrin exposure.

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 农业工人、园丁和房主经常接触杀虫剂氯菊酯。 此外，军事人员在使用国防部驱虫剂系统时会接触到氯菊酯，而非处方虱子肥皂则使用氯菊酯作为活性成分。氯菊酯的尿代谢产物丰度高，相对稳定，可能是暴露于该农药的理想生物标志物。 此外，根据给药途径，一种代谢物与另一种代谢物的比例可能不同。本研究的结果将用于确定候选人的快速，灵敏的免疫化学为基础的分析方法，可用于常规监测人类接触氯菊酯的发展。 目的：本研究的目的是确定皮肤接触后尿液中最丰富且最稳定的氯菊酯人体代谢物。加速器质谱是一种测量14 C水平的方法，比液体闪烁计数灵敏几个数量级。凭借如此高的灵敏度，我们将以生物相关剂量进行人体代谢研究。 具体目标： 开发用于分离氯菊酯及其推定的人体代谢物的LC/MS方法。 二. 使用加速器质谱法（AMS）测定氯菊酯的人体代谢物谱。 三. 开发目标II中确定的关键代谢物的免疫测定法，作为人类暴露于氯菊酯的生物标志物。 研究方法：对于特定目标I，合成代谢物标准品;开发HPLC方法，使用紫外检测分离推定的拟除虫菊酯代谢物;确定HPLC/质谱法分析拟除虫菊酯代谢物的可行性。 对于特定目标II，人体皮肤临床暴露于放射性标记的二氯苯醚菊酯，并采集尿液、血液和唾液;通过特定目标I中开发的方法分离样品，并通过加速器质谱法分析分离的样品;从所得数据中鉴定最常见的代谢物。对于特定目标III，半抗原的合成;抗体的开发;半抗原和抗体在最常见代谢物的免疫测定开发中的应用;免疫测定的验证。 预期产品（里程碑）：假定的二氯苯醚菊酯人体代谢物的文献综述;少量合成的二氯苯醚菊酯代谢物;用于分离人尿液或唾液中二氯苯醚菊酯代谢物的HPLC方法;通过加速器质谱法鉴定最丰富的人体代谢物;检测二氯苯醚菊酯靶向人体代谢物的免疫测定法 迄今为止的状态/结果：已完成文献综述，并确定了推定的主要代谢产物。 所有的主要代谢产物都已合成或获得。 使用代谢物标准品，开发了用于其分析的高效液相色谱法。 该方法随后将用于鉴定人尿样中发现的代谢物，并将液相色谱-质谱分光光度法用于验证。利用代谢物制备的化学知识，完成了用于这些分子的免疫测定检测的半抗原的合成。 已开发了3-苯氧基苯甲酸、3-苯氧基苯甲酸的甘氨酸结合物、二氯乙烯基琥珀酸（DCCA）的甘氨酸结合物和3-苯氧基苄醇的葡糖苷酸结合物的免疫测定法（见下文出版物）。 游离DCCA的测定正在进行中（手稿正在编写中）。3-苯氧基苯甲酸的含量测定已调整为灵敏、高通量方法（见下文出版物）。已完成临床暴露。所有样品都通过AMS测量了总碳-14。总吸收剂量的估计值（4例受试者）范围为0.06%至0.27%。氯菊酯从血液中消除，半衰期约为12-24小时。尿液半衰期平均为24小时。 对唾液进行了采样，但氯菊酯似乎不通过该途径排泄。尿液中代谢物模式的液相色谱分析正在进行中。 结论：本研究的结果将被用来确定候选人的快速，灵敏的免疫化学为基础的分析方法，可用于常规监测人类暴露于氯菊酯的发展。仔细监测氯菊酯吸收剂量的存在的能力将是一个有用的工具，以防止由于氯菊酯暴露对人类健康的影响的可能性。