METHODS MONITOR TOXIC SUBSTAN AND/OR INDICATORS OF PRESENCE IN HUMANS&OTHER SPE

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。人为环境毒素，即使是低剂量，也会在植物、动物、微生物甚至人体内引起一定程度的生物变化。加州大学戴维斯分校超级基金项目的目标是发现观察和量化这些环境影响的生物标志物的方法，以便了解、评估、追踪和补救这些影响的来源和原因。为此，该计划将AMS定量作为其分析核心能力之一。该分析核心服务于多个项目，包括土壤和废物运输、免疫化学生物标志物、肺生物标志物和生殖生物标志物的开发。加速器质谱法（AMS）在评估人类接触有毒物质和探索人类和其他宿主物种毒性的机制基础方面发挥着重要作用。这是我们在农业和工业活动中使用环境暴露于有毒物质的生物标志物项目中的核心技术。我们定义了特定毒性暴露的尿液、肺部、生殖和循环生物标志物，这些生物标志物可通过免疫分析、蛋白质质谱分析、色谱分析和AMS同位素标记毒素的直接定量等方法进行量化。AMS还通过同位素标签从毒素进入宿主的相关性提供其他测定的校准。然后通过AMS测量所指示的毒素摄取来校准衍生生物标志物的定量。在“运输”事件中，调查人员正在评估最近使用的燃料添加剂甲基叔丁基醚（MTBE）的生物活性，这种添加剂在过去十年中从燃料仓库泄漏到地下。目前正在研究14C-MTBE与哺乳动物蛋白的结合，以确定该化合物是否对细胞系统构成威胁。这些实验室研究可以自由地完成与细胞培养物相互作用所需的14C水平，但该计划的大部分内容涉及在不可能释放放射性示踪剂的自然环境中量化生物标志物。定量可识别的生物标记物的首选技术是免疫测定，它最终可以制成现场可用的试剂盒。重要的是要选择正确的目标免疫测定发展，如最有可能的代谢物或化学物质暴露的激素反应。AMS是发现最佳免疫测定靶点的一项特别有价值的技术，因为它可以显示同位素标记的异种生物的所有代谢物，即使在低剂量暴露下也是如此。我们发现阿特拉津的二烷基巯基代谢产物是人类中这种普遍存在的除草剂最突出的持久生物标志物。针对这些生物标记物开发了免疫测定法。生态系统中有一些对环境变化敏感的“标记”物种，就像过去几个世纪里煤矿里的金丝雀一样。越来越多的污染物被认为是荷尔蒙的仿制品，通过影响一个物种的繁殖成功，对它起着“毒药”的作用。我们以小鹌鹑为例，在它们的粪便中发现了睾酮或可的松的代谢物，这些粪便被用作样本，以避免捕获的鸟类受到应激效应。代谢产物的模式将被量化，以发现哪些可能是缓慢发展的环境压力的迹象。这种鸟很小，不能大量用药，因此需要AMS的灵敏度。需要从与环境有关剂量的呼吸来研究肺部对环境化学物质的反应。模型动物肺组织中特定蛋白质的剂量沉积很难定量，目前的方法是提供大剂量暴露，然后在二维凝胶上分离蛋白质，然后进行长期（1个月）放射自显影。AMS对合适的剂量具有敏感性，并且已经制定了顺序凝胶分离以最大限度地发现目标蛋白。AMS核心用于识别暴露的突出生物标志物，用于现场分析开发，并为项目研究人员量化暴露于标记化合物。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Anthropogenic environmental toxins, even at low doses, cause some measure of biological change to take place, within plants, animals, microbes, or even humans. The goal of the UC Davis Superfund Program is to discover ways to observe and quantify these biomarkers of environmental impacts, so that the sources and causes of of these impacts can be understood, assessed, traced, and remediated. To that end, the Program includes AMS quantitation as one of its competencies in its analytical core. This analytical core serves several of the Program projects, including Soil and Waste Transport, development of Immunochemical Biomarkers, Pulmonary Biomarkers, and Reproductive Biomarkers. Accelerator mass spectrometry (AMS) plays an important role in the assessment of human exposure to toxic substances and in probing the mechanistic basis of toxicity in humans and in other host species. It is a core technology in our program of using biomarkers of environmental exposures to toxic substances from agricultural and industrial activities. We define urinary, pulmonary, reproductive, and circulating biomarkers of specific toxic exposures that are quantifiable using assays such as immunoassays, protein mass spectrometry, chromatography, and direct quantitation of isotope labeled toxins with AMS. AMS also provides calibration of the other assays through correlation of isotope label incorporation from toxins into a host. Quantitation of a derived biomarker is then calibrated by the uptake of toxin indicated by the AMS measurements. In the case of Transport, the investigators are assessing the biological activity of the recently used fuel additive, methyl-tert-butyl-ether (MTBE), which leaked into the ground from fuel depots over the past decade. The binding of 14C-MTBE to mammalian protein is being studied to determine if the compound presents a threat to cellular systems. These laboratory studies are freely done with the levels of 14C needed to interact with cell cultures, but much of the Program is concerned with quantifying biomarkers in natural settings where radiotracer release is not possible. The preferred technology for quantifying recognizable biomarkers is the immunoassay which can eventually be made into field-usable kits. It is important to choose the right target for immunoassay development, such as the most likely metabolite or hormonal response of a chemical exposure. AMS is a particularly valuable technology for the discovery of optimal immunoassay targets because it reveals all metabolites of an isotope-labeled xenobiotic, even at low dose exposures. We found that the di-dealkyl mercapturate metabolites of atrazine were the most prominent lasting biomarkers of this ubiquitous herbicide in humans. Immunoassays are developed for these biomarkers. There are "marker" species in ecosystems which are sensitive to environmental change, much like the canaries of past centuries in coal mines. An increasing number of polutants are being seen as hormonal mimics that act as "poison" to a species by imparing its reproductive success. We are using small quail as one such example and are finding the metabolites of testosterone or cortisone in their fecal droppings, which are used as sample so as to avoid stress effects in a captured bird. The pattern of metabolites will be quantified to find which might be signs of slowly developing environmental stresses. The birds are small, and cannot be heavily dosed, so the sensitivity of AMS is needed. Pulmonary responses to environmental chemicals need to be studied from respiration of environmentally relevant doses. The dose deposition in specific proteins of lung tissue of model animals is poorly quantified by present methods that provide a large exposure followed by protein separation on two dimensional gels followed by long term (1 month) autoradiography. AMS has the sensitivity for appropriate doses and sequential gel separations have been worked out to maximize target protein discovery. The AMS core serves to identify prominent biomarkers of exposure for fieldable assay development and quantifies exposures to labeled compounds for the Program researchers.