Department of Occupational and Environmental Health, Faculty of Pharmaceutical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba, 278-8510, Japan.
Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, TN, USA.
Int Arch Occup Environ Health. 2019 Aug;92(6):873-881. doi: 10.1007/s00420-019-01430-7. Epub 2019 Apr 6.
Urinary excretion of 2,5-hexanedione is currently used to estimate the exposure levels of hexane occurring to an individual during the previous work shift. However, because hexane exposures and urinary 2,5-hexanedione levels can vary considerably from day to day, and subchronic to chronic exposures to hexane are required to produce neuropathy, this biomarker may not accurately reflect the risk of an individual for developing hexane neuropathy. This investigation examines the potential of hexane-derived pyrrole adducts produced on globin and plasma proteins as markers for integrating cumulative exposures. Because the pyrrole markers incorporate bioactivation of hexane to 2,5-hexandione and the initial step of protein adduction involved in hexane-induced neuropathy, they potentially can serve as biomarkers of effect through reflecting pathogenetic events within the nervous system. Additionally, pyrrole formation is an irreversible reaction suggesting that hexane-derived protein pyrroles can be used to assess cumulative exposures to provide a better characterization of individual susceptibilities.
To examine the utility of the proposed markers, blood samples were obtained from eleven workers who used hexane for granulating metal powders in a slurry to produce metal machining die tools and four non-exposed volunteers. Globin and plasma were isolated, and the proteins were digested using pepsin, reacted with Ehrlich's reagent and the level of pyrrole adducts were determined by absorbance at 530 nm. To determine the dose-response curve and dynamic range of the assay, erythrocytes were incubated with a range of 2,5-hexanedione concentrations and the net absorbance at 530 nm of isolated globin was measured.
Pyrrole was detected in both the globin and plasma samples of the workers exposed to hexane and the levels of pyrroles in plasma were positively correlated with the levels of pyrroles in globin for most of the workers.
This investigation demonstrates that detectable levels of hexane-derived protein pyrrole adducts are produced on peripheral proteins following occupational exposures to hexane and supports the utility of measuring pyrroles for integrating cumulative exposures to hexane.
目前，2,5-己二酮的尿排泄被用于估计个体在前一个工作班次中接触己烷的暴露水平。然而，由于己烷暴露和尿 2,5-己二酮水平可能每天都有很大变化，并且需要亚慢性到慢性暴露于己烷才能产生神经病，因此这种生物标志物可能无法准确反映个体发生己烷神经病的风险。本研究检查了己烷衍生的吡咯加合物在珠蛋白和血浆蛋白上产生作为整合累积暴露的标志物的潜力。由于吡咯标记物包含己烷生物转化为 2,5-己二酮和与己烷诱导的神经病有关的蛋白加合物的初始步骤，因此它们可以作为效应生物标志物，反映神经系统内的发病事件。此外，吡咯的形成是一个不可逆的反应，表明己烷衍生的蛋白吡咯可用于评估累积暴露，以更好地描述个体易感性。
为了检验所提出的标志物的实用性，从 11 名使用己烷作为浆液中的金属粉末造粒以生产金属机械加工模具工具的工人和 4 名非暴露志愿者中采集了血液样本。分离珠蛋白和血浆，并使用胃蛋白酶消化蛋白质，用 Ehrlich 试剂反应，通过 530nm 处的吸光度测定吡咯加合物的水平。为了确定该测定的剂量-反应曲线和动态范围，将红细胞与一系列 2,5-己二酮浓度孵育，并测量分离珠蛋白的 530nm 处的净吸光度。
在接触己烷的工人的珠蛋白和血浆样本中均检测到吡咯，并且大多数工人的血浆中吡咯的水平与珠蛋白中吡咯的水平呈正相关。
本研究表明，在职业接触己烷后，外周蛋白上会产生可检测水平的己烷衍生蛋白吡咯加合物，并支持测量吡咯来整合己烷的累积暴露的实用性。
