Pyrrole adducts in globin and plasma of workers exposed to hexane
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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.
KeywordsHexane Pyrrole Neurotoxicity Biomarker Adduct
Our sincere thanks to Dr. Doyle G. Graham (Vanderbilt University Medical Center, TN, USA), Dr. Yasutaka Ogawa, Dr. Naomi Hisanaga (National Institute of Industrial Health, Kanagawa, Japan) and Dr. Yasuhiro Takeuchi (Emeritus Professor of Nagoya University, Nagoya, Japan).
GI and WMV deigned the project and wrote the manuscript. GI, VA and HLV measured pyrrole adducts and protein. TT, KM and TS conducted the field-work. TK measured urinary 2,5-hexanedione. VA drew the figure of pyrrole formation and detection.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no competing interests.
All assays were performed with the subjects’ informed consent following the Helsinki Declaration. The study protocol was approved by the ethical committee of Tokyo University of Science.
- DeCaprio AP (1986) Mechanisms of in vitro pyrrole adduct autoxidation in 2,5-hexanedione-treated protein. Mol Pharmacol 30:452–458Google Scholar
- DeCaprio AP (1987) N-hexane neurotoxicity: a mechanism involving pyrrole adduct formation in axonal cytoskeletal protein. Neurotoxicology 8:199–210Google Scholar
- Graham DG, Anthony DC, Szakal-Quin G, Gottfried MR, Boekelheide K (1985) Covalent crosslinking of neurofilaments in the pathogenesis of n-hexane neuropathy. Neurotoxicology 6:55–63Google Scholar
- Takahashi M, Takeuchi H, Kyo S, Yorifuji S, Seki Y, Hara I (1977) N-hexane polyneuropathy—a case report with a review of the literature. Med J Osaka Univ 28:77–85Google Scholar
- Takeuchi Y, Ono Y, Hisanaga N, Kitoh J, Sugiura Y (1980) A comparative study on the neurotoxicity of n-pentane, n-hexane, and n-heptane in the rat. Br J Ind Med 37:241–247Google Scholar
- Valentine WM, Amarnath V, Amarnath K, Erve JC, Graham DG, Morgan DL et al (1998) Covalent modification of hemoglobin by carbon disulfide: III. A potential biomarker of effect. Neurotoxicology 19:99–107Google Scholar
- Yamamura Y (1969) N-hexane polyneuropathy. Folia Psychiatr Neurol Jpn 23:45–57Google Scholar