© 1992 Oxford University Press
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Ascorbate is the principal reductant of chromium(VI) in rat lung ultrafiltrates and cytosols, and mediates chromium–DNA binding in vitro
Department of Chemistry, Dartmouth College Hanover, NH 03755, USA
1Present address: Department of Experimental Pathology and Toxicology, Chemical Industry Institute of Toxicology Research Triangle Park, NC 27709, USA
2To whom correspondence should be addressed
Chromium(VI) reductase activity was measured in ultrafiltrates of rat lung after various pretreatments in vitro at 37°C and pH 7.0. Pretreatment of ultrafiltrates with L-ascorbate oxidase (EC 1.10.3.3 [EC] ), which specifically eliminated ascorbate, blocked {small tilde} 95% of chromium(VI) reductase activity in ultrafiltrates. Preincubation of ultrafiltrates with heat-denatured ascorbate oxidase or the sulfhydryl-blocking agent N-ethylmaleimide (NEM) had no significant effect on Cr(VI) reductase activity. In rat lung cytosols, L-ascorbate oxidase blocked {small tilde} 95% and NEM blocked {small tilde} 15% of Cr(VI) reductase activity. The extent of inhibition of Cr(VI) reductase activity in cytosols by L-ascorbate oxidase was significantly decreased to {small tilde}75% after addition of 1.0 mM NADPH. When Cr(VI) was incubated with salmon sperm nuclei suspended in rat lung cytosol for 15 min, Cr became bound to nuclear DNA. This Cr-DNA binding was completely inhibited by preincubation of rat lung cytosols with L-ascorbate oxidase and inhibited {small tilde} 60% by preincubation with NEM. Taken together these data suggest that ascorbate and/or ascorbate-dependent factors are the principal reductants of Cr(VT) in both ultrafiltrates and cytosols prepared from rat lung and ascorbate-dependent metabolism of Cr(VI) results in Cr binding to nuclear DNA in vitro. Although sulfhydryl-containing factors and NADPH-dependent factors only make a minor contribution to Cr(VI) reduction in rat lung cytosols, sulfhydryls may be significantly involved in the binding of Cr to nuclear DNA.
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