Ascorbate is the principal reductant of chromium(VI) in rat lung ultrafiltrates and cytosols, and mediates chromium-DNA binding in vitro.
Chromium(VI) reductase activity was measured in ultrafiltrates of rat lung after various pretreatments in vitro at 37 degrees C and pH 7.0. Pretreatment of ultrafiltrates with L-ascorbate oxidase (EC 1.10.3.3), which specifically eliminated ascorbate, blocked approximately 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 approximately 95% and NEM blocked approximately 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 approximately 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 approximately 60% by preincubation with NEM. Taken together these data suggest that ascorbate and/or ascorbate-dependent factors are the principal reductants of Cr(VI) 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.
Standeven
A M
AM
Department of Chemistry, Dartmouth College, Hanover, NH 03755.
Wetterhahn
K E
KE
D000818:Animals; D001205:Ascorbic Acid / Q000494:pharmacology*; D002857:Chromium / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D066298:In Vitro Techniques; D008168:Lung / Q000201:enzymology / Q000378:metabolism*; D008297:Male; D010084:Oxidation-Reduction / Q000187:drug effects; D010088:Oxidoreductases / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D013438:Sulfhydryl Compounds / Q000494:pharmacology; D014462:Ultrafiltration
D013438:Sulfhydryl Compounds; D002857:Chromium; D004247:DNA; D010088:Oxidoreductases; C075965:chromium(VI) reductase; D001205:Ascorbic Acid