Direct analysis and stability of methylated trivalent arsenic metabolites in cells and tissues.
Chronic ingestion of water containing inorganic arsenic (iAs) has been linked to a variety of adverse health effects, including cancer, hypertension and diabetes. Current evidence suggests that the toxic methylated trivalent metabolites of iAs, methylarsonous acid (MAs(III)) and dimethylarsinous acid (DMAs(III)) play a key role in the etiology of these diseases. Both MAs(III) and DMAs(III) have been detected in urine of subjects exposed to iAs. However, the rapid oxidation of DMAs(III) and, to a lesser extent, MAs(III) in oxygen-rich environments leads to difficulties in the analysis of these metabolites in samples of urine collected in population studies. Results of our previous work indicate that MAs(III) and DMAs(III) are relatively stable in a reducing cellular environment and can be quantified in cells and tissues. In the present study, we used the oxidation state-specific hydride generation-cryotrapping-atomic absorption spectroscopy (HG-CT-AAS) to examine the presence and stability of these trivalent metabolites in the liver of mice and in UROtsa/F35 cells exposed to iAs. Tri- and pentavalent metabolites of iAs were analyzed directly (without chemical extraction or digestion). Liver homogenates prepared in cold deionized water and cell culture medium and lysates were stored at either 0 °C or -80 °C for up to 22 days. Both MAs(III) and DMAs(III) were stable in homogenates stored at -80 °C. In contrast, DMAs(III) in homogenates stored at 0 °C began to oxidize to its pentavalent counterpart after 1 day; MAs(III) remained stable for at least 3 weeks under these conditions. MAs(III) and DMAs(III) generated in UROtsa/F35 cultures were stable for 3 weeks when culture media and cell lysates were stored at -80 °C. These results suggest that samples of cells and tissues represent suitable material for the quantitative, oxidation state-specific analysis of As in laboratory and population studies examining the metabolism or toxic effects of this metalloid.
Curriculum in Toxicology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA.
Jenna M
JM
Currier
Milan
M
Svoboda
Tomáš
T
Matoušek
Jiří
J
Dědina
Miroslav
M
Stýblo
D000818:Animals; D001151:Arsenic / Q000032:analysis / Q000378:metabolism*; D001152:Arsenicals / Q000032:analysis / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D008099:Liver / Q000737:chemistry / Q000378:metabolism*; D008745:Methylation; D051379:Mice; D008810:Mice, Inbred C57BL; D010084:Oxidation-Reduction; D013054:Spectrophotometry, Atomic
D001152:Arsenicals; D001151:Arsenic