The biomethylation of arsenic
Creators: name: Li, Hao
Publisher: name: University of British Columbia
Year: 2009

description: A semi-continuous hydride generation-gas chromatography-atomic absorption spectrometry (HG-GC-AA) system was developed and optimized for the determination of arsenite, arsenate, methylarsonate (MMA), dimethylarsinate (DMA), and trimethylarsine oxide (TMAO). Particularly, this system was used to study the pathway for the biomethylation of arsenicals in microorganisms and a marine alga. The HG-GC-AA system was used to separate and identify the extracellular arsenic metabolites produced by the microorganisms Apiotrichum humicola (previously known as Candida humicola) and Scopulariopsis brevicaulis growing in the liquid medium enriched with arsenicals. Arsenite, MMA, DMA, and TMAO were detected following incubation with arsenate. With arsenite as a substrate, the metabolites were MMA, DMA, and TMAO; MMA afforded DMA and TMAO, and DMA afforded TMAO. Trimethylarsine was not detected in these investigations. The production of the anticipated methylated intermediates from the substrates strongly support the metabolic sequence proposed by Challenger (Challenger, F Chem. Rev., 1945, 36:315). When L3-methionine-methyl-d₃ was added to the growing culture of Apiotrichum humicola grown in the presence of either arsenate, arsenite, MMA, or DMA, the CD₃ label was incorporated intact into the arsenic metabolites (DMA and TMAO) to a considerable extent, indicating that S-adenosylmethionine (SAM), or some related suiphonium compound, is involved in the biological methylation. Conclusive evidence of CD₃ incorporation into the arsenicals was provided by using a specially developed hydride generation-gas chromatography-mass spectrometry methodology (HG-GC-MS). When a unicellular marine alga Polyphysa peniculus was grown in artificial seawater enriched with arsenicals, the arsenic metabolites produced in the cells as well as in the growth medium were identified by using HG-GC-AA methodology. Arsenite and DMA were detected following incubation with arsenate. When the alga was treated with arsenite, DMA was the major metabolite in the cells and in the growth medium; trace amounts of MMA were also detected in the cells. With methylarsonate as a substrate, the metabolite was dimethylarsinate. Polyphysa peniculus did not metabolize dimethylarsinic acid when it was used as a substrate. Significant amounts of more complex arsenic species, such as arsenosugars, were not observed in the cells or medium based on the evidence given by flow injection-microwave digestion-hydride generationatomic absorption spectrometry methodology. Transfer of the exposed cells to fresh medium caused release of most cell associated arsenicals to the surrounding environment. The alga seems to follow the biomethylation pathway proposed by Challenger for microbial process, and in the case of P. peniculus, DMA is the end product of this biomethylation. When L-methionine-methyl-d₃ was added to the culture of Polyphysa peniculus enriched with 1 ppm of arsenate, the CD₃ label was incorporated intact in the DMA metabolite to a considerable extent. It thus confirmed that P. peniculus also follows the oxidation-reduction pathway involving carbonium ions originally suggested by Challenger for the alkylation of arsenic by microorganisms. The HG-GC-MS system was also used to identify the antimony hydrides produced from the trimethylantimony compoundsMe₃Sb(OH)₂ and Me₃SbCl₂.The possible causes of the molecular rearrangement of trimethyistibine were investigated. The extracts of plant samples collected from Kam Lake and Keg Lake (Yellowknife) were analyzed by using the HG-GC-MS system. The results provided conclusive evidence of the presence of methylantimony compounds in these samples.