Exploring the in vitro formation of trimethylarsine sulfide from dimethylthioarsinic acid in anaerobic microflora of mouse cecum using HPLC-ICP-MS and HPLC-ESI-MS.

Although metabolism of arsenicals to form methylated oxoarsenical species has been extensively studied, less is known about the formation of thiolated arsenical species that have recently been detected as urinary metabolites. Indeed, their presence suggests that the metabolism of ingested arsenic is more complex than previously thought. Recent reports have shown that thiolated arsenicals can be produced by the anaerobic microflora of the mouse cecum, suggesting that metabolism prior to systemic absorption may be a significant determinant of the pattern and extent of exposure to various arsenic-containing species. Here, we examined the metabolism of 34S labeled dimethylthioarsinic acid (34S-DMTA(V)) by the anaerobic microflora of the mouse cecum using HPLC-ICP-MS and HPLC-ESI-MS/MS to monitor for the presence of various oxo- and thioarsenicals. The use of isotopically enriched 34S-DMTA(V) made it possible to differentiate among potential metabolic pathways for production of the trimethylarsine sulfide (TMAS(V)). Upon in vitro incubation in an assay containing anaerobic microflora of mouse cecum, 34S-DMTA(V) underwent several transformations. Labile 34S was exchanged with more abundant 32S to produce 32S-DMTA(V), a thiol group was added to yield DMDTA(V), and a methyl group was added to yield 34S-TMAS(V). Because incubation of 34S-DMTA(V) resulted in the formation of 34S-TMAS(V), the pathway for its formation must preserve the arsenic-sulfur bond. The alternative metabolic pathway postulated for formation of TMAS(V) from dimethylarsinic acid (DMA(V)) would proceed via a dimethylarsinous acid (DMA(III)) intermediate and would necessitate the loss of 34S label. Structural confirmation of the metabolic product was achieved using HPLC-ESI-MS/MS. The data presented support the direct methylation of DMTA(V) to TMAS(V). Additionally, the detection of isotopically pure 34S-TMAS(V) raises questions about the sulfur exchange properties of TMAS(V) in the cecum material. Therefore, 34S-TMAS(V) was incubated and the exchange was monitored with respect to time. The data suggest that the As-S bond associated with TMAS(V) is less labile than the As-S bond associated with DMTA(V).

In vitro biotransformation of an arsenosugar by mouse anaerobic cecal microflora and cecal tissue as examined using IC-ICP-MS and LC-ESI-MS/MS.

This investigation examined chemical and microbiological transformations of an arsenosugar by mouse cecum. To mimic the low oxygen environment in the mammalian gastrointestinal tract, reaction mixtures were incubated under anaerobic conditions. An arsenosugar extracted from ribbon kelp, 3-[5'-deoxy-5-(dimethylarsinoyl)-beta-ribofuranosyloxy]-2-hydroxypropanesulfonic acid, As392, was added to reaction mixtures that contained either cecal microflora or cecal tissue homogenate. These reaction mixtures were incubated at 0 or 37 degrees C for up to 48 hours to monitor biotransformation of the arsenosugar. Analysis of the reaction mixtures by IC-ICP-MS and LC-ESI-MS/MS indicated that the arsenosugar was converted primarily (95%) to its sulfur analog in less than 1 h at 37 degrees C. Conversion of As392 to its sulfur analog was much slower at 0 degrees C (21% conversion after 48 h). In reaction mixtures with cecal tissue homogenate, conversion of As392 to its sulfur analog was slower (77% conversion after 48 h at 37 degrees C). A good mass balance was found in all reaction mixtures between the amount of arsenosugar added and the sum of all detected arsenic-containing products. LC-ESI-MS/MS spectra of the sulfur-containing arsenosugar formed in all reaction mixtures containing cecal microflora compared well with those of a synthetic standard. These results suggest that the anaerobic microflora of the gastrointestinal tract can rapidly convert ingested arsenosugars to sulfur analogs. This biotransformation may affect the subsequent absorption, metabolism, and disposition of arsenic present in arsenosugars.