TIME-DEPENDENCE OF ACCUMULATION AND BINDING OF INORGANIC AND ORGANIC ARSENIC SPECIES IN RABBIT ERYTHROCYTES

The uptake by rabbit erythrocytes of 0.4 mM arsenite, As(III), arsenat e, As(V), monomethyl-arsinate, MMA(V) and dimethylarsonate, DMA(V) wer e compared over 24 h. In membrane-free hemolysate, the distribution of As between proteins (10 kDa) and ultrafiltrate was determined by ultr afiltration and arsenic species in the ultrafiltrate were identified b y thin layer chromatography methods, H-1 spin-echo Fourier transform N MR was used to follow the binding of these arsenic species to glutathi one (GSH). P-31-NMR was used to observe their effects on high-energy a denine nucleotide levels (ATP, ADP). These results demonstrate that As (III) readily accumulates in cells, reaches a quasi-plateau at 78% of the total As in the incubation system after 1 h and 88% of the total A s after 24 h, On average, 20% of the total erythrocyte As(III) burden is associated with the protein fraction, particularly with hemoglobin (Hb). About 68% of the erythrocyte As(III) burden is bound to GSH. As( III) has no effect on ATP levels during a 5-h incubation. By compariso n, As(V) enters erythrocytes more slowly (53% of the total As after 5 h). Erythrocytes take up 81% of the As(V) in the reaction system after a 24 h incubation. Of the total As burden in As(V)-exposed erythrocyt es, 22% was associated with the proteins (10 kDa) and possibly reduced to As(III) and 59% was in the ultrafiltrate (8% as As(III) and 51% as As(V)). This finding indicates that, over a 24 h incubation period, t he reduction of As(V) to As(III) may account for 30% of the total As i n rabbit erythrocytes. As(V) present in the erythrocytes enters the ph osphate pool and depletes ATP. In comparison, about 65% of the total M MA(V) or about 44% of the total DMA(V) in the incubation system is tak en up by rabbit erythrocytes during a 24 h incubation. Neither organoA s species perturbed the Hb signals observed by spin-echo Fourier trans form NMR and the binding to GSH was minimal. Unlike As(V), MMA(V) and DMA(V) do not perturb phosphate metabolism, showing that, despite thei r pentavalent oxidation state, these arsenic species are not analogs f or phosphate.