Determination of trivalent methylated arsenicals in biological matrices

The enzymatically catalyzed oxidative methylation of As yields methylated a rsenicals that contain pentavalent As (As-V). Because trivalent As (As-III) is the favored substrate for this methyltransferase, methylated arsenicals containing As-V are reduced to trivalency in cells. Methylated arsenicals that contain As-III are extremely potent inhibitors of NADPH-dependent flav oprotein oxidoreductases and potent cytotoxins in many cell types. Therefor e, the formation of methylated arsenicals that contain As-III may be proper ly regarded as an activation step, rather than a means of detoxification. R ecognition of the role of methylated arsenicals that contain As-III in the toxicity and metabolism of As emphasizes the need for analytical methods to detect and quantify these species in biological samples. Hence, a method w as developed to exploit pH-dependent differences in the generation of arsin es from inorganic and methylated arsenicals that contain either As-V or As- III. Reduction with borohydride at pH 6 generated arsines from inorganic As -III, methyl As-III, and dimethyl As-III, but not from inorganic As-V, meth yl As-V, and dimethyl As-V. Reduction with borohydride at pH 2 or lower gen erated arsines from arsenicals that contained either As-V or As-III. Arsine s are trapped in a liquid nitrogen-cooled gas chromatographic trap, which i s subsequently warmed to allow separation of the hydrides by their boiling points. Atomic absorption spectrophotometry is used to detect and quantify the arsines. The detection limits (ng As ml(-1)) for inorganic As-III, meth yl As-III, and dimethyl As-III are 1.1, 1.2, and 6.5, respectively. This me thod has been applied to the analysis of arsenicals in water, human urine, and cultured cells. Both methyl As-III. and dimethyl As-III are detected in urine samples from individuals who chronically consumed inorganic As-conta minated water and in human cells exposed in vitro to inorganic As-III. The reliable quantitation of inorganic and methylated arsenicals that contain A s-III. in biological samples will aid the study of the toxicity of these sp ecies and may provide a new biomarker of the effects of chronic exposure to As. (C) 2001 Academic Press.