Urinary Dimethylarsinic Acid Research Articles

Early-Life Arsenic Exposure: Methylation Capacity and Beyond

Inorganic arsenic (iAs), a toxic metalloid, affects millions of people worldwide, mainly from drinking contaminated water. Arsenic is a human carcinogen that targets skin, lung, bladder, and possibly other sites. iAs and its methylated metabolites readily cross the placenta and reach the fetus [National Research Council (NRC) 1999], producing effects ranging from developmental toxicity to cancer (NRC 1999; Waalkes et al. 2007). Thus, early-life As exposures are drawing escalating health concerns. In this issue of Environmental Health Perspectives, Li et al. (2008) report on iAs metabolism in pregnant Bangladeshi women exposed to iAs through contaminated water. iAs is metabolized to monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA) for urinary excretion; urinary As speciation is widely used to assess As methylation capacity. Arsenic methylation capacity can be influenced by dietary intake of cysteine, methionine, folic acid, niacin, vitamin B12, and choline (Steinmaus et al. 2005), and dietary folic acid supplementation to malnourished arsenicosis patients can decrease As burden by decreasing blood MMA and increasing urinary DMA (Gamble et al. 2007). However, despite poor micronutrient status and high As exposure, the pregnant Bangladeshi women showed remarkably efficient As methylation. The median percentage of urinary DMA (74%) is in the upper range, and MMA (11%) in the lower range of what is commonly seen in urine of individuals from developed counties with much better nutrition (Vahter 2007). Women during childbearing years are more efficient at As methylation than men (Lindberg et al. 2007), particularly during pregnancy. This is likely due to the de novo synthesis of choline by the phosphatidylethanolamine methyltransferase (PEMT) pathway (Vahter 2007), which can be up-regulated by estrogen. Thus, the PEMT pathway may function in malnourished pregnant women to increase choline production needed for fetal development, and, perhaps fortuitously for As methylation (Vahter 2007). The remarkable efficiency of As methylation in malnourished pregnant women could also be an adaptive response to As exposure. Such adaptation might increase As methylation, and thereby excretion, during pregnancy, but perhaps at the expense of later toxicity. Methyl groups from S-adenosylmethionine are essential to both As and DNA methylation. DNA methylation status is a well-recognized controlling factor in gene expression. Furthermore, alterations in DNA methylation status are a recognized epigenetic mechanism in As carcinogenesis and are linked with As exposure in various systems (Pilsner et al. 2007; Waalkes et al. 2004). Gestation is a critical period of cell differentiation and genetic programming during development, and a highly sensitive time for initiation of chemical carcinogenesis (Waalkes et al. 2007). Thus, adaptation to As methylation in malnourished pregnant women could impact fetal development via altered gene expression. Activation of the PEMT pathway through estrogen signaling is clearly beneficial for As methylation (Vahter 2007). In contrast, aberrant estrogen signaling is linked with transplacental As carcinogenesis in mice (Waalkes et al. 2004, 2007). Arsenic also has potential synergistic effects with estrogenic carcinogens (Waalkes et al. 2007). Indeed, when in utero As exposure is followed by diethylstilbestrol treatment in newborn mice, tumors at multiple sites show synergistic increases in adulthood, including various estrogen sensitive targets (Waalkes et al. 2007). Recent data show that early-life As exposure has significant impact on human health (Fry et al. 2007; Smith et al. 2006). In newborns from mothers exposed to iAs through contaminated water in Thailand, altered transcript profiles in cord blood include stress-related genes and breast cancer/estrogen-signature genes (Fry et al. 2007), suggesting an adaptive response to As and pointing toward future health issues. Smith et al. (2006) observed a remarkable increase in lung cancer incidence in young adults in Chile who had been exposed to iAs in utero; this suggests a human transplacental carcinogenic potential for As. Thus, regardless of the immediate temporal tolerance to As during pregnancy, preventing early-life As exposure of the fetus and improving nutritional status of pregnant women may be critical for the health of future generations.

Clinical Manifestations and Arsenic Methylation after a Rare Subacute Arsenic Poisoning Accident

One hundred and four workers ingested excessive levels of arsenic in an accident caused by leakage of pipeline in a copper-smelting factory. Clinical examinations were performed by physicians in a local hospital. Excreted urinary arsenic species were determined by cold trap hydride generation atomic absorption spectrometry. In the initial toxic phase, gastrointestinal symptoms were predominant (83 people, 79.8%). Most patients showed leucopenia (72 people, 69.2%), and increased serum alanine aminotransferase (84 people, 80.8%) and aspartate aminotransferase (58 people, 55.8%). Thirty-five patients (33.6%) had elevated red blood cells in urine. After 17 days of admission, many subjects (45 people, 43.3%) developed peripheral neuropathy and 25 of these 45 patients (24.0%) showed a decrease in motor and sensory nerve conduction velocity. In the comparison of urinary arsenic metabolites among subacute arsenic-poisoned, chronic high arsenic-exposed and control subjects, we found that subacute arsenic-poisoned patients had significantly elevated proportions of urinary inorganic arsenic (iAs) and methylarsonic acid (MMA) but reduced proportion of urinary dimethylarsinic acid (DMA) compared with chronic high arsenic-exposed and control subjects. Chronic exposed subjects excreted higher proportions of iAs and MMA but lower proportions of DMA in urine compared with control subjects. These results suggest that gastrointestinal symptoms, leucopenia, and hepatic and urinary injury are predominant in the initial phase of subacute arsenic poisoning. Peripheral neuropathy is the most frequent manifestation after the initial phase. The biomethylation of arsenic decreases in a dose rate-dependent manner.

Nutritional status has marginal influence on the metabolism of inorganic arsenic in pregnant Bangladeshi women.

BackgroundThe interindividual variation in metabolism of inorganic arsenic (iAs), involving methylation via one-carbon metabolism, has been well documented, but the reasons remain unclear.ObjectivesIn this population-based study we aimed to elucidate the effect of nutrition on As methylation among women in Matlab, Bangladesh, where people are chronically exposed to iAs via drinking water.MethodsWe studied effects of macronutrient status using body mass index (BMI) among 442 women in early pregnancy (gestational week 8), and effects of micronutrient status (plasma folate, vitamin B12, zinc, ferritin, and selenium) among 753 women at gestational week 14. Arsenic metabolites in urine were measured by HPLC combined with hydride generation inductively coupled plasma mass spectrometry.ResultsThe median concentration of As in urine was 97 μg/L (range, 5–1,216 μg/L, adjusted by specific gravity). The average proportions of iAs, monomethylarsonic acid, and dimethylarsinic acid in urine in gestational week 8 were 15%, 11%, and 74%, respectively. Thus, the women had efficient As methylation in spite of being poorly nourished (one-third had BMIs < 18.5 kg/m2) and having elevated As exposure, both of which are known to decrease As methylation. The metabolism of iAs was only marginally influenced by micronutrient status, probably because women, especially in pregnancy and with low folate intake, have an efficient betaine-mediated remethylation of homocysteine, which is essential for an efficient As methylation.ConclusionsIn spite of the high As exposure and prevalent malnutrition, overall As methylation in women in early pregnancy was remarkably efficient. The As exposure level had the greatest impact on As methylation among the studied factors.

Developmental and genetic modulation of arsenic biotransformation: A gene by environment interaction?

The complexity of arsenic toxicology has confounded the identification of specific pathways of disease causation. One focal point of arsenic research is aimed at fully characterizing arsenic biotransformation in humans, a process that appears to be quite variable, producing a mixture of several arsenic species with greatly differing toxic potencies. In an effort to characterize genetic determinants of variability in arsenic biotransformation, a genetic association study of 135 subjects in western Sonora, Mexico was performed by testing 23 polymorphic sites in three arsenic biotransformation candidate genes. One gene, arsenic 3 methyltransferase (AS3MT), was strongly associated with the ratio of urinary dimethylarsinic acid to monomethylarsonic acid ( D/ M) in children (7–11 years) but not in adults (18–79 years). Subsequent analyses revealed that the high D/M values associated with variant AS3MT alleles were primarily due to lower levels of monomethylarsonic acid as percent of total urinary arsenic (%MMA5). In light of several reports of arsenic-induced disease being associated with relatively high %MMA5 levels, these findings raise the possibility that variant AS3MT individuals may suffer less risk from arsenic exposure than non-variant individuals. These analyses also provide evidence that, in this population, regardless of AS3MT variant status, children tend to have lower %MMA5 values than adults, suggesting that the global developmental regulation of arsenic biotransformation may interact with genetic variants in metabolic genes to result in novel genetic effects such as those in this report.

Folate, Homocysteine, and Arsenic Metabolism in Arsenic-Exposed Individuals in Bangladesh

Chronic exposure to arsenic is occurring throughout South and East Asia due to groundwater contamination of well water. Variability in susceptibility to arsenic toxicity may be related to nutritional status. Arsenic is methylated to monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA) via one-carbon metabolism, a biochemical pathway that is dependent on folate. The majority of one-carbon metabolism methylation reactions are devoted to biosynthesis of creatine, the precursor of creatinine. Our objectives of this cross-sectional study were to characterize the relationships among folate, cobalamin, homocysteine, and arsenic metabolism in Bangladeshi adults. Water arsenic, urinary arsenic, urinary creatinine, plasma folate, cobalamin, and homocysteine were assessed in 1,650 adults; urinary arsenic metabolites were analyzed for a subset of 300 individuals. The percentage of DMA in urine was positively associated with plasma folate (r = 0.14, p = 0.02) and negatively associated with total homocysteine (tHcys; r = −0.14, p = 0.01). Conversely, percent MMA was negatively associated with folate (r = −0.12, p = 0.04) and positively associated with tHcys (r = 0.21, p = 0.0002); percent inorganic arsenic (InAs) was negatively associated with folate (r = −0.12, p = 0.03). Urinary creatinine was positively correlated with percent DMA (r = 0.40 for males, p < 0.0001; 0.25 for females, p = 0.001), and with percent InAs (r = −0.45 for males, p < 0.0001; −0.20 for females, p = 0.01). Collectively, these data suggest that folate, tHcys, and other factors involved in one-carbon metabolism influence arsenic methylation. This may be particularly relevant in Bangladesh, where the prevalence of hyperhomocysteinemia is extremely high.

Intra-individual variation in the metabolism of inorganic arsenic.

Inorganic arsenic is metabolized to methylarsonic acid (MMA) and dimethylarsinic acid (DMA), which are excreted in the urine. Previous studies have shown a marked inter-individual variation in the metabolism, but the within-person variation is unknown. Therefore, we determined the variation in the relative amount of urinary arsenic metabolites, i.e., inorganic arsenic, MMA and DMA, over time in women living in an Andean village with elevated arsenic levels in drinking water. For our investigation of the individual day-to-day variation, daily spot urine samples were collected (at the same time of the day) for 5 consecutive days by 15 women. For our investigation of the within-day variation, seven women collected all the urine voided over a 72-h period, each voided into a separate container. Repeated measures analysis of variance (ANOVA) revealed no significant day-to-day variation, either in the relative distribution of inorganic arsenic metabolites (inorganic arsenic, MMA and DMA), or in the concentration of the metabolites. However, the percentage of DMA was slightly higher (on average 5.0%; P=0.003) and the percentage of inorganic arsenic lower (on average 5.8%; P=0.001) during the morning/day (03:00-15:00) compared with the evening/night (15:00-03:00). No within-day variation in the percentage of MMA was observed. The arsenic methylation efficiency of an individual is remarkably stable over time.

DMPS–Arsenic Challenge Test: II. Modulation of Arsenic Species, Including Monomethylarsonous Acid (MMAIII), Excreted in Human Urine

The administration of sodium 2,3-dimercapto-1-propane sulfonate (DMPS) to humans chronically exposed to inorganic arsenic in their drinking water resulted in the increased urinary excretion of arsenic, the appearance and identification of monomethylarsonous acid (MMAIII) in their urine, and a large decrease in the concentration and percentage of urinary dimethylarsinic acid (DMA). This is the first time that MMAIII has been detected in the urine. In vitro biochemical experiments were then designed and performed to understand the urinary appearance of MMAIII and decrease of DMA. The DMPS–MMAIII complex was not active as a substrate for the MMAIII methyltransferase. The experimental results support the hypothesis that DMPS competes with endogenous ligands for MMAIII, forming a DMPS–MMA complex that is readily excreted in the urine and points out the need for studying the biochemical toxicology of MMAIII. It should be emphasized that MMAIII was excreted in the urine only after DMPS administration. The results of these studies raise many questions about the potential central role of MMAIII in the toxicity of inorganic arsenic and to the potential involvement of MMAIII in the little-understood etiology of hyperkeratosis, hyperpigmentation, and cancer that can result from chronic inorganic arsenic exposure.

Genetic polymorphism in the biotransformation of inorganic arsenic and its role in toxicity

Arsenic is a recognized human carcinogen, but experimental cancer studies are negative. There is a variation in susceptibility among individuals, which probably is related to variation in metabolism. Inorganic arsenic is methylated to methylarsonic acid (MMA) and dimethylarsinic acid (DMA), which are less toxic and more readily excreted in urine than the inorganic arsenic. The rate of methylation of arsenic varies considerably between species. Most population groups studied so far have on average 10–30% inorganic, 10–20% MMA, and 60–70% DMA in urine, but there is a considerable inter-individual variation. Also, recent studies have identified groups with unusually low or high urinary excretion of MMA. Thus, there seems to be a genetic polymorphism in the biomethylation of arsenic. However, the methyltransferases involved in arsenic methylation have not been characterized.

Factors influencing arsenic methylation in humans

Most mammals metabolize inorganic arsenic to methylarsonic acid (MMA) and dimethylarsinic acid (DMA) by stepwise transfer of methyl groups from S-adenosylmethionine to arsenic in its trivalent oxidation state. The methylated metabolites are less reactive with tissue components, less toxic, and more readily excreted in the urine than is inorganic arsenic. There are marked differences in arsenic metabolism between mammalian species and between human population groups and individuals. We are studying arsenic metabolism in people of native Andean origin or mixed ethnic origin exposed to arsenic in drinking water (200 μg As/L) in northern Argentina. These people have a unique metabolism of arsenic in that they excrete only a few percent MMA in the urine. All other population groups studied so far have an average urinary metabolite distribution of inorganic arsenic, MMA, and DMA of about 20:15:65. However, the interindividual variation is extensive. This may indicate the existence of genetic polymorphism in the expression of arsenic methyltransferases. The toxicological significance of the observed differences in formation of MMA and DMA is not clear. In the populations studied, arsenic methylation was influenced by age, exposure level, and pregnancy. The relative percentage of urinary DMA was lower in children than in adults, which may indicate that children retain more arsenic in their tissues and, therefore, are more sensitive to arsenic exposure. In the children, there was an increase in % DMA with increasing exposure level, indicating an induction of the methylation of arsenic. Previous studies on adult populations in Chile and Mexico indicate a decrease in the relative amount of DMA in urine and a corresponding increase in inorganic arsenic and MMA, with increasing exposure levels. In the Argentinean population groups, arsenic was shown to be transferred to the fetus, in line with previously published data from experimental animal studies. There was a marked increase in the relative amount of DMA in urine at the end of pregnancy, as well as in the newborn infant, indicating induction of arsenic methylation during pregnancy. J.Trace Elem. Exp. Med. 13:173–184, 2000. © 2000 Wiley-Liss, Inc.

Determination of monomethylarsonic acid and dimethylarsinic acid in urine by liquid chromatography-tandem mass spectrometry.

A method for the simultaneous measurement of monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA) in human urine using liquid chromatography-tandem mass spectrometry with electrospray ionization (LC-ES-MS-MS) was developed. The multiple reaction monitoring mode (MRM) was used for quantitation. The protonated molecule ions (m/z 141.0 for MMA and m/z 139.0 for DMA) were selected as precursor ions, and the same fragment ion AsO+ (m/z 91.1) was monitored as the product ion. A two-step liquid-liquid extraction of MMA and DMA from urine provided recoveries of 92-100%. The coefficients of variation were lower than 7% for the within-day precision and lower than 11% for the between-day precision. The limit of quantitation was 25 microg/L as As for the two analytes. The assay was linear over the range of 25-800 microg/L.

URINARY LEVELS OF INORGANIC AND ORGANIC ARSENIC METABOLITES AMONG RESIDENTS IN AN ARSENIASIS-HYPERENDEMIC AREA IN TAIWAN

In order to elucidate whether urinary levels of inorganic and organic arsenic metabolites are associated with previous exposure to high-arsenic artesian well water, a total of 302 residents of age 30 yr or older were recruited from three arseniasis-hyperendemic villages in Taiwan. Most study subjects had stopped consuming high-arsenic artesian well water for more than 20 yr. The mean total arsenic (Ast) determined by inductively cout pled plasma mass spectrometer (ICPMS) was 267.05 +/- 20.95 mug/L, and the mean level of inorganic arsenic and its metabolites (Ast) was 86.08 +/- 3.43 mug/L. In the multivariate analysis, urinary dimethylarsinic acid (DMA) levels were significantly inversely associated with age, with women exhibiting significantly lower urinary amounts of arsenite [As(III)], arsenate [As(V)], monomethylarsonic acid (MMA), organic arsenic (Aso), and Ast compared to men. After adjustment for age and sex, previous cumulative arsenic exposure through consumption of artesian well water was significantly associated with elevated urinary levels of MMA and DMA, but not As(III) + As(V), Asot, and Ast. In the multivariate analysis, the percentage of As in As was significantly higher in men than women, but this was not significantly associated with age. The percentage of As(III) + As(V) in As increased significantly with age, while the reverse was noted with DMA in Asi. Women had a significantly higher DMA percentage but lower As(III) + As(V) and MMA percentages in Asi than men. After adjustment for age and sex, the percentages of As(III) + As(V) in Asi were significantly inversely associated with previous arsenic exposure through consumption of artesian well water. Data suggested that women seem to possess a more efficient arsenic methylation capability than men, and aging diminishes this methylation capability; furthermore, the higher the cumulative arsenic exposure, the greater is the body burden of inorganic arsenic, mainly in the form of MMA and DMA.

Arsenic methylation capacity, body retention, and null genotypes of glutathione S-transferase M1 and T1 among current arsenic-exposed residents in Taiwan.

In order to elucidate the relationships among arsenic methylation capacity, body retention, and genetic polymorphisms of glutathione S-transferase (GST) M1 and T1, a total of 115 study subjects were recruited from Lanyang Basin located on the northeast coast of Taiwan. Specimens of drinking water, blood, urine, hair and toenail were collected from each study subject. Urinary inorganic and methylated arsenic were speciated by high performance liquid chromatography combined with hydride-generation atomic absorption spectrometry. Arsenic concentration in hair and toenail were quantitated by atomic absorption spectrophotometry. The polymerase chain reaction was used to determine genetic polymorphisms of GST M1 and T1. Arsenic concentrations in urine, hair, and toenail of study subjects were positively correlated with arsenic levels in their drinking water. Percentages of various arsenic species in urine (mean +/- standard error (SE) were 11.8 +/- 1.0, 26.9 +/- 1.2 and 61.3 +/- 1.4, respectively, for inorganic arsenic, monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA). Men and women had similar arsenic methylation capability. No associations were observed between arsenic methylation capability and arsenic content in either drinking water or urine. Ratios of arsenic contents in hair and toenail to urinary arsenic content (mean +/- standard error) were 6.2 +/- 0.7 and 16.5 +/- 1.7, respectively. Genetic polymorphisms of GST M1 and T1 were significantly associated with arsenic methylation. Subjects having the null genotype of GST M1 had an increased percentage of inorganic arsenic in urine, while those with null genotype of GST T1 had an elevated percentage of DMA in urine. Arsenic contents in hair and toenail were significantly correlated with the increase in arsenic concentrations of drinking water and urine, while no significant associations were observed between arsenic contents in hair and toenail and polymorphisms of GST M1 and T1. The relationship between arsenic methylation capability and body retention was modified by genetic polymorphisms of GST M1 and T1. Arsenic contents in hair and toenail were negatively associated with MMA percentage and positively associated with DMA percentage among subjects having null genotypes of GST M1 and T1, but not among those with non-null genotypes.

Gas chromatographic determination of dimethylarsinic acid in aqueous samples

Procedures are described for the determination of dimethylarsinic acid in aqueous samples by gas chromatography of iododimethylarsine. Dimethylarsinic acid is converted into iododimethylarsine rapidly and quantitatively by treatment with hypophosphorous acid and potassium iodide, the resulting iododimethylarsine is extracted with toluene and determined with a gas chromatograph equipped with an electron-capture detector. The detection limit is 0.005 ppm as As. Recoveries from urine are over 95%. Other arsenicals do not give any response in the chromatography. The method has been applied to the determination of dimethylarsinic acid in urine and in water extracts of sea-weeds.

Confirmation of inorganic arsenic and dimethylarsinic acid in urine and plasma of dog by ion-exchange and TLC

Confirmation of inorganic arsenic and dimethylarsinic acid in urine and plasma of dog by ion-exchange and TLC