Trivalent arsenicals induce lipid peroxidation, protein carbonylation, and oxidative DNA damage in human urothelial cells

Drinking arsenic-contaminated water is associated with an increased risk of bladder cancer. Arsenate (iAs(V)), arsenite (iAs(III)), monomethylarsonous acid (MMA(III)), monomethylarsonic acid (MMA(V)), dimethylarsinous acid (DMA(III)), and dimethylarsinic acid (DMA(V)) have all been detected in the urine of people who drink arsenic-contaminated water. The aim of this research was to investigate which of these arsenicals are more hazardous to human urothelial cells. The results indicate that iAs(III), MMA(III), and DMA(III) were more potent in inducing cytotoxicity, lipid peroxidation, protein carbonylation, oxidative DNA damage, nitric oxide, superoxide, hydrogen peroxide, and cellular free iron than MMA(V), DMA(V), and iAs(V) in human urothelial carcinoma and transformed cells. However, the results did not show convincingly that the trivalent arsenicals were more potent than pentavalent arsenicals in decreasing the intracellular contents of total thiol, protein thiol, and reduced glutathione. Induction of oxidative DNA damage was observed with 0.2 microM of iAs(III), MMA(III), or DMA(III) as early as 1h. Because of its high oxidative damage, higher proportion in urine, and lower cytotoxicity, DMA(III) may be the most hazardous arsenical to human urothelial cells.     

Differential toxicity and accumulation of inorganic and methylated arsenic in rice

Background and aims

Efficient accumulation of arsenic (As) in rice (Oryza sativa L.) poses a potential health risk to rice consumers. The aim of this study was to investigate the mechanisms of uptake, transport and distribution of inorganic arsenic (As_i) and dimethylarsinic acid (DMA) in rice plants.

Methods

Rice was exposed to As_i (As(V)) and DMA in hydroponics. High-performance liquid chromatography inductively coupled plasma mass spectrometry (HPLC-ICP-MS) and synchrotron X-ray fluorescence (SXRF) microprobe were used to determine As concentration and the in situ As distribution.

Results

DMA induced abnormal florets before flowering and caused a sharp decline in the seed setting rate after flowering compared to As_i. Rice grains accumulated 2-fold higher DMA than As_i. The distribution of As_i concentration (root?>?leaf?>?husk?>?caryopsis) in As(V) treatments was different from that of the DMA concentration (caryopsis?>?husk?>?root?≥?leaf) in DMA treatments. SXRF showed that As_i mainly accumulated in the vascular trace of caryopsis with limited distribution to the endosperm, whereas DMA was observed in both tissues.

Conclusions

DMA tended to accumulate in caryopsis and induced higher toxicity to the reproductive tissues resulting in markedly reduced grain yield, whereas As_i mainly remained in the vegetative tissues and had no significant effect on yield. DMA is more toxic than As_i to the reproductive tissues when both of them are at similar concentrations in nutrient solution.     

Arsenic speciation in turnip as affected by application of chicken manure bearing roxarsone and its metabolites

Roxarsone (ROX) is widely used as a feed additive in intensive animal production. While animals are fed with ROX, the most commonly detectable As forms in fresh manures include ROX and small quantities of its metabolites such as arsenate (As(V)), arsenite (As(III)), monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA). A pot experiment was conducted to investigate the uptake, translocation and distribution of ROX, As(V), As(III), MMA and DMA in turnips, with the soil amended with 2% and 3% (w/w) chicken manure (CM) bearing ROX and its metabolites. Soil without any fertilizer was the control. The results show that only As(V) and As(III) were detected in turnip control samples. As(V), As(III) and DMA were found in all CM applied samples, but not ROX or MMA. This implies that turnip cannot take up ROX directly and accumulate MMA at detectable levels. The contents of DMA in tubers and the three As species in shoots increased with the CM rate in contrast to reduced levels of As(V) and As(III) in tubers. Increased CM rate enhanced the translocation of the three As species, especially for DMA, from tubers to shoots. DMA was the major form (42.9–61.4% in tubers and 38.1–76.3% in shoots), followed by As(III), in turnip plants fertilized with CM. The results indicate that ROX and its metabolites in animal manures can be introduced into human food chain by the way ROX → animal → manure → soil → crop.     

Arsenic biotransformation by Streptomyces sp. isolated from rice rhizosphere

Isolation and functional analysis of microbes mediating the methylation of arsenic (As) in paddy soils is important for understanding the origin of dimethylarsinic acid (DMA) in rice grains. Here, we isolated from the rice rhizosphere a unique bacterium responsible for As methylation. Strain GSRB54, which was isolated from the roots of rice plants grown in As‐contaminated paddy soil under anaerobic conditions, was classified into the genus Streptomyces by 16S ribosomal RNA sequencing. Sequence analysis of the arsenite S‐adenosylmethionine methyltransferase (arsM) gene revealed that GSRB54 arsM was phylogenetically different from known arsM genes in other bacteria. This strain produced DMA and monomethylarsonic acid when cultured in liquid medium containing arsenite [As(III)]. Heterologous expression of GSRB54 arsM in Escherichia coli promoted methylation of As(III) by converting it into DMA and trimethylarsine oxide. These results demonstrate that strain GSRB54 has a strong ability to methylate As. In addition, DMA was detected in the shoots of rice grown in liquid medium inoculated with GSRB54 and containing As(III). Since Streptomyces are generally aerobic bacteria, we speculate that strain GSRB54 inhabits the oxidative zone around roots of paddy rice and is associated with DMA accumulation in rice grains through As methylation in the rice rhizosphere.     

The Rice Aquaporin Lsi1 Mediates Uptake of Methylated Arsenic Species

Pentavalent methylated arsenic (As) species such as monomethylarsonic acid [MMA(V)] and dimethylarsinic acid [DMA(V)] are used as herbicides or pesticides, and can also be synthesized by soil microorganisms or algae through As methylation. The mechanism of MMA(V) and DMA(V) uptake remains unknown. Recent studies have shown that arsenite is taken up by rice (Oryza sativa) roots through two silicon transporters, Lsi1 (the aquaporin NIP2;1) and Lsi2 (an efflux carrier). Here we investigated whether these two transporters also mediate the uptake of MMA(V) and DMA(V). MMA(V) was partly reduced to trivalent MMA(III) in rice roots, but only MMA(V) was translocated to shoots. DMA(V) was stable in plants. The rice lsi1 mutant lost about 80% and 50% of the uptake capacity for MMA(V) and DMA(V), respectively, compared with the wild-type rice, whereas Lsi2 mutation had little effect. The short-term uptake kinetics of MMA(V) can be described by a Michaelis-Menten plus linear model, with the wild type having 3.5-fold higher V_max than the lsi1 mutant. The uptake kinetics of DMA(V) were linear with the slope being 2.8-fold higher in the wild type than the lsi1 mutant. Heterologous expression of Lsi1 in Xenopus laevis oocytes significantly increased the uptake of MMA(V) but not DMA(V), possibly because of a very limited uptake of the latter. Uptake of MMA(V) and DMA(V) by wild-type rice was increased as the pH of the medium decreased, consistent with an increasing proportion of the undissociated species. The results demonstrate that Lsi1 mediates the uptake of undissociated methylated As in rice roots.Arsenic (As) contamination affects millions of people worldwide, particularly in South Asia where As-contaminated groundwater has been extracted for drinking (Chakraborti et al., 2002; Nordstrom, 2002). Recent studies have shown that foods, especially rice (Oryza sativa), are an important source of inorganic As to populations dependent on a rice diet (Kile et al., 2007; Ohno et al., 2007; Mondal and Polya, 2008). Paddy rice is more efficient than other cereal crops in accumulating As (Williams et al., 2007). This is because anaerobic conditions in submerged paddy soils lead to mobilization of arsenite [As(III); Takahashi et al., 2004; Xu et al., 2008], which is then taken up by rice roots mainly through the highly efficient transport pathway for silicon (Si; Ma et al., 2008). The relatively high accumulation of As in rice is of concern, as it may pose a significant health risk (Zhu et al., 2008; Meharg et al., 2009).A number of As species may be present in soil depending on soil conditions and the history of As contamination. These include arsenate [As(V)], As(III), and methylated As species such as monomethylarsonic acid [MMA(V): CH₃AsO(OH)₂] and dimethylarsinic acid [DMA(V): (CH₃)₂AsO(OH)]. As(V) is the main species in aerobic soils, while As(III) dominates in anaerobic environments such as flooded paddy soils. Both MMA(V) and DMA(V) have been found in paddy soils (Takamatsu et al., 1982), which may have been derived from microbial and algal biomethylation and/or past uses of methylated As compounds. MMA(V), as sodium or calcium salt, and DMA(V), as sodium salt or free acid (also called cacodylic acid), are herbicides widely used for weed control on cotton (Gossypium hirsutum), orchards, and lawns, or as a defoliant of cotton (U.S. Environmental Protection Agency, 2006). Conversion of cotton fields for the production of paddy rice in the United States may be a reason for the high levels of methylated As reported for the U.S. rice (Meharg et al., 2009).The mechanism of As(V) uptake by plants through the phosphate transport system has been well established (for review, see Zhao et al., 2009). In contrast, As(III) is taken up into the cells by aquaglyceroporins in Escherichia coli, yeast (Saccharomyces cerevisiae), and mammalian tissues (for review, see Bhattacharjee and Rosen, 2007). Recent studies have shown that several plant aquaporin channels belonging to the Nodulin 26-like Intrinsic Protein (NIP) subfamily are permeable to As(III) when expressed heterologously in yeast (Bienert et al., 2008; Isayenkov and Maathuis, 2008; Ma et al., 2008). The rice Si transporter Lsi1 (OsNIP2;1; Ma et al., 2006) is also permeable to As(III) when expressed in yeast or Xenopus laevis oocytes (Ma et al., 2008). Furthermore, the lsi1 rice mutant lost 57% of the influx capacity for As(III) compared to the wild type in short-term assays, suggesting that Lsi1 is an important entry route for As(III) (Ma et al., 2008). In rice roots, a second Si transporter, Lsi2, functions as an efflux carrier to transport Si efflux from the exodermis and endodermis cells toward the stele for xylem loading (Ma et al., 2007). This transporter also mediates As(III) efflux; two independent lsi2 mutants had 73% to 91% lower concentrations of As(III) in the xylem sap than their wild types (Ma et al., 2008). The shared uptake pathway between Si (silicic acid) and As(III) (arsenous acid) is consistent with their physiochemical properties; both are present predominantly as undissociated neutral molecules at the normal environmental and physiological pH range (pK_a = 9.2, >99% undissociated at pH ≤ 7.0), and the two molecules have similar sizes.The uptake mechanisms of methylated As species by plant roots are not known. Previous studies showed that both MMA(V) and DMA(V) can be taken up by roots and translocated to shoots in a number of plant species (Marin et al., 1992; Carbonell-Barrachina et al., 1998, 1999; Burló et al., 1999). Marin et al. (1992) found that uptake by rice followed the order of As(III) > MMA(V) > As(V) > DMA (V), although DMA(V) was more efficiently translocated from roots to shoots than other As species. Raab et al. (2007) reported large variations in the absorption and translocation efficiencies for As(V), MMA(V), and DMA(V) among the 46 plant species tested. On average, root absorption of As(V) was 2.5- and 5-times higher than MMA(V) and DMA(V), respectively. The translocation efficiency, defined as the shoot-to-root concentration ratio after 24-h exposure, was highest for DMA(V) (0.8), followed by MMA(V) (0.3) and As(V) (0.09). The concentration-dependent uptake kinetics of MMA(V) in rice roots could be described by the Michaelis-Menten equation, whereas the limited uptake of DMA(V) appeared to be linear in relation to the increasing concentration in the uptake medium (Abedin et al., 2002). Abbas and Meharg (2008) showed that DMA(V) uptake by maize (Zea mays) seedlings was enhanced by more than 10-fold by a pretreatment of phosphorus starvation; this compared with only 2-fold increase in As(V) uptake. They thought that DMA(V) might be taken up by the phosphate transporters, or that phosphorus starvation altered expression of a range of membrane transporters or even membrane permeability itself.In addition to the root uptake of methylated As species, some plants appear to be able to biomethylate As, but the pathway and enzymology remains unclear (Wu et al., 2002; Zhao et al., 2009). In microbes, As methylation follows the Challenger pathway involving repeated steps of As reduction and oxidative methylation (Bentley and Chasteen, 2002). As(V) is first reduced to As(III), which is methylated by S-adenosylmethyltransferase using S-adenosyl-l-Met as the methyl donor. The product of this reaction is pentavalent MMA(V), which is reduced by a reductase to trivalent MMA(III) with thiols (e.g. glutathione). Methylation and reduction steps continue to produce di- and trimethyl As compounds. MMA(III) and DMA(III) are intermediates in the As methylation pathway, which is not very stable (Gong et al., 2001). In rice grain, DMA(V) is the main form of methylated As, and can account for up to 80% of the total As (Zavala et al., 2008; Meharg et al., 2009). In light of the significant presence of methylated As in rice, it is important to elucidate the transport and assimilation pathways of these As species in plants.In this study, we present evidence that MMA(V) and DMA(V) are taken up by rice roots, at least partly, through the NIP aquaporin channel Lsi1, and that this process is strongly pH dependent. We also show that MMA(V) can be reduced to MMA(III) in planta.     

Arsenic species uptake and translocation in Elodea canadensis

Abstract

The capacity of Elodea canadensis to phytofiltrate arsenic species from water was evaluated. Plants were adapted to tap water and supplemented with 15 and 250?µg L^?1 of As. Inorganic arsenic species (As III, As V), and organic arsenic compounds: monomethylarsonate (MMA) and dimethylarsinate (DMA) were analyzed. Sampling was carried out at different times after exposure in culture water and plant organs. Plants exposed to 15?µg L^?1 of As concentration showed no significant difference on As concentration (95% confidence level) in their organs compared to controls. When plants were exposed to 250?µg L^?1 of As concentration, a significant increase of As concentration in plant organs was observed. After 1?h exposure, plants reduce 63.16% the As concentration in the culture water, with a bioaccumulation factor (BF) of 4.3. Under these conditions, E. canadensis accumulate As V in roots and do not translocate it to stems (transfer factor <1). MMA was determined in stems and leaves. E. canadensis effectively phytofiltrate As from tap water of a city located in an arsenic endemic area from concentrations of 36?µg L^?1 to undetectable levels (10?ng L^?1).     

Root-microbial effects on plant iron uptake from siderophores and phytosiderophores

Collaborative experiments were conducted to determine whether microbial populations associated with plant roots may artifactually affect the rates of Fe uptake and translocation from microbial siderophores and phytosiderophores. Results showed nonaxenic maize to have 2 to 34-fold higher Fe-uptake rates than axenically grown plants when supplied with 1 μM Fe as either the microbial siderophore, ferrioxamine B (FOB), or the barley phytosiderophore, epi-hydroxymugineic acid (HMA). In experiments with nonsterile plants, inoculation of maize or oat seedlings with soil microorganisms and amendment of the hydroponic nutrient solutions with sucrose resulted in an 8-fold increase in FOB-mediated Fe-uptake rates by Fe-stressed maize and a 150-fold increase in FOB iron uptake rates by Fe-stressed oat, but had no effect on iron uptake by Fe-sufficient plants. Conversely, Fe-stressed maize and oat plants supplied with HMA showed decreased uptake and translocation in response to microbial inoculation and sucrose amendment. The ability of root-associated microorganisms to affect Fe-uptake rates from siderophores and phytosiderophores, even in short-term uptake experiments, indicates that microorganisms can be an unpredictable confounding factor in experiments examining mechanisms for utilization of microbial siderophores or phytosiderophores under nonsterile conditions.     

Individual variations in arsenic metabolism in Vietnamese: the association with arsenic exposure and GSTP1 genetic polymorphism

We investigated the association of As exposure and genetic polymorphism in glutathione S-transferase π1 (GSTP1) with As metabolism in 190 local residents from the As contaminated groundwater areas in the Red River Delta, Vietnam. Total As concentrations in groundwater ranged from <0.1 to 502 μg l(-1). Concentrations of dimethylarsinic acid (DMA(V)), monomethylarsonic acid (MMA(V)), and arsenite (As(III)) in human urine were positively correlated with total As levels in the groundwater, suggesting that people in these areas may be exposed to As through the groundwater. The concentration ratios of urinary As(III)/arsenate (As(V)) and MMA(V)/inorganic As (IA; As(III) + As(V))(M/I), which are indicators of As metabolism, increased with the urinary As level. Concentration and proportion of As(III) were high in the wild type of GSTP1 Ile105Val compared with the hetero type, and these trends were more pronounced in the higher As exposure group (>56 μg l(-1) creatinine in urine), but not in the lower exposure group. In the high As exposure group, As(III)/As(V) ratios in the urine of wild type of GSTP1 Ile105Val were significantly higher than those of the hetero type, while the opposite trend was observed for M/I. These results suggest that the excretion and metabolism of IA may depend on both the As exposure level and the GSTP1 Ile105Val genotype.     

Phloem transport of arsenic species from flag leaf to grain during grain filling

? Strategies to reduce arsenic (As) in rice grain, below concentrations that represent a serious human health concern, require that the mechanisms of As accumulation within grain be established. Therefore, retranslocation of As species from flag leaves into filling rice grain was investigated. ? Arsenic species were delivered through cut flag leaves during grain fill. Spatial unloading within grains was investigated using synchrotron X-ray fluorescence (SXRF) microtomography. Additionally, the effect of germanic acid (a silicic acid analog) on grain As accumulation in arsenite-treated panicles was examined. ? Dimethylarsinic acid (DMA) and monomethylarsonic acid (MMA) were extremely efficiently retranslocated from flag leaves to rice grain; arsenate was poorly retranslocated, and was rapidly reduced to arsenite within flag leaves; arsenite displayed no retranslocation. Within grains, DMA rapidly dispersed while MMA and inorganic As remained close to the entry point. Germanic acid addition did not affect grain As in arsenite-treated panicles. Three-dimensional SXRF microtomography gave further information on arsenite localization in the ovular vascular trace (OVT) of rice grains. ? These results demonstrate that inorganic As is poorly remobilized, while organic species are readily remobilized, from leaves to grain. Stem translocation of inorganic As may not rely solely on silicic acid transporters.     

Effects of arsenic exposure among semiconductor workers: a cautionary note on urinary 8-oxo-7,8-dihydro-2'-deoxyguanosine

Arsenic is a notorious environmental toxicant and was found to cause oxidative stress in cultured cells and animals. However, little work has been done in human studies, especially for the population occupationally exposed to arsenic. In order to investigate the effect of occupational exposure to arsenic in oxidative stress, we measured urinary 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) from 90 semiconductor workers including 50 exposed and 40 nonexposed subjects. A highly sensitive and specific isotope dilution LC-MS/MS method was used for quantification of 8-oxodGuo. The levels of inorganic arsenic (iAs3+, iAs5+), monomethylarsonic acid (MMA), and dimethylarsinic acid (DMA) in urine were determined by high-performance liquid chromatography-flow injection atomic absorption spectrometry (HPLC-FIAAS). Results showed that the mean urinary concentrations of total arsenic and 8-oxodGuo were significantly higher for exposed workers compared with the nonexposed workers. In addition, elevated urinary 8-oxodGuo concentrations of exposed workers were correlated with urinary levels of MMA (r = 0.44, P < 0.005) and the extent of primary methylation (the ratio of MMA to inorganic arsenic) (r = 0.40, P < 0.005). These findings suggested that occupational exposure to arsenic could result in the induction of oxidative stress. The presence and/or formation of MMA could play an important role in arsenic-involved injuries.     

好气条件下不同形态外源砷在土壤中的转化

在35%的田间持水量下,通过模拟试验研究了外源二甲基砷酸盐（DMA）、一甲基砷酸盐（MMA）、五价无机砷［As(V)］在土壤中的形态转化.结果表明: 外源砷进入土壤后,其含量均有随时间推延而逐渐下降的趋势,两种不同形态的有机砷DMA和MMA在土壤中主要发生脱甲基化过程,经150 d的恒温恒湿培养,其在土壤中主要转化为As(V),DMA处理仅在120 d时检测到少量MMA,而MMA处理则在7～60 d内均有少量的DMA生成.培养结束时土壤中DMA和MMA含量均显著降低(P<0.01),降幅分别为99.5%、94.3%,而两者的主要转化产物As(V)的含量则分别显著增加了4.61和5.15倍.表明外源有机态砷在土壤中基本上被转化为无机形态;与有机态外源砷相比,外源As(V)进入土壤后其形态基本上没有发生转化.     

淹水条件对土壤砷形态转化的影响

通过淹水条件下的培养试验, 探讨了外源二甲基砷酸（DMA）、一甲基砷酸（MMA）、砷酸盐［As(V)］在土壤中的动态转化规律. 结果表明: 随着培养时间的推移, 加入土壤中的DMA和MMA均主要转化为As(V), 且土壤中As(V)含量均呈增加趋势, 培养到150 d时土壤中As(V)含量均显著高于1 d时的含量(P<0.01). 外源DMA通过脱甲基化作用, 在30 d内即基本转化为As(V), 且有少量的亚砷酸盐［As(Ⅲ)］生成; 而外源MMA的转化速度相对较慢, 培养60 d后才基本完成向As(V)的转化, 同时伴随少量DMA和As(Ⅲ)的生成; 在淹水条件下外源As(V)含量随培养时间的增加而逐渐降低,该过程中除有少量As(Ⅲ)生成外,其形态基本未发生改变.     

Water management,rice varieties and mycorrhizal inoculation influence arsenic concentration and speciation in rice grains

A pot experiment was carried out to investigate the effects of water management and mycorrhizal inoculation on arsenic (As) uptake by two rice varieties, the As-resistant BRRI dhan 47 (B47) and As-sensitive BRRI dhan 29 (B29). Grain As concentration of B47 plants was significantly lower than that of B29, and grain As concentration of B47 was higher under flooding conditions than that under aerobic conditions. In general, mycorrhizal inoculation (Rhizophagus irregularis) had no significant effect on grain As concentrations, but decreased the proportion of inorganic arsenic (iAs) in grains of B47. The proportion of dimethylarsinic acid (DMA) in the total grain As was dramatically higher under flooding conditions. Results demonstrate that rice variety selection and appropriate water management along with mycorrhizal inoculation could be practical countermeasures to As accumulation and toxicity in rice grains, thus reducing health risks of As exposure in rice diets.     

Determination of trivalent methylated arsenicals in rat urine by liquid chromatography-inductively coupled plasma mass spectrometry after solvent extraction

A method for the determination of trivalent arsenicals in urine was examined. Trivalent arsenicals, extracted as complexes with diethylammonium diethyldithiocarbamate (DDDC) into carbon tetrachloride, were determined by liquid chromatography-inductively coupled plasma mass spectrometry (LC-ICP-MS). The trivalent methylated arsenicals monomethylarsonous acid (MMA(III)), dimethylarsinous acid (DMA(III)), and trimethylarsine (TMA) were detected in urine of rats that had received dimethylarsinic acid (DMA(V)) or monomethylarsonic acid (MMA(V)) at concentration of 200 microg ml(-1) in drinking water for 24 weeks. This method is the first to permit quantification of trivalent methylated arsenicals in urine without significant changes in concentration during storage or pretreatment.     

Uptake Kinetics of Arsenic in Upland Rice Cultivar Zhonghan 221 Inoculated with Arbuscular Mycorrhizal Fungi

Arbuscular mycorrhizal fungi (AMF) appear to be highly associated with arsenic (As) uptake in host plants because arsenate (As(V)) and phosphorus (P) share the same transporter, whereby AMF can enhance P uptake. A short-term experiment was conducted for low- (0 to 0.05 mM As) and high-affinity (0 to 2.5 mM As) uptake systems, to investigate the AMF role on As uptake mechanism in plants, which may explain As uptake kinetics in upland rice cultivar: Zhonghan 221. When concentration of As ranged from 0 to 0.05 mM, Funneliformis geosporum (Fg) significantly decreased arsenite (As(III)) and monomethylarsonicacid (MMA) uptake when (p < 0.05) compared to non-mycorrhizal (NM) treatment, since the major route for (As(III)) in rice roots—rice silicon transporter Lsi1 would be influenced by Fg inoculation at high As concentrations. Fg can also reduce As(V) uptake significantly (p < 0.05) under both uptake systems relative to NM treatment, whereas, Funneliformis mosseae (Fm) increased As(V) and MMA uptake in rice roots, with MMA uptake rate generally lower than As(III) and As(V). Using suitable AMF species inoculation with rice, As uptake and accumulation in rice grains can be reduced and the risk to human health, once consumed, can be minimized.     

Method for the determination of five toxicologically relevant arsenic species in human urine by liquid chromatography-hydride generation atomic absorption spectrometry

An analytical method for the simultaneous quantitation of arseneous acid (As(III)), arsenic acid (As(V)), monomethylarsonic acid (MMA), dimethylarsinic acid (DMA) and trimethylarsine oxide (TMAO) in human urine by coupling of high-performance liquid chromatography with hydride generation atomic absorption spectrometry (HPLC/HG-AAS) via a flow-injection interface is presented. After arsenic species separation by anion-exchange displacement chromatography the compounds are on-line reduced to their corresponding hydrides and detected by atomic absorption spectrometry. Detection limits range from 1.1 (TMAO) to 2.6 microg/L (As(V)). The method has been applied to determine arsenic species in the urine of a volunteer before and after consumption of seafood as well as to analyse certified reference urine samples for their arsenic species content.     

Alternative splicing variants of human arsenic (+3 oxidation state) methyltransferase

Arsenic (+3 oxidation state) methyltransferase (As3MT) catalyzes the methylation of trivalent arsenic (As(III)) to monomethylarsonate (MMA(V)) and dimethylarsinic acid (DMA(V)), and plays an important role in the detoxification of arsenicals. Here, we report the identification of two splicing variants of the human As3MT gene. One splicing variant was an exon-3 skipping (Δ3) form which produced a premature stop codon, and the other was an exon-4 and -5 skipping (Δ4,5) form which produced a 31.1 kDa As3MT protein. In addition to the full-length mRNA of As3MT, Δ4,5 mRNAs were detected in HepG2, A549, HL60, K562, and HEK293 cells. The methyltransferase activity of the recombinant Δ4,5 As3MT and wild-type As3MT proteins purified from Escherichia coli was determined. Speciation analysis by HPLC–ICP-MS showed a clear peak of MMA(V) after incubation of As(III) with the wild-type As3MT protein, but not with the Δ4,5 As3MT protein. In addition, COS-7 cells transfected with Δ4,5 As3MT cDNA did not convert As(III) to MMA(V) or DMA(V). The lack of methyltransferase activity of Δ4,5 As3MT seems to be related to the deletion of an S-adenosylmethionine-binding site and a critical cysteine residue. These data suggest that the expression pattern of splicing variants of the As3MT gene may affect the capacity for arsenic methylation in cells.     

Biological and Microcalorimetric Studies of the Toxic Effect of Organoarsenic(V) Compounds to Wild Strain of <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis>

Microcalorimetric and biological methods were carried out to determine the toxicity of dimethylarsinic acid (DMA) and monomethylarsonic acid (MMA) to wild strain of Bacillus thuringiensis. Thermokinetic parameters were obtained from the power–time curves, showing that the peak-heat output power, total heat output, and number of colonies decreased with the increases in concentration of DMA and MMA. In addition, the generation time and peak maximal time increased with the increases in the dosage of DMA and MMA. The half inhibitory concentrations of DMA and MMA were 99.02 and 142.02 μg mL⁻¹, respectively for the wild strain of B. thuringiensis. DMA shows higher toxicity to bacteria than MMA. The toxicity resistance of B. thuringiensis against organoarsenic(V) is quite high for the wild strain. Our work demonstrates that microcalorimetry is a very sensitive, simple, and useful technique for in vitro investigation of the toxic effect of organoarsenic(V) on microbial activity.     

The response of tomato (Lycopersicon esculentum) to different concentrations of inorganic and organic compounds of arsenic

Tomato plants were cultivated in greenhouse and water solutions of arsenite (As(III)), arsenate (As(V)), methylarsonic acid (MA) and dimethylarsinic acid (DMA) were applied individually into cultivation substrate at two As levels, 5 and 15 mg kg⁻¹ of the substrate. Comparing the availability of arsenic compounds increased in order arsenite = arsenate < MA < DMA where the arsenic contents in plants decreased during vegetation period. Within a single plant, the highest arsenic concentration was found in roots followed in decreasing order by leaves, stems, and fruits regardless of arsenic compound applied. Arsenic toxicity symptoms reflected in suppressed growth of plants and a lower number and size of fruits were most significant with DMA treatment. However, the highest accumulation of arsenic by plants growing in the soil containing DMA was caused by higher mobility of this compound in the soil due to its lower sorption affinity. Our results confirmed substantial role of transformation processes of arsenic compounds in soil in uptake and accumulation of arsenic by plants.