High Performance Liquid Chromatography Coupled To Inductively Coupled Plasma Mass Spectrometry Research Articles

Nitrate reductase involves in selenite reduction in Rahnella aquatilis HX2 and the characterization and anticancer activity of the biogenic selenium nanoparticles

BackgroundBiogenic selenium nanoparticles (SeNPs) show numerous advantages including their high stability, low toxicity, and high bioactivity. While metabolism of SeNPs remains not well studied and need more investigation to reveal the process. PurposeThe objective of the study was to investigate the relationship between nitrate reductase and selenite reduction in Rahnella aquatilis HX2, characterize the properties of HX2 produced SeNPs, and explore their potential applications, particularly their anticancer activity. ProceduresSelenium species were measured by high-performance liquid chromatography coupled to inductively coupled plasma - Mass spectrometry (HPLC-ICP-MS). Transcription level of nitrate reductase was determined by Real-time quantitative PCR. Morphology, particle size, crystal structure and surface chemistry of SeNPs were determined by electron microscopy, dynamic light scattering method, Raman scattering, X-ray photoelectron spectroscopy, respectively. Anti cancer cell activity was measured by CCK-8 assay. Main findingsSeNP production in R. aquatilis HX2 was correlated with the cell growth. The products of selenite reduction in HX2 detected by HPLC-ICP-MS included SeNPs, selenocysteine (SeCys), Se-Methylselenocysteine (MeSeCys), and 7 unknown compounds. Nitrate addition experiments suggested the involvement of nitrate reductase in selenite reduction in HX2. Both the cellular membrane and cytoplasm of HX2 exhibited selenite-reducing ability, indicating that membrane-associated nitrate reductase was not the sole selenite reductase in HX2. Characterization of the biogenic SeNPs revealed a spherical morphology and amorphous structure of them. Surface chemistry analysis implicated the binding of extracellular polymeric substances to the biogenic SeNPs, and the presence of Se0, Se2-, and electron-rich Se atoms on the surface of SeNPs. Finally, the IC50 values of the biogenic SeNPs were 36.49 μM for HepG2 and 3.70 μM for HeLa cells. ConclusionsThe study first revealed that the nitrate reductase is involving in selenite reduction in R. aquatilis HX2. The biogenic SeNPs coordinated with organic substances in the surface. And SeNPs produced by R. aquatilis HX2 showed excellent anticancer activities on HepG2 and HeLa cells.

Arsenic speciation and arsenic feed-to-fish transfer in Atlantic salmon fed marine low trophic feeds based blue mussel and kelp

BackgroundAquaculture aims to reduce the environmental and climate footprints of feed production. Consequently, low trophic marine (LTM) resources such as blue mussels and kelp are potential candidates to be used as ingredients in salmon feed. It is relevant to study potential undesirables associated with their use, as well as assessing food safety by investigating their transfer from feed-to-fish. The marine biota is well known to contain relatively high levels of arsenic (As), which may be present in different organic forms depending on marine biota type and trophic position. Thus, it is important to not only obtain data on the concentrations of As, but also on the As species present in the raw materials, feed and farmed salmon when being fed novel LTM feed resources. MethodsAtlantic salmon were fed experimental diets for 70 days. A total of nine diets were prepared: four diets containing up to 4 % fermented kelp, three diets containing up to 11 % blue mussel silage, and one diet containing 12 % blue mussel meal, in addition to a standard reference diet containing 25 % fish meal. Concentrations of As and As species in feeds, faeces, liver and fillet of Atlantic salmon were determined by inductively coupled plasma mass spectrometry (ICP-MS) and high-performance liquid chromatography coupled to ICP-MS (HPLC-ICP-MS), respectively. ResultsThe use of kelp or blue mussel-based feed ingredients increased the concentration of total As, but maximum level as defined in Directive 2002/32 EC and amendments was not exceeded. The concentrations found in the experimental feeds ranged from 3.4 mg kg−1 to 4.6 mg kg−1 ww. Arsenic speciation in the feed varied based on the ingredient, with arsenobetaine dominating in all feed samples (36–60 % of the total As), while arsenosugars (5.2–8.9 % of the total As) were abundant in kelp-included feed. The intestinal uptake of total As ranged from 67 % to 83 %, but retention in fillet only ranged from 2 % to 22 % and in liver from 0.3 % to 0.6 %, depending on the marine source used. Fish fed feeds containing blue mussel showed higher intestinal uptake of total As when compared with fish fed feeds containing fermented kelp. Fish fed fermented kelp-based feeds had higher retained concentrations of total As when comparing with fish fed feeds containing blue mussel. Despite relatively high intestinal uptake of total As, inorganic and organic As, the retained concentrations of As did not reflect the same trend. ConclusionAlthough the use of LTM feed ingredients increased the level of total As in this feeds, salmon reared on these diets did not show increased total As levels. The well-known toxic inorganic As forms were not detected in salmon muscle reared on LTM diets, and the non-toxic organic AsB was the dominant As species that was retained in salmon muscle, while the organic AsSug forms were not. This study shows that speciation analysis of the LTM resources provides valuable information of the feed-to-fish transfer of As, needed to assess the food safety of farmed Atlantic salmon reared on novel low trophic feeds.

Arsenic species in mesopelagic organisms and their fate during aquafeed processing.

A responsible harvest of mesopelagic species as aquafeed ingredients has the potential to address the United Nations Sustainable Development Goal 14, which calls for sustainable use of marine resources. Prior to utilization, the levels of undesirable substances need to be examined, and earlier studies on mesopelagic species have reported on total arsenic (As) content. However, the total As content does not give a complete basis for risk assessment since As can occur in different chemical species with varying toxicity. In this work, As speciation was conducted in single-species samples of the five most abundant mesopelagic organisms in Norwegian fjords. In addition, As species were studied in mesopelagic mixed biomass and in the resulting oil and meal feed ingredients after lab-scale feed processing. Water-soluble As species were determined based on ion-exchange high-performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP-MS). This was supplemented by extracting arsenolipids (AsLipids) and determining total As in this fraction. The non-toxic arsenobetaine (AB) was the dominant form in mesopelagic crustaceans and fish species, accounting for approximately 70% and 50% of total As, respectively. Other water-soluble species were present in minor fractions, including carcinogenic inorganic As, which, in most samples, was below limit of quantification. The fish species had a higher proportion of AsLipids, approximately 35% of total As, compared to crustaceans which contained 20% on average. The feed processing simulation revealed generally low levels of water-soluble As species besides AB, but considerable fractions of potentially toxic AsLipids were found in the biomass, and transferred to the mesopelagic meal and oil. This study is the first to report occurrence data of at least 12 As species in mesopelagic organisms, thereby providing valuable information for future risk assessments on the feasibility of harnessing mesopelagic biomass as feed ingredients.

A fit-for-purpose copper speciation method for the determination of exchangeable copper relevant to Wilson’s disease

Exchangeable copper (CuEXC), mainly comprised copper (Cu) bound to albumin, has been proposed as a specific marker of Cu overload in Wilson's disease (WD). To the author's knowledge, there are no methods capable of determining reliably CuEXC to meet the requirements and challenges faced by a clinical trial. The present work describes a novel speciation strategy for the determination of the main Cu-species in human serum by anion-exchange high-performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP-MS). A label-free protein quantification approach was conducted where the concentration of Cu associated to the protein fraction was based on its relative peak area distribution and the total Cu concentration in the sample. Such a methodology was characterized in terms of selectivity, sensitivity, precision, and robustness. Due to the lack of speciated Cu-reference materials, protein recovery was assessed by comparison with that of species-specific (SS) isotope dilution (ID). For this, a double SS HPLC-ICP-IDMS method for Cu-albumin was developed and presented here for the first time. Three human sera (two frozen LGC8211 and ERM®-DA250a, and the lyophilised Seronorm™ Human) were analyzed using both the relative and ID quantification methods. The validated relative approach, with relative expanded uncertainties (k = 2) between 5.7 and 10.1% for Cu-albumin concentrations ranging from 112 to 455 μg kg-1 Cu, was found to be able to discriminate between healthy and WD populations in terms of Cu-albumin content. Also, using such methodology, underestimation of CuEXC by the classical EDTA/ultrafiltration method was demonstrated. The methodology developed in this work will be invaluable for quality control assessment and WD drug monitoring. This work describes a Cu-protein quantification approach for the determination of exchangeable Cu relevant to Wilson's Disease.

Simultaneous quantitative speciation of selected toxic elements in water using high performance liquid chromatography coupled to inductively coupled plasma-mass spectrometry (HPLC-ICP-MS)

The aim of this study was to conduct multi-elemental speciation of As(III), dimethylarsinic acid (DMA), monomethylarsenic acid (MMA), As(V) and Cr(VI) in water samples using high performance liquid chromatography coupled to inductively coupled plasma-mass spectrometry (HPLC-ICP-MS) in a single analytical run. Chromatographic parameters, spectral interference identification, and elimination and the performance characteristics of the method were successfully optimized for quantification of As(III), DMA, MMA, As(V), Cr(VI) and Cr(III) in water samples. The percentage errors of 6.93–43.2% for identified potential interferences were eliminated by applying the inter-element mathematical correction method. The limit of detection (LOD) and limit of quantification (LOQ) of As species and Cr(VI) were varied from 0.040-0.298 μg L−1 and 0.133–2.98 μg L−1, respectively. The LOD and LOQ of total As, Cr and Pb determined using ICP-MS were ranged from 0.013-0.203 μg L−1 and 0.045–0.675 μg L−1, respectively. The accuracy of the method was confirmed by achieving the percentage recovery of 83.4% for As species, 90.2% for Cr(VI) and 110–118% for total concentrations of As, Cr and Pb using standard reference material for trace elements in water (SRM 1643f). Furthermore, the accuracy of the speciation analysis method was checked using the spiking and recovery studies and yielded percentage recoveries in the range of 78.3–95.4% for the analyte species. The quantities of species determined in water samples were in the range of 0.314–0.402 μg L−1 for As(III), 0.168–0.533 μg L−1 for DMA, 0.226–0.312 μg L−1 for MMA, 0.191–0.459 μg L−1 for As(V), 0.500–3.28 μg L−1 for Cr(VI) and 0.390–12.6 μg L−1 for Cr(III) in high-flow seasonal state of the river. In the low-flow seasonal state, concentrations varied from 0.314-0.391 μg L−1 for As(III), <0.094 μg L−1 for DMA, 0.315–26.7 μg L−1 for MMA, 0.290–0.744 μg L−1 for As(V), 0.426–5.42 μg L−1 for Cr(VI) and 0.424–96.8 μg L−1 Cr(III). Total concentrations of As, Cr and Pb in water samples of high-flow seasonal state ranged from 0.949-1.88 μg L−1, 1.08–14.9 μg L−1 and 0.190–33.8 μg L−1, respectively. In low-flow seasonal state, the total concentrations of As, Cr and Pb in water samples varied from 1.13-26.5 μg L−1, 3.28–97.5 μg L−1 and 0.455–0.610 μg L−1, respectively. The variations between analytes species and total As, Cr and Pb in seasonal states of the river were statistically insignificant (p > 0.05) at a 95% confidence level except for DMA. The nonpoint sources pollution such as atmospheric deposition of dust released from coal mining, power stations, sewage effluents discharge and agricultural activities are presumably the sources of potentially toxic elements in the Mokolo River.

The bioavailability of arsenic species in rice.

Rice is the principal food in many countries for billions of people and one of the most consumed cereals in the world. The rice plant has the ability to bioaccumulate essential and toxic trace elements such as arsenic. The toxicity of the elements depends not only on their concentration but also on their chemical form and their bioavailability. The inorganic forms of arsenic are more toxic than the organic forms and the toxicity increases with decreasing oxidation states. The consumers of rice in Europe who are the most exposed to inorganic arsenic are children under three, thorough diet (rice-based food). Recently, the European Commission established the maximum levels of inorganic arsenic in foodstuffs. This regulation establishes a maximum level of inorganic arsenic of 100μgkg-1 in rice destined for the production of food for infants and young children. In order to know the relation between the As ingested and the arsenic absorbed, studies of bioavailability are necessary. We proposed an in vitro digestion method with dialysis to estimate this relation. Furthermore, a bioavailability study of As species in rice was performed in order to know if a change in As species occurred during the gastrointestinal digestion process. Arsenic species were determined in rice and in the dialysate fraction by high-performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP-MS). The proposed method has been applied to different rice samples acquired in the local Spanish market. Graphical abstract.

Pharmacokinetic Properties of Arsenic Species after Intravenous and Intragastrical Administration of Arsenic Trioxide Solution in Cynomolgus Macaques Using HPLC-ICP-MS.

A rapid and sensitive method was established for arsenic (As) speciation based on high performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP-MS). This method was validated for the quantification of four arsenic species, including arsenite (AsIII), arsenate (AsV), monomethylarsonic acid (MMAV) and dimethylarsinic acid (DMAV) in cynomolgus macaque plasma. Separation was achieved in just 3.7 min with an alkyl reverse phase column and highly aqueous mobile phase containing 20 mM citric acid and 5 mM sodium hexanesulfonate (pH = 4.3). The calibration curves were linear over the range of 5–500 ng·mL−1 (measured as As), with r > 0.99. The above method was validated for selectivity, precision, accuracy, matrix effect, recovery, carryover effect and stability, and applied in a comparative pharmacokinetic study of arsenic species in cynomolgus macaque samples following intravenous and intragastrical administration of arsenic trioxide solution (0.80 mg·kg−1; 0.61 mg·kg−1 of arsenic); in addition, the absolute oral bioavailability of the active ingredient AsIII of arsenic trioxide in cynomolgus macaque samples was derived as 60.9 ± 16.1%.

Determination of hexavalent Cr in river sediments by speciated isotope dilution inductively coupled plasma mass spectrometry

During a sampling campaign in September 2015, elevated chromium (Cr) concentrations were observed in sediments of industrially exposed sites of the Sava River (stainless steel production and impact of metallurgic industry). To verify if sediments also contained hazardous hexavalent Cr (Cr(VI)), there was a need to develop a sensitive and reliable analytical procedure for its determination. In the determination of Cr(VI) in environmental samples, it is necessary to evaluate the suitability of the applied analytical methodology for each individual sample matrix. In these studies, the use of isotopically enriched Cr tracers importantly contribute to the validity of the obtained results. In the present work, an analytical procedure was optimised for the extraction of total Cr(VI) from sediments and the content of Cr(VI) determined by high performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP-MS). To leach the total amount of Cr(VI) from sediments, an ultrasound-assisted extraction procedure was applied at 80 °C, using 2% NaOH + 3% Na2CO3 as the extraction solution. The addition of 0.4 mol L−1 MgCl2 prevented oxidation of Cr(III) during the extraction step. To control for species interconversion and for an accurate calculation of Cr(VI) concentration by speciated isotope dilution (ID)-ICP-MS, the alkaline extract was doubly spiked with enriched 50Cr(VI) and 53Cr(III). The accuracy of the determination of Cr(VI) was verified by analysing the certified reference material CRM 041 (Cr(VI) in soil), and by spike recovery test. Low limits of detection (LOD) and quantification (LOQ) (1.25 and 4.0 μg Cr(VI) kg−1, respectively) and good repeatability of measurement (relative standard deviation better than ±4.8%) were obtained. The analytical data revealed that Cr(VI) concentrations in sediments of the Sava River did not represent any known environmental hazard.

Simultaneous determination and speciation analysis of arsenic and chromium in iron supplements used for iron-deficiency anemia treatment by HPLC-ICP-MS

This work proposes the use of high performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP-MS) for simultaneous speciation of arsenic and chromium in iron supplements used for the treatment of anemia. The sample preparation procedure recommended for the total determination of arsenic and chromium was established using acid digestion in a microwave assisted oven. For speciation analysis, however, the microwave-assisted extraction procedure involved the use of water as extraction solvent at 90°C for 30min. The chromatographic separation was performed using a mobile phase containing 1.0mM tetrabutylammonium hydroxide (TBAH), 0.7mM ethylenediaminetetraacetic acid (EDTA) and 5% methanol at pH 7.2. Helium was used in the collision cell for elimination of the interferences. Under optimized conditions, the separation and detection of the As(III), As(V), Cr(III) and Cr(VI) species can be performed in 5min, permitting their quantification with the external calibration technique with standards prepared in the mobile phase. The limits of quantification obtained were 0.008, 0.010, 0.5 and 0.14µgg−1, for As(III), As(V), Cr(III) and Cr(VI), respectively. The accuracy of the method was evaluated and confirmed by addition/recovery tests. The recoveries obtained varied from 81% to 110%. The proposed method was applied to the speciation analysis of arsenic and chromium in commercially available iron supplements acquired in several cities in Brazil and Spain. The content of the species ranged from 0.01 to 1.3µgg−1 for arsenic, and from 0.4 to 61.2µgg−1 for chromium.

A field deployable method for a rapid screening analysis of inorganic arsenic in seaweed

Inorganic arsenic (iAs) in 13 store-bought edible seaweed samples and 34 dried kelp (Laminaria digitata) samples was determined by a newly developed, field-deployable method (FDM) with the aid of a field test kit for arsenic in water. Results from the FDM were compared to results from speciation analysis achieved by using high performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP-MS). The FDM consisted of a simple extraction method using diluted HNO3 to quantitatively extract iAs without decomposing the organoarsenicals to iAs followed by the selective volatilisation of iAs as arsine (AsH3) and subsequent chemo-trapping on a filter paper soaked in mercury bromide (HgBr2) solution. Method optimization with a sub-set of samples showed 80–94% iAs recovery with the FDM with no matrix effect from organo-arsenic species in the form of dimethylarsinic acid (DMA) on the iAs concentration. The method displayed good reproducibility with an average error of ±19% and validation by HPLC-ICP-MS showed that the results from the FDM were comparable (slope = 1.03, R2 = 0.70) to those from speciation analysis with no bias. The FDM can be conducted within an hour and the observed limit of quantification was around 0.05 mg kg−1 (dry weight). This method is well suited for on-site monitoring of iAs in seaweed before it is harvested and can thus be recommended for use as a screening method for iAs in seaweed.Graphical abstractScreening seaweed for their inorganic arsenic concentration within one hour without bias has been made possible in the field by using a field deployable arsenic kit. Its accuracy and precision was compared to HPLC-ICPMS.

Arsenic distribution and speciation in Daphnia pulex

Rat Lake, Yellowknife, Northwest Territories, is situated on arsenic-rich tailings from a historical gold mine. The abundant zooplankton species Daphnia pulex in this lake was used to study the impact of arsenic at the base of the freshwater food web; the speciation and distribution of arsenic in D. pulex and its food sources; and the origin of formation of organoarsenicals in freshwater systems. The arsenic concentration in lake water was measured as 0.25mgL−1, while the zooplankton organisms contained up to 35mgkg−1 d.w. arsenic. Plankton samples were analyzed for arsenic speciation, by using X-ray Absorption Near Edge Structure (XANES) on the whole, dried samples and High Performance Liquid Chromatography coupled to Inductively Coupled Plasma Mass Spectrometry (HPLC-ICP-MS) on water extracts. XANES data suggest that D. pulex mainly contain inorganic arsenicals with 56% of arsenic with +5 oxidation state and 10% of arsenic with +3 oxidation state, but also 34% of organoarsenic compounds that were identified with HPLC-ICP-MS as monomethylarsonate (MMA), dimethylarsinate (DMA), and arsenosugars. The most abundant of the organoarsenicals was the glycerol sugar (Sugar 1). X-ray Fluorescence (XRF) mapping of D. pulex for arsenic distribution showed that arsenic was mainly distributed in the gut of the animal, where its concentration was ten times higher than in the surrounding tissues. Moreover, the analysis of residues from extractions targeting water-soluble and lipid-soluble arsenicals suggested that part of the measured arsenic signal comes from ingested sediments, phytoplankton, or other food sources. These food sources contain inorganic arsenic only, with AsV–O in phytoplankton and AsIII–S in sediments, suggesting the possibility that the organoarsenicals compounds detected in the tissues of the organism are created by the Daphnia.

Comparison of Two Analytical Methods for the Analysis of Methylmercury in Fish

In this study, we compared a high performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP-MS) method and an isotope dilution gas chromatography mass spectrometry (ID-GC-MS) method for the analysis of methylmercury (MeHg) in fish samples. Using HPLC-ICP-MS, the limit of detection (LOD) was 2.0 ngHg · g−1 for inorganic Hg and was 5.0 ngHg · g−1 for MeHg. In comparison, the LOD for MeHg was 3.4 ngHg · g−1 for using ID-GC-MS. Certified reference material (CRM) results agreed well with certified values for both methods. The mean recovery for analysis of CRM BCR 463 by HPLC-ICP-MS was 99 ± 2.1 % for MeHg and was 103 ± 2.1 % for total Hg. Two CRMs, BCR 463 and CRM DORM2, were analyzed by ID-GC-MS, the mean recovery of MeHg was 87 ± 1.9 % for BCR 463, and the mean recovery of MeHg was 102 ± 2.9 % for CRM DORM2. Matrix samples spiked with Hg or MeHg at various concentrations were analyzed by two methods. For HPLC-ICP-MS method, the mean recovery for MeHg spiked samples was 93 ± 4.2 %. For ID-GC-MS method, the mean recovery for MeHg spiked samples was 91 ± 5.2 %. A total of 19 predatory fish and 54 tuna fish samples were analyzed using both methods. No significant difference was observed between the MeHg data generated from both methods. The average relative standard deviation (RSD) between the two datasets for 19 predatory fish samples was 8.8 %. The average RSD between two datasets for 54 tuna fish samples was 4.4 %.The HPLC-ICP-MS method offers substantial advantages over the method of ID-GC-MS because the former method can determine the total mercury (Hg) and MeHg in a single chromatographic run and the analysis time is much shorter. The latter method is time consuming due to overnight digestion and having to conduct presample analysis for signal-matched isotope dilution spiking and requires more expensive isotope labeled reagents.

Speciation analysis of bromine-containing drug metabolites in feces samples from a human in vivo study by means of HPLC/ICP-MS combined with on-line isotope dilution

The aim of this work was speciation analysis of metabolites in feces samples collected within a clinical study during which a bromine-containing anti-tuberculosis drug (TMC207) was administered to patients with multi-drug resistant tuberculosis infection. Owing to slow elimination of the drug, no (14)C label was used within this study. Quantification of the bromine species was accomplished using high performance liquid chromatography coupled to inductively coupled plasma-mass spectrometry (HPLC/ICP-MS) in combination with on-line isotope dilution (on-line ID), while structural elucidation of the species was performed using HPLC coupled to electrospray ionization-mass spectrometry. The ICP-MS-based method developed shows a good intra- and inter-day reproducibility (relative standard deviation = 3.5%, N = 9); the limit of detection (1.5 mg TMC207 L(-1)) is of the same order of magnitude as that for HPLC/radiodetection; the dynamic range of the method covers more than two orders of magnitude. Furthermore, the column recovery was demonstrated to be quantitative (recoveries between 90.6% and 99.5%). Based on the excellent figures of merit, the "cold" HPLC/ICP-MS approach could be deployed for the actual human in vivo metabolism study, such that exposure of the human volunteers to the (14)C radiolabel was avoided.

Quantitative arsenic speciation in two species of earthworms from a former mine site

The relationship between the total arsenic concentration and the chemical speciation of arsenic in two species of earthworm (Lumbricus rubellus and Dendrodrilus rubidus) in relation to the host soil, was investigated for 13 sites of varying arsenic content, including a background level garden soil and a former mine site at the Devon Great Consols, UK. Earthworms were collected with the host soil (As soil concentration range 16-12, 466 mg kg(-1) dry weight) and measured for their total arsenic (concentration range 7-595 mg kg(-1) dry weight) using inductively coupled plasma mass spectrometry (ICP-MS). A methanol-water mixture was used to extract arsenic species from the earthworms prior to determination of the individual arsenic species by a combination of anion and cation exchange high performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP-MS). A gradient elution anion exchange method is presented whereby nine arsenic species could be measured in one sample injection. Arsenic species were identified by comparison of retention times and sample spiking with known standards and a fully characterised seaweed extract. Arsenic was generally present in the earthworm as arsenate (As(V)) or arsenite (As(III)) and arsenobetaine (AB). Methylarsonate (MA), dimethylarsinate (DMA) and three arsenosugars (glycerol, phosphate, sulfate) were present as minor constituents. These results are discussed in relation to the mechanisms for coping with exposure to soil bound arsenic.