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Modulation of cell adhesion and viability of cultured murine bone marrow cells by arsenobetaine, a major organic arsenic compound in marine animals.

1. In this study, we investigated the biological effects of trimethyl (carboxymethyl) arsonium zwitterion, namely arsenobetaine (AsBe), which is a major organic arsenic compound in marine animals using murine bone marrow (BM) cells and compared them with those of an inorganic arsenical, sodium arsenite, in vitro. 2. Sodium arsenite showed strong cytotoxicity in BM cells, and its IC(50) was 6 microM. In contrast, AsBe significantly enhanced the viability of BM cells in a dose-dependent manner during a 72-h incubation; about a twofold increase in the viability of cells compared with that of control cells cultured with the medium alone was observed with a microM level of AsBe. 3. In morphological investigations, AsBe enhanced the numbers of large mature adherent cells, especially granulocytes, during a 72-h BM culture. When BM cells were cultured together with AsBe and a low dose (1 u ml(-1)) of recombinant murine granulocyte/macrophage colony-stimulating factor (rMu GM-CSF), significant additive-like increasing effects were observed on the numbers of both granulocytes and macrophages originated from BM cells. However, AsBe did not cause proliferation of BM cells at all as determined by colony-forming assay using a gelatinous medium. 4. These findings demonstrate the unique and potent biological effects in mammalian cells of AsBe, a major organic arsenic compound in various marine animals which are ingested daily as seafood in many countries.

Animals↗

Differential influences of various arsenic compounds on glutathione redox status and antioxidative enzymes in porcine endothelial cells.

The cellular response and detoxification mechanisms in porcine endothelial cells (PAECs) to arsenic trioxide (As2O3), sodium arsenite (NaAsO2) and sodium arsenate (Na2HAsO4) were investigated. NaAsO2 at 20 microM for 72 h increased Cu/Zn superoxide dismutase activity resulting in elevated intracellular hydrogen peroxide levels, but As2O3 and Na2HAsO4 did not. Trivalent arsenic compounds increased intracellular oxidized glutathione (GSSG) and total glutathione (GSH) and cellular glutathione peroxidase (cGPX) and glutathione S-transferase (GST) activity, but not glutathione reductase activity. The increased cGPX activity resulted in an elevated cellular GSSG content. Na2HAsO4 increased the cellular GSSG level at 72 h compared to controls. These results imply that the increased GSH content responding to the oxidative stress by trivalent arsenic compounds may be mainly related to the regulation of GSH turnover. The increased GST activity implies that the elevated intracellular GSH level responding to the oxidative stress may be used to conjugate arsenic in PAECs and facilitate arsenic efflux.

Animals↗

Distribution of arsenic compounds in Mytilus galloprovincialis of the Venice lagoon (Italy).

Samples of Mytilus galloprovincialis collected in different sites of the Venice lagoon (Italy) were investigated for total arsenic concentrations by ICP-AES and for single arsenic species by HPLC-ICP-MS. For this purpose, an analytical procedure for the sensitive and efficient speciation of the arsenic species As(III), As(V), monomethylarsonic acid (MMA), dimethylarsinic acid (DMA), arsenobetaine (AB), arsenocholine (AC), and four arsenosugars was optimised. The total arsenic and the single arsenic species were determined in both the hepatopancreas (digestive gland) and the remaining soft tissues in order to verify the different arsenic accumulation in the body parts of mussels. Arsenic compounds were extracted from the mussels with a methanol/water mixture; the extracts were evaporated to dryness, redissolved in water, and chromatographed in an anion-exchange column, a Hamilton PRP-X100. Only small quantities or traces of inorganic arsenic were detected in the mussels. The majority of arsenic compounds detected in the extracts were organic species, with a predominance of arsenobetaine and of an arsenosugar. In addition, a greater arsenic accumulation in the digestive glands of mussels was observed.

Animals↗

Arsenosugars and other arsenic compounds in littoral zone algae from the Adriatic Sea.

In 10 different marine algae from the littoral zone (found between the highest and lowest tide marks on the seashore) arsenic compounds were determined by means of a high-performance liquid chromatography (anion and cation exchange)-UV photochemical digestion-hydride generation-atomic fluorescence spectrometry (HPLC-UV-HGAFS) system. Samples (Ceramium sp., Cystoseira barbata, Enteromorpha sp., Fucus virsoides, two different species of Gelidium, Padina pavonica, Polisyphonia sp. and Ulva rigida) were collected along the Adriatic Sea coast of Slovenia. The total arsenic content of the algal samples, as determined by ICP-MS, ranged from 1.35 to 28.1 microg g(-1) (fresh weight). In all algae but two, the most abundant arsenic species found were arsenosugars with minor amounts of other arsenic compounds. Cystoseira barbata and Ceramium sp. contained high amounts of mainly inorganic arsenic. A small quantity of arsenobetaine was detected in most of the investigated Adriatic algae, which probably originates from mesofauna attached to the algae in their natural habitat.

Arsenates↗

The reaction of allyl and benzylarsonic acids with thiols: mechanistic aspects and implications for dioxygen activation by trivalent arsenic compounds.

The reaction of allyl and benzylarsonic acids with thiophenol gives not only the expected diphenyl alkyldithioarsonites and diphenyl disulfide but also various other compounds arising from the decomposition at the arsenic(V) oxidation level (the arsonic acids) by thiophenol and at the arsenic(III) oxidation level (mainly the alkyldithioarsonites) by thiophenol and by dissolved dioxygen. The reaction of these arsonic acids with 4-nitrothiophenol, which is not oxidized by dioxygen, revealed that the arsenic(III) of these alkyldithioarsonites is the active atom towards dioxygen. However, the reaction of allyl, benzyl, and 2-picolylarsonic acids with DL-penicillamine gives the expected products with no or very small oxidative decomposition. The decomposition pathways of allyl and benzylarsonic acids were elucidated. The results are briefly discussed in the contexts of the use of arsonic acids in chemotherapy and the ability of arsenic(III) compounds to generate reactive oxygen species.

Allyl Compounds↗

Accurate quantification and transformation of arsenic compounds during wet ashing with nitric acid and microwave assisted heating.

Arsenous acid, dimethylarsinic acid (DMA), methylarsonic acid (MA), arsenic acid, arsenobetaine bromide (AB), trimethylarsine oxide (TMAO), arsenocholine iodide (AC), and tetramethylarsonium iodide (TETRA) were heated in a microwave autoclave with nitric acid to 100-300 degrees C. The arsenic compounds in the digests were separated with anion- and cation-exchange chromatography and determined with an inductively coupled plasma mass spectrometer as arsenic-specific detector. Arsenous acid was completely oxidized to arsenic acid at 100 degrees C. For a complete oxidation of MA and DMA to arsenic acid temperatures > 220 degrees C and > 280 degrees C were necessary. AB decomposed to arsenic acid via TMAO. Complete conversion was only obtained after heating the sample for 90 min to 300 degrees C. For a complete conversion of TMAO similar harsh conditions were necessary. AC was already substantially degraded to TMAO, TETRA and two unknown compounds at 100 degrees C. The unknown arsenic compounds were found only in the digests up to 160 degrees C. Quantitative conversion of AC to arsenic acid went also via TMAO. At temperatures above 220 degrees C TETRA started to convert to TMAO, which then was further converted to arsenic acid. To investigate whether the results obtained for the arsenic standards are transferable to real samples, the certified reference material DORM-2 was also heated in nitric acid with variable digestion temperatures and times. For an almost complete conversion of the AB present in DORM-2 90 min at 300 degrees C were necessary. Total organic carbon (TOC) was less < 0.2% when DORM-2 was heated at temperatures > or = 260 degrees C for 60 min. UV photo-oxidation of DORM-2 was investigated as an alternative sample decomposition. Only 6% of AB was converted to arsenic acid when DORM-2 was irradiated for 2 h at 1000 W. In contrast to microwave heating substantial amounts of MA were observed as degradation product.

Journal Article↗

Arsenic compounds in the haemolymph of the Dungeness crab, Cancer magister, as determined by using HPLC on-line with inductively coupled plasma mass spectrometry.

Arsenobetaine, two arsenosugars, dimethylarsinate and several unidentified arsenic species were detected in extracts of the haemolymph of the Dungeness crab, Cancer magister, by using HPLC-ICP-MS. This is the first report of the presence of arsenosugars in the haemolymph/blood of marine animals. Total, extractable and residual arsenic concentrations were determined by ICP-MS. The concentration of total arsenic was in the range of 1.4-3.8 [micro sign]g ml(-1). Nearly all (98%) the arsenic was found to be extractable, and accounted for primarily by arsenobetaine, two arsenosugars and dimethylarsinate. The results demonstrate that arsenic compounds present in the diet of crabs are not fully metabolized in the gut. They are, at least partly, taken up into the haemolymph. The concurrence of arsenobetaine and arsenosugars suggests that the use of repeated haemolymph sampling in crustaceans could facilitate investigations into the kinetics of the biotransformation pathways of arsenic compounds. Finally, the present study clearly demonstrates the unique capabilities of HPLC-ICP-MS for the detection and identification of minor arsenic components amongst the predominant arsenobetaine.

Animals↗

Cytotoxic effect of three arsenic compounds in HeLa human tumor and bacterial cells.

Numerous epidemiological studies suggest that arsenic (As) compounds are carcinogens, however, recent data have renewed the interest in their anticarcinogenic properties. The cytotoxic effects of three arsenic compounds were assessed: sodium arsenite, sodium arsenate and sodium cacodylate, representing the trivalent and pentavalent species of arsenic, along with a dimethylated pentavalent arsenic species. HeLa cells and Salmonella typhimurium (strains TA98 and TA100) were exposed to As compounds and the cytotoxic effects were evaluated. Alterations on RNA and DNA synthesis in HeLa cells were also examined. All arsenic compounds produced a dose-dependent inhibition on colony formation and DNA synthesis in HeLa cells, yet any of them significantly influenced RNA synthesis in these cells. No evidence of arsenic-induced mutagenicity or antimutagenicity was observed using the Ames assay. In bacterial cells, only sodium arsenite caused a dose-dependent inhibition of colony formation.Collectively, these results indicate that in both, HeLa and S. typhimurium cell systems, only trivalent sodium arsenite can act as an effective inhibitor of cell growth. The possible mechanism(s) of the cytotoxic effect of arsenite in these two different cell systems might be due to its reactivity with intracellular sulfhydryl groups.

Antimutagenic Agents↗

Glutathione plays different roles in the induction of the cytotoxic effects of inorganic and organic arsenic compounds in cultured BALB/c 3T3 cells.

The cytotoxicity of arsenic compounds towards BALB/c 3T3 cells in culture was investigated, together with the role of glutathione (GSH) in the induction of the cytotoxic effects. The rank order of cytotoxicity was as follows: arsenite (As3+) > arsenate (As5+) > dimethylarsinic acid (DMAA) > methylarsonic acid (MAA) > trimethylarsine oxide (TMAO). Arsenobetaine, arsenocholine and the tetramethylarsonium ion were less toxic. Depletion of GSH enhanced the cytotoxic effects of As3+, As5+, MAA and TMAO, while the cytotoxicity of DMAA was markedly reduced by depletion of GSH. These results suggest that GSH plays a role in protecting the cells against the toxic effects of As3+, As5+, MAA and TMAO while it is involved in the induction of the cytotoxic effects of DMAA.

3T3 Cells↗

Dissolution of two arsenic compounds by rabbit alveolar macrophages in vitro.

The ability of rabbit alveolar macrophages to dissolve two arsenic compounds, 74As-labeled lead arsenate and arsenic trisulfide, was studied in vitro. The solubilities in water of these two compounds are related differently to pH. The solubility of lead arsenate increases and that of arsenic trisulfide decreases with decreasing pH. The radiolabeled particles were incubated with and without macrophages for up to 3 days, whereafter the amount of 74As in soluble form and the amount in particle form and/or bound to macrophages were determined. The results strongly support the hypothesis that the dissolution of particles by macrophages is influenced by the acid milieu in the phagosomes. About 14% of the 74As-labeled lead arsenate particles incubated for 3 days with the macrophages was released into the culture medium, compared with about 2% of the particles incubated with the culture medium without macrophages. With the arsenic trisulfide particles, less soluble 74As was released into the medium in samples with macrophages than in samples without macrophages, although the solubility in all incubations was considerably greater than that for lead arsenate. The results indicate that dissolution in the phagosomes of the macrophages may be of great importance for the clearance of particles such as lead arsenate, which are more soluble at pH 4 than at pH 7.

Animals↗

Analysis of arsenic compounds by capillary electrophoresis using indirect UV and mass spectrometric detections.

CE with indirect UV and mass-spectrometric detection was used for the simultaneous determination of arsenic acid (As(V)), arsenous acid (As(III)), monomethylarsonic acid (MMA), dimethylarsinic acid (DMA), trimethylarsine oxide (TMAO), tetramethylarsonium ion (TMA(+)), arsenobetaine (AB), and arsenocholine (AC). In the CE-indirect UV analysis, a baseline separation of arsenic species was successfully achieved by using a basic background solution (BGS) for anions and an acidic BGS for cations, respectively. The LOD values in CE-indirect UV for the individual analytes were 7.8, 12.5, 7.8, 12.5, 62.5, 125, 250, and 62.5 ppm, respectively. To achieve sensitive and selective analysis, CE coupled with ESI-MS was applied to the determination of arsenic compounds. The organic arsenic species were successfully separated with a higher sensitivity by CE-MS using the acidic BGS. The LODs in CE-MS for MMA, DMA, TMAO, TMA(+), AB, and AC were 1.0, 0.1, 0.01, 0.1, 0.01, and 0.01 ppm, respectively. In contrast, the analysis of inorganic arsenic species (As(V) and As(III)) resulted in a lower detectability in CE-MS compared to that obtained with the CE-indirect UV analysis. However, the speciation of eight arsenics by CE-MS was successfully achieved in a single run by switching the ESI polarity during MS detection.

Arsenicals↗

Liquid chromatography electrospray mass spectrometry with variable fragmentor voltages gives simultaneous elemental and molecular detection of arsenic compounds.

A single quadrupole high performance liquid chromatography electrospray mass spectrometry system with a variable fragmentor voltage facility was used in the positive ion mode for simultaneous recording of elemental and molecular mass spectral data for arsenic compounds. The method was applicable to the seven organoarsenic compounds tested: four arsenic-containing carbohydrates (arsenosugars), a quaternary arsonium compound (arsenobetaine), dimethylarsinic acid, and dimethylarsinoylacetic acid. It was not suitable for the two inorganic arsenic species arsenite and arsenate. In the case of arsenosugars, qualifying ion data for a characteristic common fragment (m/z 237) was also simultaneously obtained. The method was used to identify and quantify the major arsenosugars in crude extracts of two brown algae.

Arsenicals↗

Studies on the accumulation and transformation of arsenic in freshwater organisms I. Accumulation, transformation and toxicity of arsenic compounds on the Japanese medaka, Oryzias latipes.

Accumulation, transformation and toxicity of arsenic compounds to Japanese Medaka, Oryzias latipes were investigated. For sodium arsenite [As(II)] and disodium arsenate [As(V)], the mean value for 7-day lethal concentration LC50 for O. latipes were 14.6 and 30.3 mg As/l, respectively. Direct accumulation of arsenic in O. latipes increased as a function of As(III) concentration in water. A small proportion of accumulated arsenic was transformed to methylated arsenic. As much as 70% of the total arsenic accumulated in tissue was depurated. Accumulation and transformation of As(III) by O. latipes in a simple freshwater food chain were also investigated. The transformation of As(III) to As(V) by organisms was more prevalent than biomethylation of accumulated arsenic in organisms of the three steps of the food chain.

Animals↗

[Health effects of exposure of humans to inorganic arsenic compounds].

This paper is a review of references concerning health effects of environmental and occupational exposure to inorganic arsenic compounds. Special attention is paid to epidemiological studies indicating a relationship between time and amount of arsenic absorbed via the gastrointestinal tract (drinking water, contaminated food, drugs) and an increase in skin cancer rate. Occupational and environmental exposure of humans to arsenic dust induces a higher risk of lung cancer.

Air Pollutants, Occupational↗

Solubility, retention, and metabolism of intratracheally and orally administered inorganic arsenic compounds in the hamster.

The absorption, biotransformation, and tissue retention of arsenic following intratracheal and oral administration of 74As-labeled sodium arsenite, sodium arsenate, arsenic trisulfide (suspension), and lead arsenate (suspension) have been studied in hamsters, and correlated to the in vitro and in vivo solubility of the compounds. After intratracheal instillation, the clearance of 74As from the lungs was positively correlated to the in vivo solubility. Less than 0.1% of the sodium arsenite and sodium arsenate was retained in the lungs after 3 days, compared to 1.3% of the arsenic trisulfide particles and 45% of the lead arsenate particles. The latter showed a very low solubility both in vivo and in vitro. In general, orally administered arsenic had a shorter biological half-life than intratracheally administered, especially when given in the form of arsenic trisulfide or lead arsenate particles, which seemed to be absorbed to only 20-30% in the gastrointestinal tract. Reduction, oxidation, and methylation of arsenic varied to a great extent with the arsenic compound and the route of exposure. Trivalent arsenic was methylated to a greater extent than pentavalent and less soluble compounds (suspended particles) more than dissolved compounds. The trivalent arsenic compounds caused higher concentrations than the pentavalent in the upper gastrointestinal tract but not in other tissues.

Administration, Oral↗