Arsenosugar identification in seaweed extracts using high-performance liquid chromatography/electrospray ion trap mass spectrometry

The development of analytical techniques suitable for providing structural information on a wide range of elemental species is a growing necessity. For arsenic speciation a variety of mass spectrometric techniques, mainly inductively coupled plasma mass spectrometry (ICP-MS) and electrospray tandem mass spectrometry (ES-MS/MS) coupled on-line with high-performance liquid chromatography (HPLC), are in use. In this paper we report the identification of arsenic species present in samples of marine origin (seaweed extracts) using ES ion trap mass spectrometry (IT) multistage mass spectrometry (MS(n)). Both reversed-phase and anion-exchange HPLC have been coupled on-line to ES-ITMS. Product ion scans with multiple stages of tandem MS (MS(n); n=2-4) were used to acquire diagnostic data for each arsenosugar. The spectra contain structurally characteristic fragment ions for each of the arsenosugars examined. In addition it was observed that upon successive stages of collision-induced dissociation (CID) a common product ion (m/z 237) was formed from all four arsenosugars examined. This product ion has the potential to be used as an indicator for the presence of dimethylated arsenosugars (dimethylarsinoylribosides). The HPLC/ES-ITMS(n) method developed allows the sensitive identification of arsenosugars present in crude seaweed extracts without the need for extended sample preparation. In fact, sample preparation requirements are identical to those typically employed for HPLC/ICP-MS analysis. Additionally, the resulting product ions are structurally diagnostic of the arsenosugars examined, and tandem mass spectra are reproducible and correspond well to those obtained using other low-energy CID techniques. As a result, the HPLC/ES-ITMS(n) approach minimises the potential for arsenic species misidentification and has great potential as a means of overcoming the need for characterised standards.     

Simultaneous separation of 17 inorganic and organic arsenic compounds in marine biota by means of high-performance liquid chromatography/inductively coupled plasma mass spectrometry

A method using high-performance liquid chromatography/inductively coupled plasma mass spectrometry (HPLC/ICP-MS) has been developed to determine inorganic arsenic (arsenite, arsenate) along with organic arsenic compounds (monomethylarsonic acid, dimethylarsinic acid, arsenobetaine, arsenocholine, trimethylarsine oxide, tetramethylarsonium ion and several arsenosugars) in fish, mussel, oyster and marine algae samples. The species were extracted by means of a methanol/water mixture and a dispersion unit in 2 min, with extraction efficiencies ranging from 83 to 107% in the different organisms. Up to 17 different species were determined within 15 min on an anion-exchange column, using a nitric acid gradient and an ion-pairing reagent. As all species are shown in one chromatogram, a clear overview of arsenic distribution patterns in different marine organisms is given. Arsenobetaine is the major compound in marine animals whereas arsenosugars and arsenate are dominant in marine algae. The method was validated with CRM DORM-2 (dogfish muscle). Concentrations were within the certified limits and low detection limits of 8 ng g(-1) (arsenite) to 50 ng g(-1) (arsenate) were obtained.     

Speciation of dimethylarsinyl-riboside derivatives (arsenosugars) in marine reference materials by HPLC-ICP-MS

Anion and cation exchange HPLC-ICP-MS was used to separate and detect mixtures of four dimethylarsinyl-riboside derivatives (arsenosugars), in the presence of eight other arsenic species naturally occurring in the marine environment. The separations achieved showed that two arsenosugars 11 and 13 (cf. Table 2) were present in shellfish certified reference materials (CRMs) and in a lobster hepatopancreas CRM. The concentration of the two arsenosugars in the shellfish samples amounts to 18% of the total arsenic as compared to arsenobetaine at 9–13% and dimethylarsinate at 4–9% of the total arsenic. Additionally, a chromatographic peak with the same retention time as that of 2-dimethylarsinylacetic acid was detected in the shellfish samples. Further support of the identity of this peak was gained after spiking the sample extracts with the standard substance which resulted in a single, but larger peak. The indication that this novel arsenical is present in shellfish, and the recently reported finding of arsenocholine in seafood supports a proposed marine biosynthetic pathway of arsenic that includes both of these compounds as the immediate precursors of arsenobetaine, the end-product of the marine arsenic metabolism.     

Precursor ion scanning for the non-targeted detection of individual arsenosugars in extracts of marine organisms

Arsenosugars are a group of arsenic compounds reported to be present in a wide variety of marine organisms. Numerous such compounds have been identified and characterized in marine organisms; however, unknown arsenosugar species may also be present. This indicates the need for an analytical technique suitable for their non-targeted detection. One such technique is tandem mass spectrometry operated in the precursor ion scanning mode. This technique is based on scanning for precursor ions that give specific product ions, characteristic of the compounds under investigation. In the present study two subgroups of arsenosugar species were examined, the oxo- and the thioarsenosugars, the CID behavior of which is well known from previous studies. In the case of the oxoarsenosugars characteristic product ions were observed at m/z 237 and 97, and for the thioarsenosugars at m/z 253 and 97. Validation of this approach was carried out by analyzing extracts of two commercial kelp powders with known contents of arsenosugar species. All arsenosugars reported to exist in these materials were detected successfully using the precursor ion scanning approach. The limits of detection for the oxo- and the thioarsenosugar species, and the selectivity and sensitivity of the method, strongly indicate the suitability of this approach for the non-targeted detection of arsenosugars in extracts of marine origin.     

Simultaneous determination of twelve inorganic and organic arsenic compounds by liquid chromatography-ultraviolet irradiation-hydride generation atomic fluorescence spectrometry

A coupling between column liquid chromatography (LC) and atomic fluorescence spectrometry was developed for arsenic speciation. After separation, the compounds are oxidised on-line by UV irradiation, volatilised by hydride-generation and carried to the detector by a stream of argon. A combination of anion-exchange and hydrophobic interactions in a single column (Dionex AS7) was found suitable for the simultaneous separation of organic and inorganic species. Twelve compounds (arsenite, arsenate, monomethylarsonic acid, dimethylarsinic acid, arsenobetaine, arsenocholine, trimethylarsine oxide, tetramethylarsonium ion and four arsenosugars) were separated using an acetate buffer and a nitric acid solution as mobile phases. Limits of detection are 4-22 pg. The technique was applied to three marine samples. Arsenobetaine was detected as the main species in all samples, with concentrations varying from 59 to 1947 ng(As) g(-1) of fresh mass.     

Biodegradation of arsenosugars in marine sediment

In the marine environment, arsenic accumulates in seaweed and occurs mostly in the form of arsenoribofuranosides (often called arsenosugars). This study investigated the degradation pathways of arsenosugars from decaying seaweed in a mesocosm experiment. Brown seaweed (Laminaria digitata) was placed on top of a marine sediment soaked with seawater. Seawater and porewater samples from different depths were collected and analysed for arsenic species in order to identify the degradation products using high‐performance liquid chomatography–inductively coupled plasma mass spectrometry. During the first 10 days most of the arsenic found in the seawater and the shallow sediment is in the form of the arsenosugars released from the seaweed. Dimethylarsenoylethanol (DMAE), dimethylarsinic acid (DMA(V)) and, later, monomethylarsonic acid (MMA(V)) and arsenite and arsenate were also formed. In the deeper anaerobic sediment, the arsenosugars disappear more quickly and DMAE is the main metabolite with 60–80% of the total arsenic for the first 60 days besides a constant DMA(V) contribution of 10–20% of total soluble arsenic. With the degradation of the soluble DMAE the solubility of arsenic decreases in the sediment. The final soluble degradation products (after 106 days) were arsenite, arsenate, MMA(V) and DMA(V). No arsenobetaine or arsenocholine were identified in the porewater. Copyright © 2005 John Wiley & Sons, Ltd.     

Nitrogen purity influences the occurrence of As+ ions in high-performance liquid chromatography/electrospray ionization mass spectrometric analysis of four common arsenosugars

High-performance liquid chromatography coupled to electrospray ionization mass spectrometry (HPLC/ESI-MS) can provide both elemental and molecular information and, therefore, is a very useful tool for the identification of arsenic compounds. When a method for the identification of four arsenosugars was employed in our laboratory with an HPLC/ESI-MS system equipped with a Whatman model 75-72 nitrogen generator, a signal at m/z 75 (As(+)) could not be observed. When the HPLC/ESI-MS system was operated with nitrogen 5.0 (nitrogen of a purity of at least 99.999%) all four arsenosugars gave a signal at m/z 75. Because of this observation the influence of the quality of the nitrogen drying gas on the fragmentation of the four arsenosugars was systematically investigated. Standard solutions containing the four arsenosugars (0.5 ng As each) were separated on an anion-exchange column and detected with ESI-MS in the positive ion mode by monitoring the signals for [M+H](+), m/z 237, 91, and 75. Nitrogen with defined oxygen concentrations was used as drying gas. The purity of the nitrogen ranged from 99 to 99.999% (10 400 to 10 ppm oxygen impurity). The nitrogen with 99% purity gave no signal at m/z 75, but signals were obtained at m/z 91, 237, and for [M+H](+). When higher purity nitrogen (99.9%) was used, a signal was obtained at m/z 75, which accounted for 0.8-1.1% (depending on the kind of arsenosugar) of the sum of the signals for m/z 75, 91, 237 and [M+H](+). As the level of oxygen in the nitrogen decreased, the m/z 75 signal increased to 2.0-3.1%. This was accompanied by a concomitant decrease in the m/z 91 signal from 5.2-6.6% to 0.7-1.5%, whereas the signals for [M+H](+) and m/z 237 were essentially unchanged. Signals at m/z 75 with intensities comparable with those observed for the 99.9% pure nitrogen were also obtained for all the arsenosugars when the HPLC/ESI-MS system was operated with a Domnick Hunter Nitrox UHPLCMS18 nitrogen generator. Dimethylarsinic acid, arsenobetaine, trimethylarsine oxide, arsenocholine and the tetramethylarsonium cation also gave no signal at m/z 75 when they were analyzed with the Whatman model 75-72 nitrogen generator, but clear signals at m/z 75 were observed with the Domnick Hunter Nitrox UHPLCMS18 nitrogen generator. A nitrogen quality of at least 99.9% is required to obtain elemental information (m/z 75) when arsenic compounds are investigated by HPLC/ESI-MS. Molecular and elemental information from one chromatographic run is a valuable tool for the characterization of unknown arsenic compounds.     

A systematic study on the extractability of arsenic species from algal certified reference material IAEA-140/TM (Fucus sp., Sea Plant Homogenate) using methanol/water extractant mixtures

Using methanol/water mixtures (from pure water to pure methanol), with different desorption and solubility parameters, and varying extractant volume to algal mass (V/m) ratios, the extractability of arsenic species from CRM IAEA-140/TM was investigated. A linear sorption isotherm-based model was developed to process the data obtained with variable volume extraction, allowing the unambiguous deduction of the maximal extractable species concentrations under the specific extraction conditions, even for more stable species.The maximal extractable arsenic fraction ranged from 41 to 68% of the total arsenic concentration in CRM IAEA-140/TM, depending on the extractant composition, with pure methanol giving the lowest extraction yield and pure water giving erratic extractability (probably due to bad wettability). The main arsenic species quantified in the methanol/water extracts were arsenosugars, with arsenosugars 1 (glycerol arsenosugar), 3 (sulfonate arsenosugar) and 4 (sulfate arsenosugar) making up ca. 90% of the maximal extractable arsenic. The rest accounts for DMA (dimethylarsinate), arsenosugar 2 (phosphate arsenosugar) and As(V). There is no clear extraction pattern emerging from the data although it may be seen that extraction of more polar species (e.g. arsenosugar 1) is favoured in pure methanol and less polar more ionic species (e.g. arsenosugar 2 and As(V)) in methanol extractants with a higher water percentage.The precise and highly accurate data may be used for quality control purposes under strictly followed extraction conditions since the extraction is operationally defined. Additionally, the variable volume extraction methodology presented may be applied to other elemental species in other matrices using other extractants. Although this approach does not maximise the absolute extractability but only that which is extractant-specific, experimentators are forewarned that in most cases only a fingerprint of the extractant-specific species is produced unless a quantitative extraction of all species is obtained.     

Arsenic speciation in chinese seaweeds using HPLC-ICP-MS and HPLC-ES-MS

Three common Chinese edible seaweeds, one brown (Laminaria japonica) and two red (Porphyra crispata and Eucheuma denticulatum), were examined for their total arsenic content. The As species were extracted with yields of 76.4, 69.8 and 25.0%, respectively. Anion-exchange and cation-exchange high-performance liquid chromatography (HPLC) in combination with inductively coupled plasma mass spectrometry (ICP-MS) were used for the separation of the different arsenic species in two of the three seaweed extracts (Laminaria and Porphyra). The main arsenic species in the algal extracts are arseno sugars, although it has been shown that the Laminaria seaweed contains significant amounts of dimethylarsinic acid (DMA). HPLC was coupled with electrospray mass spectrometry (ES-MS) for structural confirmation of the arsenic species. The mass spectrometer settings for the arseno sugars were optimised using standards. The conclusions drawn on the basis of HPLC-ICP-MS were confirmed by the HPLC-ES-MS data. The HPLC-ES-MS method is capable of determining both arseno sugars and DMA in the seaweeds. The unknown compounds seen in the HPLC-ICP-MS chromatogram of Laminaria could not be ascribed to trimethylarsenic oxide or tetramethylarsonium ion.     

Determination of arsenic species by inductively coupled plasma mass spectrometry with ion chromatography

Five arsenic species, trimethylarsine oxide, dimethylarsenic acid, monomethylarsonic acid, arsenobetaine and sodium arsenite, in urine were analysed by inductively coupled plasma mass spectrometry with ion chromatography (IC ICP MS). Since the toxicities of different arsenic compounds are different, speciation of arsenic compounds is very important in the investigation of metabolisms. In this paper, we applied ion chromatography (IC) as a separation device and inductively coupled plasma mass spectrometry (ICP MS) as a detection device. For separation of the five arsenic compounds, an anion-exchange column and, as mobile phase, tartaric acid were used. The eluent from the IC column was introduced directly into the nebulizer of the ICP MS and analysed at 75 amu. Detection limits were from 4 to 9 pg as arsenic.     

Arsenic metabolism in seaweed-eating sheep from Northern Scotland

Cation exchange and anion exchange liquid chromatography were coupled to an ICP-MS and optimised for the separation of 13 different arsenic species in body fluids (arsenite, arsenate, dimethylarsinic acid (DMAA), monomethylarsonic acid (MMAA), trimethylarsine oxide (TMAO), tetramethylarsonium ion (TMA), arsenobetaine (AsB), arsenocholine (AsC), dimethylarsinoyl ethanol (DMAE) and four common dimethylarsinoylribosides (arsenosugars). The arsenic species were determined in seaweed extracts and in the urine and blood serum of seaweed-eating sheep from Northern Scotland. The sheep eat 2-4 kg of seaweed daily which is washed ashore on the most northern Island of Orkney. The urine, blood and wool of 20 North Ronaldsay sheep and kidney, liver and muscle from 11 sheep were sampled and analysed for their arsenic species. In addition five Dorset Finn sheep, which lived entirely on grass, were used as a control group. The sheep have a body burden of approximately 45-90 mg arsenic daily. Since the metabolism of arsenic species varies with the arsenite and arsenate being the most toxic, and organoarsenic compounds such as arsenobetaine the least toxic compounds, the determination of the arsenic species in the diet and their body fluids are important. The major arsenic species in their diet are arsenoribosides. The major metabolite excreted into urine and blood is DMAA (95 +/- 4.1%) with minor amounts of MMAA, riboside X, TMA and an unidentified species. The occurrence of MMAA is assumed to be a precursor of the exposure to inorganic arsenic, since demethylation of dimethylated or trimethylated organoarsenic compounds is not known (max. MMAA concentration 259 microg/L). The concentrations in the urine (3179 +/- 2667 microg/L) and blood (44 +/- 19 microg/kg) are at least two orders of magnitude higher than the level of arsenic in the urine of the control sheep or literature levels of blood for the unexposed sheep. The tissue samples (liver: 292 +/- 99 microg/kg, kidney: 565 +/- 193 microg/kg, muscle: 680 +/- 224 microg/kg) and wool samples (10470 +/- 5690 microg/kg) show elevated levels which are also 100 times higher than the levels for the unexposed sheep.     

高效液相色谱-电感耦合等离子体质谱联用技术测定干海产品中砷化学形态

利用高效液相色谱-电感耦合等离子体质谱联用技术研究了海带、羊栖菜、紫菜及浒苔等干海产品中的砷含量及其化学形态.实验表明,干海产品中主要砷形态为3种未知砷化物,对3种未知砷化物进行了表征.采用高效液相色谱与飞行时间质谱联用技术对羊栖菜和海带中的3种未知砷化物的分子量,分别为329.0599, 483.0738和409.0162,通过对比,确定这3个未知峰分别为3种砷糖类物质: DMA-glycerol ribose, DMA-phosphate ribose和DMA-sulfate ribose.对样品中砷的定量分析结果显示,干海产品中砷总量虽然超出我国国标中关于海产品中砷限量标准规定的20倍以上,但大多数植物性干海产品中主要的砷形态为毒性较低的砷糖类物质DMA-sulfate ribose(U4),其含量约占可提取砷量的51.1%～80.3%,海带中的砷主要以DMA-phosphate ribose(U3)的形式存在,占总可提取砷的48.9%.     

Determination of arsenic compounds in reference materials by HPLC-(UV)-HG-AFS

Arsenic compounds were determined in six reference materials of biological origin. None of them has yet been certified for arsenic compounds but some are in the process of certification; for most of these reference materials indicative literature values are available. Eight commonly used arsenic standards were used for quantification using a recently developed hyphenated speciation system comprising high performance liquid chromatography (HPLC) and atomic fluorescence spectrometry (AFS), interfaced via a UV-photoreactor and a hydride generation (HG) unit. Absolute detection limits were ca. 0.2 and 0.4 ng As for separation on anion and cation exchange columns, respectively. Our results agree well with indicative literature values which were generated by different authors using various separation and detection methods. The HPLC-(UV)-HG-AFS system validated in this way is suitable for quantification of eight arsenic compounds. Moreover, the system is capable of separation of at least six more compounds in the mentioned reference materials, of which two could be attributed to arsenosugars (OH and phosphodiester form) but due to the lack of standards, quantification was not possible. For accurate and extensive speciation analysis the availability of certified reference materials and standards for arsenic compounds should be promoted.     

Determination of an arsenosugar in oyster extracts by liquid chromatography-electrospray mass spectrometry and liquid chromatography-ultraviolet photo-oxidation-hydride generation atomic fluorescence spectrometry

HPLC-UV-HG-AFS analysis of aqueous extracts of oysters (Crassostrea gigas) taken from the southwestern Atlantic coast of Spain showed the presence of arsenite, arsenate, dimethylarsinic acid and an unidentified arsenic peak. Subsequent analysis of the oyster samples by LC-electrospray MS and comparison with four standard dimethylarsinoylribosides (arsenosugars), showed that the previously unidentified peak was an arsenosugar (arsenosugar 2). When the arsenosugar in the oyster was quantified using the two detection methods and external calibration with standard arsenosugar, there was a large discrepancy between the two sets of results. The LC-MS analysis was strongly affected by the sample matrix and gave concentrations 50% lower than those obtained by AFS detection. When the method of standard addition was applied to the LC-MS analysis, the results were comparable to the AFS data. The matrix effects were eliminated by subjecting the extract to a clean-up procedure with anion-exchange and gel permeation preparative chromatography before the LC-MS analysis. The arsenosugars gave a small signal without photo-oxidation when they were analysed by HPLC-HG-AFS. Possibly this resulted from partial decomposition of the arsenosugar to dimethylarsinic acid under the acidic conditions employed in the hydride generation step.     

Critical review or scientific opinion paper: Arsenosugars—a class of benign arsenic species or justification for developing partly speciated arsenic fractionation in foodstuffs?

In this opinion paper the toxicokinetic behaviour of arsenosugars is reviewed and compared with that of inorganic arsenic and arsenobetaine. It is concluded that the arsenosugars are similar to inorganic arsenic in terms of metabolite formation and tissue accumulation. As a pragmatic means of generating uniform data sets which adequately represent the toxicity of arsenic in food we recommend reporting partly speciated arsenic concentrations in food commodities in three fractions: i) toxic inorganic arsenic as arsenate (after oxidation); ii) arsenobetaine as established non-toxic arsenic; and iii) potentially toxic arsenic, which includes arsenosugars and other organoarsenicals.     

Biotransformation of arsenate to arsenosugars by Chlorella vulgaris

Chlorella vulgaris was cultivated in a growth medium containing arsenate concentration of <0.01, 10, 100 and 1000 mg l^?1. Illumination was carried out in 12 h cycles for 5 days. The health status of the culture was monitored by continuous pH and dissolved oxygen (DO) readings. Destructive sampling was used for the determination of biomass, chlorophyll, total arsenic and arsenic species. The chlorophyll a content, the DO and pH cycles were not significantly different for the different arsenate concentrations in the culture. In contrast, biomass production was significantly (p < 0.05) increased for the arsenic(V) treatment at 1000 mg l^?1 compared with 100 mg l^?1. The arsenic concentration in the algae increased with the arsenate concentration in the culture. However, the bioconcentration factor decreased a hundred‐fold with increase of arsenate from the background level to 1000 mg l^?1. The arsenic species were identified by using strong anion‐exchange high‐performance liquid chromatography–inductively coupled plasma mass spectrometry analysis after methanol/water (1 : 1) extraction. The majority (87–100%) of the extractable arsenic was still arsenate; arsenite was found to be between 1 and 6% of total extractable arsenic in the algae. In addition to dimethylarsinic acid, one unknown arsenical (almost co‐eluting with methylarsonic acid) and three different arsenosugars have been identified for the first time in C. vulgaris growing in a culture containing a mixture of antibiotics and believed to be axenic. The transformation to arsenosugars in the algae is not dependent on the arsenate concentration in the culture and varies between 0.2 and 5% of total accumulated arsenic. Although no microbiological tests for bacterial contamination were made, this study supports the hypothesis that algae, and not associated bacteria, produce the arsenosugars. Copyright © 2003 John Wiley & Sons, Ltd.     

Measurement of arsenic species in marine sediments by high-performance liquid chromatography-inductively coupled plasma mass spectrometry

Extraction of sediments with phosphoric acid (0.5 M) and hydroxylamine hydrochloride (0.1 M) allowed the measurement of labile arsenic species while preserving the two redox states of arsenic. The forms and concentrations of arsenic species were measured using HPLC-ICP-MS. A Hamilton PRP X-100 strong anion exchange column using a 20 mM ammonium phosphate buffer (pH 6 and 9.2) was used to separate arsenic species. Recoveries of sediments spiked with As(V) were quantitative whereas for sediments spiked with As(III) recoveries of between 89 and 104% were obtained from four oxic certified reference sediments and an anoxic sediment. Application of the method to sediment samples from the marine Lake Macquarie, NSW, Australia, indicate that anoxic sediments can contain high concentrations of As(III), and two arsenosugars (sulfonate-ribose and sulfate-ribose). Extraction efficiencies for arsenic ranged between 6 and 82%. The arsenic species measured in sediments are strongly depended on the extraction procedure used. As(III) and arsenosugar concentrations in sediments that were freeze dried and oxidised were much less than in sediments that were not freeze dried and when exposure to air was keep to a minimum. Corresponding, As(V) concentrations tended to be higher in samples that were exposed to air.     

Arsenic metabolism in seaweed-eating sheep from Northern Scotland

Cation exchange and anion exchange liquid chromatography were coupled to an ICP-MS and optimised for the separation of 13 different arsenic species in body fluids (arsenite, arsenate, dimethylarsinic acid (DMAA), monomethylarsonic acid (MMAA), trimethylarsine oxide (TMAO), tetramethylarsonium ion (TMA), arsenobetaine (AsB), arsenocholine (AsC), dimethylarsinoyl ethanol (DMAE) and four common dimethylarsinoylribosides (arsenosugars). The arsenic species were determined in seaweed extracts and in the urine and blood serum of seaweed-eating sheep from Northern Scotland. The sheep eat 2–4 kg of seaweed daily which is washed ashore on the most northern Island of Orkney. The urine, blood and wool of 20 North Ronaldsay sheep and kidney, liver and muscle from 11 sheep were sampled and analysed for their arsenic species. In addition five Dorset Finn sheep, which lived entirely on grass, were used as a control group. The sheep have a body burden of approximately 45–90 mg arsenic daily. Since the metabolism of arsenic species varies with the arsenite and arsenate being the most toxic, and organoarsenic compounds such as arsenobetaine the least toxic compounds, the determination of the arsenic species in the diet and their body fluids are important. The major arsenic species in their diet are arsenoribosides. The major metabolite excreted into urine and blood is DMAA (95 ± 4.1%) with minor amounts of MMAA, riboside X, TMA and an unidentified species. The occurrence of MMAA is assumed to be a precursor of the exposure to inorganic arsenic, since demethylation of dimethylated or trimethylated organoarsenic compounds is not known (max. MMAA concentration 259 μg/L). The concentrations in the urine (3179 ± 2667 μg/L) and blood (44 ± 19 μg/kg) are at least two orders of magnitude higher than the level of arsenic in the urine of the control sheep or literature levels of blood for the unexposed sheep. The tissue samples (liver: 292 ± 99 μg/kg, kidney: 565 ± 193 μg/kg, muscle: 680 ± 224 μg/kg) and wool samples (10 470 ± 5690 μg/kg) show elevated levels which are also 100 times higher than the levels for the unexposed sheep. Received: 29 February 2000 / Revised: 26 April 2000 / Accepted: 1 May 2000     

Speciation of arsenic compounds by HPLC with hydride generation atomic absorption spectrometry and inductively coupled plasma mass spectrometry detection

An arsenic specific detection system utilizing on-line microwave digestion and hydride generation atomic absorption spectrometry (MD/HGAAS) is described for arsenic speciation by using high performance liquid chromatography (HPLC). Both ion exchange chromatography and ion pair chromatography have been studied for the separation of arsenite, arsenate, monomethylarsonic acid (MMAA), dimethylarsinic acid (DMAA), and arsenobetaine (AB). When the commonly used mobile phases, phosphate and carbonate buffers at pH 7.5, are used on an anion exchange column, arsenite and AB co-elute. However, selective determination of these two arsenic compounds can be achieved by using the new detection system. Partial separation between arsenite and AB can be achieved by increasing the mobile phase pH to 10.3 and by using a polymer based anion exchange column. The detection limit obtained by using anion exchange chromatography with MD/HGAAS detection is approximately 10 ng/ml (or 200 pg for a 20-mul sample injection) for arsenite, DMAA and AB, 15 ng/ml (or 300 pg) for MMAA, and 20 ng/ml (or 400 pg) for arsenate. Complete separation of the five arsenic compounds is achieved on a reversed phase C18 column by using sodium heptanesulfonate as ion pair reagent. Comparable resolution between chromatographic peaks is obtained by using MD/HGAAS detection and inductively coupled plasma mass spectrometry (ICPMS) detection.     

The identification of some water-soluble arsenic species in the marine brown algae Fucus distichus

The extraction and clean-up procedures developed to isolate the water-soluble arsenic species present in the marine macroalga Fucus distichus, from British Columbia, Canada, are described. The arsenic species were extracted into methanol and then subjected to gel-permeation and ion-exchange chromatography. Fractions high in arsenic were identified by using graphite furnace atomic absorption spectroscopy (GF-AAS), and further investigated by using high-performance liquid chromatography coupled to inductively coupled plasma–mass spectrometry (HPLC–ICP MS). By using different HPLC columns and mobile-phase conditions, the four major arsenic-containing compounds present in the macroalga were positively identified as arsenosugars; one minor compound remained unidentified. © 1997 John Wiley & Sons, Ltd.