Although it has been known for decades that arsenic forms fat-soluble arsenic compounds, only recent attempts to identify the compounds have been successful by using a combination of fractionation and elemental and molecular mass spectrometry. Here we show that arsenolipids can directly be identified and quantified in biological extracts using reversed-phase high-performance liquid chromatography (RP-HPLC) simultaneously online-coupled to high-resolution inductively coupled plasma mass spectrometry (ICPMS) and high-resolution electrospray mass spectrometry (ES-MS) without having a lipophilic arsenic standard available. Using a methanol gradient for the separation made it necessary to use a gradient-dependent arsenic response factor for the quantification of the fat-soluble arsenic species in the extract. The response factor was obtained by using the ICPMS signal of known concentration of arsenic. The arsenic response was used to determine species-specific response factors for the different arsenic species. The retention time for the arsenic species was utilized to mine the ES-MS data for accurate mass and their tandem mass spectrometry (MS/MS) fragmentation pattern to give information of molecular formula and structure information. The majority of arsenolipids, found in the hexane phase of fish meal from capelin ( Mallotus villosus ) was in the form of three dimethylarsinoyl hydrocarbons (C(23)H(38)AsO, C(17)H(38)AsO, C(19)H(42)AsO) with minor amounts of dimethylarsinoyl fatty acids (C(17)H(36)AsO(3), C(23)H(38)AsO(3), C(24)H(38)AsO(3)). One of the dimethylarsinoyl fatty acids (C(24)H(38)AsO(3)), with an even number of carbon in the fatty acid chain, was identified for the first time in this work. This molecular formula is unusual and in contrast to all previously identified arsenic-containing fatty acids with odd numbers of carbon.
Because of the toxicity of inorganic arsenic (iAs), only iAs needs to be monitored in food and feedstuff. This demands the development of easy and quick analytical methods to screen large number of samples. This work focuses on hydride generation (HG) coupled with an ICPMS as an arsenic detector where the HG is added as a selective step to determine iAs in the gaseous phase while organically bound As remains in the solution. iAs forms volatile arsine species with high efficiency when treated with NaBH4 at acidic conditions, whereas most other organoarsenic compounds do not form any or only less volatile arsines. Additionally, using high concentrations of HCl further reduces the production of the less volatile arsines and iAs is almost exclusively formed, therefore enabling to measure iAs without a prior step of species separation using chromatography. Here, we coupled a commercially available HG system to an ICPMS and optimized for determination of iAs in rice and samples of marine origin using different acid concentrations, wet and dry plasma conditions, and different reaction gas modes. Comparing this method to conventional HPLC-ICPMS, no statistical difference in iAs concentration was found and comparable limits of detections were achieved using less than half the instrument time.
The addition of an online post-column hydride generation (HG) step to the commonly used high-performance liquid chromatography inductively coupled plasma-mass spectrometry (HPLC-ICP-MS) setup for arsenic speciation proved to significantly improve the detection limits for the determination of inorganic arsenic (iAs) as arsenate in seafood samples, where the limit of detection and limit of quantification were found to be 0.0004 and 0.0014 mg kg(-1), respectively with HG. HG as an additional step further added to the selectivity of the determination of the iAs species and increased the detection and quantification of low levels of iAs (<0.002 mg kg(-1)) in samples with complicated matrices.
To evaluate the accuracy and robustness of an extraction method, utilizing an -alkaline-ethanolic solution and microwave heating, the certified reference material (CRM) TORT-2 was subjected to three different instrumental methodologies: high-performance liquid chromatography (HPLC), coupled with and without post-column hydride generation; inductively coupled plasma-mass spectrometry (ICP-MS); and HPLC-hydride generation-atomic fluorescence spectrometry (HPLC-HG-AFS). The three methods gave a consistent value of inorganic arsenic (As) which is near the mean value of the reported values in the literature, which, however, range by a factor of 10. Inorganic As, defined here as all As species that do not have an As–C bond, that is, the sum of arsenite and arsenate and any thiol-bound As, was found to be less than 4 % of total As concentration in 12 samples of fish meal when subjected to this extraction method followed by HPLC-ICP-MS. To date, there is no certified value of inorganic As in a seafood-based reference material to compare to in order to validate the findings. This illustrates the difficulties in quantitative determination of inorganic As in seafood and the need for a reference material for inorganic As and proficiency tests in order to introduce legislation for a maximum level of inorganic As in seafood and feed.
Brown macroalgae
Saccharina latissima
(30–40 individuals) and
Alaria esculenta
(15–20 individuals) were collected from natural populations in winter in Iceland. The algal thalli were sectioned into different parts (e.g. holdfast, stipe, old frond, young frond and sori-containing frond sections) that differed in age and biological function. The work elucidated that arsenic (As) was not uniformly distributed within the two brown macroalgal species, with lower levels of total As were found in the stipe/midrib compared to other thallus parts. The arsenosugars mirrored the total arsenic in the seaweed mainly due to AsSugSO
3
being the most abundant As species. However, arsenic speciation using parallel HPLC-ICP-MS/ESI-MS elucidated that the arsenic-containing lipids (AsL) had a different distribution where the arsenosugarphospholipids (AsPL) differed by approximately a factor of 4 between the sections containing the lowest and highest concentrations of AsPLs. When placing the sections in order of metabolic activity and an estimate of tissue age, there appeared to be a relationship between the activity and AsPLs, with lower levels of AsPLs in oldest parts. This is the first time such a relationship has been shown for AsLs. Hence, by applying sophisticated analytical techniques, it was possible to gain a deeper understanding of arsenolipids in seaweed.
Electronic supplementary material
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