Experimental design optimisation and validation by accuracy profile of a novel method for Hg speciation analysis by HPLC-ICP-MS and application to Total Diet Studies.

This study addresses the development and validation of an analytical method for speciation analysis of mercury (inorganic/Hg2+ and methylmercury/CH3Hg+) in fishery products. The Hg species are separated by reversed-phase (RP) high-performance liquid chromatography (HPLC) coupled to inductively coupled plasma mass spectrometry (ICP-MS). The effective separation of Hg2+ and CH3Hg+ was achieved in <8 min using a peptide mapping RP column and a mobile phase containing 2-mercaptoethanol at 0.25% (v/v) and methanol at 1% (v/v). The optimization was carried out using an experimental design through response surface methodology (RSM) with central composite design (CCD), addressing both the HPLC separation and the sample extraction. The method validation was carried out based on the accuracy profile approach. For this purpose, six series of measurements were carried out in duplicate over a time span of 2 months. The limits of quantification (LOQ) were 2.5 µg/kg (wet weight, ww) for CH3Hg+ and 1.2 µg/kg (ww) for Hg2+. The intermediate reproducibility in terms of coefficient of variation (CVR) was <6%. The bias (%) obtained for the analysis of four certified reference materials (CRMs), namely TORT-3 (lobster hepatopancreas), SRM 1566-b (oyster tissue), SQID-1 (cuttlefish) and NMIJ CRM 7402-a (cod fish tissue) was <7%. This demonstrates the method robustness and suitability for routine speciation analysis of CH3Hg+ and Hg2+ in fishery products. The method is intended to be applied for the analysis of the panel of fishery products and fish-based foods in the framework of the (ongoing) third French Total Diet Study.

Development of a Fast Method Using Inductively Coupled Plasma Mass Spectrometry Coupled with High-Performance Liquid Chromatography and Exploration of the Reduction Mechanism of Cr(VI) in Foods.

Hexavalent chromium (Cr(VI)) is known as the most hazardous species of chromium. Speciation analysis of Cr in foods is of a great significance for assessing its influences on human health. In this study, a fast HPLC-ICP-MS method for the determination of Cr(VI) was developed for determining the content of Cr(VI) and also investigating its transformation in foods. The developed method employs an alkali extraction and weak anion-exchange column separation for distinguishing the Cr species, facilitating accurate Cr(VI) quantification within 1.5 min. This technique was applied to determine the Cr(VI) levels in a range of food products, including yoghurt, milk powder, rice flour, orange juice, green tea, white vinegar, and whole wheat bread. The results showed that no Cr(VI) was detected in these food products. Spiking experiments revealed that the recovery rate of Cr(VI) decreased with the increase in its contact time with food products. A further exploration of Cr(VI) in various food components such as vitamin C, tea polyphenols, whey proteins, gelatin, fructose, and cellulose indicated the conversion of Cr(VI) to organic Cr(III) over a period from 20 min to 60 h. It was found that high temperatures and acidic conditions accelerated the rate of Cr(VI) conversion to organic Cr(III) in the six food components mentioned above. This evidence suggests that natural reducing substances in foods probably prevent the occurrence of Cr(VI).

Precise Determination of Selenium Forms and Contents in Selenium-Enriched Rapeseed Seedlings and Flowering Stalks by HPLC-ICP-MS.

Rapeseed (Brassica napus L.) has the ability of selenium (Se) enrichment. Identification of selenides in Se-rich rapeseed products will promote the development and utilization of high value. By optimizing the Se species extraction process (protease type, extraction reagent, enzyme sample ratio, extraction time, etc.) and chromatographic column, an efficient, stable, and accurate method was established for the identification of Se species and content in rapeseed seedlings and flowering stalks, which were cultured by inorganic Se hydroponics. Five Se compounds, including selenocystine (SeCys2), methylselenocysteine (MeSeCys), selenomethionine (SeMet), selenite (SeIV), and selenate (SeVI) were qualitatively and quantitatively identified. Organoselenium was absolutely dominant in both seedlings and flowering stalks among the detected rapeseed varieties, with 64.18-90.20% and 94.38-98.47%, respectively. Further, MeSeCys, a highly active selenide, predominated in rapeseed flowering stalks with a proportion of 56.36-72.93% and a content of 1707.3-5030.3 µg/kg. This study provides a new source of MeSeCys supplementation for human Se fortification.

The Fast Quantification of Vitamin B12 in Milk Powder by High-Performance Liquid Chromatography-Inductively Coupled Plasma Mass Spectrometry.

This study presents a new technique for determining vitamin B12 in milk powder using high-performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC-ICP-MS). We used ultrasonics with potassium ferrocyanide and zinc acetate solutions to extract the samples. 59Co was employed as the analytical target for cyanocobalamin. It was separated using a Phenomenex Luna 5 µm C18 (250 × 4.6 mm) chromatographic column with a mobile phase consisting of 1.6 mmol/L EDTA and 0.4 mmol/L KH2PO4 in a 60% v/v methanol solution (pH = 4.0). The sample has an excellent separating degree for free cobalt and cyanocobalamin, and isocratic elution can be finished within 4.0 min. To eliminate the matrix interference due to the presence of milk powder, we applied collision mode (KED). The linear range of cyanocobalamine ranged from 1.0 µg/L to 20 µg/L, with correlation coefficients (r2) of 0.9994. The limit of detection (LOD) was 0.63 µg/kg, and the limit of quantitation (LOQ) was 2.11 µg/kg. The mean recoveries were in the range of 87.4-103.6%. The accuracy and precision of the developed method are well suited for the fast quantification of the trace vitamin B12 in milk powder.

Speciation analysis of arsenic in honey using HPLC-ICP-MS and health risk assessment of water-soluble arsenic.

It is well known that arsenic is one of the most toxic elements. However, measuring total arsenic content is not enough, as it occurs in various forms that vary in toxicity. Since honey can be used as a bioindicator of environmental pollution, in the present study the concentration of arsenic and its species (As(III), As(V), DMA, MMA and AsB) was determined in honey samples from mostly Poland and Ukraine using HPLC-ICP-MS hyphenated technique. The accuracy of proposed methods of sample preparation and analysis was validated by analyzing certified reference materials. Arsenic concentration in honey samples ranged from 0.12 to 13 µg kg-1, with mean value of 2.3 µg kg-1. Inorganic arsenic forms, which are more toxic, dominated in honey samples, with Polish honey having the biggest mean percentage of inorganic arsenic species, and Ukrainian honey having the lowest. Furthermore, health risks resulting from the consumption of arsenic via honey were assessed. All Target Hazard Quotient (THQ) values, for total water-soluble arsenic and for each form, were below 1, and all Carcinogenic Risk (CR) values were below 10-4, which indicates no potential health risks associated with consumption of arsenic via honey at average or recommended levels.

Inorganic arsenic in seaweed: a fast HPLC-ICP-MS method without coelution of arsenosugars.

Seaweed is becoming increasingly popular in the Western diet as consumers opt for more sustainable food sources. However, seaweed is known to accumulate high levels of arsenic-which may be in the form of carcinogenic inorganic arsenic (iAs). Here we propose a fast method for the routine measurement of iAs in seaweed using HPLC-ICP-MS without coelution of arsenosugars that may complicate quantification. The developed method was optimised using design of experiments (DOE) and tested on a range of reference materials including TORT-3 (0.36 ± 0.03 mg kg-1), DORM-5 (0.02 ± 0.003 mg kg-1), and DOLT-5 (0.07 ± 0.007 mg kg-1). The use of nitric acid in the extraction solution allowed for the successful removal of interferences from arsenosugars by causing degradation to an unretained arsenosugar species, and a recovery of 99 ± 9% was obtained for iAs in Hijiki 7405-b when compared with the certified value. The method was found to be suitable for high-throughput analysis of iAs in a range of food and feed matrices including Asparagopsis taxiformis seaweed, grass silage, and insect proteins, and offers a cost-effective, fast, and robust option for routine analysis that requires minimal sample preparation. The method may be limited with regards to the quantification of dimethylarsenate (DMA) in seaweed, as the acidic extraction may lead to overestimation of this analyte by causing degradation of lipid species that are typically more abundant in seaweed than other marine matrices (i.e. arsenophospholipids). However, the concentrations of DMA quantified using this method may provide a better estimation with regard to exposure after ingestion and subsequent digestion of seaweed.

Occupational exposure to organotin substances: Development of a liquid chromatographic separation method for 11 organotin compounds in workplace air samples via HPLC-ICP-MS.

Organotin compounds (OTCs) are widely regulated but rank among the most used organometallic compounds in various industrial sectors. They are significantly more toxic than inorganic tin compounds. At workplaces, OTCs can be released as vapors or dust particles and can be absorbed by inhalation or skin contact. Occupational exposure thus represents a great risk for the absorption of OTCs for employees. Methods for OTCs speciation in workplace air monitoring currently do not exist. This study describes the development of a separation method for eleven in Germany regulated OTCs via HPLC-ICP-MS. The method allows a near baseline separation of MMT, MBT, MOT, MPhT, DMT, DBT, DPhT, TMT, TBT, TPhT and TTMT within 22 min on a C18 column and a ternary solvent and flow rate gradient using methanol, acetonitrile, and ultrapure water + 6% (v/v) acetic acid + 0.17% (m/v) α-tropolone. Ten analytes show linearity in the working range of 10 - 100 µg OTCs/L with R² > 0.999. Due to its high volatility the analyte TTMT showed a quadratic relationship between concentration and signal intensity with R² = 0.9998. The determination of the instrumental limits resulted in detection limits between 0.14 and 0.57 µg Sn/L and limits of quantification between 0.49 and 1.97 µg Sn/L. Over the course of this study thermal instability and cross reactivity of OTC in solution became apparent. Formation of two reaction products in mixed OTCs solutions have been observed. These effects will further be examined within development of appropriate sampling and sample preparation for workplace air to provide a suitable method for the determination of OTCs at workplaces according to normative references.

Certification of New Selenium-Enriched Yeast and Supplement Reference Materials for Selenomethionine Using Two Independent Measurement Strategies.

Selenium-enriched yeast possesses the unique ability of transforming chemical selenium, such as sodium selenite, into a biologically active form, which mitigates its toxic effects on the human body. The transformation product of this process, selenomethionine, can be safely and effectively absorbed and utilized by the human body; hence, it has been spiked into a selenium-enriched supplement. This study employs two distinct measurement strategies to determine the selenomethionine content in two candidate reference materials, a selenium-enriched yeast powder and supplement, using both organic and inorganic mass spectrometry. The concentrations of selenomethionine in the selenium-enriched yeast were determined using HPLC-ICP-MS and HPLC- ESI-MS/MS, with mass fractions measured at 718 mg SeMet kg-1 and 715 mg SeMet kg-1, respectively. Notably, both methods yielded consistent results for the selenium supplement, with a selenomethionine mass fraction of 59 mg SeMet kg-1. Ultimately, the certified values of these candidate reference materials were determined as 716 mg kg-1 and 59 mg SeMet kg-1 with expanded uncertainties of 36 mg SeMet kg-1 (k = 2) and 5 mg SeMet kg-1 (k = 2), respectively. The development of these candidate reference materials serves as a valuable reference for diverse methods aiming to determine the value of organic selenium speciation in complex food substrates.

Rapid, easy and catalyst-free preparation of magnetic thiourea-based covalent organic frameworks at room temperature for enrichment and speciation of mercury with HPLC-ICP-MS.

The complex and cumbersome preparation of magnetic covalent organic frameworks (COFs) nanocomposites on a small scale limits their application. Herein, a rapid and easy route was employed for the preparation of magnetic thiourea-based COFs nanocomposites. COFs were coated on Fe3O4 nanoparticles at room temperature without a catalyst within approximately 30 min. This method is suitable for the large-scale preparation of magnetic adsorbent. Using the as-prepared magnetic adsorbent (Fe3O4@COF-TpTU), we developed a simple, efficient, and sensitive magnetic solid-phase extraction-high performance liquid chromatography-inductively coupled plasma-mass spectrometry (MSPE-HPLC-ICP-MS) for the enrichment and determination of mercury species, including Hg2+, methylmercury (MeHg), and ethylmercury (EtHg). The effects of the experimental parameters on the extraction efficiency, including solution pH, adsorption and desorption time, composition and volume of the elution solvent, salinity, coexisting ions, and dissolved organic matter, were comprehensively investigated. Under optimised conditions, the limits of detection in the developed method were 0.56, 0.34, and 0.47 ng L-1 with enrichment factors of 190, 195, and 180-fold for Hg2+, MeHg, and EtHg, respectively. The satisfactory spiked recoveries (97.0-103%) in real water samples and high consistency between the certified and determined values in a certified reference material demonstrate the high accuracy and reproducibility of the developed method. The as-proposed method with simple operation, high sensitivity, and excellent anti-matrix interference performance was successfully applied to the enrichment and determination of trace levels of mercury species in the natural samples with complicated matrices, such as underground water, surface water, seawater and biological samples.

Simultaneous quantification of tin and lead species in Antarctic krill and fish by interfacing high-performance liquid chromatography with inductively coupled plasma mass spectrometry based on strong cation-exchange and Amphion columns.

Tin and lead are a global concern considering their species-dependent toxicity, bioavailability and transformation. Simultaneous speciation analysis of tin and lead is challenging for a large food capacity containing unstable species. Herein, we developed two sensitive methods for rapid quantification of tin and lead species in Antarctic seafood by high-performance liquid chromatography and inductively coupled plasma mass spectrometry based on strong cation-exchange and Amphion columns. Inorganic tin and lead, four organotin and two organolead compounds can be analysed in 16 min on a 10-cm Amphion II column (mobile phase: 4 mM sodium dodecyl benzene sulfonate at pH 2.0) with 0.02-0.24 µg L-1 detection limits. The method was applied to Antarctic krill and fish, demonstrating the presence of any tin and lead species down to µg kg-1 level. Overall, the proposed methods are sensitive, efficient and environment-friendly for routine speciation analysis of tin and lead in food samples.

Characterization of halogen species in seaweeds from the Antarctic using a multi-technique approach.

BACKGROUND: Despite the recognized importance, the determination of halogens in Antarctic seaweeds remains understudied. Limited research exists due to challenges associated with sample preparation, and reliable analytical techniques for this type of analysis. Therefore, further investigations are necessary to bridge this knowledge gap and gain a comprehensive understanding of halogen metabolism in Antarctic seaweeds. METHODS: In this study, seaweeds from the coast of the Antarctic continent were characterized concerning the total content of halogens and their species. For this purpose, different sample preparation methods, based on extraction and combustion, combining highly selective and sensitive chromatographic and spectrometric multi-technique approaches were used. RESULTS: By using optimized methods, it was possible to determine total halogens content, the distribution of bromine and iodine in different classes of species (lipids, water-soluble, proteins, carbohydrates, and residue), as well as the identification of iodinated amino acids (MIT and DIT) in ten brown and red seaweeds. Bromate and iodate were not detected in the samples, which presented only bromide and iodide species in their composition. Additionally, unknown bromine and iodine species were observed in different extracts evaluated. Furthermore, 25 halogenated polyphenols were identified in seaweeds, of which only four were already reported in the literature. CONCLUSION: The results obtained in this study comprise unprecedented data in the literature on species of halogens present in seaweeds from the Antarctic environment.

Detection of inorganic arsenic in rice using a field-deployable method with Cola extraction.

Rice is a staple food and known to accumulate inorganic arsenic (iAs), which is a class 1 carcinogen to humans. Arsenic field-deployable method kits, designed for water testing, are able to screen iAs in rice, to assure food safety and quick decision-making without the need for laboratory analysis. For the arsenic extraction within the field method, nitric acid is used. To make the field method on-site safer, cost-effective and easier to handle, the method was adapted using a Cola in the extraction process. The adapted field-deployable method was tested by screening a total of 30 rice and rice products from the Austrian market. To verify the results obtained by the Cola extraction field-deployable method, the obtained iAs concentration was compared to HPLC-ICP-MS results. The Cola extraction field method obtained an LOD of 39 µg iAs kg-1 rice, and with an average reproducibility of 14% RSD, the method was capable of recording no false-negative but 7% false-positive values at the 2023 updated European Commission (EC) limits for rice. All, but one, screened rice samples were within the EU limits for iAs in rice and rice products.

A novel analytical approach for the determination of ethylene-thiourea and propylene-thiourea in vegetal foodstuffs by high-performance liquid chromatography hyphenated to inductively coupled plasma-tandem mass spectrometry.

This study reports a novel analytical approach for the simultaneous determination of ethylene-thiourea (ETU) and propylene-thiourea (PTU) in fruits and vegetables by (reverse phase) high-performance liquid chromatography (HPLC) coupled to inductively coupled plasma-tandem mass spectrometry (ICP-QQQMS or ICP-MS/MS). A baseline separation of ETU and PTU was achieved in less than 5 min. A robust method validation by using the accuracy profile approach was performed by carrying out four measurement series in duplicate at six different levels over a timespan of 4 weeks (different days). The recovery factors ranged from 87 to 101% for ETU and from 98 to 99% for PTU (depending on the spiking level). The coefficient of variation in terms of repeatability (CVr) ranged from 1 to 4.7% for ETU and from 1.8 to 3.9% for PTU (depending also on the analyte level) while the coefficient of variation in terms of intermediate reproducibility (CVR) ranged from 3.4 to 10% for ETU and from 1.8 to 10.8% for PTU. The limit of quantification was 0.022 mg kg-1 (wet weight) for ETU and 0.010 mg kg-1 (ww) for PTU. This novel approach was proved to be highly robust and suitable for the determination of ETU and PTU in foodstuffs of vegetal origin.

Methylation and bio-accessibility assessment of arsenate in crickets (Gryllusbimaculatus).

The ability of an organism to biomethylate toxic inorganic arsenic (As) determines both, the amount of As available for uptake higher up the food chain and the toxicity of bioavailable As. An exposure study was conducted to determine ability of farmed crickets to metabolize dietary arsenate. Crickets were exposed to 1.3 ± 0.1, 5.1 ± 2.5 and 36.3 ± 5.6 mg kg-1 dietary arsenate and quantitation of total As showed retention of 0.416 ± 0.003, 1.3 ± 0.04 and 2.46 ± 0.09 mg kg-1, respectively. Speciation analysis revealed that crickets have well developed ability to biomethylate dietary arsenate and the most abundant methylated As compound was DMA followed by MMA, TMAO and an unknown compound. Arsenobetaine, although present in all feed, control and As-rich, was measured only in the control crickets. To assess the bio-accessibility of the As species, crickets were subjected to simulated gastrointestinal digestion. The results showed that majority of As was extracted in saliva, followed by gastric and intestinal juice, which mass fraction was equal to residue. Over 78% of total As was shown to be bio-accessible with methylated species reaching 100% and iAs over 79% bio-accessibility. Additionally, arsenite and arsenate have shown different distributions between sequential leachate solutions. Bioaccumulation of As was observed in the studied crickets although it does not seem to occur to the same extent at higher exposure levels.

Acclimation to medium-level non-lethal iron limitation: Adjustment of electron flow around the PSII and metalloprotein expression in Trichodesmium erythraeum IMS101.

The aim of this study was to investigate how acclimation to medium-level, long-term, non-lethal iron limitation changes the electron flux around the Photosystem II of the oceanic diazotroph Trichodesmium erythraeum IMS101. Fe availability of about 5× and 100× lower than a replete level, i.e. conditions common in the natural environment of this cyanobacterium, were applied in chemostats. The response of the cells was studied not only in terms of growth, but also mechanistically, measuring the chlorophyll fluorescence of dark-adapted filaments via imaging fluorescence kinetic microscopy (FKM) with 0.3 ms time resolution. Combining these measurements with those of metal binding to proteins via online coupling of metal-free HPLC (size exclusion chromatography SEC) to sector-field ICP-MS allowed to track the fate of the photosystems, together with other metalloproteins. General increase of fluorescence has been observed, with the consequent decrease in the quantum yields φ of the PSII, while the efficiency ψ of the electron flux between PSII and the PSI remained surprisingly unchanged. This indicates the ability of Trichodesmium to cope with a situation that makes assembling the many iron clusters in Photosystem I a particular challenge, as shown by decreasing ratios of Fe to Mg in these proteins. The negative effect of Fe limitation on PSII may also be due to its fast turnover. A broader view was obtained from metalloproteomics via HPLC-ICP-MS, revealing a differential protein expression pattern under iron limitation with a drastic down-regulation especially of iron-containing proteins and some increase in low MW metal-binding complexes.

Simultaneous speciation analysis of Hg and Se in fish by high-performance liquid chromatography and inductively coupled plasma-mass spectrometry following microwave-assisted enzymatic hydrolysis.

This study reports the development and validation of a new analytical method for simultaneous speciation analysis of Se and Hg in fish muscle. For this purpose, four Se species (selenite/Se(IV), selenate/Se(VI), selenomethionine/SeMet, and selenocysteine/SeCys) and two Hg species (inorganic mercury/iHg and methylmercury/MeHg) were extracted simultaneously by microwave-assisted enzymatic hydrolysis and then separated by HPLC in less than 15 min by using a column with both anion and cation exchange mechanisms and a mobile phase consisting of a mixture of methanol 5% (v/v), 45 mM HNO3, 0.015% 2-mercaptoethanol, and 1.5 mM sodium 3-mercapto-1-propanesulfonate. The separated species of Hg and Se were detected online by inductively coupled plasma-mass spectrometry (ICP-MS). The speciation analysis method was validated by means of the accuracy profile approach by carrying out three series of measurements in duplicate on three different days over a time-span of 3 weeks. The limits of quantification (LOQ) are in the range of 0.010-0.013 mg/kg wet weight (ww) for all selenium species, except for Se(IV) (0.15 mg/kg ww), while the coefficient of variation in terms of intermediate reproducibility (CVR) was < 7%. The LOQ for MeHg was 0.006 mg/kg ww, while the CVR was 3%. The method was successfully applied to the analysis of muscle samples from four different fish species: rainbow trout, tuna, swordfish, and dogfish.

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

BACKGROUND: Aquaculture 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. METHODS: Atlantic 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. RESULTS: The 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. CONCLUSION: Although 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 Contents, Speciation and Toxicity in Germinated Rice Alleviated by Selenium.

Rice can accumulate more organic and inorganic arsenic (iAs) than other crop plants. In this study, the localization of As in rice grains was investigated using High Performance Liquid Chromatography-Inductively Coupled Plasma Mass Spectrometry (HPLC-ICP-MS) based on 26 rice varieties collected from two provinces. In all the samples, the total As contents in polished rice were 0.03-0.37 mg/kg, with average values of 0.28 and 0.21 mg/kg for two sample sets. The results of the determination of arsenic speciation in different components of rice grain showed that in the polished and brown rice the mean value of arsenite (As(III)) was nearly twice than that of arsenate (As(V)). The regional difference was observed in both total As contents and As speciation. The reason may be that As(III) is more mobile than As(V) in a dissociated form and because of soil properties, rice varieties, and the growing environment. The proportion of iAs and the total As in rice bran was higher than that in polished rice, and this is because As tends accumulate between the husk and the endosperm. In our study, selenium could alleviate the risk of arsenic toxicity at the primary stage of rice growth. Co-exposure to As and Se in germinated rice indicated that the reduction in As accumulation in polished rice reached 73.8%, 76.8%, and 78.3% for total As, As(III), and As(V) when compared with rice treated with As alone. The addition of Se (0.3 mg/kg) along with As significantly reduced the As amount in different parts of germinated rice. Our results indicated that Se biofortification could alleviate the As accumulation and toxicity in rice crops.

Synthesis of a novel magnetic nanomaterial for the development of a multielemental speciation method of lead, mercury, and vanadium via HPLC-ICP MS.

A new magnetic functionalized material based on graphene oxide magnetic nanoparticles named by us, M@GO-TS, was designed and characterized in order to develop a magnetic solid-phase extraction method (MSPE) to enrich inorganic and organic species of lead, mercury, and vanadium. A flow injection (FI) system was used to preconcentrate the metallic and organometallic species simultaneously, while the ultra-trace separation and determination of the selected species were achieved by high-performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP MS). Therefore, preconcentration and separation/determination processes were automated and conducted separately. To the best of our knowledge, this is the first method combining an online MSPE and HPLC-ICP MS for multielemental speciation. Under the optimized conditions, the enrichment factor obtained for PbII, trimethyllead (TML), HgII, methylmercury (MetHg), and VV was 27. The calculated LOD for all studied species were as follows: 5 ng L-1, 20 ng L-1, 2 ng L-1, 10 ng L-1, and 0.4 ng L-1, respectively. The RSD values calculated with a solution containing 0.5 µg L-1 of all species were between 2.5 and 4.5%. The developed method was validated by analyzing Certified Reference Materials TMDA 64.3 for total concentration and also by recovery analysis of the species in human urine from volunteers and a seawater sample collected in Málaga. The t statistical test showed no significant differences between the certified and found values for TMDA 64.3. All the recoveries obtained from spiked human urine and seawater samples were close to 100%. All samples were analyzed using external calibration. The developed method is sensitive and promising for routine monitoring of the selected species in environmental waters and biological samples.

Selenium Biofortification Enhanced Grain Yield and Alleviated the Risk of Arsenic and Cadmium Toxicity in Rice for Human Consumption.

Arsenic (As) and Cadmium (Cd) are toxic to rice plants. However, selenium (Se) has the potential to regulate As and Cd toxicity. The present study aimed to evaluate the co-exposure to As5+ and Se6+ species in two rice cultivars, BRS Pampa and EPAGRI 108. The plants were divided into six groups and cultivated until complete maturation of the grains, under greenhouse conditions. Regarding total As and inorganic As (i-As) accumulation in grains, the highest concentrations were found for BRS Pampa. For Se, EPAGRI 108 presented the highest concentration of inorganic and organic Se (i-Se and o-Se). The exposure assessments showed that Se biofortification can mitigate the As accumulation in rice and, consequently, the risk of As and Cd toxicity in grains for human consumption. The combined effect of As and Se in rice plants could represent an alternative to biofortify this food in a safe way and with a higher percentage of bioavailable Se. Although Se is able to mitigate As toxicity in rice plants, in the present study we showed that co-exposure in different cultivars under the same growing conditions may present different responses to As and Se exposure.