Characterisation of titanium dioxide (nano)particles in foodstuffs and E171 additives by single particle inductively coupled plasma-tandem mass spectrometry using a highly efficient sample introduction system.

This study addressed primarily the characterisation and quantification of titanium dioxide (TiO2) (nano)particles (NPs) in a large variety of commercial foodstuffs. The samples were purchased from local markets in Spain before the ban of TiO2 food additive (E171) in the EU. The analyses were carried out by single particle inductively coupled plasma-tandem mass spectrometry (spICP-MS/MS) in mass shift mode (oxidation of 48Ti to 48Ti16O (m/z = 64)) and using a highly efficient sample introduction system (APEX™ Ω). This novel analytical approach allowed accurate characterisation of a large panel of TiO2 NPs sizes ranging from ∼12 to ∼800 nm without isobaric interferences from 48Ca isotope, which is highly abundant in most of the analysed foodstuffs. TiO2 NPs were extracted from foodstuffs using sodium dodecyl sulphate (0.1%, w/v) and diluted with ultra-pure water to reach ∼ 1000 particles signals per acquisition. All the analysed samples contained TiO2 NPs with concentrations ranging from 1010 to 1014 particles kg-1, but with significant low recoveries compared to the total Ti determination. A selection of samples was also analysed using a similar spICP-MS/MS approach with a conventional sample introduction system. The comparison of results highlighted the improvement of the limit of detection in size (12 nm) by the APEX™ Ω system, providing nanoparticulate fractions ranging from ∼4% (cheddar sauce) up to ∼87% (chewing gum), which is among the highest nanoparticulate fractions reported in literature using a spICP-MS approach. In addition, two commercially available E171 additives were analysed using the previous approaches and other techniques in different European laboratories with the aim of methods inter-comparison. This study provides occurrence data related to TiO2 NPs in common commercial foodstuffs but it also demonstrates the potential of the novel analytical approach based on APEX™-ICP-MS/MS to characterise nano-size TiO2 particles in complex matrices such as foodstuffs.

Assessment of TiO2 (nano)particles migration from food packaging materials to food simulants by single particle ICP-MS/MS using a high efficiency sample introduction system.

TiO2 is the most widely used white pigment in plastics and food packaging industry, thus the question of its migration towards food and hence the impact on consumers is raised. Since recent research indicate its potential toxicity, it is necessary to study TiO2 contamination as a consequence of food storage. For this purpose, plastic containers from commercially-available dairy products and custom-made TiO2-spiked polypropylene materials were put in contact with 50% (v/v) ethanol and 3% (w/v) acetic acid, which were used here as food simulants. The migration assays were carried out under standard contact conditions of packaging use (as recommended by Commission Regulation (EU) N° 10/2011 for food contact migration testing), and under conditions of extreme mechanical degradation of the packaging. The TiO2 (nano)particles released in the food simulants were analysed by single particle inductively coupled plasma-tandem mass spectrometry in mass-shift mode and using a high efficiency sample introduction system (APEX™ Ω) to avoid matrix effects from food simulants. For the dairy product containers and for the spiked polypropylene, results showed release of TiO2 particles of rather large sizes (average size: 164 and 175 nm, respectively) under mechanical degradation conditions, i.e. when the polymeric structure is damaged. The highest amounts of TiO2 were observed in 50% ethanol after 10 days of storage at 50 °C (0.62 ng cm-2) for the dairy product containers and after 1 day of storage at 50 °C (0.68 ng cm-2) for the spiked polypropylene. However, the level of Ti released in particle form was very small compared to the total Ti content in the packaging and far below the acceptable migration limits set by European legislation. Release under standard contact conditions of use of the container was not measurable, thus the migration of TiO2 particles from this packaging to dairy products among storage is expected to be negligible.

The Impact of Various Types of Cooking on the Fate of Hg and Se in Predatory Fish Species.

This study addresses the effect of various cooking approaches on total Hg (HgT) and total Se (SeT) contents in three predatory fish species. For this purpose, samples of swordfish, dogfish, and tuna from regular French (fish) markets were cooked by boiling, steaming, grilling, and frying, respectively. The levels of HgT and SeT in raw and cooked samples were determined by inductively coupled plasma-mass spectrometry. The data showed a significant increase in HgT and SeT levels between raw and cooked samples (33% of the samples for SeT and 67% for HgT) due to the water loss during the cooking. High intra-species variation related to HgT and SeT levels was found. Considering the level of exposure to HgT through fish consumption and taking also into account the possible protective effect of Se (expressed here via the Se/Hg molar ratio), the safest cooking approach corresponds to grilled swordfish, fried tuna, and steamed dogfish, which show Se/Hg molar ratios of (1.0 ± 0.5), (4.3 ± 4.2), and (1.0 ± 0.6), respectively.

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.

Trace elements and arsenic speciation in Paracentrotus lividus from North-West Mediterranean Sea.

Levels of 14 trace elements (Al, Ti, V, Cr, Mn, Co, Ni, Zn, As, Se, Cd, Sn, Hg, and Pb) and 5 arsenic species (arsenite/As(III), arsenate/As(V), monomethylarsonic acid/MA, dimethylarsinic acid/DMA, and arsenobetaine/AsB) were assessed in the gonads of sea urchin samples (Paracentrotus lividus) from North-West Mediterranean Sea (French coast). The samples were collected from 13 sites characterized by different types and levels of chemical contamination. Trace elements levels were measured by inductively coupled plasma-mass spectrometry (ICP-MS) following microwave (acid) digestion in a closed system, whereas As speciation analysis was carried out by ion-exchange liquid chromatography coupled with ICP-MS after microwave-assisted extraction. High levels of trace elements were found in sea urchins sampled from Corsica whereas the samples from Théoule were found to be the least contaminated. From all the analyzed urchin samples, none showed Cd, Hg, or Pb concentrations above the regulatory levels set by the European Community (EC) No. 1881/2006 for seafood or bivalve mollusks. Regarding arsenic speciation, AsB was confirmed to be the predominant species. Inorganic As (As(III) + As(V)) was mainly constituted by As(III), which was quantified in all samples. Methylated As forms (MA and DMA) represent 9 to 23% of the total As.

Trace elements, dioxins and PCBs in different fish species and marine regions: Importance of the taxon and regional features.

Chemical contaminant concentrations in wild organisms are used to assess environmental status under the European Marine Strategy Framework Directive. However, this approach is challenged by the complex intra- and inter-species variability, and the different regional features. In this study, concentrations in trace elements (As, Cd, Hg and Pb), polychlorinated biphenyls (PCBs), polychlorodibenzo-para-dioxines (PCDDs) and polychlorodibenzofuranes (PCDFs) were monitored in 8 fish species sampled on the continental shelf of three French regions: the Eastern English Channel (EEC) and Bay of Biscay (BoB) in the Northeast Atlantic Ocean, and the Gulf of Lions (GoL) in Western Mediterranean Sea. Our objectives were to identify species or regions more likely to be contaminated and to assess how to take this variability into account in environmental assessment. While concentrations were higher in benthic and demersal piscivores, PCB and PCDD/F concentrations (lipid-weight) were similar in most teleost species. For Cd, Hg and Pb, the trophic group accumulating the highest concentrations depended on the contaminant and region. Concentrations in Hg, PCBs and PCDD/Fs were higher in the EEC and/or GoL than in BoB. Cadmium and Pb concentrations were highest in the BoB. Lipid content accounted for 35%-84% of organic contaminant variability. Lipid normalisation was employed to enhance robustness in the identification of spatial patterns. Contaminant patterns in chondrichthyans clearly differed from that in teleosts. In addition, trophic levels accounted for ≤1% and ≤33% of the contaminant variability in teleost fishes in the EEC and BoB, respectively. Therefore, developing taxa-specific thresholds might be a more practical way forward for environmental assessment than normalisation to trophic levels.

Probabilistic determination of a maximum acceptable level of contaminant to reduce the risk of overexposure for a novel or emerging food: the case of cadmium in edible seaweed in the French population.

European and French populations are overexposed to cadmium (Cd) through their foods. The risk of increased cadmium exposure for consumers needs to be limited by reduced maximum limits (ML) for novel foodstuffs such as edible seaweed in France. The objective was to derive a low and protective cadmium concentration in edible seaweeds to limit cadmium overexposure in consumers. To do so, we applied a probabilistic approach to the data collected on French seaweed consumers, taking into account other sources of exposure for cadmium. This approach led to the identification of a cadmium concentration which should ensure that the seaweed-consuming population does not exceed the tolerable daily intake (TDI) of cadmium according to a probability of cases, when simultaneously exposed to other cadmium dietary sources. Considering the 5% of the population exceeding TDI, the estimated ML is equal to 0.35 mg Cd kg-1 dry matter of seaweed as an unprocessed food with a 95% confidence interval of [0.18,1.09]. The proposed approach is generic and could be applied to other relevant food/substance pairs when considering the setting of MLs in the regulatory system. It ensures better protection of consumer health.

Ultra-trace speciation analysis of Cr(III) and Cr(VI) in rice using species-specific isotope dilution and HPLC-ICP-MS.

The study aims to clarify the current controversy related to conflicting reports on whether presence of Cr(VI) in rice is possible or not. For this purpose, a method was employed for the single run speciation analysis of Cr(III) and Cr(VI) in rice samples using species-specific isotope dilution (SS-ID) and high performance liquid chromatography coupled to inductively coupled mass-spectrometry (HPLC-ICP-MS) and selective single run species complexation/derivatisation. The quantification limits (LOQs) were 0.014 µg kg-1 for Cr(III) and 0.047 µg kg-1 for Cr(VI), while the detection limits (LODs) were 0.004 and 0.014 µg kg-1 for Cr(III) and Cr(VI), respectively. A total of 10 rice samples of different origin and colour (depending on the type of industrial processing) were analysed in this study. The content of Cr(VI) was below the limit of quantification in all of the rice samples analysed, while the Cr(III) levels ranged between 0.59 (whole grain rice) up to 104 µg kg-1 (brown rice). All samples were also analysed for their total Cr (Crtotal) content by ICP-MS solely and the results were in all cases comparable with the Cr(III) levels determined in the same samples. To assess the stability of Cr(III) and Cr(VI) in rice, one sample was spiked with Cr(III) and Cr(VI) (individually) at different levels (5.0, 10, 15 and 20 µg kg-1), held for 2 h, and then analysed by SS-ID HPLC-ICP-MS. The results showed a complete reduction of Cr(VI) to Cr(III), while Cr(III) remained stable at all spiking levels. These findings support the general statement from the European Food Safety Authority related to the complete absence of Cr(VI) in foods and confirms that Cr in rice is found solely as Cr(III) species.

Chromium speciation analysis in raw and cooked milk and meat samples by species-specific isotope dilution and HPLC-ICP-MS.

This study aimed at the assessment of the impact of various culinary processes on the fate of chromium (Cr) species (Cr(III) and Cr(VI)) in infant formula milk, semi-skimmed milk and bovine meat samples. The cooking procedures were boiling at 70°C/100°C (milk samples) and frying without and with oil (95°C and 120°C) (bovine meat). The levels of Cr(III) and Cr(VI) in raw and cooked samples were determined by high-performance liquid chromatography (HPLC) coupled to inductively coupled plasma mass spectrometry (ICP-MS) using double spike species-specific-isotope dilution (SS-ID). The species were extracted by sequential complexation of Cr(III) with ethylenediaminetetraacetic acid and of Cr(VI) with 1,5-diphenylcarbazide in the same analytical run by heating at 70°C for 50 min. Anion exchange chromatography using a Dionex IonPac™ AG7 column and a mobile phase consisting of 10 mM HNO3, 2.5% MeOH and 30 mM EDTA at pH 2 was employed for species separation. The quantification limits were 0.013 and 0.049 µg kg-1 for Cr(III) and Cr(VI), respectively. ANOVA test used to compare the mean Cr species concentrations showed no significant differences between raw and cooked samples. The results obtained in the present study show that oxidation of Cr(III) to Cr(VI) does not occur during thermal cooking of milk and bovine meat samples. A selection of 10 samples of each type were analysed in terms of total Cr (Crtotal) as well as speciation (Cr(III) and Cr(VI)). Cr(VI) was not quantified in any of these samples, whereas Cr(III) levels ranged from 0.22 (infant formula milk) up to 80 µg kg-1 (chorizo sausage). Additionally, Cr(III) and Crtotal levels were comparable hence demonstrating that in the samples analysed in this study, Cr is found exclusively as Cr(III) species.

Optimisation and application of an analytical approach for the characterisation of TiO2 nanoparticles in food additives and pharmaceuticals by single particle inductively coupled plasma-mass spectrometry.

This study was designed to optimise an analytical method for characterising TiO2 nanoparticles (NPs) in food additives and pharmaceuticals by inductively coupled plasma-mass spectrometry in single particle mode (spICP-MS). Several parameters, including transport efficiency (TE), were assessed and optimised using the NM-100 reference material. We found that self-aspiration for sample intake and use of the concentration-based method for TE was optimal for characterising TiO2 NPs. No spectral interference was observed with either 49Ti or 48Ti isotopes. The optimised Excel spreadsheet developed for this study not only provided additional parameters but gave results closer to the NM-100 reference value than the ICP-MS software. The method was then applied to the analysis of a selection of food samples and pharmaceuticals. The average diameter of TiO2 particles ranged from 86 to 179 nm in the food samples and from 131 to 197 nm in the pharmaceuticals, while the nanoparticular fraction was between 19 and 68% in food, and between 13 and 45% in pharmaceuticals.

Development and validation of a single run method based on species specific isotope dilution and HPLC-ICP-MS for simultaneous species interconversion correction and speciation analysis of Cr(III)/Cr(VI) in meat and dairy products.

This study presents the development, validation and application of a new analytical approach for the simultaneous speciation analysis of Cr(III) and Cr(VI) in meat and dairy products by high-performance liquid chromatography (HPLC) coupled to inductively coupled plasma mass spectrometry (ICP-MS) and double spike species specific-isotope dilution (SS-ID). The species extraction was achieved by sequential complexation of Cr(III) with ethylenediaminetetraacetic acid (EDTA) and of Cr(VI) with 1,5-diphenylcarbazide (DPC) in the same analytical run. The HPLC separation of complexed species was carried out using a short (5 cm) microbore anion-exchange HPLC column and a mobile phase consisting of 0.01 mol L-1 HNO3 + 2.5% (v/v) MeOH + 0.30 mol L-1 EDTA (pH = 2) in isocratic elution mode with excellent baseline separation achieved in less than 3 min. The method was validated by means of the accuracy profile approach by carrying out 6 measurement series in duplicate on (six) different days over a timespan of two months. The quantification limit was 0.013 µg kg-1 for Cr(III) and 0.049 µg kg-1 for Cr(VI), respectively. The measurement bias corresponding to the validity domain ranged from 0.01 to 0.11%, whereas the coefficient of variation in terms of repeatability (CVr) varied from 2.9 to 11.6% (depending on the analyte level) for Cr(III) and from 6.7 to 11.8% for Cr(VI). Similarly, the coefficient of variation in terms of intermediate reproducibility (CVR) ranged from 6.8 to 13% for Cr(III) and from 6.8 to 25.9% for Cr(VI), respectively. The method was successfully applied to the analysis of a selection of food samples such as baby and semi-skimmed milk and steak beef samples. Cr(VI) was not quantified in any of these samples while Cr(III) levels ranged between 2.7 and 4.7 µg kg-1, which were comparable with the levels of total chromium analysed in the same samples by ICP-MS (accredited method). The method presented here with combined use of species specific isotope dilution and sequential species complexation is a powerful analytical tool for accurate and precise quantification of Cr(III) and Cr(VI) at trace levels and allows for correction of any species interconversion during sample preparation.

Human dietary exposure to chemicals in sub-Saharan Africa: safety assessment through a total diet study.

BACKGROUND: Human dietary exposure to chemicals can result in a wide range of adverse health effects. Some substances might cause non-communicable diseases, including cancer and coronary heart diseases, and could be nephrotoxic. Food is the main human exposure route for many chemicals. We aimed to assess human dietary exposure to a wide range of food chemicals. METHODS: We did a total diet study in Benin, Cameroon, Mali, and Nigeria. We assessed 4020 representative samples of foods, prepared as consumed, which covered more than 90% of the diet of 7291 households from eight study centres. By combining representative dietary surveys of countries with findings for concentrations of 872 chemicals in foods, we characterised human dietary exposure. FINDINGS: Exposure to lead could result in increases in adult blood pressure up to 2·0 mm Hg, whereas children might lose 8·8-13·3 IQ points (95th percentile in Kano, Nigeria). Morbidity factors caused by coexposure to aflatoxin B1 and hepatitis B virus, and sterigmatocystin and fumonisins, suggest several thousands of additional liver cancer cases per year, and a substantial contribution to the burden of chronic malnutrition in childhood. Exposure to 13 polycyclic aromatic hydrocarbons from consumption of smoked fish and edible oils exceeded levels associated with possible carcinogenicity and genotoxicity health concerns in all study centres. Exposure to aluminium, ochratoxin A, and citrinin indicated a public health concern about nephropathies. From 470 pesticides tested across the four countries, only high concentrations of chlorpyrifos in smoked fish (unauthorised practice identified in Mali) could pose a human health risk. INTERPRETATION: Risks characterised by this total diet study underscore specific priorities in terms of food safety management in sub-Saharan Africa. Similar investigations specifically targeting children are crucially needed. FUNDING: Standards and Trade Development Facility.

Seasonal and Spatial Variability of Trace Elements in Livers and Muscles of Three Fish Species from the Eastern Mediterranean.

Levels of 20 trace elements (Al, Ti, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, As, Se, Ag, Cd, Sn, Sb, Ba, Hg, Pb, and U) were assessed in livers and muscles of two demersal fish species (Siganus rivulatus and Lithognathus mormyrus) and one pelagic species (Etrumeus teres) from the Lebanese coast located in the Eastern Mediterranean. The samples were collected from three sites along the Lebanese coast during the wet and dry seasons in 2017. The trace elements were more concentrated in livers than in muscles and interspecific differences were also found. The herbivorous species S. rivulatus showed the highest levels for most trace elements, while the carnivorous species L. mormyrus showed the least contamination. Elemental seasonal differences were species dependent and were observed for Ti, Cr, Mn, Fe, Ni, As, and Hg, with higher values during the wet season. Multivariate analysis showed spatial differences mainly during the wet season, while being closely related to species that reflected different accumulation patterns in each site. Levels of most trace metals in livers were higher than those reported in other Mediterranean regions (up to 2 to 3 folds). Nevertheless, the levels of Cd, Pb, and Hg in fish muscle were below the maximum levels set by the European Commission indicating that the consumption of these fish species is not likely to have adverse effect on human health. However, exposure depends on dietary habits of the population and a continuous exposure to these elements may result in adverse effects.

Occurrence of 30 trace elements in foods from a multi-centre Sub-Saharan Africa Total Diet Study: Focus on Al, As, Cd, Hg, and Pb.

This paper reports occurrence data related to 30 trace elements in food composite samples from a multi-regional Sub-Saharan Africa Total Diet Study. Herein, 2700 samples grouped in 225 food composite samples corresponding to 13 food groups: cereals, tubers, legumes, vegetables, fruits, nuts/seeds, meat, eggs, fish, milk/dairy, oil/fats, and beverages from eight locations in four countries, namely Benin (Littoral/Borgou), Cameroon (Duala/North), Mali (Bamako/Sikasso), and Nigeria (Lagos/Kano) were prepared as consumed, pooled, and analysed using a validated method based on inductively coupled plasma-mass spectrometry. The occurrence data for Al, As, Cd, Hg, and Pb as regulated by the Codex Alimentarius are discussed herein. Although the levels of As, Cd, Hg, and Pb were above the limit of quantification, they were below the maximum limits set by the Codex in most samples analysed. A distinct feature was observed for cereals and tubers, as they were mostly contaminated with Al and Pb. A pilot study regarding the impact of using artisanal cookware (made from recycled aluminium) on the contamination of food samples was performed. Relevant contamination with Al and Pb when cooking tomato samples from Cameroon and Nigeria using artisanal aluminium cookware was compared to that when cooked using stainless-steel.

Development and application of a method for Cr(III) determination in dairy products by HPLC-ICP-MS.

This study describes the development of an analytical approach for the determination of Cr(III) in dairy products by microwave assisted extraction, complexation in situ by ethylenediaminetetraacetate (EDTA) and high performance liquid chromatography hyphenated to inductively coupled plasma-mass spectrometry (HPLC-ICP-MS). The extraction step was optimised by using an experimental design. A limit of quantification of 38µgkg-1dry weight (d.w.) was obtained whereas the bias (%) measured ranged from 10 to 18%. The repeatability and intermediate precision varied between 1.2-5.0% and 7.5-13.5%, respectively. The method was applied to the analysis of several dairy samples beforehand characterized in terms of Cr(VI) and total chromium (Crtotal). Cr(III) concentrations ranged from <13 to 255µgkg-1d.w. The results showed a good agreement between Cr(III) and Crtotal concentration levels.

Copper and cobalt mobility in soil and accumulation in a metallophyte as influenced by experimental manipulation of soil chemical factors.

The influence of Fe oxides, Mn oxides and organic matter (OM) on the Cu and Co mobility in soil and accumulation in the metallophyte Anisopappus chinensis (Ac), as compared with Helianthus annuus (Ha), was experimentally investigated. Growth and accumulation response when increasing the exchangeable Cu and Co concentrations in soil were also investigated. Plants were cultivated on soil where concentrations of Cu, Co, Fe oxides, Mn oxides and OM content were varied according to 36 treatments. The OM supply decreased the Cu mobility and increased the Co mobility, resulting in decreasing the foliar Cu of Ac and increasing the foliar Co of Ha. The Fe oxides supply could increase the Cu accumulation for Ac, but was not verified for Ha. Compared with Ha, Ac increasingly accumulated Cu and Co without negative effect on plant growth while increasing Cu and Co mobility to phytotoxic concentrations. The results revealed promising perspectives for the use of Ac in Cu-contaminated environment phytoremediation applications.

Selenium biochemistry and its role for human health.

Despite its very low level in humans, selenium plays an important and unique role among the (semi)metal trace essential elements because it is the only one for which incorporation into proteins is genetically encoded, as the constitutive part of the 21st amino acid, selenocysteine. Twenty-five selenoproteins have been identified so far in the human proteome. The biological functions of some of them are still unknown, whereas for others there is evidence for a role in antioxidant defence, redox state regulation and a wide variety of specific metabolic pathways. In relation to these functions, the selenoproteins emerged in recent years as possible biomarkers of several diseases such as diabetes and several forms of cancer. Comprehension of the selenium biochemical pathways under normal physiological conditions is therefore an important requisite to elucidate its preventing/therapeutic effect for human diseases. This review summarizes the most recent findings on the biochemistry of active selenium species in humans, and addresses the latest evidence on the link between selenium intake, selenoproteins functionality and beneficial health effects. Primary emphasis is given to the interpretation of biochemical mechanisms rather than epidemiological/observational data. In this context, the review includes the following sections: (1) brief introduction; (2) general nutritional aspects of selenium; (3) global view of selenium metabolic routes; (4) detailed characterization of all human selenoproteins; (5) detailed discussion of the relation between selenoproteins and a variety of human diseases.

Plasma selenoproteins concentrations in type 2 diabetes mellitus--a pilot study.

The association between the concentration/activity of selenium/selenoproteins in plasma and type 2 diabetes mellitus is still a matter of debate. This cross-sectional pilot study evaluates whether patients with diabetes present a different plasma selenoproteins status than a healthy control group and examines whether the introduction of clinical parameters allows the detection of correlations and further grouping criteria. For this purpose, the levels of plasma glutathione peroxidase (GPx), selenoprotein P (SelP), and seleno-albumin (SeAlb) present in 40 patients affected by type 2 diabetes mellitus were determined simultaneously and accurately by a newly developed analytical method. The results show that patients with diabetes demonstrate significantly lower levels of GPx and SeAlb with respect to healthy subjects (11 ± 3 ng/mL and 9 ± 2 ng/mL vs 18 ± 8 ng/mL and 11 ± 2 ng/mL, respectively). Significant negative correlations were revealed among GPx, SeAlb, and clinical parameters including fasting plasma glucose, hemoglobin A1c, and the albumin-to-creatinine ratio. Our findings suggest an association between the individual selenoproteins concentration and the presence of diabetes, including associated clinical parameters. It currently cannot be ascertained whether the altered selenoproteins status is a consequence or a causative factor for diabetes. This study demonstrates the potential of a method for individual selenoproteins determination for investigating the biochemical relationship between selenium and diabetes.

A systematic approach to the accurate quantification of selenium in serum selenoalbumin by HPLC-ICP-MS.

In this paper, two different methods are for the first time systematically compared for the determination of selenium in human serum selenoalbumin (SeAlb). Firstly, SeAlb was enzymatically hydrolyzed and the resulting selenomethionine (SeMet) was quantified using species-specific isotope dilution (SSID) with reversed phase-HPLC (RP-HPLC) hyphenated to (collision/reaction cell) inductively coupled plasma-quadrupole mass spectrometry (CRC ICP-QMS). In order to assess the enzymatic hydrolysis yield, SeAlb was determined as an intact protein by affinity-HPLC (AF-HPLC) coupled to CRC ICP-QMS. Using this approach, glutathione peroxidase (GPx) and selenoprotein P (SelP) (the two selenoproteins present in serum) were also determined within the same chromatographic run. The levels of selenium associated with SeAlb in three serum materials, namely BCR-637, Seronorm level 1 and Seronorm level 2, obtained using both methods were in a good agreement. Verification of the absence of free SeMet, which interferes with the SeAlb determination (down to the amino acid level), in such materials was addressed by analyzing the fraction of GPx, partially purified by AF-HPLC, using RP-HPLC (GPx only) and size exclusion-HPLC (SE-HPLC) coupled to CRC ICP-QMS. The latter methodology was also used for the investigation of the presence of selenium species other than the selenoproteins in the (AF-HPLC) SelP and SeAlb fractions; the same selenium peaks were detected in both control and BCR-637 serum with a difference in age of ca. 12 years. It is also for the first time that the concentrations of selenium associated with SeAlb, GPx and SelP species in such commercially available serums (only certified or having indicative levels of total selenium content) are reported. Such indicative values can be used for reference purposes in future validation of speciation methods for selenium in human serum and/or inter-laboratory comparisons.

Simultaneous speciation analysis of glutathione peroxidase, selenoprotein P and selenoalbumin in human serum by tandem anion exchange-affinity HPLC and on-line isotope dilution ICP-quadrupole MS.

A method based on anion exchange (AE) and affinity (AF)-HPLC (AE-AF-HPLC) hyphenated to inductively coupled plasma-(quadrupole) mass spectrometry (ICP-QMS) was developed for the speciation analysis of selenoprotein P (SelP), glutathione peroxidase (GPx) and selenoalbumin (SeAlb) in human serum. AE-HPLC is proposed here for the on-line alleviation of Cl and Br spectral interferences on (77)Se ((40)Ar(37)Cl) and (82)Se ((81)Br(1)H). Separation of GPx, SelP and SeAlb by AE-AF-HPLC was obtained within a total chromatographic runtime of <20 min. On-line (post-column) isotope dilution (ON-ID) and on-line external calibration (ON-EC)-ICP-QMS were used for the quantification of Se in GPx, SelP and SeAlb. ON-EC using a Se-L: -cystine standard was shown to be a suitable approach for the routine simultaneous speciation analysis of serum GPx, SelP and SeAlb. The method validation was carried out by direct ICP-sector field MS determination of Se in GPx, SelP and SeAlb fractions collected after AE-AF-HPLC separation. In addition, the method accuracy for the determination of total protein-bound Se was assessed by analyzing a human serum reference material (BCR-637) certified for total Se content.