Effect of selenium supplementation on thyroid antibodies

Selenium is an essential component of selenoproteins, enzymes with extensive regulatory and protective effect in organism. Immunological effects of Se are documented and are distinct even above concentrations necessary for maximal activity of selenoenzymes. Therefore, we investigated effect of supplementation by 100 μg of yeast-bound Se on concentrations of thyroid autoantibodies TPOAb and TgAb in the group of 253 seniors living in the Asylum Houses of South Bohemia. Increase of serum selenium from 59 to 150 μg Se/L serum in supplemented group and from 59 to 72 μg Se/L serum in group with placebo were detected by Instrumental Neutron Activation Analysis (INAA) and proved increased Se intake during the trial. Autoantibodies were analyzed by ELISA at the beginning of the trial and after 1 year. Statistical evaluation of results in whole groups (regardless of increased autoantibodies) by ANOVA manifested significant decrease of TPOAb and TgAb in non-supplemented group while supplementation did not effect serum autoantibodies concentrations. Evaluation of groups of seniors created from those with increased autoantibodies, ANOVA demonstrated decrease of TPOAb in both groups but Se supplementation did not affect the decrease. In opposite, TgAb increased significantly and Se supplementation led to higher increase of TgAb. Recent results of possibility to decrease serum concentration of TPOAb proved this effect only for high TPOAb concentrations and for higher Se supplements. From this point of view, it is necessary to conduct subsequent trials with the patients with autoimmune thyreoiditis with different levels of autoantibodies and detect also serum Se levels.     

Use of enriched <Superscript>74</Superscript>Se and <Superscript>77</Superscript>Se in combination with isotope pattern deconvolution to differentiate and determine endogenous and supplemented selenium in lactating rats

A quantitative methodology has been developed to differentiate between endogenous and supplemented selenium in lactating rats using two enriched selenium isotopes. Lactating rats were fed for 2 weeks with formula milk containing one enriched Se isotope, ⁷⁷Se, as the metabolic tracer. The isotopic composition of selenium in serum and urine samples was then measured by collision cell ICP-MS after the addition of a solution containing another enriched isotope, ⁷⁴Se, as quantitation tracer, before analysis. Isotope pattern deconvolution allowed the transformation of measured Se isotopic abundances into concentrations of natural abundance (endogenous) selenium and enriched ⁷⁷Se (supplemented) present in the samples. The proposed methodology was validated using serum and urine reference materials spiked with both ⁷⁷Se and ⁷⁴Se. The obtained results are discussed in terms of selenium exchange and half-life in lactating rats (11–12 days) and selenium levels in serum in comparison with non-supplemented rats and control rats after maternal feeding.     

Profile of selenium in soil and crops in seleniferous area of Punjab,India by neutron activation analysis

In the Nawanshahr–Hoshiarpur region of Punjab, India, more than 1000 hectares of agricultural land is significantly affected by high levels of selenium (Se). Studies were carried out to examine Se levels in soil and crops such as wheat grains, wheat husk, rice, maize and mustard using neutron activation analysis. The Se concentrations in soil and crop products were found to be ranging from 2.7 to 6.5 mg kg⁻¹ and 13 to 670 mg kg⁻¹, respectively, indicating significantly high selenium in these crop products. Two reference materials were analysed for Se contents by INAA as controls.     

The determination of selenium in urine

The selenium excreted in urine can be measured to assess the dietary status of selenium, an essential trace element in human nutrition. The objectives of this work were: 1) to develop a procedure, capable of high sample throughout, by which the major interferences can be reduced such that selenium concentrations can be measured in urine by Neutron Activation Analysis (NAA) using^77mSe (17.4 s; and 2) to apply the method to a human dietary selenium study in which several selenium monitors were compared. The method involves a pre-irradiation arsenic-coprecipitation separation of the selenium from urine in the presence of a high specific-activity⁷⁵Se tracer. The processed urine samples are analyzed using NAA. The procedure was applied to 58 urine specimens longitudinally collected from 12 subjects consuming three different levels of selenium. A dose-response relationship was observed in urine as well as a high correlations with both serum and whole blood selenium concentrations.     

Applicability of direct total reflection X-ray fluorescence analysis for selenium determination in solutions related to environmental and geochemical studies

A significant amount of environmental studies related to selenium determination in different environmental compartments have been published in the last years due to the narrow range between the Se nutritious requirement as essential element and toxic effects upon exposure. However, the direct analysis of complex liquid samples like natural waters and extraction solutions presents significant problems related to the low Se concentrations and the complicated matrix of this type of samples.     

Frequency of concentrations of some trace elements in scalp hair by INAA

126 samples of scalp hair from inhabitants of Central Bohemia were searched for content of zinc, cobalt, cesium, chromium, scandium, selenium, and iron, for the purpose of medical research. Simultaneous determination of the elements was made by INAA. Inorganic standards were used for calculation of concentrations of elements. Quality assurance was checked by IAEA certified reference material and by internal standard mixed human serum. Statistical results are expressed in ppm, or ppb. Both normal and, if necessary, log-normal distributions of elements are presented, and types of distribution curves are verified statistically on 95% level of probability.     

Challenges in the accurate speciation analysis of selenium in humans: first report on indicative levels of selenoproteins in a serum certified reference material for total selenium (BCR-637)

Se is one of the most investigated essential trace elements in the past years, mostly due to its cancer prevention properties. Nevertheless, the accurate determination of its biologically active species, such as the selenoproteins (SeProt) in human serum, is currently a challenging task. This is because of the lack of appropriate quantification standards, certified reference materials (CRMs), and/or reference measurement methods. Additionally, most of the methods developed so far for the determination of SeProt were applied to the analysis of control (volunteers) serums, which are not available to other laboratories, therefore making methods inter-comparison virtually impossible. We present here for the first time indicative levels of SeProt in a commercially available human serum, namely the BCR-637 CRM with certified level of total Se. The concentrations of selenium associated with glutathione peroxidase (GPx), selenoprotein P (SelP) and selenoalbumin (SeAlb) in this serum were calculated using the results obtained by 13 different analytical methods (literature and non-published data) on the basis of (affinity) high-performance liquid chromatography (AF-HPLC) coupled to inductively coupled plasma-mass spectrometry (ICP-MS). The indicative levels of SeProt in the BCR-637 serum can be used for validation of methods dealing with the determination of these proteins in human serum.     

Variation of selenium concentration in toenails following long-term storage, washing and reactor irradiation

Summary Neutron activation analysis of toenails is often used to monitor body selenium in studies looking for an association between selenium deficiency and an increased cancer risk. In this study, 87 toenail samples were analyzed for Se four times, using neutron activation with 17.4-second ^77mSe, to determine the possible systematic effects of long-term storage, the washing procedure, and irradiation in a nuclear reactor. The mean Se concentration found was 0.92 mg/g, standard deviation 0.14 mg/g. The results showed that the Se concentrations are unaffected by washing and neutron irradiation, but the samples lost moisture during storage causing a 2% increase in the mean Se concentration.     

Gas chromatographic-mass spectrometric method for the determination of selenium in biological samples.

The determination of selenium by capillary gas chromatography-mass spectrometry (GC-MS), using an enriched stable isotope 76Se as internal standard, is described. Reference values for selenium in human biological fluids (serum, red blood cells and urine) are reported. With the advent of new compact capillary GC-MS (benchtop) instruments, this method will be very simple and accurate for routine analysis.     

Internal correction of spectral interferences and mass bias for selenium metabolism studies using enriched stable isotopes in combination with multiple linear regression

The analytical methodology for the in vivo study of selenium metabolism using two enriched selenium isotopes has been modified, allowing for the internal correction of spectral interferences and mass bias both for total selenium and speciation analysis. The method is based on the combination of an already described dual-isotope procedure with a new data treatment strategy based on multiple linear regression. A metabolic enriched isotope (⁷⁷Se) is given orally to the test subject and a second isotope (⁷⁴Se) is employed for quantification. In our approach, all possible polyatomic interferences occurring in the measurement of the isotope composition of selenium by collision cell quadrupole ICP-MS are taken into account and their relative contribution calculated by multiple linear regression after minimisation of the residuals. As a result, all spectral interferences and mass bias are corrected internally allowing the fast and independent quantification of natural abundance selenium (^natSe) and enriched ⁷⁷Se. In this sense, the calculation of the tracer/tracee ratio in each sample is straightforward. The method has been applied to study the time-related tissue incorporation of ⁷⁷Se in male Wistar rats while maintaining the ^natSe steady-state conditions. Additionally, metabolically relevant information such as selenoprotein synthesis and selenium elimination in urine could be studied using the proposed methodology. In this case, serum proteins were separated by affinity chromatography while reverse phase was employed for urine metabolites. In both cases, ⁷⁴Se was used as a post-column isotope dilution spike. The application of multiple linear regression to the whole chromatogram allowed us to calculate the contribution of bromine hydride, selenium hydride, argon polyatomics and mass bias on the observed selenium isotope patterns. By minimising the square sum of residuals for the whole chromatogram, internal correction of spectral interferences and mass bias could be accomplished. As a result, the tracer/tracee ratio could be calculated for each selenium-containing species and a time relationship for synthesis and degradation established. Both selenite and selenized yeast labelled with ⁷⁷Se were employed for comparative purposes.     

Selenium speciation in human serum of cystic fibrosis patients compared to serum from healthy persons

A formerly developed Se speciation method was applied to human serum. Chomatographic performance was checked regularly by measuring control standards after four sample run, each, to ensure sufficient separation of the species and sensitivity of the method even after a considerable number of serum samples. Detection limits of investigated species were similar to those reported and showed values below 1 microg/L related to Se for each Se compound. Sera of cystic fibrosis (CF) patients were investigated in comparison to sera from healthy volunteers with respect to total selenium and Se species. Generally, CF sera showed lower values of total Se, Selenocystine (SeC), and cationic/neutral Se compounds compared to serum of healthy persons. No significant gender-specific differences were found. Total Se in sera of healthy persons was determined at 102+/-12 microg/L (n = 12 individuals, mean value from male and female, age 4-38 years), whereas CF patients showed 58+/-10 microg/L (n = 31 individuals, mean value from male and female, age 3-35 years). Se-cystine showed significant differences between the CF and healthy group with a lowered SeC value in sera of CF patients by -75% (mean ca. 26 microg/L in healthy sera compared to about 6.5 microg/L (mean) in CF sera). A similar situation is seen for neutral/cationic Se compounds, which partly may comprise of Se proteins. The lowered SeC values together with lowered cationic/neutral Se compounds (probably Se enzymes) point to a Se-depleted regulated pathway combined with a reduced capability of protective functions such as protection from peroxides.     

The distribution of selenium and other elements in corn samples studied by NAA and a biochemical technique

A biochemical technique was used to separate three kinds of proteins (albumin, globulin and gliadin) in corn samples from high selenium areas and normal areas in Erxi autonomous region of Hubei Province, China. The contents of Se and other elements in these proteins were determined by neutron activation analysis (NAA). The results show that Se is enriched in corn proteins at high selenium area, while Cu, Al, Mn, V and Cl are also enriched in varying degrees.     

Laser ablation synthesis of selenium superoxide anion SeO4- via selenium trioxide photolysis. Time-of-flight mass spectrometry and ab initio calculations

Laser desorption/ionisation and laser ablation of solid selenium trioxide, as well as the gas-phase behaviour of selenium trioxide, were studied. Selenium trioxide undergoes photochemical decomposition and, from the mass spectra obtained by laser desorption/ionisation time-of-flight mass spectrometry (LDI-TOF-MS), the following species were identified: O-, O2-, O3-, SeO-, SeO2-, SeO3-, SeO4-, Se2O7-, Se3O11-, and Se4O14-. Formation of the selenium superoxide SeO4- anion is described in this work for the first time. In addition, low-abundance selenium species such as Se2O8H2-, Se3O11H-, and Se4O15H2- were also detected. The stoichiometry of all ions was confirmed via isotopic pattern modeling and/or post-source decay (PSD) analysis. Photolysis of selenium trioxide leads partly to ozone formation. It was found that the most likely mechanisms of selenium superoxide formation are oxidation of selenium trioxide with ozone and/or reactive oxygen radicals, or photolysis of selenium trioxide tetramer (SeO3)4. Therefore, ab initio calculations were performed to support the mass spectrometric evidence and to suggest probable geometries for selenium superoxide anion SeO4- and diselenium superoxide anion Se2O7-, as well as to provide insight into and/or predict possible formation pathways. It has been found that both cyclic and non-cyclic peroxide structures of SeO4- and Se2O7- ions are possible. In addition, the SeO4 structure was also calculated guided by thermodynamic considerations using Gaussian-2 methodology, and the inferred stability of the SeO4 neutral molecule was supported by ab initio calculations.     

Capillary electrophoresis on-line coupled with hydride generation-atomic fluorescence spectrometry for speciation analysis of selenium

A new method for speciation analysis of two inorganic selenium species was developed by on-line coupling of capillary electrophoresis (CE) with hydride generation-atomic fluorescence spectrometry (HG-AFS) and on-line conversion of Se(VI) to Se(IV). Baseline separation of Se(VI) and Se(IV) was achieved by CE in a 50 cm x 75 microm inside diameter (ID) fused-silica capillary at -20 kV using a mixture of 15 mmol.L(-1) NaH2PO4 and 0.5 mmol.L(-1) cetyltrimethylammonium bromide (pH 7.5) as electrolyte buffer. Se(VI) was on-line reduced to Se(IV) by mixing the CE effluent with concentrated HCl. The precision (relative standard deviation, RSD, n=7) ranged from 0.7 to 1.3% for migration time, 6.4 to 3.7% for peak height response, and 5.9 to 6.1% for peak area for the two selenium species at the 500 microg.L(-1) (as Se) level. The detection limits were 33 and 25 microg.L(-1) (as Se) for Se(VI) and Se(IV), respectively. The recoveries of the two selenium species in five locally collected water samples ranged from 88 to 114%. The developed method was applied to speciation analysis of inorganic selenium species in spiked natural water samples.     

Analytical Strategy for the Determination of Selenium in Biological Materials by Inductively Coupled Plasma-Mass Spectrometry with a Dynamic Reaction Cell

Selenium was determined in serum, hair, and tobacco by inductively coupled plasma-mass spectrometry using ⁷⁷Se, ⁷⁸Se, and ⁸²Se. The set of standards method (SSM) and the standard addition method (SAM) were applied to calibration with and without the use of internal standards (⁷²Ge and ¹⁰³Rh). In addition, the usefulness of the dynamic reaction cell (DRC) with methane as the reaction gas was characterized. The results obtained in different conditions were evaluated in terms of precision and accuracy. It was demonstrated theoretically and experimentally that an internal standard is a potential source of systematic errors as it can be influenced multiplicatively and additively by its own interferents (independently of selenium). Furthermore, it was shown that—against common opinion—an internal standard can fail in elimination of chemical interference effects influencing selenium and in increasing of precision of selenium determinations. The DRC was shown to be effective in the elimination of additive effects, although the results obtained by both SSM and SAM with DRC were systematically positively erroneous. Finally, selenium was determined accurately in each examined sample when SAM was applied to calibration, and signals were measured either for ⁸²Se without the use of the DRC, or for ⁷⁷Se or ⁷⁸Se with the use of the DRC. In addition, it has also been shown that samples should be diluted prior to analysis to the greatest acceptable extent.     

Trocknung und Veraschung biologischer Proben bei der neutronenaktivierungsanalytischen Spurenelementbestimmung

The changes in the element content of blood serum samples due to drying and ashing procedures were investigated for those trace elements which could be determined by instrumental neutron activation analysis after long-time irradiation (Ag, Co, Cr, Cs, Fe, Rb, Sb, Sc, Se, and Zn). For some elements lower concentrations were found after oven-drying at 90° C and after low temperature ashing than after freeze-drying. In the case of selenium, which is known to form several volatile compounds, no differences could be detected between the freeze-dried, oven-dried and ashed samples.     

Determination of selenium in bread-wheat samples grown under a Se-supplementation regime in actual field conditions

Selenium is an essential micronutrient for humans and animals, yet it is deficient in at least one billion people worldwide. Plants and plant-derived products transfer the soil-uptaken selenium to humans; therefore, the cultivation of plants enriched in selenium can be an effective way to improve the selenium status on humankind. This paper focuses on determining the ability of bread wheat to accumulate selenium after supplementation. One of the methods for supplementing this element in plants is foliar application with selenium solutions. These supplemented crop of wheat samples—bread wheat; Triticum aestivum L.—were used to determine if there is an increase of selenium content in cereal grains by comparing them with cereals cultivated in 2009 and harvested in 2010 with no supplementation. The experiments were done using sodium selenate and sodium selenite at three different selenium concentrations: 4, 20 and 100 g per hectare. Total Se is assessed by cyclic neutron activation analysis (CNAA), through short irradiations on the fast pneumatic system (SIPRA) of the Portuguese Research Reactor (RPI-ITN). The short-lived nuclide ^77mSe, that features a half-lifetime of 17.5 s, was used to determine the Se content in SIPRA. The experiment was successful, since the selenium concentration increased in the cropped grains and reached values up to 35 times the non-supplemented ones.     

Selenium Determination in Biological Matrices

In this study, we discuss some relevant aspects concerning the determination of selenium in biological materials with special reference to fluorometry and hydride generation atomic absorption spectroscopy (HG-AAS) techniques. The two methods may be applied without modifications to the analysis of Se in a wide spectrum of specimen types, and we describe their reliability in serum and hair analyses. Thirty-six independent control serum samples, the concentrations of which were unknown to the analyst, were analyzed in duplicate using both techniques in the Italian External Quality Assessment Scheme (EQAS). Accuracy was assessed by comparing Se values with those previously assigned by the organizers of the scheme using graphite furnace atomic absorption spectrometry (GF-AAS), which is the most frequently used technique for selenium determination in serum among the participants in the Italian EQAS. The results confirmed that fluorometry has a higher degree of accuracy than HG-AAS: the mean differences between observed and expected values were 1.5 μg/liter (95% confidence interval, −1.06 to 3.97) for fluorometry and −1.1 μg/liter (95% confidence interval, −5.05 to 2.76) for Hg-AAS. We also report some results obtained for the determination of Se in hair. Since a critical step in hair preparation is the pretreatment for removal of external contamination, we compared six different washing procedures. In general, Se is poorly leached from hair, but the efficiency of removal differed with the substance used, ranging from 0 to 13% of the original content. A nonionic detergent like Triton X-100 offers the advantage of safe working conditions and a substantial reduction in costs compared with organic solvents. Lastly, in a consistent group (n= 131) of women, Se in hair was found to be strongly reduced by the use of dye (389.9 ng/g vs 498.7 ng/g,P< 0.001). We recommend recording information on cosmetic treatments when hair is collected to evaluate Se reference values in epidemiological studies.     

Determination of different oxidation states of arsenic and selenium by inductively coupled plasma-atomic emission spectrometry with ion chromatography

Recent regulation in Japan requires more sensitive trace analysis methods for the determination of arsenic and selenium and their oxidation states As(III) and (V), Se(IV) and (VI). The hydride generation (HG) technique is usually used in combination with AAS and ICP-AES to increase sensitivity. However, hydrochloric acid is mostly used to acidify the sample solution in HG. Isobaric interferences due to chlorine-related species cause mass spectral problems when the same solution is used for the determination of these elements by ICP-MS. In this study, different oxidation states of As and Se were determined by coupling ion chromatography (IC) to an ICP-AES instrument. An HG technique was used to introduce test samples into the ICP. Nitric acid was employed to acidify the samples for HG. The concentrations of acid and base were kept as low as possible to reduce contamination. The formation of As and Se hydrides could be achieved without HCl, if the concentrations of acid and alkaline solutions were optimized. However, HCl was necessary for additional reduction of Se(VI) to Se(IV).     

How critical is the use of commercially available enzymes for selenium speciation?

The aim of this work was to check whether commercially available enzymes are pure enough to be used for selenium speciation analysis and the contribution that impurities could make to Se determination in real samples. For this purpose, twelve commercially available enzymes with different origins and classifications (protease, amylase, cellulase, lipase) were analysed. After the dissolution of the enzyme in water, the Se species were separated by ion exchange chromatography, with inductively coupled plasma mass spectrometry used as the detection system. The results showed that the Se content was significant in several cases. The highest value was obtained for β-amylase from barley, 3100 ng Se per g of enzyme. Speciation analysis showed that Se-methionine, selenite, selenate and some unknown compounds were present in several enzymes. In general, the Se species identified represented a small fraction of the total Se. For instance, only 17% of the total Se was determined for β-amylase from barley. On the other hand, about 100% of the total Se was identified in protease from Streptomyces griseus. Upon comparing the results from different lots of the same enzyme, not all of them were found to be comparable. Thus, the presence of selenium species in commercially available enzymes could be due to the preparation procedure used for the enzyme; they could be present as degradation products. Therefore, when determining selenium species in samples with low Se contents, attention should be paid to enzyme purity in relation to selenium compounds when an enzyme is used for hydrolysis.