Glycosyldiselenides as lectin ligands detectable by NMR in biofluids.

The ability of glycosyldiselenides to act as lectin ligands and their selective detection in plasma by (77)Se NMR is reported.

Quantitative determination of small selenium species in human serum by HPLC/ICPMS following a protein-removal, pre-concentration procedure.

Protein precipitation was incorporated into a sample preparation method for the quantitative determination of small "non-protein" selenium species in human serum by high-performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC/ICPMS). The advantages of cleaner matrix and concomitant concentration of the small compounds result in quantification limits in the native serum at the sub-micrograms Se per litre level. Spiking experiments with methyl 2-acetamido-2-deoxy-1-seleno-ß-D-galactopyranoside (selenosugar 1), trimethylselenonium ion, selenomethionine, methylselenocysteine (MeSeCys) and selenate yielded recoveries from 73% to 103%. Selenite had a low recovery (44%), possibly owing to protein binding. The validated method was applied to serum samples from two volunteers before and after ingestion of a selenium food supplement. HPLC/ICPMS analysis showed, besides ingested selenate, the presence of selenosugar 1 and trace amounts of MeSeCys and methyl 2-amino-2-deoxy-1-seleno-ß-D-galactopyranoside, which have not been reported in human serum before.

Identification in human urine and blood of a novel selenium metabolite, Se-methylselenoneine, a potential biomarker of metabolization in mammals of the naturally occurring selenoneine, by HPLC coupled to electrospray hybrid linear ion trap-orbital ion trap MS.

Speciation analysis of selenium in human urine allowed for the first time the identification of a novel selenium metabolite, Se-methylselenoneine. Despite a concentration at low ppb level, its characterization was achieved after sample purification by solid phase extraction (SPE) followed by the parallel coupling of the bidimensional RP/HILIC chromatography with ICP-MS and ESI-LTQ Orbitrap MS detection. To confirm its biological significance with regards to selenoneine, the recently discovered analog of ergothioneine, and to discard the possibility of sample preparation artifacts, a new method was developed to monitor its actual presence, as well as the occurrence of its sulfur and/or non-methylated analogs, in non-preconcentrated urine and blood samples of non-supplemented humans. It consisted in a HILIC ESI-MS(3) method in high resolution mode (resolution 30 000 at m/z 400) with large isolation width windows for precursor ions. These two particular settings allowed respectively to keep observing the specific mass defect of selenium- and sulfur-containing molecules and to maintain the characteristic selenium pattern in product ions created through MS(n) fragmentations. As a result, all four metabolites were detected in blood and three of them in urine. Moreover, different ratios "methylated/non-methylated" were observed between urine and blood samples, which seemed to indicate their active metabolization. The analytical tool developed here will be of a great importance to further study the occurrence and the potential metabolic role in mammalian organelles, cells and fluids of these very particular and promising redox metabolites.

Simultaneous analysis of mercury and selenium species including chiral forms of selenomethionine in human urine and serum by HPLC column-switching coupled to ICP-MS.

The simultaneous speciation of elements is of great concern, especially in the study of the interactions of species in living organisms. Here we report a method based on the coupling of HPLC-ICP-MS that is capable of separating and analyzing different selenium and mercury species (Se-methylselenocysteine, selenite, selenate, L-selenomethionine, D-selenomethionine, methylmercury and inorganic mercury). The proposed method uses two different mobile phases that are suitable for selenium and mercury speciation and leads to a successful determination of all the species in less than 27 min with good efficiency and resolution. The method was efficiently applied for simultaneous speciation of mercury and selenium in urine and in serum, the latter from umbilical cord samples. Selenocystine has been successfully identified in the former sample. Detection limits obtained were between 0.30 and 2.46 ng. Recovery studies of samples spiked with all species were performed to check the reliability of the method, and satisfactory recoveries (93-110%) were obtained in all cases. The relative standard deviations (RSDs) for species with ten replicate determinations of 80 µg L(-1) were between 4.5 and 9.2%. The proposed method offers a deeper insight into selenium and mercury interactions in the human body.

Identification of a novel selenium-containing compound, selenoneine, as the predominant chemical form of organic selenium in the blood of bluefin tuna.

A novel selenium-containing compound having a selenium atom in the imidazole ring, 2-selenyl-N(alpha),N(alpha),N(alpha)-trimethyl-L-histidine, 3-(2-hydroseleno-1H-imidazol-5-yl)-2-(trimethylammonio)propanoate, was identified from the blood and other tissues of the bluefin tuna, Thunnus orientalis. The selenium-containing compound was purified from the tuna blood in several chromatographic steps. High resolution mass spectrometry and nuclear magnetic resonance spectroscopy showed that the exact mass of the [M+H](+) ion of the compound was 533.0562 and the molecular formula was C(18)H(29)N(6)O(4)Se(2). Its gross structure was assigned as the oxidized dimeric form of an ergothioneine selenium analog in which the sulfur of ergothioneine is replaced by selenium. Therefore, we named this novel selenium-containing compound "selenoneine." By speciation analysis of organic selenium compounds using liquid chromatography inductively coupled plasma mass spectrometry, selenoneine was found widely distributed in various tissues of the tuna, with the highest concentration in blood; mackerel blood contained similar levels. Selenoneine was measurable at 2-4 orders of magnitude lower concentration in a limited set of tissues from squid, tilapia, pig, and chicken. Quantitatively, selenoneine is the predominant form of organic selenium in tuna tissues.

Convulsant effect of diphenyl diselenide in rats and mice and its relationship to plasma levels.

Diphenyl diselenide [(PhSe)2], an organoselenium compound, presents pharmacological and toxicological properties in rodents. The aim of this study was to carry out the determination and quantification of (PhSe)2 in plasma after oral administration (p.o.) of this compound (500 mg/kg), dissolved in canola oil, in rats and mice. The second objective was to verify the involvement of different routes of administration ((p.o.), intraperitoneal (i.p.) and subcutaneous (s.c.)) and vehicle solutions (canola oil and dimethyl sulfoxide (DMSO)) in the appearance of seizure episodes and in the plasmatic levels of (PhSe)2 in rats and mice. Analysis of (PhSe)2 in blood samples was performed by gas chromatography/flame ionized detector system (GC/FID). Rat and mouse peak plasma (PhSe)2 levels were 13.13 and 10.11 microg/ml (C(max)), respectively, and occurred at 0.5h (T(max)) post-dosing. The use of different administration routes (p.o., i.p. and s.c.) and vehicle solutions (canola oil or DMSO) in rats and mice indicated that the appearance of seizures and (PhSe)2 plasmatic levels are dependent of administration routes (i.p.>p.o.>s.c.), vehicle solutions (DMSO>canola oil) and animal species (mice>rat).

Selenium metabolism in rats with long-term ingestion of Se-methylselenocysteine using enriched stable isotopes.

Se-methylselenocysteine (MeSeCys) is not only a selenium (Se) supplement but also a more promising precursor of an anti-tumor drug containing Se than selenomethionine, which is currently used as Se supplement. In this study, the metabolism of MeSeCys labeled with an Se isotope, 82Se, in rats depleted of endogenous natural abundance isotopes with another Se isotope, 78Se, was traced for 21 days when MeSeCys was continuously and perorally ingested at a supplemental dose. The tracer experiment was performed with our improved method that utilized an inductively coupled plasma-deuterium reaction-mass spectrometer. The substitution of endogenous Se with a single isotope, 78Se, facilitated the detection of exogenous labeled Se. Exogenous Se in the form of MeSeCys preferably accumulated and/or assimilated in the liver, kidneys and testes with long-term ingestion of MeSeCys and was utilized for the synthesis of selenoproteins, i.e., extracellular and cellular glutathione peroxidases and selenoprotein P. Meanwhile, intact MeSeCys was not excreted into urine although trimethylselenonium was detected in addition to selenosugar. The results suggest that MeSeCys was transformed into selenide via methylselenol by beta-lyase. Consequently, it is surmised that MeSeCys is a precursor of methylselenol under long-term ingestion.

A novel HPLC-UV/nano-TiO2-chemiluminescence system for the determination of selenocystine and selenomethionine.

Active oxygen species from the photocatalytic reaction in aqueous solution react with luminol to emit strong chemiluminescence (CL), and this can be inhibited by the UV decomposed-products of selenocystine (SeCys) or selenomethionine (SeMet). Based on this phenomenon, a novel hyphenated technique, HPLC-UV/nano-TiO(2)-CL, was established for the determination of SeCys and SeMet. The effects of pH, the UV irradiation time, the TiO(2) coated on the inner surface of the reaction tubing, and the Co(2+) catalyst concentration on the CL intensity and/or chromatographic resolution were systematically investigated. Under these optimized conditions, the inhibited CL intensity has a good linear relationship with the concentration of SeCys in the range of 0.04-10.6 microg mL(-1) or SeMet in the range of 0.05-12.4 microg mL(-1), with a limit of detection (S/N=3) of 6.4 microg L(-1) for SeCys or 12 microg L(-1) for SeMet. As an example, the method was preliminarily applied to the determination of the selenoamino acids in garlic and rabbit serum, with a recovery of 88-104%.

Metabolism of 76Se-methylselenocysteine compared with that of 77Se-selenomethionine and 82Se-selenite.

Se-Methylated selenoamino acids, Se-methylselenocysteine (MeSeCys) and selenomethionine (SeMet), are chemically inert storage forms of selenium in selenium-accumulators, and a nutritional and supplemental source. The metabolic pathway for MeSeCys was precisely traced by referring to those for SeMet and selenite by applying a new tracer method involving multiple homo-elemental stable isotopes. Male Wistar rats were depleted of endogenous natural abundance selenium with a single (80)Se-enriched isotope, and then (76)Se-MeSeCys, (77)Se-SeMet and (82)Se-selenite were orally administered simultaneously at 25 microg Se/kg body weight each. Organs and body fluids were obtained at 3, 6, 9 and 12 h, and 1 and 2 days later, and subjected to speciation analysis. The main characteristics of the metabolism were as follows; MeSeCys was incorporated into selenoprotein P slightly more than or at a comparable level to that of SeMet but less than that of selenite. MeSeCys and SeMet but not selenite was taken up by organs in their intact forms. MeSeCys and SeMet were delivered specifically to the pancreas and present in a form bound to an identical or similar protein. Trimethylselenonium (TMSe) was only produced from MeSeCys, i.e., not from SeMet or selenite, in the kidneys. Both selenosugars A and B of MeSeCys, SeMet and selenite origin were detected in the liver but only selenosugar B in the kidneys. These results suggest that MeSeCys can be a similar or better selenium source than SeMet, and supplies methylselenol much more efficiently in organs than SeMet and selenite. TMSe was produced much efficiently from MeSeCys than from SeMet and selenite, suggesting a role of methylselenol through the beta-lyase reaction in the metabolism of Se-methylated selenoamino acids.

Availability and metabolism of 77Se-methylseleninic acid compared simultaneously with those of three related selenocompounds.

Nutritional selenocompounds are considered to be transformed into the common intermediate selenide for utilization as selenoenzymes and/or for excretion as selenosugar and trimethylselenonium (TMSe). Therefore, selenocompounds can only be traced with a labeled selenium atom. Methylseleninic (MSA(IV)) has been proposed to be a third nutritional selenium source, the other two being inorganic selenocompounds and organic selenoamino acids, and to be a proximate selenochemical for producing the assumed biologically active form methylselenol. Here we applied a new tracer method to compare the availability and metabolism of MSA(IV) with those of three related selenocompounds under exactly identical host and tracing conditions. (82)Se-Selenite, (78)Se-selenate, (77)Se-MSA(IV) and (76)Se-methylselenonic acids (MSA(VI)) were simultaneously administered orally, each at the dose of 25 microg Se/kg body weight, to rats that had been depleted of endogenous natural abundance selenium with a single stable isotope ((80)Se). Time-related changes in the concentrations and/or distributions of the four labeled isotopes in the serum, liver, kidney, pancreas, lung and urine were determined simultaneously by inductively coupled argon plasma mass spectrometry (ICP MS) and/or HPLC-ICP MS. The availability with different isotope ratios was in the decreasing order of selenate>selenite=MSA(IV)>MSA(VI). Although selenate and MSA(VI) were distributed in organs and urine partly in their intact forms, MSA(IV) and selenite were not detected in the intact forms at all. MSA(IV) and MSA(VI) but not selenite or selenate produced TMSe in organs other than the liver, suggesting the transformation of MSA(IV) into methylselenol, and then either into selenide for the synthesis of selenoproteins and selenosugar or directly into TMSe. Thus, selenosugar and TMSe were produced widely in the organs. However, TMSe was not detected in the liver. The organ- and selenium source-specific production of TMSe was discussed as to the differences in selenium sources, and demethylation and methylation activity.

Analysis of the selenium species distribution in cow blood by size exclusion liquid chromatography-inductively coupled plasma collision cell mass spectrometry (SEC-ICPccMS).

A method for performing rapid semiquantitative screening of the distribution of Se species in the blood of cows fed with a diet enriched in selenized yeast was optimized. The method was based on direct injection of a blood sample onto a high resolution size exclusion chromatographic column and fractionation of the selenium species. Selenium was detected on-line by ICP-MS with a collision cell. The concentrations of selenized haemoglobin and free selenomethionine were estimated using the chromatogram. The method was applied to a study involving 15 control and 15 treated dairy cows at four different supplementation time points. The increase in the selenomethionine and selenized haemoglobin was a linear function of the total selenium concentration. A threshold value of 600 ng ml(-1) of total Se was established beyond which selenomethionine could not be incorporated into the protein. No inorganic selenium was found to be present. The total selenium in cow blood correlated well with that in milk. The selenium supplementation did not change the protein distribution profiles for other essential elements (Cu, Fe, Mn, Zn).

Organochalcogens effects on delta-aminolevulinate dehydratase activity from human erythrocytic cells in vitro.

Organochalcogens are important intermediates and useful reagents in organic synthesis, which can increase human exposure risk to these chemicals in the workplace. As well, there are a number of reported cases of acute toxicity following organochalcogen ingestion of vitamins and dietary supplements. Since, the erythrocytic delta-ALA-D activity could be an important indicator of toxicity this report investigated the organochalcogens effects on blood delta-ALA-D in vitro. To investigate a possible involvement of cysteinyl groups in the inhibitory actions of diphenyl diselenide, diphenyl ditelluride and Ebselen (4-100 micro M), the effects of thiol reducing agents (0-3 mM) or zinc chloride (0-2 mM) were examined. Diphenyl ditelluride, diphenyl diselenide and Ebselen inhibited in a concentration-dependent manner delta-ALA-D activity from human erythrocytes. Ebselen was lesser delta-ALA-D inhibitor than (PhSe)(2) and (PhTe)(2), whereas the diorganoyldichalcogenides displayed similar inhibitory potency towards delta-ALA-D. Dithiothreitol, a hydrophobic SH-reducing agent, was able to reactivate and to protect inhibited delta-ALA-D. The pre-incubation of blood with the inhibitors changed considerably the reversing potency of thiols. From these findings we suggest that organochalcogens inactivate in vitro human erythrocyte delta-ALA-D by an interaction with the sulfhydryl group essential of the enzyme activity.

Organic and inorganic selenium supplementation to lactating mothers increase the blood and milk Se concentrations and Se intake by breast-fed infants.

The aim of this study was to determine the effect of selenium (Se) supplementation to lactating women on Se concentrations and glutathione peroxidase (GSH-Px) activities in blood components of mothers and breast-fed infants and on milk Se levels and Se intake by breast-fed infants. Lactating mothers were supplied for 3 months with 200 micrograms Se/day in the form of yeast-Se (Y-Se) and sodium selenite. Initial blood and plasma Se levels of all women (n = 67) were 76.6 and 53.2 micrograms/L, respectively. After 3 months Se concentrations both in whole blood and in plasma from mothers and infants were significantly higher than the initial values. Y-Se exerts a stronger effects than selenite on blood and plasma Se levels. Initial milk Se concentration was 8.9 micrograms/L and after 1 month in both groups in reached a plateau at 14-16 micrograms/L. This resulted in an increase of Se intake in breast-fed infants from 6.1 to a plateau of 11-13 micrograms Se/day. GSH-Px activities in plasma and red cells of Y-Se group increased significantly and reached a plateau after 1 and 2 months, respectively, while in the selenite group the enzyme activities increased steadily throughout the entire period of the study. Selenite exerts a stronger effect on GSH-Px both in maternal and in infant blood components as compared with Y-Se. In milk the GSH-Px activity in the Y-Se group did not change during the study, while in the selenite group after 3 months it increased almost 2-fold compared to the initial value. In conclusion, this study shows that organic Se causes higher Se deposition than did the inorganic form.