Effect of administration route and dose on metabolism of nine bioselenocompounds.

The nutritional availability of selenium (Se) is highly dependent on its chemical form because chemical form affects absorption, distribution, metabolism, and excretion. We evaluated the effects of administration route and dose on the bioavailability of nine Se compounds found in biota, the so-called bioselenocompounds, such as selenite, selenate, selenocyanate (SeCN), Se-methylselenocysteine (MeSeCys), selenomethionine (SeMet), selenohomolanthionine (SeHLan), selenocystine (SeCys2), 1ß-methylseleno-N-acetyl-d-galactosamine (SeSug1), and trimethylselenonium ion (TMSe). We determined the bioavailability of bioselenocompounds recovered as urinary selenometabolites and serum selenoproteins from urine and serum of Se-deficient rats after the administration of bioselenocompounds by speciation analysis. Urinary Se was more easily recovered than serum selenoproteins, suggesting that the speciation of urinary Se is a better tool to indicate Se status in the body. The intravenous administration of bioselenocompounds showed different Se bioavailability from the oral administration. Intestinal microflora might be involved in the bioavailability of some bioselenocompounds, such as SeCN, MeSeCys, and SeSug1.

Detection of rare species of volatile organic selenium metabolites in male golden hamster urine.

Selenium has been considered as an essential trace element in mammals and its intake comes mainly from food. Mammals can metabolize both inorganic and organic species, and urinary excretion is the primary elimination route of selenium. Selenosugars and trimethylselenonium ion have been identified as major urinary metabolites. Other metabolites have been reported, but they were detected in some studies and not in others. Still, a large portion of the ingested selenium eliminated from the body is unknown. Volatile selenium species may account for a certain portion of the unknown species since they can easily be lost during sample analyses. While we analyzed male golden hamster urine in search of potential volatile pheromone(s), four volatile selenium compounds were detected. They were dimethyl selenenylsulfide, dimethyl diselenide, dimethyl bis(thio)selenide, and dimethyl selenodisulfide. When the urine samples were aged and dried for 48 h, dimethyl selenodisulfide tended to increase, while others decreased. The increase might be due to the formation of dimethyl selenodisulfide via reaction of dimethyl diselenide and dimethyl trisulfide whose concentration increased as urine aged. To our knowledge, dimethyl bis(thio)selenide and dimethyl selenodisulfide have never been demonstrated in urine. It remains to be determined whether these species are common metabolites in other animals or hamster-specific.

Human excretory products of selenium are natural constituents of marine fish muscle.

A selenosugar (selenosugar 1, methyl-2-acetamido-2-deoxy-1-seleno-ß-D-galactopyranoside) was identified in aqueous extracts of muscle tissue of three marine fish species, mackerel (Scomber scombrus), sardine (Sardina pilchardus), and tuna (Thunnus albacares), by high-performance liquid chromatography coupled to elemental and high-resolution molecular mass spectrometry. Selenoneine (2-selenyl-Nα, Nα, Nα-trimethyl-L-histidine), a known selenium compound in fish, was the major form of selenium in the aqueous extracts, and the methylated derivative of selenoneine, namely Se-methylselenoneine, was also identified as a minor natural constituent in the fish. Selenosugar 1, a major urinary excretion product of selenium often found in organs and body fluids related to selenium excretion, has so far not been reported in muscle tissue. Se-methylselenoneine has been proposed as the main urinary metabolite from selenoneine. This first report of selenosugar 1 and Se-methylselenoneine as natural constituents of fish muscle tissue opens up a new perspective on the role of these compounds in selenium metabolism and is relevant to selenium supplementation studies.

Determination of selenium urinary metabolites by high temperature liquid chromatography-inductively coupled plasma mass spectrometry.

The coupling of high temperature liquid chromatography (HTLC) and inductively coupled plasma mass spectrometry (ICPMS) for the determination of selenium metabolites in urine samples is reported for the first time. In order to achieve "ICPMS-friendly" chromatographic conditions, the retention on a graphite stationary phase of the major selenium urinary metabolites using only plain water with 2% methanol as the mobile phase was investigated. Under the optimal conditions (T=80°C, Ql=1.2 mL min(-1)), methyl 2-acetamido-2-deoxy-1-seleno-ß-d-galactopyranoside (selenosugar 1), methyl 2-acetamido-2-deoxy-1-seleno-ß-d-glucosopyranoside (selenosugar 2) and trimethylselenonium ion were efficiently separated in less than 7 min, without any interferences due to other common selenium species (selenite, selenate, selenocystine and selenomethionine) or detectable effect of the urine matrix. The limits of detection were 0.3-0.5 ng Se mL(-1), and the precision of the analytical procedure was better than 3% (RSD%, n=5). The HTLC-ICPMS method was applied to the analysis of urine samples from two volunteers before and after ingestion of Brazil nuts or selenium supplements. The developed procedure proved to be adequate for the analytical task, providing results consistent with previous studies.

Efficient interface for online coupling of capillary electrophoresis with inductively coupled plasma-mass spectrometry and its application in simultaneous speciation analysis of arsenic and selenium.

A simple and highly efficient online system coupling of capillary electrophoresis to inductively coupled plasma-mass spectrometry (CE-ICP-MS) for simultaneous separation and determination of arsenic and selenium compounds was developed. CE was coupled to an ICP-MS system by a sprayer with a novel direct-injection high-efficiency nebulizer (DIHEN) chamber as the interface. By using this interface, six arsenic species, including arsenite (As(III), arsenate (As(V)), monomethylarsonic acid (MMA), dimethylarsinic acid (DMA), arsenobetaine (AsB), and arsenocholine (AsC) and five selenium species (such as sodium selenite (Se(IV)), sodium selenate (Se(VI)), selenocysteine (SeCys), selenomethionine (SeMet), and Se-methylselenocysteine (MeSeCys)) were baseline-separated and determined in a single run within 9 min under the optimized conditions. Minimum dead volume, low and steady sheath flow liquid, high nebulization efficiency, and high sample transport efficiency were obtained by using this interface. Detection limits were in the range of 0.11-0.37 µg L(-1) for the six arsenic compounds (determined as (75)As at m/z 75) and 1.33-2.31 µg L(-1) for the five selenium species (determined as (82)Se at m/z 82). Repeatability expressed as the relative standard deviations (RSD, n = 6) of both migration time and peak area were better than 2.68% for arsenic compounds and 3.28% for selenium compounds, respectively. The proposed method had been successfully applied for the determination of arsenic and selenium species in the certified reference materials DORM-3, water, urine, and fish samples.

Metabolism of selenite to selenosugar and trimethylselenonium in vivo: tissue dependency and requirement for S-adenosylmethionine-dependent methylation.

Impaired S-adenosylmethionine (SAM)-dependent transmethylation and methylation capacity feature in diseases related to obesity or aging, and selenium (Se) metabolism is altered in these states. We tested the hypothesis that SAM metabolism is required for methylation and excretion of Se in a rat model. Four hours after selenite and periodate-oxidized adenosine (POA; an inhibitor of SAM metabolism) were administered, circulating markers of single-carbon status were unchanged, except for decreased circulating phosphatidylcholine (P<.05). In contrast, liver and kidney SAM and S-adenosylhomocysteine were elevated (P<.05 for all). Concentrations of total Se were significantly elevated in both liver (P<.001) and kidney (P<.01), however the degree of accumulation in liver was significantly greater than that of kidney (P<.05). Red blood cell Se levels were decreased (P=.01). Trimethylselenonium levels were decreased in liver and kidney (P=.001 for both tissues) and Se-methyl-N-acetylselenohexosamine selenosugar was decreased in liver (P=.001). Urinary output of both trimethylselenonium (P=.001) and selenosugar (P=.01) was decreased as well. Trimethylselenonium production is more inhibited by POA than is selenosugar production (P<.05). This work indicates that low molecular weight Se metabolism requires SAM-dependent methylation, and disrupting the conversion of SAM to S-adenosylhomocysteine prevents conversion of selenite and intermediate metabolites to final excretory forms, suggesting implications for selenium supplementation under conditions where transmethylation is suboptimal, such as in the case of obese or aging individuals.

Head space solid phase microextraction based on nano-structured lead dioxide: application to the speciation of volatile organoselenium in environmental and biological samples.

A novel and efficient speciation method based on the nano-structured lead dioxide as stationary phase of head space solid phase microextraction combined with gas chromatography mass spectrometry (GC-MS) was developed for the determination of volatile organoselenium compounds (dimethylselenide (DMSe) and dimethyldiselenide (DMDSe)) in different biological and environmental samples. PbO(2) particles with a diameter in the range of 50-70 nm have been grown on platinum wire via elechtrochemical deposition. The effect of different variables on the extraction efficiency was studied simultaneously using an experimental design. The variables of interest in the HS-SPME were condition of coating preparation, desorption time, stirring rate, desorption temperature, ionic strength, time and temperature of extraction. A Plackett-Burman design was performed for screening in order to determine the significant variables affecting the extraction efficiency. Then, the significant factors were optimized by a Box-Behnken design (BBD) and the response surface equations were derived. The detection limit and relative standard deviation (RSD) (n=5, c=50 µgL(-1)) for DMSe were 16 ngL(-1) and 4.3%, respectively. They were also obtained for DMDSe as 11ngL(-1) and 4.6%, respectively. The developed technique was found to be applicable to spiked environmental and biological samples.

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.

Headspace hollow fiber protected liquid-phase microextraction combined with gas chromatography-mass spectroscopy for speciation and determination of volatile organic compounds of selenium in environmental and biological samples.

A simple and novel speciation method for the determination of volatile organic compounds of selenium (dimethylselenide (DMSe) and dimethyldiselenide (DMDSe) has been developed using a headspace hollow fiber protected liquid-phase microextraction (HS-HF-LPME) combined with capillary gas chromatography-mass spectrometry (GC-MS). The organic solvent impregnated in the pores and filled inside the porous hollow fiber membrane was used as an extraction interface in the HS-HF-LPME of the compounds. The effect of different variables on the extraction efficiency was studied simultaneously using an experimental design. The variables of interest in the HS-HF-LPME were sample volume, extraction time, temperature of sample solution, ionic strength, stirring rate and dwelling time. A Plackett-Burman design was performed for screening in order to determine the significant variables affecting the extraction efficiency. Then, the significant factors were optimized by a Box-Behnken design (BBD) and the response surface equations were derived. Under optimum conditions, preconcentration factors up to 1250 and 1170 were achieved for DMSe and DMDSe respectively. The detection limit and relative standard deviation (RSD) (n=5, c=50 µg L(-1)) for DMSe were 65 ng L(-1) and 4.8%, respectively. They were also obtained for DMDSe as 57 ng L(-1) and 3.9%, respectively. The developed technique was found to be applicable to spiked environmental and biological samples.

Formation of methylselenol, dimethylselenide and dimethyldiselenide in in vitro metabolism models determined by headspace GC-MS.

The aim of this study was to identify the presence of MeSeH in metabolic reactions. An analytical method based on direct headspace GC-MS, eliminating loss of volatile species during sample pretreatment procedures, was developed for this purpose. The in vitro conversion of selenium compounds to the volatile species methylselenol, MeSeH, dimethyl selenide, DMeSe and dimethyl diselenide, DMeDSe was investigated. The analytical method was evaluated by means of standards of dimethyl diselenide, dimethyl selenide. The corresponding sulfides were found unsuitable as internal standards as they interacted with the selenides. The limit of detection was 0.25 µmol L(-1) (20 µg L(-1)) for the selenide as well as the diselenide. Formation of MeSeH was not observed in significant amount when selenomethionine was incubated with the enzyme l-methionine-γ-lyase; instead large amounts of DMeDSe were formed. In aqueous solution, methylseleninic acid, MeSeA reacted spontaneously with glutathione, GSH to form DMeDSe. In strongly reducing environments, however, MeSeH was also observed. When the formed MeSeH was trapped with iodoacetic acid, no DMeDSe was detected indicating that DMeDSe formation was due to spontaneous oxidation of MeSeH. These findings imply that DMeDSe may be a marker for the production of MeSeH in in vitro models. When MeSeA, Se-methylselenocysteine, Se-MeSeCys and SeMet were incubated with Jurkat cells, DMeDSe formation was only observed in the case of MeSeA. Trace amounts of DMeSe was observed in the vial with MeSeA as well as Se-MeSeCys. When DMeSe and DMeDSe were added to plasma, the sensitivity of only DMeDSe decreased significantly, implicating that DMeDSe underwent a reaction with plasma hindering the volatilization. This emphasizes that results from in vitro selenium metabolism studies may not be uncritically interpreted as consistent with the in vivo reality.

[LC-MSn analysis of metabolites of 1,2-[bis (1,2-benzisoselenazolone-3(2H)-ketone)]-ethane, a novel anti-cancer agent in rat].

This study is to elucidate the metabolic pathway of 1,2-[bis (1,2-benzisoselenazolone-3 (2H)-ketone)]-ethane (BBSKE) in rats. Rats were administrated with a single dose of BBSKE 200 mg x kg(-1). The metabolites in rat urine, feces, bile and plasma were identified by LC-MSn analysis. The characterization of fragment ions from LC-MSn chromatography and mass spectrometry was applied to the investigation of structures of metabolites. Three phase I metabolites were detected in rat urine and feces. Two of them were also found in plasma and one existed in bile. These products were derived from oxidized, methylated and S-methylated BBSKE, separately. One phase II glucuronide of BBSKE was also found in bile. Therefore, it is possible that BBSKE was metabolized by oxidization, methylation and glucuronidation.

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.

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.

Quantitative analysis of volatile selenium metabolites in normal urine by headspace solid phase microextraction gas chromatography-inductively coupled plasma mass spectrometry.

The combination of headspace-solid phase microextraction (HS-SPME) and gas chromatography-inductively coupled plasma mass spectrometry (GC-ICPMS) was evaluated for the determination of volatile selenium metabolites in normal urine samples, i.e. without selenium supplementation. HS-SPME operating conditions were optimised and a sampling time of 10 min was found to be suitable for simultaneous extraction of dimethylselenide (DMSe) and dimethyldiselenide (DMDSe). The amount of DMSe and DMDSe extracted onto fibre coating was calculated in clean matrix, i.e. Milli-Q water, on the basis of depletion experiments. When applied to normal urine samples, the developed method allowed the detection of four volatile selenium containing species, among which DMSe and DMDSe could be quantified by standard additions.

Consequences of vapor enhancement on selenium speciation analysis by HPLC/ICPMS.

Recent work has shown the presence of volatile selenium metabolites in human urine and suggested that these compounds could compromise quantitative selenium analyses by ICPMS. We show that with a commonly used sample introduction system (pneumatic nebulizer and spray chamber), two volatile selenium species recently identified in urine, namely, dimethyl selenide and dimethyl diselenide, gave greatly increased ICPMS responses (up to 58-fold) relative to selenite, an effect related to their volatilization in the spray chamber resulting in enhanced transport to the plasma. The quantitative consequences of this effect were demonstrated by measurement of total selenium and selenium species in certified reference material, NIES CRM 18 human urine. Direct flow injection analysis of the urine gave a total selenium concentration more than 2-fold higher than the certified value. These data suggested that NIES CRM 18 may contain part of its selenium as volatile species, and subsequent reversed-phase HPLC/ICPMS showed the presence of dimethyl selenide in addition to selenosugars and trimethylselenonium ion. Although the practice of quantifying unidentified chromatographic peaks against those of known compounds is common in speciation analysis, this approach when applied to NIES CRM 18 gave a value for the sum of selenium species which was twice the certified total selenium concentration. This work shows that the presence of volatile selenium species in urine precludes the use of flow injection analysis for total selenium measurements and imposes severe restrictions on the quantification of urinary selenium metabolites. In addition, it raises broader issues of the validity of the "dilute and shoot" approach to the determination of metals in clinical analysis of biological fluids.

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.

Susceptibility of Escherichia coli to L-selenaproline and other L-proline analogues in laboratory culture media and normal human urine.

AIMS: The aims of this study were to identify analogues of L-proline which inhibit the growth of Escherichia coli in both laboratory culture media and normal human urine and to study their mechanisms of uptake. METHODS AND RESULTS: The susceptibility of E. coli to L-proline analogues was studied by radial streak assays on agar plates and by minimal inhibitory concentration determinations in liquid media. Only L-selenaproline (SCA) inhibited growth in Mueller-Hinton medium and human urine as well as in glucose minimal medium. L-Proline did not prevent the inhibition of growth by SCA and strains defective in L-proline transport were as susceptible to SCA as wild-type strains. However, E. coli was resistant to SCA in the presence of L-cysteine and L-cystine. Spontaneous mutants selected for resistance to SCA or L-selenocystine were resistant to the other compound and had reduced growth in minimal medium containing L-cysteine or L-cystine as the sole sulfur source. CONCLUSIONS: L-selenaproline inhibited the growth of E. coli under conditions that may occur in the urinary tract and appeared to be taken up by the L-cystine transport system. SIGNIFICANCE AND IMPACT OF THE STUDY: Although urinary tract infections caused by E. coli can be treated with sulfamethoxazole/trimethoprim and quinolones, resistance to these antibiotics has been increasing. These results suggest that L-selenaproline may represent a new class of compounds that could be used to treat these infections.

Selenium speciation by high-performance anion-exchange chromatography-post-column UV irradiation coupled with atomic fluorescence spectrometry.

A technique for the speciation of selenomethylcysteine (SeMeCys), selenocystine (SeCys), selenite [Se(IV)] and selenomethionine (SeMet) was established in this paper using high-performance anion-exchange chromatography coupled with atomic fluorescence spectrometry (HPAEC-AFS). Analytes were separated on an AminoPac PA10 column and then digested by on-line ultraviolet (UV) irradiation, which destroyed organic compound structure. Hydride generation was used as an available sample introduction technique for atomic fluorescence detection. The detection limits of four compounds were 1-5 microg/L (250 microL injection, 10 times of the baseline noise). The relative standard deviations (RSDs), calculated from seven consecutive injections of 100 microg/L standard mixtures, were from 2 to 4%. Selenious yeast tablet, which had been proposed as selenium supplement, and human urine collected from a volunteer were analyzed. Good spiked recoveries from 86 to 103% were obtained.