Intravenous Infusion of High Dose Selenite in End-Stage Cancer Patients: Analysis of Systemic Exposure to Selenite and Seleno-Metabolites.

Cancer is one of the main causes of human death globally and novel chemotherapeutics are desperately required. As a simple selenium oxide, selenite is a very promising chemotherapeutic because of pronounced its dose-dependent tumor-specific cytotoxicity. We previously published a first-in-man systematic phase I clinical trial in patients with cancer (from IV to end-stage) (the SECAR trial) showing that selenite is safe and tolerable with an unexpectable high maximum tolerated dose (MTD) and short half-life. In the present study, we analyzed the selenium species in plasma samples, from the patients participating in the SECAR trial and from various time points and dose cohorts using LC-ICP-MS. In conclusion, selenite, selenosugars, and 1-2 unidentified peaks that did not correspond to any standard, herein denoted ui-selenium, were detected in the plasma. However, trimethylated selenium (trimethylselenonoium) was not detected. The unidentified ui-selenium was eluting close to the selenium-containing amino acids (selenomethionine and selenocysteine) but was not part of a protein fraction. Our data demonstrate that the major metabolite detected was selenosugar. Furthermore, the identification of selenite even long after the administration is remarkable and unexpected. The kinetic analysis did not support that dosing per the body surface area would reduce interindividual variability of the systemic exposure in terms of trough concentrations.

Comparison of external calibration and isotope dilution LC-ICP-MS/MS for quantitation of oxytocin and its selenium analogue in human plasma.

In the present study, a method for quantitation of the pharmaceutical peptide oxytocin (OT) and its diselenide-containing analogue (SeOT) in human plasma was developed using gradient elution LC-ICP-MS/MS. Plasma samples were precipitated with acetonitrile containing 1.0% TFA in a volume ratio of 1+3 (sample+precipitation agent) before analysis. Post-column isotope dilution analysis (IDA) was applied for quantitation and was compared with external calibration. Both calibration methods appeared to be fit for purpose regarding figures of merit including linearity, precision, LOD, LOQ and recovery. Analysis of OT and SeOT showed that selenium-based analysis is considerably more sensitive and selective compared to the sulfur-based analysis. Despite the relatively simpler setup of external calibration, IDA can be advantageous because it compensates for instrument drift and changes in organic solvent concentration. The method was applied for a stability study showing the degradation of OT and SeOT in plasma. The degradation of SeOT was faster than the degradation of OT in plasma. Thus, possible stability effects should be considered before replacing a disulfide bridge with a diselenide bridge or introducing a diselenide label in a potential drug.

Modulation of hypochlorous acid (HOCl) induced damage to vascular smooth muscle cells by thiocyanate and selenium analogues.

The production of hypochlorous acid (HOCl) by myeloperoxidase (MPO) plays a key role in immune defense, but also induces host tissue damage, particularly in chronic inflammatory pathologies, including atherosclerosis. This has sparked interest in the development of therapeutic approaches that decrease HOCl formation during chronic inflammation, including the use of alternative MPO substrates. Thiocyanate (SCN-) supplementation decreases HOCl production by favouring formation of hypothiocyanous acid (HOSCN), which is more selectively toxic to bacterial cells. Selenium-containing compounds are also attractive therapeutic agents as they react rapidly with HOCl and can be catalytically recycled. In this study, we examined the ability of SCN-, selenocyanate (SeCN-) and selenomethionine (SeMet) to modulate HOCl-induced damage to human coronary artery smooth muscle cells (HCASMC), which are critical to both normal vessel function and lesion formation in atherosclerosis. Addition of SCN- prevented HOCl-induced cell death, altered the pattern and extent of intracellular thiol oxidation, and decreased perturbations to calcium homeostasis and pro-inflammatory signaling. Protection was also observed with SeCN- and SeMet, though SeMet was less effective than SeCN- and SCN-. Amelioration of damage was detected with sub-stoichiometric ratios of the added compound to HOCl. The effects of SCN- are consistent with conversion of HOCl to HOSCN. Whilst SeCN- prevented HOCl-induced damage to a similar extent to SCN-, the resulting product hyposelenocyanous acid (HOSeCN), was more toxic to HCASMC than HOSCN. These results provide support for the use of SCN- and/or selenium analogues as scavengers, to decrease HOCl-induced cellular damage and HOCl production at inflammatory sites in atherosclerosis and other pathologies.

Analysis of selenium nanoparticles in human plasma by capillary electrophoresis hyphenated to inductively coupled plasma mass spectrometry.

Nanoparticles (NPs) are increasingly applied in research and development of new therapies. Characterization of NP systems most often include size, shape, size distribution, and charge but information on the chemical stability of NPs and investigation of the presence of dissolved species is most often missing in efficacy studies due to lack of appropriate methods. In this study, a method based on capillary electrophoresis coupled to inductively coupled plasma mass spectrometry (CE-ICP-MS) was established for analysis of selenium (Se) NPs and dissolved Se species in aqueous media. Peak area and migration time precisions (RSD) of 1.4-3.0% and 1.0-2.6%, respectively, were obtained. CE-ICP-MS analysis of a commercially available SeNP suspension (Q-SeNP) revealed large amounts of selenite corresponding to 32% of the total Se content in the suspension, indicating considerable NP degradation upon storage. The CE-ICP-MS method was modified using a coated fused silica capillary in order to analyze SeNPs in human plasma. Peak area and migration time precisions (RSD) in the range of 3.3-10.7% and 0.8-2.8%, respectively, were achieved. Degradation of polyvinyl alcohol (PVA)-coated SeNPs to selenite in human plasma was demonstrated using the modified method. The amounts of SeNP and selenite were estimated based on a correction factor for the ICP-MS signals of PVA-SeNP and dissolved Se. To the best of our knowledge, this is the first study of SeNPs by CE-ICP-MS and highlights the potential of CE-ICP-MS for quantitative characterization of the behavior of SeNPs in biological media.

Selenomethionine supplementation reduces lesion burden, improves vessel function and modulates the inflammatory response within the setting of atherosclerosis.

Atherosclerosis is a chronic inflammatory disease of the vasculature characterised by the infiltration of activated neutrophils and macrophages at sites of damage within the vessel wall, which contributes to lesion formation and plaque progression. Selenomethionine (SeMet) is an organic form of selenium (Se), an essential trace element that functions in the regulation of the immune response by both bolstering the endogenous thioredoxin and glutathione antioxidant defence systems and by directly scavenging damaging oxidant species. This study evaluated the effect of dietary SeMet supplementation within a high fat diet fed apolipoprotein E deficient (ApoE-/-) mouse model of atherosclerosis. Dietary supplementation with SeMet (2 mg/kg) increased the tissue concentration of Se, and the expression and activity of glutathione peroxidase, compared to non-supplemented controls. Supplementation with SeMet significantly reduced atherosclerotic plaque formation in mouse aortae, resulted in a more stable lesion phenotype and improved vessel function. Concurrent with these results, SeMet supplementation decreased lesion accumulation of M1 inflammatory type macrophages, and decreased the extent of extracellular trap release from phorbol myristate acetate (PMA)-stimulated mouse bone marrow-derived cells. Importantly, these latter results were replicated within ex-vivo experiments on cultured neutrophils isolated from acute coronary syndrome patients, indicating the ability of SeMet to alter the acute inflammatory response within a clinically-relevant setting. Together, these data highlight the potential beneficial effect of SeMet supplementation as a therapeutic strategy for atherosclerosis.

Quantification of low molecular weight selenium metabolites in human plasma after treatment with selenite in pharmacological doses by LC-ICP-MS.

The paper presents an analytical method for quantification of low molecular weight (LMW) selenium compounds in human plasma based on liquid chromatography inductively coupled plasma mass spectrometry (LC-ICP-MS) and post column isotope dilution-based quantification. Prior to analysis, samples were ultrafiltrated using a cut-off value of 3000 Da. The method was validated in aqueous solution as well as plasma using standards of selenomethionine (SeMet), Se-methylselenocysteine (MeSeCys), selenite, and the selenosugar Se-methylseleno-N-acetylgalactosamine (SeGal) for linearity, precision, recoveries, and limits of detection and quantitation with satisfactory results. The method was applied for analysis of a set of plasma samples from cancer patients receiving selenite treatment in a clinical trial. Three LMW selenium compounds were observed. The main compounds, SeGal and selenite were tentatively identified by retention time matching with standards in different chromatographic systems, while the third minor compound was not identified. The identity of the selenosugar was verified by ESI-MS-MS product ion scanning, while selenite was identified indirectly as the glutathione (GSH) reaction product, GS-Se-SG.

Selenium as an alternative peptide label - comparison to fluorophore-labelled penetratin.

In the present study, the impact on peptide properties of labelling peptides with the fluorophore TAMRA or the selenium (Se) containing amino acid SeMet was evaluated. Three differently labelled variants of the cell-penetrating peptide (CPP) penetratin (Pen) were synthesized, PenM(Se), TAMRA-PenM(Se) and TAMRA-Pen. The labelled peptides were characterized in terms of hydrodynamic radius, secondary structure during peptide-membrane interaction, effect on membrane leakage induction, uptake efficiency in HeLa cells. Furthermore, stability of peptides and identities of degradation products in cell media and cell lysate were evaluated. TAMRA-labelling increased the hydrodynamic radius of Pen and PenM(Se) significantly. Labelling with Se caused no or minimal changes in size, secondary structure and membrane leakage induction in concentration levels relevant for cellular uptake studies. Similar degradation patterns of all labelled peptides were observed in HBSS media; degradation was mainly due to oxidation. Cellular uptake was significantly higher for the TAMRA labelled peptides as compared to Se-labelled Pen. Extensive degradation was observed in media during cellular uptake studies, however, in all cell lysates, primarily the intact peptide (PenM(Se), TAMRA-PenM(Se) or TAMRA-Pen) was observed. Selenium labelling caused minimal alteration of the physicochemical properties of the peptide and allowed for absolute quantitative determination of cellular uptake by inductively coupled plasma mass spectrometry. Selenium is thus proposed as a promising alternative label for quantification of peptides in general, altering the properties of the peptide to a minor extent as compared to commonly used peptide labels.

The selenium metabolite methylselenol regulates the expression of ligands that trigger immune activation through the lymphocyte receptor NKG2D.

For decades, selenium research has been focused on the identification of active metabolites, which are crucial for selenium chemoprevention of cancer. In this context, the metabolite methylselenol (CH3SeH) is known for its action to selectively kill transformed cells through mechanisms that include increased formation of reactive oxygen species, induction of DNA damage, triggering of apoptosis, and inhibition of angiogenesis. Here we reveal that CH3SeH modulates the cell surface expression of NKG2D ligands. The expression of NKG2D ligands is induced by stress-associated pathways that occur early during malignant transformation and enable the recognition and elimination of tumors by activating the lymphocyte receptor NKG2D. CH3SeH regulated NKG2D ligands both on the transcriptional and the posttranscriptional levels. CH3SeH induced the transcription of MHC class I polypeptide-related sequence MICA/B and ULBP2 mRNA. However, the induction of cell surface expression was restricted to the ligands MICA/B. Remarkably, our studies showed that CH3SeH inhibited ULBP2 surface transport through inhibition of the autophagic transport pathway. Finally, we identified extracellular calcium as being essential for CH3SeH regulation of NKG2D ligands. A balanced cell surface expression of NKG2D ligands is considered to be an innate barrier against tumor development. Therefore, our work indicates that the application of selenium compounds that are metabolized to CH3SeH could improve NKG2D-based immune therapy.

Quantification of pharmaceutical peptides using selenium as an elemental detection label.

The aim of the present work was to demonstrate how selenium labelling of a synthetic cell-penetrating peptide may be employed in evaluation of stability and quantitative estimation of cellular uptake by inductively coupled plasma mass spectrometry (ICP-MS). Two analogues of the cell-penetrating peptide, penetratin, were synthesized, one with selenomethionine (SeMet) added at the N-terminal of the peptide (N-PenM(Se)) and the other with the internal methionine (Met) replaced with SeMet (i-PenM(Se)). The purity of the synthesized peptides was 92% for N-PenM(Se) and 89% for i-PenM(Se) as determined by liquid chromatography (LC)-ICP-MS. The selenium-labelled peptides were investigated by cell uptake studies in HeLa WT cells. The stability of the peptides was monitored in water, cell medium and during cell uptake studies. Total uptake of selenium was quantified by flow injection (FI)-ICP-MS. Speciation analysis of cell samples by LC-ICP-MS showed mainly uptake of the intact peptides, while the amount of intact peptides in cell lysates was semi-quantitatively determined. The selenium-containing penetratin analogues were to some extent degraded in pure cell medium, while an extensive degradation was observed during cell uptake studies. The major degradation products were determined by LC-electrospray ionization mass spectrometry (ESI-MS). The labelling method in combination with FI-ICP-MS, LC-ICP-MS and LC-ESI-MS techniques provided detailed information on the fate of penetratin in cellular uptake studies. Most pharmaceutical peptides, including penetratin, are synthetic analogues of endogenous peptides, and incorporation of selenium may improve the critical assessment of the native drug or drug delivery candidate early in the drug development process.

Absorption, distribution, metabolism and excretion of selenium following oral administration of elemental selenium nanoparticles or selenite in rats.

A suspension of nanoparticles of BSA-stabilized red amorphous elemental selenium (Se) or an aqueous solution of sodium selenite was repeatedly administered by oral gavage for 28 days at 0.05 mg kg(-1) bw per day (low dose) or at 0.5 mg kg(-1) bw per day (high dose) as Se to female rats. Prior to administration, the size distribution of the Se nanoparticles was characterized by dynamic light scattering and transmission electron microscopy, which showed that the particles' mean diameter was 19 nm and ranged in size from 10 to 80 nm. Following administration of the high dose of Se nanoparticles or selenite the concentration of Se was determined by ICP-MS in the liver, kidney, urine, feces, stomach, lungs, and plasma at the µg g(-1) level and in brain and muscle tissue at the sub-µg g(-1) level. In order to test if any elemental Se was present in the liver, kidney or feces, an in situ derivatization selective to elemental Se was performed by treatment with sulfite, which resulted in formation of the selenosulfate anion. This Se species was selectively and quantitatively determined by anion exchange HPLC and ICP-MS detection. The results showed that elemental Se was present in the livers, kidneys and feces of animals exposed to low and high doses of elemental Se nanoparticles or to selenite, and was also detected in the same samples from control animals. The fraction of Se present as elemental Se in livers and kidneys from the high dose animals was significantly larger than the similar fraction in samples from the low dose animals or from the controls. This suggested that the natural metabolic pathways of Se were exhausted when given the high dose of elemental Se or selenite resulting in a non-metabolized pool of elemental Se. Both dosage forms of Se were bioavailable as demonstrated by the blood biomarker selenoprotein P, which was equally up-regulated in the high-dose animals for both dosage forms of Se. Finally, the excretion of Se in urine and its occurrence as Se-methylseleno-N-acetyl-galactosamine and the trimethylselenonium-ion demonstrated that both dosage forms were metabolized and excreted. The results of the study showed that both forms of Se were equally absorbed, distributed, metabolized and excreted, but the detailed mechanism of the fate of the administered elemental Se or selenite in the gastro-intestinal tract of rats remains unclear.

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.

Development and validation of an ICP-OES method for quantitation of elemental impurities in tablets according to coming US pharmacopeia chapters.

May 1, 2014 the United States Pharmacopeia (USP) will implement two new chapters stating limit concentrations of elemental impurities in pharmaceuticals applying inductively coupled plasma methods. In the present work an inductively coupled plasma optical emission spectrometry (ICP-OES) method for quantitation of As, Cd, Cu, Cr, Fe, Hg, Ir, Mn, Mo, Ni, Os, Pb, Pd, Pt, Rh, Ru, V and Zn in tablets according to the new USP chapters was developed. Sample preparation was performed by microwave-assisted acid digestion using a mixture of 65% HNO3 and 37% HCl (3:1, v/v). Limits of detection and quantitation were at least a factor of ten below the USP limit concentrations showing that the ICP-OES technique is well suited for quantitation of elemental impurities. Excluding Os, spike recoveries in the range of 85.3-103.8% were obtained with relative standard deviations (%RSD) ranging from 1.3 to 3.2%. Due to memory effects the spike recovery and %RSD of Os were 161.5% and 13.7%, respectively, thus the method will need further development with respect to elimination of the memory effect of Os. The method was proven to be specific but with potential spectral interference for Ir, Os, Pb, Pt and Rh necessitating visual examination of the spectra. Hg memory effect was handled by using lower spike levels combined with rinsing with 0.1M HCl. The tablets had a content of Fe and Pt of 182.8 ± 18.1 and 2.8 ± 0.2 µg/g, respectively and did therefore not exceed the limit concentration defined by USP. It is suggested that the developed method is applicable to pharmaceutical products with a composition and maximal amount of daily intake (g drug product/day) similar to the tablets used in this work.

Metallomics in drug development: characterization of a liposomal cisplatin drug formulation in human plasma by CE-ICP-MS.

A capillary electrophoresis inductively coupled plasma mass spectrometry method for separation of free cisplatin from liposome-encapsulated cisplatin and protein-bound cisplatin was developed. A liposomal formulation of cisplatin based on PEGylated liposomes was used as model drug formulation. The effect of human plasma matrix on the analysis of liposome-encapsulated cisplatin and intact cisplatin was studied. The presence of 1 % of dextran and 4 mM of sodium dodecyl sulfate in HEPES buffer was demonstrated to be effective in improving the separation of liposomes and cisplatin bound to proteins in plasma. A detection limit of 41 ng/mL of platinum and a precision of 2.1 % (for 10 µg/mL of cisplatin standard) were obtained. Simultaneous measurements of phosphorous and platinum allows the simultaneous monitoring of the liposomes, liposome-encapsulated cisplatin, free cisplatin and cisplatin bound to plasma constituents in plasma samples. It was demonstrated that this approach is suitable for studies of the stability of liposome formulations as leakage of active drug from the liposomes and subsequent binding to biomolecules in plasma can be monitored. This methodology has not been reported before and will improve characterization of liposomal drugs during drug development and in studies on kinetics.

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 ((77)Se) is given orally to the test subject and a second isotope ((74)Se) 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 ((nat)Se) and enriched (77)Se. 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 (77)Se in male Wistar rats while maintaining the (nat)Se 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, (74)Se 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 (77)Se were employed for comparative purposes.

Human urinary excretion and metabolism of (82)Se-enriched selenite and selenate determined by LC-ICP-MS.

Urinary excretion of selenium after ingestion of isotope labeled selenite and selenate was studied in seven healthy volunteers, 4 men and 3 women (age 28-50 years). An aqueous solution containing 330 µL (82)Se-selenate (corresponding to 74.3 µg (82)Se) was given orally and urine samples were subsequently collected during the following 24 hours. The scheme was repeated four weeks later with a 280 µL (82)Se-selenite solution (corresponding to 74.4 µg (82)Se). The amount of total Se in the urine samples was determined by inductively coupled mass spectrometry. The mean total urinary excretion of (82)Se following (82)Se-selenate administration was 33.7% (range 15.6-42.5%) while the mean total excretion of (82)Se after (82)Se-selenite administration was 3.2% (range 2.8-3.9%) of the ingested amount. LC-ICPMS analysis of the urine samples showed that the majority of the selenium excreted after selenate ingestion was unchanged selenate for 6 of the individuals while one individual had metabolized a fraction (approx. 20%) of the selenate to selenosugar. Ingestion of 10 times larger doses of selenite in two individuals resulted in 13-23% excretion primarily excreted as selenosugar. These results show that the human metabolic pathways of selenite and selenate are different and indicate that not all selenate, although well absorbed, may be available for the beneficial health effects.

Investigation of a liposomal oxaliplatin drug formulation by capillary electrophoresis hyphenated to inductively coupled plasma mass spectrometry (CE-ICP-MS).

A capillary electrophoresis-inductively coupled plasma mass spectrometry (CE-ICP-MS) method was developed for separation of the free oxaliplatin drug substance from liposome-entrapped oxaliplatin. Simultaneous determination of phosphorous and platinum opened the possibility to simultaneously monitor the liposomes (phospholipids) and platinum-based drug. In order to suppress the interferences, argon gas was used as a collision gas in ICP-MS. A detection limit of 29 ng/mL of platinum and a precision of 2.9% (for 10 µg/mL of oxaliplatin standard) were obtained. Measurement of the total concentration of free and encapsulated oxaliplatin by CE-ICP-MS was compared with total determination by ICP-MS after microwave digestion and showed a good agreement. A liposomal formulation of oxaliplatin based on PEGylated liposomes was used as a model drug formulation. Studies of accelerated drug release induced by sonication and phospholipase A(2) catalyzed hydrolysis were performed. It was demonstrated that the CE-ICP-MS was an efficient in vitro characterization method in the development and quality assurance purposes of lipsome-based formulation of metallodrugs.

Surveying selenium speciation from soil to cell--forms and transformations.

The aim of this review is to present and evaluate the present knowledge of which selenium species are available to the general population in the form of food and common supplements and how these species are metabolized in mammals. The overview of the selenium sources takes a horizontal approach, which encompasses identification of new metabolites in yeast and food of plant and animal origin, whereas the survey of the mammalian metabolism takes a horizontal as well as a vertical approach. The vertical approach encompasses studies on dynamic conversions of selenium compounds within cells, tissues or whole organisms. New and improved sample preparation, separation and detection methods are evaluated from an analytical chemical perspective to cover the progress in horizontal speciation, whereas the analytical methods for the vertical speciation and the interpretations of the results are evaluated from a biological angle as well.

Investigation of the selenium metabolism in cancer cell lines.

The aim of this work was to compare different selenium species for their ability to induce cell death in different cancer cell lines, while investigating the underlying chemistry by speciation analysis. A prostate cancer cell line (PC-3), a colon cancer cell line (HT-29) and a leukaemia cell line (Jurkat E6-1) were incubated with five selenium compounds representing inorganic as well as organic Se compounds in different oxidation states. Selenomethionine (SeMet), Se-methylselenocysteine (MeSeCys), methylseleninic acid (MeSeA), selenite and selenate in the concentration range 5-100 µM were incubated with cells for 24 h and the induction of cell death was measured using flow cytometry. The amounts of total selenium in cell medium, cell lysate and the insoluble fractions was determined by ICP-MS. Speciation analysis of cellular fractions was performed by reversed phase, anion exchange and size exclusion chromatography and ICP-MS detection. The selenium compounds exhibited large differences in their ability to induce cell death in the three cell lines and the susceptibilities of the cell lines were different. Full recovery of selenium in the cellular fractions was observed for all Se compounds except MeSeA. Speciation analysis showed that MeSeA was completely transformed during the incubations, while metabolic conversion of the other Se compounds was limited. Production of volatile dimethyl diselenide was observed for MeSeA and MeSeCys. MeSeA, MeSeCys and selenite showed noticeable protein binding. Correlations between cell death induction and the Se compounds transformations could not be demonstrated.

Complementary use of molecular and element-specific mass spectrometry for identification of selenium compounds related to human selenium metabolism.

The aim of this paper is to give an overview of analytical data on the identification of selenium compounds in biological samples with relevance for selenium metabolism. Only studies applying the combination of element-specific inductively coupled plasma mass spectrometry as well as molecular electrospray mass spectrometry detection have been included. Hence, selenium compounds are only considered identified if molecular mass spectra obtained by analysis of the authentic biological sample have been provided. Selenium compounds identified in selenium-accumulating plants and yeast are included, as extracts from such plants and yeast have been widely used for examination of the cancer-preventive effect of selenium in cell lines, animal models and human intervention trials. Hence, these selenium compounds are available for absorption and further metabolism. Identification of selenium metabolites in simulated gastric and intestinal juice, intestinal epithelial tissue, liver and urine is described. Hence, selenium metabolites identified in relation to absorption, metabolism and excretion are included.