A high efficiency chromatographic separation on a porous graphitic carbon stationary phase was de... more A high efficiency chromatographic separation on a porous graphitic carbon stationary phase was developed for a large-scale separation of selenium metabolites in Se-rich yeast prior to their identification by electrospray hybrid quadrupole trap/Orbitrap mass spectrometry (Orbitrap MS(n)). The reversed-phase (RP) separation mode offered distinctly higher separation efficiency than the hydrophilic ion interaction (HILIC) mode. The latter was nevertheless complementary and useful to validate the detection of several compounds. The method allowed the detection of 64 metabolites including 30 SeSe or SeS conjugates (3 triple S/Se/S ones) and 14 selenoethers. 21 previously unreported metabolites were detected on the basis of the selenium isotopic pattern usually matched with the sub-ppm mass accuracy. 9 of these metabolites were subsequently identified using the multi-stage high mass accuracy (<5ppm) mass spectrometry. The identified metabolites (and their groups) were quantified on-line by ICP-MS fitted with a frequency-matching generator allowing a quasi-uniform response over the large (20-90%) acetonitrile mobile phase concentration range. The morphology of HPLC-ICP-MS chromatograms was remarkably similar to that of HPLC multi-ion extracted ESI-MS chromatograms. The detection limits obtained by ICP MS and ESI MS were 1 and 2ppb, respectively.
A proteomics approach based on 2D gel electrophoresis followed by HPLC-electrospray Orbitrap MS/M... more A proteomics approach based on 2D gel electrophoresis followed by HPLC-electrospray Orbitrap MS/MS was developed to investigate the replacement and the degree of the Se/S substitution in methionine and cysteine in Se-rich yeast. Capillary HPLC-inductively coupled plasma mass spectrometry (ICP-MS), employed in parallel to capHPLC-ESI MS, indicated the virtual independence of the ESI MS response of the peptide structure (in the elution range of 30-65% methanol), and hence, the use of ESI MS data to determine the SeCys/Cys and SeMet/Met substitution ratios. For the first time a considerable incorporation of selenocysteine (SeCys) in proteins of the yeast proteome despite the absence of the UGA codon was demonstrated. The SeMet/Met and SeCys/Cys ratios were determined in a large number of peptides (57 and 26, respectively) issued from the tryptic digestion of 19 Se-containing proteins located in the gel by laser ablation-ICP MS imaging. The average Se/S substitution in methionine was 42.9±35.0 and was protein dependent with ratios ranging from 5 to 160 for individual peptides. The substitution of sulphur in cysteine (14.1±4.8%) in the cysteine-containing peptides was relatively similar (ratios from 9 to 23). Taking into account that the cysteine/methionine average ratio (2:1) in the yeast protein fraction, the study allowed the conclusion that 10-15% of selenium present in Se-enriched yeast is in the form of selenocysteine making up the mass balance of selenium species. For the first time a considerable incorporation of selenocysteine (SeCys) in proteins of the yeast proteome despite the absence of the UGA codon was demonstrated. It was achieved using a proteomics approach based on 2D gel electrophoresis followed by HPLC-electrospray Orbitrap MS/MS in order to investigate the replacement and the degree of the Se/S substitution in methionine and cysteine in Se-rich yeast.
An accurate and precise method for the determination of arsenobetaine (AsB, (CH(3))(3)(+)AsCH(2)C... more An accurate and precise method for the determination of arsenobetaine (AsB, (CH(3))(3)(+)AsCH(2)COO(-)) in fish samples using exact matching species specific isotope dilution (ID) liquid chromatography LTQ-Orbitrap mass spectrometry (LC-LTQ-Orbitrap-MS) and standard addition LC inductively coupled plasma mass spectrometry (LC-ICPMS) is described. Samples were extracted by sonication for 30 min with high purity deionized water. An in-house synthesized (13)C enriched AsB spike was used for species specific ID analysis whereas natural abundance AsB, synthesized and characterized by quantitative (1)H NMR (nuclear magnetic resonance spectroscopy), was used for reverse ID and standard addition LC-ICPMS. With the LTQ-Orbitrap-MS instrument in scan mode (m/z 170-190) and resolution set at 7500, the intensities of [M + H](+) ions at m/z of 179.0053 and 180.0087 were used to calculate the 179.0053/180.0087 ion ratio for quantification of AsB in fish tissues. To circumvent potential difficulty in mass bias correction, an exact matching approach was applied. A quantitatively prepared mixture of the natural abundance AsB standard and the enriched spike to give a ratio near one was used for mass bias correction. Concentrations of 9.65 ± 0.24 and 11.39 ± 0.39 mg kg(-1) (expanded uncertainty, k = 2) for AsB in two fish samples of fish1 and fish2, respectively, were obtained by ID LC-LTQ-Orbitrap-MS. These results are in good agreement with those obtained by standard addition LC-ICPMS, 9.56 ± 0.32 and 11.26 ± 0.44 mg kg(-1) (expanded uncertainty, k = 2), respectively. Fish CRM DORM-2 was used for method validation and measured results of 37.9 ± 1.8 and 38.7 ± 0.66 mg kg(-1) (expanded uncertainty, k = 2) for AsB obtained by standard addition LC-ICPMS and ID LC-LTQ-Orbitrap-MS, respectively, are in good agreement with the certified value of 39.0 ± 2.6 mg kg(-1) (expanded uncertainty, k = 2). Detection limits of 0.011 and 0.033 mg kg(-1) for AsB with LC-ICPMS and ID LC-LTQ-Orbitrap-MS, respectively, were obtained demonstrating that the technique is well suited to the determination AsB in fish samples. To the best of our knowledge, this is first application of species specific isotope dilution for the accurate and precise determination of AsB in biological tissues.
ABSTRACT Orthogonal liquid chromatographic (ion exchange, reversed phase, and ion pairing) and ma... more ABSTRACT Orthogonal liquid chromatographic (ion exchange, reversed phase, and ion pairing) and mass spectrometric [electrospray ionization (ESI)-TOF-MS, ESI-Orbitrap MS, and inductively coupled plasma mass spectrometry (ICP-MS)] methods were addressed to identify and quantify selenium species from a naturally Se-enriched green bean (Phaseolus vulgaris vulgaris) sample after proteolytic digestion. While selenomethionine (10.1 mg/kg as Se) and selenate (9.5 mg/kg as Se) could be quantified in a straightforward way by anion exchange LC-ICP-MS technique, a multistep purification protocol was required to identify Se-methylselenocysteine and γ-glutamyl-Se-methylselenocysteine in an unambiguous way prior to quantification by using either in-source fragmentation (LC-ESI-TOF-MS) or collision-induced dissociation (LC-ESI-Orbitrap MS). Finally, Se-methylselenocysteine (2.6 mg/kg as Se) and γ-glutamyl-Se-methylselenocysteine (1.2 mg/kg as Se) could contribute to the overall selenium recovery of 72 %. This sample is the first of the Faboideae subfamily and Phaseolus ssp. to be speciated to such an extent for selenium including γ-glutamyl-Se-methylselenocysteine, a highly potential selenium species, which makes this bean material an ideal candidate for functional food purposes.
Tellurium (Te) is a widely used metalloid in industry because of its unique chemical and physical... more Tellurium (Te) is a widely used metalloid in industry because of its unique chemical and physical properties. However, information about the biological and toxicological activities of Te in plants and animals is limited. Although Te is expected to be metabolized in organisms via the same pathway as sulfur and selenium (Se), no precise metabolic pathways are known in organisms, particularly in plants. To reveal the metabolic pathway of Te in plants, garlic, a well-known Se accumulator, was chosen as the model plant. Garlic was hydroponically cultivated and exposed to sodium tellurate, and Te-containing metabolites in the water extract of garlic leaves were identified using HPLC coupled with inductively coupled plasma mass spectrometry (ICP-MS) or electrospray tandem mass spectrometry (ESI-MS-MS). At least three Te-containing metabolites were detected using HPLC-ICP-MS, and two of them were subjected to HPLC-ESI-MS-MS for identification. The MS spectra obtained by ESI-MS-MS indicated that the metabolite was Te-methyltellurocysteine oxide (MeTeCysO). Then, MeTeCysO was chemically synthesized and its chromatographic behavior matched with that of the Te-containing metabolite in garlic. The other was assigned as cysteine S-methyltellurosulfide. These results suggest that garlic can assimilate tellurate, an inorganic Te compound, and tellurate is transformed into a Te-containing amino acid, the so-called telluroamino acid. This is the first report addressing that telluroamino acid is de novo synthesized in a higher plant.
The identification of the ligands binding Al is essential to understand the mechanisms by which p... more The identification of the ligands binding Al is essential to understand the mechanisms by which plants detoxify Al internally. However, studies concerning the speciation of Al have been frustrated by its complex chemistry. This work describes the identification of the tri-Al tricitrate (Al3cit3) complex in Plantago almogravensis, encompassing an integrated mass spectrometry approach based on hydrophilic interaction liquid chromatography (HILIC) and parallel detection by ICP-MS and ESI-MS/MS. This work also reports that both Al and Fe are bound by tricitrate, sometimes simultaneously, and the consequences of this finding are discussed. Of the complexes separated by size exclusion chromatography, Al3cit3 is the most stable occurring in P. almogravensis as it was the only one recovered after HILIC. This approach provided new information on the mechanism of Al detoxification in P. almogravensis, namely that Al is bound by the organic acid citrate and that the relative concentration of the detected complexes is affected by the organ type and internal Al concentration, and has potential for studying the speciation of Al in less tolerant plants.
Speciation analysis of selenium in human urine allowed for the first time the identification of a... more 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.
Currently, several mathematical methods exist to address simultaneous species formation and degra... more Currently, several mathematical methods exist to address simultaneous species formation and degradation using multiple spiking isotope dilution mass spectrometry. While all of these strategies have been compared numerically, comparison of the underlying principles is lacking. Owing to recent interest in using the species inter-conversion factors, mainly to study the quality of analytical methods, this manuscript reviews the mathematical logic and inconsistencies of the existing double or triple spiking isotope dilution models. Systematic terminology is also introduced to clarify the species inter-conversion coefficient definitions.
Strong cation exchange HPLC with the parallel ICP MS and electrospray hybrid linear ion trap quad... more Strong cation exchange HPLC with the parallel ICP MS and electrospray hybrid linear ion trap quadrupole orbital trap mass spectrometry (ESI Orbitrap MS) detection was developed for the study of the metabolomic pattern of selenium in selenium-rich yeast. The mobile phase composition (gradient of ammonium formate in 20% methanol) was optimized to obtain separation in conditions guaranteeing the identical ICP MS sensitivity during the entire chromatographic run and the compatibility with electrospray ionization. Twenty seven Se-containing metabolites observed in the HPLC-ICP MS chromatogram were identified by ESI Orbitrap MS based on the Se isotopic pattern, the accurate molecular mass, and the multistage fragmentation patterns. The method allowed for the first time the correlation of the differences observed in HPLC-ICP MS chromatography of water extracts of Se-rich yeast samples from different manufacturers with the identity of the eluted compounds determined by ESI MS.
Several novel selenium containing compounds were characterized in staple crops (wheat, rice and m... more Several novel selenium containing compounds were characterized in staple crops (wheat, rice and maize) grown on soils naturally rich in selenium. A dedicated method based on the coupling of liquid chromatography with multiplexed detection (ICP-MS, ESI-Orbitrap MS(/MS)) was developed for the speciation of low-molecular weight (<5 kDa) selenium metabolites. Nine species present in different proportions as a function of the crop type were identified by cation-exchange HPLC-ESI-Orbitrap MS on the basis of the accurate molecular mass and MS/MS spectra. The natural origin of these species was then validated by varying extraction conditions and by using hydrophilic interaction LC (HILIC)-ESI-Orbitrap MS(/MS). Among the identified compounds, Se-containing monosaccharides (hexose moiety, m/z 317 and m/z 358) or Se-containing disaccharides (hexose-pentose moiety, m/z 407 and m/z 408) were the first selenosugars reported in edible plants. It is also the first report of the presence of 2,3-dihydroxypropionyl-selenolanthionine (m/z 345) in rice. Because these crops can be an important source of selenium in animal and human nutrition, the understanding of the origin and the fate of these species during metabolic processes will be of great interest.
A high efficiency chromatographic separation on a porous graphitic carbon stationary phase was de... more A high efficiency chromatographic separation on a porous graphitic carbon stationary phase was developed for a large-scale separation of selenium metabolites in Se-rich yeast prior to their identification by electrospray hybrid quadrupole trap/Orbitrap mass spectrometry (Orbitrap MS(n)). The reversed-phase (RP) separation mode offered distinctly higher separation efficiency than the hydrophilic ion interaction (HILIC) mode. The latter was nevertheless complementary and useful to validate the detection of several compounds. The method allowed the detection of 64 metabolites including 30 SeSe or SeS conjugates (3 triple S/Se/S ones) and 14 selenoethers. 21 previously unreported metabolites were detected on the basis of the selenium isotopic pattern usually matched with the sub-ppm mass accuracy. 9 of these metabolites were subsequently identified using the multi-stage high mass accuracy (<5ppm) mass spectrometry. The identified metabolites (and their groups) were quantified on-line by ICP-MS fitted with a frequency-matching generator allowing a quasi-uniform response over the large (20-90%) acetonitrile mobile phase concentration range. The morphology of HPLC-ICP-MS chromatograms was remarkably similar to that of HPLC multi-ion extracted ESI-MS chromatograms. The detection limits obtained by ICP MS and ESI MS were 1 and 2ppb, respectively.
A proteomics approach based on 2D gel electrophoresis followed by HPLC-electrospray Orbitrap MS/M... more A proteomics approach based on 2D gel electrophoresis followed by HPLC-electrospray Orbitrap MS/MS was developed to investigate the replacement and the degree of the Se/S substitution in methionine and cysteine in Se-rich yeast. Capillary HPLC-inductively coupled plasma mass spectrometry (ICP-MS), employed in parallel to capHPLC-ESI MS, indicated the virtual independence of the ESI MS response of the peptide structure (in the elution range of 30-65% methanol), and hence, the use of ESI MS data to determine the SeCys/Cys and SeMet/Met substitution ratios. For the first time a considerable incorporation of selenocysteine (SeCys) in proteins of the yeast proteome despite the absence of the UGA codon was demonstrated. The SeMet/Met and SeCys/Cys ratios were determined in a large number of peptides (57 and 26, respectively) issued from the tryptic digestion of 19 Se-containing proteins located in the gel by laser ablation-ICP MS imaging. The average Se/S substitution in methionine was 42.9±35.0 and was protein dependent with ratios ranging from 5 to 160 for individual peptides. The substitution of sulphur in cysteine (14.1±4.8%) in the cysteine-containing peptides was relatively similar (ratios from 9 to 23). Taking into account that the cysteine/methionine average ratio (2:1) in the yeast protein fraction, the study allowed the conclusion that 10-15% of selenium present in Se-enriched yeast is in the form of selenocysteine making up the mass balance of selenium species. For the first time a considerable incorporation of selenocysteine (SeCys) in proteins of the yeast proteome despite the absence of the UGA codon was demonstrated. It was achieved using a proteomics approach based on 2D gel electrophoresis followed by HPLC-electrospray Orbitrap MS/MS in order to investigate the replacement and the degree of the Se/S substitution in methionine and cysteine in Se-rich yeast.
An accurate and precise method for the determination of arsenobetaine (AsB, (CH(3))(3)(+)AsCH(2)C... more An accurate and precise method for the determination of arsenobetaine (AsB, (CH(3))(3)(+)AsCH(2)COO(-)) in fish samples using exact matching species specific isotope dilution (ID) liquid chromatography LTQ-Orbitrap mass spectrometry (LC-LTQ-Orbitrap-MS) and standard addition LC inductively coupled plasma mass spectrometry (LC-ICPMS) is described. Samples were extracted by sonication for 30 min with high purity deionized water. An in-house synthesized (13)C enriched AsB spike was used for species specific ID analysis whereas natural abundance AsB, synthesized and characterized by quantitative (1)H NMR (nuclear magnetic resonance spectroscopy), was used for reverse ID and standard addition LC-ICPMS. With the LTQ-Orbitrap-MS instrument in scan mode (m/z 170-190) and resolution set at 7500, the intensities of [M + H](+) ions at m/z of 179.0053 and 180.0087 were used to calculate the 179.0053/180.0087 ion ratio for quantification of AsB in fish tissues. To circumvent potential difficulty in mass bias correction, an exact matching approach was applied. A quantitatively prepared mixture of the natural abundance AsB standard and the enriched spike to give a ratio near one was used for mass bias correction. Concentrations of 9.65 ± 0.24 and 11.39 ± 0.39 mg kg(-1) (expanded uncertainty, k = 2) for AsB in two fish samples of fish1 and fish2, respectively, were obtained by ID LC-LTQ-Orbitrap-MS. These results are in good agreement with those obtained by standard addition LC-ICPMS, 9.56 ± 0.32 and 11.26 ± 0.44 mg kg(-1) (expanded uncertainty, k = 2), respectively. Fish CRM DORM-2 was used for method validation and measured results of 37.9 ± 1.8 and 38.7 ± 0.66 mg kg(-1) (expanded uncertainty, k = 2) for AsB obtained by standard addition LC-ICPMS and ID LC-LTQ-Orbitrap-MS, respectively, are in good agreement with the certified value of 39.0 ± 2.6 mg kg(-1) (expanded uncertainty, k = 2). Detection limits of 0.011 and 0.033 mg kg(-1) for AsB with LC-ICPMS and ID LC-LTQ-Orbitrap-MS, respectively, were obtained demonstrating that the technique is well suited to the determination AsB in fish samples. To the best of our knowledge, this is first application of species specific isotope dilution for the accurate and precise determination of AsB in biological tissues.
ABSTRACT Orthogonal liquid chromatographic (ion exchange, reversed phase, and ion pairing) and ma... more ABSTRACT Orthogonal liquid chromatographic (ion exchange, reversed phase, and ion pairing) and mass spectrometric [electrospray ionization (ESI)-TOF-MS, ESI-Orbitrap MS, and inductively coupled plasma mass spectrometry (ICP-MS)] methods were addressed to identify and quantify selenium species from a naturally Se-enriched green bean (Phaseolus vulgaris vulgaris) sample after proteolytic digestion. While selenomethionine (10.1 mg/kg as Se) and selenate (9.5 mg/kg as Se) could be quantified in a straightforward way by anion exchange LC-ICP-MS technique, a multistep purification protocol was required to identify Se-methylselenocysteine and γ-glutamyl-Se-methylselenocysteine in an unambiguous way prior to quantification by using either in-source fragmentation (LC-ESI-TOF-MS) or collision-induced dissociation (LC-ESI-Orbitrap MS). Finally, Se-methylselenocysteine (2.6 mg/kg as Se) and γ-glutamyl-Se-methylselenocysteine (1.2 mg/kg as Se) could contribute to the overall selenium recovery of 72 %. This sample is the first of the Faboideae subfamily and Phaseolus ssp. to be speciated to such an extent for selenium including γ-glutamyl-Se-methylselenocysteine, a highly potential selenium species, which makes this bean material an ideal candidate for functional food purposes.
Tellurium (Te) is a widely used metalloid in industry because of its unique chemical and physical... more Tellurium (Te) is a widely used metalloid in industry because of its unique chemical and physical properties. However, information about the biological and toxicological activities of Te in plants and animals is limited. Although Te is expected to be metabolized in organisms via the same pathway as sulfur and selenium (Se), no precise metabolic pathways are known in organisms, particularly in plants. To reveal the metabolic pathway of Te in plants, garlic, a well-known Se accumulator, was chosen as the model plant. Garlic was hydroponically cultivated and exposed to sodium tellurate, and Te-containing metabolites in the water extract of garlic leaves were identified using HPLC coupled with inductively coupled plasma mass spectrometry (ICP-MS) or electrospray tandem mass spectrometry (ESI-MS-MS). At least three Te-containing metabolites were detected using HPLC-ICP-MS, and two of them were subjected to HPLC-ESI-MS-MS for identification. The MS spectra obtained by ESI-MS-MS indicated that the metabolite was Te-methyltellurocysteine oxide (MeTeCysO). Then, MeTeCysO was chemically synthesized and its chromatographic behavior matched with that of the Te-containing metabolite in garlic. The other was assigned as cysteine S-methyltellurosulfide. These results suggest that garlic can assimilate tellurate, an inorganic Te compound, and tellurate is transformed into a Te-containing amino acid, the so-called telluroamino acid. This is the first report addressing that telluroamino acid is de novo synthesized in a higher plant.
The identification of the ligands binding Al is essential to understand the mechanisms by which p... more The identification of the ligands binding Al is essential to understand the mechanisms by which plants detoxify Al internally. However, studies concerning the speciation of Al have been frustrated by its complex chemistry. This work describes the identification of the tri-Al tricitrate (Al3cit3) complex in Plantago almogravensis, encompassing an integrated mass spectrometry approach based on hydrophilic interaction liquid chromatography (HILIC) and parallel detection by ICP-MS and ESI-MS/MS. This work also reports that both Al and Fe are bound by tricitrate, sometimes simultaneously, and the consequences of this finding are discussed. Of the complexes separated by size exclusion chromatography, Al3cit3 is the most stable occurring in P. almogravensis as it was the only one recovered after HILIC. This approach provided new information on the mechanism of Al detoxification in P. almogravensis, namely that Al is bound by the organic acid citrate and that the relative concentration of the detected complexes is affected by the organ type and internal Al concentration, and has potential for studying the speciation of Al in less tolerant plants.
Speciation analysis of selenium in human urine allowed for the first time the identification of a... more 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.
Currently, several mathematical methods exist to address simultaneous species formation and degra... more Currently, several mathematical methods exist to address simultaneous species formation and degradation using multiple spiking isotope dilution mass spectrometry. While all of these strategies have been compared numerically, comparison of the underlying principles is lacking. Owing to recent interest in using the species inter-conversion factors, mainly to study the quality of analytical methods, this manuscript reviews the mathematical logic and inconsistencies of the existing double or triple spiking isotope dilution models. Systematic terminology is also introduced to clarify the species inter-conversion coefficient definitions.
Strong cation exchange HPLC with the parallel ICP MS and electrospray hybrid linear ion trap quad... more Strong cation exchange HPLC with the parallel ICP MS and electrospray hybrid linear ion trap quadrupole orbital trap mass spectrometry (ESI Orbitrap MS) detection was developed for the study of the metabolomic pattern of selenium in selenium-rich yeast. The mobile phase composition (gradient of ammonium formate in 20% methanol) was optimized to obtain separation in conditions guaranteeing the identical ICP MS sensitivity during the entire chromatographic run and the compatibility with electrospray ionization. Twenty seven Se-containing metabolites observed in the HPLC-ICP MS chromatogram were identified by ESI Orbitrap MS based on the Se isotopic pattern, the accurate molecular mass, and the multistage fragmentation patterns. The method allowed for the first time the correlation of the differences observed in HPLC-ICP MS chromatography of water extracts of Se-rich yeast samples from different manufacturers with the identity of the eluted compounds determined by ESI MS.
Several novel selenium containing compounds were characterized in staple crops (wheat, rice and m... more Several novel selenium containing compounds were characterized in staple crops (wheat, rice and maize) grown on soils naturally rich in selenium. A dedicated method based on the coupling of liquid chromatography with multiplexed detection (ICP-MS, ESI-Orbitrap MS(/MS)) was developed for the speciation of low-molecular weight (<5 kDa) selenium metabolites. Nine species present in different proportions as a function of the crop type were identified by cation-exchange HPLC-ESI-Orbitrap MS on the basis of the accurate molecular mass and MS/MS spectra. The natural origin of these species was then validated by varying extraction conditions and by using hydrophilic interaction LC (HILIC)-ESI-Orbitrap MS(/MS). Among the identified compounds, Se-containing monosaccharides (hexose moiety, m/z 317 and m/z 358) or Se-containing disaccharides (hexose-pentose moiety, m/z 407 and m/z 408) were the first selenosugars reported in edible plants. It is also the first report of the presence of 2,3-dihydroxypropionyl-selenolanthionine (m/z 345) in rice. Because these crops can be an important source of selenium in animal and human nutrition, the understanding of the origin and the fate of these species during metabolic processes will be of great interest.
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Papers by Laurent Ouerdane