Comprehensive Vitamer Profiling of Folate Mono- and Polyglutamates in Baker’s Yeast (<i>Saccharomyces cerevisiae</i>) as a Function of Different Sample Preparation Procedures
Folates are a group of B<sub>9</sub> vitamins playing an important role in many metabolic processes such as methylation reactions, nucleotide synthesis or oxidation and reduction processes. However, humans are not able to synthesize folates de novo and thus rely on external sources there...
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| Format: | Article |
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MDPI AG
2020-07-01
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| Series: | Metabolites |
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| Online Access: | https://www.mdpi.com/2218-1989/10/8/301 |
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| author | Lena Gmelch Daniela Wirtz Michael Witting Nadine Weber Lisa Striegel Philippe Schmitt-Kopplin Michael Rychlik |
| author_facet | Lena Gmelch Daniela Wirtz Michael Witting Nadine Weber Lisa Striegel Philippe Schmitt-Kopplin Michael Rychlik |
| author_sort | Lena Gmelch |
| collection | DOAJ |
| description | Folates are a group of B<sub>9</sub> vitamins playing an important role in many metabolic processes such as methylation reactions, nucleotide synthesis or oxidation and reduction processes. However, humans are not able to synthesize folates de novo and thus rely on external sources thereof. Baker’s yeast (<i>Saccharomyces cerevisiae</i>) has been shown to produce high amounts of this vitamin but extensive identification of its folate metabolism is still lacking. Therefore, we optimized and compared different sample preparation and purification procedures applying solid phase extraction (SPE). Strong anion exchange (SAX), C18 and hydrophilic–lipophilic-balanced (HLB) materials were tested for their applicability in future metabolomics studies. SAX turned out to be the preferred material for the quantitative purification of folates. Qualification of several folate vitamers was achieved by ultra-high pressure liquid chromatography quadrupole time of flight mass spectrometry (UHPLC-Q-ToF-MS) measurements and quantification was performed by liquid chromatography tandem mass spectrometry (LC-MS/MS) applying stable isotope dilution assays (SIDAs). The oxidation product <i>s</i>-pyrazino-triazine (MeFox) was included into the SIDA method for total folate determination and validation. Applying the best protocol (SAX) in regard to folate recovery, we analyzed 32 different vitamers in different polyglutamate states up to nonaglutamates, of which we could further identify 26 vitamers based on tandem-MS (MS<sup>2</sup>) spectra. Total folate quantification revealed differences in formyl folate contents depending on the cartridge chemistry used for purification. These are supposedly a result of interconversion reactions occurring during sample preparation due to variation in pH adjustments for the different purification protocols. The occurrence of interconversion and oxidation reactions should be taken into consideration in sample preparation procedures for metabolomics analyses with a focus on folates. |
| first_indexed | 2024-03-10T18:16:45Z |
| format | Article |
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| institution | Directory Open Access Journal |
| issn | 2218-1989 |
| language | English |
| last_indexed | 2024-03-10T18:16:45Z |
| publishDate | 2020-07-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Metabolites |
| spelling | doaj.art-3b5a59b852d545759cc2576f60eb5cdc2023-11-20T07:40:09ZengMDPI AGMetabolites2218-19892020-07-0110830110.3390/metabo10080301Comprehensive Vitamer Profiling of Folate Mono- and Polyglutamates in Baker’s Yeast (<i>Saccharomyces cerevisiae</i>) as a Function of Different Sample Preparation ProceduresLena Gmelch0Daniela Wirtz1Michael Witting2Nadine Weber3Lisa Striegel4Philippe Schmitt-Kopplin5Michael Rychlik6Chair of Analytical Food Chemistry, Technical University of Munich, 85354 Freising-Weihenstephan, GermanyChair of Analytical Food Chemistry, Technical University of Munich, 85354 Freising-Weihenstephan, GermanyChair of Analytical Food Chemistry, Technical University of Munich, 85354 Freising-Weihenstephan, GermanyChair of Analytical Food Chemistry, Technical University of Munich, 85354 Freising-Weihenstephan, GermanyChair of Analytical Food Chemistry, Technical University of Munich, 85354 Freising-Weihenstephan, GermanyChair of Analytical Food Chemistry, Technical University of Munich, 85354 Freising-Weihenstephan, GermanyChair of Analytical Food Chemistry, Technical University of Munich, 85354 Freising-Weihenstephan, GermanyFolates are a group of B<sub>9</sub> vitamins playing an important role in many metabolic processes such as methylation reactions, nucleotide synthesis or oxidation and reduction processes. However, humans are not able to synthesize folates de novo and thus rely on external sources thereof. Baker’s yeast (<i>Saccharomyces cerevisiae</i>) has been shown to produce high amounts of this vitamin but extensive identification of its folate metabolism is still lacking. Therefore, we optimized and compared different sample preparation and purification procedures applying solid phase extraction (SPE). Strong anion exchange (SAX), C18 and hydrophilic–lipophilic-balanced (HLB) materials were tested for their applicability in future metabolomics studies. SAX turned out to be the preferred material for the quantitative purification of folates. Qualification of several folate vitamers was achieved by ultra-high pressure liquid chromatography quadrupole time of flight mass spectrometry (UHPLC-Q-ToF-MS) measurements and quantification was performed by liquid chromatography tandem mass spectrometry (LC-MS/MS) applying stable isotope dilution assays (SIDAs). The oxidation product <i>s</i>-pyrazino-triazine (MeFox) was included into the SIDA method for total folate determination and validation. Applying the best protocol (SAX) in regard to folate recovery, we analyzed 32 different vitamers in different polyglutamate states up to nonaglutamates, of which we could further identify 26 vitamers based on tandem-MS (MS<sup>2</sup>) spectra. Total folate quantification revealed differences in formyl folate contents depending on the cartridge chemistry used for purification. These are supposedly a result of interconversion reactions occurring during sample preparation due to variation in pH adjustments for the different purification protocols. The occurrence of interconversion and oxidation reactions should be taken into consideration in sample preparation procedures for metabolomics analyses with a focus on folates.https://www.mdpi.com/2218-1989/10/8/301folate metabolismfolate polyglutamatesbaker’s yeastsolid phase extraction<i>s</i>-pyrazino-triazine (MeFox) |
| spellingShingle | Lena Gmelch Daniela Wirtz Michael Witting Nadine Weber Lisa Striegel Philippe Schmitt-Kopplin Michael Rychlik Comprehensive Vitamer Profiling of Folate Mono- and Polyglutamates in Baker’s Yeast (<i>Saccharomyces cerevisiae</i>) as a Function of Different Sample Preparation Procedures Metabolites folate metabolism folate polyglutamates baker’s yeast solid phase extraction <i>s</i>-pyrazino-triazine (MeFox) |
| title | Comprehensive Vitamer Profiling of Folate Mono- and Polyglutamates in Baker’s Yeast (<i>Saccharomyces cerevisiae</i>) as a Function of Different Sample Preparation Procedures |
| title_full | Comprehensive Vitamer Profiling of Folate Mono- and Polyglutamates in Baker’s Yeast (<i>Saccharomyces cerevisiae</i>) as a Function of Different Sample Preparation Procedures |
| title_fullStr | Comprehensive Vitamer Profiling of Folate Mono- and Polyglutamates in Baker’s Yeast (<i>Saccharomyces cerevisiae</i>) as a Function of Different Sample Preparation Procedures |
| title_full_unstemmed | Comprehensive Vitamer Profiling of Folate Mono- and Polyglutamates in Baker’s Yeast (<i>Saccharomyces cerevisiae</i>) as a Function of Different Sample Preparation Procedures |
| title_short | Comprehensive Vitamer Profiling of Folate Mono- and Polyglutamates in Baker’s Yeast (<i>Saccharomyces cerevisiae</i>) as a Function of Different Sample Preparation Procedures |
| title_sort | comprehensive vitamer profiling of folate mono and polyglutamates in baker s yeast i saccharomyces cerevisiae i as a function of different sample preparation procedures |
| topic | folate metabolism folate polyglutamates baker’s yeast solid phase extraction <i>s</i>-pyrazino-triazine (MeFox) |
| url | https://www.mdpi.com/2218-1989/10/8/301 |
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