Dataset Information

Quantitative analysis of vitamin D and its main metabolites in human milk by supercritical fluid chromatography coupled to tandem mass spectrometry.

ABSTRACT: A novel method to quantitate vitamin D and its main metabolites (vitamin D3, vitamin D2, and their 25-hydroxy metabolites) in breast milk by supercritical fluid chromatography has been developed and fully validated. A small volume of sample (1 mL) is subjected to ethanolic protein precipitation and liquid-liquid extraction. Final extracts are derivatized with 4-phenyl-1,2,4-triazoline-3,5-dione and vitamin D derivatives analyzed by supercritical fluid chromatography hyphenated to tandem mass spectrometry with atmospheric pressure chemical ionization. Multiple reaction monitoring is used for quantitation. Separation conditions were optimized using a gradient of methanol-water-ammonium formate into carbon dioxide. Make-up solvent was methanol containing ammonium formate. The quantitation limit reached levels as low as 50 pmol/L, with intra- and inter-day relative standard deviations lower than 15% and 20% for all analytes. Accuracy was evaluated by spiking experiments and was well within acceptability ratios (± 15%). The method was then applied to a subset of commercially available human milk samples. The newly developed method provides opportunities to determine the nutritional status of mother-infant dyads from a non-invasive measure, or for interventional or observational studies building knowledge on the composition of human milk. Graphical abstract.

SUBMITTER: Oberson JM

PROVIDER: S-EPMC6992569 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Quantitative analysis of vitamin D and its main metabolites in human milk by supercritical fluid chromatography coupled to tandem mass spectrometry.

Oberson J M JM Bénet S S Redeuil K K Campos-Giménez E E

Analytical and bioanalytical chemistry 20191212 2

A novel method to quantitate vitamin D and its main metabolites (vitamin D3, vitamin D2, and their 25-hydroxy metabolites) in breast milk by supercritical fluid chromatography has been developed and fully validated. A small volume of sample (1 mL) is subjected to ethanolic protein precipitation and liquid-liquid extraction. Final extracts are derivatized with 4-phenyl-1,2,4-triazoline-3,5-dione and vitamin D derivatives analyzed by supercritical fluid chromatography hyphenated to tandem mass spe ...[more]

PMID: 31832707

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Project description:Supercritical fluid chromatography (SFC) is frequently used for the analysis and separation of non-polar metabolites, but remains relatively underutilised for the study of polar molecules, even those which pose difficulties with established reversed-phase (RP) or hydrophilic interaction liquid chromatographic (HILIC) methodologies. Here, we present a fast SFC-MS method for the analysis of medium and high-polarity (-7?cLogP?2) compounds, designed for implementation in a high-throughput metabonomics setting. Sixty polar analytes were first screened to identify those most suitable for inclusion in chromatographic test mixtures; then, a multi-dimensional method development study was conducted to determine the optimal choice of stationary phase, modifier additive and temperature for the separation of such analytes using SFC. The test mixtures were separated on a total of twelve different column chemistries at three different temperatures, using CO2-methanol-based mobile phases containing a variety of polar additives. Chromatographic performance was evaluated with a particular emphasis on peak capacity, overall resolution, peak distribution and repeatability. The results suggest that a new generation of stationary phases, specifically designed for improved robustness in mixed CO2-methanol mobile phases, can improve peak shape, peak capacity and resolution for all classes of polar analytes. A significant enhancement in chromatographic performance was observed for these urinary metabolites on the majority of the stationary phases when polar additives such as ammonium salts (formate, acetate and hydroxide) were included in the organic modifier, and the use of water or alkylamine additives was found to be beneficial for specific subsets of polar analytes. The utility of these findings was confirmed by the separation of a mixture of polar metabolites in human urine using an optimised 7min gradient SFC method, where the use of the recommended column and co-solvent combination resulted in a significant improvement in chromatographic performance.

Dataset Information

Quantitative analysis of vitamin D and its main metabolites in human milk by supercritical fluid chromatography coupled to tandem mass spectrometry.

Publications

Quantitative analysis of vitamin D and its main metabolites in human milk by supercritical fluid chromatography coupled to tandem mass spectrometry.

Similar Datasets

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