High-Throughput Analysis of Water-Soluble Forms of Choline and Related Metabolites in Human Milk by UPLC-MS/MS and Its Application
Choline and related metabolites are key factors in many metabolic processes, and insufficient supply can adversely affect reproduction and fetal development. Choline status is mainly regulated by intake, and human milk is the only choline source for exclusively breastfed infants. Further, maternal s...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2021-02-01
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| Series: | Frontiers in Nutrition |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fnut.2020.604570/full |
| _version_ | 1819057405114187776 |
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| author | Daniela Hampel Daniela Hampel Setareh Shahab-Ferdows Ngoc Nguyen Ngoc Nguyen Gilberto Kac Lindsay H. Allen Lindsay H. Allen |
| author_facet | Daniela Hampel Daniela Hampel Setareh Shahab-Ferdows Ngoc Nguyen Ngoc Nguyen Gilberto Kac Lindsay H. Allen Lindsay H. Allen |
| author_sort | Daniela Hampel |
| collection | DOAJ |
| description | Choline and related metabolites are key factors in many metabolic processes, and insufficient supply can adversely affect reproduction and fetal development. Choline status is mainly regulated by intake, and human milk is the only choline source for exclusively breastfed infants. Further, maternal status, genotype, and phenotype, as well as infant outcomes, have been related to milk choline concentrations. In order to enable the rapid assessment of choline intake for exclusively breastfed infants and to further investigate the associations between milk choline and maternal and infant status and other outcomes, we have developed a simplified method for the simultaneous analysis of human milk choline, glycerophosphocholine, phosphocholine, and the less abundant related metabolites betaine, carnitine, creatinine, dimethylglycine (DMG), methionine, and trimethylamine N-oxide (TMAO) using ultraperformance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS). These analytes have milk concentrations ranging over 3 orders of magnitude. Unlike other recently described LC-based methods, our approach does not require an ion-pairing reagent or high concentrations of solvent modifiers for successful analyte separation and thus avoid signal loss and potential permanent contamination. Milk samples (10 μl) were diluted (1:80) in water : methanol (1:4, v:v) and filtered prior to analysis with an optimized gradient of 0.1% propionic acidaq and acetonitrile, allowing efficient separation and removal of contaminants. Recovery rates ranged from 108.0 to 130.9% (inter-day variation: 3.3–9.6%), and matrix effects (MEs) from 54.1 to 114.3%. MEs were greater for carnitine, creatinine, and TMAO at lower dilution (1:40, p < 0.035 for all), indicating concentration-dependent ion suppression. Milk from Brazilian women (2–8, 28–50, and 88–119 days postpartum, ntotal = 53) revealed increasing concentration throughout lactation for glycerophosphocholine, DMG, and methionine, while carnitine decreased. Choline and phosphocholine were negatively correlated consistently at all three collection time intervals. The method is suitable for rapid analysis of human milk water-soluble forms of choline as well as previously not captured related metabolites with minimal sample volumes and preparation. |
| first_indexed | 2024-12-21T13:38:47Z |
| format | Article |
| id | doaj.art-ea7a14155a24424e972b84ca58648898 |
| institution | Directory Open Access Journal |
| issn | 2296-861X |
| language | English |
| last_indexed | 2024-12-21T13:38:47Z |
| publishDate | 2021-02-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Nutrition |
| spelling | doaj.art-ea7a14155a24424e972b84ca586488982022-12-21T19:02:06ZengFrontiers Media S.A.Frontiers in Nutrition2296-861X2021-02-01710.3389/fnut.2020.604570604570High-Throughput Analysis of Water-Soluble Forms of Choline and Related Metabolites in Human Milk by UPLC-MS/MS and Its ApplicationDaniela Hampel0Daniela Hampel1Setareh Shahab-Ferdows2Ngoc Nguyen3Ngoc Nguyen4Gilberto Kac5Lindsay H. Allen6Lindsay H. Allen7United States Department of Agriculture/Agricultural Research Service, Western Human Nutrition Research Center, Davis, CA, United StatesDepartment of Nutrition, University of California, Davis, Davis, CA, United StatesUnited States Department of Agriculture/Agricultural Research Service, Western Human Nutrition Research Center, Davis, CA, United StatesUnited States Department of Agriculture/Agricultural Research Service, Western Human Nutrition Research Center, Davis, CA, United StatesDepartment of Nutrition, University of California, Davis, Davis, CA, United StatesNutrition Institute, Rio de Janeiro Federal University, Rio de Janeiro, BrazilUnited States Department of Agriculture/Agricultural Research Service, Western Human Nutrition Research Center, Davis, CA, United StatesDepartment of Nutrition, University of California, Davis, Davis, CA, United StatesCholine and related metabolites are key factors in many metabolic processes, and insufficient supply can adversely affect reproduction and fetal development. Choline status is mainly regulated by intake, and human milk is the only choline source for exclusively breastfed infants. Further, maternal status, genotype, and phenotype, as well as infant outcomes, have been related to milk choline concentrations. In order to enable the rapid assessment of choline intake for exclusively breastfed infants and to further investigate the associations between milk choline and maternal and infant status and other outcomes, we have developed a simplified method for the simultaneous analysis of human milk choline, glycerophosphocholine, phosphocholine, and the less abundant related metabolites betaine, carnitine, creatinine, dimethylglycine (DMG), methionine, and trimethylamine N-oxide (TMAO) using ultraperformance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS). These analytes have milk concentrations ranging over 3 orders of magnitude. Unlike other recently described LC-based methods, our approach does not require an ion-pairing reagent or high concentrations of solvent modifiers for successful analyte separation and thus avoid signal loss and potential permanent contamination. Milk samples (10 μl) were diluted (1:80) in water : methanol (1:4, v:v) and filtered prior to analysis with an optimized gradient of 0.1% propionic acidaq and acetonitrile, allowing efficient separation and removal of contaminants. Recovery rates ranged from 108.0 to 130.9% (inter-day variation: 3.3–9.6%), and matrix effects (MEs) from 54.1 to 114.3%. MEs were greater for carnitine, creatinine, and TMAO at lower dilution (1:40, p < 0.035 for all), indicating concentration-dependent ion suppression. Milk from Brazilian women (2–8, 28–50, and 88–119 days postpartum, ntotal = 53) revealed increasing concentration throughout lactation for glycerophosphocholine, DMG, and methionine, while carnitine decreased. Choline and phosphocholine were negatively correlated consistently at all three collection time intervals. The method is suitable for rapid analysis of human milk water-soluble forms of choline as well as previously not captured related metabolites with minimal sample volumes and preparation.https://www.frontiersin.org/articles/10.3389/fnut.2020.604570/fullhuman milkwater-soluble cholinemetabolitesinter-relationshipsUPLC-MS/MS |
| spellingShingle | Daniela Hampel Daniela Hampel Setareh Shahab-Ferdows Ngoc Nguyen Ngoc Nguyen Gilberto Kac Lindsay H. Allen Lindsay H. Allen High-Throughput Analysis of Water-Soluble Forms of Choline and Related Metabolites in Human Milk by UPLC-MS/MS and Its Application Frontiers in Nutrition human milk water-soluble choline metabolites inter-relationships UPLC-MS/MS |
| title | High-Throughput Analysis of Water-Soluble Forms of Choline and Related Metabolites in Human Milk by UPLC-MS/MS and Its Application |
| title_full | High-Throughput Analysis of Water-Soluble Forms of Choline and Related Metabolites in Human Milk by UPLC-MS/MS and Its Application |
| title_fullStr | High-Throughput Analysis of Water-Soluble Forms of Choline and Related Metabolites in Human Milk by UPLC-MS/MS and Its Application |
| title_full_unstemmed | High-Throughput Analysis of Water-Soluble Forms of Choline and Related Metabolites in Human Milk by UPLC-MS/MS and Its Application |
| title_short | High-Throughput Analysis of Water-Soluble Forms of Choline and Related Metabolites in Human Milk by UPLC-MS/MS and Its Application |
| title_sort | high throughput analysis of water soluble forms of choline and related metabolites in human milk by uplc ms ms and its application |
| topic | human milk water-soluble choline metabolites inter-relationships UPLC-MS/MS |
| url | https://www.frontiersin.org/articles/10.3389/fnut.2020.604570/full |
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