title: Breast milk metabolome characterization in a single-phase extraction, multiplatform analytical approach.
abstract: Breast milk (BM) is a biofluid that has a fundamental role in early life nutrition and has direct impact on growth, neurodevelopment, and health. Global metabolic profiling is increasingly being utilized to characterize complex metabolic changes in biological samples. However, in order to achieve broad metabolite coverage, it is necessary to employ more than one analytical platform, typically requiring multiple sample preparation protocols. In an effort to improve analytical efficiency and retain comprehensive coverage of the metabolome, a new extraction methodology was developed that successfully retains metabolites from BM in a single-phase using an optimized methyl-tert-butyl ether solvent system. We conducted this single-phase extraction procedure on a representative pool of BM, and characterized the metabolic composition using LC-QTOF-MS and GC-Q-MS for polar and lipidic metabolites. To ensure that the extraction method was reproducible and fit-for-purpose, the analytical procedure was evaluated on both platforms using 18 metabolites selected to cover a range of chromatographic retention times and biochemical classes. Having validated the method, the metabolic signature of BM composition was mapped as a metabolic reaction network highlighting interconnected biological pathways and showing that the LC-MS and GC-MS platforms targeted largely different domains of the network. Subsequently, the same protocol was applied to ascertain compositional differences between BM at week 1 (n = 10) and 4 weeks (n = 9) post-partum. This single-phase approach is more efficient in terms of time, simplicity, cost, and sample volume than the existing two-phase methods and will be suited to high-throughput metabolic profiling studies of BM.
authors: Centre for Metabolomics and Bioanalysis (CEMBIO), Faculty of Pharmacy, Universidad San Pablo CEU, Campus Monteprincipe , Boadilla del Monte, 28668 Madrid, Spain.
authors: Alma
authors: A
authors: Villaseñor
authors: Isabel
authors: I
authors: Garcia-Perez
authors: Antonia
authors: A
authors: Garcia
authors: Joram M
authors: JM
authors: Posma
authors: Mariano
authors: M
authors: Fernández-López
authors: Andreas J
authors: AJ
authors: Nicholas
authors: Neena
authors: N
authors: Modi
authors: Elaine
authors: E
authors: Holmes
authors: Coral
authors: C
authors: Barbas
mesh_terms: D005591:Chemical Fractionation / Q000379:methods*; D002853:Chromatography, Liquid / Q000379:methods; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D055442:Metabolome; D055432:Metabolomics / Q000379:methods*; D008738:Methyl Ethers / Q000737:chemistry; D008895:Milk, Human / Q000737:chemistry* / Q000378:metabolism; D012997:Solvents / Q000737:chemistry
chemical_list: D008738:Methyl Ethers; D012997:Solvents; C043243:methyl tert-butyl ether