Targeted peak integration was performed using SCIEX OS (version 2.1.6., SCIEX) with a maximum mass error of 10 ppm. The retention times were verified against authentic standards. In case of coelution, the targets were reported using the name or abbreviation of one of the targets followed by a hashtag. Details on the abbreviations used are listed in Table S2. For the polar to semi-polar metabolites, peak area was used for further data analysis, whereas for the bile and fatty acids, the area ratio of compounds to stable isotopically labelled standards was used. Data quality inspection was performed using an in-house quality assurance software performing between batch correction and removal of metabolites with high technical variance (RSD of QC below 30%).
"],"repository":["MetaboLights"],"study_status":["Public"],"ptm_modification":[""],"instrument_platform":["Liquid Chromatography MS - - - Liquid ChromatSTUDY PROTOCOLS","Liquid Chromatography MS - positive","Liquid Chromatography MS - - -"],"chromatography_protocol":["LC-MS analysis of bile acid and fatty acids (Assay 1): Chromatographic separation was performed on UPLC-TOF/MS system (AB Sciex, USA) using a Waters Acquity UPLC HSS T3 column (1.8 μm, 2.1 mm x 100 mm) with pre-column in-line stainless steel filter (0.3 μm, Agilent Technologies, Waldbronn, Germany). The flow rate was set at 0.4 mL/min, the column was kept at 45 °C, injection volume was 2 μL. Mobile phases A consisted of 10 mM ammonium formate in water/ACN (95:5, v:v), while mobile phase B 10 mM ammonium formate in MeOH/water (99:1, v:v). The gradient was as follows: starting at 100% A; 0.0-0.2 min, 70% B; 0.2-7.5 min 100% B; 7.5-11.5 min 100% B; 11.5-11.6 min 100% A; 11.6-15.0 100% A. Isopropanol was used as an external rinsing solution (2 s sip time + rinse port). The flow was directed to waste in the first minute of the run. The autosampler temperature was set at 10 °C.
LC-MS analysis of polar to semi polar metabolites (Assay 2 & 3):Analysis of polar to semi-polar metabolites were performed with a Shimadzu Nexera X2 LC system coupled to a TripleTOF 6600 mass spectrometer (SCIEX, Foster City, CA, USA), as described previously. Briefly, the LC separation was carried out at 40 °C using a Waters Acquity UPLC HSS T3 column (1.8 μm, 2.1 mm x 100 mm) with pre-column in-line stainless steel filter (0.3 μm, Agilent Technologies, Waldbronn, Germany). The mobile phase A was 0.1% FA in water, and the mobile phase B was 0.1% FA in ACN (Actu-all chemicals). With a flow rate of 0.4 mL/min and 1 μL of injection volume, the gradient starts at 100% A; 0.0-0.5 min 80% A; 0.5-2.5 min 2% A; 2.5-7.5 min 2% A; 7.5-12.0 min 2% A; 12.0-15.0 100% A. The autosampler temperature was set at 10 °C. To decelerate the contamination of the MS, the LC flow was diverted to waste at 7 min of the gradient by an external valve (Valco instruments, USA). During the analysis, the PCI compounds were continuously pumped by a binary Agilent 1260 Infinity pump (Agilent Technologies, Santa Clara, USA) at a flow rate of 20 µL/min and combined to the LC flow with a T-piece (IDEX, PEEK Tee, 0.02 Thru hole, F-300) before entering the ESI source.
"],"publication":["Fecal metabolome alterations in infants at risk of developing allergies during the first year of life."],"submitter_name":["Michael van Vliet"],"submitter_affiliation":["Leiden University"],"organism_part":["solvent","feces"],"technology_type":["mass spectrometry assay"],"disease":[""],"extraction_protocol":["The wet sample aliquots were lyophilized at 4 mbar and -110C for 20 h (Martin Christ Gefriertrocknungsanlagen GmbH, Germany), weighed (20± 0.2mg) and stored at -80C until extraction. Liquid-liquid extraction (LLE) was performed as described previously[1], Methyl tert-butyl ether (MTBE) / methanol (MeOH) / water was carried out on the lyophilized samples, with adjusted sample amount and doubled solvent-to-feces ratio. Briefly, 72uL of water and 216uL MeOH, containing stable isotopically labelled standards (SILs), were added to the 20 mg dry-weight fecal sample (less in case of insufficient sample material). Exact weights are provided in a separate excel file. After a 3-min vortex mixing (Marshall Scientific, Cambridge, UK) 120uL ice-cold MTBE was added, followed by another 3-min vortex mixing. Following a brief centrifugation (30 s, 100 x g, 4C), 200uL of water and 168uL of MTBE were added. The samples were vortex mixed for another 3 min, incubated at 4C for 10 min until centrifugation (20 min, 16,000 x g, 4C) inducing aqueous and organic layer separation. All solvents used during the LLE were ice-cold and vortex mixing was always at maximum speed. Following layer separation, each layer was transferred to an Eppendorf tube, followed by 5.0 and 2.5 min of centrifugation (16,000 x g, 4C) for aqueous and organic layers respectively. After extraction, 150uL of the aqueous layer was aliquoted for polar to semi-polar metabolites analysis, while 48.8 uL of aqueous and 28.8 uL of organic layer was combined for the bile and fatty acids analysis. The aliquots were dried in a Speedvac (Labcono, USA) and stored at -80C. Prior to LC-MS analysis, the extracts were reconstituted in 50 uL of 0.1% FA in water for polar to semi-polar metabolites analysis and 200uL of MeOH for the bile and fatty acids analysis. The reconstitution solvents contained different SILs. Quality Control Samples were randomized into 3 batches, with those from the same subject prepared and measured in the same batch. For the preparation of the quality control sample, 72 study samples were weighed (approx. 10 mg). Every two samples were pooled to give 36 samples which were extracted. After the extraction, equal volumes of each layer were taken from each sample and pooled, resulting in pooled QC aqueous and organic layers. Those pooled layers were used to prepare QC samples for each platform. The LLE and aliquoting steps were performed as described in Sample preparation.
Hosseinkhani F, Dubbelman AC, Karu N, Harms AC, Hankemeier T. Towards Standards for Human Fecal Sample Preparation in Targeted and Untargeted LC-HRMS Studies. Metabolites. 2021 Jun 7;11(6):364. doi:10.3390/metabo11060364. PMID:34200487.</p>
"],"organism":["blank","Homo sapiens"],"full_dataset_link":["https://www.ebi.ac.uk/metabolights/MTBLS12775"],"author":["Michael Van Vliet. Leiden University. m.s.vanvliet@lacdr.leidenuniv.nl.","Thomas Hankemeier. Leiden University. hankemeier@lacdr.leidenuniv.nl."],"data_transformation_protocol":["Targeted peak integration was performed using SCIEX OS (version 2.1.6., SCIEX) with a maximum mass error of 10 ppm. The retention times were verified against authentic standards. In case of coelution, the targets were reported using the name or abbreviation of one of the targets followed by a hashtag. Details on the abbreviations used are listed in Table S2. For the polar to semi-polar metabolites, peak area was used for further data analysis, whereas for the bile and fatty acids, the area ratio of compounds to stable isotopically labelled standards was used. Data quality inspection was performed using an in-house quality assurance software performing between batch correction and removal of metabolites with high technical variance (RSD of QC below 30%).
"],"study_factor":["Allergy","Formula feeding","Visit","Delivery mode","Complementary food","Breastfeeding","Age"],"submitter_email":["m.s.vanvliet@lacdr.leidenuniv.nl"],"sample_collection_protocol":["The samples for this work arise from a randomized, double-blind, controlled, parallel-group, multi-country study called TEMPO (clinicaltrials.gov identifier: NCT03067714). TEMPO enrolled healthy term infants (less than 16 weeks) at increased risk of developing allergy based on family history. Subjects who started formula-feeding before the age of 16 weeks were randomized to one of the two intervention arms, while the subjects exclusively breastfed for at least 16 weeks were included in the “breastfed reference” group. Exclusive breastfeeding was defined as receiving only breastmilk, with no other liquids or solids except water or formula in the first 72 hours of life, disregarding vitamins, minerals, or medicines. The participants were followed for a year, during which events of allergic manifestations were diagnosed by qualified physicians and classified as skin, food, or respiratory allergies. Allergy manifestations were considered IgE-mediated if either the skin prick test to any tested allergen or specific IgE blood test was positive at 12m. In this study, we selected a subset of 72 subjects from the breastfed reference group based on the availability of fecal samples collected before 16 weeks (baseline), at 6 months (6m), and at 12 months (12m) of age.
"],"omics_type":["Metabolomics"],"study_design":["Cesarean Section","Eczema","Infant","Feces","food allergy","targeted metabolites","Feeding"],"curator_keywords":["Cesarean Section","Eczema","Infant","Feces","food allergy","targeted metabolites","Feeding"],"mass_spectrometry_protocol":["LC-MS analysis of bile acid and fatty acids (Assay 1):Analysis of bile and fatty acids was performed on an UPLC-TOF/MS system consisting of ExionLC AC UHPLC system and SCIEX ZenoTOF 7600 system (Darmstadt, Germany) equipped with an IonDrive Turbo V Source, operated in negative ESI mode. The ion source conditions were as follows: spray voltage of 4.5 kV, capillary temperature of 550 °C, ion source gas 1 50 psi, ion source gas 2 50 psi, curtain gas 35 psi, CAD gas 7 psi. The MS data was acquired under full scan mode over the m/z range of 200-900 Da. Accumulation time was set to 0.25 s, delustering potential to -70 V and collision energy to -10 eV. Data acquisition was carried out on SCIEX OS 2.1.6.
LC-MS analysis of polar to semi polar metabolites (Assay 2 Samp; 3):Analysis of polar to semi-polar metabolites were performed with a Shimadzu Nexera X2 LC system coupled to a TripleTOF 6600 mass spectrometer (SCIEX, Foster City, CA, USA), as described previously. The data were acquired under full scan mode over the m/z range of 60-800 Da with Analyst TF software 1.7.1 (SCIEX) in negative and positive ionization modes. The ESI source parameters were set as follows: spray voltage ±4.5 kV, capillary temperature 400 °C, sheath gas 40, auxiliary gas 40, curtain gas 45. The preferred ionization mode for metabolites detectable in both polarities was chosen based on lower RSD% and higher signal-to-noise ratio of the QC samples.
"],"metabolite_name":["Ascorbic acid","Guanidineacetic acid","Riboflavin","Argininosuccinic Acid","Glycylproline","O-Acetylserine","4-Guanidinobutyric acid","Saccharopine","Kynurenic acid","Caffeine","Tyramine","3-Indoleacrylic acid","Myristoylcarnitine","Threonine","AgmatineÂ","Methionine","Urocanic acid","Lauroylcarnitine","Propionylcarnitine","Urea","1-Methylxanthine","4-Aminobenzoic acid","Theophylline","4-Hydroxyproline","TMAO","Hexanoylcarnitine","Glutamic acid","3-Methylglutaric acid","5-Methylcytidine","Carnosine","Butyrylcarnitine","Guanine","N-Acetylglutamine","Targinine","Quinaldic acid","Arginine","Indoleacetic acid","Kynurenine","8-Hydroxy-2-deoxyguanosine","N-Acetyl-L-Tyrosine","Glutathione","Alanine","Serotonin","Phenylalanine","N-Acetylputrescine","5-Aminovaleric acid","Beta-alanine","5-Hydroxylysine","Pyridoxine","Xanthine","N-Acetylneuraminic Acid","Decanoylcarnitine","trans-Ferulic acid","Trimethylamine","Deoxyguanosine","Valine","Choline","Tiglylcarnitine","Uracil","4-Pyridoxic acid","Valerylcarnitine","Tryptamine","5,6-Dihydrouracil","Guanosine","Nicotinic acid","Theobromine","Indolepropionic acid","Creatine","Xanthurenic acid","3-Guanidinopropanoate","Pipecolic acid","CreatinineÂ","Isobutyrylcarnitine","5-Aminolevulinic acid","Betaine","Octanoylcarnitine","3-Methylxanthine","Asymmetric dimethylarginine","Cytosine","Tyrosine","7-Methylxanthine","N2-Gamma-Glutamylglutamine","Homoarginine","N2,N2-Dimethylguanosine","N6,N6,N6-Trimethyl-lysine","Palmitoylcarnitine","Cytidine","Carnitine","Histamine","gamma-Glutamylalanine","Paraxanthine","Hypoxanthine","Symmetric dimethylarginine","Ornithine","Homoserine","Isovalerylcarnitine","Methionine sulfoxide","Proline","Phenylethylamine","Leucine","Putrescine","Alpha-N-Acetylarginine","2-Octenoylcarnitine","5-Methyluridine","Isoleucine","Ethanolamine","Cadaverine","Deoxycarnitine","Adenosine","Thymine","N-Acetyltryptophan","Adenine","Acetylcarnitine","Sarcosine","5-Hydroxy-L-tryptophan"],"additional_accession":[]},"is_claimable":false,"name":"Fecal metabolome alterations in infants at risk of developing allergy during the first year of life","description":"Background: Disturbances in the gut microbiome (GM) during the first year of life may contribute to allergy risk. This period is characterized by rapid microbial colonization, influenced by factors like delivery mode and infant feeding practices. This study investigates changes in key GM taxa and fecal metabolites in relation to allergy development, delivery mode, age and infant feeding practices during the first year of life.
Methods: In this study, 72 infants, exclusively breastfed for at least 16 weeks and at risk of developing allergies, were followed in their first year during which allergy manifestations were recorded and fecal samples were collected. The samples were subjected to metabolic profiling covering host and microbial metabolites and fluorescent in situ hybridization to quantify Bifidobacterium spp. and the Eubacterium rectale/Clostridium coccoides group.
Results: Strong age-associated metabolic shifts were observed, particularly in aromatic amino acid metabolites, bile acids, B vitamins and short and long-chain fatty acids. Introduction of complementary feeding and the cessation of breastfeeding were significantly associated with changes to the fecal metabolome. Delivery mode had a pronounced impact on the metabolome, with differences persisting until 6 months of age. Infants who developed an allergy (n=20) had lower Bifidobacterium spp. and higher long-chain fatty acid levels before age of 16 weeks.
Conclusion: This study offers valuable insights into the longitudinal development of the fecal metabolome during infancy. It highlights potential early biomarkers for allergy risk, which could inform future dietary strategies to support gut health and reduce the risk of developing allergy.
","dates":{"publication":"2026-06-09","submission":"2025-07-25"},"accession":"MTBLS12775","cross_references":{}}