Metabolites were identified by the LIPID MAPS database (https://www.lipidmaps.org/), MS/MS fragments and authentic standard.
For MS/MS identification: AUTO files correspond to ddMS2 data acquired of sample; TARGET files correspond to target fragmentation of the ions at 20 eV of collision energy; TARGET30 files correspond to target fragmentation of the ion at 30 eV of collision energy; TARGET40 files correspond to target fragmentation of the ion at 40 eV of collision energy.
"],"repository":["MetaboLights"],"study_status":["Public"],"ptm_modification":[""],"instrument_platform":["Liquid Chromatography MS - negative - reverse phase"],"chromatography_protocol":["Chromatographic analysis was mostly replicated from literature[1] on a UHPLC (Infinity 1290; Agilent, Santa Clara, CA, USA) equipped with 100 × 2.1 mm, 100 Å, 1.7 μm, Kinetex C18 column (Phenomenex, Aschaffenburg, Germany) and performed at a constant flow rate of 0.4mL/min. Mobile phases consisted of water (A) and acetonitrile:water (95:5, v/v) (B), both containing 5 mM ammonium acetate and 0.1% formic acid. The gradient started with 25% B for 2.0 min, increased in 1.5 min to 27% B, in 2.0 min to 35% B, held for 4.5 min, increased in 1.0 min to 43% B, held for 2.0 min, increased in 2.0 min to 58% B, held for 3.0 min, increased in 0.5 to 65% B, in 0.5 min to 80% B, in 1.0 min to 100% B, held 1.0 min at 100% B, and decreased to the initial conditions followed by 2.0 min of re-equilibration. The injection volume for all samples was 3 µL, and the column oven temperature was set to 40 °C.
Ref:
[1] Reiter S, Dunkel A, Metwaly A, Panes J, Salas A, Haller D, Hofmann T. Development of a Highly Sensitive Ultra-High-Performance Liquid Chromatography Coupled to Electrospray Ionization Tandem Mass Spectrometry Quantitation Method for Fecal Bile Acids and Application on Crohn's Disease Studies. J Agric Food Chem. 2021 May 5;69(17):5238-5251. doi:10.1021/acs.jafc.1c00769. Epub 2021 Apr 23. PMID:33890469.
"],"publication":["Production of New Microbially Conjugated Bile Acids by Human Gut Microbiota. 10.3390/biom12050687. PMID:35625615"],"submitter_affiliation":["CEBAS-CSIC"],"submitter_name":["Carlos GarcÃa Hernández-Gil"],"organism_part":["feces"],"technology_type":["mass spectrometry"],"disease":[""],"extraction_protocol":["Conversion Experiments by Human Fecal Cultures
Preparation of fecal suspensions and subsequent fermentation experiments were performed under anoxic conditions in an anaerobic chamber (Concept 400, Baker Ruskinn Technologies, Ltd., Bridgend, South Wales, UK) with an atmosphere consisting of N2:H2:CO2 (85:5:10) at 37 °C. Aliquots of stool samples (10 g) were diluted 1/10 w/v in Nutrients Broth supplemented with 0.05% L-cysteine hydrochloride and homogenized by a stomacher in filter bags. Aliquots of fecal suspensions (50 µL) were inoculated into 5 mL of fermentation medium anaerobe Wilkins Chaldean containing either 50 µm of chenodeoxycholic acid (CDCA) or 50 µm 3-oxo-chenodeoxycholic acid (3-oxo-CDCA). A total of 3 replicate cultures were prepared in parallel from each fecal suspension. And 2 types of fecal control samples were prepared, one then including incubation and without incubation procedure. Fecal sample controls and either CDCA and 3-oxo-CDCA (5 mL) were incubated with fecal microbiota and samples were collected after 5 days of incubation at 37 °C and extracted with 5 mL of ethyl acetate LC-MS (Scharlau, Barcelona, Spain) using a refrigerated thermoblock shaker (VWR lnternational, LLC, Radnor, PA, USA) at 20 °C for 10 min at 1500 rpm. Samples were centrifuged at 3500 x g for 10 min at 4 °C. The organic phase was evaporated under reduced pressure in a speed vacuum concentrator (Savant SPD121P, Thermo Scientific, Waltham, MA, USA). All of these procedures were performed at CEBAS-CSIC (Murcia, Spain). Extracts were then transferred to the Department of Food Analysis and Nutrition (University of Chemistry and Technology, Prague) for instrument analysis. Samples were re-dissolved in 500 µL of methanol, and filtered through a 0.22 µm PVDF filter (Merck Millipore, Cork, Ireland), and they were diluted 1:2 in methanol before the UPLC-IM-QTOF-MS analysis.
"],"organism":["Homo sapiens"],"full_dataset_link":["https://www.ebi.ac.uk/metabolights/MTBLS4140"],"author":["Carlos Garcia. Department of food analysis and nutrition. Faculty of Food and Biochemical Technology. University of Chemistry and Technology, 16628 Prague, Czech Republic.. hernandc@vscht.cz."],"data_transformation_protocol":["The data pre-processing was performed to all samples using Profinder software (Agilent technologies) in negative polarity mode. Log tranformation for the data matrix was performed. The pre-processing operation gave a data matrix based on 10806 entities from full data set. Data matrix was imported to Mass Professional Profiler (MPP, Agilent technologies) for processing. A specific database was created, according to generic file format, for all possible MCBA combinations. The database was built using PDCL software (Agilent Technologies) to allow its use in the Agilent software package environment. ANOVA (corrected P-value cut-off: 0.05; p-value computation: Asymptotic; Multiple Testing Correction: Benjamini-Hochberg) statistics analysis and Fold change tools were used for filtering entity candidates which were not present in the control and were detected in the samples after the incubation with BAs.
"],"submitter_email":["cjgarcia@cebas.csic.es"],"sample_collection_protocol":["Chemicals
Authentic standards of 3α,7α-Dihydroxy-5β-cholan-24-oic Acid (chenodeoxycholic acid) and 3α-Hydroxy-11-oxo-5β-cholan-24-oic Acid (3-oxo-chenodeoxycholic acid) were purchased from Avanti Polar Lipids (Alabaster, Alabama, USA), Nutrient Broth (NB) from Oxoid (Basingstoke, Hampshire, UK) and L-cysteine hydrochloride (Panreac Química, Barcelona, Spain).
Collection of human fecal samples
In total, 2 healthy donors of stool samples, one male (age 44) and one female (age 30), were recruited at the Centro de Edafología y Biología Aplicada del Segura (CEBAS-CSIC, Murcia, Spain) and gave written informed consent. Each volunteer provided a fresh stool sample early in the morning to perform the fermentation experiments. Samples were stored at 4 °C and further processed within 1 h of donation.
"],"omics_type":["Metabolomics"],"study_design":["ultra-performance liquid chromatography-mass spectrometry","7alpha-hydroxy-3-oxo-5beta-cholan-24-oic acid","tandem mass spectrometry","bile acid","untargeted metabolites","microbiome","chenodeoxycholic acid"],"curator_keywords":["ultra-performance liquid chromatography-mass spectrometry","7alpha-hydroxy-3-oxo-5beta-cholan-24-oic acid","tandem mass spectrometry","bile acid","untargeted metabolites","microbiome","chenodeoxycholic acid"],"mass_spectrometry_protocol":["Metabolomics analysis were performed on a UHPLC (Infinity 1290; Agilent, Santa Clara, CA, USA) coupled to a high-resolution mass spectrometer with a hyphenated quadrupole time-of-flight mass analyzer (6560 Ion Mobility Q-TOF LC/MS; Agilent) with an Agilent Jet Stream (AJS) electrospray (ESI) source. The mass analyzer was operated in negative mode following the conditions: gas temperature, 180 °C; drying gas, 12 L/min; nebulizer pressure, 45 psig; sheath gas temperature, 350 °C; sheath gas flow, 11 L/min; capillary voltage, 3500 V; nozzle voltage, 250 V; fragmentor voltage, 350 V; octapole radiofrequency voltage, 250 V. Data were acquired over the mass range of 50-1700 m/z at the rate of 2 spectra/s. The m/z range was autocorrected on reference masses 119.0363 and 980.0164 m/z.
"],"metabolite_name":["3-oxo-5beta-cholan-24-oic-acid","Chenodeoxycholic acid (CDCA)","3-oxo-chenodeoxycholic acid (3-oxoCDCA)","Isoursochenodexycholic acid (iUDCA)","Lithocholic acid (LCA)","7-oxo-lithocholic acid (7-oxoLCA)","7alpha-Hydroxy-5beta-cholan-24-oic acid"],"pubmed_abstract":["Gut microbes have been recognized to convert human bile acids by deconjugation, dehydroxylation, dehydrogenation, and epimerization of the cholesterol core, but the ability to re-conjugate them with amino acids as an additional conversion has been recently described. These new bile acids are known as microbially conjugated bile acids (MCBAs). The aim of this study was to evaluate the MCBAs diversity produced by the gut microbiota through a metabolomics approach. In this study, fresh fecal samples from healthy donors were evaluated to explore the re-conjugation of chenodeoxycholic and 3-oxo-chenodeoxycholic acids by the human gut microbiota. No significant differences were found between the conversion trend of both BAs incubations. The in vitro results showed a clear trend to first accumulate the epimer isoursochenodeoxycholic acid and the dehydroxylated lithocholic acid derivatives in samples incubated with chenodeoxycholic and 3-oxo-chenodeoxycholic acid. They also showed a strong trend for the production of microbially conjugated dehydroxylated bile acids instead of chenodeoxycholic backbone conjugates. Different molecules and isomers of MCBAs were identified, and the new ones, valolithocholate ester and leucolithocholate ester, were identified and confirmed by MS/MS. These results document the gut microbiota's capability to produce esters of MCBAs on hydroxyls of the sterol backbone in addition to amides at the C24 acyl site. This study opens a new perspective to study the BAs diversity produced by the human gut microbiota."],"pubmed_title":["Production of New Microbially Conjugated Bile Acids by Human Gut Microbiota."],"pubmed_authors":["Garcia Carlos J CJ, Kosek Vit V, Beltrán David D, Tomás-Barberán Francisco A FA, Hajslova Jana J"],"description_synonyms":["Backbone, Salts, Bile Acids, Gallodesoxycholic Acid, Bowel, DmNa[[v]]1, HSN1E, spinal column, Quenocol, Aminosaeure, Bile Salt, Metabonomic, columna vertebralis, Amino acid, bas, Metabonomics, neutral molecular compounds, L-Leucine, L-Isomer Leucine, Bile salt, vertebral region, 17alpha)-cholest-5-en-3-ol, 3, anon-23Da, Leucin, 14beta, vertebral column skeleton, molecule, Chenique Acid, Bile, molecula, strong, bss, (3beta, amino acids, molecules, Man (Taxonomy), Spinal column, L, Gallensaeuren, Microbial Community Structure, Salt, Tissue, alimentary tract, Cholest-5-en-3beta-ol, Molekuel, backbone, Epicholesterol, DL-valine, g, Microbial Community Structures, asd, Sodium Chenodeoxycholate, Tissue Donor, Gallodesoxycholic, adult alimentary canal, alimentary system, L Valine, CXXC finger protein 9, L-Valine, Bile Acid, amides, Spine, Transplant Donor, Chenophalk, Aminokarbonsaeure, Modern, Transplant, acid, Ester, Bile Salts, Semen Donors, HYDR2, Microbial, 2-amino-3-methylbutanoic acid, alpha-amino carboxylic acids, results, par, 3-hydroxy-, Chenique, dorsal spine, Donors, l(1)14Da, 5beta-bile acid, Henohol, Microbial Communities, 7-dihydroxy-, region, 5beta, Acid, Amino Acid, ADCADN, Chenic Acid, esters, AI747421, Amino acids, UNQ203/PRO229, human gut microbiota, l(1)ESHS48, Microbial Community Compositions, MS/MS, acide, DNA (cytosine-5-)-methyltransferase 1, acids, Leuzin, Bile acid, acido, Chenodiol, human, Semen Donor, Composition, sbl-1, Semen, hyaline, gastrointestinal system, spine, Ocd, Acids, Hleu, Ovum, Ovum Donors, Organ Donors, Human Microbiomes, Community, Cholan-24-oic acid, postcranial axial skeleton, Chenofalk, digestive canal, Microbial Community Composition, (RS)-Leucine, human being, olfD, axial skeleton, Valin, Aminocarbonsaeure, L Isomer, bile acids, alpha-amino acid, Amide, Human, 5beta)-, Gus, Aim, Gur, AIM, Gut, Homo sapiens, resilient, Cholest-5-en-3-ol (3beta)-, tough, cg3488, Metabolomic, Man, DMU29170, DNMT1, study, gut, DNMT1_HUMAN, CT11757, MS2, Chenodeoxycholic, Quenobilan, (+-)-Leucine, Gallensaeure, Transplant Donors, API6, DmelCG3488, anon-23D, DmelCG9907, site, Ovum Donor, OXO GROUP, Saeure, intestines, Human Microbiome, DNA MTase HsaI, Organ, Sterol, DNA (cytosine-5)-methyltransferase 1, Community Composition, Chenodeoxycholate, DNMT, derivatives, alpha-amino acids, MCMT, DmNav1, L-Isomer, Isolithocholic, DmNa[[v]], Chenix, gut tube, oxo, 7alpha)-, analogs and derivatives, DNA methyltransferase HsaI, core, =O, Donor, Sodium, Chenic, Cholesterin, (3alpha, DmNa[[V]], tandem MS, Microbiome, CXXC9, Lithocholate, C24, Microbiomes, CT-2, Organ Donor, Leucine, Microbial Community, Saeuren, CG3488, Gus-u, Community Structure, Gus-t, Gus-s, Gus-r, Amino, CXXC-type zinc finger protein 9, CG9907, c24, clear, Isolithocholic Acid, Lithocholic, 5beta-bile acids, Modern Man, DL-Leucine, 2-amino-4-methylpentanoic acid, Microbiotas, human gut microbiota., Hval, CLEC2C, sbl, vertebral column, Leu, m.HsaI, valina"],"name_synonyms":["Bile Acid, Acid, Salts, Bile Acids, Bile salt, 5beta-bile acids, Gallensaeuren, Salt, Bile Salts, Bile Salt, Acids, bile acids, human gut microbiota., Bile acid, Gallensaeure, 5beta-bile acid, Bile"],"pubmed_title_synonyms":["Bile Acid, Acid, Salts, Bile Acids, Bile salt, 5beta-bile acids, Gallensaeuren, Salt, Bile Salts, Bile Salt, Acids, bile acids, human gut microbiota., Bile acid, Gallensaeure, 5beta-bile acid, Bile"],"pubmed_abstract_synonyms":["Backbone, Salts, Bile Acids, Gallodesoxycholic Acid, Bowel, DmNa[[v]]1, HSN1E, spinal column, Quenocol, Aminosaeure, Bile Salt, Metabonomic, columna vertebralis, Amino acid, bas, Metabonomics, neutral molecular compounds, Bile salt, vertebral region, 17alpha)-cholest-5-en-3-ol, 3, anon-23Da, 14beta, vertebral column skeleton, molecule, Chenique Acid, Bile, molecula, strong, bss, (3beta, amino acids, molecules, Man (Taxonomy), Spinal column, Gallensaeuren, Microbial Community Structure, Salt, Tissue, alimentary tract, Cholest-5-en-3beta-ol, Molekuel, backbone, Epicholesterol, g, Microbial Community Structures, asd, Sodium Chenodeoxycholate, Tissue Donor, Gallodesoxycholic, adult alimentary canal, alimentary system, CXXC finger protein 9, Bile Acid, amides, Spine, Transplant Donor, Chenophalk, Aminokarbonsaeure, Modern, Transplant, acid, Ester, Bile Salts, Semen Donors, HYDR2, Microbial, alpha-amino carboxylic acids, results, par, 3-hydroxy-, Chenique, dorsal spine, Donors, l(1)14Da, 5beta-bile acid, Henohol, Microbial Communities, 7-dihydroxy-, region, 5beta, Acid, Amino Acid, ADCADN, Chenic Acid, esters, AI747421, Amino acids, UNQ203/PRO229, human gut microbiota, l(1)ESHS48, Microbial Community Compositions, MS/MS, acide, DNA (cytosine-5-)-methyltransferase 1, acids, Bile acid, acido, Chenodiol, human, Semen Donor, Composition, sbl-1, Semen, hyaline, gastrointestinal system, spine, Ocd, Acids, Ovum, Ovum Donors, Organ Donors, Human Microbiomes, Community, Cholan-24-oic acid, postcranial axial skeleton, Chenofalk, digestive canal, Microbial Community Composition, human being, olfD, axial skeleton, Aminocarbonsaeure, bile acids, alpha-amino acid, Amide, Human, 5beta)-, Gus, Aim, Gur, AIM, Gut, Homo sapiens, resilient, Cholest-5-en-3-ol (3beta)-, tough, cg3488, Metabolomic, Man, DMU29170, DNMT1, study, gut, DNMT1_HUMAN, CT11757, MS2, Chenodeoxycholic, Quenobilan, Gallensaeure, Transplant Donors, API6, DmelCG3488, anon-23D, DmelCG9907, site, Ovum Donor, OXO GROUP, Saeure, intestines, Human Microbiome, DNA MTase HsaI, Organ, Sterol, DNA (cytosine-5)-methyltransferase 1, Community Composition, Chenodeoxycholate, DNMT, derivatives, alpha-amino acids, MCMT, DmNav1, Isolithocholic, DmNa[[v]], Chenix, gut tube, oxo, 7alpha)-, analogs and derivatives, DNA methyltransferase HsaI, core, =O, Donor, Sodium, Chenic, Cholesterin, (3alpha, DmNa[[V]], tandem MS, Microbiome, CXXC9, Lithocholate, C24, Microbiomes, CT-2, Organ Donor, Microbial Community, Saeuren, CG3488, Gus-u, Community Structure, Gus-t, Gus-s, Gus-r, Amino, CXXC-type zinc finger protein 9, CG9907, c24, clear, Isolithocholic Acid, Lithocholic, 5beta-bile acids, Modern Man, Microbiotas, human gut microbiota., CLEC2C, sbl, vertebral column, m.HsaI"],"additional_accession":[]},"is_claimable":false,"name":"Production of new microbially conjugated bile acids by human gut microbiota","description":"Gut microbes have been recognized to convert the human bile acids by deconjugation, dehydroxylation, dehydrogenation and epimerization of the cholesterol core, but the ability to re-conjugate them with amino acids as an additional conversion has only been recently described. These new bile acids are known as microbially conjugated bile acids (MCBAs). The aim of this study was evaluating the MCBAs diversity produced by the gut microbiota through a metabolomics approach. In this study, fresh fecal samples from healthy donors were evaluated to explore the re-conjugation of chenodeoxycholic and 3-oxo-chenodeoxycholic acids by the human gut microbiota. No significant differences were found between the conversion trend of both BAs incubations. The in vitro results showed a clear trend to first accumulate the epimer isoursochenodeoxycholic acid, and the dehydroxylated lithocholic acid derivatives in samples incubated with chenodeoxycholic and 3-oxo-chenodeoxycholic acid. Also showed a strong trend for the production of microbially conjugated dehydroxylated bile acids, instead of chenodeoxycholic backbone conjugates. Different molecules and isomers of microbial conjugates with valine and leucine were also identified and confirmed by MS/MS. These results document the gut microbiota capability to produce esters of MCBAs on hydroxyls of the sterol backbone in addition to amides at the C24 acyl site. This study opens a new perspective to study the BAs diversity produced by the human gut microbiota.
","dates":{"publication":"2024-01-16","submission":"2022-01-17"},"accession":"MTBLS4140","cross_references":{"MetaboLights":["MTBLC16755","MTBLC43419","MTBLC88097","MTBLC82679","MTBLC16325","MTBLC73676","MTBLC17639"],"pubmed":["35625615"],"ChEBI":["CHEBI:16755","CHEBI:43419","CHEBI:88097","CHEBI:82679","CHEBI:16325","CHEBI:73676","CHEBI:17639"]}}