For bile acid targeted metabolomics, identification and quantification were performed using an in-house library of 65 authentic standards, with each compound confirmed by matching both retention time and optimized MRM transitions (precursor-to-fragment ion pairs) on a SCIEX QTRAP 6500+ system controlled by Analyst 1.6.3. Peak integration and absolute quantification (expressed as ng/g tissue) were carried out in MultiQuant 3.0.3 using internal standard correction and external calibration curves (0.1-1000 ng/mL), and the resulting concentration matrix was normalized by unit variance (Z-score) scaling before statistical analysis. Multivariate explorations including PCA and OPLS-DA, along with VIP extraction and 200-fold permutation validation, were executed with the MetaboAnalystR package (version 1.0.1) in R; differential metabolites were screened by VIP > 1, P < 0.05 (t-test or ANOVA), and fold change >= 2 or <= 0.5. Functional annotation and pathway enrichment were performed by mapping the identified bile acids to the KEGG Compound and Pathway databases, with enrichment significance assessed via hypergeometric tests. Throughout the analytical workflow, pooled QC samples were injected every 10 runs to monitor retention time stability and signal reproducibility, ensuring that >80% of detected metabolites exhibited a coefficient of variation (CV) below 0.3.
"],"repository":["MetaboLights"],"study_status":["Public"],"ptm_modification":[""],"instrument_platform":["Liquid Chromatography MS - negative - reverse-phase"],"chromatography_protocol":["Analysis was performed on a Sciex ExionLC™ AD UHPLC system coupled to an Applied Biosystems 6500 QTRAP mass spectrometer. Chromatographic separation was achieved on a Waters ACQUITY UPLC HSS T3 C18 column (100 mm × 2.1 mm, 1.8 µm) using gradient elution with mobile phase A (ultrapure water + 0.01% acetic acid + 5 mmol/L ammonium acetate) and mobile phase B (acetonitrile + 0.01% acetic acid). Chromatographic conditions: flow rate 0.35 mL/min, column temperature 40 °C, injection volume 3 μL.
"],"publication":["Dietary supplementation with three probiotics ameliorates high-carbohydrate-high-fat diet-induced metabolic disorders and intestinal barrier impairment in common carp via remodeling gut microbiota."],"submitter_affiliation":["Henan normal university"],"submitter_name":["Peng Pang"],"determination_of_bile_acid_concentrations_in_hepatopancreatic_tissue_protocol":["Ball-milled tissue samples (20 mg) were extracted with 200 μL methanol-acetonitrile (2:8, v/v) containing 10 μL mixed internal standard (1 μg/mL) for quantitative analysis. After protein precipitation at −20 °C for 10 min, samples were centrifuged at 12,000 rpm and 4 °C for 10 min. The supernatant was vacuum-dried and reconstituted in 100 μL 50% methanol for LC-ESI-MS/MS analysis.
Analysis was performed on a Sciex ExionLC™ AD UHPLC system coupled to an Applied Biosystems 6500 QTRAP mass spectrometer. Chromatographic separation was achieved on a Waters ACQUITY UPLC HSS T3 C18 column (100 mm × 2.1 mm, 1.8 µm) using gradient elution with mobile phase A (ultrapure water + 0.01% acetic acid + 5 mmol/L ammonium acetate) and mobile phase B (acetonitrile + 0.01% acetic acid). Chromatographic conditions: flow rate 0.35 mL/min, column temperature 40 °C, injection volume 3 μL. Mass spectrometry was operated in negative ESI mode with scheduled multiple reaction monitoring (MRM) detection. Source parameters: ion source temperature 550 °C, ion spray voltage −4500 V, curtain gas 35 psi. Declustering potential (DP) and collision energy (CE) were optimized for each MRM transition.
Data were acquired with Analyst 1.6.3 software and quantified using MultiQuant 3.0.3 software. Bile acid profiles were analyzed by PCA and OPLS-DA, and differentially abundant metabolites were identified based on OPLS-DA VIP values and univariate P values.
"],"organism_part":["hepatopancreas"],"technology_type":["mass spectrometry assay"],"disease":[""],"extraction_protocol":["Samples were mixed with beads and an extraction mixture containing deuterated internal standards. The mixture was vortexed for 30 s, incubated at −40 °C for 1 h for protein precipitation, and centrifuged at 12,000 rpm for 15 min at 4 °C. The supernatant was transferred to a new glass vial for analysis. A quality control (QC) sample was prepared by pooling equal aliquots of all experimental supernatants to ensure analytical reliability.
"],"organism":["Cyprinus carpio"],"full_dataset_link":["https://www.ebi.ac.uk/metabolights/MTBLS15295"],"author":["Peng Pang. Henan Normal University. pangp1998@163.com."],"data_transformation_protocol":["For targeted bile acid analysis: Raw data were acquired on a SCIEX QTRAP 6500+ system using Analyst 1.6.3. Peak integration and absolute quantification were performed with MultiQuant 3.0.3 using internal standard correction and external calibration curves (0.1-1000 ng/mL). The concentration matrix was normalized by unit variance scaling (Z-score) before statistical analysis. Multivariate analyses including PCA and OPLS-DA, VIP extraction, and 200-fold permutation validation were executed with the MetaboAnalystR package (version 1.0.1) in R. Differential metabolites were screened by VIP > 1, P < 0.05 (t-test or ANOVA), and fold change >= 2 or <= 0.5.
For untargeted metabolomics of intestinal contents: Raw data from the Orbitrap Exploris 120 were converted to mzXML format using ProteoWizard. Peak detection, alignment, and integration were performed with an in-house R script based on XCMS. Metabolite identification was achieved using the BiotreeDB (V3.0) database. The same R packages (MetaboAnalystR, etc.) were used for statistical analyses. KEGG pathway enrichment was performed via hypergeometric tests.
All QC samples were injected every 10 runs to monitor stability. CV values were calculated; only metabolites with >80% of QC samples having CV < 0.3 were retained.
"],"study_factor":["Probiotics"],"submitter_email":["pangp1998@163.com"],"sample_collection_protocol":["The hepatopancreas was collected for the analysis of bile acid concentrations. The sample was stored at −80 °C for later analysis.
"],"omics_type":["Metabolomics"],"study_design":["Gut microbiota","Applied Biosystems 6500 QTRAP","Metabolomics","Common carp","Probiotics","molecular docking","targeted analysis","hepatopancreas","Sciex ExionLC™ AD UHPLC","Cyprinus carpio","glycolipid metabolism","experimental sample"],"curator_keywords":["Gut microbiota","Applied Biosystems 6500 QTRAP","Metabolomics","Common carp","Probiotics","molecular docking","targeted analysis","Sciex ExionLC™ AD UHPLC","hepatopancreas","Cyprinus carpio","glycolipid metabolism","experimental sample"],"mass_spectrometry_protocol":["Instrument (UHPLC): ExionLC™ AD system (SCIEX, Framingham, MA, USA)
Instrument (MS/MS): QTRAP® 6500+ triple quadrupole-linear ion trap mass spectrometer (SCIEX, Framingham, MA, USA)
Ion Source: Electrospray Ionization (ESI) Turbo Ion-Spray interface
Ionization Mode: Negative ion mode
m/z Range: Not explicitly stated in the report; analysis was performed using scheduled Multiple Reaction Monitoring (MRM) with optimized transitions for each bile acid. Full MS scanning was not performed.
Chromatographic Column: Waters ACQUITY UPLC HSS T3 C18 column (100 mm × 2.1 mm i.d., 1.8 µm particle size)
Column Temperature: 40 °C
Mobile Phase A: Ultrapure water with 0.01% acetic acid and 5 mmol/L ammonium acetate
Mobile Phase B: Acetonitrile with 0.01% acetic acid
Flow Rate: 0.35 mL/min
Injection Volume: 3 µL
Gradient Program:
0 min: 95% A / 5% B
0.5 min: 60% A / 40% B
4.5 min: 50% A / 50% B
7.5 min: 25% A / 75% B
10 min: 5% A / 95% B
12.0 min: 95% A / 5% B (re-equilibration)
Ion Source Temperature: 550 °C
Ion Spray Voltage: −4500 V
Curtain Gas (CUR): 35 psi
Declustering Potential (DP) and Collision Energy (CE): Optimized individually for each MRM transition (specific values not listed for individual compounds in the report)
Scan Type: Scheduled Multiple Reaction Monitoring (MRM)
Software for Data Acquisition: Analyst 1.6.3 (SCIEX)
Software for Quantification: MultiQuant 3.0.3 (SCIEX)
"],"additional_accession":[]},"is_claimable":false,"name":"Bile Acid Metabolome in Hepatopancreas Tissue of Common Carp","description":"This study investigated the regulatory effects of three probiotics (P. pentosaceus, B. coagulans, and E. faecalis) on bile acid metabolism in common carp (Cyprinus carpio) fed a high-carbohydrate-high-fat (HCHF) diet for 56 days. A targeted metabolomics approach using UHPLC-QTRAP-MS/MS with scheduled multiple reaction monitoring (MRM) was employed to quantify 10 bile acid components in hepatopancreatic tissues, including free bile acids (CDCA, CA, DCA, LCA, alpha-MCA) and their taurine-conjugated forms (TCDCA, TCA, TDCA, TLCA, Talpha-MCA, Tbeta-MCA, Tomega-MCA). The analysis aimed to elucidate whether probiotic supplementation alleviates HCHF-induced glycolipid metabolic disorders through modulating bile acid profiles and FXR signaling pathway. Furthermore, molecular docking was performed to predict the binding affinities of these bile acids to common carp FXR protein, with CDCA showing the strongest binding energy of -6.859 kcal/mol. The results revealed that all three probiotics significantly increased the contents of FXR-activating free bile acids (CDCA, CA, DCA, LCA) while decreasing conjugated bile acid levels, thereby inhibiting gluconeogenesis and lipogenesis through the FXR-SHP-CYP7A1/CYP8B1 and FXR-SHP-HNF4 alpha/FOXO1 pathways.
","dates":{"publication":"2026-08-09","submission":"2026-08-08"},"accession":"MTBLS15295","cross_references":{}}