Summary of Study ST003931
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR002462. The data can be accessed directly via it's Project DOI: 10.21228/M8D83H This work is supported by NIH grant, U2C- DK119886. See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST003931 |
| Study Title | Acylated putrescine therapeutic discovery for Inflammatory Bowel Diseases: HILIC-neg, C8-pos and C18-neg profiling of mouse fecal samples |
| Study Type | Study 2: HILIC-neg, C8-pos and C18-neg profiling of mouse fecal samples. |
| Study Summary | The data generated in this study using the HILIC-neg, C8-pos, and C18-neg profiling method were used to prioritize both disease-associated and microbe-associated metabolic features with potential bioactivity from untargeted metabolomics data. This mouse fecal microbiome data was used to develop a filter and prioritization tool: Metabolome Analysis and Combined Annotation Ranks to pRioritize Novel bioactives, that we call micro-MACARRoN. This workflow annotates the microbial association of gut metabolic features by comparing fecal metabolomics data among germ-free (GF), Altered Schaedler flora community (ASF), and specific pathogen free (SPF) mice. Application of this analysis pipeline to a publicly available dataset of 546 IBD fecal metabolomes from the Integrative Human Microbiome Project (HMP2/iHMP) cohort study enabled us to narrow our focus to metabolic features that are gut microbiome dependent. Using this tool we determined ~79% of gnotobiotic mouse-curated, IBD-associated HMP2 features are microbially associated (n=2,289 out of 2,883) indicating a substantial contribution of microbes to the gut metabolomes in IBD as would be expected. |
| Institute | Broad Institute of MIT and Harvard |
| Last Name | Avila-Pacheco |
| First Name | Julian |
| Address | 415 Main Street |
| jravilap@broadinstitute.org | |
| Phone | (617) 714-1729 |
| Submit Date | 2025-05-16 |
| Num Groups | 3 |
| Total Subjects | 21 |
| Num Males | 10 |
| Num Females | 11 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML, raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2025-10-05 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR002462 |
| Project DOI: | doi: 10.21228/M8D83H |
| Project Title: | Acylated putrescine therapeutic discovery for Inflammatory Bowel Diseases |
| Project Summary: | Gut microbial metabolites influence immune responses and play a key role in regulating host metabolism. Inflammatory bowel diseases (IBD) patients have altered gut microbial metabolic signatures driven by shifts in the microbiome. However, the microbial metabolites driving inflammation and remission and their immunomodulatory roles remain largely uncharacterized. We prioritized metabolic features from untargeted IBD fecal metabolomics and selected candidate features based on their microbial association for structural elucidation. We uncovered four acylated putrescines that are IBD-associated and microbially-dependent and investigated their bioactivity. One acylated putrescine, N-oleoylputrescine (NOP), has anti-inflammatory activity and suppressed type 1 immune responses. NOP ameliorated inflammation in several colitis models, modulating myeloid and lymphocyte cell populations and dampening proinflammatory responses, relevant for resolution of inflammation and maintenance of remission in IBD. We present an approach to identify clinically-relevant microbial metabolites from untargeted metabolomics useful for the discovery of bioactive molecules in IBD and other diseases involving host-microbial interactions. |
| Institute: | Broad Institute of MIT and Harvard |
| Last Name: | Avila-Pacheco |
| First Name: | Julian |
| Address: | 415 Main Street, Cambridge, MA 02142 USA |
| Email: | jravilap@broadinstitute.org |
| Phone: | +1 (617) 714-1729 |
| Contributors: | Sena Bae, Julian Avila-Pacheco, Slater L. Clay, Sunghee Bang, Monia Michaud, Diogo Fonseca-Pereira, Eunyoung Chun, Y. Grace Cao, Yancong Zhang, Amrisha Bhosle, Rosa M. Perez, Gleb Pishchany, Hera Vlamakis, Natalia Andreeva, Geniver El Tekle, Xochitl C. Morgan, Jonathan N. Glickman, Ramnik J. Xavier, Daniel B. Graham, Clary B. Clish, Jon Clardy, Eric A. Franzosa, Curtis Huttenhower, and Wendy S. Garrett |
Subject:
| Subject ID: | SU004067 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Genotype Strain: | C57BL/6 wild-type |
| Age Or Age Range: | 6-9 weeks |
| Gender: | Male and female |
| Animal Animal Supplier: | Jackson Laboratories |
Factors:
Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Sample source | Type | Sex |
|---|---|---|---|---|
| SA446280 | M13 | Fecal Samples | Altered Schaedler Flora (ASF) | Female |
| SA446281 | M15 | Fecal Samples | Altered Schaedler Flora (ASF) | Female |
| SA446282 | M14 | Fecal Samples | Altered Schaedler Flora (ASF) | Female |
| SA446283 | M12 | Fecal Samples | Altered Schaedler Flora (ASF) | Female |
| SA446284 | M8 | Fecal Samples | Altered Schaedler Flora (ASF) | Male |
| SA446285 | M9 | Fecal Samples | Altered Schaedler Flora (ASF) | Male |
| SA446286 | M10 | Fecal Samples | Altered Schaedler Flora (ASF) | Male |
| SA446287 | M11 | Fecal Samples | Altered Schaedler Flora (ASF) | Male |
| SA446288 | M6 | Fecal Samples | germ-free (GF) | Female |
| SA446289 | M7 | Fecal Samples | germ-free (GF) | Female |
| SA446290 | M4 | Fecal Samples | germ-free (GF) | Female |
| SA446291 | M5 | Fecal Samples | germ-free (GF) | Female |
| SA446292 | M2 | Fecal Samples | germ-free (GF) | Male |
| SA446293 | M3 | Fecal Samples | germ-free (GF) | Male |
| SA446294 | M1 | Fecal Samples | germ-free (GF) | Male |
| SA446295 | M19 | Fecal Samples | specified-pathogen free (SPF) | Female |
| SA446296 | M20 | Fecal Samples | specified-pathogen free (SPF) | Female |
| SA446297 | M21 | Fecal Samples | specified-pathogen free (SPF) | Female |
| SA446298 | M16 | Fecal Samples | specified-pathogen free (SPF) | Male |
| SA446299 | M17 | Fecal Samples | specified-pathogen free (SPF) | Male |
| SA446300 | M18 | Fecal Samples | specified-pathogen free (SPF) | Male |
| Showing results 1 to 21 of 21 |
Collection:
| Collection ID: | CO004060 |
| Collection Summary: | Stool pellets were collected from germ-free (GF), Altered Schaedler Flora (ASF) and specified-pathogen free (SPF) C57BL/6 mice (6-9wks), snap-frozen immediately, and stored at -80°C until untargeted metabolomics sample preparation |
| Sample Type: | Feces |
| Storage Conditions: | -80℃ |
Treatment:
| Treatment ID: | TR004076 |
| Treatment Summary: | No treatments were applied in this section of the study |
Sample Preparation:
| Sampleprep ID: | SP004073 |
| Sampleprep Summary: | Snap-frozen stool samples were thawed on ice, and then aqueous homogenates were generated by homogenizing samples in 10 volumes of water (mg: µL) using a TissueLyser II (QIAGEN) bead mill set to two 2 min intervals at 20Hz. The homogenate for each sample was divided into two 10 µL and two 30 µL aliquots in 1.5mL centrifuge tubes for LC-MS sample preparation. Metabolites were extracted from these homogenates for each method as follows; C8-pos: Bacterial cultures (10 μL) using 190 μL isopropanol containing 1-dodecanoyl-2-tridecanoyl-sn-glycero-3-phosphocholine as an internal standard (Avanti Polar Lipids; Alabaster, AL). After centrifugation (10 min, 9,000g, ambient temperature), supernatants (2 μL) were injected directly onto column. - C18-neg: Bacterial cultures (30 μL) were extracted using 90 μl methanol containing 15R-15-methyl Prostaglandin A2,15R-15-methyl Prostaglandin F2α, 15S-15-methyl Prostaglandin D2, 15S-15-methyl Prostaglandin E1, and 15S-15-methyl Prostaglandin E2 as internal standards (Cayman Chemical Co.) and centrifuged (10 min, 9,000g, 4°C). The supernatants (10 μL) were injected onto column. - HILIC-neg: Bacterial cultures (30 μL) were extracted with the addition of four volumes of 80% methanol containing inosine-15N4, thymine-d4 and glycocholate-d4 internal standards (Cambridge Isotope Laboratories). The samples were centrifuged (10 min, 9,000g, 4°C) and the supernatants 10 μL) were injected directly onto column. |
Combined analysis:
| Analysis ID | AN006454 | AN006455 | AN006456 |
|---|---|---|---|
| Chromatography ID | CH004900 | CH004901 | CH004902 |
| MS ID | MS006153 | MS006154 | MS006155 |
| Analysis type | MS | MS | MS |
| Chromatography type | Reversed phase | Reversed phase | HILIC |
| Chromatography system | Shimadzu Nexera X2 | Shimadzu Nexera X2 | Shimadzu Nexera X2 |
| Column | Waters ACQUITY UPLC BEH C18 (150 x 2.1mm, 1.7um) | Waters Acquity BEH C8 (100 x 2.1mm, 1.7um) | Phenomenex Luna NH2 (150 x 2mm, 5um) |
| MS Type | ESI | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap | Orbitrap |
| MS instrument name | Thermo Q Exactive Plus Orbitrap | Thermo Q Exactive Plus Orbitrap | Thermo Q Exactive Plus Orbitrap |
| Ion Mode | NEGATIVE | POSITIVE | NEGATIVE |
| Units | Abudances | Abudances | Abudances |
Chromatography:
| Chromatography ID: | CH004900 |
| Instrument Name: | Shimadzu Nexera X2 |
| Column Name: | Waters ACQUITY UPLC BEH C18 (150 x 2.1mm, 1.7um) |
| Column Temperature: | 45°C |
| Flow Gradient: | The column was eluted isocratically at a flow rate of 450 µL/min with 20% mobile phase A for 3 minutes followed by a linear gradient to 100% mobile phase B over 12 minutes. |
| Flow Rate: | 450 µL/min |
| Solvent A: | 100% Water; 0.01% Formic acid |
| Solvent B: | 100% Acetonitrile; 0.01% Acetic acid |
| Chromatography Type: | Reversed phase |
| Chromatography ID: | CH004901 |
| Instrument Name: | Shimadzu Nexera X2 |
| Column Name: | Waters Acquity BEH C8 (100 x 2.1mm, 1.7um) |
| Column Temperature: | 40°C |
| Flow Gradient: | The column was eluted at a flow rate of 450 µL/min isocratically for 1 minute at 80% mobile phase A, followed by a linear gradient to 80% mobile-phase B over 2 minutes, a linear gradient to 100% mobile phase B over 7 minutes, and then 3 minutes at 100% mobile-phase B. |
| Flow Rate: | 450 µL/min |
| Solvent A: | 95% Water/5% Methanol; 10 mM Ammonium acetate; 0.1% Acetic acid |
| Solvent B: | 100% Methanol; 0.1% Acetic acid |
| Chromatography Type: | Reversed phase |
| Chromatography ID: | CH004902 |
| Instrument Name: | Shimadzu Nexera X2 |
| Column Name: | Phenomenex Luna NH2 (150 x 2mm, 5um) |
| Column Temperature: | 30°C |
| Flow Gradient: | The column was eluted with initial conditions of 10% mobile phase A and 90% mobile phase B followed by a 10 min linear gradient to 100% mobile phase A. |
| Flow Rate: | 400 µL/min |
| Solvent A: | 100% water; 20 mM ammonium acetate; 20 mM ammonium hydroxide |
| Solvent B: | 75% acetonitrile/25% methanol; 10 mM ammonium hydroxide |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS006153 |
| Analysis ID: | AN006454 |
| Instrument Name: | Thermo Q Exactive Plus Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Raw data were processed using TraceFinder 3.3 software (Thermo Fisher Scientific; Waltham, MA) and Progenesis QI (Nonlinear Dynamics; Newcastle upon Tyne, UK). Metabolite identities were confirmed using authentic reference standards or reference samples. |
| Ion Mode: | NEGATIVE |
| MS ID: | MS006154 |
| Analysis ID: | AN006455 |
| Instrument Name: | Thermo Q Exactive Plus Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Raw data were processed using TraceFinder 3.3 software (Thermo Fisher Scientific; Waltham, MA) and Progenesis QI (Nonlinear Dynamics; Newcastle upon Tyne, UK). Metabolite identities were confirmed using authentic reference standards or reference samples. |
| Ion Mode: | POSITIVE |
| MS ID: | MS006155 |
| Analysis ID: | AN006456 |
| Instrument Name: | Thermo Q Exactive Plus Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Raw data were processed using TraceFinder 3.3 software (Thermo Fisher Scientific; Waltham, MA) and Progenesis QI (Nonlinear Dynamics; Newcastle upon Tyne, UK). Metabolite identities were confirmed using authentic reference standards or reference samples. |
| Ion Mode: | NEGATIVE |
