Summary of Study ST003131

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 PR001945. The data can be accessed directly via it's Project DOI: 10.21228/M89M7J This work is supported by NIH grant, U2C- DK119886.

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This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study IDST003131
Study TitleUntargeted Metabolomics on mouse caecal contents
Study TypeMouse
Study SummaryWhile there is strong evidence for interactions between the microbiota-gut-brain axis and host physiology in the context of chronic stress, limited research has investigated the role of the microbiome in host response to acute stress. Determining the underlying mechanisms by which stress-induced microbiota changes may provoke functional changes in the gut and brain is critical for developing future therapeutics to alleviate the adverse consequences of traumatic stress. Here, we aimed to identify a biological signature of gut metabolites that are significantly altered following exposure to acute restraint stress. Adult male C57Bl/6 conventional, germ-free and colonized germ-free mice underwent a 15-minute restraint stress exposure. Caecal contents were collected from naïve mice and stressed mice, either immediately or 45 minutes following stress. Caecal contents underwent untargeted metabolomics analysis.
Institute
University College Cork
DepartmentPsychiatry
LaboratoryMicrobiota-Gut-Brain Axis Group
Last NameClarke
First NameGerard
AddressGaol Walk, Cork
EmailG.Clarke@ucc.ie
Phone+353-21-4901408
Submit Date2024-03-19
Num Groups9
Total Subjects63
Num Males63
Analysis Type DetailOther
Release Date2024-03-22
Release Version1
Gerard Clarke Gerard Clarke
https://dx.doi.org/10.21228/M89M7J
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:

Project:

Project ID:PR001945
Project DOI:doi: 10.21228/M89M7J
Project Title:The microbiota drives diurnal rhythms in tryptophan metabolism in the stressed gut
Project Type:Untargeted metabolomics by Metabolon
Project Summary:Adult male C57Bl/6 conventional, germ-free and colonized germ-free mice underwent a 15-minute restraint stress exposure. Caecal contents and colonic mucosal scrapings were collected from naïve mice and stressed mice, either immediately or 45 minutes following stress. Caecal contents and colonic mucosal scrapings underwent untargeted metabolomics analysis.
Institute:University College Cork
Department:Psychiatry
Laboratory:Microbiota-Gut-Brain Axis Group
Last Name:Clarke
First Name:Gerard
Address:Gaol Walk, Cork
Email:G.Clarke@ucc.ie
Phone:+353-21-4901408
Funding Source:The research was conducted in the APC Microbiome Ireland which is funded by Science Foundation Ireland (SFI/12/RC/2273). This project in part is a collaborative agreement (FA9550-17-1-0016) funded by European Office of Aerospace Research and Development, Air Force Office of Scientific Research and 711 Human Performance Wing, Air Force Research Laboratory.

Subject:

Subject ID:SU003248
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090
Genotype Strain:C57Bl/6J
Age Or Age Range:8-10 weeks
Weight Or Weight Range:20-29g
Height Or Height Range:-
Gender:Male
Animal Animal Supplier:Taconic
Animal Housing:Gnotobiotic isolators/conventional housing
Animal Light Cycle:12/12
Animal Feed:RM1A (P) and RM3A (P) from Special Diet Services
Species Group:Mammals

Factors:

Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id local_sample_id Factor
SA339176ColGF10ColGF_0min
SA339177ColGF11ColGF_0min
SA339178ColGF8ColGF_0min
SA339179ColGF6ColGF_0min
SA339180ColGF5ColGF_0min
SA339181ColGF12ColGF_0min
SA339182ColGF7ColGF_0min
SA339183ColGF20ColGF_45min
SA339184ColGF21ColGF_45min
SA339185ColGF19ColGF_45min
SA339186ColGF18ColGF_45min
SA339187ColGF17ColGF_45min
SA339188ColGF9ColGF_45min
SA339189ColGF16ColGF_45min
SA339190ColGF3ColGF_Pre
SA339191ColGF1ColGF_Pre
SA339192ColGF4ColGF_Pre
SA339193ColGF2ColGF_Pre
SA339194ColGF14ColGF_Pre
SA339195ColGF13ColGF_Pre
SA339196ColGF15ColGF_Pre
SA339197Conv17Conv_0min
SA339198Conv16Conv_0min
SA339199Conv19Conv_0min
SA339200Conv22Conv_0min
SA339201Conv13Conv_0min
SA339202Conv21Conv_0min
SA339203Conv18Conv_0min
SA339204Conv8Conv_45min
SA339205Conv7Conv_45min
SA339206Conv9Conv_45min
SA339207Conv14Conv_45min
SA339208Conv15Conv_45min
SA339209Conv6Conv_45min
SA339210Conv10Conv_45min
SA339211Conv3Conv_Pre
SA339212Conv1Conv_Pre
SA339213Conv20Conv_Pre
SA339214Conv12Conv_Pre
SA339215Conv2Conv_Pre
SA339216Conv4Conv_Pre
SA339217Conv5Conv_Pre
SA339218GF7GF_0min
SA339219GF15GF_0min
SA339220GF16GF_0min
SA339221GF6GF_0min
SA339222GF17GF_0min
SA339223GF5GF_0min
SA339224GF4GF_0min
SA339225GF18GF_45min
SA339226GF20GF_45min
SA339227GF8GF_45min
SA339228GF11GF_45min
SA339229GF19GF_45min
SA339230GF10GF_45min
SA339231GF9GF_45min
SA339232GF3GF_Pre
SA339233GF2GF_Pre
SA339234GF12GF_Pre
SA339235GF1GF_Pre
SA339236GF21GF_Pre
SA339237GF14GF_Pre
SA339238GF13GF_Pre
Showing results 1 to 63 of 63

Collection:

Collection ID:CO003241
Collection Summary:The acute restraint stress procedure was performed using a clean perforated polypropylene screw-cap 50 mL conical tubes. Cages were randomly assigned to either non-stress or stress groups. Each mouse that underwent stress was placed into the 50 mL tube and restrained for 15 minutes. After 15 minutes of restraint stress, mice were removed from the restrainer and either transported immediately to the cull room or returned to their home cage and left undisturbed for 45 minutes. To control for the variable of transport stress, mice from the non-stress control group were placed into new cages containing fresh bedding and transported the same distance before entering the cull room. Upon entering the cull room, mice were immediately decapitated, trunk blood collected, and tissues harvested. Caecal contents and intestinal samples were manually dissected and stored in PCR-grade microfuge tubes at -80°C until analyses.
Sample Type:Cecum
Storage Conditions:-80?

Treatment:

Treatment ID:TR003257
Treatment Summary:All animal work carried was approved by the Animal Experimentation Ethics Committee of University College Cork and Health Products Regulatory Authority (HPRA) before beginning this study. All experimentation was carried out in accordance with European Directive 2010/63/EU and was approved by both the Animal Experimentation Ethics Committee of University College Cork (Project Authorization AE19130/P160) and United States Air Force Surgeon General's Office of Research Oversight and Compliance. C57/BL6 mice breeding pairs were acquired from Taconic Biosciences, and F1-generation male and female offspring were used in all experiments. Germ-free, ex-germ-free, and conventional mice were housed 2-4 mice/cage under a 12-hour light/dark cycle and maintained on ad libitum autoclaved water and autoclaved, pelleted diet (Special Diet Services). Housing conditions for germ-free, conventional, and colonized germ-free adhered to the same environmental conditions of temperature (21 ± 1°C) and humidity (55%-60%). Germ-free mice were housed in gnotobiotic flexible-film isolators. Colonized germ-free mice were born and maintained as germ-free mice in gnotobiotic flexible-film isolators until postnatal day 21 when they were removed from the isolators and, for the remaining duration of this study, re-located to the standard animal facility and housed in wire-top cages that contained used-bedding from age- and sex-matched conventional mice. The acute restraint stress procedure was performed using a clean perforated polypropylene screw-cap 50 mL conical tubes. Cages were randomly assigned to either non-stress or stress groups. Each mouse that underwent stress was placed into the 50 mL tube and restrained for 15 minutes.

Sample Preparation:

Sampleprep ID:SP003255
Sampleprep Summary:Samples were prepared using the automated MicroLab STAR® system from Hamilton Company. Several recovery standards were added prior to the first step in the extraction process for QC purposes. To remove protein, dissociate small molecules bound to protein or trapped in the precipitated protein matrix, and to recover chemically diverse metabolites, proteins were precipitated with methanol under vigorous shaking for 2 min (Glen Mills GenoGrinder 2000) followed by centrifugation. The resulting extract was divided into five fractions: two for analysis by two separate reverse phase (RP)/UPLC-MS/MS methods with positive ion mode electrospray ionization (ESI), one for analysis by RP/UPLC-MS/MS with negative ion mode ESI, one for analysis by HILIC/UPLC-MS/MS with negative ion mode ESI, and one sample was reserved for backup. Samples were placed briefly on a TurboVap® (Zymark) to remove the organic solvent. The sample extracts were stored overnight under nitrogen before preparation for analysis.

Combined analysis:

Analysis ID AN005135 AN005136 AN005137 AN005138
Analysis type MS MS MS MS
Chromatography type Reversed phase Reversed phase Reversed phase HILIC
Chromatography system Waters Acquity Waters Acquity Waters Acquity Waters Acquity
Column Waters Acquity BEH C18 (100 x 2.1mm, 1.7um) Waters Acquity BEH C18 (100 x 2.1mm, 1.7um) Waters Acquity BEH C18 (100 x 2.1mm, 1.7um) Waters Acquity BEH Amide (150 x 2.1mm, 1.7um)
MS Type ESI ESI ESI ESI
MS instrument type Orbitrap Orbitrap Orbitrap Orbitrap
MS instrument name Thermo Q Exactive Orbitrap Thermo Q Exactive Orbitrap Thermo Q Exactive Orbitrap Thermo Q Exactive Orbitrap
Ion Mode POSITIVE POSITIVE NEGATIVE NEGATIVE
Units Raw Peak Area Raw Peak Area Raw Peak Area Raw Peak Area

Chromatography:

Chromatography ID:CH003885
Chromatography Summary:Low pH polar (LC/MS Pos early)
Instrument Name:Waters Acquity
Column Name:Waters Acquity BEH C18 (100 x 2.1mm, 1.7um)
Column Temperature:40-50
Flow Gradient:Linear gradient from 5% B to 80% B over 3.35 minutes
Flow Rate:0.35 mL/min
Solvent A:100% water; 0.1% formic acid; 0.05% PFPA, pH ~2.5
Solvent B:100% methanol; 0.1% formic acid; 0.05% PFPA, pH ~2.5
Chromatography Type:Reversed phase
  
Chromatography ID:CH003886
Chromatography Summary:Low pH Lipophilic (LC/MS Pos late)
Instrument Name:Waters Acquity
Column Name:Waters Acquity BEH C18 (100 x 2.1mm, 1.7um)
Column Temperature:40-50
Flow Gradient:Linear gradient from 40% B to 99.5% B over 1.0 minute, hold 99.5% B for 2.4 minutes.
Flow Rate:0.60 mL/min
Solvent A:100% water; 0.1% formic acid; 0.05% PFPA, pH ~2.5
Solvent B:50% methanol/50% acetonitrile; 0.1% formic acid; 0.05% PFPA, pH ~2.5
Chromatography Type:Reversed phase
  
Chromatography ID:CH003887
Chromatography Summary:High pH (LC/MS Neg)
Instrument Name:Waters Acquity
Column Name:Waters Acquity BEH C18 (100 x 2.1mm, 1.7um)
Column Temperature:40-50
Flow Gradient:Linear gradient from 0.5 to 70% B over 4.0 minutes, then rapid gradient to 99% B in 0.5 minutes.
Flow Rate:0.35 mL/min
Solvent A:100% water; 6.5 mM ammonium bicarbonate, pH 8
Solvent B:95% methanol/5% water; 6.5 mM ammonium bicarbonate
Chromatography Type:Reversed phase
  
Chromatography ID:CH003888
Chromatography Summary:HILIC (LC/MS Polar Neg)
Instrument Name:Waters Acquity
Column Name:Waters Acquity BEH Amide (150 x 2.1mm, 1.7um)
Column Temperature:40-50
Flow Gradient:Linear gradient from 5% B to 50% B in 3.5 minutes, then linear gradient from 50% B to 95% B in 2 minutes.
Flow Rate:0.50 mL/min
Solvent A:15% water/5% methanol/80% acetonitrile; 10 mM ammonium formate, (effective pH 10.16 with NH4OH)
Solvent B:50% water/50% acetonitrile; 10 mM ammonium formate, (effective pH 10.60 with NH4OH)
Chromatography Type:HILIC

MS:

MS ID:MS004871
Analysis ID:AN005135
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Metabolon (LC/MS Pos early)
Ion Mode:POSITIVE
  
MS ID:MS004872
Analysis ID:AN005136
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Metabolon (LC/MS Pos late)
Ion Mode:POSITIVE
  
MS ID:MS004873
Analysis ID:AN005137
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Metabolon (LC/MS Neg)
Ion Mode:NEGATIVE
  
MS ID:MS004874
Analysis ID:AN005138
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Metabolon (LC/MS Polar)#_1 MS_METABOLITE_DATA
Ion Mode:NEGATIVE
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