Summary of Study ST003288

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 PR002040. The data can be accessed directly via it's Project DOI: 10.21228/M8XB94 This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

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.

Study ID	ST003288
Study Title	Quantification of amoxicillin in mouse intestinal contents
Study Summary	Conventionally housed mice were orally treated with amoxicillin for 24 hours with and without cholic acid (48h) and small intestinal and cecal contents were collected for targeted MS analysis. Samples were flash frozen and stored at -80°C prior to analysis. We found that amoxicillin levels in the small intestine were 0-40 pmol/mg of contents and no significant difference in amoxicillin concentration was detected between small intestine and cecum or between mice receiving cholic acid vs amoxicillin alone.
Institute	Brown University
Department	Molecular Microbiology & Immunology
Laboratory	Molecular Microbiology & Immunology, Belenky Lab
Last Name	Beekman
First Name	Chapman
Address	BMC 613, 171 Meeting Street, Providence RI 02912
Email	Chapman_Beekman@brown.edu
Phone	4018635953
Submit Date	2024-06-21
Raw Data Available	Yes
Raw Data File Type(s)	raw(Thermo)
Analysis Type Detail	LC-MS
Release Date	2024-08-05
Release Version	1

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Project:

Project ID:	PR002040
Project DOI:	doi: 10.21228/M8XB94
Project Title:	Quantification of amoxicillin in intestinal contents of mice
Project Summary:	Antibiotics cause collateral damage to resident microbes that is associated with various health risks. To-date, studies have largely focused on impacts of antibiotics on large intestinal and fecal microbiota. Here, we employ a GI-wide integrated multiomic approach to reveal that amoxicillin (AMX) treatment reduces overall bacterial abundance, bile salt hydrolase activity and unconjugated bile acids in the small intestine (SI). An accompanying loss of fatty acids and increase in acyl-carnitines in the large intestine corresponded with spatially-distinct expansions of proteobacteria. Parasuterella excrementihominis utilized fatty acid biosynthesis, becoming dominant in the SI while multiple Klebsiella species employed fatty acid oxidation during expansion in the large intestine. We subsequently demonstrate that restoration of unconjugated bile acids can mitigate losses of commensals in the large intestine while also inhibiting the expansion of Proteobacteria during AMX treatment.
Institute:	Brown University
Last Name:	Beekman
First Name:	Chapman
Address:	BMC 613, 171 Meeting Street, Providence RI 02912
Email:	Chapman_Beekman@brown.edu
Phone:	4018635953

Subject:

Subject ID:	SU003408
Subject Type:	Mammal
Subject Species:	Mus musculus
Taxonomy ID:	10090

Factors:

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

mb_sample_id	local_sample_id	Treatment	Sample source
SA356323	Cec_40	AMX + CA	mouse cecum
SA356324	Cec_38	AMX + CA	mouse cecum
SA356325	Cec_37	AMX + CA	mouse cecum
SA356326	Cec_06	AMX + CA	mouse cecum
SA356327	Cec_32	AMX + CA	mouse cecum
SA356328	Cec_31	AMX + CA	mouse cecum
SA356329	Cec_30	AMX + CA	mouse cecum
SA356330	Cec_29	AMX + CA	mouse cecum
SA356331	Cec_39	AMX + CA	mouse cecum
SA356332	Cec_07	AMX + CA	mouse cecum
SA356333	Cec_08	AMX + CA	mouse cecum
SA356334	SI_30	AMX + CA	mouse small intestine
SA356335	SI_31	AMX + CA	mouse small intestine
SA356336	SI_32	AMX + CA	mouse small intestine
SA356337	SI_37	AMX + CA	mouse small intestine
SA356338	SI_38	AMX + CA	mouse small intestine
SA356339	SI_39	AMX + CA	mouse small intestine
SA356340	SI_08	AMX + CA	mouse small intestine
SA356341	SI_40	AMX + CA	mouse small intestine
SA356342	SI_06	AMX + CA	mouse small intestine
SA356343	SI_29	AMX + CA	mouse small intestine
SA356344	SI_07	AMX + CA	mouse small intestine
SA356345	Cec_16	AMX + std. diet	mouse cecum
SA356346	Cec_23	AMX + std. diet	mouse cecum
SA356347	Cec_34	AMX + std. diet	mouse cecum
SA356348	Cec_33	AMX + std. diet	mouse cecum
SA356349	Cec_13	AMX + std. diet	mouse cecum
SA356350	Cec_14	AMX + std. diet	mouse cecum
SA356351	Cec_21	AMX + std. diet	mouse cecum
SA356352	Cec_15	AMX + std. diet	mouse cecum
SA356353	Cec_22	AMX + std. diet	mouse cecum
SA356354	Cec_35	AMX + std. diet	mouse cecum
SA356355	Cec_24	AMX + std. diet	mouse cecum
SA356356	SI_36	AMX + std. diet	mouse small intestine
SA356357	SI_35	AMX + std. diet	mouse small intestine
SA356358	SI_34	AMX + std. diet	mouse small intestine
SA356359	SI_33	AMX + std. diet	mouse small intestine
SA356360	SI_24	AMX + std. diet	mouse small intestine
SA356361	SI_23	AMX + std. diet	mouse small intestine
SA356362	SI_21	AMX + std. diet	mouse small intestine
SA356363	SI_16	AMX + std. diet	mouse small intestine
SA356364	SI_15	AMX + std. diet	mouse small intestine
SA356365	SI_14	AMX + std. diet	mouse small intestine
SA356366	SI_13	AMX + std. diet	mouse small intestine
SA356367	SI_22	AMX + std. diet	mouse small intestine
SA356368	Cec_05	untreated + CA	mouse cecum
SA356369	SI_05	untreated + CA	mouse small intestine
SA356370	SI_10	untreated + std. diet	mouse small intestine
SA356371	SI_09	untreated + std. diet	mouse small intestine

Showing results 1 to 49 of 49

Showing results 1 to 49 of 49

Collection:

Collection ID:	CO003401
Collection Summary:	SI and cecal contents were homogenized at a concentration of 50-100 mg/mL in 70% methanol (LCMS grade) containing 50 µM 13C6-labeled AMX as an internal standard on the Bead Ruptor (Omni Inc.) without beads at a frequency setting of 15 for 5 minutes. Homogenized samples were then centrifuged at 16,000 X g for 10 minutes and supernatants were used for analysis. AMX was quantitated with HPLC-MS on an Ultimate 3000, Dionex coupled to Q Executive Classic (Thermo) with ESI interface.
Sample Type:	Intestine

Treatment:

Treatment ID:	TR003417
Treatment Summary:	AMX treatment was administered in drinking water (166.7 µg/mL) for a 24-hour period. This concentration is based on previous studies38,39 and determined to achieve a dosage of approximately 25mg/kg/day. Untreated mice received pH-matched water and both AMX and vehicle solutions were sterile filtered prior to administration. For cholic acid supplementation experiments standard mouse chow was first blended into a course powder using a blender (Vitamix) and 5mg of cholic acid was added per gram of powdered chow. Cholic acid was mixed evenly by re-blending and provided in the powdered chow for a 48-hour period and 24 hours prior to AMX treatment.

Treatment ID:

TR003417

Treatment Summary:

AMX treatment was administered in drinking water (166.7 µg/mL) for a 24-hour period. This concentration is based on previous studies38,39 and determined to achieve a dosage of approximately 25mg/kg/day. Untreated mice received pH-matched water and both AMX and vehicle solutions were sterile filtered prior to administration. For cholic acid supplementation experiments standard mouse chow was first blended into a course powder using a blender (Vitamix) and 5mg of cholic acid was added per gram of powdered chow. Cholic acid was mixed evenly by re-blending and provided in the powdered chow for a 48-hour period and 24 hours prior to AMX treatment.

Sample Preparation:

Sampleprep ID:	SP003415
Sampleprep Summary:	SI and cecal contents were homogenized at a concentration of 50-100 mg/mL in 70% methanol (LCMS grade) containing 50 µM 13C6-labeled AMX as an internal standard on the Bead Ruptor (Omni Inc.) without beads at a frequency setting of 15 for 5 minutes. Homogenized samples were then centrifuged at 16,000 X g for 10 minutes and supernatants were used for analysis. AMX was quantitated with HPLC-MS on an Ultimate 3000, Dionex coupled to Q Executive Classic (Thermo) with ESI interface.

Sampleprep ID:

SP003415

Sampleprep Summary:

SI and cecal contents were homogenized at a concentration of 50-100 mg/mL in 70% methanol (LCMS grade) containing 50 µM 13C6-labeled AMX as an internal standard on the Bead Ruptor (Omni Inc.) without beads at a frequency setting of 15 for 5 minutes. Homogenized samples were then centrifuged at 16,000 X g for 10 minutes and supernatants were used for analysis. AMX was quantitated with HPLC-MS on an Ultimate 3000, Dionex coupled to Q Executive Classic (Thermo) with ESI interface.

Combined analysis:

Analysis ID	AN005385
Analysis type	MS
Chromatography type	Reversed phase
Chromatography system	Thermo Dionex Ultimate 3000
Column	Waters ACQUITY UPLC BEH C18 (50 x 2.1mm,1.7um)
MS Type	ESI
MS instrument type	Orbitrap
MS instrument name	Thermo Q Exactive Orbitrap
Ion Mode	POSITIVE
Units	nmol / mg sample

Chromatography:

Chromatography ID:	CH004082
Instrument Name:	Thermo Dionex Ultimate 3000
Column Name:	Waters ACQUITY UPLC BEH C18 (50 x 2.1mm,1.7um)
Column Temperature:	30°C
Flow Gradient:	mobile phase A/B set to 90%/10% from 0.00 to 0.50 min and 90%/10% to 5%/95% from 0.5 to 1.6 min and held at A/B ratio of 5%/95% for 0.5 min from 1.6 to 2.1 min (wash) and set back to the starting composition A/B 90%/10% from 2.1 to 4.5 (for equilibration)
Flow Rate:	0.35 mL/min
Solvent A:	100% Water; 0.2% Formic acid
Solvent B:	100% Methanol; 0.2% Formic acid
Chromatography Type:	Reversed phase

MS:

MS ID:	MS005112
Analysis ID:	AN005385
Instrument Name:	Thermo Q Exactive Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	The mass spectrometer was operated in the positive ion mode in Full MS; at resolution 70,000; AGT target 5 E5 and Scan range 200-500 m/z. Spray voltage and source temperature were set at 3,500 volts, and 320°C, respectively.
Ion Mode:	POSITIVE