Summary of Study ST000224

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 PR000182. The data can be accessed directly via it's Project DOI: 10.21228/M8M888 This work is supported by NIH grant, U2C- DK119886.

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

Study ID	ST000224
Study Title	Vitamin targeted metabolomics of the small intestine in malnourished and control mice
Study Type	Targeted metabolomics
Study Summary	A total of 8 samples from 6 week old, female C57BL/6 mice, treated for 3 weeks with a malnourished diet or a control-fed isocaloric diet. Samples were taken from the small intestinal fecal content at the terminus of the ileum for targeted analysis of vitamin concentrations.
Institute	University of Victoria
Department	The Uvic Proteomics and Metabolomics Innovation Centre
Last Name	Borchers
First Name	Christoph
Address	#3101-4464 Markham St Victoria, BC Canada, V8Z 7X8
Email	christoph@proteincentre.com
Submit Date	2015-06-08
Num Groups	2
Total Subjects	8
Raw Data Available	No
Analysis Type Detail	LC-MS
Release Date	2015-06-08
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR000182
Project DOI:	doi: 10.21228/M8M888
Project Title:	Metabolomic analysis of the small intestinal content of malnourished mice
Institute:	University of British Columbia
Department:	Microbiology and Immunology
Laboratory:	Dr. B. Brett Finlay
Last Name:	Finlay
First Name:	Brett
Address:	2185 East Mall, Vancouver, BC, Canada V6T 1Z4
Email:	bfinlay@msl.ubc.ca
Funding Source:	Canadian Institutes for Health Research

Subject:

Subject ID:	SU000243
Subject Type:	Animal
Subject Species:	Mus musculus
Taxonomy ID:	10090
Genotype Strain:	C57BL/6
Age Or Age Range:	6 weeks at collection
Gender:	Female
Animal Animal Supplier:	Jackson Labs
Animal Housing:	Conventional
Animal Feed:	Low Protein, Low Fat Malnourished Diet/Isocaloric Control Diet
Cell Primary Immortalized:	Malnourished/Control
Species Group:	Mammal

Factors:

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

mb_sample_id	local_sample_id	Diet
SA010788	CON2	control
SA010789	CON3	control
SA010790	CON4	control
SA010791	CON1	control
SA010792	MAL4	malnourished
SA010793	MAL2	malnourished
SA010794	MAL3	malnourished
SA010795	MAL1	malnourished

Showing results 1 to 8 of 8

Showing results 1 to 8 of 8

Collection:

Collection ID:	CO000231
Collection Summary:	-
Sample Type:	Small intestinal fecal contents
Collection Time:	6 weeks of age
Storage Conditions:	-80 degrees
Tissue Cell Quantity Taken:	15-40mg

Treatment:

Treatment ID:	TR000251
Treatment Summary:	4 mice were given a malnourished diet and 4 mice were untreated, fed a control diet

Sample Preparation:

Sampleprep ID:	SP000245
Sampleprep Summary:	Vitamin-targeted metabolomics. The samples were analyzed by UPLC-MRM/MS on a Dionex UltiMate 3400 RSLC system coupled to an AB Sciex 4000 QTRAP triple-quadrupole mass spectrometer equipped with an electrospray ionization source. The standard substances of vitamin A (retinal, retinol, retinoic acid), B1 (thiamine), B2 (riboflavin), B3 (niacinamide), B6 (pydidoximine, pyridoxine, pyridoxal, pyridoxal-mono-phosphate), B7 (biotin), B9 (folic acid), D2, D3, E (?-tocopherol, ?-tocopherol, and ?-tocotrienol), K1 and K2, were purchased either from Sigma-Aldrich or from Cayman Chemicals Inc. The MRM transitions of individual analytes were optimized by direct infusion of a standard solution of each compound into the MS instrument. Each sample was added with a methanolic BHT (2 mg/mL) solution at a ratio of 15 ?L per mg of the small intestine digestate. Vitamins were extracted by homogenizing the samples at a shaking frequency of 30 Hz for 1 min twice using a Retsch MM400 mixer mill and with the aid of two 3-mm stainless steel metal balls, followed by 5-min sonication in an icy water bath. The samples were then centrifuged in a micro-centrifuge at 12,500 rpm and 4oC for 10 min. A 300-?L aliquot of the supernatant was transferred into a 3-mL borosilicate glass test tube and mixed with 300 ?L of water and 900 ?L of hexane. After 1 min vortex mixing, the tubes were centrifuged at 4000 rpm and 10 oC in a Beckman R22 centrifuge to separate the supernatant organic phase from the lower aqueous phase. The supernatants were carefully pipetted out to another sets of 3-mL test tubes. The fat-soluble vitamins were further extracted from the aqueous phase with 900 ?L of hexane two more times. After liquid-liquid extraction, the pooled organic phase for each sample was dried down in a speed-vacuum concentrator at room temperature. The dried residue was reconstituted in 100 ?L of ethanol. A 20-?L aliquot was injected for quantitation of the fat-soluble vitamins by LC-(+)ESI-MRM/MS on Waters BEH C18 (2.1 x 50 mm, 1.7 ?m) UPLC column and with 0.1% formic acid in water and acetonitrile as the mobile phase for binary solvent gradient elution. An efficient elution gradient was 50% to 100% B in 10 min. The column temperature was 50oC and the flow rate was 300 ?L/min. The aqueous phases were loaded onto reversed-phase polymeric HLB cartridges (60 mg/1mL, Waters Inc.), which have been activated with 1 mL of methanol and equilibrated with 1 mL of 50% methanol before use. Under a 5-inch Hg vaccum, the flow-throw fractions were collected and the resins were washed with 1 mL of 50% methanol with the flow-through fractions collected. The pooled flow-through fractions were dried in a nitrogen evaporator at 30 oC. The residue from each sample was reconstituted in 100 ?L of 2% methanol. A 20-?L aliquot was injected for quantitation of the water-soluble vitamins by UPLC-MRM/MS with (+) or (-) ESI and on a Waters BEH C18 (2.1 x 150 mm, 1.7 ?m) UPLC column and using 0.01% formic acid in water and methanol as the mobile phase for binary solvent gradient elution. The efficient elution gradient was 2% B for 0.5 min and 2% to 50% B in 8 min. The column temperature was 30 oC and the flow rate was 250 ?L/min. The concentrations of all the detected vitamins were calculated from the standard calibration curves of individual vitamins, which were prepared with the use of their authentic compounds.

Sampleprep ID:

SP000245

Sampleprep Summary:

Vitamin-targeted metabolomics. The samples were analyzed by UPLC-MRM/MS on a Dionex UltiMate 3400 RSLC system coupled to an AB Sciex 4000 QTRAP triple-quadrupole mass spectrometer equipped with an electrospray ionization source. The standard substances of vitamin A (retinal, retinol, retinoic acid), B1 (thiamine), B2 (riboflavin), B3 (niacinamide), B6 (pydidoximine, pyridoxine, pyridoxal, pyridoxal-mono-phosphate), B7 (biotin), B9 (folic acid), D2, D3, E (?-tocopherol, ?-tocopherol, and ?-tocotrienol), K1 and K2, were purchased either from Sigma-Aldrich or from Cayman Chemicals Inc. The MRM transitions of individual analytes were optimized by direct infusion of a standard solution of each compound into the MS instrument. Each sample was added with a methanolic BHT (2 mg/mL) solution at a ratio of 15 ?L per mg of the small intestine digestate. Vitamins were extracted by homogenizing the samples at a shaking frequency of 30 Hz for 1 min twice using a Retsch MM400 mixer mill and with the aid of two 3-mm stainless steel metal balls, followed by 5-min sonication in an icy water bath. The samples were then centrifuged in a micro-centrifuge at 12,500 rpm and 4oC for 10 min. A 300-?L aliquot of the supernatant was transferred into a 3-mL borosilicate glass test tube and mixed with 300 ?L of water and 900 ?L of hexane. After 1 min vortex mixing, the tubes were centrifuged at 4000 rpm and 10 oC in a Beckman R22 centrifuge to separate the supernatant organic phase from the lower aqueous phase. The supernatants were carefully pipetted out to another sets of 3-mL test tubes. The fat-soluble vitamins were further extracted from the aqueous phase with 900 ?L of hexane two more times. After liquid-liquid extraction, the pooled organic phase for each sample was dried down in a speed-vacuum concentrator at room temperature. The dried residue was reconstituted in 100 ?L of ethanol. A 20-?L aliquot was injected for quantitation of the fat-soluble vitamins by LC-(+)ESI-MRM/MS on Waters BEH C18 (2.1 x 50 mm, 1.7 ?m) UPLC column and with 0.1% formic acid in water and acetonitrile as the mobile phase for binary solvent gradient elution. An efficient elution gradient was 50% to 100% B in 10 min. The column temperature was 50oC and the flow rate was 300 ?L/min. The aqueous phases were loaded onto reversed-phase polymeric HLB cartridges (60 mg/1mL, Waters Inc.), which have been activated with 1 mL of methanol and equilibrated with 1 mL of 50% methanol before use. Under a 5-inch Hg vaccum, the flow-throw fractions were collected and the resins were washed with 1 mL of 50% methanol with the flow-through fractions collected. The pooled flow-through fractions were dried in a nitrogen evaporator at 30 oC. The residue from each sample was reconstituted in 100 ?L of 2% methanol. A 20-?L aliquot was injected for quantitation of the water-soluble vitamins by UPLC-MRM/MS with (+) or (-) ESI and on a Waters BEH C18 (2.1 x 150 mm, 1.7 ?m) UPLC column and using 0.01% formic acid in water and methanol as the mobile phase for binary solvent gradient elution. The efficient elution gradient was 2% B for 0.5 min and 2% to 50% B in 8 min. The column temperature was 30 oC and the flow rate was 250 ?L/min. The concentrations of all the detected vitamins were calculated from the standard calibration curves of individual vitamins, which were prepared with the use of their authentic compounds.

Combined analysis:

Analysis ID	AN000333	AN000334
Analysis type	MS	MS
Chromatography type
Chromatography system
Column
MS Type	ESI	ESI
MS instrument type	Triple quadrupole	Triple quadrupole
MS instrument name	ABI Sciex API 4000 QTrap	ABI Sciex API 4000 QTrap
Ion Mode	NEGATIVE	POSITIVE
Units	nmol/g	nmol/g

Chromatography:

Chromatography ID:	CH000250

Chromatography ID:

CH000250

MS:

MS ID:	MS000281
Analysis ID:	AN000333
Instrument Name:	ABI Sciex API 4000 QTrap
Instrument Type:	Triple quadrupole
MS Type:	ESI
Ion Mode:	NEGATIVE

MS ID:	MS000282
Analysis ID:	AN000334
Instrument Name:	ABI Sciex API 4000 QTrap
Instrument Type:	Triple quadrupole
MS Type:	ESI
Ion Mode:	POSITIVE