#METABOLOMICS WORKBENCH kcbauer_20200420_130612 DATATRACK_ID:1990 STUDY_ID:ST001367 ANALYSIS_ID:AN002278 PROJECT_ID:PR000935
VERSION             	1
CREATED_ON             	April 28, 2020, 4:26 pm
#PROJECT
PR:PROJECT_TITLE                 	Malnutrition Gut microbes and Liver Metabolomics
PR:PROJECT_SUMMARY               	Examining the role of malnutrition and gut microbes on fatty liver using the
PR:PROJECT_SUMMARY               	MAL-BG murine model. RP-UPLC-FTMS and HILIC-FTMS were conducted on murine livers
PR:PROJECT_SUMMARY               	prior and following dietary intervention. This submission contains prior
PR:PROJECT_SUMMARY               	intervention raw data.
PR:INSTITUTE                     	University of British Columbia
PR:DEPARTMENT                    	Microbiology and Immunology
PR:LABORATORY                    	Finlay
PR:LAST_NAME                     	Bauer
PR:FIRST_NAME                    	Kylynda
PR:ADDRESS                       	#301 – 2185 East Mall, University of British Columbia Vancouver, British
PR:ADDRESS                       	Columbia Canada, V6T 1Z4
PR:EMAIL                         	kcbauer@msl.ubc.ca
PR:PHONE                         	(604) 822-2210
PR:PROJECT_COMMENTS              	Pre-Intervention
PR:CONTRIBUTORS                  	B. Brett Finlay
#STUDY
ST:STUDY_TITLE                   	Malnutrition and Liver Metabolomics Pre-Intervention (part-I)
ST:STUDY_SUMMARY                 	RP-UPLC-FTMS (+/- ion detection) and HILIC-FTMS (+/- ion detection) were
ST:STUDY_SUMMARY                 	conducted on murine livers from healthy (CON) and malnourished (MAL) mice. To
ST:STUDY_SUMMARY                 	examine the impact of gut microbes and malnutrition, data was also collected
ST:STUDY_SUMMARY                 	from a third group (MBG).
ST:INSTITUTE                     	University of British Columbia
ST:DEPARTMENT                    	Microbiology and Immunology
ST:LABORATORY                    	B. Brett Finlay
ST:LAST_NAME                     	Bauer
ST:FIRST_NAME                    	Kylynda
ST:ADDRESS                       	Michael Smith Laboratories, #301 – 2185 East Mall, University of British
ST:ADDRESS                       	Columbia Vancouver, British Columbia Canada, V6T 1Z4
ST:EMAIL                         	kcbauer@msl.ubc.ca
ST:PHONE                         	(604) 822-2210
ST:NUM_GROUPS                    	3
ST:TOTAL_SUBJECTS                	12
ST:NUM_FEMALES                   	12
#SUBJECT
SU:SUBJECT_TYPE                  	Mammal
SU:SUBJECT_SPECIES               	Mus musculus
SU:TAXONOMY_ID                   	10090
SU:GENOTYPE_STRAIN               	C57BL/6J
SU:AGE_OR_AGE_RANGE              	~7 weeks (Pre-intervention)
SU:GENDER                        	Female
SU:ANIMAL_ANIMAL_SUPPLIER        	Jackson Laboratories
#SUBJECT_SAMPLE_FACTORS:         	SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data
SUBJECT_SAMPLE_FACTORS           	-	CONA_1	Treatment:CON | Start Diet:Healthy | Final Diet:Healthy	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	CONA_2	Treatment:CON | Start Diet:Healthy | Final Diet:Healthy	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	CONA_3	Treatment:CON | Start Diet:Healthy | Final Diet:Healthy	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	CONA_4	Treatment:CON | Start Diet:Healthy | Final Diet:Healthy	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	MALA_1	Treatment:MAL | Start Diet:Malnourished | Final Diet:Malnourished	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	MALA_2	Treatment:MAL | Start Diet:Malnourished | Final Diet:Malnourished	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	MALA_3	Treatment:MAL | Start Diet:Malnourished | Final Diet:Malnourished	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	MALA_4	Treatment:MAL | Start Diet:Malnourished | Final Diet:Malnourished	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	MBGA_1	Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	MBGA_2	Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	MBGA_3	Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished	Study=Pre-Intervention
SUBJECT_SAMPLE_FACTORS           	-	MBGA_4	Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished	Study=Pre-Intervention
#COLLECTION
CO:COLLECTION_SUMMARY            	Following sacrifice, murine liver lobes were isolated and stored at -70/80 C
CO:COLLECTION_SUMMARY            	prior to untargeted metabolomics.
CO:SAMPLE_TYPE                   	Liver
#TREATMENT
TR:TREATMENT_SUMMARY             	CON = control mice placed on a healthy (standard mouse chow), MAL = mice placed
TR:TREATMENT_SUMMARY             	on an isocaloric malnourished diet (protein/fat deficient), MBG = MAL mice with
TR:TREATMENT_SUMMARY             	repeated exposure to fecal microbes, mice were placed on the respective
TR:TREATMENT_SUMMARY             	treatments for one month following weaning.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	RP-UPLC-FTMS: Individual mouse liver samples were mixed with water, 5 μL per mg
SP:SAMPLEPREP_SUMMARY            	of the tissue, and two 4-mm metal balls were added to an Eppendorf tube. The
SP:SAMPLEPREP_SUMMARY            	tissue was homogenized on a MM 400 mill mixer at a vibrating frequency of 30 Hz
SP:SAMPLEPREP_SUMMARY            	for 1 min twice. After 5-s spin-down, a mixture of methanol-chloroform (4:1) was
SP:SAMPLEPREP_SUMMARY            	added, at 25 μL per mg tissue, to each tube. The sample was homogenized again
SP:SAMPLEPREP_SUMMARY            	for metabolite extraction using the same setup for 1 min twice, followed by
SP:SAMPLEPREP_SUMMARY            	sonication in an ice-water bath for 5 min. The tube was centrifuged at 15,000
SP:SAMPLEPREP_SUMMARY            	rpm and at 10 0C for 20 min. The clear supernatant was transferred to a 1.5-mL
SP:SAMPLEPREP_SUMMARY            	Eppendorf tube. A 60-μL aliquot from each sample was dried down inside the same
SP:SAMPLEPREP_SUMMARY            	nitrogen evaporator and the residue was reconstituted in 40 μL of 80% methanol.
SP:SAMPLEPREP_SUMMARY            	10 μL was injected for reversed-phase RP-UPLC-FTMS. HILIC-FTMS: Individual
SP:SAMPLEPREP_SUMMARY            	sample supernatants were mixed with 120 μL of water, 180 μL of methanol and
SP:SAMPLEPREP_SUMMARY            	195 μL of chloroform. The mixture was vortex mixed at 3000 rpm for 30 s before
SP:SAMPLEPREP_SUMMARY            	centrifugal clarification. 300 μL of the upper, aqueous phase was precisely
SP:SAMPLEPREP_SUMMARY            	taken out and transferred to a “V”-shape LC injection microvial and was
SP:SAMPLEPREP_SUMMARY            	dried down under a gentle nitrogen gas flow in the nitrogen evaporator. The
SP:SAMPLEPREP_SUMMARY            	residue was reconstituted in 50 μL of 80% acetonitrile. 10 μL was injected for
SP:SAMPLEPREP_SUMMARY            	HILIC-FTMS. *See Methods
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_TYPE           	HILIC
CH:INSTRUMENT_NAME               	Thermo Dionex Ultimate 3000
CH:COLUMN_NAME                   	Waters HILIC column (2.1 x 100 mm, 1.8 µm)
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Thermo LTQ-Orbitrap Velos Pro
MS:INSTRUMENT_TYPE               	Orbitrap
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	NEGATIVE
MS:MS_COMMENTS                   	The MS instrument was run in the survey scan mode with FTMS detection at a mass
MS:MS_COMMENTS                   	resolution of 60,000 FWHM at m/z 400. Two HILIC-FTMS datasets were acquired for
MS:MS_COMMENTS                   	each sample, one with positive-ion detection and the other with negative-ion
MS:MS_COMMENTS                   	detection. The mass scan range was m/z 80 to 800 (from publication methods).
MS:MS_RESULTS_FILE               	ST001367_AN002278_Results.txt	UNITS:abundance	Has m/z:Yes	Has RT:Yes	RT units:Minutes
#END