#METABOLOMICS WORKBENCH kcbauer_20200428_133208_mwtab.txt DATATRACK_ID:1997 STUDY_ID:ST001368 ANALYSIS_ID:AN002279 PROJECT_ID:000000 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 intervention PR:PROJECT_SUMMARY 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 Intervention PR:CONTRIBUTORS B. Brett Finlay #STUDY ST:STUDY_TITLE Malnutrition and Liver Metabolomics Intervention (part-II) 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 model (MBG) mice. ST:STUDY_SUMMARY To examine the impact of dietary intervention the same untargeted metabolomics ST:STUDY_SUMMARY was conducted following an intervention treatment (CON, MBG, C-MBG, MBG-R ST:STUDY_SUMMARY groups). C-MBG = healthy to malnourished, MBG-R = malnourished model reversed on ST:STUDY_SUMMARY a healthy diet. 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 4 ST:TOTAL_SUBJECTS 32 ST:NUM_FEMALES 32 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN C57BL/6J SU:AGE_OR_AGE_RANGE ~11 weeks (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 - CON_1 Study:Intervention | Treatment:CON | Start Diet:Healthy | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - CON_2 Study:Intervention | Treatment:CON | Start Diet:Healthy | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - CON_3 Study:Intervention | Treatment:CON | Start Diet:Healthy | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - CON_4 Study:Intervention | Treatment:CON | Start Diet:Healthy | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - CON_5 Study:Intervention | Treatment:CON | Start Diet:Healthy | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - CON_6 Study:Intervention | Treatment:CON | Start Diet:Healthy | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - CON_7 Study:Intervention | Treatment:CON | Start Diet:Healthy | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - CON_8 Study:Intervention | Treatment:CON | Start Diet:Healthy | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - C-MBG_1 Study:Intervention | Treatment:C-MBG | Start Diet:Healthy | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - C-MBG_2 Study:Intervention | Treatment:C-MBG | Start Diet:Healthy | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - C-MBG_3 Study:Intervention | Treatment:C-MBG | Start Diet:Healthy | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - C-MBG_4 Study:Intervention | Treatment:C-MBG | Start Diet:Healthy | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - C-MBG_5 Study:Intervention | Treatment:C-MBG | Start Diet:Healthy | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - C-MBG_6 Study:Intervention | Treatment:C-MBG | Start Diet:Healthy | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - C-MBG_7 Study:Intervention | Treatment:C-MBG | Start Diet:Healthy | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - C-MBG_8 Study:Intervention | Treatment:C-MBG | Start Diet:Healthy | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - MBG_1 Study:Intervention | Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - MBG_2 Study:Intervention | Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - MBG_3 Study:Intervention | Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - MBG_4 Study:Intervention | Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - MBG_5 Study:Intervention | Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - MBG_6 Study:Intervention | Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - MBG_7 Study:Intervention | Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - MBG_8 Study:Intervention | Treatment:MBG | Start Diet:Malnourished | Final Diet:Malnourished SUBJECT_SAMPLE_FACTORS - MBG-R_1 Study:Intervention | Treatment:MBG-R | Start Diet:Malnourished | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - MBG-R_2 Study:Intervention | Treatment:MBG-R | Start Diet:Malnourished | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - MBG-R_3 Study:Intervention | Treatment:MBG-R | Start Diet:Malnourished | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - MBG-R_4 Study:Intervention | Treatment:MBG-R | Start Diet:Malnourished | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - MBG-R_5 Study:Intervention | Treatment:MBG-R | Start Diet:Malnourished | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - MBG-R_6 Study:Intervention | Treatment:MBG-R | Start Diet:Malnourished | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - MBG-R_7 Study:Intervention | Treatment:MBG-R | Start Diet:Malnourished | Final Diet:Healthy SUBJECT_SAMPLE_FACTORS - MBG-R_8 Study:Intervention | Treatment:MBG-R | Start Diet:Malnourished | Final Diet:Healthy #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) for 2 months, MBG = TR:TREATMENT_SUMMARY malnourished model for 2 months, C-MBG = control diet one month and then TR:TREATMENT_SUMMARY malnourished model for one month, MBG-R = malnourished model for one month and TR:TREATMENT_SUMMARY then reversed on the healthy diet #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 Reversed phase CH:INSTRUMENT_NAME Thermo Dionex Ultimate 3000 CH:COLUMN_NAME Waters BEH C8 (2.1 x 50 mm, 1.7 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 POSITIVE MS:MS_COMMENTS For relative quantitation, the MS instrument was run in the survey scan mode MS:MS_COMMENTS with FTMS detection at a mass resolution of 60,000 FWHM at m/z 400. The mass MS:MS_COMMENTS scan range was m/z 80 to 1800, with a reference lock-mass for real-time MS:MS_COMMENTS calibration. Two UPLC-FTMS datasets were acquired for each sample, one with MS:MS_COMMENTS positive-ion detection and the other with negative-ion detection. LC-MS/MS data MS:MS_COMMENTS was also acquired from each sample set with collision induced dissociation (CID) MS:MS_COMMENTS at different levels of normalized collision energy (from publication methods). MS:MS_RESULTS_FILE ST001368_AN002279_Results.txt UNITS:Abundance Has m/z:Yes Has RT:Yes RT units:Minutes #END