#METABOLOMICS WORKBENCH Reeddom_20210522_192918_mwtab.txt DATATRACK_ID:2652 STUDY_ID:ST001911 ANALYSIS_ID:AN003108 PROJECT_ID:PR001205
VERSION             	1
CREATED_ON             	August 20, 2021, 10:13 am
#PROJECT
PR:PROJECT_TITLE                 	Metabolome changes induced by bitter melon (Momordica charantia)- intake in a
PR:PROJECT_TITLE                 	high-fat diet induced obesity model
PR:PROJECT_TYPE                  	LCMS quanitative analysis
PR:PROJECT_SUMMARY               	Quanitative analysis of metabolomics on samples from murine samples (KEGG
PR:PROJECT_SUMMARY               	Pathway MAPS, LIPIDMAPS, Human Metabolome Database)
PR:INSTITUTE                     	University of Colorado Anschutz Medical Campus
PR:DEPARTMENT                    	Pharmaceutical Sciences
PR:LABORATORY                    	Dr. Rajesh Agarwal's Laboratory and
PR:LAST_NAME                     	Agarwal
PR:FIRST_NAME                    	Rajesh
PR:ADDRESS                       	12850 E. Montview Blvd, C238, Room V20-2118, Aurora, CO 80045, USA
PR:EMAIL                         	Rajesh.Agarwal@cuanschutz.edu
PR:PHONE                         	303-724-4055
PR:FUNDING_SOURCE                	This work was supported by the National Institutes of Health/ National Cancer
PR:FUNDING_SOURCE                	Institute grant R01CA195708, the National Institutes of Health Office of Dietary
PR:FUNDING_SOURCE                	Supplements supplemental funds R01CA195708-02S1, and the pilot funding from the
PR:FUNDING_SOURCE                	office of the Associate Dean for Research and Graduate Education (ADR), School
PR:FUNDING_SOURCE                	of Pharmacy, UC AMC (to RA), and the National Cancer Institute diversity
PR:FUNDING_SOURCE                	supplement grant R01CA195708-04S1 (to DR and RA).
PR:CONTRIBUTORS                  	Dominique Reed, Dileep Kumar, Sushil Kumar, Komal Raina, Reenu Punia, Charmion
PR:CONTRIBUTORS                  	Cruickshank-Quinn, Boris Tabakoff, Nichole Reisdorph, Michael Wempe, Chapla
PR:CONTRIBUTORS                  	Agarwal
#STUDY
ST:STUDY_TITLE                   	A high-fat diet induced obesity murine model treated with bitter melon
ST:STUDY_TITLE                   	(Momordica charantia)
ST:STUDY_TYPE                    	A 40-day Intervention experiment
ST:STUDY_SUMMARY                 	Metabolic effects of lyophilized bitter melon juice (BMJ) extract (oral gavage
ST:STUDY_SUMMARY                 	200mg/kg/body weight-daily for 40 days) intake were evaluated in diet-induced
ST:STUDY_SUMMARY                 	obese C57BL/6J male mice [fed-high fat diet (HFD), 60 kcal% fat]. Changes in
ST:STUDY_SUMMARY                 	metabolite abundance levels in lipid-phase plasma [liquid chromatography mass
ST:STUDY_SUMMARY                 	spectrometry (LC-MS)-based metabolomics] after BMJ intervention were assessed.
ST:INSTITUTE                     	University of Colorado Anschutz Medical Campus
ST:DEPARTMENT                    	Pharmaceutical Sciences
ST:LABORATORY                    	Dr. Rajesh Agarwal's Laboratory
ST:LAST_NAME                     	Agarwal
ST:FIRST_NAME                    	Rajesh
ST:ADDRESS                       	12850 E. Montview Blvd, C238, Room V20-2118, Aurora, CO 80045, USA
ST:EMAIL                         	Rajesh.Agarwal@cuanschutz.edu
ST:PHONE                         	303-724-4055
ST:NUM_GROUPS                    	2
ST:TOTAL_SUBJECTS                	8
ST:NUM_MALES                     	8
#SUBJECT
SU:SUBJECT_TYPE                  	Mammal
SU:SUBJECT_SPECIES               	Mus musculus
SU:TAXONOMY_ID                   	10090
SU:GENOTYPE_STRAIN               	wildtype
SU:AGE_OR_AGE_RANGE              	4 weeks
SU:GENDER                        	Male
SU:ANIMAL_ANIMAL_SUPPLIER        	Jackson Laboratory
SU:ANIMAL_HOUSING                	UC Denver
SU:ANIMAL_FEED                   	High fat diet (HFD), 60 kcal% fat from Research diets Inc., # D12492
SU:ANIMAL_WATER                  	ad libitum
#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           	-	OB_07(raw)	Factor:High Fat	RAW_FILE_NAME=OB_07
SUBJECT_SAMPLE_FACTORS           	-	OB_11(raw)	Factor:High Fat	RAW_FILE_NAME=OB_11
SUBJECT_SAMPLE_FACTORS           	-	OB_12_A(raw)	Factor:High Fat	RAW_FILE_NAME=OB_12_A
SUBJECT_SAMPLE_FACTORS           	-	OB+BMJ_03(raw)	Factor:Treated	RAW_FILE_NAME=OB+BMJ_03
SUBJECT_SAMPLE_FACTORS           	-	OB+BMJ_04(raw)	Factor:Treated	RAW_FILE_NAME=OB+BMJ_04
SUBJECT_SAMPLE_FACTORS           	-	OB+BMJ_05(raw)	Factor:Treated	RAW_FILE_NAME=OB+BMJ_05
#COLLECTION
CO:COLLECTION_SUMMARY            	Blood was collected and plasma/serum flash frozen in liquid nitrogen and stored
CO:COLLECTION_SUMMARY            	in -80 degrees Celsius for biochemical/metabolic assessments.
CO:SAMPLE_TYPE                   	Blood (plasma)
#TREATMENT
TR:TREATMENT_SUMMARY             	Sixteen-week-old diet-induced obese (DIO) C57BL/6J male mice [fed-high fat diet
TR:TREATMENT_SUMMARY             	(HFD), 60 kcal% fat from Research diets Inc., # D12492) from 4 weeks of age]
TR:TREATMENT_SUMMARY             	were purchased from Jackson labs (Bar Harbor, ME). Once received at the animal
TR:TREATMENT_SUMMARY             	house facility in UC Denver, the mice were randomized into two groups (n=4/
TR:TREATMENT_SUMMARY             	group): a) Control-DIO group maintained on the 60 kcal% HFD for additional 40
TR:TREATMENT_SUMMARY             	days, and b) BMJ+DIO group maintained on 60 kcal% HFD fat for additional 40 days
TR:TREATMENT_SUMMARY             	plus orally gavage with lyophilized bitter melon juice extract [(5mg per mouse;
TR:TREATMENT_SUMMARY             	total volume of 100 μl, 200mg/kg body weight/ day), reconstituted in sterile
TR:TREATMENT_SUMMARY             	deionized water immediately before use)] throughout the study.
TR:TREATMENT                     	Intervention
TR:TREATMENT_COMPOUND            	Bitter melon juice
TR:TREATMENT_ROUTE               	Oral gavage
TR:TREATMENT_DOSE                	200mg/kg body weight/ day
TR:ANIMAL_FASTING                	On day 40 of study initiation, mice in control and treament groups were
TR:ANIMAL_FASTING                	subjected to fasting for six hours prior to sacrifice.
TR:ANIMAL_ENDP_EUTHANASIA        	On day 40, mice were sacrificed after CO2 asphyxiation and exsanguination.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	Frozen plasma was stored at -80°C until sample preparation for untargeted
SP:SAMPLEPREP_SUMMARY            	liquid-chromatography mass spectrometry (LC-MS)-based metabolomics. Briefly,
SP:SAMPLEPREP_SUMMARY            	100µL of sample underwent protein precipitation using methanol, followed by
SP:SAMPLEPREP_SUMMARY            	liquid-liquid extraction using methyl-tert butyl ether as previously
SP:SAMPLEPREP_SUMMARY            	described[1, 2] to obtain an aqueous fraction and a lipid fraction. The lipid
SP:SAMPLEPREP_SUMMARY            	fraction was dried down and resuspended in 100% methanol for LC-MS analysis.
SP:PROCESSING_STORAGE_CONDITIONS 	Described in summary
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_SUMMARY        	The samples from the lipid fraction were analyzed an Agilent 1290 series pump
CH:CHROMATOGRAPHY_SUMMARY        	with an Agilent Zorbax Rapid Resolution HD (RRHD) SB-C18, 1.8 micron (2.1 x 100
CH:CHROMATOGRAPHY_SUMMARY        	mm) analytical column and an Agilent Zorbax SB-C18, 1.8 micron (2.1 x 5 mm)
CH:CHROMATOGRAPHY_SUMMARY        	guard column. The autosampler tray temperature was set at 4°C, column
CH:CHROMATOGRAPHY_SUMMARY        	temperature was set at 60°C, and the sample injection volume was 4µL. The flow
CH:CHROMATOGRAPHY_SUMMARY        	rate was 0.7 mL/min with the following mobile phases: mobile phase A was water
CH:CHROMATOGRAPHY_SUMMARY        	with 0.1% formic acid, and mobile phase B was 60:36:4 isopropyl
CH:CHROMATOGRAPHY_SUMMARY        	alcohol:acetonitrile:water with 0.1% formic acid. Gradient elution was as
CH:CHROMATOGRAPHY_SUMMARY        	follows: 0-1 minute 30-70% B, 1-7.92 minutes 70-100% B, 7.92-10.4 minutes 100%
CH:CHROMATOGRAPHY_SUMMARY        	B, 10.4-10.5 minutes 100-30% B, followed by column re-equilibration with 30% B
CH:CHROMATOGRAPHY_SUMMARY        	from 10.5-15.1 minutes.
CH:CHROMATOGRAPHY_TYPE           	Reversed phase
CH:INSTRUMENT_NAME               	Agilent 1290
CH:COLUMN_NAME                   	Agilent Zorbax RRHD SB-C18 (100 x 2.1mm, 1.8um)
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Agilent 6210 TOF
MS:INSTRUMENT_TYPE               	TOF
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	POSITIVE
MS:MS_COMMENTS                   	The mass spectrometry conditions were as follows: Agilent 6210 Time-of-Flight
MS:MS_COMMENTS                   	mass spectrometer (TOF-MS) in positive ionization mode with dual electrospray
MS:MS_COMMENTS                   	(ESI) source, mass range 60-1600m/z, scan rate 2.03, gas temperature 300°C, gas
MS:MS_COMMENTS                   	flow 12.0L/min, nebulizer 30psi, skimmer 60V, capillary voltage 4000V,
MS:MS_COMMENTS                   	fragmentor 120V, reference masses 121.050873 and 922.009798 (Agilent reference
MS:MS_COMMENTS                   	mix). Proprietary analytical software for integration and peak picking
MS:MS_RESULTS_FILE               	ST001911_AN003108_Results.txt	UNITS:Abundance (mass spectral counts)	Has m/z:Neutral masses	Has RT:Yes	RT units:Minutes
#END