#METABOLOMICS WORKBENCH erinzhang_0220_20211027_143938 DATATRACK_ID:2910 STUDY_ID:ST001959 ANALYSIS_ID:AN003194
VERSION                          	1
CREATED_ON                       	03-03-2022
#PROJECT
PR:PROJECT_TITLE                 	Serum metabolomics study in intestinal AMPK KO mice
PR:PROJECT_TYPE                  	Serum untargeted metabolomics
PR:PROJECT_SUMMARY               	Serum untargeted metabolomics studies on serum samples from intestinal AMPK KO
PR:PROJECT_SUMMARY               	mice and control mice
PR:INSTITUTE                     	City of Hope National Medical Center
PR:DEPARTMENT                    	Diabetes Complications & Metabolism
PR:LAST_NAME                     	Zhang
PR:FIRST_NAME                    	Eryun
PR:ADDRESS                       	1500 Duarte Rd, Duarte, CA, 91010, USA
PR:EMAIL                         	erzhang@coh.org
PR:PHONE                         	6268735517
PR:DOI                           	http://dx.doi.org/10.21228/M8NM5W
#STUDY
ST:STUDY_TITLE                   	Investigation of serum metabolites in the AMPK intestinal KO mice
ST:STUDY_SUMMARY                 	Conducted serum untargeted metabolomics analysis in AMP-activated protein kinase
ST:STUDY_SUMMARY                 	(AMPK) intestinal KO mice and control mice under high-fat diet (HFD) conditions
ST:INSTITUTE                     	City of Hope National Medical Center
ST:DEPARTMENT                    	Diabetes Complications & Metabolism
ST:LAST_NAME                     	Zhang
ST:FIRST_NAME                    	Eryun
ST:ADDRESS                       	1500 Duarte Rd, Duarte, CA91010
ST:EMAIL                         	zhangey0220@126.com
ST:PHONE                         	16268735517
ST:SUBMIT_DATE                   	2021-10-27
#SUBJECT
SU:SUBJECT_TYPE                  	Mammal
SU:SUBJECT_SPECIES               	Mus musculus
SU:TAXONOMY_ID                   	10090
#SUBJECT_SAMPLE_FACTORS:         	SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data
SUBJECT_SAMPLE_FACTORS           	-	A328pos	Genotype:AMPKfl/fl(control) mice | Treatment:HFD	Sample_Data:RAW_FILE_NAME=A328pos.mzXML
SUBJECT_SAMPLE_FACTORS           	-	A329pos	Genotype:AMPKfl/fl(control) mice | Treatment:HFD	Sample_Data:RAW_FILE_NAME=A329pos.mzXML
SUBJECT_SAMPLE_FACTORS           	-	A330pos	Genotype:AMPKfl/fl(control) mice | Treatment:HFD	Sample_Data:RAW_FILE_NAME=A330pos.mzXML
SUBJECT_SAMPLE_FACTORS           	-	A331pos	Genotype:AMPKfl/fl(control) mice | Treatment:HFD	Sample_Data:RAW_FILE_NAME=A331pos.mzXML
SUBJECT_SAMPLE_FACTORS           	-	B332pos	Genotype:AMPK intestinal KO mice | Treatment:HFD	Sample_Data:RAW_FILE_NAME=B332pos.mzXML
SUBJECT_SAMPLE_FACTORS           	-	B333pos	Genotype:AMPK intestinal KO mice | Treatment:HFD	Sample_Data:RAW_FILE_NAME=B333pos.mzXML
SUBJECT_SAMPLE_FACTORS           	-	B334pos	Genotype:AMPK intestinal KO mice | Treatment:HFD	Sample_Data:RAW_FILE_NAME=B334pos.mzXML
SUBJECT_SAMPLE_FACTORS           	-	B335pos	Genotype:AMPK intestinal KO mice | Treatment:HFD	Sample_Data:RAW_FILE_NAME=B335pos.mzXML
#COLLECTION
CO:COLLECTION_SUMMARY            	Collect whole blood in a covered test tube,leave the tube in a standing position
CO:COLLECTION_SUMMARY            	and wait 30 min.Centrifuge 1500 g 10 min at 4ºC.Take out the serum.
CO:SAMPLE_TYPE                   	Blood (serum)
#TREATMENT
TR:TREATMENT_SUMMARY             	C57BL/6-AMPKa1flox/flox mice (#014141), and B6.Cg-Tg (Vil-cre) 1000Gum/J
TR:TREATMENT_SUMMARY             	(#021504) were purchased from Jackson Laboratory (Bar Harbor, ME). AMPK
TR:TREATMENT_SUMMARY             	intestinal KO (AMPK-IKO) mice were generated by cross-breeding
TR:TREATMENT_SUMMARY             	C57BL/6-AMPKa1flox/flox mice with B6.Cg-Tg (Vil-cre) 1000Gum/J. All animal
TR:TREATMENT_SUMMARY             	procedures were approved by the City of Hope Institutional Animal Care and Use
TR:TREATMENT_SUMMARY             	Committee and conducted in accordance with the National Institutes of Health
TR:TREATMENT_SUMMARY             	Guidelines for the Care and Use of Laboratory Animals. 6-week-old male mice were
TR:TREATMENT_SUMMARY             	housed in a temperature (22-23°C) and light-controlled vivarium with free
TR:TREATMENT_SUMMARY             	access to water and high-fat diet (HFD, 60% kcal fat; D12492, Research Diets,
TR:TREATMENT_SUMMARY             	New Brunswick, NJ) for 12 weeks.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	50 µL of serum was used for metabolites extraction. 200 μL extract solution
SP:SAMPLEPREP_SUMMARY            	(acetonitrile: methanol = 1: 1) containing isotopically labeled internal
SP:SAMPLEPREP_SUMMARY            	standard mixture was added to each sample. After vortexing for 1 min, the
SP:SAMPLEPREP_SUMMARY            	samples were sonicated for 10 min in cold water bath. Then, the samples were
SP:SAMPLEPREP_SUMMARY            	incubated at -40 °C for 1 h, and centrifuged at 12000 rpm for 15 min at 4°C.
SP:SAMPLEPREP_SUMMARY            	200 µL of supernatant were transferred to a fresh tube, and dried in a vacuum
SP:SAMPLEPREP_SUMMARY            	concentrator at 37 °C. Then, the dried samples were reconstituted in 200 µL of
SP:SAMPLEPREP_SUMMARY            	50% acetonitrile by sonication on ice for 10 min. The constitution was then
SP:SAMPLEPREP_SUMMARY            	centrifuged at 13000 rpm for 15 min at 4°C, and 75 µL of supernatant were
SP:SAMPLEPREP_SUMMARY            	transferred to a fresh glass vial for LC/MS analysis.
#CHROMATOGRAPHY
CH:INSTRUMENT_NAME               	Agilent 1290 Infinity
CH:COLUMN_NAME                   	Waters Acquity BEH Amide (150 x 2.1mm, 1.7um)
CH:CHROMATOGRAPHY_TYPE           	HILIC
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
AN:UNITS                         	Peak area
#MS
MS:INSTRUMENT_NAME               	ABI Sciex 5600 TripleTOF
MS:INSTRUMENT_TYPE               	QTOF
MS:MS_TYPE                       	ESI
MS:MS_COMMENTS                   	UHPLC-MS raw data files were converted into mzXML format using the
MS:MS_COMMENTS                   	“msconvert” program from Pro-teoWizard and then analyzed by the XCMS 67 and
MS:MS_COMMENTS                   	CAMERA toolbox 68 with R statistical software. The CentWave algorithm in XCMS
MS:MS_COMMENTS                   	was used for peak detection. The parameter “peak-width” was set as (5, 20)
MS:MS_COMMENTS                   	in units of seconds, referring to the minimum and maximum peak widths for peak
MS:MS_COMMENTS                   	detection. The pa-rameter ‘‘snthresh’’ is set as 3 for sensitive peak
MS:MS_COMMENTS                   	detection. For multiple UHPLC–MS data files, an or-dered bijective
MS:MS_COMMENTS                   	interpolated warping (OBI-Warp) algorithm in XCMS was used for peak alignment.
MS:ION_MODE                      	POSITIVE
MS:MS_RESULTS_FILE               	ST001959_AN003194_Results.txt	UNITS:Peak area	Has m/z:Yes	Has RT:Yes	RT units:Seconds
#END