#METABOLOMICS WORKBENCH dougguzior_20250326_082930 DATATRACK_ID:5784 STUDY_ID:ST004084 ANALYSIS_ID:AN006768 PROJECT_ID:PR002564
VERSION             	1
CREATED_ON             	April 4, 2025, 5:20 pm
#PROJECT
PR:PROJECT_TITLE                 	Dietary methionine restriction in high-fat diet-fed male mice
PR:PROJECT_SUMMARY               	25-week-old male mice (n=31) with diet-induced obesity were divided into four
PR:PROJECT_SUMMARY               	groups. The first group (n=8) was fed a high-fat diet (HFD, 60% kcal fat) for 15
PR:PROJECT_SUMMARY               	weeks. The second, third, and fourth groups were fed a modified high-fat diet
PR:PROJECT_SUMMARY               	(HFD-MetR, 60% fat/0.12% methionine) for 3 days (MetR3, n=8), 5 days (MetR5,
PR:PROJECT_SUMMARY               	n=8), or 10 days (MetR10, n=7), respectively. The HFD group (60% fat/0.86%
PR:PROJECT_SUMMARY               	methionine) served as the control. Mice were fasted for 4 hours prior to sample
PR:PROJECT_SUMMARY               	collection. The goal of this study was to investigate the effect of methionine
PR:PROJECT_SUMMARY               	restriction on plasma, brown adipose, and liver metabolomes.
PR:INSTITUTE                     	Panome Bio
PR:LAST_NAME                     	Guzior
PR:FIRST_NAME                    	Douglas
PR:ADDRESS                       	4340 Duncan Ave, St. Louis, MO 63110
PR:EMAIL                         	douglas.guzior@panomebio.com
PR:PHONE                         	(314) 632-6588
#STUDY
ST:STUDY_TITLE                   	Dietary methionine restriction in high-fat diet-fed male mice, liver
ST:STUDY_TITLE                   	metabolomics
ST:STUDY_SUMMARY                 	25-week-old male mice (n=31) with diet-induced obesity were divided into four
ST:STUDY_SUMMARY                 	groups. The first group (n=8) was fed a high-fat diet (HFD, 60% kcal fat) for 15
ST:STUDY_SUMMARY                 	weeks. The second, third, and fourth groups were fed a modified high-fat diet
ST:STUDY_SUMMARY                 	(HFD-MetR, 60% fat/0.12% methionine) for 3 days (MetR3, n=8), 5 days (MetR5,
ST:STUDY_SUMMARY                 	n=8), or 10 days (MetR10, n=7), respectively. The HFD group (60% fat/0.86%
ST:STUDY_SUMMARY                 	methionine) served as the control. Mice were fasted for 4 hours prior to sample
ST:STUDY_SUMMARY                 	collection. The goal of this study was to investigate the effect of methionine
ST:STUDY_SUMMARY                 	restriction on plasma, brown adipose, and liver metabolomes.
ST:INSTITUTE                     	Panome Bio
ST:LAST_NAME                     	Guzior
ST:FIRST_NAME                    	Douglas
ST:ADDRESS                       	4340 Duncan Ave, St. Louis, MO 63110
ST:EMAIL                         	douglas.guzior@panomebio.com
ST:PHONE                         	(314) 632-6588
ST:NUM_GROUPS                    	4
ST:TOTAL_SUBJECTS                	31
ST:NUM_MALES                     	31
ST:STUDY_COMMENTS                	Part 2 of 3
#SUBJECT
SU:SUBJECT_TYPE                  	Mammal
SU:SUBJECT_SPECIES               	Mus musculus
SU:TAXONOMY_ID                   	10090
SU:GENOTYPE_STRAIN               	C57BL/6J
SU:AGE_OR_AGE_RANGE              	25 wk
SU:WEIGHT_OR_WEIGHT_RANGE        	>45g bodyweight at 25 w/o
SU:GENDER                        	Male
SU:ANIMAL_LIGHT_CYCLE            	12-hour light/12-hour dark photoperiod (0700 – 1900)
SU:ANIMAL_FEED                   	Research Diets D12492; Research Diets A14032002; Research Diets A14032001
SU:ANIMAL_INCLUSION_CRITERIA     	>45g bodyweight at 25 w/o
#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           	-	LIV-CF-5	group:Control | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-015_039.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-CF-6	group:Control | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-016_042.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-CF-7	group:Control | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-017_028.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-CF-8	group:Control | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-018_029.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR3-13	group:MetR3 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-019_040.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR3-14	group:MetR3 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-020_035.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR3-15	group:MetR3 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-021_030.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR3-16	group:MetR3 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-022_032.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR5-21	group:MetR5 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-023_038.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR5-22	group:MetR5 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-024_037.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR5-23	group:MetR5 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-025_031.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR5-24	group:MetR5 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-026_036.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR10-29	group:MetR10 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-027_041.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR10-30	group:MetR10 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-028_034.mzML
SUBJECT_SAMPLE_FACTORS           	-	LIV-MR10-31	group:MetR10 | Sample source:Liver	RAW_FILE_NAME(filename)=060-0116-029_033.mzML
SUBJECT_SAMPLE_FACTORS           	-	blank1	group:blank | Sample source:blank	RAW_FILE_NAME(filename)=060-0116-Blank_4_049.mzML
SUBJECT_SAMPLE_FACTORS           	-	qc1	group:qc | Sample source:pooled_qc	RAW_FILE_NAME(filename)=060-0116-L-QC_1_026.mzML
SUBJECT_SAMPLE_FACTORS           	-	qc2	group:qc | Sample source:pooled_qc	RAW_FILE_NAME(filename)=060-0116-L-QC_2_027.mzML
SUBJECT_SAMPLE_FACTORS           	-	qc3	group:qc | Sample source:pooled_qc	RAW_FILE_NAME(filename)=060-0116-L-QC_3_043.mzML
#COLLECTION
CO:COLLECTION_SUMMARY            	Liver samples were collected upon completion of the time course after fasting
CO:COLLECTION_SUMMARY            	the mice for 4 h.
CO:SAMPLE_TYPE                   	Liver
CO:STORAGE_CONDITIONS            	-80℃
#TREATMENT
TR:TREATMENT_SUMMARY             	Mice were weight-matched and divided into two groups fed either HFD-control (60%
TR:TREATMENT_SUMMARY             	fat/0.86% methionine; Research Diets A14032002) or HFD-MetR (60% fat/0.12%
TR:TREATMENT_SUMMARY             	methionine; Research Diets A14032001). The HFD-MetR group was split into 3
TR:TREATMENT_SUMMARY             	groups, where the HFD-MetR was administered for 3 days (MetR3, n=8), 5 days
TR:TREATMENT_SUMMARY             	(MetR5, n=8), or 10 days (MetR10, n=7). HFD-MetR groups were compared to each
TR:TREATMENT_SUMMARY             	other and the control group (HFD, n=8).
TR:TREATMENT                     	Dietary intervention and duration of altered diet
TR:TREATMENT_ROUTE               	Diet
TR:TREATMENT_DOSE                	Research Diets A14032002; Research Diets A14032001
TR:TREATMENT_DOSEVOLUME          	HFD-control (60% fat/0.86% methionine; Research Diets A14032002) or HFD-MetR
TR:TREATMENT_DOSEVOLUME          	(60% fat/0.12% methionine; Research Diets A14032001)
TR:TREATMENT_DOSEDURATION        	3-10 days
TR:TREATMENT_VEHICLE             	Food
TR:ANIMAL_FASTING                	4h prior to sample collection
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	Frozen tissue samples were homogenized with 80 μL 2:2:1
SP:SAMPLEPREP_SUMMARY            	methanol:acetonitrile:water per mg tissue, centrifuged, and the supernatant was
SP:SAMPLEPREP_SUMMARY            	collected after a cold incubation and wash steps, then stored at -80°C. Plasma
SP:SAMPLEPREP_SUMMARY            	samples underwent solid-phase extraction (SPE) with sequential additions of
SP:SAMPLEPREP_SUMMARY            	acetonitrile:methanol and methanol:acetonitrile:water to separate polar and
SP:SAMPLEPREP_SUMMARY            	non-polar metabolites; polar metabolites were eluted and stored at -80°C, while
SP:SAMPLEPREP_SUMMARY            	non-polar metabolites were eluted using methyl tert-butyl ether:methanol, dried
SP:SAMPLEPREP_SUMMARY            	under nitrogen, reconstituted in isopropanol:methanol, and stored at -80°C.
SP:PROCESSING_STORAGE_CONDITIONS 	-80℃
SP:EXTRACT_STORAGE               	-80℃
SP:SAMPLE_RESUSPENSION           	2:2:1 methanol:acetonitrile:water (polar), isopropanol:methanol (lipid)
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_TYPE           	Reversed phase
CH:INSTRUMENT_NAME               	Agilent 1290 Infinity II
CH:COLUMN_NAME                   	Waters ACQUITY UPLC HSS T3 (100 x 2.1mm,1.8um)
CH:SOLVENT_A                     	50% water/30% acetonitrile/20% isopropanol; 10 mM ammonium formate; 5 µM
CH:SOLVENT_A                     	InfinityLab Deactivator Additive (Agilent)
CH:SOLVENT_B                     	90% isopropanol/10% acetonitrile; 10 mM ammonium formate; 0.1% formic acid; 2.5
CH:SOLVENT_B                     	mM medronic acid
CH:FLOW_GRADIENT                 	0-2 min: 30% B, 2-17 min: 30-75% B, 17-20 min: 75-85% B, 20-23 min: 85-100% B,
CH:FLOW_GRADIENT                 	23-26 min: 100% B, 26-27 min: 100-30% B
CH:FLOW_RATE                     	400 µL/min
CH:COLUMN_TEMPERATURE            	45
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Agilent 6546 QTOF
MS:INSTRUMENT_TYPE               	QTOF
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	NEGATIVE
MS:MS_COMMENTS                   	MS/MS data were acquired in a data-dependent iterative fashion with a 1.3 m/z
MS:MS_COMMENTS                   	isolation window. Features were detected in LC/MS data through in-house peak
MS:MS_COMMENTS                   	detection and curation software.
MS:MS_RESULTS_FILE               	ST004084_AN006768_Results.txt	UNITS:log2-transformed normalized relative abundance	Has m/z:Yes	Has RT:Yes	RT units:Minutes
#END