#METABOLOMICS WORKBENCH juliehaines_20211122_115205 DATATRACK_ID:2938 STUDY_ID:ST001997 ANALYSIS_ID:AN003258 PROJECT_ID:000000
VERSION             	1
CREATED_ON             	November 22, 2021, 12:06 pm
#PROJECT
PR:PROJECT_TITLE                 	Polyamine import and accumulation causes immunomodulation in macrophages
PR:PROJECT_TITLE                 	engulfing apoptotic cells
PR:PROJECT_SUMMARY               	Phagocytosis of apoptotic cells, termed efferocytosis, is critical for tissue
PR:PROJECT_SUMMARY               	homeostasis and drives anti-inflammatory programming in engulfing macrophages.
PR:PROJECT_SUMMARY               	Here, we assess metabolites in naïve and inflammatory macrophages following
PR:PROJECT_SUMMARY               	engulfment of multiple cellular and non-cellular targets. Efferocytosis leads to
PR:PROJECT_SUMMARY               	unique increases in the arginine-derived polyamines, spermidine and spermine, in
PR:PROJECT_SUMMARY               	vitro and in vivo. Surprisingly, polyamine accumulation after efferocytosis does
PR:PROJECT_SUMMARY               	not arise from retention of apoptotic cell metabolites or de novo synthesis, but
PR:PROJECT_SUMMARY               	from enhanced polyamine import that is dependent on Rac1, actin, and PI3 kinase.
PR:PROJECT_SUMMARY               	Blocking polyamine import prevents efferocytosis from suppressing macrophage
PR:PROJECT_SUMMARY               	IL-1or IL-6. This identifies efferocytosis as a trigger for polyamine
PR:PROJECT_SUMMARY               	import and accumulation, and imported polyamines as mediators of
PR:PROJECT_SUMMARY               	efferocytosis-induced immune reprogramming.
PR:INSTITUTE                     	University of Colorado Denver
PR:LAST_NAME                     	Haines
PR:FIRST_NAME                    	Julie
PR:ADDRESS                       	12801 E 17th Ave, Room 1303, Aurora, Colorado, 80045, USA
PR:EMAIL                         	julie.haines@cuanschutz.edu
PR:PHONE                         	3037243339
#STUDY
ST:STUDY_TITLE                   	Polyamine import and accumulation causes immunomodulation in macrophages
ST:STUDY_TITLE                   	engulfing apoptotic cells (Part 2)
ST:STUDY_SUMMARY                 	Phagocytosis of apoptotic cells, termed efferocytosis, is critical for tissue
ST:STUDY_SUMMARY                 	homeostasis and drives anti-inflammatory programming in engulfing macrophages.
ST:STUDY_SUMMARY                 	Here, we assess metabolites in naïve and inflammatory macrophages following
ST:STUDY_SUMMARY                 	engulfment of multiple cellular and non-cellular targets. Efferocytosis leads to
ST:STUDY_SUMMARY                 	unique increases in the arginine-derived polyamines, spermidine and spermine, in
ST:STUDY_SUMMARY                 	vitro and in vivo. Surprisingly, polyamine accumulation after efferocytosis does
ST:STUDY_SUMMARY                 	not arise from retention of apoptotic cell metabolites or de novo synthesis, but
ST:STUDY_SUMMARY                 	from enhanced polyamine import that is dependent on Rac1, actin, and PI3 kinase.
ST:STUDY_SUMMARY                 	Blocking polyamine import prevents efferocytosis from suppressing macrophage
ST:STUDY_SUMMARY                 	IL-1or IL-6. This identifies efferocytosis as a trigger for polyamine
ST:STUDY_SUMMARY                 	import and accumulation, and imported polyamines as mediators of
ST:STUDY_SUMMARY                 	efferocytosis-induced immune reprogramming.
ST:INSTITUTE                     	University of Colorado Denver
ST:LAST_NAME                     	Haines
ST:FIRST_NAME                    	Julie
ST:ADDRESS                       	12801 E 17th Ave, Room 1303, Aurora, Colorado, 80045, USA
ST:EMAIL                         	julie.haines@cuanschutz.edu
ST:PHONE                         	3037243339
#SUBJECT
SU:SUBJECT_TYPE                  	Cultured cells
SU:SUBJECT_SPECIES               	Mus musculus
SU:TAXONOMY_ID                   	10090
#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           	-	4h 1	category:4h unexposed	RAW_FILE_NAME=37
SUBJECT_SAMPLE_FACTORS           	-	4h 2	category:4h unexposed	RAW_FILE_NAME=40
SUBJECT_SAMPLE_FACTORS           	-	4h 3	category:4h unexposed	RAW_FILE_NAME=43
SUBJECT_SAMPLE_FACTORS           	-	4h 1-	category:4h non-engulfing	RAW_FILE_NAME=38
SUBJECT_SAMPLE_FACTORS           	-	4h 2-	category:4h non-engulfing	RAW_FILE_NAME=41
SUBJECT_SAMPLE_FACTORS           	-	4h 3-	category:4h non-engulfing	RAW_FILE_NAME=44
SUBJECT_SAMPLE_FACTORS           	-	4h 1+	category:4h engulfing	RAW_FILE_NAME=39
SUBJECT_SAMPLE_FACTORS           	-	4h 2+	category:4h engulfing	RAW_FILE_NAME=42
SUBJECT_SAMPLE_FACTORS           	-	4h 3+	category:4h engulfing	RAW_FILE_NAME=45
SUBJECT_SAMPLE_FACTORS           	-	8h 1	category:8h unexposed	RAW_FILE_NAME=46
SUBJECT_SAMPLE_FACTORS           	-	8h 2	category:8h unexposed	RAW_FILE_NAME=49
SUBJECT_SAMPLE_FACTORS           	-	8h 3	category:8h unexposed	RAW_FILE_NAME=52
SUBJECT_SAMPLE_FACTORS           	-	8h 1-	category:8h non-engulfing	RAW_FILE_NAME=47
SUBJECT_SAMPLE_FACTORS           	-	8h 2-	category:8h non-engulfing	RAW_FILE_NAME=50
SUBJECT_SAMPLE_FACTORS           	-	8h 3-	category:8h non-engulfing	RAW_FILE_NAME=53
SUBJECT_SAMPLE_FACTORS           	-	8h 1+	category:8h engulfing	RAW_FILE_NAME=48
SUBJECT_SAMPLE_FACTORS           	-	8h 2+	category:8h engulfing	RAW_FILE_NAME=51
SUBJECT_SAMPLE_FACTORS           	-	8h 3+	category:8h engulfing	RAW_FILE_NAME=54
#COLLECTION
CO:COLLECTION_SUMMARY            	Experimental animals This study was approved and performed in accordance with
CO:COLLECTION_SUMMARY            	the ethical guidelines of the Institutional Animal Care and Use Committee at
CO:COLLECTION_SUMMARY            	National Jewish Health. C57BL/6J (000664), B6-CD45.1 (002014) and CCR2KO
CO:COLLECTION_SUMMARY            	(004999) mice were obtained from Jackson Laboratories (Bar Harbor, ME, USA).
CO:COLLECTION_SUMMARY            	DsRed.T3 mice (available as 006051 at Jackson Labs) were generously shared by
CO:COLLECTION_SUMMARY            	the Nagy laboratory (Vintersten et al., 2004). CCR2KO mice were crossed with
CO:COLLECTION_SUMMARY            	DsRed mice until double homozygous. Mice used in experiments were between 8-16
CO:COLLECTION_SUMMARY            	weeks of age. Both male and female sex were used in experiments except bone
CO:COLLECTION_SUMMARY            	marrow chimera experiments, which used only male mice. Human tissue This study
CO:COLLECTION_SUMMARY            	used deidentified donor peripheral blood mononuclear cells (PBMC) collected and
CO:COLLECTION_SUMMARY            	isolated (Young et al., 2011) by the National Jewish Blood Prep Core (IRB No.
CO:COLLECTION_SUMMARY            	HS-1285). Age and sex of deidentified blood donors in this study is not known.
CO:COLLECTION_SUMMARY            	Cell Lines Jurkat (human T lymphocytes, clone E6-1) were obtained from ATCC and
CO:COLLECTION_SUMMARY            	cultured in RPMI-1640 (GIBCO) supplemented with 10% heat-inactivated fetal
CO:COLLECTION_SUMMARY            	bovine serum (v/v) (GeminiBio), 1X Pen/Strep/Glut, 1mM HEPES, and 100M Sodium
CO:COLLECTION_SUMMARY            	Pyruvate. Cells were cultured in a humidified CO2 incubator at 37C. The Jurkat
CO:COLLECTION_SUMMARY            	cell line was established from peripheral blood of a 14-year-old male diagnosed
CO:COLLECTION_SUMMARY            	with T cell leukemia. Murine primary cell cultures Peritoneal cells were
CO:COLLECTION_SUMMARY            	isolated from naïve mice by lavage with 10mL of PBS containing 0.5mM EDTA.
CO:COLLECTION_SUMMARY            	Lavage was estimated to contain 50% macrophages. Peritoneal macrophages were
CO:COLLECTION_SUMMARY            	isolated by adhesion purification of lavage cells in RPMI-10 (RPMI-1640
CO:COLLECTION_SUMMARY            	containing 10% heat-inactivated fetal bovine serum (v/v), 1X Pen/Strep/Glut, 1mM
CO:COLLECTION_SUMMARY            	HEPES, and 100M Sodium Pyruvate) plated 1.3x106 macrophages/well in 6-well
CO:COLLECTION_SUMMARY            	plates for FACS sorting or 4x106 macrophage/well in 24-well plates for flow
CO:COLLECTION_SUMMARY            	cytometry. Non-adherent cells were removed by washing after 3-5 hours, and
CO:COLLECTION_SUMMARY            	macrophages were cultured in fresh RPMI-10 overnight. 400ng/mL purified E. coli
CO:COLLECTION_SUMMARY            	O111:B4 LPS (List Biological Laboratories Inc.) was added to generate
CO:COLLECTION_SUMMARY            	inflammatory macrophages 12h before the addition of target cells. Bone
CO:COLLECTION_SUMMARY            	marrow-derived macrophages were generated by 8-day culture of murine bone marrow
CO:COLLECTION_SUMMARY            	cells grown in media containing M-CSF (High glucose DMEM containing 10% v/v FBS,
CO:COLLECTION_SUMMARY            	20% v/v L929-conditioned media, 1X Pen/Strep/Glut, 100M Sodium Pyruvate,
CO:COLLECTION_SUMMARY            	55M beta-Mercaptoethanol). All macrophages were removed from culture dishes
CO:COLLECTION_SUMMARY            	by 3-minute incubation with 1X Trypsin-EDTA (Sigma) plus gentle scraping.
CO:SAMPLE_TYPE                   	Macrophages
#TREATMENT
TR:TREATMENT_SUMMARY             	LPS-primed murine peritoneal macrophages were fed fluorescently-tagged
TR:TREATMENT_SUMMARY             	IgG-opsonized live Jurkat cells for 1h. Unengulfed targets were washed off and
TR:TREATMENT_SUMMARY             	macrophages were left to degrade targets for a further 3 or 7 hours. Macrophages
TR:TREATMENT_SUMMARY             	were then collected and sorted into non-engulfing or engulfing populations,
TR:TREATMENT_SUMMARY             	alongside control unexposed macrophages. Sorted cells were pelleted at 400xg and
TR:TREATMENT_SUMMARY             	dry pellets were snap frozen and stored at -80C until metabolite extraction.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	To process cells for assessment of intracellular metabolites, cells were
SP:SAMPLEPREP_SUMMARY            	pelleted at 400xg in tubes coated with 0.06% BSA. Supernatant was aspirated and
SP:SAMPLEPREP_SUMMARY            	discarded; residual liquid was carefully wicked away from the pellet with a
SP:SAMPLEPREP_SUMMARY            	kimwipe. Dry pellets were immediately snap frozen and stored at -80C until
SP:SAMPLEPREP_SUMMARY            	processing. To process culture supernatants for assessment of metabolites,
SP:SAMPLEPREP_SUMMARY            	supernatant was centrifuged at 400xg to pellet any cells. Cell-free supernatant
SP:SAMPLEPREP_SUMMARY            	was then transferred to a fresh tube, snap frozen, and stored at -80C until
SP:SAMPLEPREP_SUMMARY            	processing.
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_SUMMARY        	Isocratic flow of 100% phase A (95% water, 5% acetonitrile, 10 mM ammonium
CH:CHROMATOGRAPHY_SUMMARY        	acetate) at 250 uL/min.
CH:CHROMATOGRAPHY_TYPE           	Reversed phase
CH:INSTRUMENT_NAME               	Thermo Vanquish
CH:COLUMN_NAME                   	Phenomenex Kinetex C18 (150 x 2.1mm, 2.6 um)
CH:FLOW_GRADIENT                 	isocratic
CH:FLOW_RATE                     	250 ul/min
CH:COLUMN_TEMPERATURE            	25
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Thermo Q Exactive Orbitrap
MS:INSTRUMENT_TYPE               	Orbitrap
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	NEGATIVE
MS:MS_COMMENTS                   	Global metabolomics analyses were performed using a 3 min isocratic run in
MS:MS_COMMENTS                   	positive and negative ion modes (separate runs) as described previously (Nemkov
MS:MS_COMMENTS                   	et al., 2015, Nemkov et al., 2017); stable isotope tracing samples were analyzed
MS:MS_COMMENTS                   	using a 5 min C18 gradient in positive and negative ion modes (separate runs) as
MS:MS_COMMENTS                   	described (Nemkov et al., 2019, Gehrke et al., 2019). For all analyses, the MS
MS:MS_COMMENTS                   	scanned in MS1 mode across the m/z range of 65 to 950. Peaks were annotated (in
MS:MS_COMMENTS                   	conjunction with the KEGG database), integrated, and quality control performed
MS:MS_COMMENTS                   	using Maven (Princeton University) as described. Stable isotope tracing results
MS:MS_COMMENTS                   	were isotopically corrected for the natural abundance of 13C1, 13C2, 15N1, and
MS:MS_COMMENTS                   	15N2 (Nemkov et al., 2017).
#MS_METABOLITE_DATA
MS_METABOLITE_DATA:UNITS	peak area
MS_METABOLITE_DATA_START
Samples	4h 1	4h 2	4h 3	4h 1-	4h 2-	4h 3-	4h 1+	4h 2+	4h 3+	8h 1	8h 2	8h 3	8h 1-	8h 2-	8h 3-	8h 1+	8h 2+	8h 3+
Factors	category:4h unexposed	category:4h unexposed	category:4h unexposed	category:4h non-engulfing	category:4h non-engulfing	category:4h non-engulfing	category:4h engulfing	category:4h engulfing	category:4h engulfing	category:8h unexposed	category:8h unexposed	category:8h unexposed	category:8h non-engulfing	category:8h non-engulfing	category:8h non-engulfing	category:8h engulfing	category:8h engulfing	category:8h engulfing
ATP	1.46E+04	1.37E+05	6.61E+04	4.26E+04	5.42E+04	1.00E+05	2.49E+05	1.23E+05	2.14E+04	1.17E+04	1.10E+05	2.84E+05	2.06E+05	1.87E+05	1.94E+05	4.83E+05	3.20E+04	2.50E+05
UTP	4.37E+04	9.11E+04	4.92E+04	1.46E+04	6.39E+04	9.77E+04	9.00E+04	7.77E+04	4.51E+04	4.07E+04	6.24E+04	9.99E+04	6.55E+04	6.71E+04	7.48E+04	1.10E+05	5.74E+04	9.72E+04
UDP	5.48E+04	2.79E+05	1.97E+05	6.44E+04	2.79E+05	2.21E+05	2.35E+05	2.68E+05	6.71E+04	5.46E+04	1.85E+05	2.27E+05	2.51E+05	1.62E+05	2.21E+05	2.55E+05	5.47E+04	1.76E+05
IDP	1.95E+06	3.09E+06	3.08E+06	3.44E+06	2.87E+06	2.26E+06	2.42E+06	3.21E+06	2.67E+06	2.34E+06	2.62E+06	2.31E+06	2.40E+06	2.58E+06	2.60E+06	2.43E+06	2.56E+06	2.40E+06
Pyridoxal	6.95E+04	7.53E+04	5.61E+04	5.87E+04	8.11E+04	6.39E+04	7.31E+04	7.10E+04	6.94E+04	6.41E+04	9.29E+04	7.68E+04	7.99E+04	8.69E+04	7.62E+04	9.88E+04	6.39E+04	7.87E+04
UDP-glucose	4.87E+04	3.37E+04	3.19E+04	3.78E+04	2.69E+04	5.54E+04	5.93E+04	5.30E+04	4.14E+04	4.41E+04	5.37E+04	4.86E+04	5.26E+04	6.10E+04	5.32E+04	6.86E+04	7.38E+04	7.14E+04
Phosphate	9.53E+07	1.10E+08	1.09E+08	1.06E+08	1.12E+08	1.08E+08	8.97E+07	8.38E+07	1.02E+08	1.00E+08	9.23E+07	1.11E+08	1.06E+08	1.06E+08	1.13E+08	9.98E+07	1.39E+08	1.12E+08
Diphosphate	6.24E+06	7.36E+06	7.27E+06	6.54E+06	6.21E+06	7.21E+06	5.08E+06	3.91E+06	7.52E+06	6.52E+06	6.39E+06	7.50E+06	6.98E+06	7.41E+06	8.00E+06	6.25E+06	1.32E+07	8.40E+06
D-Glucose	1.38E+07	1.76E+07	1.40E+07	1.49E+07	1.50E+07	1.28E+07	1.67E+07	1.76E+07	1.23E+07	1.17E+07	1.23E+07	1.45E+07	1.48E+07	1.25E+07	1.43E+07	1.74E+07	1.06E+07	1.22E+07
D-Glucose 6-phosphate	4.15E+05	4.30E+05	3.95E+05	4.72E+05	3.68E+05	5.92E+05	6.53E+05	5.01E+05	6.92E+05	3.26E+05	3.89E+05	3.90E+05	3.59E+05	3.59E+05	4.08E+05	4.68E+05	3.81E+05	5.53E+05
D-Fructose 1-6-bisphosphate	8.59E+05	1.56E+06	1.04E+06	6.75E+05	7.81E+05	1.32E+06	1.72E+06	1.48E+06	1.48E+06	6.40E+05	8.15E+05	8.60E+05	6.92E+05	8.34E+05	9.11E+05	8.36E+05	9.94E+05	1.20E+06
D-Glyceraldehyde 3-phosphate/Glycerone phosphate	9.57E+06	1.19E+06	3.35E+06	2.22E+06	2.54E+06	4.80E+06	7.98E+06	9.07E+06	2.91E+06	4.04E+06	4.37E+06	1.13E+06	2.82E+06	2.92E+06	6.21E+06	2.02E+06	3.33E+06	4.92E+06
2/3-Phospho-D-glycerate	3.18E+05	4.14E+05	3.07E+05	3.55E+05	3.28E+05	4.55E+05	3.84E+05	3.33E+05	3.58E+05	2.95E+05	3.46E+05	4.23E+05	4.08E+05	3.79E+05	4.36E+05	3.79E+05	3.87E+05	4.46E+05
Phosphoenolpyruvate	7.66E+04	1.45E+05	7.80E+04	9.82E+04	1.24E+05	1.79E+05	1.35E+05	7.48E+04	5.68E+04	8.52E+04	2.14E+05	9.52E+04	9.22E+04	9.08E+04	9.59E+04	1.13E+05	6.67E+04	7.78E+04
Pyruvate	5.73E+05	7.40E+05	5.72E+05	5.83E+05	6.46E+05	4.76E+05	7.05E+05	8.20E+05	6.11E+05	5.64E+05	5.70E+05	5.69E+05	6.95E+05	5.42E+05	7.08E+05	7.24E+05	3.76E+05	5.54E+05
Lactate	5.03E+06	4.64E+06	4.01E+06	4.87E+06	4.79E+06	4.08E+06	5.36E+06	5.92E+06	4.61E+06	3.86E+06	4.67E+06	4.69E+06	5.38E+06	4.30E+06	5.94E+06	6.33E+06	3.57E+06	5.03E+06
D-Ribose	3.55E+05	3.63E+05	3.83E+05	3.98E+05	3.59E+05	3.17E+05	3.96E+05	3.83E+05	2.76E+05	2.85E+05	3.46E+05	3.42E+05	3.30E+05	3.21E+05	3.38E+05	4.18E+05	2.80E+05	3.02E+05
D-Rhamnose	1.38E+06	1.51E+06	9.77E+05	1.42E+06	1.39E+06	1.39E+06	1.52E+06	9.70E+05	1.42E+06	1.07E+06	1.21E+06	1.37E+06	9.99E+05	1.18E+06	1.31E+06	1.35E+06	9.80E+05	1.20E+06
Citrate	3.46E+06	4.52E+06	3.74E+06	4.20E+06	3.64E+06	5.94E+06	7.22E+06	4.31E+06	5.68E+06	3.59E+06	3.95E+06	5.94E+06	5.24E+06	5.13E+06	6.99E+06	8.19E+06	6.10E+06	8.81E+06
2-Oxoglutarate	5.94E+04	7.45E+04	4.97E+04	5.24E+04	5.99E+04	6.89E+04	8.65E+04	8.72E+04	6.12E+04	8.52E+04	8.07E+04	5.68E+04	8.61E+04	6.90E+04	7.48E+04	1.13E+05	8.88E+04	6.52E+04
Succinate	4.86E+05	5.87E+05	5.20E+05	7.58E+05	5.18E+05	5.18E+05	6.31E+05	6.51E+05	5.14E+05	5.27E+05	6.51E+05	5.21E+05	8.44E+05	6.81E+05	5.96E+05	1.06E+06	7.58E+05	6.37E+05
Fumarate	7.61E+04	8.16E+04	5.05E+04	5.41E+04	6.59E+04	8.58E+04	1.27E+05	8.40E+04	1.28E+05	6.09E+04	6.42E+04	5.89E+04	7.18E+04	8.50E+04	9.50E+04	1.16E+05	1.15E+05	1.29E+05
Malate	7.44E+05	1.09E+06	7.85E+05	6.10E+05	6.40E+05	1.13E+06	1.44E+06	9.91E+05	1.85E+06	7.47E+05	9.27E+05	8.20E+05	9.01E+05	9.35E+05	1.34E+06	1.37E+06	1.52E+06	1.72E+06
Itaconate	4.72E+06	4.93E+06	3.44E+06	4.23E+06	3.56E+06	4.68E+06	1.00E+07	6.32E+06	5.74E+06	6.21E+06	5.87E+06	4.66E+06	7.97E+06	7.58E+06	5.51E+06	1.27E+07	1.07E+07	7.29E+06
2-Hydroxyglutarate/Citramalate	4.47E+05	5.09E+05	4.26E+05	4.46E+05	3.63E+05	6.08E+05	5.23E+05	3.44E+05	5.38E+05	4.07E+05	5.20E+05	4.32E+05	5.64E+05	5.46E+05	4.40E+05	6.26E+05	6.63E+05	5.05E+05
Sedoheptulose 1-phosphate	6.71E+04	8.51E+04	5.12E+04	5.88E+04	4.84E+04	9.16E+04	9.21E+04	7.73E+04	1.04E+05	4.34E+04	5.11E+04	4.94E+04	4.55E+04	7.17E+04	2.57E+04	1.00E+05	4.91E+04	9.69E+04
alpha-D-Ribose 1-phosphate	1.73E+05	1.98E+05	2.11E+05	1.86E+05	2.11E+05	1.49E+05	2.08E+05	1.94E+05	1.65E+05	1.40E+05	1.26E+05	2.08E+05	2.67E+05	9.82E+04	2.11E+05	2.38E+05	1.37E+05	1.69E+05
Glutathione	1.99E+05	2.35E+05	1.15E+05	1.32E+05	1.51E+05	3.82E+05	6.03E+05	1.42E+05	2.10E+05	3.66E+05	4.41E+05	4.56E+05	5.32E+05	3.88E+05	7.74E+05	9.09E+05	4.50E+05	5.93E+05
Ascorbate	1.90E+05	2.84E+05	2.17E+05	2.34E+05	2.69E+05	1.87E+05	2.59E+05	2.83E+05	1.79E+05	1.74E+05	1.31E+05	1.99E+05	2.28E+05	1.48E+05	2.20E+05	2.50E+05	1.50E+05	1.65E+05
Dehydroascorbate	6.93E+04	7.20E+04	6.98E+04	6.90E+04	5.77E+04	7.42E+04	1.26E+05	8.01E+04	7.54E+04	6.68E+04	9.45E+04	8.84E+04	7.34E+04	9.00E+04	1.13E+05	1.46E+05	1.02E+05	1.30E+05
L-gamma-Glutamyl-L-hypoglycin	3.61E+04	7.03E+04	2.00E+04	3.52E+04	4.62E+04	2.67E+04	6.21E+04	2.77E+04	5.74E+04	3.18E+04	3.84E+04	4.99E+04	4.68E+04	3.40E+04	7.73E+04	4.93E+04	4.03E+04	2.99E+04
1-4-beta-D-Xylan	3.82E+05	4.83E+05	3.81E+05	4.59E+05	3.90E+05	3.57E+05	5.59E+05	5.21E+05	4.17E+05	3.70E+05	3.65E+05	3.84E+05	4.19E+05	3.87E+05	3.77E+05	5.49E+05	2.96E+05	3.71E+05
UDP-N-acetyl-D-glucosamine	5.03E+04	6.17E+04	6.43E+04	6.79E+04	4.75E+04	7.72E+04	8.87E+04	6.20E+04	7.94E+04	9.09E+04	8.69E+04	6.88E+04	7.64E+04	8.47E+04	7.42E+04	9.36E+04	9.16E+04	6.78E+04
Glycolate	5.39E+06	1.64E+06	4.14E+06	6.30E+06	1.39E+06	2.86E+06	1.99E+06	1.79E+06	5.43E+06	5.91E+06	5.06E+06	2.10E+06	2.18E+06	3.84E+06	1.89E+06	2.27E+06	5.89E+06	2.97E+06
Phosphocreatine	1.19E+05	8.86E+04	1.10E+05	1.09E+05	7.17E+04	1.24E+05	1.40E+05	6.05E+04	5.71E+04	2.10E+05	2.53E+05	2.78E+05	2.66E+05	2.77E+05	2.63E+05	3.70E+05	2.02E+05	2.38E+05
Taurine	7.50E+05	6.20E+05	4.38E+05	5.49E+05	5.17E+05	1.24E+06	9.78E+05	4.28E+05	5.89E+05	1.12E+06	1.34E+06	1.38E+06	1.52E+06	1.47E+06	1.69E+06	1.41E+06	8.96E+05	1.14E+06
L-Cysteate	2.42E+05	9.51E+04	1.25E+05	2.73E+05	9.40E+04	1.69E+05	9.26E+04	1.01E+05	2.02E+05	2.51E+05	1.78E+05	1.25E+05	1.05E+05	1.78E+05	1.22E+05	1.11E+05	2.35E+05	1.40E+05
Indole-3-acetaldehyde	1.67E+04	2.43E+04	1.86E+04	1.70E+04	2.11E+04	2.99E+04	5.73E+04	8.18E+04	6.40E+04	1.30E+04	3.11E+04	2.09E+04	2.47E+04	1.14E+04	2.65E+04	7.72E+04	6.74E+04	6.35E+04
Anthranilate	7.58E+04	7.05E+04	5.27E+04	5.06E+04	4.97E+04	6.88E+04	7.49E+04	7.62E+04	6.41E+04	5.97E+04	6.57E+04	1.02E+05	7.54E+04	7.74E+04	8.70E+04	7.52E+04	6.52E+04	1.30E+05
Glycerol 3-phosphate	4.76E+04	8.30E+04	8.84E+04	6.27E+04	5.56E+04	1.01E+05	8.48E+04	1.04E+05	1.49E+05	7.73E+04	5.17E+04	7.29E+04	6.71E+04	6.11E+04	7.80E+04	9.24E+04	8.14E+04	9.38E+04
Ethanolamine phosphate	9.99E+05	9.74E+05	8.90E+05	6.98E+05	7.28E+05	1.63E+06	1.17E+06	6.49E+05	1.01E+06	1.21E+06	1.21E+06	1.32E+06	1.23E+06	1.46E+06	1.48E+06	1.56E+06	1.28E+06	1.44E+06
Sphinganine 1-phosphate	3.46E+05	3.81E+05	3.81E+05	3.97E+05	4.24E+05	4.36E+05	4.97E+05	3.60E+05	3.81E+05	3.61E+05	5.42E+05	3.36E+05	3.53E+05	4.52E+05	4.36E+05	3.96E+05	3.58E+05	3.12E+05
Acetyl phosphate	7.91E+04	8.47E+04	8.84E+04	6.79E+04	8.76E+04	9.33E+04	9.29E+04	8.56E+04	6.65E+04	7.14E+04	6.50E+04	8.87E+04	7.14E+04	7.33E+04	8.42E+04	7.21E+04	7.73E+04	9.69E+04
2-Methyleneglutarate	9.37E+05	1.10E+06	7.80E+05	1.01E+06	9.10E+05	7.98E+05	1.02E+06	1.06E+06	8.34E+05	8.11E+05	7.16E+05	9.67E+05	9.20E+05	7.84E+05	8.70E+05	1.15E+06	6.49E+05	7.92E+05
6-Pyruvoyltetrahydropterin	2.79E+04	4.85E+04	3.57E+04	1.71E+04	3.93E+04	4.60E+04	2.70E+04	3.48E+04	2.44E+04	1.60E+04	4.91E+04	1.08E+04	5.19E+04	3.73E+04	2.80E+04	2.60E+04	2.34E+04	2.71E+04
D-glucono-1,5-lactone	4.31E+05	7.63E+05	5.66E+05	5.81E+05	5.95E+05	3.98E+05	4.61E+05	8.47E+05	4.06E+05	3.10E+05	3.34E+05	5.06E+05	4.95E+05	3.22E+05	5.62E+05	5.31E+05	3.23E+05	4.20E+05
Glucosinolate	1.30E+06	9.05E+05	1.02E+06	1.27E+06	8.47E+05	1.21E+06	1.11E+06	9.49E+05	1.19E+06	1.09E+06	1.15E+06	1.21E+06	1.12E+06	1.28E+06	1.30E+06	1.21E+06	1.32E+06	1.48E+06
9-Oxononanoic acid	3.52E+05	6.42E+05	4.46E+05	2.98E+05	6.43E+05	3.82E+05	3.86E+05	4.90E+05	3.70E+05	4.05E+05	5.64E+05	2.99E+05	3.81E+05	3.46E+05	6.96E+05	4.58E+05	4.40E+05	4.56E+05
6-Carboxyhexanoate	9.35E+05	8.45E+05	7.84E+05	8.43E+05	1.01E+06	7.39E+05	9.87E+05	7.71E+05	9.89E+05	7.34E+05	7.77E+05	8.72E+05	8.30E+05	8.48E+05	9.40E+05	9.75E+05	7.28E+05	9.11E+05
MS_METABOLITE_DATA_END
#METABOLITES
METABOLITES_START
metabolite_name	KEGG ID	m/z	r.t.
ATP	C00002	505.9878	0.59
UTP	C00075	482.9611	0.58
UDP	C00015	402.9999	0.91
IDP	C00104	427.0070	0.54
Pyridoxal	C00250	166.0498	2.03
UDP-glucose	C00029	565.0474	0.56
Phosphate	C00009	96.9680	0.61
Diphosphate	C00013	176.9348	0.61
D-Glucose	C00031	179.0551	0.66
D-Glucose 6-phosphate	C00092	259.0221	0.57
D-Fructose 1-6-bisphosphate	C00354	338.9884	0.53
D-Glyceraldehyde 3-phosphate/Glycerone phosphate	C00118	168.9883	1.74
2/3-Phospho-D-glycerate	C00631	184.9846	0.57
Phosphoenolpyruvate	C00074	166.9740	0.57
Pyruvate	C00022	87.0071	0.67
Lactate	C01432	89.0228	0.66
D-Ribose	C00121	149.0443	0.66
D-Rhamnose	C01684	163.0614	3.59
Citrate	C00158	191.0187	0.58
2-Oxoglutarate	C00026	145.0130	0.57
Succinate	C00042	117.0178	0.61
Fumarate	C00122	115.0022	0.59
Malate	C00149	133.0128	0.57
Itaconate	C00490	129.0178	0.59
2-Hydroxyglutarate/Citramalate	C02630	147.0286	0.56
Sedoheptulose 1-phosphate	C06222	289.0328	0.58
alpha-D-Ribose 1-phosphate	C00620	229.0117	0.66
Glutathione	C00051	306.0764	0.64
Ascorbate	C00072	175.0237	0.66
Dehydroascorbate	C05422	173.0081	0.56
L-gamma-Glutamyl-L-hypoglycin	C08280	269.1157	3.61
1-4-beta-D-Xylan	C02352	131.0336	0.67
UDP-N-acetyl-D-glucosamine	C00043	606.0742	0.55
Glycolate	C00160	75.0071	0.68
Phosphocreatine	C02305	210.0278	0.56
Taurine	C00245	124.0059	0.66
L-Cysteate	C00506	167.9960	0.73
Indole-3-acetaldehyde	C00637	158.0599	1.98
Anthranilate	C00108	136.0390	1.80
Glycerol 3-phosphate	C00093	171.0056	0.60
Ethanolamine phosphate	C00346	140.0105	0.62
Sphinganine 1-phosphate	C01120	380.2551	3.61
Acetyl phosphate	C00227	138.9788	0.60
2-Methyleneglutarate	C02930	143.0336	0.67
6-Pyruvoyltetrahydropterin	C03684	236.0778	1.75
D-glucono-1,5-lactone	C00198	177.0394	0.66
Glucosinolate	C17260	316.9841	0.74
9-Oxononanoic acid	C16322	171.1015	2.57
6-Carboxyhexanoate	C02656	159.0664	3.51
METABOLITES_END
#END