#METABOLOMICS WORKBENCH amaynard_20231219_194558 DATATRACK_ID:4541 STUDY_ID:ST003020 ANALYSIS_ID:AN004954 PROJECT_ID:PR001773 VERSION 1 CREATED_ON December 20, 2023, 3:43 pm #PROJECT PR:PROJECT_TITLE Folate depletion induces erythroid differentiation through perturbation of de PR:PROJECT_TITLE novo purine synthesis PR:PROJECT_SUMMARY Folate, an essential vitamin, is a one-carbon acceptor and donor in key PR:PROJECT_SUMMARY metabolic reactions. Erythroid cells harbor a unique sensitivity to folate PR:PROJECT_SUMMARY deprivation, as revealed by the primary pathological manifestation of PR:PROJECT_SUMMARY nutritional folate deprivation: megaloblastic anemia. To study this metabolic PR:PROJECT_SUMMARY sensitivity, we applied mild folate depletion to human and mouse erythroid cell PR:PROJECT_SUMMARY lines, and primary murine erythroid progenitors. We show that folate depletion PR:PROJECT_SUMMARY induces early blockade of purine synthesis and accumulation of the purine PR:PROJECT_SUMMARY synthesis intermediate and signaling molecule, AICAR, followed by enhanced heme PR:PROJECT_SUMMARY metabolism, hemoglobin synthesis, and erythroid differentiation. This is PR:PROJECT_SUMMARY phenocopied by inhibition of folate metabolism using the SHMT1/2 inhibitor - PR:PROJECT_SUMMARY SHIN1, and by AICAR supplementation. Mechanistically, the metabolically-driven PR:PROJECT_SUMMARY differentiation is independent of nucleotide sensing through mTORC1 and AMPK, PR:PROJECT_SUMMARY and is instead mediated by protein kinase C (PKC). Our findings suggest that PR:PROJECT_SUMMARY folate deprivation-induced premature differentiation of erythroid progenitor PR:PROJECT_SUMMARY cells is a molecular etiology to folate-deficiency induced anemia. PR:INSTITUTE Boston Children's Hospital, Harvard Medical School PR:DEPARTMENT pathology PR:LABORATORY Kanarek Lab PR:LAST_NAME Kanarek PR:FIRST_NAME Naama PR:ADDRESS Enders 1116.2, 300 Longwood Ave, Boston, MA 02115 PR:EMAIL naama.kanarek@childrens.harvard.edu PR:PHONE (617) 355-7433 #STUDY ST:STUDY_TITLE Polar metabolite levels in MEL cells following PKC inhibition in 2,000 nM or 100 ST:STUDY_TITLE nM FA conditions ST:STUDY_SUMMARY Culture of MEL cells in RPMI media containing 2000 nM or 100 nM folic acid for 6 ST:STUDY_SUMMARY days in the presence or absence of the PKC inhibitor, GF109203X. These samples ST:STUDY_SUMMARY were followed up by metabolomics targeting polar metabolites. ST:INSTITUTE Boston Children's Hospital, Harvard Medical School ST:DEPARTMENT pathology ST:LABORATORY Kanarek Lab ST:LAST_NAME Kanarek ST:FIRST_NAME Naama ST:ADDRESS Enders 1116.2, 300 Longwood Ave, Boston, MA 02115 ST:EMAIL naama.kanarek@childrens.harvard.edu ST:PHONE 6173557433 ST:NUM_GROUPS 4 #SUBJECT SU:SUBJECT_TYPE Cultured cells SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:CELL_STRAIN_DETAILS MEL #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 MEL MEL_longterm_PKCi_HILIC_2000nM_FA_1 Treatment:2000nM_FA_Veh RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1129.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_2000nM_FA_2 Treatment:2000nM_FA_Veh RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1130.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_2000nM_FA_3 Treatment:2000nM_FA_Veh RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1131.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_2000nM_FA_4 Treatment:2000nM_FA_Veh RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1132.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_2000nM_FA_GFX_1 Treatment:2000nM_FA_GFX RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1137.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_2000nM_FA_GFX_2 Treatment:2000nM_FA_GFX RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1138.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_2000nM_FA_GFX_3 Treatment:2000nM_FA_GFX RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1139.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_2000nM_FA_GFX_4 Treatment:2000nM_FA_GFX RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1140.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_100nM_FA_1 Treatment:100nM_FA_Veh RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1141.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_100nM_FA_2 Treatment:100nM_FA_Veh RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1142.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_100nM_FA_3 Treatment:100nM_FA_Veh RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1143.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_100nM_FA_4 Treatment:100nM_FA_Veh RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1144.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_100nM_FA_GFX_1 Treatment:100nM_FA_GFX RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1149.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_100nM_FA_GFX_2 Treatment:100nM_FA_GFX RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1150.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_100nM_FA_GFX_3 Treatment:100nM_FA_GFX RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1151.zip SUBJECT_SAMPLE_FACTORS MEL MEL_longterm_PKCi_HILIC_100nM_FA_GFX_4 Treatment:100nM_FA_GFX RAW_FILE_NAME=Copy of 20231002_QEF1_HILIC_AM_revision_Exp4_AM1152.zip #COLLECTION CO:COLLECTION_SUMMARY One million cells from culture were collected via centrifugation for 20 seconds CO:COLLECTION_SUMMARY at 18,000xG, washed with 0.9% NaCl, and collected via centrifugation for 20 CO:COLLECTION_SUMMARY seconds at 18,000xG CO:SAMPLE_TYPE Cultured cells #TREATMENT TR:TREATMENT_SUMMARY MEL cells were cultured for 48hr in 2000 nM folic acid + vehicle, 100 nM folic TR:TREATMENT_SUMMARY acid + vehicle, 2000 nM folic acid + AICAR (500 nM), and 2000 nM folic acid + TR:TREATMENT_SUMMARY SHIN1 (1.25 uM). #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Pellet was resuspended in extraction buffer (80% Methanol, 25 mM Ammonium SP:SAMPLEPREP_SUMMARY Acetate and 2.5 mM Na-Ascorbate prepared in LC-MS water, supplemented with SP:SAMPLEPREP_SUMMARY isotopically-labelled amino acid standards [Cambridge Isotope Laboratories, SP:SAMPLEPREP_SUMMARY MSK-A2-1.2], aminopterin, and reduced glutathione standard [Cambridge Isotope SP:SAMPLEPREP_SUMMARY Laboratories, CNLM-6245-10]). Samples were vortexed for 10 sec, then centrifuged SP:SAMPLEPREP_SUMMARY for 10 minutes at 18,000 g to pellet cell debris. The supernatant was divided SP:SAMPLEPREP_SUMMARY into two tubes and dried on ice using a liquid nitrogen dryer. One tube of dried SP:SAMPLEPREP_SUMMARY sample was used for polar metabolite detection. Dried samples were resuspended SP:SAMPLEPREP_SUMMARY in 25 μL water #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Sample was injected into a ZIC-pHILIC 150 x 2.1 mm (5 μm particle size) column CH:CHROMATOGRAPHY_SUMMARY (EMD Millipore). operated on a Vanquish™ Flex UHPLC Systems (Thermo Fisher CH:CHROMATOGRAPHY_SUMMARY Scientific, San Jose, CA, USA). Chromatographic separation was achieved using CH:CHROMATOGRAPHY_SUMMARY the following conditions: buffer A was acetonitrile; buffer B was 20 mM ammonium CH:CHROMATOGRAPHY_SUMMARY carbonate, 0.1% ammonium hydroxide in water; resulting pH is around 9 without pH CH:CHROMATOGRAPHY_SUMMARY adjustment. Gradient conditions we used were: linear gradient from 20% to 80% B; CH:CHROMATOGRAPHY_SUMMARY 20–20.5 min: from 80% to 20% B; 20.5–28 min: hold at 20% B at 150 mL/min CH:CHROMATOGRAPHY_SUMMARY flow rate. The column oven and autosampler tray were held at 25 °C and 4 °C, CH:CHROMATOGRAPHY_SUMMARY respectively. CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME Thermo Vanquish CH:COLUMN_NAME EMD Millipore ZIC-HILIC (100 x 2.1 mm,3.5 um) CH:SOLVENT_A 100% acetonitrile CH:SOLVENT_B 100% water; 20 mM ammonium carbonate; 0.1% ammonium hydroxide CH:FLOW_GRADIENT linear gradient from 20% to 80% B; 20–20.5 min: from 80% to 20% B; 20.5–28 CH:FLOW_GRADIENT min: hold at 20% B at 150 mL/min flow rate CH:FLOW_RATE 150 mL/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 UNSPECIFIED MS:MS_COMMENTS Polar metabolites were relatively quantified while referencing an in-house MS:MS_COMMENTS library of chemical standards and using TraceFinder 4.1 (Thermo Fisher MS:MS_COMMENTS Scientific, Waltham, MA, USA), with a 5 ppm mass tolerance. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS Arbitrary Units MS_METABOLITE_DATA_START Samples MEL_longterm_PKCi_HILIC_2000nM_FA_1 MEL_longterm_PKCi_HILIC_2000nM_FA_2 MEL_longterm_PKCi_HILIC_2000nM_FA_3 MEL_longterm_PKCi_HILIC_2000nM_FA_4 MEL_longterm_PKCi_HILIC_2000nM_FA_GFX_1 MEL_longterm_PKCi_HILIC_2000nM_FA_GFX_2 MEL_longterm_PKCi_HILIC_2000nM_FA_GFX_3 MEL_longterm_PKCi_HILIC_2000nM_FA_GFX_4 MEL_longterm_PKCi_HILIC_100nM_FA_1 MEL_longterm_PKCi_HILIC_100nM_FA_2 MEL_longterm_PKCi_HILIC_100nM_FA_3 MEL_longterm_PKCi_HILIC_100nM_FA_4 MEL_longterm_PKCi_HILIC_100nM_FA_GFX_1 MEL_longterm_PKCi_HILIC_100nM_FA_GFX_2 MEL_longterm_PKCi_HILIC_100nM_FA_GFX_3 MEL_longterm_PKCi_HILIC_100nM_FA_GFX_4 Factors Treatment:2000nM_FA_Veh Treatment:2000nM_FA_Veh Treatment:2000nM_FA_Veh Treatment:2000nM_FA_Veh Treatment:2000nM_FA_GFX Treatment:2000nM_FA_GFX Treatment:2000nM_FA_GFX Treatment:2000nM_FA_GFX Treatment:100nM_FA_Veh Treatment:100nM_FA_Veh Treatment:100nM_FA_Veh Treatment:100nM_FA_Veh Treatment:100nM_FA_GFX Treatment:100nM_FA_GFX Treatment:100nM_FA_GFX Treatment:100nM_FA_GFX 3-phosphoglycerate 13239801.79 15291925.6 12614306.25 18582681.16 31582152.99 18482255.56 26635773.57 26347093.37 18395808.66 19248609.42 26913138.46 20799483.33 16215257.82 17986737.07 9888863.991 12586026.31 4-Hydroxyphenylpyruvic acid 2083100.068 2829910.823 2877671.7 2477766.986 2252593.2 4701450.967 1947561.565 3576179.168 NA NA 1025223.414 725541.4027 NA 2247288.556 NA 482191.5818 Acetyl CoA 2500246.491 2677669.974 2730156.794 2769075.629 2477396.164 2426417.936 2154044.83 2612551.212 2304871.448 1843754.989 2674509.827 2268305.775 1075783.004 1348977.502 884484.7558 1120605.309 ADP 24617573.07 28172272.79 25195522.51 35226385.28 51274744.38 32803904.63 40151131.8 45502266.45 33154816.51 31013346.95 49332928.68 34492038.68 27379717.19 45360821.57 14856288.09 22472177.29 AICAR NA NA NA 73340.61604 NA NA NA NA 37080082.82 33110730.94 38567692.68 33319753.55 1917618.528 3507558.234 1858690.805 2541887.616 alanine 55213984.9 54731481.41 51177265.25 48269904.65 45034288.3 47447548.58 52901867.18 60206822.84 44632558.85 47395072.19 40448492.7 43810473.26 57932026.08 66229309.74 65839810.75 100592751.6 arginine 8528833392 7978917658 8087368797 7321429436 6372151005 6961186111 5209405772 6256604912 5735019390 7752581932 5745038791 6128696160 6873919586 4142838083 4558718868 4567440669 argininosuccinate 51577330.49 51234303.87 55720986.81 56204558.93 20175314.24 18807906.37 19141761.47 23126071.85 22123028.93 22958950.64 20741337.95 21348348.33 9671666.049 9141380.243 7283597.16 11866787.02 asparagine 325065945.7 350289968.7 345927486.1 326050775.9 365264457.2 327951430.9 336426625.4 406596294 349551001 382295366.2 352302097.9 372273515.1 265750053.1 381516803.3 211537719.3 271278166.3 ATP 244770812.2 249799868.1 274209100.6 370565285.8 463410630.6 334650450.7 387519505.9 384258192.5 313481464.1 308736262.3 460337531.8 317212254.7 262217308.4 366830810.6 191131656.3 196052455.9 beta.alanine 37796101.14 36352667.22 36946838.39 36224857.95 23576044.25 25933730.25 24344862.38 25537582.89 43821491.82 52050927.38 38911599.54 44322062.93 17904871.81 21942783 16085392.21 26870498.58 CDP 4077719.991 3568548.327 3824034.185 4064489.745 1858534.266 2910577.969 2221521.341 1739996.436 3835988.395 3606375.678 3769415.899 3179051.781 1231307.698 1940055.238 1280523.432 1606102.158 choline 163145403.9 188208586 191086424.2 176418523 568403187 666123188.9 508139016.4 655870778.4 229093667.8 169063470.8 215141797.5 189269670.2 667103005.9 1046579940 673543231.9 591828509 cis.aconitate 31042288.64 21202981.24 29021896.51 26750908.76 5695872.374 16406412.96 10186009.11 16185278.37 27930640.85 24824252.65 30338034.85 24539538.56 11176962.77 41823619.98 18322904.01 12505750.02 citrate 1498715603 1220353112 1364668138 1605680103 660345509.7 380506011.2 559862378.2 502619352.9 1238625389 1694936617 1821261462 1632584508 144526375.7 170395986.8 148714465.8 414773399.2 citrulline 4115105.394 4048342.85 10687674.27 2781393.88 5204171.104 10731778.93 88920792.58 7695138.02 2776741.562 5112814.812 2983258.78 32524055.46 149186793.1 22302523.93 196424507.9 98071276.71 Coenzyme.A 4032300.249 3751726.029 2511057.378 3910208.905 1627652.064 1497708.159 1296046.986 2119115.559 4697713.773 4431693.579 4471214.406 3538672.835 216544.9586 NA 406793.7942 NA CTP 23749394.34 17071603.67 22455304.27 30361618.07 26174064.68 20722650.2 23992303.38 20489770.18 24595999.93 28375405.68 37495108.63 25660248.37 20512804.71 24119618.48 11390789.8 13920428.55 cystathionine 6252198.786 7033604.322 7129601.628 6431541.239 9425998.218 9336665.461 9697661.177 12285908.45 6526801.047 7889983.274 6334191.049 6799769.724 2981017.749 2756616.26 1697197.935 5265303.33 cystine 66282450.8 80212833.82 66318520.73 44830117.29 68543573.27 81303584.58 34456105.61 63335654.36 34874063.63 50394125.86 22506874.61 32618023.57 91462852.04 86763145.82 56560607.11 42457040.55 cytidine 2187357.676 1194852.64 2051709.284 4302951.076 89635534.09 75114894.11 76428260.62 70392629.62 423513.3723 4038044.371 5898834.849 7001867.61 167181568.8 127026103.8 137135552.2 123944380 dihydroxyacetone phosphate 8649437.674 8382271.222 6039677.762 6786789.701 6709878.194 6035685.574 5804233.224 6462835.91 2686789.451 2199041.359 2148466.209 1423533.473 2292142.853 5737366.4 2676601.796 4339508.004 dTTP 10168923.51 8194208.869 9771666.004 8077733.13 8120781.542 17058018.61 6379408.064 7669685.514 7673794.316 9301484.279 4406169.052 5505950.313 6743565.381 14392959.05 15131232.33 6925048.509 dUMP 265658.8479 352317.747 158399.3594 187482.5755 141269.881 NA 512939.6881 543078.1247 2860624.985 2806354.816 4043257.376 3392438.749 118170.9179 NA 334166.7911 NA F6P/G1P 14750515.37 15606522.87 15374342.35 16071436.41 6854680.621 7322727.26 7849237.981 6856188.06 7167716.877 5814346.697 6507152.945 5480741.605 3728643.282 6733700.517 3382506.511 4139967.937 FAD 1379094.064 1684532.975 1336681.206 1788635.42 2076933.046 2250096.413 1650390.217 1528726.526 3235688.32 1827105.351 2643934.039 2770003.646 840620.9658 1071009.518 1378602.039 NA Fructose-1,6-bisphosphate 5204072.341 5994559.656 4339328.536 4182441.801 3743149.008 3482167.423 2774567.608 3153676.653 2560761.439 2046395.237 2118406.856 2097304.509 2599311.381 4895645.531 NA 2184070.23 gamma-aminobutyric acid 88200667.43 NA 93851054.64 70671793.11 92170407.09 103298244.3 134129506.1 83365162.36 61051395.26 83037506.12 57215992.77 68478133.55 87856325.96 134762028.6 87004234.69 69681701.8 GAR NA NA NA NA NA NA NA NA 33875175.37 32172588.71 34433384.26 32095865.63 3868996.231 5885035.603 3356748.804 4555395.822 GDP 6041263.767 4876249.92 3741802.074 5539477.118 4205074.234 4963469.992 3843775.846 3644882.166 5419580.341 5433633.767 5320582.473 4857806.459 3043220.879 4059019.863 2958566.223 2637241.295 Glucose-6-phosphate 9299693.446 11305826.67 9725882.876 10026285 7408041.17 7753300.639 6828125.804 7933929.699 7966730.142 7219713.754 6972562.838 7219100.651 5933179.535 10719282.12 4931861.618 6138536.798 glucose 5298954.291 6455528.78 3633319.997 2115151.9 3606503.27 5501566.087 2162292.541 3661380.305 3736514.739 2374342.768 1598260.503 2333316.631 9087579.447 4119591.819 3067201.812 12637814.95 glutamine 7112833524 6950240040 7102768344 6300128554 7532397470 7669808972 6624235660 8240069723 7584466890 8522072144 6951667464 7701144474 6252780802 7100612104 5077203632 5750347769 Glycerophosphocholine 13758249191 14487997789 14517181721 14586955797 43481743435 44851030455 36228774603 41761803141 15896012162 14995938269 13991064716 14032441605 52816962481 60072856349 48141367192 40978882447 glycine 29685600.89 27587962.41 27544033.22 25549763.66 22105810.22 21330567.61 20635135.76 20315937.01 19027334.08 20772839.45 18362052.85 18396625.31 6917156.105 11045576.63 11050887.74 6233424.226 GSH 2500020030 2906151582 2726998949 2710775299 2613390369 2045727450 2364820908 2532923350 3300535774 3370631837 3232944956 3233379756 1388030936 85360531.24 1062678419 10829558.63 GSSG 72823415.64 16520778.08 32233362.9 27454630.42 22692975.04 26253348.84 23094993.67 16969609.14 55431013.12 98309002.54 62971153.25 57563378.61 28651314.87 285573311.8 20018675.84 87374286.91 GTP 22198263.5 12776553.24 18632735.46 29292074.75 29682933.32 21279013.89 23693519.22 15712094.67 21222412.03 26924842.91 30468857.76 18343517.06 28229480.06 24697431.29 15143258.13 17219073.71 guanine 2032440.915 372671.9138 1621546.693 2775703.999 19763812.88 19008391.58 15129745.69 16000942.68 6914405.748 933039.5705 628427.2292 627920.4999 34203002.73 35258663.65 36434488.05 24432556.02 guanosine 1061750.64 1121156.03 1025857.632 1414742.561 17521916.45 16729920.42 13085809.46 13834313.36 350553.5851 1191024.652 405832.384 223527.9026 30361822.73 28765835.12 32303065.85 24334740.77 histidine 797363484 767473934.5 780296010.9 660499063.5 470882865.6 608893584.1 590792044.5 508222507.7 648011540 706189215.5 541631283.7 629388561.1 473956677.2 NA 582817886.6 362972198.2 hydroxyproline-gammaALA 65514574.82 91489512.65 82850079.41 85199912.11 86410417.97 80570487.94 78069403.61 103983753.7 116710505.6 92869314.25 119640643.1 122592382.4 56411495.25 99639182.74 48881216.87 64525285.99 hypoxanthine 25485424.79 30416580.75 30263105.79 30408904.62 25969913.94 24925491.33 23738210.45 32384347.94 9561591.626 9274945.787 10639888.71 10468988.25 23323024.81 9425547.694 31318436.89 82204130.68 IMP 3519893.049 3165218.31 1932239.878 2865182.32 643163.4325 535039.1433 581624.9726 814852.092 4123413.554 3816960.733 2991317.997 3378259.35 528971.457 595623.0963 430676.9186 797151.5051 inosine 16557926.93 20165206.19 13647342.43 20669015 11311572.38 10006210.48 8347631.201 10356263.85 1036661.109 806456.6205 1104664.573 875303.2711 13394684.33 10873237.85 13351641.77 14023840.75 isocitrate 1580443735 1319446093 1445743414 1708514869 740588589.9 NA 617138464.8 527853354.6 1302232426 1832579218 1944199570 1729556158 149192671.7 179735381.1 173861663.5 432058145.8 isoleucine 653756993.5 780858759.7 716670535.3 642741617.3 781525351 673524444 637996022.7 812952455.8 555533303 605082699.9 543488895.1 607423619.5 625064278.7 390341541.1 463742792.2 539634942.6 kynurenine 1304597.197 1904019.993 1754260.704 1864561.325 1816545.771 1922163.097 2446725.017 2077297.813 1203452.741 1386249.176 1821288.305 1719022.292 1272519.84 403070.7587 1063194.119 3055777.522 leucine 621467296.9 795479729.6 737464215.7 653763110.4 791409714.4 691487813.4 659424493.6 871407053.1 580581340.4 591798927.8 575178289.2 624058974.2 562959657.9 411938613.3 473077648.4 547927319.6 lysine 225984911.7 206990684.5 211967904 204074488.7 211678270.5 188299609 198540534.4 189067641.7 138616805.4 196456495.3 150430995.1 160233657.4 261366188.8 131844705 182811765.3 282973635.5 Methionine sulfoxide 144308597.5 116663759.2 127257528.1 120230871.8 112412160.1 104823096 138738587.1 116267817.9 119034782.8 140278846.4 150181859.8 165763591.1 159965529.3 81163899.7 122072590.2 268789099.3 methionine 365317850.9 432282708.2 398065340.5 365956407.9 421078209.9 336742488.7 347469916.7 464221146.4 272021528.6 323947427.3 312920163.3 337720463.3 304716147.3 133500716.5 193715770 259667815.5 methylthioadenosine 24102059.98 27618793.64 27182208.58 27580052.96 16385541.58 14517174.48 18432620.88 17225504 17931144.39 19178747.16 21702386.17 20548736.79 5099441.061 11846464.86 4779322.843 19217680.22 Mevalonate 41979218.92 43116955.86 42617247.41 38673415.76 43062882.2 45669993.97 41065516.83 50752172.04 44822156.24 52534909.37 42946957.58 46313345.12 36835634.7 28035782.55 30477358.2 33640804.79 N-acetylglutamate 3823301.32 4063626.169 4082904.501 4255363.232 1055321.293 1072250.371 1033344.611 1041108.122 1947310.048 2534834.724 2395142.493 2531037.371 934699.0718 359216.2863 1165129.857 2724642.64 N-acetylglutamine 4646837.702 5426602.942 6243109.02 4619441.724 5061573.902 4467104.842 5572086.334 6860407.023 11607558.54 8964289.392 NA 13335242.12 6093290.512 5104908.48 NA 11898264.29 NAD+ 142692566.3 130276854.2 119095029.1 122315132.2 132267406.1 132427857.9 119274669.6 133414299.4 164339929.1 168244312.5 145641436 138884481 101015368.8 165975549 82920350.94 80293038.46 NADP+ 5391052.419 3789143.81 3556783.115 4642428.621 4738082.166 7094020.084 4025806.487 4501485.555 8211984.308 7184447.215 5976607.145 4743362.315 4508624.299 8736838.657 3921296.982 3920552.826 niacinamide 409323328.6 485957982.2 497044724 471497182.1 357975230.5 320002831.7 348727641.3 453566308.5 376550339.1 385126342.3 396267401.1 438068225.1 180615972.6 78400976.3 168478148.6 435990367.5 ornithine 30726264.32 29042249.28 38886859.71 26502968.98 42975749.38 48857924.12 154851591.2 50180233.83 50444303.1 48121683.62 45466350.12 50503658.56 155404163.8 18290169.6 251739405.4 65275913.73 palmitate 763225287.4 625911918.1 1769335634 1095573349 847086041.5 1015004642 1269191392 1219358636 532158234 509172689.5 243655191.5 373317294.6 210800069.2 808301376.6 475445309.6 1742957444 Pantothenate 415907279.8 455991510.2 423987629.7 441210120.1 685128149 730301433.9 627477399.3 731173468.5 593022079.9 526042542.7 569970063.2 540214375.9 486945356.9 297359243.6 413835754.2 653239074.9 phenylalanine 51594253.07 97983531.74 87224794.68 80766943.36 94583273.44 97199261.23 83883716.63 107366013.3 84113406.29 51508547.02 76182067.39 76730311.55 56507893.08 86975989.62 70542747.93 77156467.23 phosphocholine 24073769800 23407414137 23669558126 23907119706 16805311711 8646642442 14961174521 17071360491 12691457805 11585528619 11634098531 10990719993 6058584942 6274346034 5256438819 2884642267 phosphoenolpyruvate 7867704.286 9709503.8 8552853.654 9441490.386 31476739.5 15263436.73 30211228.36 25534386.32 11389315.54 11687374.02 16103208.19 14721674.03 14910200.52 18205560.39 11095388.89 11343055.65 phosphoserine 1326490.184 1847025.495 1258328.454 1524848.924 23758124.01 21894630.4 19573678.81 25140158.67 14346197.2 13420084.99 16610659.44 15467532.76 20358327.93 32250707.79 14754714.22 17219341.11 proline 6632957962 6880711985 6595284749 6232570417 5506579640 5343529144 5025965950 6156287938 5444513500 5756482014 5353225786 5573919157 3600225260 2533630522 2532601808 3733723714 pseudouridine 217972.3141 324722.6231 321180.4918 400877.9247 2578915.82 2980514.246 2546289.684 2486984.892 432617.7229 207428.8818 574597.7135 995664.5858 4182337.737 5262424.434 5125451.366 4391159.562 Pyridoxine 131584160.7 161798521.5 139961172 119796373 108959090.7 126779996.8 79123416.47 115087221.7 136494411.8 146052203.2 113712855.3 131508475.1 107047251.5 97820184.36 82462843.77 77634346.71 Ribose-1-phosphate 1177508.79 1984189.909 1539174.624 1415470.521 1275256.68 1972930.276 1613590.513 1356129.084 4016617.867 3478019.748 4290843.436 4471808.159 61876.9986 1704099.757 706138.4442 54511.81175 Ribose Ribulose-5-phosphate-3Me-Lys 2322981.411 1984189.909 2237456.744 1818457.473 1286126.39 1972930.276 1613590.513 1703252.17 3535391.678 2328930.776 2587729.63 2708015.21 813471.3132 1867102.675 1026476.04 992567.7106 S-Adenosyl-L-Homocysteine 20897209.01 19186957.25 20659647 19811447.59 11589600.35 11227020.23 11154675.42 11470450.15 17498606.77 15823192.56 18041098.34 16999596.75 4951590.163 1733422.367 4160191.107 5592645.457 S-Adenosyl-L-methionine 643074265.9 543743611.1 555814604.2 511804308.6 432867393 593650127.2 407749489.9 444561586.4 750950743.6 684479698.9 556009691.3 564658714.9 272951798 249210240.3 265261357.7 226148089.3 Sedoheptulose-7-phosphate 19303202.86 20692436.06 20169186.28 20894980.3 2624395.004 2407770.682 2546524.538 2854467.185 2618423.206 2246548.913 2236721.429 1857494.192 2002801.125 2309817.034 1278867.156 2045058.836 serine 100442992.1 103549072.9 101188777.9 97312168.84 116372076.5 109676421.9 127403759.8 130547681.3 144520430.2 149635208.8 140721051.8 154797703.3 147542062.6 157746127.3 149027413 121305628.9 succinate 38775493.33 22396414 64363467.64 31296506.48 42543245.51 61272641.91 31751346.35 NA 33393346.27 35692112.81 15983691.67 6828730.524 20493773.58 29163082.13 39341773.78 39107709.15 threonine 236291349.4 234593434.8 219946302.8 197193492.7 168389910.1 188493189.2 176147143.7 204777677.6 242952993.5 295309087.9 249691715.7 269534852.9 127210787.2 141726591.7 143934914 131251392.5 trans-urocanate 67729604.14 85405500.5 144970273.1 49150870.65 83506897.89 67928961.28 529901210.2 86828320.18 43591470.54 59456551.1 76838503.8 208405818.6 1382333200 91423076.43 2262830434 281718027.8 tryptophan 67349200.23 84707129.25 71932875.83 67506239.13 96945531.97 76503039.07 89443977.9 103496001.2 52124366.49 58338832.02 60107865.88 66211724.21 89535972 44068700.01 78093713.88 104424894.3 tyrosine 329432709.4 388044510.2 339255239.6 324408027.3 430611964.5 312300703.6 388494055.8 456067595.2 234495989.2 288233398.5 269291705.9 283341234.4 346528607 246681247.1 249183060.4 349365792.2 UDP 13618526.28 10858408.48 11362452.96 12230507.27 7242812.324 9938108.976 9477090.021 6484674.494 15615249.85 14454954.22 16309858.63 15262728.97 5499897.415 10152528.02 5857982.264 7055809.284 UMP 3472612.489 3729973.758 2364870.281 4082691.564 1215263.987 1529876.487 825994.2791 1043321.941 2449328.201 2220425.868 2648712.832 2211716.736 650402.6642 1453174.83 633857.8828 2629319.851 UTP 82293069.1 65198725.77 73939263.73 105248979.9 111473321.4 81990414 91760205.44 78131789.88 111760749 120577503.9 162419690.6 118015188.5 89569976.54 114203188.4 61213616.34 57675271.1 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name Standardized name Formula Exact mass Sub class 3-phosphoglycerate 3-Phosphoglyceric acid C3H7O7P 185.9929 Short-chain acids 4-Hydroxyphenylpyruvic acid 4-Hydroxyphenylpyruvic acid C9H8O4 180.0423 Cinnamic acids Acetyl CoA Acetyl-CoA C23H38N7O17P3S 809.1258 Acyl CoAs ADP ADP C10H15N5O10P2 427.0294 Purine rNDP AICAR AICAR C9H14N4O5 258.0964 Imidazole ribonucleosides and ribonucleotides alanine Alanine C3H7NO2 89.0477 Amino acids arginine Arginine C6H14N4O2 174.1117 Amino acids argininosuccinate Argininosuccinic acid C10H18N4O6 290.1226 Amino acids asparagine Asparagine C4H8N2O3 132.0535 Amino acids ATP ATP C10H16N5O13P3 506.9958 Purine rNTP beta.alanine beta-Alanine C3H7NO2 89.0477 Amino acids CDP CDP C9H15N3O11P2 403.0182 Pyrimidine rNDP choline Choline C5H14NO 104.1075 Cholines cis.aconitate cis-Aconitic acid C6H6O6 174.0164 TCA acids citrate Citric acid C6H8O7 192.0270 TCA acids citrulline Citrulline C6H13N3O3 175.0957 Amino acids Coenzyme.A Coenzyme A C21H36N7O16P3S 767.1152 Other purines CTP CTP C9H16N3O14P3 482.9845 Pyrimidine rNTP cystathionine Cystathionine C7H14N2O4S 222.0674 Amino acids cystine Cystine C6H12N2O4S2 240.0239 Amino acids cytidine Cytidine C9H13N3O5 243.0855 Pyrimidine ribonucleosides dihydroxyacetone phosphate Dihydroxyacetone phosphate C3H7O6P 169.9980 Short-chain acids dTTP dTTP C10H17N2O14P3 481.9893 Pyrimidine dNTP dUMP dUMP C9H13N2O8P 308.0410 Pyrimidine dNMP F6P/G1P - FAD FAD C27H33N9O15P2 785.1571 Flavin nucleotides Fructose-1,6-bisphosphate Fructose 1,6-bisphosphate C6H14O12P2 339.9961 Monosaccharide phosphates gamma-aminobutyric acid gamma-Aminobutyric acid C4H9NO2 103.0633 Amino FA GAR GAR C7H15N2O8P 286.0566 Glycinamide ribonucleotides GDP GDP C10H15N5O11P2 443.0243 Purine rNDP Glucose-6-phosphate Glucose 6-phosphate C6H13O9P 260.0297 Monosaccharide phosphates glucose Glucose C6H12O6 180.0634 Hexoses glutamine Glutamine C5H10N2O3 146.0691 Amino acids Glycerophosphocholine sn-Glycero-3-phosphocholine C8H20NO6P 257.1028 Organic phosphoric acids glycine Glycine C2H5NO2 75.0320 Amino acids GSH Glutathione C10H17N3O6S 307.0838 Tripeptides GSSG Oxidized glutathione C20H32N6O12S2 612.1520 Tripeptides GTP GTP C10H16N5O14P3 522.9907 Purine rNTP guanine Guanine C5H5N5O 151.0494 Hypoxanthines guanosine Guanosine C10H13N5O5 283.0917 Purine ribonucleosides histidine Histidine C6H9N3O2 155.0695 Amino acids hydroxyproline-gammaALA - hypoxanthine Hypoxanthine C5H4N4O 136.0385 Hypoxanthines IMP IMP C10H13N4O8P 348.0471 Purine rNMP inosine Inosine C10H12N4O5 268.0808 Purine ribonucleosides isocitrate Isocitric acid C6H8O7 192.0270 TCA acids isoleucine Isoleucine C6H13NO2 131.0946 Amino acids kynurenine Kynurenine C10H12N2O3 208.0848 Amino acids leucine Leucine C6H13NO2 131.0946 Amino acids lysine Lysine C6H14N2O2 146.1055 Amino acids Methionine sulfoxide Methionine sulfoxide C5H11NO3S 165.0460 Amino acids methionine Methionine C5H11NO2S 149.0511 Amino acids methylthioadenosine Methylthioadenosine C11H15N5O3S 297.0896 Purine ribonucleosides Mevalonate Mevalonic acid C6H12O4 148.0736 Hydroxy FA N-acetylglutamate N-Acetylglutamic acid C7H11NO5 189.0637 Amino acids N-acetylglutamine N-Acetylglutamine C7H12N2O4 188.0797 Amino acids NAD+ NAD+ C21H27N7O14P2 663.1091 Nicotinamide dinucleotides NADP+ NADP+ C21H28N7O17P3 743.0755 Nicotinamide dinucleotides niacinamide Niacinamide C6H6N2O 122.0480 Pyridine alkaloids ornithine Ornithine C5H12N2O2 132.0899 Amino acids palmitate Palmitic acid C16H32O2 256.2402 Saturated FA Pantothenate Pantothenic acid C9H17NO5 219.1107 Amino acids phenylalanine Phenylalanine C9H11NO2 165.0790 Amino acids phosphocholine Choline phosphate C5H14NO4P 183.0660 Cholines phosphoenolpyruvate Phosphoenolpyruvic acid C3H5O6P 167.9824 Short-chain acids phosphoserine Phosphoserine C3H8NO6P 185.0089 Amino acids proline Proline C5H9NO2 115.0633 Amino acids pseudouridine Pseudouridine C9H12N2O6 244.0695 Pyrimidine ribonucleosides Pyridoxine Pyridoxine C8H11NO3 169.0739 Pyridine alkaloids Ribose-1-phosphate Ribose 1-phosphate C5H11O8P 230.0192 Monosaccharide phosphates Ribose Ribulose-5-phosphate-3Me-Lys - S-Adenosyl-L-Homocysteine S-Adenosylhomocysteine C14H20N6O5S 384.1216 Other purines S-Adenosyl-L-methionine S-Adenosylmethionine C15H23N6O5S 399.1451 Other purines Sedoheptulose-7-phosphate Sedoheptulose 7-phosphate C7H15O10P 290.0403 Monosaccharide phosphates serine Serine C3H7NO3 105.0426 Amino acids succinate Succinic acid C4H6O4 118.0266 TCA acids threonine Threonine C4H9NO3 119.0582 Amino acids trans-urocanate Urocanic acid C6H6N2O2 138.0429 Imidazole alkaloids tryptophan Tryptophan C11H12N2O2 204.0899 Amino acids tyrosine Tyrosine C9H11NO3 181.0739 Amino acids UDP UDP C9H14N2O12P2 404.0022 Pyrimidine rNDP UMP UMP C9H13N2O9P 324.0359 Pyrimidine rNMP UTP UTP C9H15N2O15P3 483.9685 Pyrimidine rNTP METABOLITES_END #END