#METABOLOMICS WORKBENCH oilkay_20241205_115113 DATATRACK_ID:5430 STUDY_ID:ST003647 ANALYSIS_ID:AN005993 PROJECT_ID:PR002257 VERSION 1 CREATED_ON 01-03-2025 #PROJECT PR:PROJECT_TITLE Integration of metabolomic and transcriptomic analyses reveals novel regulatory PR:PROJECT_TITLE functions of the ChREBP transcription factor in energy metabolism. PR:PROJECT_SUMMARY In this study, we have integrated transcriptomic and metabolomic analyses to PR:PROJECT_SUMMARY better understand the role of the metabolic regulatory transcription factor PR:PROJECT_SUMMARY ChREBP in coordinated regulation of key pathways of intermediary metabolism in PR:PROJECT_SUMMARY the liver. We have uncovered regulatory effects of ChREBP on metabolic PR:PROJECT_SUMMARY homeostasis beyond itâs historical role in control of core glucose and lipid PR:PROJECT_SUMMARY metabolic pathways, to now include effects on co-factors, transporters for amino PR:PROJECT_SUMMARY acids and other small molecules, nucleotide metabolism, and control of PR:PROJECT_SUMMARY mitochondrial substrate supply. PR:INSTITUTE Duke University PR:LAST_NAME Ilkayeva PR:FIRST_NAME Olga PR:ADDRESS 300 N Duke St, Durham, NC, 27701, USA PR:EMAIL olga.ilkayeva@duke.edu PR:PHONE 919-479-2370 PR:DOI http://dx.doi.org/10.21228/M8WC2H #STUDY ST:STUDY_TITLE Targeted mass spec-based metabolomic and clinical analyte analyses of liver and ST:STUDY_TITLE plasma samples from rats with and without hepatic knockdown of ChREBP ST:STUDY_TITLE expression. ST:STUDY_SUMMARY The transcription factor Carbohydrate Response Element-Binding Protein (ChREBP) ST:STUDY_SUMMARY activates genes of glucose, fructose and lipid metabolism in response to ST:STUDY_SUMMARY carbohydrate feeding. Integrated transcriptomic and metabolomic analyses in rats ST:STUDY_SUMMARY with GalNac-siRNA-mediated suppression of ChREBP expression (GalNac-siChREBP ST:STUDY_SUMMARY treatment) in liver revealed novel ChREBP functions relative to rats treated ST:STUDY_SUMMARY with a GalNac vector expressing a non-targeting siRNA (GalNac-siCtrl treatment). ST:STUDY_SUMMARY GalNac-siChREBP treatment reduced expression of genes involved in coenzyme A ST:STUDY_SUMMARY (CoA) biosynthesis, and lowered CoA and short chain acyl CoA levels. Despite ST:STUDY_SUMMARY suppression of pyruvate kinase, pyruvate levels were maintained, possibly via ST:STUDY_SUMMARY increased expression of pyruvate and amino acid transporters. In addition, ST:STUDY_SUMMARY expression of multiple anaplerotic enzymes was decreased by GalNac-siChREBP ST:STUDY_SUMMARY treatment, affecting TCA cycle intermediates. Finally, GalNAc-siChREBP treatment ST:STUDY_SUMMARY suppressed late steps in purine and NAD synthesis, with increases in precursors ST:STUDY_SUMMARY and lowering of end products in both pathways. In sum, our studies have revealed ST:STUDY_SUMMARY functions of ChREBP beyond its canonical roles in carbohydrate and lipid ST:STUDY_SUMMARY metabolism to include regulation of substrate transport, mitochondrial function, ST:STUDY_SUMMARY and cellular energy balance. ST:INSTITUTE Duke University ST:LAST_NAME Ilkayeva ST:FIRST_NAME Olga ST:ADDRESS 300 N Duke St, Durham, NC, 27701, USA ST:EMAIL olga.ilkayeva@duke.edu ST:PHONE 919-479-2370 ST:SUBMIT_DATE 2024-12-05 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Rattus norvegicus SU:TAXONOMY_ID 10116 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS R01E4-1 1 Treatment:GalNAc-siChrebp | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a032; RAW_FILE_NAME(AA raw file name)=AnJie_Newgard_AA_013123_a032; RAW_FILE_NAME(OA raw file name)=AnJie_Newgard_27Liver_OA_052924_a020; RAW_FILE_NAME(BCKA raw file name)=AnJie_Newgard_KA_012723_a032; RAW_FILE_NAME(Nucleotides raw file name)=AnJie_Newgard_Liver_NucleoTides_061024_a028; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=AnJie_Newgard_27_Liver_061124_a012; RAW_FILE_NAME(SC Acyl CoAs raw file name)=LC_SC_CoA_032123_a018 SUBJECT_SAMPLE_FACTORS R01E4-19 19 Treatment:GalNAc-siChrebp | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a050; RAW_FILE_NAME(AA raw file name)=AnJie_Newgard_AA_013123_a050; RAW_FILE_NAME(OA raw file name)=AnJie_Newgard_27Liver_OA_052924_a038; RAW_FILE_NAME(BCKA raw file name)=AnJie_Newgard_KA_012723_a050; RAW_FILE_NAME(Nucleotides raw file name)=AnJie_Newgard_Liver_NucleoTides_061024_a023; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=AnJie_Newgard_27_Liver_061124_a030; RAW_FILE_NAME(SC Acyl CoAs raw file name)=LC_SC_CoA_032123_a036 SUBJECT_SAMPLE_FACTORS R01E4-21 21 Treatment:GalNAc-siChrebp | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a052; RAW_FILE_NAME(AA raw file name)=AnJie_Newgard_AA_013123_a052; RAW_FILE_NAME(OA raw file name)=AnJie_Newgard_27Liver_OA_052924_a040; RAW_FILE_NAME(BCKA raw file name)=AnJie_Newgard_KA_012723_a052; RAW_FILE_NAME(Nucleotides raw file name)=AnJie_Newgard_Liver_NucleoTides_061024_a038; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=AnJie_Newgard_27_Liver_061124_a032; RAW_FILE_NAME(SC Acyl CoAs raw file name)=LC_SC_CoA_032123_a038 SUBJECT_SAMPLE_FACTORS R01E4-3 3 Treatment:GalNAc-siChrebp | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a034; RAW_FILE_NAME(AA raw file name)=AnJie_Newgard_AA_013123_a034; RAW_FILE_NAME(OA raw file name)=AnJie_Newgard_27Liver_OA_052924_a022; RAW_FILE_NAME(BCKA raw file name)=AnJie_Newgard_KA_012723_a034; RAW_FILE_NAME(Nucleotides raw file name)=AnJie_Newgard_Liver_NucleoTides_061024_a033; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=AnJie_Newgard_27_Liver_061124_a014; RAW_FILE_NAME(SC Acyl CoAs raw file name)=LC_SC_CoA_032123_a020 SUBJECT_SAMPLE_FACTORS R01E4-6 6 Treatment:GalNAc-siChrebp | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a037; RAW_FILE_NAME(AA raw file name)=AnJie_Newgard_AA_013123_a037; RAW_FILE_NAME(OA raw file name)=AnJie_Newgard_27Liver_OA_052924_a025; RAW_FILE_NAME(BCKA raw file name)=AnJie_Newgard_KA_012723_a037; RAW_FILE_NAME(Nucleotides raw file name)=AnJie_Newgard_Liver_NucleoTides_061024_a031; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=AnJie_Newgard_27_Liver_061124_a017; RAW_FILE_NAME(SC Acyl CoAs raw file name)=LC_SC_CoA_032123_a023 SUBJECT_SAMPLE_FACTORS R01E4-7 7 Treatment:GalNAc-siChrebp | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a038; RAW_FILE_NAME(AA raw file name)=AnJie_Newgard_AA_013123_a038; RAW_FILE_NAME(OA raw file name)=AnJie_Newgard_27Liver_OA_052924_a026; RAW_FILE_NAME(BCKA raw file name)=AnJie_Newgard_KA_012723_a038; RAW_FILE_NAME(Nucleotides raw file name)=AnJie_Newgard_Liver_NucleoTides_061024_a042; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=AnJie_Newgard_27_Liver_061124_a018; RAW_FILE_NAME(SC Acyl CoAs raw file name)=LC_SC_CoA_032123_a024 SUBJECT_SAMPLE_FACTORS R01E4-15 15 Treatment:GalNAc-siCtrl | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a046; RAW_FILE_NAME(AA raw file name)=AnJie_Newgard_AA_013123_a046; RAW_FILE_NAME(OA raw file name)=AnJie_Newgard_27Liver_OA_052924_a034; RAW_FILE_NAME(BCKA raw file name)=AnJie_Newgard_KA_012723_a046; RAW_FILE_NAME(Nucleotides raw file name)=AnJie_Newgard_Liver_NucleoTides_061024_a029; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=AnJie_Newgard_27_Liver_061124_a026; RAW_FILE_NAME(SC Acyl CoAs raw file name)=LC_SC_CoA_032123_a032 SUBJECT_SAMPLE_FACTORS R01E4-17 17 Treatment:GalNAc-siCtrl | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a048; RAW_FILE_NAME(AA raw file name)=AnJie_Newgard_AA_013123_a048; RAW_FILE_NAME(OA raw file name)=AnJie_Newgard_27Liver_OA_052924_a036; RAW_FILE_NAME(BCKA raw file name)=AnJie_Newgard_KA_012723_a048; RAW_FILE_NAME(Nucleotides raw file name)=AnJie_Newgard_Liver_NucleoTides_061024_a021; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=AnJie_Newgard_27_Liver_061124_a028; RAW_FILE_NAME(SC Acyl CoAs raw file name)=LC_SC_CoA_032123_a034 SUBJECT_SAMPLE_FACTORS R01E4-18 18 Treatment:GalNAc-siCtrl | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a049; RAW_FILE_NAME(AA raw file name)=AnJie_Newgard_AA_013123_a049; RAW_FILE_NAME(OA raw file name)=AnJie_Newgard_27Liver_OA_052924_a037; RAW_FILE_NAME(BCKA raw file name)=AnJie_Newgard_KA_012723_a049; RAW_FILE_NAME(Nucleotides raw file name)=AnJie_Newgard_Liver_NucleoTides_061024_a045; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=AnJie_Newgard_27_Liver_061124_a029; RAW_FILE_NAME(SC Acyl CoAs raw file name)=LC_SC_CoA_032123_a035 SUBJECT_SAMPLE_FACTORS R01E4-2 2 Treatment:GalNAc-siCtrl | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a033; RAW_FILE_NAME(AA raw file name)=AnJie_Newgard_AA_013123_a033; RAW_FILE_NAME(OA raw file name)=AnJie_Newgard_27Liver_OA_052924_a021; RAW_FILE_NAME(BCKA raw file name)=AnJie_Newgard_KA_012723_a033; RAW_FILE_NAME(Nucleotides raw file name)=AnJie_Newgard_Liver_NucleoTides_061024_a026; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=AnJie_Newgard_27_Liver_061124_a013; RAW_FILE_NAME(SC Acyl CoAs raw file name)=LC_SC_CoA_032123_a019 SUBJECT_SAMPLE_FACTORS R01E4-10 10 Treatment:Sal | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a041; RAW_FILE_NAME(AA raw file name)=-; RAW_FILE_NAME(OA raw file name)=-; RAW_FILE_NAME(BCKA raw file name)=-; RAW_FILE_NAME(Nucleotides raw file name)=-; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=-; RAW_FILE_NAME(SC Acyl CoAs raw file name)=- SUBJECT_SAMPLE_FACTORS R01E4-11 11 Treatment:Sal | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a042; RAW_FILE_NAME(AA raw file name)=-; RAW_FILE_NAME(OA raw file name)=-; RAW_FILE_NAME(BCKA raw file name)=-; RAW_FILE_NAME(Nucleotides raw file name)=-; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=-; RAW_FILE_NAME(SC Acyl CoAs raw file name)=- SUBJECT_SAMPLE_FACTORS R01E4-13 13 Treatment:Sal | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a044; RAW_FILE_NAME(AA raw file name)=-; RAW_FILE_NAME(OA raw file name)=-; RAW_FILE_NAME(BCKA raw file name)=-; RAW_FILE_NAME(Nucleotides raw file name)=-; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=-; RAW_FILE_NAME(SC Acyl CoAs raw file name)=- SUBJECT_SAMPLE_FACTORS R01E4-4 4 Treatment:Sal | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a035; RAW_FILE_NAME(AA raw file name)=-; RAW_FILE_NAME(OA raw file name)=-; RAW_FILE_NAME(BCKA raw file name)=-; RAW_FILE_NAME(Nucleotides raw file name)=-; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=-; RAW_FILE_NAME(SC Acyl CoAs raw file name)=- SUBJECT_SAMPLE_FACTORS R01E4-9 9 Treatment:Sal | Sample source:liver RAW_FILE_NAME(AC raw file name)=AnJie_Newgard_AC_013123_a040; RAW_FILE_NAME(AA raw file name)=-; RAW_FILE_NAME(OA raw file name)=-; RAW_FILE_NAME(BCKA raw file name)=-; RAW_FILE_NAME(Nucleotides raw file name)=-; RAW_FILE_NAME(Creatine/Phosphocreatine raw file name)=-; RAW_FILE_NAME(SC Acyl CoAs raw file name)=- #COLLECTION CO:COLLECTION_SUMMARY On day 28 between 8 AM-noon, animals were anesthetized and sacrificed for CO:COLLECTION_SUMMARY collection of plasma and tissue samples. To minimize tissue harvest time, a team CO:COLLECTION_SUMMARY of 3 scientists collected tissues from the animals via the following procedure. CO:COLLECTION_SUMMARY Animals were anesthetized with 250 mg/kg Nembutal, and the abdomen and diaphragm CO:COLLECTION_SUMMARY were surgically opened. Five ml of blood was drawn from the heart, and one CO:COLLECTION_SUMMARY operator centrifuged the sample, collected the plasma, and transferred it to CO:COLLECTION_SUMMARY Eppendorf tubes for rapid freezing by submersion in liquid nitrogen. The other CO:COLLECTION_SUMMARY two team members proceeded to immediate surgical excision of the heart and CO:COLLECTION_SUMMARY liver, which were briefly rinsed with ice-cold PBS, quickly wrapped in aluminum CO:COLLECTION_SUMMARY foil, and then frozen by submersion in liquid nitrogen. The time elapsed between CO:COLLECTION_SUMMARY beginning of surgery to removal and freezing of the heart and liver was less CO:COLLECTION_SUMMARY than 2 minutes. Other tissues (skeletal muscle, adipose, kidney) were collected CO:COLLECTION_SUMMARY after excision of the heart and liver and rapidly frozen in liquid nitrogen. All CO:COLLECTION_SUMMARY tissues were stored at -80C until processing for metabolomic, transcriptomic, CO:COLLECTION_SUMMARY and proteomic analyses. CO:SAMPLE_TYPE Liver #TREATMENT TR:TREATMENT_SUMMARY All procedures were approved by Duke University Institutional Animal Care and TR:TREATMENT_SUMMARY Use Committee and performed according to the regulations of the committee. TR:TREATMENT_SUMMARY Breeding pairs of Obese Prone CD (OP/CD) Sprague Dawley rats were gifts from Dr. TR:TREATMENT_SUMMARY Warren Grill and Dr. Eric Gonzalez, Duke University, and a colony was TR:TREATMENT_SUMMARY established and maintained by Duke Laboratory Animal Resources (DLAR). Starting TR:TREATMENT_SUMMARY at 4 weeks of age, male OP/CD rats were single-housed with a light cycle of 7 AM TR:TREATMENT_SUMMARY on/7 PM off, and fed ad libitum with a high-fat/high-sucrose (HF/HS) diet TR:TREATMENT_SUMMARY (D12451i, Research Diets) containing 47% fat (kcal) and 17% sucrose (kcal). Body TR:TREATMENT_SUMMARY weight and food intake were monitored weekly. After 9 weeks of feeding of the TR:TREATMENT_SUMMARY HF/HS diet, plasma samples were collected via saphenous vein bleeding. One week TR:TREATMENT_SUMMARY later, animals received an initial subcutaneous injection of one of two TR:TREATMENT_SUMMARY GalNAc-siRNA constructs at a dose of 9 mg/kg body weight, or an equal volume of TR:TREATMENT_SUMMARY the diluent (PBS), (see below for description of the two GalNAc-siRNA reagents). TR:TREATMENT_SUMMARY Additional doses of each GalNAc-siRNA construct were injected at 10, 18 and 25 TR:TREATMENT_SUMMARY days after the first injection. Animals were fasted overnight one day after the TR:TREATMENT_SUMMARY third injection (day 19), and subjected to an intraperitoneal glucose tolerance TR:TREATMENT_SUMMARY test (IPGTT) on the following day. Animals were weighed and a glucose solution TR:TREATMENT_SUMMARY (1g/kg body weight) was administered via intraperitoneal injection. Tail blood TR:TREATMENT_SUMMARY samples were obtained and glucose levels measured with a blood glucose meter TR:TREATMENT_SUMMARY (CVSHealth) immediately before and at 30,60, 90, 120, and 180 minutes after TR:TREATMENT_SUMMARY bolus injection of glucose. One day after the fourth GalNAc-siRNA or saline TR:TREATMENT_SUMMARY injection on day 25, plasma samples were collected via saphenous vein bleeding. TR:TREATMENT_SUMMARY A bolus of deuterium oxide (D2O, 10 ml/kg body weight, Sigma Aldrich) was then TR:TREATMENT_SUMMARY given by intraperitoneal injection and followed by free access to drinking water TR:TREATMENT_SUMMARY supplemented with 4% D2O for the rest of the experimental period. Saphenous TR:TREATMENT_SUMMARY plasma samples were collected again one day after the bolus delivery of D2O (day TR:TREATMENT_SUMMARY 27). #SAMPLEPREP SP:SAMPLEPREP_SUMMARY To prepare samples for analyses, frozen livers were pulverized under liquid SP:SAMPLEPREP_SUMMARY nitrogen, and weighed aliquots of the powder (50 mg powdered tissue/aliquot) SP:SAMPLEPREP_SUMMARY were transferred into pre-frozen Eppendorf tubes for further homogenization at SP:SAMPLEPREP_SUMMARY 50 mg of wet tissue per 1 ml of homogenate using the following buffers: 50% SP:SAMPLEPREP_SUMMARY acetonitrile/0.3% formic acid for the analysis of amino acids, acylcarnitines, SP:SAMPLEPREP_SUMMARY organic acids, creatine, and phosphocreatine; 3M perchloric acid for the SP:SAMPLEPREP_SUMMARY analysis of branched-chain keto acids; 0.3M perchloric acid for the analysis of SP:SAMPLEPREP_SUMMARY short-chain acyl CoAs; 100% methanol for the analysis of nucleotides. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY MS/MS analysis of amino acid and acylcarnitines CH:INSTRUMENT_NAME Waters Acquity I-Class CH:COLUMN_NAME N/A CH:COLUMN_TEMPERATURE N/A CH:FLOW_GRADIENT isocratic CH:FLOW_RATE 0.1 ml/min CH:SOLVENT_A 80% methanol CH:SOLVENT_B 80% methanol CH:CHROMATOGRAPHY_TYPE None (Direct infusion) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Waters Xevo-TQ MS:INSTRUMENT_TYPE Triple quadrupole MS:MS_TYPE ESI MS:MS_COMMENTS Mass spectra for acylcarnitine and amino acid esters were obtained using MS:MS_COMMENTS precursor ion (parents of m/z = 99) and neutral loss (NTL 102 amu for acidic and MS:MS_COMMENTS neutral amino acids; NTL 119 amu for basic amino acids; and NTL 161 amu for MS:MS_COMMENTS arginine) scanning methods, respectively. The spectra were acquired in a MS:MS_COMMENTS multi-channel analyzer (MCA) mode to improve signal-to-noise. Metabolite MS:MS_COMMENTS concentrations were computed using a ratio to heavy isotope-labeled internal MS:MS_COMMENTS standards. NeoLynx was used for data processing. MS:ION_MODE POSITIVE #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS µM MS_METABOLITE_DATA_START Samples 1 19 21 3 6 7 15 17 18 2 10 11 13 4 9 Factors Treatment:GalNAc-siChrebp | Sample source:liver Treatment:GalNAc-siChrebp | Sample source:liver Treatment:GalNAc-siChrebp | Sample source:liver Treatment:GalNAc-siChrebp | Sample source:liver Treatment:GalNAc-siChrebp | Sample source:liver Treatment:GalNAc-siChrebp | Sample source:liver Treatment:GalNAc-siCtrl | Sample source:liver Treatment:GalNAc-siCtrl | Sample source:liver Treatment:GalNAc-siCtrl | Sample source:liver Treatment:GalNAc-siCtrl | Sample source:liver Treatment:Sal | Sample source:liver Treatment:Sal | Sample source:liver Treatment:Sal | Sample source:liver Treatment:Sal | Sample source:liver Treatment:Sal | Sample source:liver Alanine 33.7732 41.4567 44.4321 44.8868 67.9468 53.1948 38.3372 36.3803 46.6420 47.3370 Arginine 2.1093 0.8292 1.2361 1.7942 1.5316 1.3926 1.0069 0.8594 1.1199 1.6982 Aspartic acid/Asparagine 17.5308 28.6530 23.5896 25.6973 28.8286 33.1171 38.8894 49.7706 68.6853 73.1212 CAR 10:0 0.0300 0.0272 0.0229 0.0280 0.0233 0.0242 0.0134 0.0142 0.0192 0.0226 0.0169 0.0187 0.0119 0.0138 0.0120 CAR 10:1 0.0055 0.0096 0.0079 0.0064 0.0074 0.0079 0.0064 0.0056 0.0100 0.0075 0.0084 0.0065 0.0056 0.0046 0.0065 CAR 10:2 0.0015 0.0013 0.0021 0.0024 0.0024 0.0028 0.0033 0.0010 0.0016 0.0024 0.0024 0.0019 CAR 10:3 0.0029 0.0071 0.0038 0.0041 0.0059 0.0042 0.0021 0.0020 0.0058 0.0033 0.0046 0.0055 0.0023 0.0014 0.0045 CAR 12:0 0.0649 0.0463 0.0210 0.0503 0.0201 0.0400 0.0127 0.0075 0.0141 0.0189 0.0183 0.0131 0.0116 0.0120 0.0091 CAR 12:1 0.0203 0.0169 0.0098 0.0135 0.0092 0.0143 0.0060 0.0061 0.0065 0.0074 0.0051 0.0094 0.0053 0.0033 0.0072 CAR 12:2 0.0016 0.0018 0.0008 0.0021 0.0022 0.0016 0.0022 0.0010 0.0018 0.0006 CAR 14:0 0.1400 0.1171 0.0562 0.1219 0.0454 0.1021 0.0234 0.0128 0.0266 0.0444 0.0275 0.0229 0.0171 0.0239 0.0282 CAR 14:1 0.0374 0.0360 0.0150 0.0362 0.0152 0.0323 0.0068 0.0072 0.0116 0.0128 0.0082 0.0092 0.0051 0.0048 0.0077 CAR 14:2 0.0087 0.0058 0.0036 0.0080 0.0048 0.0101 0.0024 0.0041 0.0043 0.0027 0.0014 0.0039 0.0017 0.0014 0.0010 CAR 16:0 0.4969 0.3979 0.1681 0.4168 0.1692 0.3568 0.0869 0.0459 0.0946 0.1163 0.1143 0.0880 0.0601 0.0797 0.1041 CAR 16:1 0.1211 0.1010 0.0497 0.1069 0.0434 0.0873 0.0253 0.0132 0.0270 0.0334 0.0234 0.0291 0.0164 0.0140 0.0185 CAR 16:2 0.0289 0.0274 0.0113 0.0298 0.0123 0.0268 0.0051 0.0028 0.0083 0.0083 0.0040 0.0057 0.0028 0.0019 0.0049 CAR 16:3 0.0021 0.0017 0.0006 0.0013 0.0023 0.0013 0.0013 0.0009 0.0011 CAR 18:0 0.2415 0.2140 0.0732 0.1891 0.0672 0.1460 0.0531 0.0348 0.0679 0.0928 0.0898 0.0536 0.0455 0.0644 0.0480 CAR 18:1 0.5595 0.4524 0.1861 0.4549 0.1709 0.3860 0.1082 0.0510 0.1336 0.1711 0.1270 0.1592 0.0681 0.0903 0.0982 CAR 18:2 0.1912 0.1511 0.0696 0.1617 0.0794 0.1464 0.0348 0.0239 0.0431 0.0595 0.0354 0.0457 0.0226 0.0241 0.0365 CAR 18:3 0.0222 0.0152 0.0096 0.0145 0.0066 0.0162 0.0043 0.0019 0.0036 0.0068 0.0045 0.0053 0.0030 0.0034 0.0034 CAR 2:0 6.7974 9.7698 11.4220 8.6832 8.6182 9.2038 5.6872 4.8869 5.6995 6.3434 5.3935 6.2722 5.5667 4.9855 5.9960 CAR 20:0 0.0098 0.0102 0.0038 0.0092 0.0019 0.0075 0.0026 0.0013 0.0034 0.0030 0.0036 0.0019 0.0023 0.0026 0.0026 CAR 20:1 0.0489 0.0386 0.0143 0.0405 0.0100 0.0294 0.0087 0.0034 0.0134 0.0098 0.0152 0.0073 0.0053 0.0077 0.0073 CAR 20:2 0.0373 0.0284 0.0111 0.0297 0.0087 0.0230 0.0073 0.0023 0.0087 0.0085 0.0109 0.0064 0.0043 0.0068 0.0051 CAR 20:3 0.0117 0.0122 0.0075 0.0105 0.0043 0.0102 0.0064 0.0019 0.0062 0.0073 0.0047 0.0066 0.0043 0.0030 0.0036 CAR 20:4 0.0075 0.0085 0.0049 0.0073 0.0051 0.0066 0.0060 0.0053 0.0045 0.0100 0.0075 0.0143 0.0036 0.0055 0.0055 CAR 22:0 0.0011 0.0009 0.0006 0.0011 0.0013 0.0006 0.0011 0.0004 0.0011 0.0019 0.0009 0.0009 CAR 22:1 0.0011 0.0013 0.0041 0.0009 0.0013 0.0009 0.0004 0.0011 0.0021 0.0015 0.0009 0.0013 0.0019 0.0015 CAR 22:2 0.0019 0.0009 0.0015 0.0004 0.0002 0.0009 0.0004 0.0006 CAR 22:3 0.0006 0.0015 0.0006 0.0004 0.0002 0.0015 CAR 22:4 0.0009 0.0023 0.0009 0.0004 0.0009 0.0011 0.0021 0.0009 0.0013 CAR 22:5 0.0009 0.0004 0.0011 0.0004 CAR 3:0 0.1184 0.1580 0.2065 0.1931 0.2264 0.1714 0.2294 0.1590 0.1883 0.2365 0.1721 0.1397 0.1921 0.1655 0.1350 CAR 4:0 0.1386 0.1755 0.1617 0.1685 0.1521 0.1821 0.3609 0.2634 0.3659 0.2650 0.4052 0.2529 0.3078 0.3834 0.3771 CAR 4:0;OH 0.0676 0.0810 0.0860 0.0817 0.0585 0.0720 0.0549 0.0320 0.0488 0.0411 0.0494 0.0497 0.0379 0.0398 0.0358 CAR 5:0 0.0375 0.0489 0.0401 0.0423 0.0357 0.0402 0.0434 0.0299 0.0312 0.0383 0.0500 0.0343 0.0521 0.0279 CAR 5:1 0.0445 0.0354 0.0415 0.0401 0.0343 0.0380 0.0490 0.0258 0.0489 0.0492 0.0501 0.0584 0.0486 0.0428 0.0424 CAR 6:0 0.0254 0.0219 0.0125 0.0235 0.0096 0.0209 0.0075 0.0041 0.0121 0.0087 0.0289 0.0106 0.0093 0.0301 0.0093 CAR 8:0 0.0430 0.0324 0.0337 0.0375 0.0273 0.0377 0.0172 0.0159 0.0202 0.0230 0.0178 0.0120 0.0115 0.0101 0.0118 CAR 8:1 0.0157 0.0277 0.0219 0.0193 0.0197 0.0195 0.0173 0.0187 0.0244 0.0222 0.0135 0.0168 0.0156 0.0094 0.0095 CAR DC10:0/CAR 12:0;OH [Methyl esters] 0.0157 0.0174 0.0063 0.0159 0.0043 0.0150 0.0029 0.0041 0.0051 0.0056 0.0024 0.0051 0.0010 0.0014 CAR DC10:1/CAR 12:1;OH [Methyl esters] 0.0092 0.0060 0.0029 0.0065 0.0027 0.0060 0.0029 0.0019 0.0019 0.0031 0.0027 0.0017 0.0022 0.0012 0.0031 CAR DC10:2/CAR 12:2;OH [Methyl esters] 0.0024 0.0027 0.0041 0.0034 0.0029 0.0022 0.0017 0.0019 0.0007 0.0017 0.0022 0.0022 0.0019 0.0012 CAR DC12:0/CAR 14:0;OH [Methyl esters] 0.0348 0.0355 0.0171 0.0338 0.0089 0.0340 0.0063 0.0080 0.0051 0.0082 0.0039 0.0065 0.0027 0.0022 0.0036 CAR DC12:1/CAR 14:1;OH [Methyl esters] 0.0270 0.0196 0.0077 0.0205 0.0058 0.0176 0.0027 0.0068 0.0056 0.0053 0.0041 0.0041 0.0014 0.0022 0.0048 CAR DC14:0/CAR 16:0;OH [Methyl esters] 0.0669 0.0686 0.0283 0.0591 0.0185 0.0625 0.0102 0.0087 0.0108 0.0166 0.0074 0.0113 0.0057 0.0032 0.0057 CAR DC14:1/CAR 16:1;OH [Methyl esters] 0.0453 0.0357 0.0189 0.0361 0.0123 0.0383 0.0072 0.0066 0.0089 0.0108 0.0040 0.0055 0.0032 0.0030 0.0055 CAR DC14:2/CAR 16:2;OH [Methyl esters] 0.0157 0.0102 0.0038 0.0138 0.0049 0.0145 0.0032 0.0026 0.0026 0.0038 0.0021 0.0036 0.0006 0.0009 0.0030 CAR DC16:0/CAR 18:0;OH [Methyl esters] 0.0361 0.0378 0.0111 0.0320 0.0085 0.0258 0.0087 0.0062 0.0087 0.0085 0.0064 0.0066 0.0041 0.0041 0.0041 CAR DC16:1/CAR 18:1;OH [Methyl esters] 0.0849 0.0706 0.0356 0.0715 0.0220 0.0734 0.0169 0.0100 0.0162 0.0273 0.0083 0.0132 0.0070 0.0049 0.0102 CAR DC16:2/CAR 18:2;OH [Methyl esters] 0.0275 0.0256 0.0137 0.0271 0.0107 0.0248 0.0079 0.0055 0.0062 0.0081 0.0041 0.0064 0.0036 0.0017 0.0043 CAR DC16:3/CAR 18:3;OH [Methyl esters] 0.0043 0.0026 0.0011 0.0032 0.0013 0.0049 0.0017 0.0013 0.0011 0.0006 CAR DC18:0/CAR 20:0;OH [Methyl esters] 0.0013 0.0017 0.0006 0.0017 0.0013 0.0009 0.0009 0.0004 0.0006 0.0009 0.0009 0.0006 CAR DC18:1/CAR 20:1;OH [Methyl esters] 0.0098 0.0081 0.0038 0.0060 0.0023 0.0079 0.0017 0.0013 0.0028 0.0036 0.0019 0.0021 0.0030 0.0017 0.0030 CAR DC18:2/CAR 20:2;OH [Methyl esters] 0.0064 0.0062 0.0030 0.0060 0.0026 0.0060 0.0021 0.0013 0.0017 0.0032 0.0017 0.0009 0.0013 0.0017 0.0019 CAR DC18:3/CAR 20:3;OH [Methyl esters] 0.0017 0.0030 0.0009 0.0023 0.0013 0.0011 0.0013 0.0011 0.0006 0.0006 0.0015 CAR DC3:0/CAR 5:0;OH [Methyl esters] 0.0319 0.0465 0.0409 0.0381 0.0481 0.0436 0.0410 0.0297 0.0340 0.0345 0.0463 0.0470 0.0315 0.0356 0.0296 CAR DC4:0/CAR DC3:0;2Me [Methyl esters] 0.0119 0.0110 0.0144 0.0125 0.0085 0.0096 0.0066 0.0080 0.0079 0.0072 0.0109 0.0079 0.0101 0.0076 0.0055 CAR DC5:0 0.0253 0.0263 0.0225 0.0258 0.0199 0.0260 0.0218 0.0277 0.0281 0.0213 0.0218 0.0209 CAR DC6:0/CAR 8:0;OH [Methyl esters] 0.0377 0.0369 0.0339 0.0333 0.0233 0.0313 0.0150 0.0140 0.0253 0.0195 0.0137 0.0173 0.0194 0.0101 0.0109 CAR DC6:1/CAR 8:1;OH [Methyl esters] 0.0395 0.0463 0.0493 0.0448 0.0436 0.0476 0.0286 0.0262 0.0430 0.0298 0.0232 0.0288 0.0177 0.0180 0.0151 CAR DC7:0 0.0118 0.0101 0.0186 0.0106 0.0097 0.0119 0.0070 0.0064 0.0072 0.0093 0.0076 0.0050 0.0054 0.0033 0.0048 CAR DC8:0/CAR 10:0;OH [Methyl esters] 0.0123 0.0116 0.0131 0.0124 0.0055 0.0143 0.0042 0.0049 0.0055 0.0068 0.0074 0.0038 0.0032 0.0041 CAR DC8:1 0.0093 0.0114 0.0139 0.0097 0.0089 0.0084 0.0057 0.0120 0.0096 0.0110 0.0090 0.0093 0.0041 0.0049 Citrulline 0.8818 0.8893 0.9125 0.8543 0.7562 0.8031 0.8780 0.9896 1.2030 0.8052 Glutamic acid/Glutamine 99.7087 148.7152 153.9213 146.2536 140.1309 156.7284 119.6568 101.9325 142.2709 168.4019 Glycine 35.6760 43.3744 36.4055 42.6712 40.7994 41.3441 67.7966 46.6062 51.1586 61.1678 Histidine 22.7312 25.1919 26.4922 26.8011 26.3119 26.4866 28.3304 23.6851 25.2918 27.7064 Leucine/Isoleucine 8.7492 9.9800 9.4324 9.3075 9.7869 9.4388 13.8691 11.5147 11.9346 14.2802 Methionine 2.9620 3.2153 3.0481 2.9190 3.1755 3.5471 2.9593 2.8765 2.5533 3.4062 Ornithine 7.0657 7.7265 8.8644 7.6108 6.8622 7.7402 13.9591 11.0449 11.9108 14.4505 Phenylalanine 2.6778 2.8554 3.1009 3.0903 3.3426 3.2030 4.1163 3.2107 3.4423 4.3989 Proline 5.0353 7.4882 6.1660 6.4772 6.3878 6.0123 8.1055 8.0389 6.5219 8.3780 Serine 39.0075 45.5671 41.6468 46.6492 45.4578 46.2399 57.8684 33.1565 59.6916 58.1083 Tyrosine 2.7602 2.3838 2.6058 3.1877 2.2610 2.9201 4.4530 4.9252 5.5140 4.8321 Valine 9.3391 10.7152 10.4068 10.5580 10.7300 10.6458 13.2323 11.7388 10.7687 13.6200 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name pubchem_id inchi_key kegg_id other_id other_id_type ri ri_type moverz_quant Alanine Arginine Aspartic acid/Asparagine CAR 10:0 CAR 10:1 CAR 10:2 CAR 10:3 CAR 12:0 CAR 12:1 CAR 12:2 CAR 14:0 CAR 14:1 CAR 14:2 CAR 16:0 CAR 16:1 CAR 16:2 CAR 16:3 CAR 18:0 CAR 18:1 CAR 18:2 CAR 18:3 CAR 2:0 CAR 20:0 CAR 20:1 CAR 20:2 CAR 20:3 CAR 20:4 CAR 22:0 CAR 22:1 CAR 22:2 CAR 22:3 CAR 22:4 CAR 22:5 CAR 3:0 CAR 4:0 CAR 4:0;OH CAR 5:0 CAR 5:1 CAR 6:0 CAR 8:0 CAR 8:1 CAR DC10:0/CAR 12:0;OH [Methyl esters] CAR DC10:1/CAR 12:1;OH [Methyl esters] CAR DC10:2/CAR 12:2;OH [Methyl esters] CAR DC12:0/CAR 14:0;OH [Methyl esters] CAR DC12:1/CAR 14:1;OH [Methyl esters] CAR DC14:0/CAR 16:0;OH [Methyl esters] CAR DC14:1/CAR 16:1;OH [Methyl esters] CAR DC14:2/CAR 16:2;OH [Methyl esters] CAR DC16:0/CAR 18:0;OH [Methyl esters] CAR DC16:1/CAR 18:1;OH [Methyl esters] CAR DC16:2/CAR 18:2;OH [Methyl esters] CAR DC16:3/CAR 18:3;OH [Methyl esters] CAR DC18:0/CAR 20:0;OH [Methyl esters] CAR DC18:1/CAR 20:1;OH [Methyl esters] CAR DC18:2/CAR 20:2;OH [Methyl esters] CAR DC18:3/CAR 20:3;OH [Methyl esters] CAR DC3:0/CAR 5:0;OH [Methyl esters] CAR DC4:0/CAR DC3:0;2Me [Methyl esters] CAR DC5:0 CAR DC6:0/CAR 8:0;OH [Methyl esters] CAR DC6:1/CAR 8:1;OH [Methyl esters] CAR DC7:0 CAR DC8:0/CAR 10:0;OH [Methyl esters] CAR DC8:1 Citrulline Glutamic acid/Glutamine Glycine Histidine Leucine/Isoleucine Methionine Ornithine Phenylalanine Proline Serine Tyrosine Valine METABOLITES_END #END