#METABOLOMICS WORKBENCH nmatulionis_20230907_122351 DATATRACK_ID:4296 STUDY_ID:ST002852 ANALYSIS_ID:AN004672 PROJECT_ID:PR001786 VERSION 1 CREATED_ON September 7, 2023, 1:44 pm #PROJECT PR:PROJECT_TITLE MYC is a regulator of androgen receptor inhibition-induced metabolic PR:PROJECT_TITLE requirements in prostate cancer PR:PROJECT_SUMMARY Advanced prostate cancers are treated with therapies targeting the androgen PR:PROJECT_SUMMARY receptor (AR) signaling pathway. While many tumors initially respond to AR PR:PROJECT_SUMMARY inhibition, nearly all develop resistance. It is critical to understand how PR:PROJECT_SUMMARY prostate tumor cells respond to AR inhibition in order to exploit PR:PROJECT_SUMMARY therapy-induced phenotypes prior to the outgrowth of treatment-resistant PR:PROJECT_SUMMARY disease. Here, we comprehensively characterize the effect of AR blockade on PR:PROJECT_SUMMARY prostate cancer metabolism using transcriptomics, metabolomics and bioenergetics PR:PROJECT_SUMMARY approaches. The metabolic response to AR inhibition is defined by reduced PR:PROJECT_SUMMARY glycolysis, robust elongation of mitochondria, and increased reliance on PR:PROJECT_SUMMARY mitochondrial oxidative metabolism. We establish DRP1 activity and MYC signaling PR:PROJECT_SUMMARY as mediators of AR blockade-induced metabolic phenotypes. Rescuing DRP1 PR:PROJECT_SUMMARY phosphorylation after AR inhibition restores mitochondrial fission, while PR:PROJECT_SUMMARY rescuing MYC restores glycolytic activity and prevents sensitivity to complex I PR:PROJECT_SUMMARY inhibition. Our study provides new insight into the regulation of PR:PROJECT_SUMMARY treatment-induced metabolic phenotypes and vulnerabilities in prostate cancer. PR:INSTITUTE University of California, Los Angeles PR:DEPARTMENT Biological Chemistry PR:LABORATORY Heather Christofk PR:LAST_NAME Matulionis PR:FIRST_NAME Nedas PR:ADDRESS 615 Charles E Young Dr S, BSRB 354-05 PR:EMAIL nmatulionis@mednet.ucla.edu PR:PHONE 3102060163 #STUDY ST:STUDY_TITLE MYC is a regulator of androgen receptor inhibition-induced metabolic ST:STUDY_TITLE requirements in prostate cancer ST:STUDY_SUMMARY Advanced prostate cancers are treated with therapies targeting the androgen ST:STUDY_SUMMARY receptor (AR) signaling pathway. While many tumors initially respond to AR ST:STUDY_SUMMARY inhibition, nearly all develop resistance. It is critical to understand how ST:STUDY_SUMMARY prostate tumor cells respond to AR inhibition in order to exploit ST:STUDY_SUMMARY therapy-induced phenotypes prior to the outgrowth of treatment-resistant ST:STUDY_SUMMARY disease. Here, we comprehensively characterize the effect of AR blockade on ST:STUDY_SUMMARY prostate cancer metabolism using transcriptomics, metabolomics and bioenergetics ST:STUDY_SUMMARY approaches. The metabolic response to AR inhibition is defined by reduced ST:STUDY_SUMMARY glycolysis, robust elongation of mitochondria, and increased reliance on ST:STUDY_SUMMARY mitochondrial oxidative metabolism. We establish DRP1 activity and MYC signaling ST:STUDY_SUMMARY as mediators of AR blockade-induced metabolic phenotypes. Rescuing DRP1 ST:STUDY_SUMMARY phosphorylation after AR inhibition restores mitochondrial fission, while ST:STUDY_SUMMARY rescuing MYC restores glycolytic activity and prevents sensitivity to complex I ST:STUDY_SUMMARY inhibition. Our study provides new insight into the regulation of ST:STUDY_SUMMARY treatment-induced metabolic phenotypes and vulnerabilities in prostate cancer. ST:INSTITUTE University of California, Los Angeles ST:DEPARTMENT Biological Chemistry ST:LABORATORY Heather Christofk ST:LAST_NAME Matulionis ST:FIRST_NAME Nedas ST:ADDRESS 615 Charles E Young Dr S, BSRB 354-05 ST:EMAIL nmatulionis@mednet.ucla.edu ST:PHONE 3102060163 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:GENOTYPE_STRAIN NSG #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 - tumor-veh-1-01 Treatment:Vehicle | Technical Replicate (Region of Tumor):1 RAW_FILE_NAME=tumor-veh-1-01.RAW SUBJECT_SAMPLE_FACTORS - tumor-veh-1-02 Treatment:Vehicle | Technical Replicate (Region of Tumor):2 RAW_FILE_NAME=tumor-veh-1-02.RAW SUBJECT_SAMPLE_FACTORS - tumor-veh-2-01 Treatment:Vehicle | Technical Replicate (Region of Tumor):1 RAW_FILE_NAME=tumor-veh-2-01.RAW SUBJECT_SAMPLE_FACTORS - tumor-veh-2-02 Treatment:Vehicle | Technical Replicate (Region of Tumor):2 RAW_FILE_NAME=tumor-veh-2-02.RAW SUBJECT_SAMPLE_FACTORS - tumor-veh-3-01 Treatment:Vehicle | Technical Replicate (Region of Tumor):1 RAW_FILE_NAME=tumor-veh-3-01.RAW SUBJECT_SAMPLE_FACTORS - tumor-veh-3-02 Treatment:Vehicle | Technical Replicate (Region of Tumor):2 RAW_FILE_NAME=tumor-veh-3-02.RAW SUBJECT_SAMPLE_FACTORS - tumor-enzalutamide-1-01 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):1 RAW_FILE_NAME=tumor-enzalutamide-1-01.RAW SUBJECT_SAMPLE_FACTORS - tumor-enzalutamide-1-02 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):2 RAW_FILE_NAME=tumor-enzalutamide-1-02.RAW SUBJECT_SAMPLE_FACTORS - tumor-enzalutamide-2-01 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):1 RAW_FILE_NAME=tumor-enzalutamide-2-01.RAW SUBJECT_SAMPLE_FACTORS - tumor-enzalutamide-2-02 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):2 RAW_FILE_NAME=tumor-enzalutamide-2-02.RAW SUBJECT_SAMPLE_FACTORS - tumor-enzalutamide-3-01 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):1 RAW_FILE_NAME=tumor-enzalutamide-3-01.RAW SUBJECT_SAMPLE_FACTORS - tumor-enzalutamide-3-02 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):2 RAW_FILE_NAME=tumor-enzalutamide-3-02.RAW SUBJECT_SAMPLE_FACTORS - tumor-enzalutamide-4-01 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):1 RAW_FILE_NAME=tumor-enzalutamide-4-01.RAW SUBJECT_SAMPLE_FACTORS - tumor-enzalutamide-4-02 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):2 RAW_FILE_NAME=tumor-enzalutamide-4-02.RAW SUBJECT_SAMPLE_FACTORS - tumor-enzalutamide-5-01 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):1 RAW_FILE_NAME=tumor-enzalutamide-5-01.RAW SUBJECT_SAMPLE_FACTORS - tumor-enzalutamide-5-02 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):2 RAW_FILE_NAME=tumor-enzalutamide-5-02.RAW #COLLECTION CO:COLLECTION_SUMMARY Tumors were collected after 10 days of treatment and prepared for histology, CO:COLLECTION_SUMMARY protein extraction, and metabolite extraction. CO:SAMPLE_TYPE Prostate #TREATMENT TR:TREATMENT_SUMMARY 7 million 16D cells were implanted subcutaneously with 100 μl Matrigel TR:TREATMENT_SUMMARY (Corning) into NSG mice to form primary tumors. Primary tumors were harvested, TR:TREATMENT_SUMMARY minced, and re-implanted (20 - 80 mg of minced tumor tissue with 100 μl TR:TREATMENT_SUMMARY Matrigel per mouse) into NSG mice. 16D tumor-bearing mice were treated by oral TR:TREATMENT_SUMMARY gavage with 10 mg/kg/day of Enzalutamide in the vehicle (1% carboxymethyl TR:TREATMENT_SUMMARY cellulose, 0.5% Tween 80, and 5% dimethylsulfoxide) or the vehicle only, with a TR:TREATMENT_SUMMARY two-days-on/one-day-off schedule. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY After tumor dissection, a maximum of 30mg of tissue was weighed, snap frozen, SP:SAMPLEPREP_SUMMARY and stored at -80C until metabolite extraction. To extract metabolites, SP:SAMPLEPREP_SUMMARY weighed tumor tissue was added to a bead tube (Thermo Fisher Scientific) SP:SAMPLEPREP_SUMMARY containing 1ml 80% methanol plus 10mM potassium trifluoromethanesulfonate (TMSO) SP:SAMPLEPREP_SUMMARY internal standard on ice. Samples were homogenized for 1 minute at max speed on SP:SAMPLEPREP_SUMMARY a bead homogenizer (Thermo Fisher Scientific). Bead tubes were spun at 17000g at SP:SAMPLEPREP_SUMMARY 4C for 10 minutes. The supernatant was transferred to an Eppendorf tube and SP:SAMPLEPREP_SUMMARY spun at 17000g at 4C for 10 minutes. A volume of extraction equivalent to 3mg SP:SAMPLEPREP_SUMMARY of tumor tissue was transferred to an ABC vial (Thermo Fisher Scientific). All SP:SAMPLEPREP_SUMMARY volumes were normalized to 500 µl with 80% methanol containing TMSO internal SP:SAMPLEPREP_SUMMARY standard. 80% MeOH was evaporated from the ABC vials using the EZ-2Elite SP:SAMPLEPREP_SUMMARY evaporator (Genevac) and samples were stored at -80°C until analysis. Dried SP:SAMPLEPREP_SUMMARY metabolites were reconstituted in 100 µL of a 50% acetonitrile (ACN) 50% dH20 SP:SAMPLEPREP_SUMMARY solution. Samples were vortexed and spun down for 10 minutes at 17,000g. 70 µL SP:SAMPLEPREP_SUMMARY of the supernatant was then transferred to HPLC glass vials. 10 µL of these SP:SAMPLEPREP_SUMMARY metabolite solutions were injected per analysis. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Samples were run on a Vanquish (Thermo Fisher Scientific) UHPLC system with CH:CHROMATOGRAPHY_SUMMARY mobile phase A (20mM ammonium carbonate, pH 9.7) and mobile phase B (100% ACN) CH:CHROMATOGRAPHY_SUMMARY at a flow rate of 150 µL/min on a SeQuant ZIC-pHILIC Polymeric column (2.1 × CH:CHROMATOGRAPHY_SUMMARY 150 mm 5 μm, EMD Millipore) at 35°C. Separation was achieved with a linear CH:CHROMATOGRAPHY_SUMMARY gradient from 20% A to 80% A in 20 minutes followed by a linear gradient from CH:CHROMATOGRAPHY_SUMMARY 80% A to 20% A from 20 minutes to 20.5 minutes. 20% A was then held from 20.5 CH:CHROMATOGRAPHY_SUMMARY minutes to 28 minutes. CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME Thermo Vanquish CH:COLUMN_NAME SeQuant ZIC- pHILIC (150 x 2.1mm,5um) CH:SOLVENT_A 20 mM Ammonium carbonate, pH 9.7 CH:SOLVENT_B 100% Acetonitrile CH:FLOW_GRADIENT Check Summary CH:FLOW_RATE 150 µL/min CH:COLUMN_TEMPERATURE 35°C #ANALYSIS AN:ANALYSIS_TYPE MS AN:LABORATORY_NAME Heather Christofk AN:OPERATOR_NAME Nedas Matulionis #MS MS:INSTRUMENT_NAME Thermo Q Exactive Orbitrap MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS The UHPLC was coupled to a Q-Exactive (Thermo Fisher Scientific) mass analyzer MS:MS_COMMENTS running in polarity switching mode with spray-voltage=3.2kV, sheath-gas=40, MS:MS_COMMENTS aux-gas=15, sweep-gas=1, aux-gas-temp=350°C, and capillary-temp=275°C. For MS:MS_COMMENTS both polarities mass scan settings were kept at full-scan-range=(70-1000), MS:MS_COMMENTS ms1-resolution=70,000, max-injection-time=250ms, and AGC-target=1E6. MS2 data MS:MS_COMMENTS was also collected from the top three most abundant singly-charged ions in each MS:MS_COMMENTS scan with normalized-collision-energy=35. Each of the resulting “.RAW” files MS:MS_COMMENTS was then centroided and converted into two “.mzXML” files (one for positive MS:MS_COMMENTS scans and one for negative scans) using msconvert from ProteoWizard12. These MS:MS_COMMENTS “.mzXML” files were imported into the MZmine 2 software package13. Ion MS:MS_COMMENTS chromatograms were generated from MS1 spectra via the built-in Automated Data MS:MS_COMMENTS Analysis Pipeline (ADAP) chromatogram module14 and peaks were detected via the MS:MS_COMMENTS ADAP wavelets algorithm. Peaks were aligned across all samples via the Random MS:MS_COMMENTS sample consensus aligner module, gap-filled, and assigned identities using an MS:MS_COMMENTS exact mass MS1(+/-15ppm) and retention time RT (+/-0.5min) search of our MS:MS_COMMENTS in-house MS1-RT database. Peak boundaries and identifications were then further MS:MS_COMMENTS refined by manual curation. Peaks were quantified by area under the curve MS:MS_COMMENTS integration and exported as CSV files. If stable isotope tracing was used in the MS:MS_COMMENTS experiment, the peak areas were additionally processed via the R package MS:MS_COMMENTS AccuCor15 to correct for natural isotope abundance. Peak areas for each sample MS:MS_COMMENTS were normalized by the measured area of the internal standard MS:MS_COMMENTS trifluoromethanesulfonate (present in the extraction buffer) and by the number MS:MS_COMMENTS of cells present in the extracted well. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS Peak Area MS_METABOLITE_DATA_START Samples tumor-enzalutamide-1-01 tumor-enzalutamide-1-02 tumor-enzalutamide-2-01 tumor-enzalutamide-2-02 tumor-enzalutamide-3-01 tumor-enzalutamide-3-02 tumor-enzalutamide-4-01 tumor-enzalutamide-4-02 tumor-enzalutamide-5-01 tumor-enzalutamide-5-02 tumor-veh-1-01 tumor-veh-1-02 tumor-veh-2-01 tumor-veh-2-02 tumor-veh-3-01 tumor-veh-3-02 Factors Treatment:Enzalutamide | Technical Replicate (Region of Tumor):1 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):2 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):1 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):2 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):1 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):2 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):1 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):2 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):1 Treatment:Enzalutamide | Technical Replicate (Region of Tumor):2 Treatment:Vehicle | Technical Replicate (Region of Tumor):1 Treatment:Vehicle | Technical Replicate (Region of Tumor):2 Treatment:Vehicle | Technical Replicate (Region of Tumor):1 Treatment:Vehicle | Technical Replicate (Region of Tumor):2 Treatment:Vehicle | Technical Replicate (Region of Tumor):1 Treatment:Vehicle | Technical Replicate (Region of Tumor):2 3-hydroxy-trimethyllysine 1312316.718 647802.298 1292392.066 698006.8246 167884.0447 683301.1378 1711586.998 739687.917 356027.1388 825561.2761 250650.3541 832683.1756 1549524.63 1756604.776 319567.286 266805.1144 5'-methylthioadenosine 4064835.233 2976798.684 5816087.13 4217810.022 6170014.125 3526258.535 7678740.506 3810675.021 5015010.547 6800873.709 864900.6026 1799790.437 2994224.605 3486801.921 3252875.178 3492262.558 acetyl-carnitine 6.08E+08 4.50E+08 1.13E+09 6.67E+08 7.18E+08 6.54E+08 1.26E+09 8.68E+08 7.40E+08 9.22E+08 2.45E+08 3.62E+08 6.81E+08 8.31E+08 4.47E+08 3.82E+08 AIR 1504774.5 1721530.088 1678955.464 1813852.199 1667619.592 1591378.193 1639595.091 1901898.362 1488784.281 1771029.354 470861.9297 940613.6363 1271603.68 1581062.101 1055739.742 1877741.075 alanine 2.90E+07 2.34E+07 3.54E+07 2.52E+07 3.97E+07 2.87E+07 5.34E+07 3.23E+07 3.82E+07 4.95E+07 1.67E+07 1.96E+07 3.60E+07 4.35E+07 4.50E+07 3.73E+07 arginine 6.02E+07 4.17E+07 7.34E+07 5.79E+07 1.02E+08 6.39E+07 8.36E+07 5.43E+07 5.38E+07 6.59E+07 2.10E+07 2.83E+07 4.78E+07 5.32E+07 5.00E+07 4.02E+07 asparagine 1.18E+07 6613895.9 1.47E+07 9077568.528 1.80E+07 1.23E+07 2.21E+07 1.18E+07 1.68E+07 2.14E+07 3110170.765 4265958.409 8474477.925 1.26E+07 1.35E+07 7290741.155 betaine 8.84E+07 1.28E+08 3.20E+08 1.48E+08 9.03E+07 1.09E+08 1.43E+08 1.04E+08 1.26E+08 1.92E+08 9.34E+07 1.05E+08 4.92E+07 9.30E+07 9.75E+07 1.39E+08 carnitine 9.76E+08 8.00E+08 2.10E+09 1.20E+09 1.43E+09 9.84E+08 1.93E+09 1.03E+09 8.29E+08 1.38E+09 2.46E+08 4.42E+08 6.06E+08 9.14E+08 9.43E+08 9.70E+08 CDP-choline 4539646.153 3042834.699 1.01E+07 5286932.496 1.34E+07 6476831.876 1.78E+07 7450382.737 8067363.222 7637937.253 981577.5532 1504770.432 5200458.775 6120016.578 5468865.735 6743871.259 choline 1.98E+08 1.79E+08 3.17E+08 2.08E+08 3.22E+08 2.35E+08 3.64E+08 2.50E+08 2.49E+08 2.94E+08 6.98E+07 8.03E+07 8.42E+07 1.43E+08 1.03E+08 1.62E+08 citrulline 1.42E+07 1.58E+07 1.45E+07 1.04E+07 1.64E+07 1.63E+07 1.02E+07 7912524.719 1.19E+07 1.39E+07 1.36E+07 1.59E+07 1.85E+07 2.08E+07 4.28E+07 2.73E+07 creatine 3.15E+09 2.59E+09 3.38E+09 2.75E+09 2.95E+09 2.59E+09 3.33E+09 2.66E+09 2.84E+09 3.14E+09 1.86E+09 2.46E+09 3.46E+09 3.75E+09 3.82E+09 3.66E+09 creatinine 1.11E+07 8360789.694 1.69E+07 1.03E+07 1.87E+07 1.32E+07 2.11E+07 1.08E+07 1.32E+07 1.82E+07 6359142.641 6147158.581 1.01E+07 1.60E+07 1.26E+07 1.63E+07 cystathionine 9631730.665 5884225.307 1.44E+07 1.06E+07 2.41E+07 7198701.099 1.84E+07 8333992.46 1.38E+07 1.75E+07 2024114.304 5051831.181 1.89E+07 1.59E+07 2.31E+07 5.15E+07 cysteine-glutathione disulfide 1123196.573 1053262.672 1351679.673 1403431.773 2489982.093 1982866.377 902803.8833 1169626.767 1555719.416 1284928.517 1170034.048 845379.047 1225223.595 1869788.029 917883.436 957584.5394 gamma-butyrobetaine 2.38E+07 2.26E+07 3.85E+07 2.05E+07 2.82E+07 2.50E+07 3.51E+07 1.66E+07 1.34E+07 2.14E+07 6631001.649 1.34E+07 1.28E+07 2.03E+07 1.90E+07 1.90E+07 gamma-glutamylcysteine 187058.7627 179131.4502 141783.7557 151129.7203 150255.7917 192385.348 211688.4243 139339.3128 156240.0927 178622.5486 12404.11121 91343.78353 117054.6516 184626.8302 0 146509.9484 glycerophosphocholine 7.38E+08 7.71E+08 1.58E+09 1.24E+09 2.85E+09 1.38E+09 2.12E+09 1.31E+09 1.68E+09 1.95E+09 1.71E+08 2.06E+08 5.56E+08 5.58E+08 3.48E+08 1.08E+09 glycine 3936427.623 2574561.455 4760783.638 3100845.598 7178951.365 3852515.294 5552982.345 3068573.837 3401280.483 4453352.341 2050736.213 2868563.519 5300355.456 8112374.927 6798496.605 6594900.371 kynurenic acid 27107.21095 38268.23742 109428.247 127333.611 120762.1011 181538.8622 148764.9303 101922.3763 70983.3756 100741.0784 12768.40723 7500.910156 118275.9026 107010.4912 2526.980469 130298.7682 kynurenine 466514.1448 296671.493 653876.8824 249736.1479 167122.8073 194185.9563 397697.6268 331029.3587 92261.219 350649.3403 248173.7715 322644.4686 421960.4201 689098.9758 110161.4733 206206.0638 L-2-aminobutyric acid 1419548.284 1138338.969 1678027.594 1543955.188 1898375.376 1082745.11 1373651.516 930554.9756 690221.6034 830897.037 8160964.545 2132841.617 1641428.486 2064455.625 1686786.178 2143099.697 leucine / isoleucine 4.70E+07 3.94E+07 6.98E+07 5.30E+07 8.70E+07 7.40E+07 8.28E+07 5.45E+07 5.14E+07 5.79E+07 2.49E+07 1.59E+07 3.52E+07 5.58E+07 4.39E+07 5.84E+07 lysine 3.62E+07 3.16E+07 5.69E+07 3.89E+07 7.79E+07 5.23E+07 5.89E+07 3.44E+07 4.41E+07 4.80E+07 1.59E+07 2.04E+07 3.10E+07 5.68E+07 4.37E+07 5.82E+07 methionine 2.73E+07 2.50E+07 3.94E+07 3.00E+07 5.99E+07 4.37E+07 3.20E+07 2.45E+07 1.89E+07 2.45E+07 1.37E+07 1.11E+07 1.88E+07 3.13E+07 2.61E+07 3.54E+07 methioninesulfoxide 3861847.559 2748344.2 4377804.095 3585067.284 6878669.881 3430593.559 3250986.409 2047409.455 2094907.178 2558324.036 371578.3461 694585.802 1127758.371 1483866.869 1072537.387 1555352.155 NAD+ 5540622.975 3693003.379 8192040.33 5307513.7 8411715.656 4297764.435 1.02E+07 3949958.454 5523746.254 9628978.335 770924.4883 3121451.272 4191589.688 6142301.625 7129242.875 4176312.438 NADP+ 539307.6957 315389.2439 901608.7809 491765.2036 803859.8056 387107.7708 892161.7842 406810.6283 495180.76 817449.1538 49046.42919 194445.7727 219594.5091 545440.2252 434576.0352 348062.231 nicotinamide 2.37E+08 2.26E+08 2.73E+08 2.27E+08 3.29E+08 3.39E+08 2.94E+08 3.70E+08 2.45E+08 2.44E+08 1.16E+08 1.21E+08 2.92E+08 2.93E+08 3.13E+08 3.43E+08 N-trimethyllysine 6588826.901 5078466.493 8678801.558 7423019.201 1.89E+07 1.03E+07 1.18E+07 8762414.957 1.22E+07 1.42E+07 4361856.082 4840433.732 7751566.895 1.27E+07 1.55E+07 1.68E+07 ophthalmate 0 0 0 0 19171.4342 0 0 22985.0428 0 0 0 0 0 35517.59583 15493.53882 22856.33011 ornithine 1.10E+07 1.30E+07 1.39E+07 1.26E+07 2.05E+07 2.23E+07 1.72E+07 1.33E+07 1.60E+07 1.94E+07 9402357.457 1.01E+07 1.07E+07 1.85E+07 1.55E+07 2.68E+07 phenylalanine 3.17E+07 2.49E+07 5.24E+07 3.45E+07 6.31E+07 5.27E+07 6.30E+07 3.96E+07 3.36E+07 3.60E+07 1.92E+07 9338889.129 2.50E+07 3.98E+07 3.06E+07 5.00E+07 phosphocholine 1.68E+09 1.16E+09 2.25E+09 1.58E+09 1.50E+09 1.18E+09 2.17E+09 1.44E+09 1.43E+09 1.79E+09 6.43E+08 1.14E+09 1.57E+09 2.06E+09 1.66E+09 1.25E+09 proline 3.12E+08 2.60E+08 4.88E+08 3.32E+08 4.65E+08 3.21E+08 5.82E+08 3.72E+08 4.12E+08 5.22E+08 1.53E+08 2.04E+08 3.15E+08 4.53E+08 4.38E+08 4.25E+08 propionyl-carnitine 2.22E+07 1.18E+07 4.41E+07 2.05E+07 2.47E+07 1.57E+07 3.90E+07 2.19E+07 1.67E+07 2.69E+07 3415354.163 6329496.604 1.01E+07 1.18E+07 1.33E+07 1.25E+07 riboflavin 57112.8626 67509.8161 60437.52423 16278.6595 65483.98956 43995.93629 73210.49248 68139.82483 50319.99499 50639.63216 35794.91305 19490.76765 14878.80413 32792.16403 0 26421.87483 S-adenosyl-methionine 1.08E+07 8168448.943 1.24E+07 1.13E+07 1.90E+07 1.15E+07 1.75E+07 1.24E+07 1.58E+07 1.09E+07 2335164.792 4422128.536 1.01E+07 8559027.319 8805813.087 1.15E+07 serine 1.27E+07 7155330.371 1.58E+07 9855248.415 2.16E+07 1.22E+07 2.20E+07 1.05E+07 1.37E+07 1.65E+07 4138291.701 6008346.943 1.10E+07 1.61E+07 1.68E+07 1.36E+07 threonine 7.10E+07 4.78E+07 1.01E+08 6.52E+07 1.04E+08 6.80E+07 7.55E+07 3.90E+07 3.81E+07 5.12E+07 2.17E+07 3.87E+07 7.05E+07 1.07E+08 1.11E+08 1.02E+08 valine 2.75E+07 2.24E+07 4.31E+07 3.14E+07 5.00E+07 4.04E+07 4.76E+07 3.19E+07 2.87E+07 3.19E+07 1.48E+07 1.03E+07 2.19E+07 3.49E+07 2.61E+07 3.87E+07 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name quantified m/z retention time, min PubChem ID 3-hydroxy-trimethyllysine 137.072 4.42 439460 5'-methylthioadenosine 298.097 2.38 439176 acetyl-carnitine 204.123 4.81 7045767 AIR 296.064 7.2 161500 alanine 90.055 7.43 5950 arginine 175.119 12.83 6322 asparagine 133.061 7.82 6267 betaine 118.086 5.02 247 carnitine 162.112 6.41 288 CDP-choline 489.115 8.53 13804 choline 104.107 9.81 305 citrulline 176.103 8.28 9750 creatine 132.077 7.43 586 creatinine 114.066 3.96 588 cystathionine 223.075 9.19 834 cysteine-glutathione disulfide 427.095 9.84 3080690 gamma-butyrobetaine 146.118 6.47 725 gamma-glutamylcysteine 251.07 7.64 123938 glycerophosphocholine 258.11 7.18 657272 glycine 76.039 8.13 750 kynurenic acid 190.05 2.84 3845 kynurenine 209.092 4.67 846 L-2-aminobutyric acid 104.071 6.55 80283 leucine / isoleucine 132.102 4.68 6106 lysine 147.113 12.34 5962 methionine 150.058 5.12 6137 methioninesulfoxide 166.053 6.62 158980 NAD+ 664.116 7.78 5892 NADP+ 744.083 10.04 5886 nicotinamide 123.055 2.63 936 N-trimethyllysine 189.16 11 440121 ophthalmate 290.135 7.07 7018721 ornithine 133.097 11.26 6262 phenylalanine 166.086 4.22 6140 phosphocholine 184.074 8.18 1014 proline 116.071 6.01 614 propionyl-carnitine 218.139 4.07 107738 riboflavin 377.146 3.09 493570 S-adenosyl-methionine 399.145 8.38 34756 serine 106.05 8.08 5951 threonine 120.066 7.13 6288 valine 118.086 5.76 6287 METABOLITES_END #END