#METABOLOMICS WORKBENCH H256_20210922_192207 DATATRACK_ID:2855 STUDY_ID:ST001920 ANALYSIS_ID:AN003119 PROJECT_ID:PR001211 VERSION 1 CREATED_ON September 23, 2021, 7:48 am #PROJECT PR:PROJECT_TITLE Comprehensive metabolic and lipidomic profiling of radioresistant MDA-MB-231 PR:PROJECT_TITLE human breast cancer cells PR:PROJECT_TYPE Untargeted MS PR:PROJECT_SUMMARY Metabolic and lipidomic characteristics of radioresistant MDA-MB-231 human PR:PROJECT_SUMMARY breast cancer cells were revealed using GC-MS and nanoESI-MS analyses. PR:INSTITUTE ChungAng University PR:DEPARTMENT College of Pharmacy PR:LABORATORY Natural product biotechnology and Metabolomics PR:LAST_NAME Lee PR:FIRST_NAME Hwanhui PR:ADDRESS College of Pharmacy, Chung-Ang University, Seoul 06974, Republic of Korea PR:EMAIL hwanhui56@gmail.com PR:PHONE +82-2-820-5605 #STUDY ST:STUDY_TITLE Metabolic and lipidomic characterization of radioresistant MDA-MB-231 human ST:STUDY_TITLE breast cancer cells to investigate potential therapeutic targets ST:STUDY_SUMMARY To provide preliminary insights into metabolic and lipidomic characteristics in ST:STUDY_SUMMARY radioresistant triple-negative breast cancer (TNBC) cells and suggest potential ST:STUDY_SUMMARY therapeutic targets, we performed a comprehensive metabolic and lipidomic ST:STUDY_SUMMARY profiling of radioresistant MDA-MB-231 (MDA-MB-231/RR) TNBC cells and their ST:STUDY_SUMMARY parental cells using gas chromatography-mass spectrometry and nano electrospray ST:STUDY_SUMMARY ionization-mass spectrometry, followed by multivariate statistical analysis. ST:STUDY_SUMMARY Buthionine sulfoximine (BSO) and radiation were co-treated to radioresistant ST:STUDY_SUMMARY TNBC cells. The level of glutathione (GSH) was significantly increased, and the ST:STUDY_SUMMARY levels of GSH synthesis-related metabolites, such as cysteine, glycine, and ST:STUDY_SUMMARY glutamine were also increased in MDA-MB-231/RR cells. In contrast, the level of ST:STUDY_SUMMARY lactic acid was significantly reduced. In addition, reactive oxygen species ST:STUDY_SUMMARY (ROS) level was decreased in MDA-MB-231/RR cells. In the lipidomic profiles of ST:STUDY_SUMMARY MDA-MB-231/RR cells, the levels of phosphatidylcholine (PC) and ST:STUDY_SUMMARY phosphatidylethanolamine (PE) were significantly increased, whereas those of ST:STUDY_SUMMARY most of the phosphatidylinositol species were significantly decreased. BSO ST:STUDY_SUMMARY sensitized MDA-MB-231/RR cells to radiotherapy, which resulted in decreased GSH ST:STUDY_SUMMARY level and increased ROS level and apoptosis. Radioresistant TNBC cells showed ST:STUDY_SUMMARY distinct metabolic and lipidomic characteristics compared to their parental ST:STUDY_SUMMARY cells. We suggested activated GSH, PC, and PE biosynthesis pathways as potential ST:STUDY_SUMMARY targets for treating radioresistant TNBC cells. Particularly, enhanced ST:STUDY_SUMMARY radiosensitivity was achieved by inhibition of GSH biosynthesis in MDA-MB-231/RR ST:STUDY_SUMMARY cells. ST:INSTITUTE ChungAng University ST:DEPARTMENT College of Pharmacy ST:LABORATORY Natural product biotechnology and Metabolomics ST:LAST_NAME Lee ST:FIRST_NAME Hwanhui ST:ADDRESS College of Pharmacy, Chung-Ang University, Seoul 06974, Republic of Korea ST:EMAIL hwanhui56@gmail.com ST:PHONE +8228205605 #SUBJECT SU:SUBJECT_TYPE Cultured cells SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 #FACTORS #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 - MDAMB231_1_1 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_1_1.D nanoESI-MS/NEG/MDAMB231_1_1.raw nanoESI-MS/POS/MDAMB231_1_1.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_1_2 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_1_2.D nanoESI-MS/NEG/MDAMB231_1_2.raw nanoESI-MS/POS/MDAMB231_1_2.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_1_3 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_1_3.D nanoESI-MS/NEG/MDAMB231_1_3.raw nanoESI-MS/POS/MDAMB231_1_3.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_2_1 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_2_1.D nanoESI-MS/NEG/MDAMB231_2_1.raw nanoESI-MS/POS/MDAMB231_2_1.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_2_2 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_2_2.D nanoESI-MS/NEG/MDAMB231_2_2.raw nanoESI-MS/POS/MDAMB231_2_2.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_2_3 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_2_3.D nanoESI-MS/NEG/MDAMB231_2_3.raw nanoESI-MS/POS/MDAMB231_2_3.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_3_1 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_3_1.D nanoESI-MS/NEG/MDAMB231_3_1.raw nanoESI-MS/POS/MDAMB231_3_1.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_3_2 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_3_2.D nanoESI-MS/NEG/MDAMB231_3_2.raw nanoESI-MS/POS/MDAMB231_3_2.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_3_3 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_3_3.D nanoESI-MS/NEG/MDAMB231_3_3.raw nanoESI-MS/POS/MDAMB231_3_3.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_4_1 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_4_1.D nanoESI-MS/NEG/MDAMB231_4_1.raw nanoESI-MS/POS/MDAMB231_4_1.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_4_2 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_4_2.D nanoESI-MS/NEG/MDAMB231_4_2.raw nanoESI-MS/POS/MDAMB231_4_2.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_4_3 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_4_3.D nanoESI-MS/NEG/MDAMB231_4_3.raw nanoESI-MS/POS/MDAMB231_4_3.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_5_1 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_5_1.D nanoESI-MS/NEG/MDAMB231_5_1.raw nanoESI-MS/POS/MDAMB231_5_1.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_5_2 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_5_2.D nanoESI-MS/NEG/MDAMB231_5_2.raw nanoESI-MS/POS/MDAMB231_5_2.raw SUBJECT_SAMPLE_FACTORS - MDAMB231_5_3 Factors:Factors;Control RAW_FILE_NAME=GC-MS/MDAMB231_5_3.D nanoESI-MS/NEG/MDAMB231_5_3.raw nanoESI-MS/POS/MDAMB231_5_3.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_1_1 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_1_1.D nanoESI-MS/NEG/MDAMB231RR_1_1.raw nanoESI-MS/POS/MDAMB231RR_1_1.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_1_2 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_1_2.D nanoESI-MS/NEG/MDAMB231RR_1_2.raw nanoESI-MS/POS/MDAMB231RR_1_2.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_1_3 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_1_3.D nanoESI-MS/NEG/MDAMB231RR_1_3.raw nanoESI-MS/POS/MDAMB231RR_1_3.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_2_1 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_2_1.D nanoESI-MS/NEG/MDAMB231RR_2_1.raw nanoESI-MS/POS/MDAMB231RR_2_1.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_2_2 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_2_2.D nanoESI-MS/NEG/MDAMB231RR_2_2.raw nanoESI-MS/POS/MDAMB231RR_2_2.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_2_3 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_2_3.D nanoESI-MS/NEG/MDAMB231RR_2_3.raw nanoESI-MS/POS/MDAMB231RR_2_3.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_3_1 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_3_1.D nanoESI-MS/NEG/MDAMB231RR_3_1.raw nanoESI-MS/POS/MDAMB231RR_3_1.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_3_2 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_3_2.D nanoESI-MS/NEG/MDAMB231RR_3_2.raw nanoESI-MS/POS/MDAMB231RR_3_2.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_3_3 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_3_3.D nanoESI-MS/NEG/MDAMB231RR_3_3.raw nanoESI-MS/POS/MDAMB231RR_3_3.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_4_1 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_4_1.D nanoESI-MS/NEG/MDAMB231RR_4_1.raw nanoESI-MS/POS/MDAMB231RR_4_1.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_4_2 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_4_2.D nanoESI-MS/NEG/MDAMB231RR_4_2.raw nanoESI-MS/POS/MDAMB231RR_4_2.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_4_3 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_4_3.D nanoESI-MS/NEG/MDAMB231RR_4_3.raw nanoESI-MS/POS/MDAMB231RR_4_3.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_5_1 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_5_1.D nanoESI-MS/NEG/MDAMB231RR_5_1.raw nanoESI-MS/POS/MDAMB231RR_5_1.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_5_2 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_5_2.D nanoESI-MS/NEG/MDAMB231RR_5_2.raw nanoESI-MS/POS/MDAMB231RR_5_2.raw SUBJECT_SAMPLE_FACTORS - MDAMB231RR_5_3 Factors:Factors;radioresistant cells RAW_FILE_NAME=GC-MS/MDAMB231RR_5_3.D nanoESI-MS/NEG/MDAMB231RR_5_3.raw nanoESI-MS/POS/MDAMB231RR_5_3.raw SUBJECT_SAMPLE_FACTORS - QC_1 Factors:Factors;Quality control RAW_FILE_NAME=GC-MS/QC_1.D nanoESI-MS/NEG/QC_1.raw nanoESI-MS/POS/QC_1.raw SUBJECT_SAMPLE_FACTORS - QC_2 Factors:Factors;Quality control RAW_FILE_NAME=GC-MS/QC_2.D nanoESI-MS/NEG/QC_2.raw nanoESI-MS/POS/QC_2.raw SUBJECT_SAMPLE_FACTORS - QC_3 Factors:Factors;Quality control RAW_FILE_NAME=GC-MS/QC_3.D nanoESI-MS/NEG/QC_3.raw nanoESI-MS/POS/QC_3.raw SUBJECT_SAMPLE_FACTORS - QC_4 Factors:Factors;Quality control RAW_FILE_NAME=GC-MS/QC_4.D nanoESI-MS/NEG/QC_4.raw nanoESI-MS/POS/QC_4.raw SUBJECT_SAMPLE_FACTORS - QC_5 Factors:Factors;Quality control RAW_FILE_NAME=GC-MS/QC_5.D nanoESI-MS/NEG/QC_5.raw nanoESI-MS/POS/QC_5.raw SUBJECT_SAMPLE_FACTORS - QC_6 Factors:Factors;Quality control RAW_FILE_NAME=GC-MS/QC_6.D nanoESI-MS/NEG/QC_6.raw nanoESI-MS/POS/QC_6.raw #COLLECTION CO:COLLECTION_SUMMARY The cells were seeded at a density of 1 × 10^6 cells/mL in a culture flask. The CO:COLLECTION_SUMMARY cells which had grown to 80-90% confluence, were harvested by treatment with CO:COLLECTION_SUMMARY trypsin- ethylenediaminetetraacetic acid and centrifuged. To remove the residual CO:COLLECTION_SUMMARY medium, the cell pellet was washed with 1× phosphate buffered saline twice. The CO:COLLECTION_SUMMARY cells were freeze-dried and stored at −70 °C until further analysis. CO:SAMPLE_TYPE Breast cancer cells CO:STORAGE_CONDITIONS Described in summary #TREATMENT TR:TREATMENT_SUMMARY Cells at 70–80% confluency were subjected to irradiation using a ^60CO TR:TREATMENT_SUMMARY Theratron-780 teletherapy unit at a dose rate of 1.52 Gy per minute. The cells TR:TREATMENT_SUMMARY were subjected to 25 cycles of 2 Gy irradiation over 5 weeks, and the surviving TR:TREATMENT_SUMMARY cells were designated as MDA-MB-231/RR cells. TR:CELL_MEDIA DMEM supplemented with 10% FBS and 1% penicillin-streptomycin TR:CELL_ENVIR_COND A humidified incubator at 37 °C (95% air and 5% CO2) #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Extraction of metabolites and intact lipid species was performed using a SP:SAMPLEPREP_SUMMARY modified Folch procedur. Briefly, 1 mL of ice-cold chloroform containing 0.1% SP:SAMPLEPREP_SUMMARY BHT and 0.5 mL of ice-cold methanol containing 0.1% BHT were added to the SP:SAMPLEPREP_SUMMARY freeze-dried cells and vortexed. The mixture were sonicated for 30 min at 4 °C SP:SAMPLEPREP_SUMMARY and incubated for 40 min on ice with shaking. Ice-cold water containing 0.1% BHT SP:SAMPLEPREP_SUMMARY (0.38 mL) was added to the mixture for phase separation, and then centrifuged. SP:SAMPLEPREP_SUMMARY The upper (methanol) and lower (chloroform) phases were dried under nitrogen gas SP:SAMPLEPREP_SUMMARY and used GC-MS and nanoESI-MS analyses, respectively. To conduct metabolic SP:SAMPLEPREP_SUMMARY profiling using GC-MS, derivatization reaction was conducted by adding 30 μL of SP:SAMPLEPREP_SUMMARY 20,000 μg/mL methoxylamine hydrochloride in pyridine, 10 μL of myristic-d27 SP:SAMPLEPREP_SUMMARY acid (300 μg/mL as an internal standard), and 50 μL of BSTFA containing 1% SP:SAMPLEPREP_SUMMARY TMCS into dried (methanol phase) sample and incubated for 60 min at 65 °C. SP:SAMPLEPREP_SUMMARY After derivatization, 15 μL of each sample was pooled for quality control (QC) SP:SAMPLEPREP_SUMMARY and analyzed in sextuplicate. To conduct metabolic profiling using nanoESI-MS, SP:SAMPLEPREP_SUMMARY dried (chloroform phase) sample was resuspended with 130 μL of buffer solution SP:SAMPLEPREP_SUMMARY (7.5 mM ammonium acetate in methanol-chloroform (9:1, v/v)) and 10 μL of each SP:SAMPLEPREP_SUMMARY resuspended sample was pooled for QC and analyzed in sextuplicate. SP:PROCESSING_STORAGE_CONDITIONS Described in summary SP:EXTRACTION_METHOD A modified Folch method SP:EXTRACT_STORAGE -80℃ SP:SAMPLE_RESUSPENSION Described in summary SP:SAMPLE_DERIVATIZATION Described in summary #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Agilent 7890A CH:COLUMN_NAME Agilent DB5-MS (30m x 0.25mm, 0.25um) CH:INTERNAL_STANDARD myristic-d27 acid (300 μg/mL as an internal standard #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Agilent 5975C MS:INSTRUMENT_TYPE Triple axis detector MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:MS_COMMENTS The initial oven temperature was set at 70 °C and then increased to 190°C (5 MS:MS_COMMENTS °C/min), 240 °C (6 °C/min), and 280 °C (5 °C/min). Data was processed using MS:MS_COMMENTS Expressionist® MSX software (version 2013.0.39, Genedata, Basel, Switzerland). #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS relative intensity/µg proteins MS_METABOLITE_DATA_START Samples MDAMB231_1_1 MDAMB231_1_2 MDAMB231_1_3 MDAMB231_2_1 MDAMB231_2_2 MDAMB231_2_3 MDAMB231_3_1 MDAMB231_3_2 MDAMB231_3_3 MDAMB231_4_1 MDAMB231_4_2 MDAMB231_4_3 MDAMB231_5_1 MDAMB231_5_2 MDAMB231_5_3 MDAMB231RR_1_1 MDAMB231RR_1_2 MDAMB231RR_1_3 MDAMB231RR_2_1 MDAMB231RR_2_2 MDAMB231RR_2_3 MDAMB231RR_3_1 MDAMB231RR_3_2 MDAMB231RR_3_3 MDAMB231RR_4_1 MDAMB231RR_4_2 MDAMB231RR_4_3 MDAMB231RR_5_1 MDAMB231RR_5_2 MDAMB231RR_5_3 QC_1 QC_2 QC_3 QC_4 QC_5 QC_6 Factors Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;Control Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;radioresistant cells Factors:Factors;Quality control Factors:Factors;Quality control Factors:Factors;Quality control Factors:Factors;Quality control Factors:Factors;Quality control Factors:Factors;Quality control Glycerol 0.001050203 0.001039122 0.001311954 0.001483459 0.001486793 0.001415172 0.001202856 0.001248246 0.001202433 0.001087092 0.001174208 0.001027961 0.001054026 0.001016333 0.001040004 0.001273519 0.001293623 0.001310109 0.001355197 0.001365679 0.001388249 0.001667333 0.001521455 0.001623297 0.001278551 0.001253276 0.001276527 0.001577235 0.001580394 0.001543145 0.000960928 0.001392792 0.001304884 0.00125901 0.001103549 0.001026557 Myo-inositol 0.00150639 0.001461845 0.001387743 0.001132933 0.001089274 0.001090273 0.001016463 0.001030106 0.000990157 0.001298198 0.001341743 0.001397005 0.001584211 0.001519777 0.001497076 0.000204949 0.000206251 0.00020141 0.000256641 0.000217805 0.000220133 0.000252846 0.000280456 0.000275526 0.000251586 0.00025162 0.000239859 0.000309284 0.000301506 0.000310278 0.000697323 0.00072649 0.000749891 0.000728482 0.000775408 0.000756045 β-Alanine 3.55141E-05 3.63884E-05 3.68359E-05 1.5431E-05 1.65895E-05 1.73523E-05 1.90322E-05 1.7544E-05 1.65214E-05 2.5622E-05 3.12005E-05 2.60715E-05 3.69883E-05 4.34893E-05 3.80203E-05 0 5.89321E-06 9.14787E-06 1.09009E-05 7.68753E-06 1.13727E-05 1.70273E-05 1.68099E-05 1.84183E-05 1.59932E-05 1.68632E-05 1.61638E-05 2.12099E-05 2.003E-05 1.93653E-05 3.54044E-05 5.03897E-06 6.96067E-06 6.48984E-06 3.13621E-05 3.82213E-05 Alanine 0.001355878 0.001353144 0.001535876 0.000705373 0.000776438 0.000704902 0.000801855 0.000805831 0.000702074 0.000983031 0.000990525 0.000868073 0.001033634 0.000888214 0.000834101 0.000883992 0.000715882 0.000926685 0.000903314 0.000941367 0.00088457 0.001343701 0.001167404 0.001091505 0.000641088 0.000625079 0.000621321 0.001067158 0.00112104 0.000884835 0.000643637 0.000564205 0.000525586 0.000534505 0.000564823 0.00064443 Aspartic acid 0.000604215 0.000598767 0.000614566 0.00039694 0.00039022 0.000381133 0.000358386 0.000370308 0.000360249 0.000515345 0.000539757 0.000526378 0.00064857 0.000690747 0.000642606 0.000226654 0.000234538 0.000250079 0.000241246 0.000228514 0.000223566 0.000406231 0.000389675 0.000406469 0.000310348 0.000317699 0.000327765 0.000402329 0.000396294 0.000411173 0.00039988 0.000410542 0.000406399 0.000382063 0.000401015 0.000406345 Cysteine 9.69503E-05 9.86376E-05 0.000103537 2.60287E-05 2.21226E-05 2.46906E-05 4.46612E-05 3.8345E-05 4.25552E-05 3.92893E-05 3.92434E-05 3.89609E-05 7.21145E-05 7.81755E-05 7.53092E-05 0.000116823 0.000113653 0.000124355 0.000144247 0.00013996 0.000124632 0.000265166 0.000261996 0.000269997 0.000187538 0.000196827 0.000188669 0.000275348 0.000263399 0.000288363 0.000101644 0.000113701 0.000116954 0.000104086 9.79819E-05 0.00010314 Glutamic acid 0.002605636 0.002633822 0.002653764 0.002024037 0.002149195 0.002002944 0.001892876 0.001882065 0.001872393 0.002689369 0.00271408 0.002607998 0.003273552 0.003246106 0.003290918 0.001991549 0.002082833 0.002168011 0.002216854 0.002135415 0.002064804 0.003610131 0.003697738 0.00388574 0.002692449 0.002894414 0.002794855 0.003288133 0.003241785 0.003354185 0.002532685 0.002456065 0.002482912 0.002464408 0.002422867 0.002536798 Glutamine 7.15466E-06 8.08819E-06 7.69756E-06 8.07126E-06 7.09071E-06 8.17016E-06 6.57431E-06 5.88633E-06 7.09131E-06 5.73367E-06 6.60957E-06 6.019E-06 8.89004E-06 7.93539E-06 6.75994E-06 1.39023E-05 1.40057E-05 1.38899E-05 1.58283E-05 1.60012E-05 1.06992E-05 2.29196E-05 2.34041E-05 2.52641E-05 1.82541E-05 1.87103E-05 1.54791E-05 1.78107E-05 1.64934E-05 1.73492E-05 1.41059E-05 1.19544E-05 1.13804E-05 1.17219E-05 1.27122E-05 1.49957E-05 Glycine 0.001134285 0.001129705 0.001265088 0.001060519 0.001092178 0.001054385 0.000953748 0.000955499 0.000923817 0.001097541 0.00115709 0.001035303 0.00111707 0.001140449 0.001097028 0.001419863 0.001282455 0.001444712 0.001441679 0.001471585 0.001374213 0.002237146 0.002033384 0.002029859 0.001433687 0.001469407 0.001501911 0.001795924 0.001814296 0.001797298 0.001280726 0.001149644 0.00113418 0.001100456 0.001303394 0.001265552 Isoleucine 0.000301394 0.00029569 0.00035117 0.00027379 0.000293764 0.000277143 0.000248159 0.000239272 0.000227131 0.000277904 0.000288332 0.000269228 0.000302817 0.000285754 0.000298311 0.000212295 0.000277565 0.000273426 0.000303821 0.000290407 0.000273449 0.000443188 0.000418093 0.00043909 0.000282553 0.000275528 0.000275118 0.000397353 0.000395338 0.000384918 0.000215666 0.000290245 0.000281479 0.000279772 0.000227187 0.000217392 Lysine 0.00025716 0.000273422 0.000231601 0.00020145 0.000229486 0.000245423 0.000179814 0.000184056 0.000186424 0.000257433 0.000250029 0.000259917 0.000269227 0.000305567 0.000263976 0.000168237 0.000161222 0.000177547 0.000171743 0.000166596 0.000164128 0.000246232 0.000279079 0.000272105 0.000240918 0.000270887 0.000235978 0.000312699 0.000319682 0.000332771 0.000279602 0.000153574 0.000160356 0.000169969 0.000261732 0.000290453 Proline 0.001936787 0.001952289 0.002194437 0.001382734 0.001381939 0.001324746 0.001197228 0.001194999 0.001101793 0.001649083 0.001787534 0.001607705 0.001867882 0.001788336 0.001789867 0.000787403 0.000922042 0.000912138 0.001030094 0.000956987 0.000917489 0.001671372 0.001584282 0.001666791 0.000953022 0.000921247 0.000817915 0.001391887 0.001403094 0.001452304 0.00099173 0.001353199 0.001288803 0.001207165 0.001046694 0.001039042 Pyroglutamic acid 0.000124458 0.000124175 0.000130639 7.27686E-05 6.95659E-05 7.55201E-05 7.30313E-05 7.07346E-05 7.64992E-05 0.000121066 0.00012808 0.000123281 0.000159169 0.000160509 0.000159202 3.70347E-05 3.45697E-05 3.27191E-05 4.34224E-05 3.52448E-05 3.66804E-05 7.74873E-05 7.92829E-05 7.8845E-05 5.05476E-05 5.58331E-05 5.41018E-05 6.12626E-05 5.65124E-05 5.9479E-05 8.54572E-05 8.82868E-05 8.30627E-05 7.65985E-05 7.49792E-05 8.73961E-05 Serine 0.000488795 0.000514762 0.000545595 0.000570676 0.000595455 0.000554568 0.000490469 0.000464987 0.000456301 0.000486718 0.000508071 0.000484131 0.000518573 0.000527569 0.000532478 0.000532532 0.000632393 0.000625512 0.000557202 0.000567551 0.000552856 0.000751269 0.000738058 0.000784833 0.000496828 0.000501088 0.000513441 0.00072186 0.000708961 0.000746114 0.000517936 0.000568915 0.000581495 0.000544741 0.00047855 0.000543104 Threonine 0.000233801 0.000241017 0.000242654 0.000203462 0.000210179 0.000196112 0.000168046 0.000166105 0.000171859 0.000220169 0.000224899 0.000221395 0.000258012 0.000253673 0.000241674 0.000207286 0.000209638 0.000216316 0.0002014 0.000209946 0.00019379 0.000329084 0.000284099 0.000314965 0.000215861 0.00023937 0.000229589 0.00026807 0.000276872 0.000296319 0.000216643 0.000236123 0.000221625 0.000223803 0.000207498 0.000232305 Tyrosine 0.000262339 0.0002456 0.000232678 0.00021039 0.000212141 0.000225925 0.000165561 0.000165196 0.000167344 0.000230991 0.000230908 0.000227886 0.000243738 0.000274975 0.000239797 0.000224734 0.000227123 0.0002499 0.000207015 0.000181087 0.000175147 0.000287987 0.000290414 0.000294232 0.00022855 0.000272112 0.000247299 0.000291759 0.000281496 0.000311146 0.000281608 0.000236842 0.000239045 0.000241183 0.000245013 0.000268778 Valine 0.000560615 0.00056 0.000613202 0.000542887 0.000551661 0.000523696 0.000482283 0.00045923 0.000421419 0.00051108 0.000542245 0.000476256 0.000562245 0.000523206 0.00050863 0.000550898 0.000567671 0.00057837 0.000508301 0.000516661 0.000533165 0.000761367 0.000669078 0.000710653 0.000423414 0.000436196 0.000438313 0.000686884 0.000700431 0.000691797 0.000357691 0.000491804 0.000527522 0.000425807 0.000326334 0.00036837 Aminomalonic acid 9.7895E-05 0.00010496 0.000105565 5.01949E-05 5.15637E-05 5.3567E-05 4.24684E-05 4.33345E-05 4.36475E-05 8.37696E-05 9.26303E-05 9.06316E-05 0.000114123 0.000118316 0.000125088 2.34063E-05 2.45226E-05 2.90262E-05 2.74616E-05 2.38485E-05 2.46766E-05 6.07124E-05 6.16634E-05 7.03212E-05 5.89124E-05 5.76615E-05 6.47159E-05 8.76613E-05 9.0075E-05 0.000107576 0.000112849 1.10259E-05 1.80717E-05 2.07122E-05 0.000103152 0.000105842 Lactic acid 0.004090887 0.00419574 0.005258725 0.003112344 0.003206526 0.002994243 0.00286826 0.002910353 0.002868594 0.003973494 0.004302514 0.003887947 0.00468466 0.004116714 0.004525233 0.001055548 0.001019629 0.001070267 0.001270344 0.001281386 0.001311203 0.00257451 0.002444432 0.002647994 0.001809669 0.001839524 0.00188588 0.001780998 0.001832313 0.00188163 0.001949621 0.002756847 0.00266575 0.002564563 0.002181932 0.002150683 Malic acid 4.13605E-05 4.34787E-05 4.11242E-05 2.90926E-05 2.78126E-05 2.83201E-05 3.17051E-05 2.88392E-05 2.84369E-05 4.24279E-05 4.48659E-05 4.56783E-05 5.54046E-05 5.76692E-05 5.10618E-05 1.01013E-05 1.13401E-05 9.19276E-06 1.2746E-05 1.40733E-05 1.18145E-05 2.78823E-05 2.5979E-05 2.50069E-05 1.9477E-05 2.15871E-05 1.93986E-05 2.04302E-05 1.96613E-05 2.40574E-05 3.1161E-05 2.68712E-05 2.81405E-05 2.70364E-05 2.56533E-05 2.98256E-05 Adenine 1.59734E-05 1.90401E-05 1.53229E-05 1.76101E-05 2.14358E-05 2.11142E-05 2.00738E-05 2.23158E-05 2.09362E-05 1.43337E-05 1.27094E-05 1.57227E-05 1.63346E-05 1.56749E-05 1.4939E-05 5.26668E-05 6.19646E-05 6.62912E-05 6.16132E-05 5.9167E-05 5.65857E-05 5.95891E-05 5.57531E-05 5.67062E-05 4.99717E-05 5.1731E-05 4.83297E-05 4.63461E-05 4.07276E-05 4.30158E-05 2.67956E-05 3.44446E-05 3.48406E-05 3.57355E-05 2.96218E-05 2.88358E-05 Guanine 0.000725767 0.000689189 0.000666527 0.000695539 0.000681709 0.000721857 0.000625915 0.000598219 0.000569976 0.0007369 0.000741103 0.000775706 0.000719035 0.00078232 0.000750801 0.000300786 0.00029212 0.00029776 0.00032465 0.000312806 0.000312528 0.000415621 0.000427079 0.000476595 0.00044494 0.00046686 0.000455134 0.000521141 0.000558766 0.000555463 0.000552642 0.000486615 0.000515685 0.000441051 0.000586784 0.000557439 Hypoxanthine 0.001100514 0.001136175 0.001135544 0.001065361 0.001044852 0.001030649 0.000861282 0.000831432 0.000818371 0.001117372 0.001135359 0.001114369 0.001186346 0.001196193 0.001184412 0.00047685 0.00049059 0.000481481 0.000470085 0.000460171 0.000437741 0.000853054 0.000853187 0.000881866 0.000641994 0.000684401 0.000666528 0.000844905 0.000819455 0.000856194 0.000832895 0.000843195 0.000873918 0.000828858 0.000843914 0.000860666 Inosine 2.6695E-05 2.74986E-05 3.03497E-05 3.13089E-05 3.10893E-05 2.85232E-05 2.07225E-05 2.04506E-05 2.43134E-05 2.60951E-05 2.72853E-05 2.52401E-05 2.67075E-05 3.00088E-05 2.44683E-05 8.36908E-06 9.93264E-06 9.5295E-06 8.33131E-06 7.46649E-06 8.10351E-06 1.3026E-05 1.16384E-05 1.39145E-05 9.70544E-06 1.18423E-05 8.3599E-06 1.57798E-05 1.46687E-05 1.34617E-05 1.61872E-05 2.05051E-05 1.89371E-05 1.73154E-05 1.76216E-05 1.57582E-05 Uracil 0.000408835 0.000399268 0.000439519 0.000435383 0.000411725 0.000402892 0.000321692 0.00031494 0.000311677 0.000393586 0.000416533 0.000401341 0.000447032 0.000474289 0.000453981 0.000278496 0.000274635 0.000304764 0.000269772 0.000276948 0.000282537 0.000409664 0.000382064 0.00040223 0.000315635 0.000343686 0.000356797 0.00039416 0.000385213 0.00039866 0.000342073 0.000382683 0.000356967 0.000348187 0.000333441 0.000341492 Uridine 6.41513E-05 5.88995E-05 6.77059E-05 6.74046E-05 6.76136E-05 6.08435E-05 6.21219E-05 6.19768E-05 6.00233E-05 6.29988E-05 6.42828E-05 6.49525E-05 6.24143E-05 6.04474E-05 5.87831E-05 1.83765E-05 1.59861E-05 1.98671E-05 1.94699E-05 1.93048E-05 1.74819E-05 2.76425E-05 2.32161E-05 2.34404E-05 2.31496E-05 2.55407E-05 2.19965E-05 2.41802E-05 2.58197E-05 2.41478E-05 4.00613E-05 3.86155E-05 4.30389E-05 3.96939E-05 3.97505E-05 4.13209E-05 Glucose 0.000155414 0.000147737 0.000138884 0.000106862 0.000105502 0.000108759 5.45017E-05 5.08615E-05 5.31781E-05 9.65306E-05 9.89357E-05 9.39836E-05 0.000148867 0.000160515 0.000139088 0.000138664 0.000135956 0.000143784 0.000127098 0.000121441 0.000116996 0.000152008 0.000163302 0.000153739 0.000117759 0.000146014 0.000133678 0.000271811 0.000256842 0.000284015 0.00015228 0.000131762 0.000137185 0.000132301 0.000140397 0.000148211 Glucose-6- phosphate 0.000463639 0.000459947 0.000487748 0.000535009 0.000554869 0.000553049 0.000457184 0.000454052 0.000459314 0.000541307 0.000545372 0.000567719 0.000537173 0.00056581 0.000530822 0.000414708 0.000412502 0.000418265 0.000363895 0.000326806 0.000341483 0.000506854 0.000519711 0.000528406 0.000395978 0.000439185 0.000389347 0.000545303 0.000530435 0.000562207 0.00045861 0.00043411 0.000470338 0.000441824 0.000471225 0.000479638 Glyceric acid 2.92912E-05 2.65819E-05 2.8699E-05 1.61015E-05 1.48726E-05 1.35816E-05 1.66931E-05 1.26207E-05 1.48829E-05 2.29032E-05 2.7566E-05 2.46415E-05 2.61552E-05 2.66787E-05 2.76148E-05 5.22807E-06 6.20218E-06 3.50201E-06 6.30508E-06 4.96128E-06 6.55097E-06 8.06554E-06 9.94832E-06 1.03322E-05 9.05401E-06 7.92578E-06 7.99599E-06 1.04054E-05 1.05531E-05 1.12376E-05 1.24048E-05 1.52267E-05 1.35212E-05 1.42132E-05 1.34419E-05 1.34895E-05 Glycerol-3-phosphate 1.51873E-05 1.86491E-05 1.91087E-05 1.63343E-05 1.34109E-05 1.66635E-05 1.77896E-05 1.53096E-05 1.68309E-05 1.53732E-05 1.32026E-05 1.47051E-05 1.79985E-05 1.55209E-05 1.43525E-05 4.41099E-05 4.28545E-05 4.53689E-05 5.10406E-05 4.12008E-05 4.52509E-05 5.81754E-05 5.80707E-05 6.03346E-05 4.69317E-05 5.12206E-05 4.91687E-05 5.69464E-05 5.12608E-05 5.33855E-05 2.68092E-05 2.93258E-05 2.83731E-05 2.80214E-05 2.62649E-05 2.65598E-05 Mannose-6-phosphate 8.56408E-05 7.90988E-05 8.69223E-05 0.000107505 0.000104697 0.000110039 5.42002E-05 5.56996E-05 5.48621E-05 6.95159E-05 7.15442E-05 6.94951E-05 5.68349E-05 5.94144E-05 5.98527E-05 5.96897E-05 6.32576E-05 6.08586E-05 5.64967E-05 5.1811E-05 5.66614E-05 4.54144E-05 4.70901E-05 4.80096E-05 6.80892E-05 8.41024E-05 7.93938E-05 7.74416E-05 6.93176E-05 7.86002E-05 7.27542E-05 6.09366E-05 5.64577E-05 6.72918E-05 6.99475E-05 7.32491E-05 Ribose 5.36147E-05 5.55632E-05 5.75056E-05 6.66409E-05 6.7957E-05 6.85615E-05 4.61414E-05 4.10717E-05 4.1208E-05 5.74546E-05 5.79395E-05 5.5628E-05 4.90888E-05 4.45544E-05 5.49673E-05 2.67769E-05 3.5508E-05 3.48292E-05 3.26234E-05 3.02098E-05 3.2048E-05 3.8914E-05 3.45914E-05 4.20545E-05 3.70657E-05 3.90535E-05 4.01113E-05 5.18783E-05 5.70406E-05 5.75985E-05 4.72601E-05 4.7546E-05 4.5021E-05 4.45912E-05 4.70395E-05 5.08943E-05 Creatinine 7.10678E-05 7.33497E-05 7.37077E-05 3.68499E-05 3.85649E-05 3.89787E-05 4.10247E-05 4.17345E-05 3.95695E-05 5.87473E-05 6.12477E-05 6.04964E-05 7.08542E-05 7.48198E-05 6.65666E-05 1.03726E-05 9.69233E-06 1.16958E-05 1.23498E-05 1.13962E-05 1.10319E-05 1.91225E-05 2.01294E-05 1.77041E-05 1.42183E-05 1.43202E-05 1.42634E-05 1.33293E-05 1.73593E-05 1.43476E-05 3.18728E-05 3.30577E-05 3.06902E-05 3.02215E-05 2.65125E-05 2.91031E-05 Putrescine 3.39617E-05 3.67321E-05 3.5867E-05 2.62708E-05 2.61841E-05 2.55119E-05 2.71001E-05 2.60367E-05 2.43225E-05 2.99401E-05 3.37651E-05 3.19309E-05 4.35165E-05 4.68258E-05 4.31472E-05 0 0 0 0 1.53873E-05 1.26223E-05 2.37296E-05 2.41674E-05 2.66195E-05 2.24509E-05 2.4789E-05 2.16731E-05 2.52848E-05 2.62084E-05 3.05125E-05 2.80903E-05 0 0 1.12583E-05 2.73863E-05 2.62853E-05 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name m/z RT(min) Fragmentation ion (m/z) TMS Glycerol 205 11.48 103, 117, 205, 218 3 Myo-inositol 305 29.18 191, 217, 305, 318 6 β-Alanine 174 15.36 133, 174, 248, 290 3 Alanine 116 7.19 100, 116, 190, 218 2 Aspartic acid 232 17.65 100, 202, 218, 232 3 Cysteine 220 18.45 100, 132, 220, 294 3 Glutamic acid 246 20.01 128, 156, 246, 348 3 Glutamine 156 23.33 128, 156, 245, 347 3 Glycine 102 7.66 86, 102, 176, 204 2 Isoleucine 158 11.94 100, 158, 218, 232 2 Lysine 84 21.94 84, 102, 156, 362 3 Proline 142 12.03 100, 142, 216, 244 2 Pyroglutamic acid 156 17.59 133, 156, 230, 258 2 Serine 204 13.68 100, 204, 218, 278 3 Threonine 218 14.31 101, 117, 218, 291 3 Tyrosine 218 26.59 100, 179, 218, 280 3 Valine 144 9.95 100, 144, 218, 246 2 Aminomalonic acid 218 16.40 133, 174, 218, 320 3 Lactic acid 117 6.21 117, 133, 191, 219 2 Malic acid 233 16.89 133, 233, 245, 335 3 Adenine 264 25.16 84, 192, 264, 279 2 Guanine 352 29.77 238, 264, 352, 367 3 Hypoxanthine 265 23.94 193, 206, 265, 280 2 Inosine 217 36.31 217, 230, 245, 281 4 Uracil 241 13.07 99, 113, 241, 255 2 Uridine 217 34.53 103, 169, 217, 259 3 Glucose 204 25.73 129, 191, 204, 217 5 Glucose-6- phosphate 387 33.54 129, 204, 299, 387 6 Glyceric acid 189 12.91 103, 133, 189, 292 3 Glycerol-3-phosphate 357 23.10 299, 315, 357, 445 4 Mannose-6-phosphate 387 32.71 217, 299, 357, 387 6(MEOX) Ribose 217 20.69 133, 191, 204, 217 4 Creatinine 115 18.33 100, 115, 314, 329 3 Putrescine 174 22.49 86, 174, 200, 214 4 METABOLITES_END #END