#METABOLOMICS WORKBENCH Cristina79_20190307_131539 DATATRACK_ID:1665 STUDY_ID:ST001152 ANALYSIS_ID:AN001902 PROJECT_ID:PR000771 VERSION 1 CREATED_ON March 19, 2019, 4:59 pm #PROJECT PR:PROJECT_TITLE Metabolomic Analysis of Liver Tissues for Characterization of Hepatocellular PR:PROJECT_TITLE Carcinoma PR:PROJECT_SUMMARY Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer PR:PROJECT_SUMMARY causing more than half a million annual deaths world-wide. Understanding the PR:PROJECT_SUMMARY molecular mechanisms contributing to HCC development and progression is highly PR:PROJECT_SUMMARY desirable for improved surveillance, diagnosis and treatment. Liver tissue PR:PROJECT_SUMMARY metabolomics has the potential to reflect the physiological changes behind HCC PR:PROJECT_SUMMARY development. Also, it allows researchers to investigate racial disparities in PR:PROJECT_SUMMARY HCC. The use of both gas chromatography – mass spectrometry (GC-MS) and liquid PR:PROJECT_SUMMARY chromatography – mass spectrometry (LC-MS) platforms helps increase the PR:PROJECT_SUMMARY metabolome coverage, allowing researchers to better unravel the relationships of PR:PROJECT_SUMMARY metabolites and HCC. The objective of this study is to identify HCC-associated PR:PROJECT_SUMMARY metabolites by analysis of liver tissues from HCC patients using both GC-MS and PR:PROJECT_SUMMARY LC-MS platforms. Paired tumor and non-tumor tissues from 40 patients were PR:PROJECT_SUMMARY analyzed by GC-MS and LC-MS. The patients consist of 14 African-Americans (AA), PR:PROJECT_SUMMARY 10 Asian-Americans (AS), and 16 European-Americans (EA). The levels of the PR:PROJECT_SUMMARY metabolites extracted from the solid liver tissue of the HCC area and adjacent PR:PROJECT_SUMMARY non-HCC area were compared. Among the analytes detected by GC-MS and LC-MS with PR:PROJECT_SUMMARY significant alterations, 17 were selected based on availability of putative PR:PROJECT_SUMMARY metabolite identifications. These metabolites belong to TCA cycle, glycolysis, PR:PROJECT_SUMMARY purines, and lipid metabolism, and have been previously reported in liver PR:PROJECT_SUMMARY metabolomics studies where high correlation with HCC progression was implied. We PR:PROJECT_SUMMARY demonstrated that metabolites that are related to HCC pathogenesis can be PR:PROJECT_SUMMARY identified through metabolomics analysis of liver tissues by both GC-MS and PR:PROJECT_SUMMARY LC-MS. In addition, this analysis has led to the identification of metabolites PR:PROJECT_SUMMARY associated with HCC in a race-specific manner. PR:INSTITUTE Georgetown University PR:DEPARTMENT Oncology, Georgetown University Medical Center PR:LABORATORY Ressom Lab PR:LAST_NAME Ressom PR:FIRST_NAME Habtom PR:ADDRESS 3970 Reservoir Rd., NW, Research Bldg, Room W325, Washington, DC, 20007, USA PR:EMAIL hwr@georgetown.edu PR:PHONE 2026872283 #STUDY ST:STUDY_TITLE Metabolomic Analysis of Liver Tissues for Characterization of Hepatocellular ST:STUDY_TITLE Carcinoma ST:STUDY_SUMMARY Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer ST:STUDY_SUMMARY causing more than half a million annual deaths world-wide. Understanding the ST:STUDY_SUMMARY molecular mechanisms contributing to HCC development and progression is highly ST:STUDY_SUMMARY desirable for improved surveillance, diagnosis and treatment. Liver tissue ST:STUDY_SUMMARY metabolomics has the potential to reflect the physiological changes behind HCC ST:STUDY_SUMMARY development. Also, it allows researchers to investigate racial disparities in ST:STUDY_SUMMARY HCC. The use of both gas chromatography – mass spectrometry (GC-MS) and liquid ST:STUDY_SUMMARY chromatography – mass spectrometry (LC-MS) platforms helps increase the ST:STUDY_SUMMARY metabolome coverage, allowing researchers to better unravel the relationships of ST:STUDY_SUMMARY metabolites and HCC. The objective of this study is to identify HCC-associated ST:STUDY_SUMMARY metabolites by analysis of liver tissues from HCC patients using both GC-MS and ST:STUDY_SUMMARY LC-MS platforms. Paired tumor and non-tumor tissues from 40 patients were ST:STUDY_SUMMARY analyzed by GC-MS and LC-MS. The patients consist of 14 African-Americans (AA), ST:STUDY_SUMMARY 10 Asian-Americans (AS), and 16 European-Americans (EA). The levels of the ST:STUDY_SUMMARY metabolites extracted from the solid liver tissue of the HCC area and adjacent ST:STUDY_SUMMARY non-HCC area were compared. Among the analytes detected by GC-MS and LC-MS with ST:STUDY_SUMMARY significant alterations, 17 were selected based on availability of putative ST:STUDY_SUMMARY metabolite identifications. These metabolites belong to TCA cycle, glycolysis, ST:STUDY_SUMMARY purines, and lipid metabolism, and have been previously reported in liver ST:STUDY_SUMMARY metabolomics studies where high correlation with HCC progression was implied. We ST:STUDY_SUMMARY demonstrated that metabolites that are related to HCC pathogenesis can be ST:STUDY_SUMMARY identified through metabolomics analysis of liver tissues by both GC-MS and ST:STUDY_SUMMARY LC-MS. In addition, this analysis has led to the identification of metabolites ST:STUDY_SUMMARY associated with HCC in a race-specific manner. ST:INSTITUTE Georgetown University ST:DEPARTMENT Oncology ST:LABORATORY Ressom Lab ST:LAST_NAME Di Poto ST:FIRST_NAME Cristina ST:ADDRESS 3970 Reservoir Rd. NW, Research Bldg., Room W325 ST:EMAIL cd329@georgetown.edu ST:PHONE 2026872926 ST:NUM_GROUPS 4 ST:TOTAL_SUBJECTS 40 #SUBJECT SU:SUBJECT_TYPE Human 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]Additional sample data SUBJECT_SAMPLE_FACTORS HCC/CIRR 01 HCC/CIRR 01_HCC/C Group:HCC/C | RACE:Black GCMS Batch=1; GCMS Data Name=A18_1; LCMS Batch=1; LCMSNeg Data Name=NEG_22; LCMSPos Data Name=POS_22 SUBJECT_SAMPLE_FACTORS HCC/CIRR 01 HCC/CIRR 01_ADJ/C Group:ADJ/C | RACE:Black GCMS Batch=1; GCMS Data Name=A20_1; LCMS Batch=1; LCMSNeg Data Name=NEG_21; LCMSPos Data Name=POS_21 SUBJECT_SAMPLE_FACTORS HCC/CIRR 02 HCC/CIRR 02_HCC/C Group:HCC/C | RACE:Black GCMS Batch=1; GCMS Data Name=A21_1; LCMS Batch=1; LCMSNeg Data Name=NEG_28; LCMSPos Data Name=POS_28 SUBJECT_SAMPLE_FACTORS HCC/CIRR 02 HCC/CIRR 02_ADJ/C Group:ADJ/C | RACE:Black GCMS Batch=1; GCMS Data Name=A14_1; LCMS Batch=1; LCMSNeg Data Name=NEG_27; LCMSPos Data Name=POS_27 SUBJECT_SAMPLE_FACTORS HCC/CIRR 03 HCC/CIRR 03_HCC/C Group:HCC/C | RACE:White GCMS Batch=1; GCMS Data Name=A13_1; LCMS Batch=1; LCMSNeg Data Name=NEG_34; LCMSPos Data Name=POS_34 SUBJECT_SAMPLE_FACTORS HCC/CIRR 03 HCC/CIRR 03_ADJ/C Group:ADJ/C | RACE:White GCMS Batch=1; GCMS Data Name=A07_1; LCMS Batch=1; LCMSNeg Data Name=NEG_33; LCMSPos Data Name=POS_33 SUBJECT_SAMPLE_FACTORS HCC/CIRR 04 HCC/CIRR 04_HCC/C Group:HCC/C | RACE:White GCMS Batch=1; GCMS Data Name=A05_1; LCMS Batch=1; LCMSNeg Data Name=NEG_37; LCMSPos Data Name=POS_37 SUBJECT_SAMPLE_FACTORS HCC/CIRR 04 HCC/CIRR 04_ADJ/C Group:ADJ/C | RACE:White GCMS Batch=1; GCMS Data Name=A06_1; LCMS Batch=1; LCMSNeg Data Name=NEG_36; LCMSPos Data Name=POS_36 SUBJECT_SAMPLE_FACTORS HCC/CIRR 05 HCC/CIRR 05_HCC/C Group:HCC/C | RACE:White GCMS Batch=1; GCMS Data Name=A03_1; LCMS Batch=1; LCMSNeg Data Name=NEG_41; LCMSPos Data Name=POS_41 SUBJECT_SAMPLE_FACTORS HCC/CIRR 05 HCC/CIRR 05_ADJ/C Group:ADJ/C | RACE:White GCMS Batch=1; GCMS Data Name=A02_1; LCMS Batch=1; LCMSNeg Data Name=NEG_40; LCMSPos Data Name=POS_40 SUBJECT_SAMPLE_FACTORS HCC/CIRR 06 HCC/CIRR 06_HCC/C Group:HCC/C | RACE:Black GCMS Batch=2; GCMS Data Name=B-13_1; LCMS Batch=1; LCMSNeg Data Name=NEG_48; LCMSPos Data Name=POS_48 SUBJECT_SAMPLE_FACTORS HCC/CIRR 06 HCC/CIRR 06_ADJ/C Group:ADJ/C | RACE:Black GCMS Batch=2; GCMS Data Name=B-19_1; LCMS Batch=1; LCMSNeg Data Name=NEG_47; LCMSPos Data Name=POS_47 SUBJECT_SAMPLE_FACTORS HCC/CIRR 07 HCC/CIRR 07_HCC/C Group:HCC/C | RACE:Black GCMS Batch=2; GCMS Data Name=B-09_1; LCMS Batch=1; LCMSNeg Data Name=NEG_54; LCMSPos Data Name=POS_54 SUBJECT_SAMPLE_FACTORS HCC/CIRR 07 HCC/CIRR 07_ADJ/C Group:ADJ/C | RACE:Black GCMS Batch=2; GCMS Data Name=B-21_1; LCMS Batch=1; LCMSNeg Data Name=NEG_53; LCMSPos Data Name=POS_53 SUBJECT_SAMPLE_FACTORS HCC/CIRR 08 HCC/CIRR 08_HCC/C Group:HCC/C | RACE:Black GCMS Batch=2; GCMS Data Name=B-01_1; LCMS Batch=1; LCMSNeg Data Name=NEG_60; LCMSPos Data Name=POS_60 SUBJECT_SAMPLE_FACTORS HCC/CIRR 08 HCC/CIRR 08_ADJ/C Group:ADJ/C | RACE:Black GCMS Batch=2; GCMS Data Name=B-10_1; LCMS Batch=1; LCMSNeg Data Name=NEG_59; LCMSPos Data Name=POS_59 SUBJECT_SAMPLE_FACTORS HCC/CIRR 09 HCC/CIRR 09_HCC/C Group:HCC/C | RACE:White GCMS Batch=2; GCMS Data Name=B-06_1; LCMS Batch=1; LCMSNeg Data Name=NEG_63; LCMSPos Data Name=POS_63 SUBJECT_SAMPLE_FACTORS HCC/CIRR 09 HCC/CIRR 09_ADJ/C Group:ADJ/C | RACE:White GCMS Batch=2; GCMS Data Name=B-17_1; LCMS Batch=1; LCMSNeg Data Name=NEG_62; LCMSPos Data Name=POS_62 SUBJECT_SAMPLE_FACTORS HCC/CIRR 10 HCC/CIRR 10_HCC/C Group:HCC/C | RACE:White GCMS Batch=2; GCMS Data Name=B-11_1; LCMS Batch=1; LCMSNeg Data Name=NEG_67; LCMSPos Data Name=POS_67 SUBJECT_SAMPLE_FACTORS HCC/CIRR 10 HCC/CIRR 10_ADJ/C Group:ADJ/C | RACE:White GCMS Batch=2; GCMS Data Name=B-05_1; LCMS Batch=1; LCMSNeg Data Name=NEG_66; LCMSPos Data Name=POS_66 SUBJECT_SAMPLE_FACTORS HCC/NOR 01 HCC/NOR 01_HCC/N Group:HCC/N | RACE:White GCMS Batch=1; GCMS Data Name=A09_1; LCMS Batch=1; LCMSNeg Data Name=NEG_25; LCMSPos Data Name=POS_25 SUBJECT_SAMPLE_FACTORS HCC/NOR 01 HCC/NOR 01_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=1; GCMS Data Name=A08_1; LCMS Batch=1; LCMSNeg Data Name=NEG_24; LCMSPos Data Name=POS_24 SUBJECT_SAMPLE_FACTORS HCC/NOR 02 HCC/NOR 02_HCC/N Group:HCC/N | RACE:White GCMS Batch=1; GCMS Data Name=A04_1; LCMS Batch=1; LCMSNeg Data Name=NEG_31; LCMSPos Data Name=POS_31 SUBJECT_SAMPLE_FACTORS HCC/NOR 02 HCC/NOR 02_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=1; GCMS Data Name=A15_1; LCMS Batch=1; LCMSNeg Data Name=NEG_30; LCMSPos Data Name=POS_30 SUBJECT_SAMPLE_FACTORS HCC/NOR 03 HCC/NOR 03_HCC/N Group:HCC/N | RACE:Black GCMS Batch=1; GCMS Data Name=A16_1; LCMS Batch=1; LCMSNeg Data Name=NEG_39; LCMSPos Data Name=POS_39 SUBJECT_SAMPLE_FACTORS HCC/NOR 03 HCC/NOR 03_ADJ/N Group:ADJ/N | RACE:Black GCMS Batch=1; GCMS Data Name=A09_2; LCMS Batch=1; LCMSNeg Data Name=NEG_38; LCMSPos Data Name=POS_38 SUBJECT_SAMPLE_FACTORS HCC/NOR 04 HCC/NOR 04_HCC/N Group:HCC/N | RACE:Black GCMS Batch=2; GCMS Data Name=B-14_1; LCMS Batch=1; LCMSNeg Data Name=NEG_51; LCMSPos Data Name=POS_51 SUBJECT_SAMPLE_FACTORS HCC/NOR 04 HCC/NOR 04_ADJ/N Group:ADJ/N | RACE:Black GCMS Batch=2; GCMS Data Name=B-07_1; LCMS Batch=1; LCMSNeg Data Name=NEG_50; LCMSPos Data Name=POS_50 SUBJECT_SAMPLE_FACTORS HCC/NOR 05 HCC/NOR 05_HCC/N Group:HCC/N | RACE:White GCMS Batch=2; GCMS Data Name=B-15_1; LCMS Batch=1; LCMSNeg Data Name=NEG_57; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS HCC/NOR 05 HCC/NOR 05_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=2; GCMS Data Name=B-12_1; LCMS Batch=1; LCMSNeg Data Name=NEG_56; LCMSPos Data Name=POS_56 SUBJECT_SAMPLE_FACTORS HCC/NOR 06 HCC/NOR 06_HCC/N Group:HCC/N | RACE:White GCMS Batch=2; GCMS Data Name=B-08_1; LCMS Batch=1; LCMSNeg Data Name=NEG_65; LCMSPos Data Name=POS_65 SUBJECT_SAMPLE_FACTORS HCC/NOR 06 HCC/NOR 06_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=2; GCMS Data Name=B-18_1; LCMS Batch=1; LCMSNeg Data Name=NEG_64; LCMSPos Data Name=POS_64 SUBJECT_SAMPLE_FACTORS HCC/NOR 07 HCC/NOR 07_HCC/N Group:HCC/N | RACE:Black GCMS Batch=3; GCMS Data Name=C-19_1; LCMS Batch=1; LCMSNeg Data Name=NEG_74; LCMSPos Data Name=POS_74 SUBJECT_SAMPLE_FACTORS HCC/NOR 07 HCC/NOR 07_ADJ/N Group:ADJ/N | RACE:Black GCMS Batch=3; GCMS Data Name=C-02_1; LCMS Batch=1; LCMSNeg Data Name=NEG_73; LCMSPos Data Name=POS_73 SUBJECT_SAMPLE_FACTORS HCC/NOR 08 HCC/NOR 08_HCC/N Group:HCC/N | RACE:Black GCMS Batch=3; GCMS Data Name=C-08_1; LCMS Batch=1; LCMSNeg Data Name=NEG_77; LCMSPos Data Name=POS_77 SUBJECT_SAMPLE_FACTORS HCC/NOR 08 HCC/NOR 08_ADJ/N Group:ADJ/N | RACE:Black GCMS Batch=3; GCMS Data Name=C-03_1; LCMS Batch=1; LCMSNeg Data Name=NEG_76; LCMSPos Data Name=POS_76 SUBJECT_SAMPLE_FACTORS HCC/NOR 09 HCC/NOR 09_HCC/N Group:HCC/N | RACE:White GCMS Batch=3; GCMS Data Name=C-06_1; LCMS Batch=1; LCMSNeg Data Name=NEG_80; LCMSPos Data Name=POS_80 SUBJECT_SAMPLE_FACTORS HCC/NOR 09 HCC/NOR 09_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=3; GCMS Data Name=C-07_1; LCMS Batch=1; LCMSNeg Data Name=NEG_79; LCMSPos Data Name=POS_79 SUBJECT_SAMPLE_FACTORS HCC/NOR 10 HCC/NOR 10_HCC/N Group:HCC/N | RACE:Black GCMS Batch=3; GCMS Data Name=C-11_1; LCMS Batch=1; LCMSNeg Data Name=NEG_83; LCMSPos Data Name=POS_83 SUBJECT_SAMPLE_FACTORS HCC/NOR 10 HCC/NOR 10_ADJ/N Group:ADJ/N | RACE:Black GCMS Batch=3; GCMS Data Name=C-14_1; LCMS Batch=1; LCMSNeg Data Name=NEG_82; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS HCC/NOR 11 HCC/NOR 11_HCC/N Group:HCC/N | RACE:White GCMS Batch=3; GCMS Data Name=C-17_1; LCMS Batch=1; LCMSNeg Data Name=NEG_85; LCMSPos Data Name=POS_85 SUBJECT_SAMPLE_FACTORS HCC/NOR 11 HCC/NOR 11_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=3; GCMS Data Name=C-16_1; LCMS Batch=1; LCMSNeg Data Name=NEG_84; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS HCC/NOR 12 HCC/NOR 12_HCC/N Group:HCC/N | RACE:Black GCMS Batch=3; GCMS Data Name=C-09_1; LCMS Batch=1; LCMSNeg Data Name=NEG_88; LCMSPos Data Name=POS_88 SUBJECT_SAMPLE_FACTORS HCC/NOR 12 HCC/NOR 12_ADJ/N Group:ADJ/N | RACE:Black GCMS Batch=3; GCMS Data Name=C-01_1; LCMS Batch=1; LCMSNeg Data Name=NEG_87; LCMSPos Data Name=POS_87 SUBJECT_SAMPLE_FACTORS HCC/NOR 13 HCC/NOR 13_HCC/N Group:HCC/N | RACE:White GCMS Batch=3; GCMS Data Name=C-12_1; LCMS Batch=1; LCMSNeg Data Name=NEG_91; LCMSPos Data Name=POS_91 SUBJECT_SAMPLE_FACTORS HCC/NOR 13 HCC/NOR 13_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=3; GCMS Data Name=C-18_1; LCMS Batch=1; LCMSNeg Data Name=NEG_90; LCMSPos Data Name=POS_90 SUBJECT_SAMPLE_FACTORS HCC/NOR 14 HCC/NOR 14_HCC/N Group:HCC/N | RACE:White GCMS Batch=3; GCMS Data Name=C-05_1; LCMS Batch=1; LCMSNeg Data Name=NEG_93; LCMSPos Data Name=POS_93 SUBJECT_SAMPLE_FACTORS HCC/NOR 14 HCC/NOR 14_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=3; GCMS Data Name=C-20_1; LCMS Batch=1; LCMSNeg Data Name=NEG_92; LCMSPos Data Name=POS_92 SUBJECT_SAMPLE_FACTORS HCC/NOR 15 HCC/NOR 15_HCC/N Group:HCC/N | RACE:White GCMS Batch=4; GCMS Data Name=D-04_1; LCMS Batch=1; LCMSNeg Data Name=NEG_100; LCMSPos Data Name=POS_100 SUBJECT_SAMPLE_FACTORS HCC/NOR 15 HCC/NOR 15_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=4; GCMS Data Name=D-07_1; LCMS Batch=1; LCMSNeg Data Name=NEG_99; LCMSPos Data Name=POS_99 SUBJECT_SAMPLE_FACTORS HCC/NOR 16 HCC/NOR 16_HCC/N Group:HCC/N | RACE:Black GCMS Batch=4; GCMS Data Name=D-08_1; LCMS Batch=1; LCMSNeg Data Name=NEG_103; LCMSPos Data Name=POS_103 SUBJECT_SAMPLE_FACTORS HCC/NOR 16 HCC/NOR 16_ADJ/N Group:ADJ/N | RACE:Black GCMS Batch=4; GCMS Data Name=D-16_1; LCMS Batch=1; LCMSNeg Data Name=NEG_102; LCMSPos Data Name=POS_102 SUBJECT_SAMPLE_FACTORS HCC/NOR 17 HCC/NOR 17_HCC/N Group:HCC/N | RACE:Asian GCMS Batch=4; GCMS Data Name=D-18_1; LCMS Batch=1; LCMSNeg Data Name=NEG_106; LCMSPos Data Name=POS_106 SUBJECT_SAMPLE_FACTORS HCC/NOR 17 HCC/NOR 17_ADJ/N Group:ADJ/N | RACE:Asian GCMS Batch=4; GCMS Data Name=D-19_1; LCMS Batch=1; LCMSNeg Data Name=NEG_105; LCMSPos Data Name=POS_105 SUBJECT_SAMPLE_FACTORS HCC/NOR 18 HCC/NOR 18_HCC/N Group:HCC/N | RACE:White GCMS Batch=4; GCMS Data Name=D-21_1; LCMS Batch=1; LCMSNeg Data Name=NEG_108; LCMSPos Data Name=POS_108 SUBJECT_SAMPLE_FACTORS HCC/NOR 18 HCC/NOR 18_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=4; GCMS Data Name=D-15_1; LCMS Batch=1; LCMSNeg Data Name=NEG_107; LCMSPos Data Name=POS_107 SUBJECT_SAMPLE_FACTORS HCC/NOR 19 HCC/NOR 19_HCC/N Group:HCC/N | RACE:Black GCMS Batch=4; GCMS Data Name=D-12_1; LCMS Batch=1; LCMSNeg Data Name=NEG_111; LCMSPos Data Name=POS_111 SUBJECT_SAMPLE_FACTORS HCC/NOR 19 HCC/NOR 19_ADJ/N Group:ADJ/N | RACE:Black GCMS Batch=4; GCMS Data Name=D-14_1; LCMS Batch=1; LCMSNeg Data Name=NEG_110; LCMSPos Data Name=POS_110 SUBJECT_SAMPLE_FACTORS HCC/NOR 20 HCC/NOR 20_HCC/N Group:HCC/N | RACE:White GCMS Batch=4; GCMS Data Name=D-17_1; LCMS Batch=1; LCMSNeg Data Name=NEG_114; LCMSPos Data Name=POS_114 SUBJECT_SAMPLE_FACTORS HCC/NOR 20 HCC/NOR 20_ADJ/N Group:ADJ/N | RACE:White GCMS Batch=4; GCMS Data Name=D-10_1; LCMS Batch=1; LCMSNeg Data Name=NEG_113; LCMSPos Data Name=POS_113 SUBJECT_SAMPLE_FACTORS HCC/NOR 21 HCC/NOR 21_HCC/N Group:HCC/N | RACE:Black GCMS Batch=4; GCMS Data Name=D-11_1; LCMS Batch=1; LCMSNeg Data Name=NEG_117; LCMSPos Data Name=POS_117 SUBJECT_SAMPLE_FACTORS HCC/NOR 21 HCC/NOR 21_ADJ/N Group:ADJ/N | RACE:Black GCMS Batch=4; GCMS Data Name=D-05_1; LCMS Batch=1; LCMSNeg Data Name=NEG_116; LCMSPos Data Name=POS_116 SUBJECT_SAMPLE_FACTORS HCC/NOR 22 HCC/NOR 22_HCC/N Group:HCC/N | RACE:Asian GCMS Batch=4; GCMS Data Name=D-09_1; LCMS Batch=1; LCMSNeg Data Name=NEG_119; LCMSPos Data Name=POS_119 SUBJECT_SAMPLE_FACTORS HCC/NOR 22 HCC/NOR 22_ADJ/N Group:ADJ/N | RACE:Asian GCMS Batch=4; GCMS Data Name=D-03_1; LCMS Batch=1; LCMSNeg Data Name=NEG_118; LCMSPos Data Name=POS_118 SUBJECT_SAMPLE_FACTORS HCC/NOR 23 HCC/NOR 23_HCC/N Group:HCC/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-08_1; LCMS Batch=1; LCMSNeg Data Name=NEG_126; LCMSPos Data Name=POS_126 SUBJECT_SAMPLE_FACTORS HCC/NOR 23 HCC/NOR 23_ADJ/N Group:ADJ/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-20_1; LCMS Batch=1; LCMSNeg Data Name=NEG_125; LCMSPos Data Name=POS_125 SUBJECT_SAMPLE_FACTORS HCC/NOR 24 HCC/NOR 24_HCC/N Group:HCC/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-04_1; LCMS Batch=1; LCMSNeg Data Name=NEG_129; LCMSPos Data Name=POS_129 SUBJECT_SAMPLE_FACTORS HCC/NOR 24 HCC/NOR 24_ADJ/N Group:ADJ/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-10_1; LCMS Batch=1; LCMSNeg Data Name=NEG_128; LCMSPos Data Name=POS_128 SUBJECT_SAMPLE_FACTORS HCC/NOR 25 HCC/NOR 25_HCC/N Group:HCC/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-02_1; LCMS Batch=1; LCMSNeg Data Name=NEG_132; LCMSPos Data Name=POS_132 SUBJECT_SAMPLE_FACTORS HCC/NOR 25 HCC/NOR 25_ADJ/N Group:ADJ/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-07_1; LCMS Batch=1; LCMSNeg Data Name=NEG_131; LCMSPos Data Name=POS_131 SUBJECT_SAMPLE_FACTORS HCC/NOR 26 HCC/NOR 26_HCC/N Group:HCC/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-01_1; LCMS Batch=1; LCMSNeg Data Name=NEG_135; LCMSPos Data Name=POS_135 SUBJECT_SAMPLE_FACTORS HCC/NOR 26 HCC/NOR 26_ADJ/N Group:ADJ/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-16_1; LCMS Batch=1; LCMSNeg Data Name=NEG_134; LCMSPos Data Name=POS_134 SUBJECT_SAMPLE_FACTORS HCC/NOR 27 HCC/NOR 27_HCC/N Group:HCC/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-19_1; LCMS Batch=1; LCMSNeg Data Name=NEG_138; LCMSPos Data Name=POS_138 SUBJECT_SAMPLE_FACTORS HCC/NOR 27 HCC/NOR 27_ADJ/N Group:ADJ/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-17_1; LCMS Batch=1; LCMSNeg Data Name=NEG_137; LCMSPos Data Name=POS_137 SUBJECT_SAMPLE_FACTORS HCC/NOR 28 HCC/NOR 28_HCC/N Group:HCC/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-13_1; LCMS Batch=1; LCMSNeg Data Name=NEG_140; LCMSPos Data Name=POS_140 SUBJECT_SAMPLE_FACTORS HCC/NOR 28 HCC/NOR 28_ADJ/N Group:ADJ/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-06_1; LCMS Batch=1; LCMSNeg Data Name=NEG_139; LCMSPos Data Name=POS_139 SUBJECT_SAMPLE_FACTORS HCC/NOR 29 HCC/NOR 29_HCC/N Group:HCC/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-15_1; LCMS Batch=1; LCMSNeg Data Name=NEG_143; LCMSPos Data Name=POS_143 SUBJECT_SAMPLE_FACTORS HCC/NOR 29 HCC/NOR 29_ADJ/N Group:ADJ/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-11_1; LCMS Batch=1; LCMSNeg Data Name=NEG_142; LCMSPos Data Name=POS_142 SUBJECT_SAMPLE_FACTORS HCC/NOR 30 HCC/NOR 30_HCC/N Group:HCC/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-21_1; LCMS Batch=1; LCMSNeg Data Name=NEG_145; LCMSPos Data Name=POS_145 SUBJECT_SAMPLE_FACTORS HCC/NOR 30 HCC/NOR 30_ADJ/N Group:ADJ/N | RACE:Asian GCMS Batch=5; GCMS Data Name=E-05_1; LCMS Batch=1; LCMSNeg Data Name=NEG_144; LCMSPos Data Name=POS_144 SUBJECT_SAMPLE_FACTORS - Blank_A Group:Blank | RACE:- GCMS Batch=1; GCMS Data Name=Blank_A_1; LCMS Batch=1; LCMSNeg Data Name=NEG_4; LCMSPos Data Name=POS_4 SUBJECT_SAMPLE_FACTORS - Blank_B Group:Blank | RACE:- GCMS Batch=2; GCMS Data Name=Blank_B_1; LCMS Batch=1; LCMSNeg Data Name=NEG_5; LCMSPos Data Name=POS_5 SUBJECT_SAMPLE_FACTORS - Blank_C Group:Blank | RACE:- GCMS Batch=3; GCMS Data Name=Blank_C_1; LCMS Batch=1; LCMSNeg Data Name=NEG_6; LCMSPos Data Name=POS_6 SUBJECT_SAMPLE_FACTORS - Blank_D Group:Blank | RACE:- GCMS Batch=4; GCMS Data Name=Blank_D_1; LCMS Batch=1; LCMSNeg Data Name=NEG_7; LCMSPos Data Name=POS_7 SUBJECT_SAMPLE_FACTORS - Blank_E Group:Blank | RACE:- GCMS Batch=5; GCMS Data Name=Blank_E_1; LCMS Batch=1; LCMSNeg Data Name=NEG_8; LCMSPos Data Name=POS_8 SUBJECT_SAMPLE_FACTORS - QC_HCC/C_1 Group:QC_HCC/C | RACE:- GCMS Batch=1; GCMS Data Name=QC_HCC_C_1_1; LCMS Batch=1; LCMSNeg Data Name=NEG_17; LCMSPos Data Name=POS_17 SUBJECT_SAMPLE_FACTORS - QC_HCC/C_2 Group:QC_HCC/C | RACE:- GCMS Batch=1; GCMS Data Name=QC_HCC_C_2_1; LCMS Batch=1; LCMSNeg Data Name=NEG_43; LCMSPos Data Name=POS_43 SUBJECT_SAMPLE_FACTORS - QC_HCC/C_3 Group:QC_HCC/C | RACE:- GCMS Batch=1; GCMS Data Name=QC_HCC_C_3_1; LCMS Batch=1; LCMSNeg Data Name=NEG_69; LCMSPos Data Name=POS_69 SUBJECT_SAMPLE_FACTORS - QC_HCC/C_4 Group:QC_HCC/C | RACE:- GCMS Batch=2; GCMS Data Name=QC_HCC_C_4_1; LCMS Batch=1; LCMSNeg Data Name=NEG_95; LCMSPos Data Name=POS_95 SUBJECT_SAMPLE_FACTORS - QC_HCC/C_5 Group:QC_HCC/C | RACE:- GCMS Batch=2; GCMS Data Name=QC_HCC_C_5_1; LCMS Batch=1; LCMSNeg Data Name=NEG_121; LCMSPos Data Name=POS_121 SUBJECT_SAMPLE_FACTORS - QC_HCC/C_6 Group:QC_HCC/C | RACE:- GCMS Batch=2; GCMS Data Name=QC_HCC_C_6_1; LCMS Batch=1; LCMSNeg Data Name=NEG_147; LCMSPos Data Name=POS_147 SUBJECT_SAMPLE_FACTORS - QC_HCC/N_1 Group:QC_HCC/N | RACE:- GCMS Batch=1; GCMS Data Name=QC_HCC_N_1_1; LCMS Batch=1; LCMSNeg Data Name=NEG_20; LCMSPos Data Name=POS_20 SUBJECT_SAMPLE_FACTORS - QC_HCC/N_2 Group:QC_HCC/N | RACE:- GCMS Batch=1; GCMS Data Name=QC_HCC_N_2_1; LCMS Batch=1; LCMSNeg Data Name=NEG_46; LCMSPos Data Name=POS_46 SUBJECT_SAMPLE_FACTORS - QC_HCC/N_3 Group:QC_HCC/N | RACE:- GCMS Batch=1; GCMS Data Name=QC_HCC_N_3_1; LCMS Batch=1; LCMSNeg Data Name=NEG_72; LCMSPos Data Name=POS_72 SUBJECT_SAMPLE_FACTORS - QC_HCC/N_4 Group:QC_HCC/N | RACE:- GCMS Batch=2; GCMS Data Name=QC_HCC_N_4_1; LCMS Batch=1; LCMSNeg Data Name=NEG_98; LCMSPos Data Name=POS_98 SUBJECT_SAMPLE_FACTORS - QC_HCC/N_5 Group:QC_HCC/N | RACE:- GCMS Batch=2; GCMS Data Name=QC_HCC_N_5_1; LCMS Batch=1; LCMSNeg Data Name=NEG_124; LCMSPos Data Name=POS_124 SUBJECT_SAMPLE_FACTORS - QC_HCC/N_6 Group:QC_HCC/N | RACE:- GCMS Batch=2; GCMS Data Name=QC_HCC_N_6_1; LCMS Batch=1; LCMSNeg Data Name=NEG_150; LCMSPos Data Name=POS_150 SUBJECT_SAMPLE_FACTORS - QC_HCC/N_7 Group:QC_HCC/N | RACE:- GCMS Batch=3; GCMS Data Name=QC_HCC_N_7_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_HCC/N_8 Group:QC_HCC/N | RACE:- GCMS Batch=3; GCMS Data Name=QC_HCC_N_8_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_HCC/N_9 Group:QC_HCC/N | RACE:- GCMS Batch=3; GCMS Data Name=QC_HCC_N_9_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_HCC/N_10 Group:QC_HCC/N | RACE:- GCMS Batch=4; GCMS Data Name=QC_HCC_N_10_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_HCC/N_11 Group:QC_HCC/N | RACE:- GCMS Batch=4; GCMS Data Name=QC_HCC_N_11_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_HCC/N_12 Group:QC_HCC/N | RACE:- GCMS Batch=4; GCMS Data Name=QC_HCC_N_12_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_HCC/N_13 Group:QC_HCC/N | RACE:- GCMS Batch=5; GCMS Data Name=QC_HCC_N_13_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_HCC/N_14 Group:QC_HCC/N | RACE:- GCMS Batch=5; GCMS Data Name=QC_HCC_N_14_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_HCC/N_15 Group:QC_HCC/N | RACE:- GCMS Batch=5; GCMS Data Name=QC_HCC_N_15_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_ADJ/C_1 Group:QC_ADJ/C | RACE:- GCMS Batch=1; GCMS Data Name=QC_ADJ_C_1_1; LCMS Batch=1; LCMSNeg Data Name=NEG_16; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_ADJ/C_2 Group:QC_ADJ/C | RACE:- GCMS Batch=1; GCMS Data Name=QC_ADJ_C_2_1; LCMS Batch=1; LCMSNeg Data Name=NEG_42; LCMSPos Data Name=POS_42 SUBJECT_SAMPLE_FACTORS - QC_ADJ/C_3 Group:QC_ADJ/C | RACE:- GCMS Batch=1; GCMS Data Name=QC_ADJ_C_3_1; LCMS Batch=1; LCMSNeg Data Name=NEG_68; LCMSPos Data Name=POS_68 SUBJECT_SAMPLE_FACTORS - QC_ADJ/C_4 Group:QC_ADJ/C | RACE:- GCMS Batch=2; GCMS Data Name=QC_ADJ_C_4_1; LCMS Batch=1; LCMSNeg Data Name=NEG_94; LCMSPos Data Name=POS_94 SUBJECT_SAMPLE_FACTORS - QC_ADJ/C_5 Group:QC_ADJ/C | RACE:- GCMS Batch=2; GCMS Data Name=QC_ADJ_C_5_1; LCMS Batch=1; LCMSNeg Data Name=NEG_120; LCMSPos Data Name=POS_120 SUBJECT_SAMPLE_FACTORS - QC_ADJ/C_6 Group:QC_ADJ/C | RACE:- GCMS Batch=2; GCMS Data Name=QC_ADJ_C_6_1; LCMS Batch=1; LCMSNeg Data Name=NEG_146; LCMSPos Data Name=POS_146 SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_1 Group:QC_ADJ/N | RACE:- GCMS Batch=1; GCMS Data Name=QC_ADJ_N_1_1; LCMS Batch=1; LCMSNeg Data Name=NEG_19; LCMSPos Data Name=POS_19 SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_2 Group:QC_ADJ/N | RACE:- GCMS Batch=1; GCMS Data Name=QC_ADJ_N_2_1; LCMS Batch=1; LCMSNeg Data Name=NEG_45; LCMSPos Data Name=POS_45 SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_3 Group:QC_ADJ/N | RACE:- GCMS Batch=1; GCMS Data Name=QC_ADJ_N_3_1; LCMS Batch=1; LCMSNeg Data Name=NEG_71; LCMSPos Data Name=POS_71 SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_4 Group:QC_ADJ/N | RACE:- GCMS Batch=2; GCMS Data Name=QC_ADJ_N_4_1; LCMS Batch=1; LCMSNeg Data Name=NEG_97; LCMSPos Data Name=POS_97 SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_5 Group:QC_ADJ/N | RACE:- GCMS Batch=2; GCMS Data Name=QC_ADJ_N_5_1; LCMS Batch=1; LCMSNeg Data Name=NEG_123; LCMSPos Data Name=POS_123 SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_6 Group:QC_ADJ/N | RACE:- GCMS Batch=2; GCMS Data Name=QC_ADJ_N_6_1; LCMS Batch=1; LCMSNeg Data Name=NEG_149; LCMSPos Data Name=POS_149 SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_7 Group:QC_ADJ/N | RACE:- GCMS Batch=3; GCMS Data Name=QC_ADJ_N_7_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_8 Group:QC_ADJ/N | RACE:- GCMS Batch=3; GCMS Data Name=QC_ADJ_N_8_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_9 Group:QC_ADJ/N | RACE:- GCMS Batch=3; GCMS Data Name=QC_ADJ_N_9_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_10 Group:QC_ADJ/N | RACE:- GCMS Batch=4; GCMS Data Name=QC_ADJ_N_10_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_11 Group:QC_ADJ/N | RACE:- GCMS Batch=4; GCMS Data Name=QC_ADJ_N_11_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_12 Group:QC_ADJ/N | RACE:- GCMS Batch=4; GCMS Data Name=QC_ADJ_N_12_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_13 Group:QC_ADJ/N | RACE:- GCMS Batch=5; GCMS Data Name=QC_ADJ_N_13_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_14 Group:QC_ADJ/N | RACE:- GCMS Batch=5; GCMS Data Name=QC_ADJ_N_14_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- SUBJECT_SAMPLE_FACTORS - QC_ADJ/N_15 Group:QC_ADJ/N | RACE:- GCMS Batch=5; GCMS Data Name=QC_ADJ_N_15_1; LCMS Batch=1; LCMSNeg Data Name=-; LCMSPos Data Name=- #COLLECTION CO:COLLECTION_SUMMARY Adult patients were recruited at MedStar Georgetown University Hospital (MGUH). CO:COLLECTION_SUMMARY All participants provided informed consent to a protocol approved by the CO:COLLECTION_SUMMARY Institutional Review Board (IRB) at Georgetown University. Following the CO:COLLECTION_SUMMARY participant’s informed consent signature and enrollment, tissue samples were CO:COLLECTION_SUMMARY collected at the time of the surgical procedure and stored under liquid nitrogen CO:COLLECTION_SUMMARY until the day of metabolite extraction. HCC cases were diagnosed based on CO:COLLECTION_SUMMARY well-established diagnostic imaging criteria and/or histology. Clinical stages CO:COLLECTION_SUMMARY for HCC cases were determined based on the TNM staging system. CO:SAMPLE_TYPE Liver #TREATMENT TR:TREATMENT_SUMMARY Tissue samples were collected at the time of the surgical procedure and stored TR:TREATMENT_SUMMARY under liquid nitrogen until the day of metabolite extraction. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY 10 mg of liver tissue was homogenized on ice with 1 mL of pre-chilled SP:SAMPLEPREP_SUMMARY methanol:water (1:1) containing five internal standards: 0.001 ppm debrisoquine, SP:SAMPLEPREP_SUMMARY 0.004 ppm 4-nitro benzoic acid, 0.0004 ppb stearoyl SP:SAMPLEPREP_SUMMARY (d35)-2-hydroxy-glycerophosphocholine, 0.2 ppb D-erythro-sphingosine SP:SAMPLEPREP_SUMMARY (d7)-1-phosphate, and 2 ppm Myristic-d27 acid. Homogenized samples were SP:SAMPLEPREP_SUMMARY centrifuged at 14.500 g, at 4°C for 15 minutes. Supernatant (S1) was collected SP:SAMPLEPREP_SUMMARY and divided in two aliquots (one for GC-MS and one for LC-MS analysis), while SP:SAMPLEPREP_SUMMARY the remaining pellet (P1) was kept at -80°C until further extraction. For the SP:SAMPLEPREP_SUMMARY extraction of medium to polar compounds, 1:1 volume of pre-chilled acetonitrile SP:SAMPLEPREP_SUMMARY was added to the two aliquots (GC&LC), vortex-mixed and kept on ice for 20 SP:SAMPLEPREP_SUMMARY minutes. Samples were centrifuged again at 14.500 g, at 4°C for 15 minutes, and SP:SAMPLEPREP_SUMMARY supernatant (S2) was collected and concentrated to dryness in a speedvac system SP:SAMPLEPREP_SUMMARY operated at room temperature. Pellets (P2 and P3) and supernatants were stored SP:SAMPLEPREP_SUMMARY at -80°C. For the extraction of low-polar compounds, P1 was resuspended with SP:SAMPLEPREP_SUMMARY 500μl of pre-chilled dichloromethane:methanol (3:1) while P2, and P3 with 125 SP:SAMPLEPREP_SUMMARY μL of the same mix. Pellets were sonicated on ice for 90sec, combined and SP:SAMPLEPREP_SUMMARY centrifuged at 14.500 g, at 4°C for 20 minutes. Supernatant (S3) was collected, SP:SAMPLEPREP_SUMMARY split in two aliquots (one for GC-MS and one for LC-MS analysis) and a 1:1 SP:SAMPLEPREP_SUMMARY volume of pre-chilled acetonitrile was added to both aliquots (GC&LC), vortexed SP:SAMPLEPREP_SUMMARY and kept on ice for 20 minutes. Samples were then centrifuged at 14.500 g for 15 SP:SAMPLEPREP_SUMMARY min at 4°C while pellets were kept at -80 °C for protein quantitation. SP:SAMPLEPREP_SUMMARY Finally, supernatants (S4) were concentrated to dryness by speedvac and kept at SP:SAMPLEPREP_SUMMARY -80°C until the day of analysis. Blank samples were prepared together with the SP:SAMPLEPREP_SUMMARY human samples by adding all the reagents to an empty tube and following the same SP:SAMPLEPREP_SUMMARY sample preparation steps. Dried supernatant (S2) collected for GC-MS analysis SP:SAMPLEPREP_SUMMARY underwent derivatization step. Samples were derivatized in each batch prior to SP:SAMPLEPREP_SUMMARY injection following a two-stage process of oximation and trimethylsilylation SP:SAMPLEPREP_SUMMARY (-Si(CH3)3). Briefly, 20µL of a 20mg/mL methoxyamine hydrochloride in pyridine SP:SAMPLEPREP_SUMMARY were added to the dried extracts, vortexed and incubated at 30°C for 90 SP:SAMPLEPREP_SUMMARY minutes. After returning the samples to room temperature, 80µL of MSTFA were SP:SAMPLEPREP_SUMMARY added, vortex-mixed and incubated at 30°C for 30 minutes. Samples were then SP:SAMPLEPREP_SUMMARY centrifuged at 14,500 g for 15 minutes, and 60µL of the supernatant were SP:SAMPLEPREP_SUMMARY transferred into 250µL clear glass autosampler vials. Finally, 20µL of 0.006 SP:SAMPLEPREP_SUMMARY µg/µL C18 Methyl Stearate in hexane were added to the vial prior to injection. SP:SAMPLEPREP_SUMMARY For quality assessment, Myristic-d27 acid was spiked into the working solution SP:SAMPLEPREP_SUMMARY to verify tissue metabolites extraction and derivatization steps. C18 Methyl SP:SAMPLEPREP_SUMMARY Stearate was added just before GC-MS analysis to monitor each sample injection. SP:SAMPLEPREP_SUMMARY QC samples (QCs) were generated by pooling together the supernatant obtained SP:SAMPLEPREP_SUMMARY after derivatization of samples of each biological group in each batch SP:SAMPLEPREP_SUMMARY separately. A retention index (RI) standard sample was prepared by mixing fatty SP:SAMPLEPREP_SUMMARY acid methyl esters (FAMEs) with alkanes. Specifically, FAMEs C8, C9, C10, C12, SP:SAMPLEPREP_SUMMARY C14, C16, C18, C20, C22, C24, C26, C28 and C30 linear chain length were SP:SAMPLEPREP_SUMMARY individually dissolved in chloroform at concentrations of 0.8 mg/mL (C8 – C16) SP:SAMPLEPREP_SUMMARY and 0.4 mg/ml (C18 – C30) to generate FAME-1 stock solutions. 100 µL of each SP:SAMPLEPREP_SUMMARY FAME-1 were mixed together and 1.2 mL of chloroform were added for a final SP:SAMPLEPREP_SUMMARY volume of 2.5mL generating FAME-2 solution. Alkanes, containing all even CnH2n+2 SP:SAMPLEPREP_SUMMARY from C10 to C40, were purchased as a mixture at a concentration of 50mg/L in SP:SAMPLEPREP_SUMMARY n-heptane. Alkane mixture were mixed with FAME-2 markers and hexane at a ratio SP:SAMPLEPREP_SUMMARY of 1:2:17 and vortex-mixed prior to injection into the GC-MS. Dried supernatants SP:SAMPLEPREP_SUMMARY (S2 & S4) collected from the first and second extraction for LC-MS analysis were SP:SAMPLEPREP_SUMMARY reconstituted with 125 μL of methanol:acetonitrile:water (50:25:25) each, and SP:SAMPLEPREP_SUMMARY combined for a total volume of 250 μL. d35-lysophosphocholine and SP:SAMPLEPREP_SUMMARY d7-sphingosine-1-phosphate were included to determine the quality of the SP:SAMPLEPREP_SUMMARY metabolite extraction. Debrisoquine and 4-nitrobenzoic acid were used to assess SP:SAMPLEPREP_SUMMARY equipment performance. QCs were generated by pooling together the supernatant SP:SAMPLEPREP_SUMMARY obtained after resuspension in appropriate solvent of each biological group. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Waters Acquity CH:COLUMN_NAME Waters Acquity CSH C18 (100 x 2.1mm, 1.7um) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Waters Synapt G2 Si QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE NEGATIVE MS:MS_COMMENTS Software XCMS MS:MS_RESULTS_FILE ST001152_AN001902_Results.txt UNITS:Da Has m/z:Yes Has RT:Yes RT units:Seconds #END