#METABOLOMICS WORKBENCH NicholasDevanney_20220920_144226 DATATRACK_ID:3467 STUDY_ID:ST002450 ANALYSIS_ID:AN003998 VERSION 1 CREATED_ON 01-23-2023 #PROJECT PR:PROJECT_TITLE APOE modulates microglial immunometabolism in response to age, amyloid PR:PROJECT_TITLE pathology, and inflammatory challenge PR:PROJECT_SUMMARY The E4 allele of Apolipoprotein E (APOE) is associated with both metabolic PR:PROJECT_SUMMARY dysfunction and a heightened pro-inflammatory response – two findings that may PR:PROJECT_SUMMARY be intrinsically linked through the concept of immunometabolism. Here, we PR:PROJECT_SUMMARY combined bulk, single-cell, and spatial transcriptomics with cell-specific and PR:PROJECT_SUMMARY spatially resolved metabolic analyses to systematically address the role of APOE PR:PROJECT_SUMMARY across age, neuroinflammation, and AD pathology. RNAseq highlighted PR:PROJECT_SUMMARY immunometabolic changes across the APOE4 glial transcriptome, specifically in PR:PROJECT_SUMMARY subsets of metabolically distinct microglia enriched in the E4 brain during PR:PROJECT_SUMMARY aging or following an inflammatory challenge. E4 microglia display increased PR:PROJECT_SUMMARY Hif1α expression, a disrupted TCA cycle, and are inherently pro-glycolytic, PR:PROJECT_SUMMARY while spatial transcriptomics and MALDI mass spectrometry imaging highlight an PR:PROJECT_SUMMARY E4-specific response to amyloid that is characterized by widespread alterations PR:PROJECT_SUMMARY in lipid metabolism. Taken together, our findings emphasize a central role for PR:PROJECT_SUMMARY APOE in regulating microglial immunometabolism. PR:INSTITUTE University of Kentucky, Department of Physiology PR:LAST_NAME Devanney PR:FIRST_NAME Nicholas PR:ADDRESS Physiology, 760 Press Ave, Healthy Kentucky Research Bldg, Rm152, Lexington, PR:ADDRESS Kentucky, 40508, USA PR:EMAIL Nicholas.Devanney@uky.edu PR:PHONE 8593238083 PR:PROJECT_COMMENTS Part 2 of 3 PR:DOI http://dx.doi.org/10.21228/M8G711 #STUDY ST:STUDY_TITLE APOE modulates microglial immunometabolism in response to age, amyloid ST:STUDY_TITLE pathology, and inflammatory challenge (Part 1 of 3) ST:STUDY_SUMMARY The E4 allele of Apolipoprotein E (APOE) is associated with both metabolic ST:STUDY_SUMMARY dysfunction and a heightened pro-inflammatory response – two findings that may ST:STUDY_SUMMARY be intrinsically linked through the concept of immunometabolism. Here, we ST:STUDY_SUMMARY combined bulk, single-cell, and spatial transcriptomics with cell-specific and ST:STUDY_SUMMARY spatially resolved metabolic analyses to systematically address the role of APOE ST:STUDY_SUMMARY across age, neuroinflammation, and AD pathology. RNAseq highlighted ST:STUDY_SUMMARY immunometabolic changes across the APOE4 glial transcriptome, specifically in ST:STUDY_SUMMARY subsets of metabolically distinct microglia enriched in the E4 brain during ST:STUDY_SUMMARY aging or following an inflammatory challenge. E4 microglia display increased ST:STUDY_SUMMARY Hif1α expression, a disrupted TCA cycle, and are inherently pro-glycolytic, ST:STUDY_SUMMARY while spatial transcriptomics and MALDI mass spectrometry imaging highlight an ST:STUDY_SUMMARY E4-specific response to amyloid that is characterized by widespread alterations ST:STUDY_SUMMARY in lipid metabolism. Taken together, our findings emphasize a central role for ST:STUDY_SUMMARY APOE in regulating microglial immunometabolism. ST:INSTITUTE University of Kentucky, Department of Physiology ST:LAST_NAME Devanney ST:FIRST_NAME Nicholas ST:ADDRESS Physiology, 760 Press Ave, Healthy Kentucky Research Bldg, Rm152, Lexington, ST:ADDRESS Kentucky, 40508, USA ST:EMAIL Nicholas.Devanney@uky.edu ST:PHONE 8593238083 ST:SUBMIT_DATE 2022-09-20 #SUBJECT SU:SUBJECT_TYPE Cultured cells SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN Primary microglia isolated from APOE-targeted replacement mice homozygous for SU:GENOTYPE_STRAIN human E3 (B6.129P2-Apoe^tm2(APOE*3)Mae N8, Taconic #1548-F) or human E4 SU:GENOTYPE_STRAIN (B6.129P2- Apoe^tm3(APOE*4)Mae N8, Taconic #1549-F) alleles SU:AGE_OR_AGE_RANGE P0-P3 SU:GENDER Pooled #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS - 15_3_2 Genotype:E3 RAW_FILE_NAME=15_3_2.CDF; Protein Amount (BCA)=725.828 SUBJECT_SAMPLE_FACTORS - 15_3_3 Genotype:E3 RAW_FILE_NAME=15_3_3.CDF; Protein Amount (BCA)=1044.435 SUBJECT_SAMPLE_FACTORS - 16_3_2 Genotype:E3 RAW_FILE_NAME=16_3_2.CDF; Protein Amount (BCA)=1534.876 SUBJECT_SAMPLE_FACTORS - 16_3_3_1 Genotype:E3 RAW_FILE_NAME=16_3_3_1.CDF; Protein Amount (BCA)=791.458 SUBJECT_SAMPLE_FACTORS - 16_3_3_2 Genotype:E3 RAW_FILE_NAME=16_3_3_2.CDF; Protein Amount (BCA)=1050.401 SUBJECT_SAMPLE_FACTORS - 16_3_4 Genotype:E3 RAW_FILE_NAME=16_3_4.CDF; Protein Amount (BCA)=727.021 SUBJECT_SAMPLE_FACTORS - 16_3_5 Genotype:E3 RAW_FILE_NAME=16_3_5.CDF; Protein Amount (BCA)=742.534 SUBJECT_SAMPLE_FACTORS - 16_3_6 Genotype:E3 RAW_FILE_NAME=16_3_6.CDF; Protein Amount (BCA)=834.417 SUBJECT_SAMPLE_FACTORS - 3_3 Genotype:E3 RAW_FILE_NAME=3_3.CDF; Protein Amount (BCA)=1361.61 SUBJECT_SAMPLE_FACTORS - 3_4 Genotype:E3 RAW_FILE_NAME=3_4.CDF; Protein Amount (BCA)=1174.855 SUBJECT_SAMPLE_FACTORS - 3_5 Genotype:E3 RAW_FILE_NAME=3_5.CDF; Protein Amount (BCA)=894.722 SUBJECT_SAMPLE_FACTORS - 3_6 Genotype:E3 RAW_FILE_NAME=3_6.CDF; Protein Amount (BCA)=1162.522 SUBJECT_SAMPLE_FACTORS - 3_7 Genotype:E3 RAW_FILE_NAME=3_7.CDF; Protein Amount (BCA)=1305.231 SUBJECT_SAMPLE_FACTORS - 3_8 Genotype:E3 RAW_FILE_NAME=3_8.CDF; Protein Amount (BCA)=1078.834 SUBJECT_SAMPLE_FACTORS - 3_9 Genotype:E3 RAW_FILE_NAME=3_9.CDF; Protein Amount (BCA)=762.435 SUBJECT_SAMPLE_FACTORS - MG14_1 Genotype:E3 RAW_FILE_NAME=MG14_1.CDF; Protein Amount (BCA)=767.228 SUBJECT_SAMPLE_FACTORS - MG14_2 Genotype:E3 RAW_FILE_NAME=MG14_2.CDF; Protein Amount (BCA)=687.234 SUBJECT_SAMPLE_FACTORS - MG15_2 Genotype:E3 RAW_FILE_NAME=MG15_2.CDF; Protein Amount (BCA)=395.081 SUBJECT_SAMPLE_FACTORS - MG15_3 Genotype:E3 RAW_FILE_NAME=MG15_3.CDF; Protein Amount (BCA)=424.644 SUBJECT_SAMPLE_FACTORS - MG16_E3_1 Genotype:E3 RAW_FILE_NAME=MG16_E3_1.CDF; Protein Amount (BCA)=312.99 SUBJECT_SAMPLE_FACTORS - MG16_E3_1_1 Genotype:E3 RAW_FILE_NAME=MG16_E3_1_1.CDF; Protein Amount (BCA)=380.592 SUBJECT_SAMPLE_FACTORS - MG16_E3_2 Genotype:E3 RAW_FILE_NAME=MG16_E3_2.CDF; Protein Amount (BCA)=435.457 SUBJECT_SAMPLE_FACTORS - 15_4_4 Genotype:E4 RAW_FILE_NAME=15_4_4.CDF; Protein Amount (BCA)=693.609 SUBJECT_SAMPLE_FACTORS - 15_4_5 Genotype:E4 RAW_FILE_NAME=15_4_5.CDF; Protein Amount (BCA)=795.038 SUBJECT_SAMPLE_FACTORS - 16_4_3 Genotype:E4 RAW_FILE_NAME=16_4_3.CDF; Protein Amount (BCA)=914.367 SUBJECT_SAMPLE_FACTORS - 16_4_4 Genotype:E4 RAW_FILE_NAME=16_4_4.CDF; Protein Amount (BCA)=828.45 SUBJECT_SAMPLE_FACTORS - 16_4_5_1 Genotype:E4 RAW_FILE_NAME=16_4_5_1.CDF; Protein Amount (BCA)=1150.638 SUBJECT_SAMPLE_FACTORS - 16_4_5_2 Genotype:E4 RAW_FILE_NAME=16_4_5_2.CDF; Protein Amount (BCA)=815.324 SUBJECT_SAMPLE_FACTORS - 16_4_6_1 Genotype:E4 RAW_FILE_NAME=16_4_6_1.CDF; Protein Amount (BCA)=742.534 SUBJECT_SAMPLE_FACTORS - 16_4_6_2 Genotype:E4 RAW_FILE_NAME=16_4_6_2.CDF; Protein Amount (BCA)=976.418 SUBJECT_SAMPLE_FACTORS - 4_1 Genotype:E4 RAW_FILE_NAME=4_1.CDF; Protein Amount (BCA)=1678.742 SUBJECT_SAMPLE_FACTORS - 4_2 Genotype:E4 RAW_FILE_NAME=4_2.CDF; Protein Amount (BCA)=1381.871 SUBJECT_SAMPLE_FACTORS - 4_3 Genotype:E4 RAW_FILE_NAME=4_3.CDF; Protein Amount (BCA)=968.719 SUBJECT_SAMPLE_FACTORS - 4_4 Genotype:E4 RAW_FILE_NAME=4_4.CDF; Protein Amount (BCA)=959.029 SUBJECT_SAMPLE_FACTORS - 4_5 Genotype:E4 RAW_FILE_NAME=4_5.CDF; Protein Amount (BCA)=1005.718 SUBJECT_SAMPLE_FACTORS - 4_6 Genotype:E4 RAW_FILE_NAME=4_6.CDF; Protein Amount (BCA)=1177.497 SUBJECT_SAMPLE_FACTORS - 4_7 Genotype:E4 RAW_FILE_NAME=4_7.CDF; Protein Amount (BCA)=1149.308 SUBJECT_SAMPLE_FACTORS - 4_8 Genotype:E4 RAW_FILE_NAME=4_8.CDF; Protein Amount (BCA)=1336.944 SUBJECT_SAMPLE_FACTORS - MG15_4 Genotype:E4 RAW_FILE_NAME=MG15_4.CDF; Protein Amount (BCA)=254.222 SUBJECT_SAMPLE_FACTORS - MG15_5 Genotype:E4 RAW_FILE_NAME=MG15_5.CDF; Protein Amount (BCA)=389.864 SUBJECT_SAMPLE_FACTORS - MG16_E4_1 Genotype:E4 RAW_FILE_NAME=MG16_E4_1.CDF; Protein Amount (BCA)=300.254 SUBJECT_SAMPLE_FACTORS - MG16_E4_2 Genotype:E4 RAW_FILE_NAME=MG16_E4_2.CDF; Protein Amount (BCA)=338.463 SUBJECT_SAMPLE_FACTORS - MG16_E4_4 Genotype:E4 RAW_FILE_NAME=MG16_E4_4.CDF; Protein Amount (BCA)=468.768 #COLLECTION CO:COLLECTION_SUMMARY Primary microglia were plated at 7x106 cells/well in 6-well plates (VWR CO:COLLECTION_SUMMARY #10062-894) and incubated at 5% CO2 37°C. Upon reaching confluence, cells were CO:COLLECTION_SUMMARY removed from the incubator washed with warm 0.9% NaCl solution. Culture plates CO:COLLECTION_SUMMARY were placed on a bed of crushed dry ice and 1mL of ice cold 50% methanol CO:COLLECTION_SUMMARY (HPLC-grade, Sigma #A456-4) was added to quench cellular metabolic activity CO:COLLECTION_SUMMARY followed by a 10 minute incubation at -80°C to ensure cell lysis. After CO:COLLECTION_SUMMARY removing from the freezer, cells were detached with a cell scraper (VWR CO:COLLECTION_SUMMARY #10062-906) and the entire contents collected into a microcentrifuge tube, CO:COLLECTION_SUMMARY vortexed briefly, and placed on ice until all samples were collected. The tubes CO:COLLECTION_SUMMARY were then placed on a Disruptor Genie Cell Disruptor Homogenizer (Scientific CO:COLLECTION_SUMMARY Industries) for 5 min at 3,000 rpm. Tubes were then centrifuged at 20,000 x g CO:COLLECTION_SUMMARY for 10 min at 4 °C. The supernatant containing polar metabolites was isolated CO:COLLECTION_SUMMARY to a new tube and stored at -80C, and the resulting pellet was briefly dried at CO:COLLECTION_SUMMARY 10-3 mbar using a CentriVap vacuum concentrator (LabConco) to evaporate CO:COLLECTION_SUMMARY remaining methanol, followed by determination of protein content via BCA assay CO:COLLECTION_SUMMARY (ThermoFisher #23225) to normalize metabolite concentrations to total protein CO:COLLECTION_SUMMARY amount of each sample. CO:SAMPLE_TYPE Cultured cells CO:COLLECTION_METHOD Ice-cold methanol was added to primary microglia cultures in 6-well plates after CO:COLLECTION_METHOD which cells were harvested using a cell scraper CO:COLLECTION_LOCATION University of Kentucky CO:VOLUMEORAMOUNT_COLLECTED 1ml CO:STORAGE_CONDITIONS -80℃ CO:COLLECTION_TUBE_TEMP -80 #TREATMENT TR:TREATMENT_SUMMARY Cells were not treated, as we were just comparing APOE3/3 vs. APOE4/4 genotypes TR:CELL_GROWTH_CONTAINER 6-well plate TR:CELL_MEDIA DMEM/F12 with 10% FBS, 10% LCCM supplement (see full methods in associated TR:CELL_MEDIA publication for preparation details), and 1% Penicillin/Streptomycin TR:CELL_ENVIR_COND 37C, 5% CO2 TR:CELL_HARVESTING Cell scraper TR:CELL_PCT_CONFLUENCE 100 TR:CELL_MEDIA_LASTCHANGED 2-4 days prior to harvest #SAMPLEPREP SP:SAMPLEPREP_SUMMARY The supernatant fraction containing polar metabolites was thawed gently on ice SP:SAMPLEPREP_SUMMARY and dried at 10-3 mbar using a CentriVap vacuum concentrator (LabConco) to SP:SAMPLEPREP_SUMMARY evaporate methanol. The dried polar metabolite pellet was derivatized by a SP:SAMPLEPREP_SUMMARY two-step methoxyamine protocol first by addition of 70µL methoxyamine HCl SP:SAMPLEPREP_SUMMARY (Sigma-Aldrich #226904-5G) in pyridine (20 mg/mL; Sigma-Aldrich #TS25730) to SP:SAMPLEPREP_SUMMARY each pellet followed by 90 min dry heat incubation at 30°C. Samples were then SP:SAMPLEPREP_SUMMARY centrifuged at 20,000 x g for 10 minutes after which 50µL of each sample was SP:SAMPLEPREP_SUMMARY transferred to an amber V-shaped glass chromatography vial (Agilent #5184-3554) SP:SAMPLEPREP_SUMMARY containing 80µL N-methyl-trimethylsilyl-trifluoroacetamide (MSTFA; ThermoFisher SP:SAMPLEPREP_SUMMARY #TS48915) and gently vortexed followed by 30 min dry heat incubation at 37°C. SP:EXTRACTION_METHOD 50% methanol SP:EXTRACT_STORAGE -80℃ SP:SAMPLE_DERIVATIZATION MSTFA #CHROMATOGRAPHY CH:INSTRUMENT_NAME Agilent 8890 CH:COLUMN_NAME Agilent HP5-MS (30m x 0.25mm, 0.25 um) CH:CHROMATOGRAPHY_TYPE GC #ANALYSIS AN:ANALYSIS_TYPE MS AN:DATA_FORMAT CDF #MS MS:INSTRUMENT_NAME Agilent 5977 MS:INSTRUMENT_TYPE Single quadrupole MS:MS_TYPE EI MS:MS_COMMENTS Metabolites were identified and quantified using DExSI v1.11 MS:MS_COMMENTS (https://github.com/DExSI/DExSI) MS:ION_MODE POSITIVE #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS Relative Abundance MS_METABOLITE_DATA_START Samples 15_3_2 15_3_3 16_3_2 16_3_3_1 16_3_3_2 16_3_4 16_3_5 16_3_6 3_3 3_4 3_5 3_6 3_7 3_8 3_9 MG14_1 MG14_2 MG15_2 MG15_3 MG16_E3_1 MG16_E3_1_1 MG16_E3_2 15_4_4 15_4_5 16_4_3 16_4_4 16_4_5_1 16_4_5_2 16_4_6_1 16_4_6_2 4_1 4_2 4_3 4_4 4_5 4_6 4_7 4_8 MG15_4 MG15_5 MG16_E4_1 MG16_E4_2 MG16_E4_4 Factors Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E3 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Genotype:E4 Acetohydroxamic acid 1.3345 1.0373 0.5474 1.1751 0.5793 1.0481 1.2073 1.0710 0.5315 0.7902 1.4558 1.1594 0.7151 0.8379 1.5102 0.7432 0.8547 0.9862 1.4159 1.1855 0.9312 0.8833 1.2909 0.7705 0.8610 1.1485 0.5210 1.1634 1.3718 0.8648 0.7988 0.9061 1.0773 0.9953 0.9360 0.5815 1.0597 0.9722 2.3431 1.5107 1.1899 1.0460 0.7369 Aspartic acid 1.0142 0.8628 0.6882 1.2965 1.0850 1.1359 0.9436 0.9739 0.9636 1.1698 0.9430 0.9842 0.8245 0.9229 1.1921 0.7622 0.8149 1.1186 1.3043 0.5068 1.0345 1.4586 0.6156 0.7150 0.6669 1.0248 0.7365 1.2805 0.8570 1.0158 0.9647 1.4236 1.5652 1.7816 1.1412 0.8085 0.8499 0.6275 1.1973 0.8238 1.4898 1.7740 0.7970 beta-Alanine 1.4838 1.2599 0.8805 0.8291 1.3391 0.6480 0.8313 0.7284 0.7213 1.0095 1.1697 0.9649 0.7818 0.9918 1.3610 1.3626 1.2679 0.6507 0.7187 0.6384 1.1249 1.2367 1.0723 1.0780 0.4629 0.6349 0.4693 1.5554 0.5831 1.4889 0.6862 0.6235 0.9915 0.8193 0.9350 0.6522 0.7752 0.6379 0.7266 0.4601 1.4535 1.4560 1.6303 Citric acid 1.1368 0.7991 0.9021 1.1105 1.2151 0.8280 1.0420 0.9663 0.8921 1.1870 0.9818 1.2211 0.8109 0.7779 1.1293 0.8997 1.0806 0.9102 1.1095 0.7060 1.0080 1.2860 0.8373 0.8931 0.7427 1.0180 0.8623 1.8494 0.8952 1.1427 0.9644 1.4346 1.4859 2.0321 1.0845 0.7974 0.6252 0.7142 1.4661 1.0549 1.6460 1.9192 1.9707 Creatinine 1.1609 0.8719 0.9952 0.9920 1.1720 0.9307 1.0296 0.8477 0.9422 1.0582 1.1850 0.9644 0.8995 0.8752 1.0754 1.0406 0.9205 1.0361 1.0028 0.6391 1.0616 1.2993 0.9712 1.1875 0.6302 0.9297 0.6895 1.3105 0.7162 0.8087 0.6487 0.6906 1.0646 1.0924 1.1204 0.9454 0.7284 0.5637 1.4830 1.0252 1.5846 2.0545 1.9450 Cysteine 0.6525 0.5500 0.5912 1.4628 0.8833 1.2529 1.3798 1.2275 1.0325 1.2437 0.8000 1.2651 0.6043 0.8468 1.2075 0.6296 0.6657 1.2884 1.4162 1.2942 0.7092 0.9965 0.7604 0.9007 0.8411 1.0463 0.9353 1.0429 0.9312 0.5878 0.7074 1.1695 1.1199 1.7704 0.7593 0.7630 0.6548 0.5755 3.4285 2.3785 0.6192 0.7531 0.5869 Fructose 0.6842 0.5281 0.6746 1.2858 0.8119 1.2941 1.4432 1.2780 0.9162 1.3189 0.8111 1.3897 0.8074 0.6645 1.0922 0.5096 0.5061 1.2766 1.7078 1.0957 0.9002 1.0041 0.7608 0.5634 1.0434 1.7131 1.4452 1.5812 1.6033 0.9802 1.3941 2.6872 2.2173 3.0898 1.0500 0.7439 0.4607 0.5729 3.2935 2.1108 1.3431 1.5130 1.1109 Fumaric acid 1.1316 0.9282 0.5144 1.3230 0.7410 1.0399 1.2509 1.0711 1.0209 1.1733 0.9404 0.8288 0.7956 0.9394 1.3016 0.7163 0.7017 1.2895 1.2924 0.3175 1.2050 1.4775 0.9641 1.0163 0.8072 0.9426 0.7648 0.7219 0.8867 0.6899 0.7243 0.8327 1.1801 1.1537 1.0034 0.8424 0.8344 0.6904 1.2774 0.8868 1.4909 1.5261 0.5717 gamma-Aminobutyric acid 1.0476 0.8240 0.8273 1.1210 1.2406 0.9000 1.1177 0.9218 0.9786 1.1038 1.0917 0.9587 0.7502 0.8883 1.2286 1.1264 1.3195 0.7577 0.7964 0.3859 1.2456 1.3685 0.9586 0.9107 0.7729 0.9709 0.7774 1.3540 0.8161 1.1509 0.6708 0.7028 1.0254 0.9860 0.9362 0.7950 0.7899 0.6355 0.7751 0.4022 2.1853 2.1601 1.6832 Glucose 0.4416 0.3017 0.6663 1.6385 0.7575 1.4157 1.5260 1.2527 0.9637 1.3534 0.7282 1.3887 0.7894 0.6651 1.1115 0.3616 0.3487 1.4591 1.8307 0.9363 1.0362 1.0275 0.5015 0.2778 1.1083 1.9859 1.4885 1.6575 1.5033 0.9197 1.3021 2.2494 2.1796 2.9068 0.9633 0.7229 0.3688 0.4923 4.0613 2.6810 1.2432 1.4276 0.9158 Glutamic acid 1.1314 0.9913 0.7366 1.1631 1.0558 0.9075 1.0296 0.9846 0.3325 1.0845 1.1300 1.1482 0.8164 1.0199 1.4685 1.1455 1.2873 0.7196 0.8476 0.2976 1.1297 1.5726 0.9275 1.0134 0.5752 0.8420 0.6214 1.4547 0.7296 1.0478 0.9897 1.2426 1.5595 1.7164 1.2723 0.7400 0.9700 0.8217 0.8495 0.5307 2.1112 2.3732 1.0794 Glutamine 0.7625 0.7643 0.5753 1.3089 1.0573 1.2515 1.0962 1.1842 0.0525 1.6299 0.8770 1.4628 1.0086 0.8156 1.1535 0.6923 0.6663 1.2395 1.4019 0.9550 0.8862 1.1587 0.8037 0.8689 0.8712 1.4323 1.2822 1.2450 1.2949 1.0089 1.4760 2.4446 2.5997 3.5637 1.3484 0.9979 0.7508 0.8888 2.5931 1.8534 1.4926 1.8370 1.0688 Glycine 1.1249 1.0893 0.7426 0.9871 1.3602 0.9289 0.9095 0.8574 0.9972 1.2354 0.9300 1.0877 0.7753 0.8307 1.1438 0.9146 0.8985 1.0017 1.1853 1.0724 0.9635 0.9641 0.9492 1.0637 0.7044 0.9915 0.8095 1.5015 0.9063 1.3404 0.9783 1.5551 1.6335 1.8781 1.0063 0.8339 0.7217 0.6713 1.5076 0.9836 1.4603 1.5298 1.1762 Glyoxylic acid 1.0056 1.1406 0.5193 1.1347 0.7457 0.9030 1.1969 1.3543 0.5795 1.0758 0.8789 1.1805 0.5330 0.9738 1.7785 0.8604 1.0282 0.7946 1.3167 1.2534 0.8681 0.8786 1.0881 0.8255 0.7666 1.3159 0.6812 0.9476 1.2996 1.0295 1.0088 1.0926 1.1816 1.2402 0.7358 0.8398 1.1994 0.9362 1.5558 0.9090 1.4058 1.0402 1.1566 Hypotaurine 1.0786 0.9347 0.8176 1.2206 0.9090 0.7016 1.1534 1.1844 0.5881 0.9175 1.2169 1.0987 0.7392 1.0095 1.4301 1.1731 1.2377 0.7133 0.8759 0.6918 1.0709 1.2374 1.0309 0.7333 0.5035 0.8927 0.4727 1.3804 0.8211 1.0319 0.5896 0.5370 0.7897 0.8330 0.8346 0.4559 0.6948 0.6496 1.2525 0.8827 1.3337 1.3213 1.0779 Isoleucine 0.9056 0.6220 0.5756 1.2433 0.9094 1.1500 1.3433 1.2508 1.1230 1.1406 0.8105 1.4429 0.8333 0.7309 0.9188 0.6995 0.8014 1.3737 1.1253 0.9111 1.0066 1.0823 1.0872 0.9774 1.0759 1.3431 1.1710 1.3329 1.2851 0.7067 0.5997 1.9173 1.5713 1.9218 0.8356 0.8635 0.5119 0.6499 1.6231 1.4552 1.3953 1.6315 0.9337 Itaconic acid 1.6489 1.5350 0.3964 0.8727 0.7664 0.8140 1.1429 0.8236 1.0480 1.2313 1.1615 1.1994 0.5665 0.7002 1.0931 0.2654 0.3166 1.7305 1.6876 1.1500 0.7369 1.1131 0.4169 0.4546 0.5718 0.3869 0.6095 0.3232 0.5254 0.5697 0.2071 0.2403 0.3637 0.3216 0.3202 0.2391 0.2506 0.2192 1.5388 1.3652 0.7667 0.7724 0.7227 Lactic acid 0.9082 0.7525 0.5734 1.2449 0.8463 1.2099 1.3156 1.1492 1.0007 1.2766 0.8808 1.2753 0.7522 0.7047 1.1096 0.9219 0.9180 1.0122 1.1479 0.9931 1.0211 0.9858 1.1278 0.9706 0.9798 1.3231 0.9848 1.3085 1.2967 0.9858 1.2682 2.3781 2.0805 2.8081 1.0974 0.8893 0.6808 0.7146 1.5257 1.1021 1.7369 1.6660 1.2518 Leucine 0.8047 0.6290 0.7174 1.1709 1.0425 1.1965 1.2139 1.2251 1.0198 1.2320 0.7753 1.4078 0.7958 0.7794 0.9898 0.7344 0.9817 0.9803 1.3035 1.1497 1.0037 0.8466 0.8058 0.9363 0.9403 1.2559 1.1139 1.4757 1.1644 0.9861 0.7593 2.0283 1.7679 2.4892 0.9471 0.8646 0.5595 0.5934 1.6892 0.9533 1.3212 1.1905 0.8498 Malic acid 1.1341 0.9292 0.4529 1.2114 0.6866 1.1218 1.2788 1.1852 1.0438 1.1089 0.9674 0.9072 0.8045 0.9537 1.2144 0.8242 0.8108 1.1470 1.2179 0.2122 1.3105 1.4773 1.0723 1.0908 0.7013 0.9010 0.8347 0.6579 0.9113 0.5668 0.9547 1.0690 1.5075 1.4651 1.2873 1.0743 0.9349 0.9421 1.1345 0.8854 1.5287 1.7076 0.5769 Ornithine 0.9656 0.2484 0.4833 1.5822 1.2939 0.8300 1.2735 1.3232 0.7576 1.1559 0.9013 1.5025 0.7741 0.7417 1.1669 0.1613 0.9087 0.6106 2.3193 1.3065 0.4999 1.1936 0.6017 0.2427 0.9520 1.3715 2.0472 2.2967 1.7407 0.9588 1.2064 1.6950 1.5502 2.0536 0.9307 0.7436 0.5148 0.6640 2.9741 2.1388 1.7085 1.6846 1.7880 Palmitic acid 1.7990 0.6053 1.0006 0.7422 0.7742 1.2162 0.7340 1.1285 0.6704 0.7409 2.0768 0.7014 1.5075 0.5655 0.7375 0.5156 0.4813 1.3876 1.6156 1.1296 1.0005 0.8699 0.8072 0.9774 0.6809 1.4750 0.9520 1.4584 1.0517 0.7798 0.7286 0.6070 0.9021 0.8914 1.8934 0.5151 0.4426 0.4305 2.8331 1.2257 1.3212 1.4872 1.4532 Pyroglutamic acid 0.9650 0.8303 0.6899 1.2326 0.9678 1.0716 1.0810 1.1618 1.0986 1.2614 0.8036 1.2279 0.7441 0.7311 1.1333 0.7538 0.7758 1.0295 1.4409 0.8713 1.0183 1.1104 0.9698 0.8453 0.7704 1.3386 0.9615 1.4799 1.1790 1.0497 1.1481 2.0994 1.9644 2.5596 0.9946 0.7812 0.6233 0.6377 1.9666 1.3768 1.5607 1.6641 1.1383 Pyruvic acid 0.7905 0.5263 0.5221 1.3723 0.7047 1.4821 1.3945 1.2075 1.0750 1.3076 0.8502 1.1439 0.7572 0.7713 1.0949 0.3065 0.3092 1.6404 1.7438 1.2162 0.8879 0.8959 0.7440 0.6381 1.1499 1.3665 1.1739 1.2419 0.8762 0.6295 1.0580 1.3863 1.9497 1.7378 0.8760 0.7598 0.5519 0.4352 2.8961 2.1103 1.1694 1.1889 1.0522 sn-Glycero-3-phosphate 0.6539 1.5238 0.0002 0.8209 2.3494 0.0065 1.1983 1.4470 1.8302 0.9715 0.7776 0.7753 0.6502 0.9528 1.0424 1.0006 0.8866 1.2536 0.8592 1.3877 0.6949 0.9174 1.2580 2.0898 0.6684 1.2741 1.5102 1.6450 1.2359 1.7738 0.3995 0.3614 0.6327 0.7445 0.7775 1.0243 0.7170 0.9410 1.1835 0.8476 1.1836 1.2102 2.2148 Stearic acid 2.3885 0.4537 1.2999 0.6162 0.6026 1.0079 0.6903 0.9410 0.4690 0.5395 2.7033 0.4825 1.9515 0.3751 0.4791 0.5869 0.5478 1.4571 1.4082 1.0922 1.1134 0.7944 0.6274 0.8342 0.6162 1.3655 0.9012 1.5136 0.9034 0.6710 0.4968 0.4049 0.5893 0.5355 2.4716 0.3560 0.3421 0.2860 3.1764 1.3827 1.3242 1.3599 1.2915 Succinic acid 1.1232 1.0872 0.7436 0.9885 1.3590 0.9286 0.9112 0.8587 0.9702 1.1342 0.8687 1.0672 0.7169 0.9125 1.3302 0.4927 0.7426 1.1261 1.6387 0.7166 1.0222 1.2612 0.9496 1.0618 0.7046 0.9933 0.8099 1.5044 0.9046 1.3398 1.8820 3.0807 3.0483 3.4936 1.7545 1.2751 1.0559 0.8761 1.9960 1.5605 1.5686 1.5335 0.8833 Taurine 1.1417 0.9334 0.6888 1.1021 1.0852 0.8505 1.1479 1.0503 0.6419 1.0310 1.1281 1.0192 0.7408 1.0168 1.4222 1.7336 1.9433 0.0673 0.2558 0.8732 0.9610 1.1657 1.0569 1.1776 0.6610 0.7062 0.7986 1.4956 0.7330 1.0743 0.6627 0.5391 0.9353 0.9016 0.9652 0.6900 0.7871 0.7556 0.4378 0.4219 1.3158 1.3531 1.2001 Threonic acid 0.9776 0.7949 0.5970 1.1698 0.9981 0.9186 1.3652 1.1788 1.1018 1.2692 0.9296 1.0564 0.8104 0.7811 1.0515 0.7829 0.6668 1.2701 1.2803 0.6630 1.1997 1.1373 1.0655 1.2069 0.9887 1.1948 1.2532 1.4642 1.3491 1.0434 1.0286 1.4175 1.6592 1.8622 1.1279 0.9193 0.7421 0.7562 1.6086 1.1293 1.8675 2.1036 1.6865 Threonine 0.8747 0.7903 0.7089 1.2060 1.0130 1.4714 0.9954 0.9402 0.8238 1.4479 0.7312 1.2461 0.7614 0.7906 1.1990 0.6186 0.6120 1.1673 1.6022 1.0341 0.8227 1.1432 0.7023 0.8454 0.9783 1.3276 1.1354 1.3851 1.1643 1.1178 1.2491 2.3696 2.2608 3.0477 1.1190 0.9419 0.6226 0.5594 2.1399 1.4758 1.3694 1.5051 1.3699 Tyrosine 0.7729 0.6589 0.7264 1.2138 1.0130 1.2523 1.1730 1.1895 0.7368 1.3201 0.8620 1.4186 0.8001 0.7154 1.1469 0.6564 0.6474 1.1403 1.5559 1.0659 0.8241 1.1100 0.8777 0.9138 0.9651 1.3358 1.2972 1.5813 1.1893 1.1734 1.3486 2.5315 2.1848 3.0487 1.0361 0.7419 0.5436 0.6100 2.6909 1.5874 1.3415 1.7014 1.4715 Urea 0.9230 0.7216 0.6644 1.1064 0.7794 1.3979 1.1589 1.2484 1.2190 1.3218 0.7560 1.2496 0.7591 0.7314 0.9632 0.4996 0.5974 1.2621 1.6409 1.0307 0.9039 1.0654 0.7460 0.7867 1.0223 1.4840 1.0040 1.3701 1.1420 1.2022 1.2272 2.4708 2.3256 2.8177 0.9321 0.7687 0.5770 0.5710 1.8213 1.2022 1.3291 1.4340 1.3364 Valine 0.7550 0.7687 0.8270 1.0373 1.0214 1.1041 1.1647 1.3218 1.1014 1.6240 0.8444 0.9324 0.7560 0.8857 0.8561 0.6988 0.6572 1.1834 1.4606 1.3324 0.7148 0.9528 0.8999 0.9996 0.9417 1.3067 1.0681 1.4240 1.1397 1.7511 1.1480 1.4213 2.0877 1.8556 0.9196 0.6600 0.5465 0.5128 1.8618 1.6707 0.9445 1.2632 1.0850 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 Acetohydroxamic acid 1990 C06808 249 Aspartic acid 5960 C00049 232 beta-Alanine 239 C00099 248 Citric acid 311 C00158 273 Creatinine 588 C00791 115 Cysteine 5862 C00097 218 Fructose 439709 C02336 307 Fumaric acid 444972 C00122 245 gamma-Aminobutyric acid 223130 C00334 174 Glucose 5793 C00221 319 Glutamic acid 33032 C00025 246 Glutamine 5961 C00064 156 Glycine 750 C00037 248 Glyoxylic acid 760 C00048 184 Hypotaurine 107812 C00519 188 Isoleucine 6306 C00407 86 Itaconic acid 811 C00490 259 Lactic acid 107689 C00186 219 Leucine 6106 C00123 86 Malic acid 525 C03668 233 Ornithine 6262 C00077 420 Palmitic acid 985 C00249 313 Pyroglutamic acid 7405 C01879 156 Pyruvic acid 1060 C00022 174 sn-Glycero-3-phosphate 439162 C00093 357 Stearic acid 5281 C01530 341 Succinic acid 1110 C00042 247 Taurine 1123 C00245 326 Threonic acid 151152 C01620 292 Threonine 6288 C00188 218 Tyrosine 6057 C00082 218 Urea 1176 C00086 189 Valine 6287 C00183 218 METABOLITES_END #END