#METABOLOMICS WORKBENCH boudreaulab_20240604_145401 DATATRACK_ID:4899 STUDY_ID:ST003247 ANALYSIS_ID:AN005318 PROJECT_ID:PR002016 VERSION 1 CREATED_ON June 6, 2024, 8:24 am #PROJECT PR:PROJECT_TITLE Effects of mitoregulin loss on cardiac and mitochondrial lipids in mice PR:PROJECT_SUMMARY We and others discovered a highly-conserved mitochondrial transmembrane PR:PROJECT_SUMMARY microprotein, named Mitoregulin (Mtln), that supports lipid metabolism. We PR:PROJECT_SUMMARY reported that Mtln strongly binds cardiolipin (CL), increases mitochondrial PR:PROJECT_SUMMARY respiration and Ca2+ retention capacities, and reduces reactive oxygen species PR:PROJECT_SUMMARY (ROS). Here we extend our observation of Mtln-CL binding and examine Mtln PR:PROJECT_SUMMARY influence on cristae structure and mitochondrial membrane integrity during PR:PROJECT_SUMMARY stress. We demonstrate that mitochondria from constitutive- and inducible PR:PROJECT_SUMMARY Mtln-knockout (KO) mice are susceptible to membrane freeze-damage and that this PR:PROJECT_SUMMARY can be rescued by acute Mtln re-expression. In mitochondrial-simulated lipid PR:PROJECT_SUMMARY monolayers, we show that synthetic Mtln decreases lipid packing and monolayer PR:PROJECT_SUMMARY elasticity. Lipidomics revealed that Mtln-KO heart tissues show broad decreases PR:PROJECT_SUMMARY in 22:6-containing lipids and increased cardiolipin damage/remodeling. Lastly, PR:PROJECT_SUMMARY we demonstrate that Mtln-KO mice suffer worse myocardial ischemia-reperfusion PR:PROJECT_SUMMARY injury, hinting at a translationally-relevant role for Mtln in cardioprotection. PR:PROJECT_SUMMARY Our work supports a model in which Mtln binds cardiolipin and stabilizes PR:PROJECT_SUMMARY mitochondrial membranes to broadly influence diverse mitochondrial functions, PR:PROJECT_SUMMARY including lipid metabolism, while also protecting against stress. PR:INSTITUTE University of Iowa PR:LAST_NAME Boudreau PR:FIRST_NAME Ryan PR:ADDRESS 4334 PBDB, 169 Newton Rd, Iowa City, IA 52242 PR:EMAIL ryan-boudreau@uiowa.edu PR:PHONE 3193535573 #STUDY ST:STUDY_TITLE Effects of mitoregulin loss in aged female mice ST:STUDY_SUMMARY Cardiac lipidome analysis in aged (21 to 23-months old) female wildtype and mtln ST:STUDY_SUMMARY knockout mice ST:INSTITUTE University of Iowa ST:LAST_NAME Boudreau ST:FIRST_NAME Ryan ST:ADDRESS 4334 PBDB, 169 Newton Rd, Iowa City, IA 52242 ST:EMAIL ryan-boudreau@uiowa.edu ST:PHONE 3193535573 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENDER Female #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 - 1066-23 Genotype:WT | Sample source:heart tissue RAW_FILE_NAME(Raw file name POS mode)=6530_20190815_25_+_1066-23_WT.mzML; RAW_FILE_NAME(Raw file name NEG mode)=6530_20190815_118_-_1066-23_WT.mzML SUBJECT_SAMPLE_FACTORS - 1066-4 Genotype:WT | Sample source:heart tissue RAW_FILE_NAME(Raw file name POS mode)=6530_20190815_11_+_1066-4_WT.mzML; RAW_FILE_NAME(Raw file name NEG mode)=6530_20190815_125_-_1066-4_WT.mzML SUBJECT_SAMPLE_FACTORS - 1074-34 Genotype:WT | Sample source:heart tissue RAW_FILE_NAME(Raw file name POS mode)=6530_20190815_19_+_1074-34_WT.mzML; RAW_FILE_NAME(Raw file name NEG mode)=6530_20190815_115_-_1074-34_WT.mzML SUBJECT_SAMPLE_FACTORS - 1103-12 Genotype:WT | Sample source:heart tissue RAW_FILE_NAME(Raw file name POS mode)=6530_20190815_24_+_1103-12_WT.mzML; RAW_FILE_NAME(Raw file name NEG mode)=6530_20190815_112_-_1103-12_WT.mzML SUBJECT_SAMPLE_FACTORS - 1133-1 Genotype:KO | Sample source:heart tissue RAW_FILE_NAME(Raw file name POS mode)=6530_20190815_14_+_1133-1_KO.mzML; RAW_FILE_NAME(Raw file name NEG mode)=6530_20190815_114_-_1133-1_KO.mzML SUBJECT_SAMPLE_FACTORS - 1143-2 Genotype:KO | Sample source:heart tissue RAW_FILE_NAME(Raw file name POS mode)=6530_20190815_15_+_1143-2_KO.mzML; RAW_FILE_NAME(Raw file name NEG mode)=6530_20190815_107_-_1143-2_KO.mzML SUBJECT_SAMPLE_FACTORS - 1143-3 Genotype:KO | Sample source:heart tissue RAW_FILE_NAME(Raw file name POS mode)=6530_20190815_13_+_1143-3_KO.mzML; RAW_FILE_NAME(Raw file name NEG mode)=6530_20190815_123_-_1143-3_KO.mzML SUBJECT_SAMPLE_FACTORS - 1145-2 Genotype:KO | Sample source:heart tissue RAW_FILE_NAME(Raw file name POS mode)=6530_20190815_26_+_1145-2_KO.mzML; RAW_FILE_NAME(Raw file name NEG mode)=6530_20190815_124_-_1145-2_KO.mzML SUBJECT_SAMPLE_FACTORS - 1152-1 Genotype:KO | Sample source:heart tissue RAW_FILE_NAME(Raw file name POS mode)=6530_20190815_07_+_1152-1_KO.mzML; RAW_FILE_NAME(Raw file name NEG mode)=6530_20190815_108_-_1152-1_KO.mzML SUBJECT_SAMPLE_FACTORS - WTF Genotype:WT | Sample source:heart tissue RAW_FILE_NAME(Raw file name POS mode)=6530_20190815_18_+_WTF_WT.mzML; RAW_FILE_NAME(Raw file name NEG mode)=6530_20190815_106_-_WTF_WT.mzML #COLLECTION CO:COLLECTION_SUMMARY Mtln-KO and WT mice were fasted for 4-6 h, and while under deep CO:COLLECTION_SUMMARY ketamine/xylazine anesthesia, the heart apex was snipped off and immediately CO:COLLECTION_SUMMARY snap-frozen by LN2 immersion. CO:SAMPLE_TYPE Heart #TREATMENT TR:TREATMENT_SUMMARY No treatment. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Lipid extraction. Lipid extraction, based on Matyash et al.72, was performed as SP:SAMPLEPREP_SUMMARY follows. All solutions were pre-chilled on ice. Tissues or mitochondrial pellets SP:SAMPLEPREP_SUMMARY were transferred to labeled bead-mill tubes (1.4 mm, MoBio Cat# 13113-50) where SP:SAMPLEPREP_SUMMARY lipids were extracted in a solution of 250 µL PBS, 225 µL MeOH containing SP:SAMPLEPREP_SUMMARY internal standards, and 750 µL MTBE (methyl tert-butyl ether). Internal SP:SAMPLEPREP_SUMMARY standards were Avanti SPLASH LipidoMix (Lot#12) at 10 µL per sample and SP:SAMPLEPREP_SUMMARY Cambridge Isotope laboratories NSK-B and NSK-B-G1 (deuterated carnitines) at 10 SP:SAMPLEPREP_SUMMARY µL per sample. The samples were homogenized in one 30 s cycle using the Omni SP:SAMPLEPREP_SUMMARY Bead Ruptor followed by a rest on ice for 1 h. An addition of 188 µL PBS was SP:SAMPLEPREP_SUMMARY made to induce phase separation. After centrifugation at 16,000 g for 5 minutes SP:SAMPLEPREP_SUMMARY at 4 °C, the upper phases were collected and evaporated to dryness under a SP:SAMPLEPREP_SUMMARY gentle nitrogen stream at room temperature. Lipid samples were reconstituted in SP:SAMPLEPREP_SUMMARY 500 µL IPA (isopropyl alcohol) and transferred to an LC-MS vial with insert SP:SAMPLEPREP_SUMMARY (Agilent 5182-0554 and 5183-2086) for analysis. Concurrently, a process blank SP:SAMPLEPREP_SUMMARY sample and pooled quality control (QC) sample was prepared by taking equal SP:SAMPLEPREP_SUMMARY volumes (~50 µL) from each sample after final resuspension. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Positive Mode RP LCMS CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Agilent 1290 Infinity CH:COLUMN_NAME Waters ACQUITY UPLC CSH C18 (100 x 2.1mm,1.7um) CH:SOLVENT_A 40% water/60% acetonitrile; 0.1% formic acid; 10 mM ammonium formate CH:SOLVENT_B 90% isopropanol/9% acetonitrile/1% water; 0.1% formic acid; 10 mM ammonium CH:SOLVENT_B formate CH:FLOW_GRADIENT Started at 15% mobile phase B then increased to 30% B over 2.4 min. It CH:FLOW_GRADIENT sequentially increased to 48% B from 2.4 – 3.0 min, 82% B from 3 – 13.2 min, CH:FLOW_GRADIENT and 99% B from 13.2 – 13.8 min where it’s held until 16.7 min and returned CH:FLOW_GRADIENT to the initial conditions and equilibrated for 5 min. CH:FLOW_RATE 0.4 mL min CH:COLUMN_TEMPERATURE 65 #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Agilent 6530 QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS For positive mode, the source gas temperature was set to 225 °C, with a drying MS:MS_COMMENTS gas flow of 11 L/minute, nebulizer pressure of 40 psig, sheath gas temp of 350 MS:MS_COMMENTS °C and sheath gas flow of 11 L/minute. VCap voltage is set at 3500 V, nozzle MS:MS_COMMENTS voltage 500V, fragmentor at 110 V, skimmer at 85 V and octopole RF peak at 750 MS:MS_COMMENTS V. For data processing, Agilent MassHunter (MH) Workstation and software MS:MS_COMMENTS packages MH Qualitiative and MH Quantitative were used. The pooled QC (n=8) and MS:MS_COMMENTS process blank (n=4) were injected throughout the sample queue to ensure the MS:MS_COMMENTS reliability of acquired lipidomics data. For lipid annotation, accurate mass and MS:MS_COMMENTS MS/MS matching was used with the Agilent Lipid Annotator library. Results from MS:MS_COMMENTS the positive and negative ionization modes from Lipid Annotator were merged MS:MS_COMMENTS based on the class of lipid identified. Data exported from MH Quantitative was MS:MS_COMMENTS evaluated using Excel where initial lipid targets are parsed based on the MS:MS_COMMENTS following criteria. Only lipids with relative standard deviations (RSD) less MS:MS_COMMENTS than 30% in QC samples are used for data analysis. Additionally, only lipids MS:MS_COMMENTS with background AUC counts in process blanks that are less than 30% of QC are MS:MS_COMMENTS used for data analysis. The parsed excel data tables are normalized based on the MS:MS_COMMENTS ratio to class-specific internal standards, then to sum prior to statistical MS:MS_COMMENTS analysis. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS pmol lipid per mg tissue MS_METABOLITE_DATA_START Samples WTF 1066-23 1066-4 1074-34 1103-12 1133-1 1143-2 1143-3 1145-2 1152-1 Factors Genotype:WT | Sample source:heart tissue Genotype:WT | Sample source:heart tissue Genotype:WT | Sample source:heart tissue Genotype:WT | Sample source:heart tissue Genotype:WT | Sample source:heart tissue Genotype:KO | Sample source:heart tissue Genotype:KO | Sample source:heart tissue Genotype:KO | Sample source:heart tissue Genotype:KO | Sample source:heart tissue Genotype:KO | Sample source:heart tissue CAR 14:0 9.89 6.32 6.06 16.09 36.79 20.09 7.73 31.21 7.73 26.15 CAR 14:1 4.71 3.54 3.51 8.77 26.94 6.97 3.58 17.64 5.22 13.61 CAR 15:0 0.35 0.39 0.28 0.61 1.73 1.19 0.49 1.50 0.39 1.00 CAR 16:0 48.33 27.52 31.54 67.85 240.33 141.39 54.24 207.14 37.65 104.82 CAR 16:1 10.70 8.27 8.40 21.46 87.22 31.22 10.89 59.40 14.58 25.91 CAR 16:2 3.38 4.39 3.66 7.44 25.46 9.26 3.49 20.01 3.80 14.39 CAR 18:0 17.46 8.93 10.47 18.98 64.01 29.51 12.67 41.35 7.15 35.72 CAR 18:1 37.66 25.51 32.00 64.25 424.32 121.28 48.93 254.76 58.18 94.17 CAR 18:2 15.70 16.16 20.23 30.57 208.09 69.53 23.14 170.05 19.82 75.68 CAR 20:1 1.59 0.51 0.40 0.93 11.05 5.08 0.73 6.76 0.63 2.46 CAR 20:2 0.46 0.72 0.43 0.94 3.42 3.54 0.42 3.75 0.42 1.30 CAR 20:3 0.33 0.28 0.46 0.56 1.99 0.79 0.32 1.37 0.20 1.20 CAR 20:4 2.79 2.07 1.99 2.90 5.10 1.98 2.06 3.55 1.60 4.61 CL 18:1_18:2_18:2_18:2 16.81 14.58 20.49 21.57 14.23 14.19 20.68 14.80 25.59 19.77 CL 70:7 1.03 1.27 1.19 1.17 0.50 0.99 1.49 0.95 2.14 1.63 CL 72:6 3.65 3.25 5.43 7.41 6.95 3.30 3.75 3.57 6.22 4.31 CL 72:8 34.22 33.11 30.16 26.72 11.56 25.06 51.01 28.94 39.89 44.36 CL 72:9 0.39 0.49 0.50 0.41 0.16 0.37 0.49 0.33 0.81 0.62 CL 74:10 0.89 1.14 1.41 1.33 0.74 0.78 1.18 1.00 1.93 1.22 CL 74:11 0.34 0.57 0.70 0.63 0.32 0.37 0.32 0.38 1.28 0.58 CL 74:9 2.62 3.79 3.85 3.02 1.82 2.34 4.50 3.62 5.13 4.03 CL 76:10 0.53 0.62 1.54 1.62 1.61 0.55 0.43 0.69 1.85 0.67 CL 76:11 5.84 7.67 14.44 13.96 10.98 5.93 5.37 6.98 17.93 8.40 CL 76:12 17.76 22.81 27.19 23.25 12.87 14.36 18.07 17.27 36.39 26.65 CL 76:13 0.14 0.20 0.26 0.22 0.11 0.14 0.13 0.14 0.42 0.21 CL 78:12 0.68 0.94 1.44 1.17 0.94 0.53 0.45 0.68 1.69 1.09 CL 78:13 0.67 1.29 1.91 1.34 0.97 0.68 0.75 1.19 2.66 1.21 CL 78:14 0.17 0.30 0.47 0.42 0.24 0.18 0.16 0.25 0.68 0.28 CL 78:15 0.03 0.07 0.11 0.11 0.08 0.04 0.02 0.04 0.22 0.07 CL 80:15 0.36 0.61 1.65 1.43 1.22 0.41 0.23 0.60 1.91 0.64 CL 80:16 3.15 5.41 8.88 7.30 5.48 2.78 2.24 4.33 10.85 5.58 CL 82:17 0.06 0.13 0.28 0.18 0.15 0.05 0.04 0.14 0.36 0.11 DG 18:0_22:6 12.58 12.90 11.15 13.21 12.33 10.42 8.79 10.80 8.06 11.98 DG 18:1_18:2 41.31 86.36 54.36 73.71 46.17 37.95 60.89 63.61 114.32 40.99 OxPC O-18:1/20:3+1O 31.82 30.19 28.09 28.54 36.81 25.14 38.72 42.12 32.11 31.01 PC O-16:0/20:4 13.68 13.68 13.04 14.80 17.79 11.40 11.31 13.23 12.77 13.31 PC O-16:0/22:5 15.14 19.15 21.24 22.74 23.99 14.18 13.65 18.85 20.54 18.88 PC O-16:0/22:6 135.56 149.13 152.38 163.85 174.33 111.63 102.02 121.19 138.05 145.36 PC O-16:1/18:2 12.55 13.68 12.06 12.73 10.90 13.31 17.11 13.59 13.37 12.27 PC O-16:1/20:4 40.94 57.67 43.69 49.37 48.60 44.80 50.58 51.96 47.08 40.66 PC O-16:1/22:6 258.24 326.27 234.24 297.00 296.57 239.53 213.32 236.60 229.97 289.15 PC O-18:0/22:6 14.60 9.41 13.19 12.02 18.30 8.99 7.33 9.78 11.80 16.83 PC O-18:1/22:6 21.81 25.28 28.73 28.47 38.61 18.59 16.05 22.05 26.27 25.08 PC O-18:3/22:6 4.23 6.31 6.47 5.46 5.70 4.57 4.35 4.89 5.14 5.59 PC O-22:6/22:6 1.19 1.48 1.56 1.14 1.18 1.14 0.95 1.04 1.67 1.48 PC O-32:0 8.64 4.73 5.68 5.09 10.34 5.08 4.94 6.11 5.64 7.51 PC O-32:1 6.66 7.35 6.54 6.50 5.47 6.32 6.50 7.25 7.52 7.28 PC O-34:0 2.42 0.57 1.21 0.79 3.34 0.84 0.86 1.16 1.09 2.02 PC O-34:1 10.58 8.15 10.15 9.62 14.89 8.81 8.55 10.05 10.97 9.70 PC O-34:4 0.54 0.49 0.58 0.60 0.64 0.46 0.50 0.50 0.60 0.53 PC O-36:0 0.56 0.12 0.21 0.16 0.65 0.16 0.19 0.25 0.23 0.38 PC O-36:2 5.59 1.59 2.88 1.86 8.49 2.12 2.21 2.98 2.16 5.07 PC O-38:0 0.06 0.03 0.05 0.04 0.10 0.05 0.05 0.05 0.07 0.08 PC O-38:2 1.04 0.39 0.49 0.41 1.45 0.40 0.39 0.52 0.44 0.77 PC O-39:7 4.30 7.64 6.60 5.69 5.00 5.47 4.80 4.77 5.53 5.99 PC O-40:5 1.73 1.09 1.69 1.52 2.49 1.08 1.06 1.48 1.48 1.87 PC O-40:7 0.57 0.43 0.50 0.45 0.73 0.41 0.41 0.46 0.48 0.56 PC O-40:8 28.70 38.00 39.09 39.32 44.78 28.26 27.02 33.41 33.23 34.24 PC O-40:9 1.80 2.02 1.71 1.93 2.04 1.45 1.24 1.46 1.28 1.92 PC O-42:6 0.93 0.26 0.49 0.44 1.21 0.39 0.47 0.47 0.63 0.77 PC O-42:7 0.42 0.28 0.34 0.36 0.42 0.34 0.38 0.32 0.43 0.40 PC O-44:7 0.76 0.13 0.44 0.26 0.86 0.36 0.28 0.33 0.56 0.66 LPC 16:0 94.22 103.22 117.42 119.40 120.68 97.39 110.43 129.09 84.54 102.10 LPC 17:0 1.05 1.55 1.64 1.48 1.67 1.39 1.28 1.49 1.15 1.52 LPC 18:0 134.80 120.11 127.53 198.95 254.17 137.65 134.01 145.58 126.04 143.20 LPC 18:1 5.68 6.87 9.27 8.02 9.54 6.30 7.55 10.04 6.37 6.72 LPC 18:2 10.89 15.44 16.49 12.84 18.09 14.83 15.32 21.32 11.73 16.71 LPC 19:0 0.83 1.30 1.41 0.84 1.36 1.11 1.03 1.15 0.81 1.18 LPC 20:4 2.77 3.57 3.69 5.08 6.31 3.33 4.20 5.24 3.20 3.31 LPC 22:6 7.77 9.25 7.87 9.38 9.97 6.66 8.10 8.20 6.94 7.32 PC 12:0_21:0 3.68 4.81 5.40 4.61 3.74 4.84 4.37 4.46 4.67 4.96 PC 14:0_16:0 6.15 5.04 4.96 5.01 3.02 5.20 4.91 5.18 5.06 6.38 PC 14:0_21:1 6.56 7.17 10.76 7.85 8.00 9.32 8.67 8.71 9.00 8.07 PC 14:0_22:6 6.03 6.15 6.14 5.68 4.75 6.10 3.04 3.96 5.77 5.41 PC 15:0_18:4 0.66 0.65 0.70 0.70 0.40 0.69 1.25 0.95 0.79 0.72 PC 15:0_22:6 13.57 21.16 20.88 16.97 18.09 18.79 13.91 14.27 17.54 18.59 PC 16:0_16:0 159.35 179.06 171.69 180.70 147.60 164.44 167.93 177.89 175.60 196.90 PC 16:0_16:1 20.73 19.99 22.74 23.57 18.26 20.53 20.91 22.39 31.34 24.84 PC 16:0_18:0 49.23 50.50 53.35 58.17 46.56 48.77 54.60 56.59 61.45 52.57 PC 16:0_18:1 355.70 335.69 432.18 415.70 387.53 395.92 388.74 398.97 438.97 405.45 PC 16:0_18:2 355.73 363.81 400.27 369.47 336.55 342.16 424.01 435.06 420.76 436.19 PC 16:0_20:4 493.81 545.20 449.37 534.33 468.47 421.60 535.47 541.51 464.11 494.57 PC 16:0_20:5 28.06 21.66 28.44 21.36 14.16 20.83 26.38 29.21 32.18 24.27 PC 16:0_22:5 71.25 70.88 77.74 79.02 91.62 63.95 65.86 76.85 82.33 74.02 PC 16:0_22:6 1333.88 1485.19 1317.51 1433.63 1339.71 1211.94 1293.14 1338.26 1231.94 1478.23 PC 16:1_18:2 14.02 17.88 21.05 20.09 16.61 17.96 19.33 20.97 35.10 20.74 PC 16:1_18:3 0.97 1.05 1.04 1.05 0.82 0.94 0.84 0.93 1.25 0.95 PC 16:1_20:4 8.07 7.84 9.59 8.00 5.89 7.19 9.38 8.28 11.93 7.80 PC 16:1_22:6 23.92 29.20 30.33 33.77 33.20 26.61 20.72 25.69 41.42 28.40 PC 17:0_18:2 6.95 8.61 12.01 8.43 8.22 9.10 9.96 10.74 11.36 9.50 PC 17:0_20:4 20.54 28.92 23.81 25.36 22.76 21.12 28.77 28.50 24.96 21.45 PC 17:0_22:5 3.80 5.72 6.21 5.43 4.68 4.80 4.86 5.61 5.71 5.76 PC 17:0_22:6 43.16 61.87 59.34 57.46 57.86 48.52 47.47 51.23 52.91 53.91 PC 17:1_22:6 8.86 11.86 13.66 10.74 12.49 10.63 8.98 10.63 11.40 11.56 PC 18:0_18:1 194.71 155.36 238.85 219.57 207.87 226.60 239.98 230.17 245.23 182.49 PC 18:0_18:2 349.06 322.98 361.05 347.31 351.63 288.30 414.90 423.67 435.02 380.06 PC 18:0_20:1 6.84 2.41 5.46 4.33 6.24 3.66 4.85 5.69 5.81 5.92 PC 18:0_20:2 12.96 11.19 12.87 10.74 14.13 10.50 12.34 13.47 12.68 14.40 PC 18:0_20:3 31.09 30.13 27.90 28.54 36.81 25.63 39.39 42.12 32.12 30.24 PC 18:0_20:4 502.14 588.38 457.82 540.95 486.36 433.06 593.26 600.30 463.17 466.30 PC 18:0_22:4 13.21 11.86 15.35 22.34 25.57 13.14 13.21 15.55 17.19 14.20 PC 18:0_22:5 58.31 54.38 58.38 65.65 77.01 47.40 47.52 55.10 58.22 53.57 PC 18:0_22:6 1275.41 1397.50 1244.10 1358.31 1292.21 1130.25 1263.56 1291.57 1195.89 1341.23 PC 18:1_18:2 144.84 145.74 183.93 160.76 158.07 151.79 201.99 202.95 240.73 170.80 PC 18:1_22:5 13.72 15.96 18.02 17.54 17.47 14.21 14.02 16.56 18.91 16.99 PC 18:1_22:6 187.34 209.65 230.43 224.46 220.31 189.60 223.92 233.75 231.86 209.81 PC 18:2_18:2 45.43 54.52 70.10 51.96 43.08 56.78 75.73 78.13 84.98 66.91 PC 18:2_18:3 3.04 3.96 4.20 3.63 2.38 4.56 3.94 4.09 5.43 4.06 PC 18:2_20:4 67.74 66.34 74.14 68.93 52.33 68.40 95.25 85.20 72.89 57.80 PC 18:2_22:5 15.35 14.22 18.34 16.80 12.36 16.43 15.66 16.70 18.13 16.96 PC 18:2_22:6 404.85 374.40 418.06 401.88 364.83 386.28 431.47 405.99 429.23 386.58 PC 18:3_22:6 7.11 8.28 6.12 6.90 6.81 6.51 6.76 6.96 5.21 7.69 PC 19:0_18:2 3.56 4.09 6.07 3.44 4.42 4.30 5.76 6.06 5.58 4.33 PC 19:0_20:4 4.76 6.97 6.62 4.82 5.85 5.86 7.77 8.40 6.44 4.80 PC 19:0_22:5 1.10 1.29 1.45 1.03 1.27 1.08 1.10 1.22 1.27 1.16 PC 19:0_22:6 29.11 45.74 46.38 33.49 40.80 37.64 34.41 40.08 39.26 36.53 PC 20:0_22:6 11.20 8.48 12.63 10.23 12.97 8.10 8.98 11.56 12.30 11.10 PC 20:1_18:3 5.20 5.47 4.61 6.16 4.99 4.43 6.44 6.77 5.54 4.32 PC 20:1_22:6 5.46 2.58 5.64 3.19 5.73 3.37 3.69 5.63 4.38 5.92 PC 20:2_22:6 4.09 5.66 5.01 4.27 6.33 3.77 3.58 4.53 4.13 5.06 PC 20:3_22:6 6.71 8.17 8.45 8.68 9.09 6.60 6.28 7.71 6.86 5.47 PC 20:4_22:6 7.38 7.58 7.04 9.77 8.99 6.69 5.60 6.33 6.75 4.22 PC 20:5_22:6 0.61 0.49 0.57 0.61 0.30 0.53 0.39 0.44 0.34 0.39 PC 22:2_20:3 0.88 0.78 1.01 1.07 1.28 0.76 0.82 1.01 1.06 1.01 PC 22:5_22:6 2.88 1.91 2.70 2.84 1.55 2.41 1.38 1.67 2.20 1.90 PC 22:6_22:6 7.33 5.12 5.96 7.90 5.88 5.60 3.26 4.02 5.22 3.39 PC 24:0_18:1 0.19 0.19 0.29 0.27 0.26 0.24 0.23 0.26 0.29 0.26 PC 31:0 4.81 5.68 5.95 5.09 3.95 5.89 5.36 5.07 5.03 6.26 PC 32:2 2.82 2.97 3.38 2.96 2.23 3.35 2.75 3.10 3.94 3.34 PC 33:1 5.61 6.12 8.19 6.39 5.89 7.27 6.56 6.73 8.02 7.18 PC 33:2 1.15 1.54 1.91 1.33 1.12 1.62 1.75 1.69 1.61 1.67 PC 35:3 1.24 1.49 1.71 1.36 1.41 1.51 1.76 1.75 1.55 1.64 PC 36:6 0.83 1.07 1.00 1.08 1.01 0.92 0.93 1.02 1.38 1.25 PC 37:1 1.65 1.72 2.81 1.70 2.11 2.37 2.50 2.32 2.56 1.91 PC 37:5 1.61 2.03 2.35 1.98 1.72 1.85 2.11 2.26 2.31 1.97 PC 38:4 18.36 19.94 21.52 21.45 25.67 19.01 21.63 24.40 24.46 21.91 PC 38:5 181.50 215.90 212.37 220.94 197.03 190.55 192.96 214.63 210.25 237.15 PC 38:7 4.52 5.59 5.76 5.24 4.62 5.14 6.17 6.69 6.24 6.30 PC 40:0 0.20 0.24 0.30 0.28 0.25 0.26 0.22 0.27 0.30 0.31 PC 40:1 0.31 0.24 0.39 0.34 0.38 0.29 0.32 0.37 0.37 0.43 PC 40:2 0.30 0.26 0.38 0.31 0.31 0.30 0.29 0.33 0.33 0.37 PC 40:3 0.81 0.61 0.92 0.74 0.99 0.70 0.79 0.87 0.85 0.89 PC 41:7 1.79 2.55 2.84 1.95 2.25 2.23 2.33 2.52 2.56 2.33 PC 42:10 1.00 0.72 0.81 0.68 0.84 0.69 0.62 0.55 0.63 0.56 PC 42:2 0.43 0.46 0.60 0.44 0.42 0.47 0.45 0.52 0.39 0.58 PC 44:6 0.31 0.28 0.36 0.35 0.42 0.29 0.27 0.32 0.33 0.36 PC 46:6 0.12 0.11 0.12 0.14 0.14 0.11 0.10 0.12 0.11 0.13 SM 18:0;OH/16:0 14.14 10.50 9.75 11.33 17.22 11.83 9.13 9.91 11.67 13.74 SM 18:1;O2/18:0 59.00 63.71 54.52 60.41 50.94 66.25 58.84 58.05 62.11 62.98 SM 18:1;O2/21:0 17.52 26.96 31.85 21.69 23.71 30.90 25.28 25.97 38.87 25.40 SM 32:1;O2 0.69 0.83 0.80 0.85 0.61 0.86 0.72 0.79 1.13 0.81 SM 33:1;O2 3.86 5.19 5.75 4.39 5.89 5.13 4.10 4.24 5.98 5.32 SM 34:1;O2 127.54 116.12 111.93 129.41 140.69 121.74 113.89 121.75 127.29 126.60 SM 34:2;O2 3.59 3.94 3.78 3.83 4.47 4.18 3.10 3.42 5.33 3.75 SM 35:1;O2 5.28 6.95 6.60 6.01 5.98 7.00 5.80 5.93 6.63 6.41 SM 36:2;O2 16.50 17.78 14.12 15.38 13.95 16.18 14.57 15.57 16.15 15.77 SM 38:1;O2 192.68 147.64 183.85 160.39 164.27 168.67 154.79 183.61 226.88 199.43 SM 38:2;O2 34.71 29.15 34.11 27.80 32.59 26.68 25.47 35.59 35.58 37.48 SM 39:2;O2 6.56 10.07 10.65 7.63 9.08 10.32 8.35 9.93 10.62 8.13 SM 40:1;O2 156.92 144.77 163.26 167.61 158.77 162.32 133.96 159.77 222.07 190.05 SM 40:2;O2 43.77 43.28 49.63 49.58 50.42 43.23 36.16 47.42 61.03 53.19 SM 41:1;O2 23.58 32.39 30.25 30.20 22.16 32.62 23.40 25.67 40.77 26.90 SM 41:2;O2 9.31 10.30 10.98 10.45 9.80 10.68 8.31 9.05 13.86 9.57 SM 42:1;O2 31.90 34.70 36.11 38.29 28.66 37.51 30.27 33.61 52.12 37.54 SM 42:2;O2 100.14 77.76 105.20 101.78 124.88 101.70 94.16 105.73 148.56 119.84 SM 42:3;O2 32.97 36.76 34.38 35.15 39.43 36.38 30.03 33.64 47.99 37.42 SM 43:1;O2 0.95 1.33 1.35 1.41 1.00 1.30 1.02 1.24 1.64 1.16 SM 43:2;O2 0.68 0.93 0.95 0.88 0.78 1.00 0.80 0.79 1.20 0.82 SM 44:1;O2 0.65 0.80 0.81 0.78 0.77 0.94 0.64 0.83 1.16 0.69 SM 44:2;O2 0.27 0.28 0.32 0.29 0.28 0.31 0.29 0.32 0.44 0.31 TG 12:0_18:2_18:2 0.42 1.66 3.39 1.77 0.85 0.85 1.01 1.41 6.83 2.10 TG 14:0_16:0_18:2 5.61 23.43 37.84 23.66 6.52 14.21 14.78 18.17 56.41 23.19 TG 14:0_18:2_18:2 4.57 28.02 43.79 23.11 10.53 12.54 14.19 22.26 90.40 25.87 TG 15:0_18:2_18:2 0.45 3.35 5.27 1.98 1.29 1.30 1.84 2.70 12.15 2.96 TG 16:0_16:0_16:0 5.91 10.17 15.34 9.94 2.71 8.90 8.31 8.03 18.05 11.05 TG 16:0_16:0_18:0 3.81 6.80 10.98 7.31 2.61 6.23 6.37 6.15 16.75 7.96 TG 16:0_16:0_18:1 40.49 101.09 126.83 88.61 48.24 65.85 86.74 94.09 234.19 81.71 TG 16:0_16:1_18:2 18.18 87.63 119.62 78.60 33.88 44.75 55.08 73.08 229.78 71.17 TG 16:0_17:1_18:2 1.23 9.15 14.40 5.78 4.35 3.83 5.83 8.80 35.57 7.50 TG 16:0_18:0_18:1 17.29 43.41 62.62 46.40 24.49 31.31 42.37 47.61 125.68 43.12 TG 16:0_22:6_22:6 12.59 6.27 15.10 29.41 6.20 7.49 1.88 4.04 6.54 10.30 TG 16:1_18:1_18:2 85.75 354.87 399.16 242.20 175.26 161.23 262.09 309.67 777.46 286.42 TG 17:0_18:1_18:2 2.71 17.18 27.84 12.07 12.19 7.99 14.41 20.51 78.84 15.24 TG 17:1_18:2_18:2 0.75 6.08 8.45 3.30 2.93 2.14 4.01 6.24 26.02 4.86 TG 18:0_18:2_22:6 56.35 95.43 144.88 115.62 101.45 57.15 71.73 96.57 213.16 103.99 TG 18:1_18:1_20:1 20.26 22.63 42.90 28.57 48.16 18.87 32.61 48.64 78.73 41.70 TG 18:1_18:1_22:5 36.06 59.96 91.44 85.00 83.82 41.25 45.78 66.53 177.68 72.12 TG 18:1_18:2_19:1 2.21 10.08 13.16 6.23 9.09 4.73 9.10 9.79 39.43 9.44 TG 18:1_18:2_22:5 42.71 79.68 108.70 96.48 79.24 47.01 47.72 73.44 185.82 92.97 TG 18:1_22:6_22:6 10.70 3.93 10.87 22.16 6.11 5.42 1.35 2.52 5.75 7.11 TG 18:2_22:6_22:6 18.02 7.55 15.57 31.09 6.57 8.38 2.13 3.72 5.46 12.84 TG 22:6_22:6_22:6 1.30 0.18 0.54 1.74 0.17 0.31 0.04 0.07 0.13 0.33 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name Formula m/z Retention time CAR 14:0 C21H41NO4 372.3114 1.766 CAR 14:1 C21H39NO4 370.2955 1.47 CAR 15:0 C22H43NO4 386.3264 2.117 CAR 16:0 C23H45NO4 400.3435 2.551 CAR 16:1 C23H43NO4 398.3274 1.977 CAR 16:2 C23H41NO4 396.3113 1.573 CAR 18:0 C25H49NO4 428.3744 3.596 CAR 18:1 C25H47NO4 426.3597 2.766 CAR 18:2 C25H45NO4 424.344 2.162 CAR 20:1 C27H51NO4 454.3884 3.741 CAR 20:2 C27H49NO4 452.3726 3.02 CAR 20:3 C27H47NO4 450.3585 2.462 CAR 20:4 C27H45NO4 448.3426 2.117 CL 18:1_18:2_18:2_18:2 C81H144O17P2 1449.9803 12.631 CL 70:7 C79H140O17P2 1440.9918 12.119 CL 72:6 C81H146O17P2 1471.0306 12.995 CL 72:8 C81H142O17P2 1467.0105 12.205 CL 72:9 C81H140O17P2 1464.9881 11.833 CL 74:10 C83H142O17P2 1491.007 12.109 CL 74:11 C83H140O17P2 1488.989 11.847 CL 74:9 C83H144O17P2 1493.0245 12.328 CL 76:10 C85H146O17P2 1519.0317 12.75 CL 76:11 C85H144O17P2 1517.0264 12.338 CL 76:12 C85H142O17P2 1515.0114 11.919 CL 76:13 C85H140O17P2 1512.9889 11.574 CL 78:12 C87H146O17P2 1543.04 12.407 CL 78:13 C87H144O17P2 1541.0245 12.044 CL 78:14 C87H142O17P2 1539.0021 11.825 CL 78:15 C87H140O17P2 1536.9887 11.579 CL 80:15 C89H144O17P2 1565.0226 12.045 CL 80:16 C89H142O17P2 1563.0099 11.629 CL 82:17 C91H144O17P2 1589.0189 11.743 DG 18:0_22:6 C43H72O5 686.5727 9.326 DG 18:1_18:2 C39H70O5 636.5582 8.968 OxPC O-18:1/20:3+1O C46H86NO8P 856.6064 8.239 PC O-16:0/20:4 C44H82NO7P 768.5918 7.501 PC O-16:0/22:5 C46H84NO7P 794.6088 7.524 PC O-16:0/22:6 C46H82NO7P 792.5902 7.272 PC O-16:1/18:2 C42H80NO7P 742.5729 7.506 PC O-16:1/20:4 C44H80NO7P 810.5649 7.366 PC O-16:1/22:6 C46H80NO7P 834.5668 7.126 PC O-18:0/22:6 C48H86NO7P 820.6236 8.129 PC O-18:1/22:6 C48H84NO7P 818.6088 7.322 PC O-18:3/22:6 C48H80NO7P 814.5758 6.567 PC O-22:6/22:6 C52H82NO7P 864.5915 6.147 PC O-32:0 C40H82NO7P 720.5921 8.228 PC O-32:1 C40H80NO7P 718.5713 8.104 PC O-34:0 C42H86NO7P 748.6224 9.123 PC O-34:1 C42H84NO7P 746.6075 8.329 PC O-34:4 C42H78NO7P 740.5585 6.632 PC O-36:0 C44H90NO7P 776.653 9.979 PC O-36:2 C44H86NO7P 772.6233 8.515 PC O-38:0 C46H94NO7P 804.6837 10.816 PC O-38:2 C46H90NO7P 800.6573 9.393 PC O-39:7 C47H82NO7P 804.5899 7.501 PC O-40:5 C48H88NO7P 822.6341 8.42 PC O-40:7 C48H84NO7P 818.5991 8.4 PC O-40:8 C48H82NO7P 816.5919 7.209 PC O-40:9 C48H80NO7P 814.5717 7.262 PC O-42:6 C50H90NO7P 848.6514 9.01 PC O-42:7 C50H88NO7P 846.6366 8.864 PC O-44:7 C52H92NO7P 874.6688 8.978 LPC 16:0 C24H50NO7P 496.3409 2.192 LPC 17:0 C25H52NO7P 510.3561 2.929 LPC 18:0 C26H54NO7P 524.3738 3.509 LPC 18:1 C26H52NO7P 522.354 2.623 LPC 18:2 C26H50NO7P 520.3395 2.009 LPC 19:0 C27H56NO7P 538.3869 3.936 LPC 20:4 C28H50NO7P 544.3404 1.937 LPC 22:6 C30H50NO7P 612.3292 1.831 PC 12:0_21:0 C41H82NO8P 748.5879 8.13 PC 14:0_16:0 C38H76NO8P 706.5409 6.83 PC 14:0_21:1 C43H84NO8P 774.6034 8.183 PC 14:0_22:6 C44H76NO8P 778.5405 5.968 PC 15:0_18:4 C41H74NO8P 740.5251 6.812 PC 15:0_22:6 C45H78NO8P 792.5566 6.367 PC 16:0_16:0 C40H80NO8P 734.5732 7.699 PC 16:0_16:1 C40H78NO8P 732.5571 6.928 PC 16:0_18:0 C42H84NO8P 762.6058 8.562 PC 16:0_18:1 C42H82NO8P 760.5894 7.765 PC 16:0_18:2 C42H80NO8P 758.5714 7.131 PC 16:0_20:4 C44H80NO8P 782.5726 6.982 PC 16:0_20:5 C44H78NO8P 780.5562 6.443 PC 16:0_22:5 C46H82NO8P 808.5885 7.331 PC 16:0_22:6 C46H80NO8P 806.5728 6.767 PC 16:1_18:2 C42H78NO8P 756.5561 6.445 PC 16:1_18:3 C42H76NO8P 754.54 6.163 PC 16:1_20:4 C44H78NO8P 780.5553 6.252 PC 16:1_22:6 C46H78NO8P 804.5572 6.105 PC 17:0_18:2 C43H82NO8P 772.5883 7.523 PC 17:0_20:4 C45H82NO8P 796.5888 7.435 PC 17:0_22:5 C47H84NO8P 822.6032 7.469 PC 17:0_22:6 C47H82NO8P 820.5885 7.198 PC 17:1_22:6 C47H80NO8P 818.5727 6.503 PC 18:0_18:1 C44H86NO8P 788.6193 8.652 PC 18:0_18:2 C44H84NO8P 786.6045 8.002 PC 18:0_20:1 C46H90NO8P 860.6422 9.47 PC 18:0_20:2 C46H88NO8P 814.6346 8.775 PC 18:0_20:3 C46H86NO8P 812.6197 8.207 PC 18:0_20:4 C46H84NO8P 810.6046 7.854 PC 18:0_22:4 C48H88NO8P 838.6355 8.471 PC 18:0_22:5 C48H86NO8P 836.6195 7.914 PC 18:0_22:6 C48H84NO8P 834.6044 7.617 PC 18:1_18:2 C44H82NO8P 828.5773 7.223 PC 18:1_22:5 C48H84NO8P 878.5904 7.144 PC 18:1_22:6 C48H82NO8P 832.5888 6.877 PC 18:2_18:2 C44H80NO8P 782.5735 6.571 PC 18:2_18:3 C44H78NO8P 780.5562 6.025 PC 18:2_20:4 C46H80NO8P 806.5725 6.442 PC 18:2_22:5 C48H82NO8P 832.5874 6.485 PC 18:2_22:6 C48H80NO8P 830.5723 6.239 PC 18:3_22:6 C48H78NO8P 828.5558 5.736 PC 19:0_18:2 C45H86NO8P 800.6177 8.433 PC 19:0_20:4 C47H86NO8P 824.6194 8.315 PC 19:0_22:5 C49H88NO8P 850.6337 8.611 PC 19:0_22:6 C49H86NO8P 848.6208 8.062 PC 20:0_22:6 C50H88NO8P 862.6354 8.482 PC 20:1_18:3 C46H84NO8P 810.6025 8.382 PC 20:1_22:6 C50H86NO8P 860.6176 7.68 PC 20:2_22:6 C50H84NO8P 858.6032 6.966 PC 20:3_22:6 C50H82NO8P 856.5867 6.442 PC 20:4_22:6 C50H80NO8P 854.5723 6.082 PC 20:5_22:6 C50H78NO8P 852.5555 5.606 PC 22:2_20:3 C50H90NO8P 864.6462 8.737 PC 22:5_22:6 C52H82NO8P 880.5872 6.145 PC 22:6_22:6 C52H80NO8P 878.572 5.894 PC 24:0_18:1 C50H98NO8P 872.7109 11.09 PC 31:0 C39H78NO8P 720.555 7.252 PC 32:2 C40H76NO8P 730.5386 6.291 PC 33:1 C41H80NO8P 746.5714 7.379 PC 33:2 C41H78NO8P 744.5556 6.67 PC 35:3 C43H80NO8P 770.5706 6.886 PC 36:6 C44H76NO8P 778.5397 6.423 PC 37:1 C45H88NO8P 802.6351 9.075 PC 37:5 C45H80NO8P 794.571 6.662 PC 38:4 C46H84NO8P 810.6006 7.424 PC 38:5 C46H82NO8P 808.5884 7.047 PC 38:7 C46H78NO8P 804.554 6.577 PC 40:0 C48H96NO8P 846.6953 11.056 PC 40:1 C48H94NO8P 844.6794 10.287 PC 40:2 C48H92NO8P 842.6627 9.502 PC 40:3 C48H90NO8P 840.6464 8.856 PC 41:7 C49H84NO8P 846.6011 7.254 PC 42:10 C50H80NO8P 854.5681 5.877 PC 42:2 C50H96NO8P 870.6954 10.533 PC 44:6 C52H92NO8P 890.663 9.341 PC 46:6 C54H96NO8P 918.6921 10.164 SM 18:0;OH/16:0 C39H81N2O6P 705.5913 7.054 SM 18:1;O2/18:0 C41H83N2O6P 731.6089 7.659 SM 18:1;O2/21:0 C44H89N2O6P 773.6562 8.987 SM 32:1;O2 C37H75N2O6P 675.5436 5.882 SM 33:1;O2 C38H77N2O6P 689.5598 6.301 SM 34:1;O2 C39H79N2O6P 703.5781 6.704 SM 34:2;O2 C39H77N2O6P 701.5607 6.036 SM 35:1;O2 C40H81N2O6P 717.5917 7.187 SM 36:2;O2 C41H81N2O6P 729.5929 6.901 SM 38:1;O2 C43H87N2O6P 759.6418 8.537 SM 38:2;O2 C43H85N2O6P 757.6247 7.769 SM 39:2;O2 C44H87N2O6P 771.635 8.224 SM 40:1;O2 C45H91N2O6P 787.6727 9.417 SM 40:2;O2 C45H89N2O6P 785.6565 8.683 SM 41:1;O2 C46H93N2O6P 801.6875 9.842 SM 41:2;O2 C46H91N2O6P 799.671 9.114 SM 42:1;O2 C47H95N2O6P 815.7035 10.268 SM 42:2;O2 C47H93N2O6P 813.6878 9.385 SM 42:3;O2 C47H91N2O6P 811.6693 8.664 SM 43:1;O2 C48H97N2O6P 829.7156 10.647 SM 43:2;O2 C48H95N2O6P 827.6988 9.811 SM 44:1;O2 C49H99N2O6P 843.7291 11.068 SM 44:2;O2 C49H97N2O6P 841.7112 10.233 TG 12:0_18:2_18:2 C51H90O6 816.7089 12.347 TG 14:0_16:0_18:2 C51H94O6 820.7389 13.402 TG 14:0_18:2_18:2 C53H94O6 844.7385 12.955 TG 15:0_18:2_18:2 C54H96O6 858.7521 13.25 TG 16:0_16:0_16:0 C51H98O6 824.775 14.434 TG 16:0_16:0_18:0 C53H102O6 852.806 14.608 TG 16:0_16:0_18:1 C53H100O6 850.7934 14.593 TG 16:0_16:1_18:2 C53H96O6 846.7548 13.437 TG 16:0_17:1_18:2 C54H98O6 860.7689 13.718 TG 16:0_18:0_18:1 C55H104O6 878.8229 14.613 TG 16:0_22:6_22:6 C63H98O6 968.7705 12.755 TG 16:1_18:1_18:2 C55H98O6 872.7712 13.504 TG 17:0_18:1_18:2 C56H102O6 888.8076 14.212 TG 17:1_18:2_18:2 C56H98O6 884.7682 13.285 TG 18:0_18:2_22:6 C61H102O6 948.7989 13.676 TG 18:1_18:1_20:1 C59H108O6 930.855 14.621 TG 18:1_18:1_22:5 C61H104O6 950.8198 13.81 TG 18:1_18:2_19:1 C58H104O6 914.8228 14.24 TG 18:1_18:2_22:5 C61H102O6 948.8017 13.373 TG 18:1_22:6_22:6 C65H100O6 994.7854 12.772 TG 18:2_22:6_22:6 C65H98O6 992.7733 12.273 TG 22:6_22:6_22:6 C69H98O6 1040.7702 11.865 METABOLITES_END #END