#METABOLOMICS WORKBENCH KasparasP_20240820_075329 DATATRACK_ID:5130 STUDY_ID:ST003424 ANALYSIS_ID:AN005623 PROJECT_ID:PR002115 VERSION 1 CREATED_ON August 20, 2024, 7:59 am #PROJECT PR:PROJECT_TITLE TRAM–LAG1–CLN8 family proteins are acyltransferases regulating phospholipid PR:PROJECT_TITLE composition PR:PROJECT_SUMMARY The diversity of cellular phospholipids, crucial for membrane homeostasis and PR:PROJECT_SUMMARY function, arises from enzymatic remodeling of their fatty acyl chains. In this PR:PROJECT_SUMMARY work, we reveal that poorly understood TRAM–LAG1–CLN8 domain-containing PR:PROJECT_SUMMARY (TLCD) proteins are phospholipid remodeling enzymes. We demonstrate that TLCD1 PR:PROJECT_SUMMARY is an evolutionarily conserved lysophosphatidylethanolamine acyltransferase, PR:PROJECT_SUMMARY which regulates cellular phospholipid composition and generates novel fatty acid PR:PROJECT_SUMMARY and thiamine (vitamin B1) esters as its secondary products. Furthermore, we PR:PROJECT_SUMMARY establish that human TLCD protein CLN8, mutations in which cause fatal PR:PROJECT_SUMMARY neurodegenerative Batten disease, is a lysophosphatidylglycerol acyltransferase. PR:PROJECT_SUMMARY We show that CLN8 catalyzes the essential step in the biosynthesis of PR:PROJECT_SUMMARY bis(monoacylglycero)phosphate, a phospholipid critical for lysosome function. PR:PROJECT_SUMMARY Our study unveils a new family of acyltransferases integral to cellular membrane PR:PROJECT_SUMMARY phospholipid homeostasis and human disease. PR:INSTITUTE University of Cambridge PR:LAST_NAME Petkevicius PR:FIRST_NAME Kasparas PR:ADDRESS The Keith Peters Building, Cambridge, Cambridgeshire, CB2 0XY, United Kingdom PR:EMAIL kp416@mrc-mbu.cam.ac.uk PR:PHONE +447500233355 PR:PUBLICATIONS TRAM–LAG1–CLN8 family proteins are acyltransferases regulating phospholipid PR:PUBLICATIONS composition #STUDY ST:STUDY_TITLE Analysis of lipid composition of CLN8 KO cultured cells ST:STUDY_SUMMARY Untargeted lipidomics experiments were conducted to analyze the differences in ST:STUDY_SUMMARY lipid composition between CLN8 KO and isogenic control HeLa and U2OS cell lines. ST:STUDY_SUMMARY See a reference publication (Sheokand et al) for details. ST:INSTITUTE University of Cambridge ST:LAST_NAME Petkevicius ST:FIRST_NAME Kasparas ST:ADDRESS The Keith Peters Building ST:EMAIL kp416@mrc-mbu.cam.ac.uk ST:PHONE 07500233355 ST:PUBLICATIONS TRAM–LAG1–CLN8 family proteins are acyltransferases regulating phospholipid ST:PUBLICATIONS composition #SUBJECT SU:SUBJECT_TYPE Cultured cells SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 #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 - Cell process blank Genotype:Process blank | Sample source:Process blank | Transfection:Process blank | Replicate:Process blank RAW_FILE_NAME(Raw file name)=Process blank.raw SUBJECT_SAMPLE_FACTORS - HeLa Wild-Type poolReplicate 1 Genotype:Wild-type | Sample source:HeLa | Transfection:None | Replicate:1 RAW_FILE_NAME(Raw file name)=HeLa WT r1.raw SUBJECT_SAMPLE_FACTORS - HeLa Wild-Type poolReplicate 2 Genotype:Wild-type | Sample source:HeLa | Transfection:None | Replicate:2 RAW_FILE_NAME(Raw file name)=HeLa WT r2.raw SUBJECT_SAMPLE_FACTORS - HeLa Wild-Type poolReplicate 3 Genotype:Wild-type | Sample source:HeLa | Transfection:None | Replicate:3 RAW_FILE_NAME(Raw file name)=HeLa WT r3.raw SUBJECT_SAMPLE_FACTORS - HeLa CLN8 KO poolReplicate 1 Genotype:CLN8 KO | Sample source:HeLa | Transfection:None | Replicate:1 RAW_FILE_NAME(Raw file name)=HeLa CLN8 KO r1.raw SUBJECT_SAMPLE_FACTORS - HeLa CLN8 KO poolReplicate 2 Genotype:CLN8 KO | Sample source:HeLa | Transfection:None | Replicate:2 RAW_FILE_NAME(Raw file name)=HeLa CLN8 KO r2.raw SUBJECT_SAMPLE_FACTORS - HeLa CLN8 KO poolReplicate 3 Genotype:CLN8 KO | Sample source:HeLa | Transfection:None | Replicate:3 RAW_FILE_NAME(Raw file name)=HeLa CLN8 KO r3.raw SUBJECT_SAMPLE_FACTORS - HeLa CLN8 KO pool + EV Replicate 1 Genotype:CLN8 KO | Sample source:HeLa | Transfection:Empty vector | Replicate:1 RAW_FILE_NAME(Raw file name)=HeLa CLN8 KO EV r1.raw SUBJECT_SAMPLE_FACTORS - HeLa CLN8 KO pool + EV Replicate 2 Genotype:CLN8 KO | Sample source:HeLa | Transfection:Empty vector | Replicate:2 RAW_FILE_NAME(Raw file name)=HeLa CLN8 KO EV r2.raw SUBJECT_SAMPLE_FACTORS - HeLa CLN8 KO pool + EV Replicate 3 Genotype:CLN8 KO | Sample source:HeLa | Transfection:Empty vector | Replicate:3 RAW_FILE_NAME(Raw file name)=HeLa CLN8 KO EV r3.raw SUBJECT_SAMPLE_FACTORS - HeLa CLN8 KO pool + CLN8-HA Replicate 1 Genotype:CLN8 KO | Sample source:HeLa | Transfection:CLN8-HA | Replicate:1 RAW_FILE_NAME(Raw file name)=HeLa CLN8 KO HA CLN8 r1.raw SUBJECT_SAMPLE_FACTORS - HeLa CLN8 KO pool + CLN8-HA Replicate 2 Genotype:CLN8 KO | Sample source:HeLa | Transfection:CLN8-HA | Replicate:2 RAW_FILE_NAME(Raw file name)=HeLa CLN8 KO HA CLN8 r2.raw SUBJECT_SAMPLE_FACTORS - HeLa CLN8 KO pool + CLN8-HA Replicate 3 Genotype:CLN8 KO | Sample source:HeLa | Transfection:CLN8-HA | Replicate:3 RAW_FILE_NAME(Raw file name)=HeLa CLN8 KO HA CLN8 r3.raw SUBJECT_SAMPLE_FACTORS - U-2OS Wild-Type poolReplicate 1 Genotype:Wild-type | Sample source:U2OS | Transfection:None | Replicate:1 RAW_FILE_NAME(Raw file name)=U2OS WT r1.raw SUBJECT_SAMPLE_FACTORS - U-2OS Wild-Type poolReplicate 2 Genotype:Wild-type | Sample source:U2OS | Transfection:None | Replicate:2 RAW_FILE_NAME(Raw file name)=U2OS WT r2.raw SUBJECT_SAMPLE_FACTORS - U-2OS Wild-Type poolReplicate 3 Genotype:Wild-type | Sample source:U2OS | Transfection:None | Replicate:3 RAW_FILE_NAME(Raw file name)=U2OS WT r3.raw SUBJECT_SAMPLE_FACTORS - U-2OS CLN8 KO poolReplicate 1 Genotype:CLN8 KO | Sample source:U2OS | Transfection:None | Replicate:1 RAW_FILE_NAME(Raw file name)=U2OS CLN8 KO r1.raw SUBJECT_SAMPLE_FACTORS - U-2OS CLN8 KO poolReplicate 2 Genotype:CLN8 KO | Sample source:U2OS | Transfection:None | Replicate:2 RAW_FILE_NAME(Raw file name)=U2OS CLN8 KO r2.raw SUBJECT_SAMPLE_FACTORS - U-2OS CLN8 KO poolReplicate 3 Genotype:CLN8 KO | Sample source:U2OS | Transfection:None | Replicate:3 RAW_FILE_NAME(Raw file name)=U2OS CLN8 KO r3.raw SUBJECT_SAMPLE_FACTORS - U-2OS CLN8 KO pool + EV Replicate 1 Genotype:CLN8 KO | Sample source:U2OS | Transfection:Empty vector | Replicate:1 RAW_FILE_NAME(Raw file name)=U2OS CLN8 KO EV r1.raw SUBJECT_SAMPLE_FACTORS - U-2OS CLN8 KO pool + EV Replicate 2 Genotype:CLN8 KO | Sample source:U2OS | Transfection:Empty vector | Replicate:2 RAW_FILE_NAME(Raw file name)=U2OS CLN8 KO EV r2.raw SUBJECT_SAMPLE_FACTORS - U-2OS CLN8 KO pool + EV Replicate 3 Genotype:CLN8 KO | Sample source:U2OS | Transfection:Empty vector | Replicate:3 RAW_FILE_NAME(Raw file name)=U2OS CLN8 KO EV r3.raw SUBJECT_SAMPLE_FACTORS - U-2OS CLN8 KO pool + CLN8-HA Replicate 1 Genotype:CLN8 KO | Sample source:U2OS | Transfection:CLN8-HA | Replicate:1 RAW_FILE_NAME(Raw file name)=U2OS CLN8 KO HA CLN8 r1.raw SUBJECT_SAMPLE_FACTORS - U-2OS CLN8 KO pool + CLN8-HA Replicate 2 Genotype:CLN8 KO | Sample source:U2OS | Transfection:CLN8-HA | Replicate:2 RAW_FILE_NAME(Raw file name)=U2OS CLN8 KO HA CLN8 r2.raw SUBJECT_SAMPLE_FACTORS - U-2OS CLN8 KO pool + CLN8-HA Replicate 3 Genotype:CLN8 KO | Sample source:U2OS | Transfection:CLN8-HA | Replicate:3 RAW_FILE_NAME(Raw file name)=U2OS CLN8 KO HA CLN8 r3.raw #COLLECTION CO:COLLECTION_SUMMARY All cell lines were maintained in Dulbecco’s Modified Eagle Medium (DMEM, CO:COLLECTION_SUMMARY 11965084, Thermo Fisher) supplemented with 10% fetal bovine serum (FBS, Sigma) CO:COLLECTION_SUMMARY at 37°C and 5% CO2. To collect samples for lipid measurements, confluent cells CO:COLLECTION_SUMMARY in 6-well plates were swiftly washed twice with 3 ml of ice-cold PBS on ice, CO:COLLECTION_SUMMARY then scraped into 500 μl of ice-cold PBS on ice using inverted 200 μl pipette CO:COLLECTION_SUMMARY tip. The resulting cell suspension was transferred into pre-chilled 2 ml plastic CO:COLLECTION_SUMMARY vials used for lipid extraction (3469-11, Thermo Scientific), and centrifuged at CO:COLLECTION_SUMMARY 500 g, 4°C for 5 min. The supernatant was aspirated and the cell pellet was CO:COLLECTION_SUMMARY snap-frozen and stored at -70°C. One well of a 6-well plate yielded one CO:COLLECTION_SUMMARY replicate in analysis. CO:SAMPLE_TYPE Cultured cells CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY Cells were either untreated (collected under standard culturing conditions) or TR:TREATMENT_SUMMARY transfected using FuGENE® HD Transfection Reagent (E2311, Promega) according to TR:TREATMENT_SUMMARY the manufacturer’s protocol. For a single well of a 6-well plate, 20 μl of TR:TREATMENT_SUMMARY Opti-MEM (31985062, Thermo Fisher), 0.4 μg plasmid DNA and 1.2 μl of FuGENE HD TR:TREATMENT_SUMMARY was used. Medium was refreshed the next day and cells were analyzed 48-72 hours TR:TREATMENT_SUMMARY post transfection. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY The extraction of total lipids from cellular matrices, immunoprecipitated SP:SAMPLEPREP_SUMMARY mitochondria and in vitro assays was conducted employing the butanol-methanol SP:SAMPLEPREP_SUMMARY (BUME) method, as described in detail (37). Note that the chloroform-methanol SP:SAMPLEPREP_SUMMARY lipid extraction methods result in the partitioning of acyl-thiamines into the SP:SAMPLEPREP_SUMMARY polar phase. 2 ml screw cap plastic tubes were used (3469-11, Thermo SP:SAMPLEPREP_SUMMARY Scientific), and a blank extraction was always performed in parallel to account SP:SAMPLEPREP_SUMMARY for the plastic-related contaminants. The extraction commenced with the SP:SAMPLEPREP_SUMMARY homogenization of frozen cell pellets, mitochondrial beads or in vitro assay SP:SAMPLEPREP_SUMMARY mixtures in a 0.5 ml ice-cold solution of butanol to methanol in a 3:1 ratio. SP:SAMPLEPREP_SUMMARY For lipidomics samples, BUME solution was enriched with SPLASH internal standard SP:SAMPLEPREP_SUMMARY mix (330707, Avanti Polar Lipids). For the extraction, a further 0.5 ml of 1% SP:SAMPLEPREP_SUMMARY acetic acid and 0.5 ml of a heptane:ethyl acetate 3:1 mixture were added, SP:SAMPLEPREP_SUMMARY followed by vigorous vortexing for a total of 5 minutes. The mixture was then SP:SAMPLEPREP_SUMMARY centrifuged at 6000 g for 5 minutes, allowing for the separation of phases, SP:SAMPLEPREP_SUMMARY after which the upper organic phase was carefully decanted into glass vials. A SP:SAMPLEPREP_SUMMARY second extraction was conducted on the remaining aqueous phase, with the newly SP:SAMPLEPREP_SUMMARY acquired upper phase being combined in the same glass vials as the first. Post SP:SAMPLEPREP_SUMMARY extraction, the solvents were evaporated under a stream of nitrogen, and the SP:SAMPLEPREP_SUMMARY resultant dry lipid extracts were preserved at −70°C pending further SP:SAMPLEPREP_SUMMARY analysis. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Dried samples were reconstituted in 100 μL of isopropanol, acetonitrile, and CH:CHROMATOGRAPHY_SUMMARY water (2:1:1 ratio) and thoroughly vortexed. Liquid chromatography was conducted CH:CHROMATOGRAPHY_SUMMARY using a Shimadzu HPLC System, with 10 μL of the sample introduced onto a Waters CH:CHROMATOGRAPHY_SUMMARY Acquity Premier UPLC® CSH column (1.7 μm pore size, 2.1 mm × 50 mm), which CH:CHROMATOGRAPHY_SUMMARY was maintained at 55 °C. The mobile phase A comprised a 6:4 ratio of CH:CHROMATOGRAPHY_SUMMARY acetonitrile to water with 10 mM ammonium formate, and mobile phase B consisted CH:CHROMATOGRAPHY_SUMMARY of a 9:1 ratio of isopropanol to acetonitrile with 10 mM ammonium formate. A CH:CHROMATOGRAPHY_SUMMARY flow rate of 500 μL per minute was maintained with a gradient protocol for CH:CHROMATOGRAPHY_SUMMARY mobile phase B as follows: 0.00 minutes_40% mobile phase B; 1.5 minutes_40% CH:CHROMATOGRAPHY_SUMMARY mobile phase B; 8.00 minutes_99% mobile phase B; 10.00 minutes_99% mobile phase CH:CHROMATOGRAPHY_SUMMARY B; 10.10 minutes_40% mobile phase B; 12.00 minutes_40% mobile phase. The sample CH:CHROMATOGRAPHY_SUMMARY injection needle was rinsed with a 9:1 isopropanol and acetonitrile solution CH:CHROMATOGRAPHY_SUMMARY (strong wash) and isopropanol, acetonitrile, and water (2:1:1, weak wash). CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Shimadzu 10A CH:COLUMN_NAME Waters ACQUITY UPLC CSH C18 (100 x 2.1mm,1.7um) CH:SOLVENT_A 6:4 of acetonitrile to water with 10 mM ammonium formate CH:SOLVENT_B 9:1 of isopropanol to acetonitrile with 10 mM ammonium formate CH:FLOW_GRADIENT Gradient protocol for mobile phase B as follows: 0.00 minutes_40% mobile phase CH:FLOW_GRADIENT B; 1.5 minutes_40% mobile phase B; 8.00 minutes_99% mobile phase B; 10.00 CH:FLOW_GRADIENT minutes_99% mobile phase B; 10.10 minutes_40% mobile phase B; 12.00 minutes_40% CH:FLOW_GRADIENT mobile phase. CH:FLOW_RATE 500 μL per minute CH:COLUMN_TEMPERATURE 55 #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Thermo Q Exactive Orbitrap MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE ESI MS:ION_MODE UNSPECIFIED MS:MS_COMMENTS Mass spectrometric detection was carried out on a ThermoFisher Scientific MS:MS_COMMENTS Q-Exactive Orbitrap equipped with a heated electrospray ionization source. The MS:MS_COMMENTS mass spectrometer was calibrated immediately before sample analysis using MS:MS_COMMENTS positive and negative ionization calibration solution (recommended by Thermo MS:MS_COMMENTS Scientific). The electrospray ionization parameters were optimized with a 50:50 MS:MS_COMMENTS mix of mobile phase A and B for spray stability, setting the capillary MS:MS_COMMENTS temperature at 300 °C, source heater temperature at 420 °C, with the sheath, MS:MS_COMMENTS auxiliary, and spare gas flows at specific arbitrary units (40, 15 and 3, MS:MS_COMMENTS respectively), and source voltage at 4 kV. The mass spectrometer operated at a MS:MS_COMMENTS scan rate of 4 Hz, yielding a resolution of 35,000 at m/z 200, over a full-scan MS:MS_COMMENTS range from m/z 120 to 1800, with continuous switching between positive and MS:MS_COMMENTS negative modes. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS nmol MS_METABOLITE_DATA_START Samples Cell process blank HeLa Wild-Type poolReplicate 1 HeLa Wild-Type poolReplicate 2 HeLa Wild-Type poolReplicate 3 HeLa CLN8 KO poolReplicate 1 HeLa CLN8 KO poolReplicate 2 HeLa CLN8 KO poolReplicate 3 HeLa CLN8 KO pool + EV Replicate 1 HeLa CLN8 KO pool + EV Replicate 2 HeLa CLN8 KO pool + EV Replicate 3 HeLa CLN8 KO pool + CLN8-HA Replicate 1 HeLa CLN8 KO pool + CLN8-HA Replicate 2 HeLa CLN8 KO pool + CLN8-HA Replicate 3 U-2OS Wild-Type poolReplicate 1 U-2OS Wild-Type poolReplicate 2 U-2OS Wild-Type poolReplicate 3 U-2OS CLN8 KO poolReplicate 1 U-2OS CLN8 KO poolReplicate 2 U-2OS CLN8 KO poolReplicate 3 U-2OS CLN8 KO pool + EV Replicate 1 U-2OS CLN8 KO pool + EV Replicate 2 U-2OS CLN8 KO pool + EV Replicate 3 U-2OS CLN8 KO pool + CLN8-HA Replicate 1 U-2OS CLN8 KO pool + CLN8-HA Replicate 2 U-2OS CLN8 KO pool + CLN8-HA Replicate 3 Factors Genotype:Process blank | Sample source:Process blank | Transfection:Process blank | Replicate:Process blank Genotype:Wild-type | Sample source:HeLa | Transfection:None | Replicate:1 Genotype:Wild-type | Sample source:HeLa | Transfection:None | Replicate:2 Genotype:Wild-type | Sample source:HeLa | Transfection:None | Replicate:3 Genotype:CLN8 KO | Sample source:HeLa | Transfection:None | Replicate:1 Genotype:CLN8 KO | Sample source:HeLa | Transfection:None | Replicate:2 Genotype:CLN8 KO | Sample source:HeLa | Transfection:None | Replicate:3 Genotype:CLN8 KO | Sample source:HeLa | Transfection:Empty vector | Replicate:1 Genotype:CLN8 KO | Sample source:HeLa | Transfection:Empty vector | Replicate:2 Genotype:CLN8 KO | Sample source:HeLa | Transfection:Empty vector | Replicate:3 Genotype:CLN8 KO | Sample source:HeLa | Transfection:CLN8-HA | Replicate:1 Genotype:CLN8 KO | Sample source:HeLa | Transfection:CLN8-HA | Replicate:2 Genotype:CLN8 KO | Sample source:HeLa | Transfection:CLN8-HA | Replicate:3 Genotype:Wild-type | Sample source:U2OS | Transfection:None | Replicate:1 Genotype:Wild-type | Sample source:U2OS | Transfection:None | Replicate:2 Genotype:Wild-type | Sample source:U2OS | Transfection:None | Replicate:3 Genotype:CLN8 KO | Sample source:U2OS | Transfection:None | Replicate:1 Genotype:CLN8 KO | Sample source:U2OS | Transfection:None | Replicate:2 Genotype:CLN8 KO | Sample source:U2OS | Transfection:None | Replicate:3 Genotype:CLN8 KO | Sample source:U2OS | Transfection:Empty vector | Replicate:1 Genotype:CLN8 KO | Sample source:U2OS | Transfection:Empty vector | Replicate:2 Genotype:CLN8 KO | Sample source:U2OS | Transfection:Empty vector | Replicate:3 Genotype:CLN8 KO | Sample source:U2OS | Transfection:CLN8-HA | Replicate:1 Genotype:CLN8 KO | Sample source:U2OS | Transfection:CLN8-HA | Replicate:2 Genotype:CLN8 KO | Sample source:U2OS | Transfection:CLN8-HA | Replicate:3 BMP 34:2 0 0.001979175 0.001846901 0.001887963 0 0 0 0 0 0 0.00434931 0.003435167 0.003768793 3.00815E-05 5.09167E-07 2.92472E-07 0 0 0 0 0 0 3.60341E-06 1.73813E-05 6.31094E-05 BMP 36:2 0 0.076903964 0.072210493 0.066649553 0.000697887 0.0005463 0.000630411 0.000938488 0.000257884 0.00073907 0.136747849 0.143200221 0.140275136 0.005388911 0.006556766 0.005813989 0.000134615 0 0 0.00034222 0.000290747 0.000199798 0.005356538 0.005778491 0.005735632 BMP 36:3 0 0.003142498 0.003059713 0.002909188 0 0 0 0 0 0 0.004585128 0.002543133 0.002401598 9.18852E-05 5.39894E-05 6.21194E-05 0 0 0 0 0 0 1.86341E-05 1.08443E-05 1.88328E-06 BMP 38:3 0 0.014196666 0.014533889 0.014373245 0 0 3.41608E-05 4.89846E-05 3.75149E-05 0 0.017698866 0.014138371 0.015034998 0.000415345 0.001092538 0.000363681 2.67507E-05 0 0 0 0 1.14496E-05 0.000165362 0.000328843 0.000354681 BMP 38:4 0 0.00747242 0.006107573 0.007221468 0 0 0 4.50726E-07 6.48863E-05 3.61808E-07 0.004291127 0.007310284 0.009471669 0.000981946 0.001339008 0.001273941 0 2.62123E-05 0 0 0 8.75828E-05 0.000790545 0.000746517 0.001063262 BMP 38:6 0 0.000725614 0.00076165 0.000744648 0 0 0 0 0 0 0.002342641 0.001523471 0.001640936 0.00039397 0.000626406 0.000444427 0 0 0 5.28867E-06 0 0 0.000162224 0.000141892 0.000174349 BMP 40:5 0 0.00211292 0.002442741 0.001864268 0 0 3.89453E-05 0 0 0 0.007237991 0.007203466 0.007405031 0.000788115 0.000842977 0.000855789 0 0 0 0 0 5.0914E-05 0.001061819 0.001199745 0.00120464 BMP 40:6 0 0.00086181 0.000913309 0.000874578 0 0 0 0 0 0 0.004279608 0.003383182 0.003785405 0.000710109 0.001053717 0.000826141 0 0 0 0 0 0 0.000387615 0.00050464 0.00057416 BMP 40:7 0 0.008974582 0.008991298 0.008761278 0 0 0 0 0 0 0.018278499 0.013061676 0.015029568 0.004920685 0.0066904 0.005488749 0 0 0 0 0 0 0.00068596 0.000589376 0.000477621 BMP 40:8 0 0.001096164 0.001084809 0.001078388 0 0 0 0 0 0 0.001471846 0.001180117 0.001393412 0.001021513 0.001583994 0.001103219 0 0 0 0 0 0 1.88079E-05 1.47546E-05 8.17948E-05 BMP 42:10 0 0.000203079 0.00022967 0.00020872 0 0 0 0 0 0 0.001841872 0.001456995 0.002020884 0.000939691 0.001813178 0.000880427 0 0 0 0 0 0 0.000172594 2.99677E-07 3.24247E-05 BMP 42:6 0 0.001243509 0.001231659 0.001428837 0 0 0 0 0 0 0.002689842 0.002491494 0.002903071 0.000239932 0.000380248 0.000232164 0 0 0 0 0 0 3.19765E-05 2.67146E-05 8.64023E-05 BMP 42:8 0 0.001048903 0.001253852 0.000972374 0 0 0 4.55896E-06 0 1.22362E-05 0.000958261 0.000706246 0.000801645 0.000532318 0.000730435 0.000632833 0 0 0 0 0 0 3.74044E-05 3.28399E-05 3.98416E-05 BMP 44:10 0 0.000130786 0.000168369 0.00016143 0 0 0 0 0 0 0.000580588 0.000420789 0.000521244 0.000454415 0.000631696 0.000533766 0 0 0 0 0 0 3.13329E-05 4.94287E-06 2.28236E-05 BMP 44:12 0 0.005290346 0.005503094 0.00509946 3.32599E-06 0 0 0 0 0 0.00516533 0.004655337 0.005477303 0.018196639 0.023036182 0.019745675 6.80461E-05 3.57394E-05 6.73413E-05 4.97223E-05 2.6829E-05 0.000126956 0.000588641 0.00051931 0.000731286 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name m/z [M-H]- Average Rt(min) BMP 34:2 745.50348 2.327 BMP 36:2 773.53351 3.313 BMP 36:3 771.51788 2.713 BMP 38:3 799.54944 3.381 BMP 38:4 797.53259 2.953 BMP 38:6 793.50244 2.089 BMP 40:5 823.54883 3.323 BMP 40:6 821.53314 2.524 BMP 40:7 819.51941 2.227 BMP 40:8 817.50244 1.661 BMP 42:10 841.50067 1.535 BMP 42:6 849.5639 3.345 BMP 42:8 845.53326 2.446 BMP 44:10 869.53406 2.105 BMP 44:12 865.5025 1.441 METABOLITES_END #END