#METABOLOMICS WORKBENCH TruxalCarlson_20200909_105704 DATATRACK_ID:2158 STUDY_ID:ST001480 ANALYSIS_ID:AN002456 PROJECT_ID:PR001004 VERSION 1 CREATED_ON September 14, 2020, 4:17 pm #PROJECT PR:PROJECT_TITLE Large diversity in nitrogen- and sulfur-containing compatible solute profiles in PR:PROJECT_TITLE polar and temperate diatoms PR:PROJECT_TYPE Marine Metabolomics PR:PROJECT_SUMMARY Intense bottom-ice algal blooms, often dominated by diatoms, are an important PR:PROJECT_SUMMARY source of food for grazers, organic matter for export during sea ice melt, and PR:PROJECT_SUMMARY dissolved organic carbon. Sea-ice diatoms have a number of adaptations, PR:PROJECT_SUMMARY including accumulation of compatible solutes, that allow them to inhabit this PR:PROJECT_SUMMARY highly variable environment characterized by extremes in temperature, salinity, PR:PROJECT_SUMMARY and light. In addition to protecting them from extreme conditions, these PR:PROJECT_SUMMARY compounds present a labile, nutrient-rich source of organic matter and include PR:PROJECT_SUMMARY precursors to climate active compounds (e.g. DMS), which are likely regulated PR:PROJECT_SUMMARY with environmental change. Here, intracellular concentrations of 45 metabolites PR:PROJECT_SUMMARY were quantified in three sea-ice diatom species and were compared to two PR:PROJECT_SUMMARY temperate diatom species, with a focus on compatible solutes and free amino acid PR:PROJECT_SUMMARY pools. There was a large diversity of metabolite concentrations between diatoms PR:PROJECT_SUMMARY with no clear pattern identifiable for sea-ice species. Concentrations of some PR:PROJECT_SUMMARY compatible solutes (isethionic acid, homarine) approached 1 M in the sea-ice PR:PROJECT_SUMMARY diatoms, Fragilariopsis cylindrus and Navicula cf. perminuta, but not in the PR:PROJECT_SUMMARY larger sea-ice diatom, Nitzschia lecointei or in the temperate diatom species. PR:PROJECT_SUMMARY The differential use of compatible solutes in sea-ice diatoms suggest different PR:PROJECT_SUMMARY adaptive strategies and highlights which small organic compounds may be PR:PROJECT_SUMMARY important in polar biogeochemical cycles. PR:INSTITUTE University of Washington PR:DEPARTMENT Oceanography PR:LABORATORY Ingalls Lab PR:LAST_NAME Dawson PR:FIRST_NAME Hannah PR:ADDRESS 1501 NE Boat Street, Marine Science Building, Room G, Seattle, WA, 98195, USA PR:EMAIL hmdawson@uw.edu PR:PHONE 206-543-0744 PR:PUBLICATIONS Dawson et al, 2020, Integrative and Comparative Biology #STUDY ST:STUDY_TITLE Large diversity in nitrogen- and sulfur-containing compatible solute profiles in ST:STUDY_TITLE polar and temperate diatoms ST:STUDY_TYPE Intracellular metabolites were quantified in diatom species ST:STUDY_SUMMARY Intense bottom-ice algal blooms, often dominated by diatoms, are an important ST:STUDY_SUMMARY source of food for grazers, organic matter for export during sea ice melt, and ST:STUDY_SUMMARY dissolved organic carbon. Sea-ice diatoms have a number of adaptations, ST:STUDY_SUMMARY including accumulation of compatible solutes, that allow them to inhabit this ST:STUDY_SUMMARY highly variable environment characterized by extremes in temperature, salinity, ST:STUDY_SUMMARY and light. In addition to protecting them from extreme conditions, these ST:STUDY_SUMMARY compounds present a labile, nutrient-rich source of organic matter and include ST:STUDY_SUMMARY precursors to climate active compounds (e.g. DMS), which are likely regulated ST:STUDY_SUMMARY with environmental change. Here, intracellular concentrations of 45 metabolites ST:STUDY_SUMMARY were quantified in three sea-ice diatom species and were compared to two ST:STUDY_SUMMARY temperate diatom species, with a focus on compatible solutes and free amino acid ST:STUDY_SUMMARY pools. There was a large diversity of metabolite concentrations between diatoms ST:STUDY_SUMMARY with no clear pattern identifiable for sea-ice species. Concentrations of some ST:STUDY_SUMMARY compatible solutes (isethionic acid, homarine) approached 1 M in the sea-ice ST:STUDY_SUMMARY diatoms, Fragilariopsis cylindrus and Navicula cf. perminuta, but not in the ST:STUDY_SUMMARY larger sea-ice diatom, Nitzschia lecointei or in the temperate diatom species. ST:STUDY_SUMMARY The differential use of compatible solutes in sea-ice diatoms suggest different ST:STUDY_SUMMARY adaptive strategies and highlights which small organic compounds may be ST:STUDY_SUMMARY important in polar biogeochemical cycles. ST:INSTITUTE University of Washington ST:DEPARTMENT Oceanography ST:LABORATORY Ingalls Lab ST:LAST_NAME Dawson ST:FIRST_NAME Hannah ST:ADDRESS 1501 NE Boat Street, Marine Science Building, Room G, Seattle, WA, 98195, USA ST:EMAIL hmdawson@uw.edu ST:PHONE 206-543-0744 ST:PUBLICATIONS Dawson et al, 2020, Integrative and Comparative Biology #SUBJECT SU:SUBJECT_TYPE Other SU:SUBJECT_SPECIES Nitzschia lecointei;Fragilariopsis cylindrus;Navicula cf. perminuta;Navicula pelliculosa #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 - Nl_32ppt-1C_1 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.2756163; Vol_filtered_mL=70; Replicate=1; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_32ppt-1C_A 200309_Smp_32ppt-1C_A SUBJECT_SAMPLE_FACTORS - Nl_32ppt-1C_2 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.174789; Vol_filtered_mL=70; Replicate=2; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_32ppt-1C_B 200309_Smp_32ppt-1C_ SUBJECT_SAMPLE_FACTORS - Nl_32ppt-1C_3 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.1610501; Vol_filtered_mL=70; Replicate=3; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_32ppt-1C_C 200309_Smp_32ppt-1C_C SUBJECT_SAMPLE_FACTORS - Nl_32ppt4C_1 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:4 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.532752987; Vol_filtered_mL=70; Replicate=1; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_32ppt4C_A 200309_Smp_32ppt4C_A SUBJECT_SAMPLE_FACTORS - Nl_32ppt4C_2 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:4 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.75542558; Vol_filtered_mL=70; Replicate=2; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_32ppt4C_B 200309_Smp_32ppt4C_B SUBJECT_SAMPLE_FACTORS - Nl_32ppt4C_3 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:4 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.66524318; Vol_filtered_mL=70; Replicate=3; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_32ppt4C_C 200309_Smp_32ppt4C_C SUBJECT_SAMPLE_FACTORS - Nl_41ppt-1C_1 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.746388756; Vol_filtered_mL=70; Replicate=1; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_41ppt-1C_A 200309_Smp_41ppt-1C_A SUBJECT_SAMPLE_FACTORS - Nl_41ppt-1C_2 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.399316304; Vol_filtered_mL=70; Replicate=2; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_41ppt-1C_B 200309_Smp_41ppt-1C_B SUBJECT_SAMPLE_FACTORS - Nl_41ppt-1C_3 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.657191524; Vol_filtered_mL=70; Replicate=3; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_41ppt-1C_C 200309_Smp_41ppt-1C_C SUBJECT_SAMPLE_FACTORS - Nl_41ppt4C_1 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:4 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.11758985; Vol_filtered_mL=70; Replicate=1; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_41ppt4C_A 200309_Smp_41ppt4C_A SUBJECT_SAMPLE_FACTORS - Nl_41ppt4C_2 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:4 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.310557636; Vol_filtered_mL=70; Replicate=2; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_41ppt4C_B 200309_Smp_41ppt4C_B SUBJECT_SAMPLE_FACTORS - Nl_41ppt4C_3 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:4 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=1.272660153; Vol_filtered_mL=70; Replicate=3; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_41ppt4C_C 200309_Smp_41ppt4C_C SUBJECT_SAMPLE_FACTORS - Fc_1 Species:Fragilariopsis cylindrus | Salinity:31 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=0.232254; Vol_filtered_mL=69; Replicate=1; Type=Smp; Strain=CCMP1102; RAW_FILE_NAME=200309_Smp_Fc_1 200309_Smp_Fc_1 SUBJECT_SAMPLE_FACTORS - Fc_2 Species:Fragilariopsis cylindrus | Salinity:31 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=0.23529; Vol_filtered_mL=69; Replicate=2; Type=Smp; Strain=CCMP1102; RAW_FILE_NAME=200309_Smp_Fc_2 200309_Smp_Fc_2 SUBJECT_SAMPLE_FACTORS - Nl_1 Species:Nitzschia lecointei | Salinity:31 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=0.51129; Vol_filtered_mL=69; Replicate=1; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_Nl_1 200309_Smp_Nl_1 SUBJECT_SAMPLE_FACTORS - Nl_2 Species:Nitzschia lecointei | Salinity:31 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=0.490314; Vol_filtered_mL=69; Replicate=2; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_Nl_2 200309_Smp_Nl_2 SUBJECT_SAMPLE_FACTORS - Nperm_1 Species:Navicula cf. perminuta | Salinity:31 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=0.203895; Vol_filtered_mL=69; Replicate=1; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_Np_1 200309_Smp_Np_1 SUBJECT_SAMPLE_FACTORS - Nperm_2 Species:Navicula cf. perminuta | Salinity:31 | Temp_degC:-1 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=0.193545; Vol_filtered_mL=69; Replicate=2; Type=Smp; Strain=NA; RAW_FILE_NAME=200309_Smp_Np_2 200309_Smp_Np_2 SUBJECT_SAMPLE_FACTORS - Npell_1 Species:Navicula pelliculosa | Salinity:35 | Temp_degC:13 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=0.6318675; Vol_filtered_mL=69; Replicate=1; Type=Smp; Strain=CCMP543; RAW_FILE_NAME=200309_Smp_NpB12SL_AB 200309_Smp_NpB12SL_AB SUBJECT_SAMPLE_FACTORS - Npell_2 Species:Navicula pelliculosa | Salinity:35 | Temp_degC:13 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=0.6090975; Vol_filtered_mL=69; Replicate=2; Type=Smp; Strain=CCMP543; RAW_FILE_NAME=200309_Smp_NpB12SL_.D 200309_Smp_NpB12SL_CD SUBJECT_SAMPLE_FACTORS - Npell_3 Species:Navicula pelliculosa | Salinity:35 | Temp_degC:13 Light=Saturating; Cobalamin=Replete; Vol_intracellular_µL=0.59409; Vol_filtered_mL=69; Replicate=3; Type=Smp; Strain=CCMP543; RAW_FILE_NAME=200309_Smp_NpB12SL_EF 200309_Smp_NpB12SL_EF #COLLECTION CO:COLLECTION_SUMMARY Axenic cultures of three Antarctic sea-ice diatoms (N. lecointei, N. cf. CO:COLLECTION_SUMMARY perminuta, and F. cylindrus) and two temperate diatoms (T. pseudonana and N. CO:COLLECTION_SUMMARY pelliculosa) were chosen for study. Cells were harvested during exponential CO:COLLECTION_SUMMARY growth onto 47 mm 0.2 µm PTFE filters (Omnipore) using combusted glassware and CO:COLLECTION_SUMMARY gentle filtration and stored at –80 °C until extraction. For each biological CO:COLLECTION_SUMMARY replicate (n = 2 for Antarctic species, n = 3 for temperate species), two 35 mL CO:COLLECTION_SUMMARY cultures were harvested onto each filter . An un-inoculated media blank was CO:COLLECTION_SUMMARY prepared and treated in the same manner as samples. CO:SAMPLE_TYPE Cultured diatom cells CO:STORAGE_CONDITIONS Described in summary #TREATMENT TR:TREATMENT_SUMMARY Antarctic species were grown at −1°C and a PAR irradiance of 45 𝜇mol TR:TREATMENT_SUMMARY photons m−2 s−1 (16:8 light:dark cycle) using cool white lights. Temperate TR:TREATMENT_SUMMARY species were grown at 13°C and a PAR irradiance of 120 𝜇mol photons m−-2 TR:TREATMENT_SUMMARY s−-1(12:12 light:dark cycle). In both cases, light was saturating. Cultures TR:TREATMENT_SUMMARY were grown in artificial seawater (ESAW, salinity 31, for Antarctic species and TR:TREATMENT_SUMMARY Instant Ocean, salinity ~35 for temperate species). Cobalamin (vitamin B12) was TR:TREATMENT_SUMMARY replete in all cultures. To explore the effect of growth conditions on metabolic TR:TREATMENT_SUMMARY profiles using non-metric dimensional scaling analysis, samples were included of TR:TREATMENT_SUMMARY N. lecointei grown at temperatures of −1 and 4˚C and salinities of 32 and 41. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Each sample was extracted using a modified Bligh-Dyer extraction. Briefly, SP:SAMPLEPREP_SUMMARY filters were cut up and put into 15 mL teflon centrifuge tubes containing a SP:SAMPLEPREP_SUMMARY mixture of 100 µm and 400 µm silica beads. Heavy isotope-labeled internal SP:SAMPLEPREP_SUMMARY standards were added along with ~2 mL of cold aqueous solvent (50:50 SP:SAMPLEPREP_SUMMARY methanol:water) and ~3 mL of cold organic solvent (dichloromethane). The samples SP:SAMPLEPREP_SUMMARY were shaken on a FastPrep-24 Homogenizer for 30 seconds and chilled in a -20 °C SP:SAMPLEPREP_SUMMARY freezer repeatedly for three cycles of bead-beating and a total of 30 minutes of SP:SAMPLEPREP_SUMMARY chilling. The organic and aqueous layers were separated by spinning samples in a SP:SAMPLEPREP_SUMMARY centrifuge at 4,300 rpm for 2 minutes at 4 °C. The aqueous layer was removed to SP:SAMPLEPREP_SUMMARY a new glass centrifuge tube. The remaining organic fraction was rinsed three SP:SAMPLEPREP_SUMMARY more times with additions of 1 to 2 mL of 50:50 methanol:water. All aqueous SP:SAMPLEPREP_SUMMARY rinses were combined for each sample and dried down under N2 gas. The remaining SP:SAMPLEPREP_SUMMARY organic layer was transferred into a clean glass centrifuge tube and the SP:SAMPLEPREP_SUMMARY remaining bead beating tube was rinsed two more times with cold organic solvent. SP:SAMPLEPREP_SUMMARY The combined organic rinses were centrifuged, transferred to a new tube, and SP:SAMPLEPREP_SUMMARY dried under N2 gas. Dried aqueous fractions were re-dissolved in 380 µL of SP:SAMPLEPREP_SUMMARY water. Dried organic fractions were re-dissolved in 380 µL of 1:1 SP:SAMPLEPREP_SUMMARY water:acetonitrile. 20 µL of isotope-labeled injection standards in water were SP:SAMPLEPREP_SUMMARY added to both fractions. An un-inoculated media blank was prepared and treated SP:SAMPLEPREP_SUMMARY in the same manner as the samples. SP:PROCESSING_STORAGE_CONDITIONS On ice SP:EXTRACTION_METHOD Bligh-Dyer SP:EXTRACT_STORAGE -80℃ #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY See attached summary. CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME Waters Acquity I-Class CH:COLUMN_NAME SeQuant ZIC- pHILIC (150 x 2.1mm, 5um) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Thermo Q Exactive HF hybrid Orbitrap MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS See attached protocol. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS mM (intracellular concentration) MS_METABOLITE_DATA_START Samples Fc_1 Fc_2 Nperm_1 Nperm_2 Nl_1 Nl_2 Nl_32ppt-1C_1 Nl_32ppt-1C_2 Nl_32ppt-1C_3 Nl_32ppt4C_1 Nl_32ppt4C_2 Nl_32ppt4C_3 Nl_41ppt-1C_1 Nl_41ppt-1C_2 Nl_41ppt-1C_3 Nl_41ppt4C_1 Nl_41ppt4C_2 Nl_41ppt4C_3 Npell_1 Npell_2 Npell_3 Factors Species:Fragilariopsis cylindrus | Salinity:31 | Temp_degC:-1 Species:Fragilariopsis cylindrus | Salinity:31 | Temp_degC:-1 Species:Navicula cf. perminuta | Salinity:31 | Temp_degC:-1 Species:Navicula cf. perminuta | Salinity:31 | Temp_degC:-1 Species:Nitzschia lecointei | Salinity:31 | Temp_degC:-1 Species:Nitzschia lecointei | Salinity:31 | Temp_degC:-1 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:-1 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:-1 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:-1 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:4 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:4 Species:Nitzschia lecointei | Salinity:32 | Temp_degC:4 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:-1 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:-1 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:-1 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:4 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:4 Species:Nitzschia lecointei | Salinity:41 | Temp_degC:4 Species:Navicula pelliculosa | Salinity:35 | Temp_degC:13 Species:Navicula pelliculosa | Salinity:35 | Temp_degC:13 Species:Navicula pelliculosa | Salinity:35 | Temp_degC:13 (3-Carboxypropyl)trimethylammonium (TMAB) 0.248815281 0.171516943 0.21151051 0.248663501 0.012909635 0.016212709 0.006140422 0.005179659 0.006493089 0.005116949 0.004738335 0.004687017 0.006743508 0.008753397 0.007055501 0.011375511 0.007684584 0.008104792 0.002429566 0.002042382 0.002961815 (Iso)leucine 17.99796092 14.37141543 1.748252771 1.930261678 2.440341229 2.519345807 6.040888566 5.862310876 Adenosyl Homocysteine 0.075340969 0.00720887 0.056844644 0.0154331 0.002136796 0.000963847 0.000395059 0.000285267 0.007369635 0.00067298 0.024005766 0.017384601 0.019844159 Adenosyl Methionine 5.447321351 3.341175962 5.197879483 3.024898497 1.07887326 0.700995948 0.251151901 0.243854912 0.259607181 0.62938532 0.530829929 0.658134347 0.281100462 0.284509484 0.280094036 2.121457126 0.87611646 0.62265043 0.318922329 0.080702918 0.080227925 Alanine 115.3977767 94.52566536 58.4859535 77.59800842 6.594668311 6.149927843 4.190378107 3.881396317 4.537531383 2.59635995 2.560149947 3.006205982 3.54806395 4.375822542 3.715192458 3.664944081 2.894606598 2.706637102 13.72656816 12.24768314 14.53306748 Aminobutyric Acid Arginine 115.5972202 96.93536426 87.23228276 77.37448095 26.72676214 21.06124364 28.53166674 26.42229246 20.44844147 25.13535503 17.47647729 18.15793294 23.32315028 26.10985085 19.96571758 25.98232417 17.35235917 11.78669513 51.95047188 59.21365383 52.54628171 Arsenobetaine 0.00919615 0.000220524 Asparagine 11.16800048 10.31249064 5.817056239 9.150491156 2.130991731 1.979705175 1.302906804 1.426576686 1.755055384 2.984956686 2.266126136 2.757567234 1.743684191 2.220634722 1.925736717 4.266018339 4.203986679 3.699470755 2.745259114 2.740621854 2.697907101 Aspartic acid 77.97242283 84.13548785 85.21973243 97.60524638 23.96253576 28.50587439 10.87992612 11.36971993 14.60663184 5.918365941 4.953371121 6.972848571 8.147541293 10.16563525 9.602003074 7.384798467 5.907500085 6.158585274 6.104857135 7.268865684 6.034089768 Betaine 42.07370373 31.0325419 23.73411343 27.21888387 19.71719408 19.99111709 10.66444189 9.778588711 10.36964232 11.41644942 10.0903652 9.663529023 11.42484794 13.52280456 10.82624944 22.0826484 15.99057315 15.50689458 Betonicine 5.902125275 5.124310869 4.165029347 5.173092182 3.979797768 2.457236438 2.093817754 1.924276234 3.798892514 3.834748842 3.26939037 3.057889915 2.644434825 2.177696303 Carnitine 0.429307503 0.29900502 1.659347514 1.406841238 0.230726556 0.277300605 0.062509721 0.059065354 0.074462164 0.113313519 0.105881708 0.111605886 0.033155947 0.047108942 0.035919702 0.15278835 0.112332332 0.120558016 0.033296552 0.024774949 0.031500704 Choline 4.770674115 3.272052321 45.66424781 51.78792922 3.951568521 4.634924283 1.022806608 1.059079221 1.073246777 0.436390809 0.373038052 0.465698129 0.773900582 0.922208944 0.631337117 0.453163013 0.449072608 0.067072296 0.041684836 0.043831705 Dimethyl Glycine Dimethylsulfonioacetate (DMS-Ac) 0.0069948 0.004065706 0.015342538 0.008233684 0.467304331 0.471909286 0.070665143 0.061187255 0.065924711 0.123512351 0.106021 0.141887905 0.076655597 0.088788568 0.083263328 0.253063481 0.178426824 0.157361697 DMSP 89.8777507 68.97266464 140.9467369 153.3157991 88.37325556 88.67025049 37.95663725 37.58829396 38.30212743 23.96755717 21.0584327 22.52772464 30.92430756 37.99172308 30.0755893 43.26257981 30.8138578 31.29516834 Ectoine 0.01700636 0.016427704 0.007241716 0.008813689 0.001471897 0.001509537 0.000633046 0.00063142 0.000719141 0.000498341 0.000443044 0.000531398 0.000581219 0.000707807 0.000630729 0.000602925 0.000570238 0.000672207 0.001983513 0.002212526 0.002111085 Glucosylglycerol 2.50511671 1.615387733 5.388097342 7.018704633 34.32912157 34.01752345 6.259840228 5.967978802 5.833165142 4.15798683 4.499687999 4.072855777 6.608330458 8.206972403 7.102185871 6.189157499 4.59088179 5.548703417 2.218960031 1.700842918 2.004095066 Glutamic acid 302.0915854 283.7047477 334.0308693 458.8433828 54.57660205 39.64272683 22.90730634 22.46969517 21.72099184 18.72658199 12.88193096 16.19762727 15.61010467 17.57289255 13.72664966 19.13220893 15.08138564 11.82588916 23.6734599 19.80395469 22.58672285 Glutamine 37.66792161 24.32588963 63.28293945 80.81126053 13.35272272 14.45864878 6.126812368 5.695553759 7.844001597 7.768276843 6.789113502 6.713672788 6.018329509 7.202774702 7.220147778 8.210710111 7.068063323 7.127979069 7.92899374 6.22474725 7.233120994 Glycine 2.312670949 2.84999908 2.115711918 3.898350633 0.622083013 0.561763864 0.304464724 0.324036964 0.327732138 0.331086644 0.265585847 0.304253743 0.408906704 0.39128665 0.293033762 0.472216934 0.44709393 0.447376412 1.048009077 1.344349548 1.18601216 Gonyol 0.025207463 0.028342237 0.025346081 0.148702315 0.139213007 0.145548754 0.018689939 0.023795484 0.021106855 0.089444585 0.078051846 0.080833388 Histidine 2.409738267 2.032875845 1.684686552 2.24359818 0.473453798 0.418023972 0.25147613 0.23692709 0.265473763 0.320857088 0.323532851 0.360210055 0.339005533 0.325008923 0.270828863 0.717360092 0.552913649 0.530830155 0.700732122 0.642637717 Homarine 862.4923308 607.932279 769.6164364 891.0053175 1.887391004 0.650735595 0.033501585 0.012899952 0.041835906 0.016509337 0.015295869 0.029108126 0.021179169 0.022500066 0.024914314 Homoserine 20.84894987 14.03313276 13.98934095 17.59618644 0.950979511 1.068829361 0.477683081 0.455407817 0.503873199 0.192017224 0.164866499 0.162013411 0.612250067 0.746016956 0.613334974 0.197183856 0.173228022 0.151110414 Hydroxyectoine 0.00031965 0.000304142 9.69E-05 0.00011249 8.13E-05 8.82E-05 8.04E-05 9.61E-05 8.95E-05 8.04E-05 6.67E-05 Hypotaurine 0.035485301 0.042888439 0.040738839 0.282728821 0.221173046 0.2959834 0.029079225 0.032511409 0.03879985 0.293127136 0.320667948 0.225599825 Lysine 165.0799719 150.0720932 12.02951646 10.78352973 2.839971135 2.436932997 4.005434477 3.739616272 3.245282109 4.072976772 3.378098117 3.600703221 4.261423804 4.601022557 3.161398756 5.886349176 3.772776569 2.503819484 1.839436502 1.95621987 1.57585151 Methionine 23.75027903 19.22067639 5.296842663 6.991325682 0.359729913 0.31624283 0.149983521 0.150381896 0.157834478 0.111676006 0.091893264 0.108254152 0.162060413 0.218354868 0.201307871 0.135628181 0.108211568 0.104946958 0.290334469 0.280916427 0.234085448 N(e)-Acetyl-Lysine 0.074005409 0.047423564 0.037795946 0.042315962 0.006143427 0.006047415 0.008451573 0.007560621 0.01106451 0.025666574 0.018678883 0.037288553 0.008731437 0.011815273 0.013596832 0.074543768 0.041408519 0.039930865 0.011883518 0.005309388 0.003776951 Ornithine 608.5175125 761.8505989 45.42360635 40.60684423 5.076173009 3.55842725 3.065552103 2.98860457 2.651858049 3.15816805 2.90760239 2.132451257 2.909561933 2.957590214 2.033273698 2.318729867 1.603262813 1.377854968 9.711387222 10.01795102 8.782030411 Proline 142.1000007 109.3283207 507.0386759 746.2533583 49.98175468 48.73741274 15.97365495 18.09035288 18.09414198 10.18484278 9.18333706 12.47604754 36.74672403 47.20485391 37.64533372 34.8523381 25.44475295 24.63783559 96.31382043 75.32686382 85.55505973 Sarcosine 1.99053822 1.371358684 0.734677767 1.141674546 0.093405965 0.119922698 0.030388539 0.037313644 0.040109894 0.066077861 0.061943533 0.063684924 0.05033598 0.067030882 0.050905221 0.06888162 0.084967836 0.068747847 Serine 15.82638061 15.21094623 7.989346215 12.40826212 2.185259313 2.044081131 0.979578519 0.937879934 1.058747599 0.906645323 0.806381107 0.838532335 1.087841344 0.972108704 0.788941017 1.048539513 0.961472333 0.932628764 1.262154275 1.190246846 1.180419837 Stachydrine hydrochloride 0.007058172 0.005377246 0.007190518 0.007845373 0.001739727 0.002018675 0.000865559 0.000776554 0.001043972 0.000952532 0.001483818 0.001594551 Threonine 24.02459536 11.72196338 18.03336544 12.32421931 1.511483459 0.795405283 0.485983405 0.41712759 0.320408678 0.441345053 0.395347699 0.284526693 0.474624292 0.443974189 0.260200106 0.893273767 0.458171573 0.338956682 0.644885831 0.485735728 0.575301256 trans Hydroxyl proline 0.323084956 0.218096316 0.617123473 0.655679767 0.061375604 0.072788566 0.027429782 0.028414903 0.029797436 0.026860999 0.025547819 0.028767643 0.066840091 0.081426528 0.065185607 0.092149503 0.067301717 0.069403075 0.241290814 0.163394615 0.212167865 Trigonelline 0.00573808 0.005801546 0.002169511 0.001654133 0.001794472 0.004108829 0.003206721 0.002107947 Trimethylamine N-oxide 0.042354075 0.034632872 Trimethylammonium Propionate (TMAP) 0.363460627 0.377091736 0.045608564 0.049451994 0.048597982 0.047535738 0.041348011 0.046095041 0.027409411 0.032709761 0.031731604 0.042605032 0.04893376 0.042479758 Tyrosine 35.01637628 30.52153879 1.014429595 1.175633707 0.589623025 0.423731929 0.12870241 0.129554987 0.146958989 0.963184436 0.764916229 1.030313422 0.113933296 0.107692157 0.094317329 1.515959192 1.579190821 1.511412055 0.143140853 0.100319436 0.099308939 Valine MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name quantitated m/z KEGG_ID CHEBI KEGGNAME MS_method (3-Carboxypropyl)trimethylammonium (TMAB) 146.118104 CHEBI:1941 HILIC_QE_Pos (Iso)leucine 132.102454 C00407 CHEBI:17191 L-Isoleucine; 2-Amino-3-methylvaleric acid HILIC_QE_Pos Adenosyl Homocysteine 385.129416 C00021 CHEBI:16680 S-Adenosyl-L-homocysteine; S-Adenosylhomocysteine HILIC_QE_Pos Adenosyl Methionine 399.145066 C00019 CHEBI:15414 S-Adenosyl-L-methionine; S-Adenosylmethionine; AdoMet; SAM HILIC_QE_Pos Alanine 90.055504 C00041 CHEBI:16977 L-Alanine; L-2-Aminopropionic acid; L-alpha-Alanine HILIC_QE_Pos Aminobutyric Acid 104.071154 C00334 CHEBI:16865 4-Aminobutanoate; 4-Aminobutanoic acid; 4-Aminobutyrate; 4-Aminobutyric acid; gamma-Aminobutyric acid; GABA HILIC_QE_Pos Arginine 175.119501 C00062 CHEBI:16467 L-Arginine; (S)-2-Amino-5-guanidinovaleric acid; L-Arg HILIC_QE_Pos Arsenobetaine 179.005326 C19331 CHEBI:82392 Arsenobetaine HILIC_QE_Pos Asparagine 133.061318 C00152 CHEBI:17196 L-Asparagine; 2-Aminosuccinamic acid HILIC_QE_Pos Aspartic acid 134.045334 C00049 CHEBI:17053 L-Aspartate; L-Aspartic acid; 2-Aminosuccinic acid; L-Asp HILIC_QE_Pos Betaine 118.086804 C00719 CHEBI:17750 Betaine; Trimethylaminoacetate; Glycine betaine; N,N,N-Trimethylglycine; Trimethylammonioacetate HILIC_QE_Pos Betonicine 160.097369 C08269 CHEBI:85533 (-)-Betonicine; Betonicine; trans-4-Hydroxy-L-proline betaine HILIC_QE_Pos Carnitine 162.113019 C00487 CHEBI:17126 Carnitine; gamma-Trimethyl-hydroxybutyrobetaine; 3-Hydroxy-4-trimethylammoniobutanoate HILIC_QE_Pos Choline 104.107539 C00114 CHEBI:15354 Choline; Bilineurine HILIC_QE_Pos Dimethyl Glycine 104.071154 C01026 CHEBI:17724 N,N-Dimethylglycine; Dimethylglycine HILIC_QE_Pos Dimethylsulfonioacetate (DMS-Ac) 121.032327 C03392 CHEBI:4623 Dimethylsulfonioacetate HILIC_QE_Pos DMSP 135.047977 C04022 CHEBI:16457 S,S-Dimethyl-beta-propiothetin; S-Dimethylsulfonium propionic acid; Dimethylpropiothetin; DMPT; DMSP HILIC_QE_Pos Ectoine 143.082053 C06231 CHEBI:27592 Ectoine; L-Ectoine HILIC_QE_Pos Glucosylglycerol 255.107995 C11546 CHEBI:82766 2-O-(alpha-D-Glucopyranosyl)glycerol; 2-O-alpha-D-Glucosylglycerol HILIC_QE_Pos Glutamic acid 148.060984 C00025 CHEBI:16015 L-Glutamate; L-Glutamic acid; L-Glutaminic acid; Glutamate HILIC_QE_Pos Glutamine 147.076968 C00064 CHEBI:18050 L-Glutamine; L-2-Aminoglutaramic acid HILIC_QE_Pos Glycine 76.039854 C00037 CHEBI:15428 Glycine; Aminoacetic acid; Gly HILIC_QE_Pos Gonyol 179.074191 HILIC_QE_Pos Histidine 156.077302 C00135 CHEBI:15971 L-Histidine; (S)-alpha-Amino-1H-imidazole-4-propionic acid HILIC_QE_Pos Homarine 138.055503 CHEBI:69061 HILIC_QE_Pos Homoserine 120.066069 C00263 CHEBI:15699 L-Homoserine; 2-Amino-4-hydroxybutyric acid HILIC_QE_Pos Hydroxyectoine 159.076968 C16432 CHEBI:49432 5-Hydroxyectoine HILIC_QE_Pos Hypotaurine 110.027576 C00519 CHEBI:16668 Hypotaurine; 2-Aminoethanesulfinic acid HILIC_QE_Pos Lysine 147.113353 C00047 CHEBI:18019 L-Lysine; Lysine acid; 2,6-Diaminohexanoic acid HILIC_QE_Pos Methionine 150.058876 C00073 CHEBI:16811 L-Methionine; Methionine; L-2-Amino-4methylthiobutyric acid HILIC_QE_Pos N(e)-Acetyl-Lysine 189.123918 C02727 CHEBI:17752 N6-Acetyl-L-lysine HILIC_QE_Pos Ornithine 133.097703 C00077 CHEBI:15729 L-Ornithine; (S)-2,5-Diaminovaleric acid; (S)-2,5-Diaminopentanoic acid; (S)-2,5-Diaminopentanoate HILIC_QE_Pos Proline 116.071154 C00148 CHEBI:17203 L-Proline; 2-Pyrrolidinecarboxylic acid HILIC_QE_Pos Sarcosine 90.055504 C00213 CHEBI:15611 Sarcosine; N-Methylglycine HILIC_QE_Pos Serine 106.050419 C00065 CHEBI:17115 L-Serine; L-2-Amino-3-hydroxypropionic acid; L-3-Hydroxy-alanine; Serine HILIC_QE_Pos Stachydrine hydrochloride 144.102454 C10172 CHEBI:35280 Stachydrine; Proline betaine; L-Proline betaine HILIC_QE_Pos Threonine 120.066069 C00188 CHEBI:16857 L-Threonine; 2-Amino-3-hydroxybutyric acid HILIC_QE_Pos trans Hydroxyl proline 132.066069 C01157 CHEBI:18095 Hydroxyproline; L-Hydroxyproline; trans-4-Hydroxy-L-proline HILIC_QE_Pos Trigonelline 138.055503 C01004 CHEBI:18123 N-Methylnicotinate; Trigonelline; Trigenelline; 1-Methylpyridinio-3-carboxylate; Betaine nicotinate; Caffearin; Gynesine HILIC_QE_Pos Trimethylamine N-oxide 76.076239 C01104 CHEBI:15724 Trimethylamine N-oxide; (CH3)3NO HILIC_QE_Pos Trimethylammonium Propionate (TMAP) 132.1018 C08263 CHEBI:28825 beta-Alaninebetaine; beta-Alanine betaine; Propiobetaine; Trimethylalanine HILIC_QE_Pos Tyrosine 182.081719 C00082 CHEBI:17895 L-Tyrosine; (S)-3-(p-Hydroxyphenyl)alanine; (S)-2-Amino-3-(p-hydroxyphenyl)propionic acid; Tyrosine HILIC_QE_Pos Valine 118.086804 C00183 CHEBI:16414 L-Valine; 2-Amino-3-methylbutyric acid HILIC_QE_Pos METABOLITES_END #END