#METABOLOMICS WORKBENCH dongf01_20230127_194251 DATATRACK_ID:3712 STUDY_ID:ST002463 ANALYSIS_ID:AN004018 PROJECT_ID:PR001590 VERSION 1 CREATED_ON January 29, 2023, 7:29 am #PROJECT PR:PROJECT_TITLE Tryptophan metabolites in mouse serum PR:PROJECT_TYPE MS quantitative study PR:PROJECT_SUMMARY quantify tryptophan metabolites in mouse serum PR:INSTITUTE The Pennsylvania State University PR:LAST_NAME DONG PR:FIRST_NAME FANGCONG PR:ADDRESS 309 Life Sciences Building, State college, PA, 16802, USA PR:EMAIL fxd93@psu.edu PR:PHONE 8146990203 #STUDY ST:STUDY_TITLE tryptophan metabolites in mouse serum ST:STUDY_SUMMARY tryptophan metabolites in mouse serum ST:INSTITUTE Pennsylvania State University ST:LAST_NAME DONG ST:FIRST_NAME FANGCONG ST:ADDRESS 309 Life Sciences Building, University Park, PA 16802 ST:EMAIL fxd93@psu.edu ST:PHONE 8146990203 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 #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 - Batch1_SCH1 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch1_SCH1.mzML SUBJECT_SAMPLE_FACTORS - Batch1_SCH2 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch1_SCH2.mzML SUBJECT_SAMPLE_FACTORS - Batch1_SCH3 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch1_SCH3.mzML SUBJECT_SAMPLE_FACTORS - Batch1_SCH4 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch1_SCH4.mzML SUBJECT_SAMPLE_FACTORS - Batch1_SCH5 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch1_SCH5.mzML SUBJECT_SAMPLE_FACTORS - Batch1_SCH6 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch1_SCH6.mzML SUBJECT_SAMPLE_FACTORS - Batch1_std1 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch1_10nM_std1.mzML SUBJECT_SAMPLE_FACTORS - Batch1_std2 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch1_30nM_std2.mzML SUBJECT_SAMPLE_FACTORS - Batch1_std3 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch1_50nM_std3.mzML SUBJECT_SAMPLE_FACTORS - Batch1_std4 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch1_100nM_std4.mzML SUBJECT_SAMPLE_FACTORS - Batch1_std5 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch1_200nM_std5.mzML SUBJECT_SAMPLE_FACTORS - Batch1_std6 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch1_500nM_std6.mzML SUBJECT_SAMPLE_FACTORS - Batch1_std7 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch1_1000nM_std7.mzML SUBJECT_SAMPLE_FACTORS - Batch1_std8 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch1_2000nM_std8.mzML SUBJECT_SAMPLE_FACTORS - Batch1_std9 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch1_5000nM_std9.mzML SUBJECT_SAMPLE_FACTORS - Batch1_std10 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch1_10000nM_std10.mzML SUBJECT_SAMPLE_FACTORS - Batch2_SGF1 diluted factor:2.5 Genotype=Germ free; Treatment=chow diet; RAW_FILE_NAME=Batch2_SGF1.mzML SUBJECT_SAMPLE_FACTORS - Batch2_SGF2 diluted factor:2.5 Genotype=Germ free; Treatment=chow diet; RAW_FILE_NAME=Batch2_SGF2.mzML SUBJECT_SAMPLE_FACTORS - Batch2_SGF3 diluted factor:2.5 Genotype=Germ free; Treatment=chow diet; RAW_FILE_NAME=Batch2_SGF3.mzML SUBJECT_SAMPLE_FACTORS - Batch2_SGF4 diluted factor:2.5 Genotype=Germ free; Treatment=chow diet; RAW_FILE_NAME=Batch2_SGF4.mzML SUBJECT_SAMPLE_FACTORS - Batch2_SGF5 diluted factor:2.5 Genotype=Germ free; Treatment=chow diet; RAW_FILE_NAME=Batch2_SGF5.mzML SUBJECT_SAMPLE_FACTORS - Batch2_SGF6 diluted factor:2.5 Genotype=Germ free; Treatment=chow diet; RAW_FILE_NAME=Batch2_SGF6.mzML SUBJECT_SAMPLE_FACTORS - Batch2_SGF7 diluted factor:2.5 Genotype=Germ free; Treatment=chow diet; RAW_FILE_NAME=Batch2_SGF7.mzML SUBJECT_SAMPLE_FACTORS - Batch2_Sconv1 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch2_Sconv1.mzML SUBJECT_SAMPLE_FACTORS - Batch2_Sconv2 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch2_Sconv2.mzML SUBJECT_SAMPLE_FACTORS - Batch2_Sconv3 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch2_Sconv3.mzML SUBJECT_SAMPLE_FACTORS - Batch2_Sconv4 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch2_Sconv4.mzML SUBJECT_SAMPLE_FACTORS - Batch2_Sconv5 diluted factor:2.5 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch2_Sconv5.mzML SUBJECT_SAMPLE_FACTORS - Batch2_std1 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch2_10nM_std1.mzML SUBJECT_SAMPLE_FACTORS - Batch2_std2 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch2_30nM_std2.mzML SUBJECT_SAMPLE_FACTORS - Batch2_std3 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch2_50nM_std3.mzML SUBJECT_SAMPLE_FACTORS - Batch2_std4 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch2_100nM_std4.mzML SUBJECT_SAMPLE_FACTORS - Batch2_std5 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch2_200nM_std5.mzML SUBJECT_SAMPLE_FACTORS - Batch2_std6 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch2_500nM_std6.mzML SUBJECT_SAMPLE_FACTORS - Batch2_std7 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch2_1000nM_std7.mzML SUBJECT_SAMPLE_FACTORS - Batch2_std8 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch2_2000nM_std8.mzML SUBJECT_SAMPLE_FACTORS - Batch2_std9 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch2_5000nM_std9.mzML SUBJECT_SAMPLE_FACTORS - Batch2_std10 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch2_10000nM_std10.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SP1 diluted factor:1.25 Genotype=wide type; Treatment=purified diet; RAW_FILE_NAME=Batch3_SP1.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SP2 diluted factor:1.25 Genotype=wide type; Treatment=purified diet; RAW_FILE_NAME=Batch3_SP2.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SP3 diluted factor:1.25 Genotype=wide type; Treatment=purified diet; RAW_FILE_NAME=Batch3_SP3.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SP4 diluted factor:1.25 Genotype=wide type; Treatment=purified diet; RAW_FILE_NAME=Batch3_SP4.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SP5 diluted factor:1.25 Genotype=wide type; Treatment=purified diet; RAW_FILE_NAME=Batch3_SP5.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SP6 diluted factor:1.25 Genotype=wide type; Treatment=purified diet; RAW_FILE_NAME=Batch3_SP6.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SWT7 diluted factor:1.25 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch3_SWT7.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SWT8 diluted factor:1.25 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch3_SWT8.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SWT9 diluted factor:1.25 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch3_SWT9.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SWT10 diluted factor:1.25 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch3_SWT10.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SWT11 diluted factor:1.25 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch3_SWT11.mzML SUBJECT_SAMPLE_FACTORS - Batch3_SWT12 diluted factor:1.25 Genotype=wide type; Treatment=chow diet; RAW_FILE_NAME=Batch3_SWT12.mzML SUBJECT_SAMPLE_FACTORS - Batch3_std1 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch3_10nM_std1.mzML SUBJECT_SAMPLE_FACTORS - Batch3_std2 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch3_30nM_std2.mzML SUBJECT_SAMPLE_FACTORS - Batch3_std3 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch3_50nM_std3.mzML SUBJECT_SAMPLE_FACTORS - Batch3_std4 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch3_100nM_std4.mzML SUBJECT_SAMPLE_FACTORS - Batch3_std5 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch3_200nM_std5.mzML SUBJECT_SAMPLE_FACTORS - Batch3_std6 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch3_500nM_std6.mzML SUBJECT_SAMPLE_FACTORS - Batch3_std7 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch3_1000nM_std7.mzML SUBJECT_SAMPLE_FACTORS - Batch3_std8 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch3_2000nM_std8.mzML SUBJECT_SAMPLE_FACTORS - Batch3_std9 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch3_5000nM_std9.mzML SUBJECT_SAMPLE_FACTORS - Batch3_std10 diluted factor:1 Genotype=-; Treatment=-; RAW_FILE_NAME=Batch3_10000nM_std10.mzML #COLLECTION CO:COLLECTION_SUMMARY Serum samples from conventional and GF mice were collected and stored at -80°C. CO:SAMPLE_TYPE Blood (serum) #TREATMENT TR:TREATMENT_SUMMARY Thawed on ice, 25 µL of serum sample, was mixed with 100 µL extraction solvent TR:TREATMENT_SUMMARY of ice-cold methanol containing indole-3-acetic acid-d4 and kynurenic acid-d5. TR:TREATMENT_SUMMARY Mixture was vortexed and incubated at −20 °C for 30 min. Following TR:TREATMENT_SUMMARY centrifugation at 12,000 × g for 15 min at 4℃, 90 µL supernatant was TR:TREATMENT_SUMMARY collected and subsequently evaporated to dryness (Thermo Scientific, Waltham, TR:TREATMENT_SUMMARY MA) and dissolved in 45 µL of 10% acetonitrile containing 1 µM chlorpropamide. TR:TREATMENT_SUMMARY After centrifugation at 12,000 × g for 15 min at 4℃, supernatants were TR:TREATMENT_SUMMARY transferred to autosampler vials for LC-MS analysis. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY GF mice were reared in standard, sterile (autoclaved cages containing sterile SP:SAMPLEPREP_SUMMARY (autoclaved) bedding inside a hermetically sealed isolator with HEPA-filtered SP:SAMPLEPREP_SUMMARY airflow and maintained on sterile (autoclaved) water and sterile (autoclaved) SP:SAMPLEPREP_SUMMARY food (LabDiet® 5013, Land O’Lakes) ad libitum. One of two groups of SP:SAMPLEPREP_SUMMARY conventional C57BL/6J mice with ad libitum access to a standard laboratory chow SP:SAMPLEPREP_SUMMARY were switched onto a semi-purified diet (AIN-93G) while the second remained on SP:SAMPLEPREP_SUMMARY standard chow. After one week, portal vein blood was collected. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Shimadzu 20AD CH:COLUMN_NAME Waters ACQUITY UPLC BEH C18 (100 x 2.1mm,1.7um) CH:SOLVENT_A Water with 0.1% formic acid CH:SOLVENT_B Acetonitrile with 0.1% formic acid CH:FLOW_GRADIENT The initial condition were 97% A and 3 % B, increasing to 45% B at 10 min, 75% B CH:FLOW_GRADIENT at 12 min where it was held at 75% B until 17.5 min before returning to the CH:FLOW_GRADIENT initial conditions. CH:FLOW_RATE 0.25 mL/min CH:COLUMN_TEMPERATURE 55 #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME ABI Sciex 5600 TripleTOF MS:INSTRUMENT_TYPE Triple TOF MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS Peakview was used for data analysis. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS uM MS_METABOLITE_DATA_START Samples Batch2_SGF1 Batch2_SGF2 Batch2_SGF3 Batch2_SGF4 Batch2_SGF5 Batch2_SGF6 Batch2_SGF7 Batch2_Sconv1 Batch2_Sconv2 Batch2_Sconv3 Batch2_Sconv4 Batch2_Sconv5 Batch1_SCH1 Batch1_SCH2 Batch1_SCH3 Batch1_SCH4 Batch1_SCH5 Batch1_SCH6 Batch3_SP1 Batch3_SP2 Batch3_SP3 Batch3_SP4 Batch3_SP5 Batch3_SP6 Batch3_SWT7 Batch3_SWT8 Batch3_SWT9 Batch3_SWT10 Batch3_SWT11 Batch3_SWT12 Factors diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:1.25 diluted factor:1.25 diluted factor:1.25 diluted factor:1.25 diluted factor:1.25 diluted factor:1.25 diluted factor:1.25 diluted factor:1.25 diluted factor:1.25 diluted factor:1.25 diluted factor:1.25 diluted factor:1.25 indole-3-acetic acid 0.514132922 1.338609191 0.818466719 0.886584282 1.185463502 0.962337944 0.759968364 0.490730219 0.341126759 0.338016013 0.497053252 0.684028333 1.329244799 1.274139964 2.026068862 1.263826796 1.554504132 1.018393653 0.409614549 0.418421187 0.38531446 0.488603092 0.339141919 0.459557431 0.960921935 0.81930508 0.616515286 0.461825736 0.437711931 0.773665704 kynurenic acid 0.256955658 0.521982016 0.124151783 0.208607953 0.428180034 0.233335368 0.519988452 0.722066905 0.741645583 0.444203119 0.684695993 0.69319022 0.101992524 0.115135493 0.377144461 0.312293614 0.540536039 0.432148455 0.099146318 0.225981099 0.195656677 0.224251127 0.116095287 0.264382773 0.303456889 0.382001076 0.234545787 0.156770838 0.16910886 0.211006638 2-oxindole 0 0 0 0 0 0 0 0 0.093894402 0.039511313 0.282157867 0.202691312 0.088032065 0.079431381 0.087318744 0.134961333 0.071200638 0.072451548 0.23143409 0.023283957 0.030009291 0.04432406 0.014095096 0.026261464 0.106041972 0.051533444 0.161192715 0.042649914 0.030552481 0.134432891 indole-3-carboxaldehyde 0.437248663 0.121226613 0.535137606 0.667533872 0.908535858 0.304871897 0.712118747 0.07493086 0.368030744 0.295475424 0.365401623 0.237983004 0.628433502 0.44835185 0.624104163 0.729612149 0.45771167 0.621492118 0.254232875 0.294554497 0.134753173 0.183319878 0.164310776 0.119056116 0.206206981 0.174777966 0.218447176 0.201761422 0.159401834 0.170609255 indole-3-propionic acid 0 0 0 0 0 0 0 1.616432643 2.039424064 1.605984236 2.871811498 2.684295649 11.75428777 8.928927222 4.755583543 10.96966672 9.463394096 8.434854202 0.114591984 0.08170122 0.039903542 0.127848101 0.053199364 0.320535998 6.509461745 5.269438034 5.897730019 4.596820172 2.752775029 3.245398418 indole-3-lactic acid 1.856958296 2.573463893 2.439657629 1.763473055 4.669059888 2.491677124 3.06824567 1.570081523 2.71017839 2.430671026 3.879917367 1.943955042 1.994997676 1.901861407 2.151726276 1.702536992 1.886241139 1.486314509 1.508647812 1.219181624 1.233100702 1.341683489 1.009926596 1.069360181 2.286749683 1.777888281 2.211711848 1.315848539 0.952493469 1.439568172 5-hydroxyindole-3-acetic acid 0.592289135 1.110968862 1.024182859 0.769555517 0.741972039 0.823119537 0.720040787 0.679105711 0.552411115 0.663989005 0.433011842 0.322661094 1.08319054 1.649490019 1.360418297 1.028053161 1.340785411 1.075695368 0.496667104 0.403302429 0.634261674 0.634084909 0.329544514 0.619875846 0.818148455 0.521160108 0.712989819 0.538348485 0.526727828 0.860375273 kynurenine 0.863275864 0.882985061 1.426898485 1.114536519 2.106893649 1.380506898 1.546074845 1.20564108 0.960076507 1.194084634 0.945043806 0.771529752 1.127368346 2.588643618 2.194209069 3.392111981 1.858135234 2.050654068 1.458511047 1.616089412 1.773706441 1.649299138 1.33078649 1.94623977 1.165816765 1.0913281 1.005825713 1.540442329 1.141790806 0.682042694 3-indoleacrylic acid 0 0 0 0 0 0 0 0.189347573 0.190059757 0.081132755 0.047443372 0.474275654 0.208031165 0.341983713 0.070466277 0.0277149 0.074720264 0.167332618 0.109454852 0.131195231 0.072510075 0.099732337 0.067996144 0.104797782 0.420608026 0.296204461 0.44073677 0.244958041 0.166171623 0.291355226 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name quantified m/z Pubchem ID indole-3-acetic acid 130.06 802 kynurenic acid 144.03 3845 2-oxindole 77.01 321710 indole-3-carboxaldehyde 91.05 10256 indole-3-propionic acid 130.09 3744 indole-3-lactic acid 130.09 92904 5-hydroxyindole-3-acetic acid 146.03 1826 kynurenine 146.05 846 3-indoleacrylic acid 115.02 5375048 METABOLITES_END #END