#METABOLOMICS WORKBENCH dongf01_20230127_175842 DATATRACK_ID:3711 STUDY_ID:ST002462 ANALYSIS_ID:AN004017 PROJECT_ID:PR001589 VERSION 1 CREATED_ON February 1, 2023, 9:25 am #PROJECT PR:PROJECT_TITLE Tryptophan metabolites in human serum PR:PROJECT_TYPE MS quantitative study PR:PROJECT_SUMMARY Quantify tryptophan metabolites in human 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 human sera on a defined diet ST:STUDY_SUMMARY Quantify tryptophan metabolites in human sera on a defined diet 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 Human 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 - 16DP3 diluted factor:2.5 RAW_FILE_NAME=16DP3.mzML SUBJECT_SAMPLE_FACTORS - 14DP2 diluted factor:2.5 RAW_FILE_NAME=14DP2.mzML SUBJECT_SAMPLE_FACTORS - 52DP1 diluted factor:2.5 RAW_FILE_NAME=52DP1.mzML SUBJECT_SAMPLE_FACTORS - 25DP3 diluted factor:2.5 RAW_FILE_NAME=25DP3.mzML SUBJECT_SAMPLE_FACTORS - 12DP3 diluted factor:2.5 RAW_FILE_NAME=12DP3.mzML SUBJECT_SAMPLE_FACTORS - 17DP1 diluted factor:2.5 RAW_FILE_NAME=17DP1.mzML SUBJECT_SAMPLE_FACTORS - 20DP2 diluted factor:2.5 RAW_FILE_NAME=20DP2.mzML SUBJECT_SAMPLE_FACTORS - 29DP1 diluted factor:2.5 RAW_FILE_NAME=29DP1.mzML SUBJECT_SAMPLE_FACTORS - 62DP3 diluted factor:2.5 RAW_FILE_NAME=62DP3.mzML SUBJECT_SAMPLE_FACTORS - 58DP3 diluted factor:2.5 RAW_FILE_NAME=58DP3.mzML SUBJECT_SAMPLE_FACTORS - 58DP2 diluted factor:2.5 RAW_FILE_NAME=58DP2.mzML SUBJECT_SAMPLE_FACTORS - 37DP2 diluted factor:2.5 RAW_FILE_NAME=37DP2.mzML SUBJECT_SAMPLE_FACTORS - 54DP3 diluted factor:2.5 RAW_FILE_NAME=54DP3.mzML SUBJECT_SAMPLE_FACTORS - 61DP2 diluted factor:2.5 RAW_FILE_NAME=61DP2.mzML SUBJECT_SAMPLE_FACTORS - 54DP2 diluted factor:2.5 RAW_FILE_NAME=54DP2.mzML SUBJECT_SAMPLE_FACTORS - 55DP2 diluted factor:2.5 RAW_FILE_NAME=55DP2.mzML SUBJECT_SAMPLE_FACTORS - 40DP1 diluted factor:2.5 RAW_FILE_NAME=40DP1.mzML SUBJECT_SAMPLE_FACTORS - 50DP3 diluted factor:2.5 RAW_FILE_NAME=50DP3.mzML SUBJECT_SAMPLE_FACTORS - 37DP3 diluted factor:2.5 RAW_FILE_NAME=37DP3.mzML SUBJECT_SAMPLE_FACTORS - 61DP1 diluted factor:2.5 RAW_FILE_NAME=61DP1.mzML SUBJECT_SAMPLE_FACTORS - 47DP2 diluted factor:2.5 RAW_FILE_NAME=47DP2.mzML SUBJECT_SAMPLE_FACTORS - 60DP2 diluted factor:2.5 RAW_FILE_NAME=60DP2.mzML SUBJECT_SAMPLE_FACTORS - 17DP2 diluted factor:2.5 RAW_FILE_NAME=17DP2.mzML SUBJECT_SAMPLE_FACTORS - 20DP3 diluted factor:2.5 RAW_FILE_NAME=20DP3.mzML SUBJECT_SAMPLE_FACTORS - 55DP1 diluted factor:2.5 RAW_FILE_NAME=55DP1.mzML SUBJECT_SAMPLE_FACTORS - 25DP2 diluted factor:2.5 RAW_FILE_NAME=25DP2.mzML SUBJECT_SAMPLE_FACTORS - 47DP1 diluted factor:2.5 RAW_FILE_NAME=47DP1.mzML SUBJECT_SAMPLE_FACTORS - 40DP3 diluted factor:2.5 RAW_FILE_NAME=40DP3.mzML SUBJECT_SAMPLE_FACTORS - 63DP3 diluted factor:2.5 RAW_FILE_NAME=63DP3.mzML SUBJECT_SAMPLE_FACTORS - 63DP2 diluted factor:2.5 RAW_FILE_NAME=63DP2.mzML SUBJECT_SAMPLE_FACTORS - 62DP1 diluted factor:2.5 RAW_FILE_NAME=62DP1.mzML SUBJECT_SAMPLE_FACTORS - 16DP1 diluted factor:2.5 RAW_FILE_NAME=16DP1.mzML SUBJECT_SAMPLE_FACTORS - 30DP1 diluted factor:2.5 RAW_FILE_NAME=30DP1.mzML SUBJECT_SAMPLE_FACTORS - 51DP1 diluted factor:2.5 RAW_FILE_NAME=51DP1.mzML SUBJECT_SAMPLE_FACTORS - 52DP3 diluted factor:2.5 RAW_FILE_NAME=52DP3.mzML SUBJECT_SAMPLE_FACTORS - 12DP1 diluted factor:2.5 RAW_FILE_NAME=12DP1.mzML SUBJECT_SAMPLE_FACTORS - 14DP1 diluted factor:2.5 RAW_FILE_NAME=14DP1.mzML SUBJECT_SAMPLE_FACTORS - 29DP2 diluted factor:2.5 RAW_FILE_NAME=29DP2.mzML SUBJECT_SAMPLE_FACTORS - 60DP3 diluted factor:2.5 RAW_FILE_NAME=60DP3.mzML SUBJECT_SAMPLE_FACTORS - 30DP2 diluted factor:2.5 RAW_FILE_NAME=30DP2.mzML SUBJECT_SAMPLE_FACTORS - std1 diluted factor:1 RAW_FILE_NAME=10nM_std1 SUBJECT_SAMPLE_FACTORS - std2 diluted factor:1 RAW_FILE_NAME=30nM_std2 SUBJECT_SAMPLE_FACTORS - std3 diluted factor:1 RAW_FILE_NAME=50nM_std3 SUBJECT_SAMPLE_FACTORS - std4 diluted factor:1 RAW_FILE_NAME=100nM_std4 SUBJECT_SAMPLE_FACTORS - std5 diluted factor:1 RAW_FILE_NAME=200nM_std5 SUBJECT_SAMPLE_FACTORS - std6 diluted factor:1 RAW_FILE_NAME=500nM_std6 SUBJECT_SAMPLE_FACTORS - std7 diluted factor:1 RAW_FILE_NAME=1000nM_std7 SUBJECT_SAMPLE_FACTORS - std8 diluted factor:1 RAW_FILE_NAME=2000nM_std8 SUBJECT_SAMPLE_FACTORS - std9 diluted factor:1 RAW_FILE_NAME=5000nM_std9 SUBJECT_SAMPLE_FACTORS - std10 diluted factor:1 RAW_FILE_NAME=10000nM_std10 #COLLECTION CO:COLLECTION_SUMMARY human sera were processed by ice-cold methanol. 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 40 selected trial participants were fed a standardized isocaloric diet SP:SAMPLEPREP_SUMMARY (representative of an average American macronutrient intake) to achieve weight SP:SAMPLEPREP_SUMMARY maintenance for four weeks At the end of the diet period and following a 12 h SP:SAMPLEPREP_SUMMARY fast (and avoidance of over-the-counter medication), serum was collected, stored SP:SAMPLEPREP_SUMMARY at -80℃ and subjected to analysis by metabolomics. #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 100% water; 0.1% formic acid CH:SOLVENT_B 100% acetonitrile; 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 16DP3 14DP2 52DP1 25DP3 12DP3 17DP1 20DP2 29DP1 62DP3 58DP3 58DP2 37DP2 54DP3 61DP2 54DP2 55DP2 40DP1 50DP3 37DP3 61DP1 47DP2 60DP2 17DP2 20DP3 55DP1 25DP2 47DP1 40DP3 63DP3 63DP2 62DP1 16DP1 30DP1 51DP1 52DP3 12DP1 14DP1 29DP2 60DP3 30DP2 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: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:2.5 diluted factor:2.5 diluted factor:2.5 diluted factor:2.5 indole-3-acetic acid 2.0161 2.1811 2.8042 2.3732 0.8777 1.1019 2.7672 2.7405 1.6428 1.4507 1.4810 1.2241 1.3835 2.3948 2.6223 1.3966 2.2169 2.3166 1.4447 2.2327 1.9026 2.4017 1.6661 2.4417 1.4516 2.6014 1.7945 0.9762 1.8940 1.6203 1.5641 2.3224 1.7830 1.1988 2.6305 0.9767 1.9751 1.4050 2.4545 2.3885 kynurenic acid 0.1969 0.1513 0.1053 0.0843 0.0147 0.0000 0.0000 0.1761 0.1005 0.0376 0.0252 0.0484 0.0826 0.0973 0.0393 0.0474 0.0587 0.0518 0.1016 0.1191 0.0150 0.0996 0.0957 0.0155 0.0089 0.0058 0.0686 0.0129 0.0348 0.0719 0.1368 0.1061 0.0490 0.0412 0.1570 0.0445 0.1386 0.2362 0.1431 0.0304 indole-3-carboxaldehyde 0.1072 0.1501 0.1800 0.1595 0.0875 0.0868 0.1414 0.2174 0.1087 0.1743 0.0699 0.1364 0.0781 0.0980 0.1231 0.0635 0.0774 0.0796 0.1293 0.1717 0.1269 0.1084 0.1080 0.0931 0.1082 0.0738 0.1005 0.0956 0.0604 0.1559 0.1288 0.0827 0.1139 0.0834 0.1568 0.0549 0.1174 0.1659 0.1663 0.0406 indole-3-propionic acid 1.1900 2.4394 3.2499 0.3381 3.9686 1.4906 1.9418 3.6792 2.6916 1.1466 0.9735 1.9573 1.3559 2.9911 1.5449 6.4280 3.4239 0.9883 1.0581 2.0743 2.1750 1.0903 3.2329 3.2146 3.8239 0.4902 3.6770 1.5159 1.6245 0.9311 4.6761 1.8167 1.0527 2.2060 2.5409 2.2787 3.7029 2.9363 1.4721 0.8343 indole-3-lactic acid 1.5855 1.2334 2.0980 4.2225 1.1817 0.8442 0.9790 1.6573 1.1362 1.4475 2.1760 1.4333 1.1363 1.5781 1.1779 0.8050 1.4281 0.7490 1.6720 1.7932 1.1335 1.4277 1.0788 1.1995 0.7352 0.9517 1.1882 1.1839 1.1085 1.3455 1.5967 1.4032 1.2272 1.4622 2.5416 1.0224 1.2819 1.1807 1.8655 1.2114 kynurenine 2.4808 3.1575 1.4821 0.6956 0.8624 0.8452 2.2991 1.7963 2.0698 1.7218 1.1687 1.4402 1.5353 2.1799 1.3238 1.6614 0.8964 1.1002 1.9644 1.5516 1.8046 2.3827 1.1561 2.0050 0.6376 0.7744 2.2607 1.2048 1.0904 1.4224 1.7166 1.5344 1.2075 1.4107 1.8062 0.7307 1.8426 2.5953 2.1397 1.6517 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 indole-3-carboxaldehyde 91.05 10256 indole-3-propionic acid 130.09 3744 indole-3-lactic acid 130.09 92904 kynurenine 145.99 846 METABOLITES_END #END