#METABOLOMICS WORKBENCH xin_hu_emory_20210507_074434_mwtab.txt DATATRACK_ID:2629 STUDY_ID:ST001804 ANALYSIS_ID:AN002926 PROJECT_ID:000000 VERSION 1 CREATED_ON May 7, 2021, 7:51 am #PROJECT PR:PROJECT_TITLE A scalable workflow for the human exposome PR:PROJECT_TYPE Untargeted GC-MS quantitative analysis PR:PROJECT_SUMMARY Complementing the genome with an understanding of the human exposome is an PR:PROJECT_SUMMARY important challenge for contemporary science and technology. Tens of thousands PR:PROJECT_SUMMARY of chemicals are used in commerce, yet cost for targeted environmental chemical PR:PROJECT_SUMMARY analysis limits surveillance to a few hundred known hazards. To overcome PR:PROJECT_SUMMARY limitations which prevent scaling to thousands of chemicals, we developed a PR:PROJECT_SUMMARY single-step express liquid extraction (XLE), gas chromatography high-resolution PR:PROJECT_SUMMARY mass spectrometry (GC-HRMS) analysis and computational pipeline to PR:PROJECT_SUMMARY operationalize the human exposome. We show that the workflow supports PR:PROJECT_SUMMARY quantification of environmental chemicals in human plasma (200 µL) and tissue PR:PROJECT_SUMMARY (≤ 100 mg) samples. The method also provides high resolution, sensitivity and PR:PROJECT_SUMMARY selectivity for exposome epidemiology of mass spectral features without a priori PR:PROJECT_SUMMARY knowledge of chemical identity. The simplicity of the method can facilitate PR:PROJECT_SUMMARY harmonization of environmental biomonitoring between laboratories and enable PR:PROJECT_SUMMARY population level human exposome research with limited sample volume. PR:INSTITUTE Emory University PR:DEPARTMENT Medicine/Pulmonary PR:LABORATORY Dean Jones PR:LAST_NAME Hu PR:FIRST_NAME Xin PR:ADDRESS Emory University Whitehead building (Rm 225), 615 Michael Street, Atlanta, PR:ADDRESS Georgia, 30322, USA PR:EMAIL xin.hu2@emory.edu PR:PHONE 4047275091 PR:FUNDING_SOURCE This study was supported by the NIEHS, U2C ES030163 (DPJ), U2C ES030859 (DIW) PR:FUNDING_SOURCE and P30 ES019776 (CJM), NIDDK RC2 DK118619 (KNL), NHLBI R01 HL086773 (DPJ), US PR:FUNDING_SOURCE Department of Defense W81XWH2010103 (DPJ), and the Chris M. Carlos and Catharine PR:FUNDING_SOURCE Nicole Jockisch Carlos Endowment Fund in Primary Sclerosing Cholangitis (PSC) PR:FUNDING_SOURCE (KNL). PR:CONTRIBUTORS Xin Hu, Douglas I. Walker, Yongliang Liang, M. Ryan Smith, Michael L. Orr, Brian PR:CONTRIBUTORS D. Juran, Chunyu Ma, Karan Uppal, Michael Koval, Greg S. Martin, David C. PR:CONTRIBUTORS Neujahr, Carmen J. Marsit, Young-Mi Go, Kurt Pennell, Gary W. Miller, PR:CONTRIBUTORS Konstantinos N. Lazaridis, Dean P. Jones #STUDY ST:STUDY_TITLE Human stool exposomics analysis ST:STUDY_TYPE Untargeted MS anlaysis ST:STUDY_SUMMARY Human stool samples, as a noninvasive matrix, have unique value in exposome ST:STUDY_SUMMARY research but have not been extensively studied for environmental chemical ST:STUDY_SUMMARY exposures. For lipophilic and unabsorbed dietary environmental chemicals, stool ST:STUDY_SUMMARY is a primary route of elimination and can therefore provide useful information ST:STUDY_SUMMARY on body burden and clearance of chemicals. In a pilot analysis of six human ST:STUDY_SUMMARY stool samples, we detected 52 and quantified 21 environmental chemicals, with ST:STUDY_SUMMARY HCB found in all samples. Quantification of HCB showed a median concentration of ST:STUDY_SUMMARY 0.057 ng/g. HCA of correlation matrix showed co-exposures of chemicals are ST:STUDY_SUMMARY likely as shown in the plasma, lung and thyroid. The high correlations of these ST:STUDY_SUMMARY persistent chemicals are not surprising as they likely derive from similar ST:STUDY_SUMMARY environmental exposure events. ST:INSTITUTE Emory University ST:DEPARTMENT Medicine/Pulmonary ST:LABORATORY Dean Jones ST:LAST_NAME Hu ST:FIRST_NAME Xin ST:ADDRESS Emory University Whitehead building (Rm 225), 615 Michael Street ST:EMAIL xin.hu2@emory.edu ST:PHONE 4047275091 #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 6097001_1 BL_12172019_001 type:stool RAW_FILE_NAME=BL_12172019_001.mzXML SUBJECT_SAMPLE_FACTORS 6097001_2 BL_12172019_002 type:stool RAW_FILE_NAME=BL_12172019_002.mzXML SUBJECT_SAMPLE_FACTORS 6879001_1 BL_12172019_003 type:stool RAW_FILE_NAME=BL_12172019_003.mzXML SUBJECT_SAMPLE_FACTORS 6879001_2 BL_12172019_004 type:stool RAW_FILE_NAME=BL_12172019_004.mzXML SUBJECT_SAMPLE_FACTORS 6081001_1 BL_12172019_005 type:stool RAW_FILE_NAME=BL_12172019_005.mzXML SUBJECT_SAMPLE_FACTORS 6081001_2 BL_12172019_006 type:stool RAW_FILE_NAME=BL_12172019_006.mzXML SUBJECT_SAMPLE_FACTORS 5531001_1 BL_12172019_007 type:stool RAW_FILE_NAME=BL_12172019_007.mzXML SUBJECT_SAMPLE_FACTORS 5531001_2 BL_12172019_008 type:stool RAW_FILE_NAME=BL_12172019_008.mzXML SUBJECT_SAMPLE_FACTORS 6157001_1 BL_12172019_009 type:stool RAW_FILE_NAME=BL_12172019_009.mzXML SUBJECT_SAMPLE_FACTORS 6157001_2 BL_12172019_010 type:stool RAW_FILE_NAME=BL_12172019_010.mzXML SUBJECT_SAMPLE_FACTORS 6154001_1 BL_12172019_011 type:stool RAW_FILE_NAME=BL_12172019_011.mzXML SUBJECT_SAMPLE_FACTORS 6154001_2 BL_12172019_012 type:stool RAW_FILE_NAME=BL_12172019_012.mzXML SUBJECT_SAMPLE_FACTORS - Exstd5_1019_1 type:external std RAW_FILE_NAME=Exstd5_1019_1.mzXML SUBJECT_SAMPLE_FACTORS - Exstd5_1019_2 type:external std RAW_FILE_NAME=Exstd5_1019_2.mzXML SUBJECT_SAMPLE_FACTORS - Exstd5_1019 type:external std RAW_FILE_NAME=Exstd5_1019.mzXML SUBJECT_SAMPLE_FACTORS - Exstd5_1219_1 type:external std RAW_FILE_NAME=Exstd5_1219_1.mzXML SUBJECT_SAMPLE_FACTORS - Exstd5_1219_2 type:external std RAW_FILE_NAME=Exstd5_1219_2.mzXML SUBJECT_SAMPLE_FACTORS - Exstd5_1219 type:external std RAW_FILE_NAME=Exstd5_1219.mzXML SUBJECT_SAMPLE_FACTORS - Isooctane_1 type:wash RAW_FILE_NAME=Isooctane_1.mzXML SUBJECT_SAMPLE_FACTORS - Isooctane_3 type:wash RAW_FILE_NAME=Isooctane_3.mzXML SUBJECT_SAMPLE_FACTORS - NIST1957_1 type:SRM1957 RAW_FILE_NAME=NIST1957_1.mzXML SUBJECT_SAMPLE_FACTORS - NIST1957_2 type:SRM1957 RAW_FILE_NAME=NIST1957_2.mzXML SUBJECT_SAMPLE_FACTORS - NIST1957_3 type:SRM1957 RAW_FILE_NAME=NIST1957_3.mzXML SUBJECT_SAMPLE_FACTORS - NIST1957_4 type:SRM1957 RAW_FILE_NAME=NIST1957_4.mzXML SUBJECT_SAMPLE_FACTORS - NIST1958_1 type:SRM1958 RAW_FILE_NAME=NIST1958_1.mzXML SUBJECT_SAMPLE_FACTORS - NIST1958_2 type:SRM1958 RAW_FILE_NAME=NIST1958_2.mzXML SUBJECT_SAMPLE_FACTORS - NIST1958_3 type:SRM1958 RAW_FILE_NAME=NIST1958_3.mzXML SUBJECT_SAMPLE_FACTORS - NIST1958_4 type:SRM1958 RAW_FILE_NAME=NIST1958_4.mzXML #COLLECTION CO:COLLECTION_SUMMARY Human stool samples were obtained from PROGRESS, a biobank for cholestatic liver CO:COLLECTION_SUMMARY disease (IRB 670-02; Mayo Clinic, Rochester, MN), to test suitability of XLE for CO:COLLECTION_SUMMARY environmental chemical measurements in stool of patients with primary sclerosing CO:COLLECTION_SUMMARY cholangitis (PSC). CO:SAMPLE_TYPE Feces #TREATMENT TR:TREATMENT_SUMMARY Tissue materials were processed similarly with a consistent ratio of 1:5 (sample TR:TREATMENT_SUMMARY to hexane-ethyl acetate mixture), i.e., 100 mg lung was homogenized in 300 µL TR:TREATMENT_SUMMARY water and extracted with 150 µL formic acid and 400 µL hexane-ethyl acetate TR:TREATMENT_SUMMARY mixture, while 40 mg thyroid was homogenized in 250 µL water and extracted with TR:TREATMENT_SUMMARY 50 µL formic acid and 200 µL hexane-ethyl acetate mixture. Stool samples (100 TR:TREATMENT_SUMMARY mg) were homogenized and extracted directly in 50 µL formic acid and 500 µL TR:TREATMENT_SUMMARY hexane-ethyl acetate mixture and then processed as plasma samples. The variation TR:TREATMENT_SUMMARY of 1:4 from 1:5 in lung extraction was arbitrary in consideration of the lower TR:TREATMENT_SUMMARY density of lung as an organ. The internal standards were spiked at final TR:TREATMENT_SUMMARY concentration: 1 ng/mL. The sample mixture was shaken vigorously on ice using TR:TREATMENT_SUMMARY multi-tube vortexer (VWR VX-2500) for 1 h and centrifuged at 1000 g, 4 °C for TR:TREATMENT_SUMMARY 10 min. The sample mixture was chilled during entire extraction procedure. The TR:TREATMENT_SUMMARY organic supernatant was transferred to a new tube with 25 mg MgSO4 (≥99.99% TR:TREATMENT_SUMMARY pure, Sigma-Aldrich) for testing of QuEChERS based procedure, and vortexed TR:TREATMENT_SUMMARY vigorously to remove water. After 10 min centrifugation at 1000 g, 80 µL of the TR:TREATMENT_SUMMARY final supernatant was spiked with instrumental internal standards (final TR:TREATMENT_SUMMARY concentration: 1 ng/mL) for analysis. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Same as treatment #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Samples were analyzed with three injections using GC-HRMS with a Thermo CH:CHROMATOGRAPHY_SUMMARY Scientific Q Exactive GC hybrid quadrupole Orbitrap mass spectrometer with 2 µL CH:CHROMATOGRAPHY_SUMMARY per injection. A capillary DB-5MS column (15 m × 0.25 mm × 0.25 µm film CH:CHROMATOGRAPHY_SUMMARY thickness) was used with the following temperature program: hold 75 °C for 1 CH:CHROMATOGRAPHY_SUMMARY min, 25 °C/min to 180 °C, 6 °C/min to 250 °C, 20 °C/min to 350 °C and hold CH:CHROMATOGRAPHY_SUMMARY for 5 min. The flow rate of the helium carrier gas was 1 mL/min. Ion source and CH:CHROMATOGRAPHY_SUMMARY transfer line temperatures were 250°C and 280°C, respectively. Data were CH:CHROMATOGRAPHY_SUMMARY collected from 3 to 24.37 min with positive electron ionization (EI) mode (+70 CH:CHROMATOGRAPHY_SUMMARY eV), scanning from m/z 85.0000 to 850.0000 with a resolution of 60,000. CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Thermo Trace 1310 CH:COLUMN_NAME Agilent DB5-MS (15m x 0.25mm, 0.25um) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Thermo Q Exactive Orbitrap MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:MS_COMMENTS Data were collected from 3 to 24.37 min with positive electron ionization (EI) MS:MS_COMMENTS mode (+70 eV), scanning from m/z 85.0000 to 850.0000 with a resolution of MS:MS_COMMENTS 60,000. Raw data were examined by checking signal-to-noise ratio, peak shape and MS:MS_COMMENTS spectral information for surrogate and internal standards using a 5 ppm m/z MS:MS_COMMENTS tolerance and 30 s retention time window in xCalibur Qualbrowser software. Data MS:MS_COMMENTS extraction was performed by XCMS to generate about 40,000 chemical features MS:MS_COMMENTS identified by spectral m/z and retention time. MS:MS_RESULTS_FILE ST001804_AN002926_Results.txt UNITS:raw intensity Has m/z:Yes Has RT:Yes RT units:Seconds #END