#METABOLOMICS WORKBENCH douglas_walker_20200619_083741 DATATRACK_ID:2058 STUDY_ID:ST001406 ANALYSIS_ID:AN002349 PROJECT_ID:PR000963 VERSION 1 CREATED_ON June 22, 2020, 7:12 pm #PROJECT PR:PROJECT_TITLE Environmental chemical burden in metabolic tissues and systemic biological PR:PROJECT_TITLE pathways in adolescent bariatric surgery patients: A pilot untargeted PR:PROJECT_TITLE metabolomic approach PR:PROJECT_TYPE Pilot Study PR:PROJECT_SUMMARY Background: Advances in untargeted metabolomic technologies have great potential PR:PROJECT_SUMMARY for insight into adverse metabolic effects underlying exposure to environmental PR:PROJECT_SUMMARY chemicals. However, important challenges need to be addressed, including how PR:PROJECT_SUMMARY biological response corresponds to the environmental chemical burden in PR:PROJECT_SUMMARY different target tissues. Aim: We performed a pilot study using state-of-the-art PR:PROJECT_SUMMARY ultra-high-resolution mass spectrometry (UHRMS) to characterize the burden of PR:PROJECT_SUMMARY lipophilic persistent organic pollutants (POPs) in metabolic tissues and PR:PROJECT_SUMMARY associated alterations in the plasma metabolome. Methods: We studied 11 PR:PROJECT_SUMMARY adolescents with severe obesity at the time of bariatric surgery. We measured 18 PR:PROJECT_SUMMARY POPs that can act as endocrine and metabolic disruptors (i.e. 2 dioxins, 11 PR:PROJECT_SUMMARY organochlorine compounds [OCs] and 5 polybrominated diphenyl ethers [PBDEs]) in PR:PROJECT_SUMMARY visceral and subcutaneous abdominal adipose tissue (vAT and sAT), and liver PR:PROJECT_SUMMARY samples using gas chromatography with UHRMS. Biological pathways were evaluated PR:PROJECT_SUMMARY by measuring the plasma metabolome using high-resolution metabolomics. Network PR:PROJECT_SUMMARY and pathway enrichment analysis assessed correlations between the PR:PROJECT_SUMMARY tissue-specific burden of three frequently detected POPs (i.e. PR:PROJECT_SUMMARY p,p’-dichlorodiphenyldichloroethene [DDE], hexachlorobenzene [HCB] and PR:PROJECT_SUMMARY PBDE-47) and plasma metabolic pathways. Results: Concentrations of 4 OCs and 3 PR:PROJECT_SUMMARY PBDEs were quantifiable in at least one metabolic tissue for >80% of PR:PROJECT_SUMMARY participants. All POPs had the highest median concentrations in adipose tissue, PR:PROJECT_SUMMARY especially sAT, except for PBDE-154, which had comparable average concentrations PR:PROJECT_SUMMARY across all tissues. Pathway analysis showed high correlations between PR:PROJECT_SUMMARY tissue-specific POPs and metabolic alterations in pathways of amino acid PR:PROJECT_SUMMARY metabolism, lipid and fatty acid metabolism, and carbohydrate metabolism. PR:PROJECT_SUMMARY Conclusions: Most of the measured POPs appear to accumulate preferentially in PR:PROJECT_SUMMARY adipose tissue compared to liver. Findings of plasma metabolic pathways PR:PROJECT_SUMMARY potentially associated with tissue-specific POPs concentrations merit further PR:PROJECT_SUMMARY investigation in larger populations. Keywords: persistent organic pollutants, PR:PROJECT_SUMMARY adipose tissue, liver, bariatric surgery, exposome, high-resolution metabolomics PR:INSTITUTE Icahn School of Medicine at Mount Sinai PR:DEPARTMENT Environmental Medicine and Public Health PR:LABORATORY High Resolution Exposomics Research Group PR:LAST_NAME Walker PR:FIRST_NAME Douglas PR:ADDRESS One Gustave L. Levy Place, Box 1057, New York, NY 10029 PR:EMAIL douglas.walker@mssm.edu PR:PHONE 212-241-9891 PR:FUNDING_SOURCE NIEHS: R21ES028903, R21ES029328, R21ES029681, R01ES029944, R01ES030364, PR:FUNDING_SOURCE U2CES026561, U2CES030163, P30ES023515, P30 ES019776, P30ES007048, P01ES022845, PR:FUNDING_SOURCE R01ES024946; EPA: RD-83544101 PR:PUBLICATIONS Valvi D, Walker DI, Inge T, Bartell SM, Jenkins T, Helmrath M, Ziegler TR, La PR:PUBLICATIONS Merrill MA, Eckel SP, Conti D, Liang Y, Jones DP, McConnell R, Chatzi L. (2020). PR:PUBLICATIONS Environmental chemical burden in metabolic tissues and systemic biological PR:PUBLICATIONS pathways in adolescent bariatric surgery patients: A pilot untargeted PR:PUBLICATIONS metabolomic approach. Environment International. In Press. PR:CONTRIBUTORS Valvi D, Walker DI, Inge T, Bartell SM, Jenkins T, Helmrath M, Ziegler TR, La PR:CONTRIBUTORS Merrill MA, Eckel SP, Conti D, Liang Y, Jones DP, McConnell R, Chatzi L #STUDY ST:STUDY_TITLE Environmental chemical burden in metabolic tissues and systemic biological ST:STUDY_TITLE pathways in adolescent bariatric surgery patients: A pilot untargeted ST:STUDY_TITLE metabolomic approach (part-II) ST:STUDY_TYPE Subcutaneous adipose tissue (AT); Visceral AT; Liver Tissue; Plasma ST:STUDY_SUMMARY Background: Advances in untargeted metabolomic technologies have great potential ST:STUDY_SUMMARY for insight into adverse metabolic effects underlying exposure to environmental ST:STUDY_SUMMARY chemicals. However, important challenges need to be addressed, including how ST:STUDY_SUMMARY biological response corresponds to the environmental chemical burden in ST:STUDY_SUMMARY different target tissues. Aim: We performed a pilot study using state-of-the-art ST:STUDY_SUMMARY ultra-high-resolution mass spectrometry (UHRMS) to characterize the burden of ST:STUDY_SUMMARY lipophilic persistent organic pollutants (POPs) in metabolic tissues and ST:STUDY_SUMMARY associated alterations in the plasma metabolome. Methods: We studied 11 ST:STUDY_SUMMARY adolescents with severe obesity at the time of bariatric surgery. We measured 18 ST:STUDY_SUMMARY POPs that can act as endocrine and metabolic disruptors (i.e. 2 dioxins, 11 ST:STUDY_SUMMARY organochlorine compounds [OCs] and 5 polybrominated diphenyl ethers [PBDEs]) in ST:STUDY_SUMMARY visceral and subcutaneous abdominal adipose tissue (vAT and sAT), and liver ST:STUDY_SUMMARY samples using gas chromatography with UHRMS. Biological pathways were evaluated ST:STUDY_SUMMARY by measuring the plasma metabolome using high-resolution metabolomics. Network ST:STUDY_SUMMARY and pathway enrichment analysis assessed correlations between the ST:STUDY_SUMMARY tissue-specific burden of three frequently detected POPs (i.e. ST:STUDY_SUMMARY p,p’-dichlorodiphenyldichloroethene [DDE], hexachlorobenzene [HCB] and ST:STUDY_SUMMARY PBDE-47) and plasma metabolic pathways. Results: Concentrations of 4 OCs and 3 ST:STUDY_SUMMARY PBDEs were quantifiable in at least one metabolic tissue for >80% of ST:STUDY_SUMMARY participants. All POPs had the highest median concentrations in adipose tissue, ST:STUDY_SUMMARY especially sAT, except for PBDE-154, which had comparable average concentrations ST:STUDY_SUMMARY across all tissues. Pathway analysis showed high correlations between ST:STUDY_SUMMARY tissue-specific POPs and metabolic alterations in pathways of amino acid ST:STUDY_SUMMARY metabolism, lipid and fatty acid metabolism, and carbohydrate metabolism. ST:STUDY_SUMMARY Conclusions: Most of the measured POPs appear to accumulate preferentially in ST:STUDY_SUMMARY adipose tissue compared to liver. Findings of plasma metabolic pathways ST:STUDY_SUMMARY potentially associated with tissue-specific POPs concentrations merit further ST:STUDY_SUMMARY investigation in larger populations. ST:INSTITUTE Icahn School of Medicine at Mount Sinai ST:DEPARTMENT Environmental Medicine and Public Health ST:LABORATORY High Resolution Exposomics Research Group ST:LAST_NAME Walker ST:FIRST_NAME Doug ST:ADDRESS One Gustave L. Levy Place, Box 1057, New York, NY 10029 ST:EMAIL douglas.walker@mssm.edu ST:PHONE 212-241-9891 ST:NUM_GROUPS 4 ST:TOTAL_SUBJECTS 11 ST:NUM_MALES 1 ST:NUM_FEMALES 10 ST:STUDY_COMMENTS Upload #1: Visceral and subcutaneous abdominal adipose tissue, liver tissue. ST:STUDY_COMMENTS Plasma metabolomics are in upload #2 ST:PUBLICATIONS Valvi D, Walker DI, Inge T, Bartell SM, Jenkins T, Helmrath M, Ziegler TR, La ST:PUBLICATIONS Merrill MA, Eckel SP, Conti D, Liang Y, Jones DP, McConnell R, Chatzi L. (2020). ST:PUBLICATIONS Environmental chemical burden in metabolic tissues and systemic biological ST:PUBLICATIONS pathways in adolescent bariatric surgery patients: A pilot untargeted ST:PUBLICATIONS metabolomic approach. Environment International. In Press. #SUBJECT SU:SUBJECT_TYPE Human SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 SU:AGE_OR_AGE_RANGE 11-20 years SU:GENDER Male and female #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 POTR_02 POTR_02_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.3613; RAW_FILE_NAME=DW_20180308_003;DW_20180308_004 SUBJECT_SAMPLE_FACTORS POTR_03 POTR_03_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.1314; RAW_FILE_NAME=DW_20180308_007;DW_20180308_008 SUBJECT_SAMPLE_FACTORS POTR_04 POTR_04_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.3944; RAW_FILE_NAME=DW_20180308_009;DW_20180308_010 SUBJECT_SAMPLE_FACTORS POTR_05 POTR_05_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.266; RAW_FILE_NAME=DW_20180308_011;DW_20180308_012 SUBJECT_SAMPLE_FACTORS POTR_06 POTR_06_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.2335; RAW_FILE_NAME=DW_20180308_013;DW_20180308_014 SUBJECT_SAMPLE_FACTORS POTR_07 POTR_07_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.2454; RAW_FILE_NAME=DW_20180308_015;DW_20180308_016 SUBJECT_SAMPLE_FACTORS POTR_08 POTR_08_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.2901; RAW_FILE_NAME=DW_20180308_017;DW_20180308_018 SUBJECT_SAMPLE_FACTORS POTR_09 POTR_09_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.3367; RAW_FILE_NAME=DW_20180308_019;DW_20180308_020 SUBJECT_SAMPLE_FACTORS POTR_10 POTR_10_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.6257; RAW_FILE_NAME=DW_20180308_021;DW_20180308_022 SUBJECT_SAMPLE_FACTORS POTR_11 POTR_11_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.1377; RAW_FILE_NAME=DW_20180308_023;DW_20180308_024 SUBJECT_SAMPLE_FACTORS POTR_12 POTR_12_sAT Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Batch=1; Tissue Weight (g)=0.5808; RAW_FILE_NAME=DW_20180309_005;DW_20180309_006 SUBJECT_SAMPLE_FACTORS POTR_02 POTR_02_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.7665; RAW_FILE_NAME=DW_20180309_007;DW_20180309_008 SUBJECT_SAMPLE_FACTORS POTR_03 POTR_03_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.139; RAW_FILE_NAME=DW_20180309_009;DW_20180309_010 SUBJECT_SAMPLE_FACTORS POTR_04 POTR_04_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.2564; RAW_FILE_NAME=DW_20180309_011;DW_20180309_012 SUBJECT_SAMPLE_FACTORS POTR_05 POTR_05_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.2598; RAW_FILE_NAME=DW_20180309_013;DW_20180309_014 SUBJECT_SAMPLE_FACTORS POTR_06 POTR_06_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.3776; RAW_FILE_NAME=DW_20180309_015;DW_20180309_016 SUBJECT_SAMPLE_FACTORS POTR_07 POTR_07_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.1319; RAW_FILE_NAME=DW_20180309_017;DW_20180309_018 SUBJECT_SAMPLE_FACTORS POTR_08 POTR_08_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.2847; RAW_FILE_NAME=DW_20180309_019;DW_20180309_020 SUBJECT_SAMPLE_FACTORS POTR_09 POTR_09_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.2846; RAW_FILE_NAME=DW_20180309_021;DW_20180309_022 SUBJECT_SAMPLE_FACTORS POTR_10 POTR_10_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.2978; RAW_FILE_NAME=DW_20180309_023;DW_20180309_024 SUBJECT_SAMPLE_FACTORS POTR_11 POTR_11_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.254; RAW_FILE_NAME=DW_20180309_025;DW_20180309_026 SUBJECT_SAMPLE_FACTORS POTR_12 POTR_12_vAT Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Batch=3; Tissue Weight (g)=0.1495; RAW_FILE_NAME=DW_20180312_005;DW_20180312_006 SUBJECT_SAMPLE_FACTORS POTR_02 POTR_02_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.4708; RAW_FILE_NAME=DW_20180312_007;DW_20180312_008 SUBJECT_SAMPLE_FACTORS POTR_03 POTR_03_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.1482; RAW_FILE_NAME=DW_20180312_009;DW_20180312_010 SUBJECT_SAMPLE_FACTORS POTR_04 POTR_04_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.2413; RAW_FILE_NAME=DW_20180312_011;DW_20180312_012 SUBJECT_SAMPLE_FACTORS POTR_05 POTR_05_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.38; RAW_FILE_NAME=DW_20180312_013;DW_20180312_014 SUBJECT_SAMPLE_FACTORS POTR_06 POTR_06_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.3593; RAW_FILE_NAME=DW_20180312_015;DW_20180312_016 SUBJECT_SAMPLE_FACTORS POTR_07 POTR_07_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.0918; RAW_FILE_NAME=DW_20180312_017;DW_20180312_018 SUBJECT_SAMPLE_FACTORS POTR_08 POTR_08_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.5442; RAW_FILE_NAME=DW_20180312_019;DW_20180312_020 SUBJECT_SAMPLE_FACTORS POTR_09 POTR_09_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.2081; RAW_FILE_NAME=DW_20180312_021;DW_20180312_022 SUBJECT_SAMPLE_FACTORS POTR_10 POTR_10_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.1846; RAW_FILE_NAME=DW_20180312_023;DW_20180312_024 SUBJECT_SAMPLE_FACTORS POTR_11 POTR_11_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.1493; RAW_FILE_NAME=DW_20180312_025;DW_20180312_026 SUBJECT_SAMPLE_FACTORS POTR_12 POTR_12_Liver Tissue Type:LIVER Batch=2; Tissue Weight (g)=0.1051; RAW_FILE_NAME=DW_20180312_025 #COLLECTION CO:COLLECTION_SUMMARY Eleven adolescents 12–20 years of age undergoing bariatric surgery at CO:COLLECTION_SUMMARY Cincinnati Children’s Hospital between 2006 and 2012 were offered enrollment CO:COLLECTION_SUMMARY in a prospective biospecimen repository protocol (Pediatric Obesity Tissue CO:COLLECTION_SUMMARY Repository [POTR]). Sample recruitment and other POTR features have been CO:COLLECTION_SUMMARY reported previously (Davidson et al. 2017). Intraoperatively, visceral adipose CO:COLLECTION_SUMMARY tissue (vAT) samples from the omentum, abdominal subcutaneous AT (sAT), and CO:COLLECTION_SUMMARY liver samples were obtained by the surgeon and processed immediately in an area CO:COLLECTION_SUMMARY adjacent to the operating room. All samples were snap-frozen in liquid nitrogen, CO:COLLECTION_SUMMARY then stored at −80°C. Plasma was collected pre-operatively after overnight CO:COLLECTION_SUMMARY fasting and stored at -80°C. Written informed consent was obtained from CO:COLLECTION_SUMMARY participants equal to or above 18 years old or from the parent or guardian if CO:COLLECTION_SUMMARY participants were less than 18 years old. The study was approved by the CO:COLLECTION_SUMMARY Institutional Review Board at Cincinnati Children’s Hospital. CO:SAMPLE_TYPE Adipose tissue CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY The objective of the observational study was to evaluate the relationship TR:TREATMENT_SUMMARY between adipose and liver tissue POPs and the plasma metabolome. All TR:TREATMENT_SUMMARY participants underwent bariatric surgery at the time of tissue collection. No TR:TREATMENT_SUMMARY other treatment or intervention was evaluated. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Tissue POPs concentrations were measured in vAT, sAT and liver tissues collected SP:SAMPLEPREP_SUMMARY during surgery. All tissue samples were prepared in batches of 11 study samples SP:SAMPLEPREP_SUMMARY and 3 method blanks using a modified version of the QuECHERS method described by SP:SAMPLEPREP_SUMMARY (Zamariola et al. 2017). Briefly, 0.2-0.5g of tissue was weighed, placed in an SP:SAMPLEPREP_SUMMARY amber glass vial and treated with 3.5mL of LC-MS grade water. Each sample was SP:SAMPLEPREP_SUMMARY then spiked with 50μL internal standard solution prepared in 2-proponal that SP:SAMPLEPREP_SUMMARY was designed to represent environmental chemicals with a range of physiochemical SP:SAMPLEPREP_SUMMARY properties to monitor analysis QA/QC, and included 500 ng/mL [13C6]-Anthracene, SP:SAMPLEPREP_SUMMARY [13C12]-PCB28, [DIETHYL-D10]-Chlorpyrifos, [13C12]-PCB101, [13C12]-4,4'-DDE, SP:SAMPLEPREP_SUMMARY [13C12]-PCB153, [13C12]-PCB180, [13C12]-PBDE47, [13C10]-Mirex, SP:SAMPLEPREP_SUMMARY [13C6]-cis-Permethrin, [13C12]-PBDE99 and [13C12]-PBB153. Following addition of SP:SAMPLEPREP_SUMMARY the internal standard solution, the sample was then homogenized for 1 min and SP:SAMPLEPREP_SUMMARY placed in a sonicating bath for 10 min. The resulting homogenate was transferred SP:SAMPLEPREP_SUMMARY to a 50 mL conical tube containing 10mL acetonitrile, 4000mg MgSO4 and1000mg SP:SAMPLEPREP_SUMMARY NaCl, and vortexed for 5 min. After centrifuging, a 1.5mL aliquout was SP:SAMPLEPREP_SUMMARY transferred to a cleanup tube containing 50 mg primary and secondary amine SP:SAMPLEPREP_SUMMARY exchange material (PSA), 50 mg C18 and 150 mg MgSO4, vortex-mixed for 1 min and SP:SAMPLEPREP_SUMMARY centrifuged at max speed for 5 min. From the supernatant, a 1 mL aliquot was SP:SAMPLEPREP_SUMMARY transferred to a clean, glass tube and dried completely in a vacuum centrifuge SP:SAMPLEPREP_SUMMARY operated at 35°C. The residue was then resuspended in 50μL isooctane and SP:SAMPLEPREP_SUMMARY transferred to a GC vial containing a low volume insert and capped with a Teflon SP:SAMPLEPREP_SUMMARY lined cap until analysis. SP:SAMPLEPREP_PROTOCOL_ID douglas_walker_Protocol_for_adipose_tissue_exposomics_v3_08Mar2018.pdf SP:SAMPLEPREP_PROTOCOL_FILENAME douglas_walker_Protocol_for_adipose_tissue_exposomics_v3_08Mar2018.pdf SP:PROCESSING_STORAGE_CONDITIONS Room temperature #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Tissue extracts were analyzed using a Thermo Scientific 1310 gas chromatograph CH:CHROMATOGRAPHY_SUMMARY connected to a Q Exactive GC Orbitrap GC-MS/MS ultra-high-resolution mass CH:CHROMATOGRAPHY_SUMMARY spectrometer and Triplus RSH autosampler. A 2 µL aliquot of extract was CH:CHROMATOGRAPHY_SUMMARY injected into an inlet maintained at 250ºC in pulsed split-less mode. The CH:CHROMATOGRAPHY_SUMMARY analytes were separated on an Agilent DB-5MSUI capillary column (30m length × CH:CHROMATOGRAPHY_SUMMARY 0.25mm inner diameter × 0.25µm film thickness) using high purity helium CH:CHROMATOGRAPHY_SUMMARY (99.999% purity) as the carrier gas at a constant flow rate of 1 mL/min. The CH:CHROMATOGRAPHY_SUMMARY oven temperature program consisted of an initial temperature of 100ºC for 1 CH:CHROMATOGRAPHY_SUMMARY min, increased to 180ºC at 25ºC/min; followed by a temperature ramp to 215ºC CH:CHROMATOGRAPHY_SUMMARY at 5ºC/min, and finally increased to 300ºC at 25ºC/min and held for 10 min, CH:CHROMATOGRAPHY_SUMMARY resulting in a total run time of 26.6 min. CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Thermo Trace 1310 CH:COLUMN_NAME Agilent DB5-MS (30m x 0.25mm, 0.25um) CH:FLOW_RATE 1 mL/min CH:INJECTION_TEMPERATURE 250C CH:INTERNAL_STANDARD [13C6]-Anthracene, [13C12]-PCB28, [DIETHYL-D10]-Chlorpyrifos, [13C12]-PCB101, CH:INTERNAL_STANDARD [13C12]-4,4'-DDE, [13C12]-PCB153, [13C12]-PCB180, [13C12]-PBDE47, [13C10]-Mirex, CH:INTERNAL_STANDARD [13C6]-cis-Permethrin, [13C12]-PBDE99 and [13C12]-PBB153 CH:SAMPLE_INJECTION 2 uL CH:ANALYTICAL_TIME 26.6 CH:OVEN_TEMPERATURE The oven temperature program consisted of an initial temperature of 100ºC for 1 CH:OVEN_TEMPERATURE min, increased to 180ºC at 25ºC/min; followed by a temperature ramp to 215ºC CH:OVEN_TEMPERATURE at 5ºC/min, and finally increased to 300ºC at 25ºC/min and held for 10 min CH:TRANSFERLINE_TEMPERATURE 280 CH:SAMPLE_SYRINGE_SIZE 10uL #ANALYSIS AN:ANALYSIS_TYPE MS AN:LABORATORY_NAME Clinical Biomarkers Laboratory AN:OPERATOR_NAME Bill Liang AN:ACQUISITION_DATE March 2018 AN:DATA_FORMAT .Raw #MS MS:INSTRUMENT_NAME Thermo Q Exactive GC Orbitrap GC-MS/MS MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:MS_COMMENTS Targeted peak assignment and integration was completed using TraceFinder MS:ION_SOURCE_TEMPERATURE 250C MS:IONIZATION Postive MS:IONIZATION_ENERGY -70eV #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS pg/g MS_METABOLITE_DATA_START Samples POTR_02_sAT POTR_03_sAT POTR_04_sAT POTR_05_sAT POTR_06_sAT POTR_07_sAT POTR_08_sAT POTR_09_sAT POTR_10_sAT POTR_11_sAT POTR_12_sAT POTR_02_vAT POTR_03_vAT POTR_04_vAT POTR_05_vAT POTR_06_vAT POTR_07_vAT POTR_08_vAT POTR_09_vAT POTR_10_vAT POTR_11_vAT POTR_12_vAT POTR_02_Liver POTR_03_Liver POTR_04_Liver POTR_05_Liver POTR_06_Liver POTR_07_Liver POTR_08_Liver POTR_09_Liver POTR_10_Liver POTR_11_Liver POTR_12_Liver Factors Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:INTRA-ABDOMINAL ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:SUBCUTANEOUS ADIPOSE TISSUE Tissue Type:LIVER Tissue Type:LIVER Tissue Type:LIVER Tissue Type:LIVER Tissue Type:LIVER Tissue Type:LIVER Tissue Type:LIVER Tissue Type:LIVER Tissue Type:LIVER Tissue Type:LIVER Tissue Type:LIVER PCB-52 ND ND ND ND ND ND 39.54 ND 37.75 91.13 197.52 28.40 ND 45.58 258.42 ND ND ND ND ND ND 184.78 ND ND ND ND ND ND ND 55.89 ND ND ND PBDE-28 30.86 ND ND ND ND 1028.69 60.51 86.72 131.26 ND 1010.40 89.58 ND ND ND 48.82 2051.42 ND 60.41 ND 2092.36 1190.90 ND ND ND ND ND ND ND ND ND ND ND p'p-DDE 7236.64 351.90 3532.15 5992.24 5557.92 6284.58 5303.77 5725.23 5645.20 3996.85 6263.13 9170.66 2345.33 7472.32 14144.22 4964.45 14439.87 6433.22 8091.20 11240.46 13810.68 10624.08 677.63 540.75 160.58 1042.72 433.93 396.56 455.69 476.46 403.97 1013.40 2898.82 o.p-DDE ND ND ND ND ND ND ND ND ND ND ND ND ND ND 564.74 ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND PCB-28 93.83 265.02 330.62 149.54 107.04 168.08 323.98 514.22 519.12 143.43 162.96 203.56 668.18 1133.43 1696.27 243.80 894.91 417.30 1054.62 492.93 1195.06 818.93 38.98 116.58 ND 38.95 127.53 404.42 83.49 87.44 599.10 152.82 407.05 Hexachlorobenzene 658.13 ND 693.73 835.45 1112.86 1151.36 1048.40 1150.35 887.96 952.75 713.87 730.44 392.16 2083.04 1267.13 1333.00 2059.26 1804.25 1263.66 1525.63 1288.12 2558.82 116.14 164.51 ND 160.80 195.39 207.39 37.23 80.66 508.70 170.77 277.34 PCB-77 ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND 2,3,7,8-TCDF ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND 2,3,7,8-TCDD ND 222.54 ND ND ND 134.42 91.22 ND ND ND ND ND 441.63 200.60 ND ND 544.86 ND ND ND ND ND 91.16 ND 96.45 ND 127.33 ND ND 118.69 ND ND ND PBDE-47 3611.93 263.81 2092.88 937.25 1881.23 26349.95 4261.06 2112.89 3144.95 3409.47 19397.14 5316.95 3323.21 3904.27 1764.52 1838.94 56613.64 6754.79 2658.03 7541.23 32391.98 34236.85 201.66 287.64 ND ND ND 538.29 99.14 ND ND 1532.40 5447.87 PCB-101 ND ND ND ND ND ND ND ND ND ND 208.27 ND ND ND 126.49 ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND PCB-153 325.82 ND 91.65 247.62 202.15 72.83 109.68 575.21 556.30 142.42 271.46 442.58 ND 199.43 371.10 236.16 111.74 54.40 841.42 707.61 626.09 176.66 ND 98.98 ND ND ND ND 18.42 ND ND ND 103.53 PCB-138 54.13 ND ND ND 69.48 ND ND 107.39 ND ND ND ND ND ND ND 18.74 286.77 ND 86.73 98.07 ND ND ND ND ND ND ND ND ND ND ND ND ND PCB-180 ND ND ND ND ND ND ND 303.17 388.93 161.01 ND 42.94 ND ND ND ND ND ND 252.76 815.71 81.62 ND ND ND ND ND ND ND ND ND ND ND ND PBDE-100 370.30 ND 367.04 ND 464.72 3203.87 637.15 140.04 739.21 359.35 3331.21 719.59 ND 638.13 159.82 381.79 7048.19 651.24 138.82 1468.84 5656.45 7495.14 ND ND ND ND ND ND ND ND ND ND 587.92 PBDE-99 240.81 ND 118.12 ND ND 5120.01 314.91 ND 225.83 ND 2634.26 847.22 ND 168.64 ND ND 10099.36 351.60 ND 235.99 4549.32 5030.53 ND ND ND ND ND ND ND ND ND ND 532.59 Trans-Nonachlor 554.02 ND ND 429.06 563.49 511.19 645.06 641.45 357.56 109.97 487.00 ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND Cis-Nonachlor ND ND ND ND ND ND ND ND ND ND ND 226.69 ND 251.14 650.65 222.81 1303.82 71.82 648.95 273.86 822.30 361.09 ND ND ND ND ND ND ND ND ND ND ND PCB-170 ND ND ND ND 995.81 ND ND 142.95 ND ND ND ND ND ND 2177.00 ND ND ND ND ND ND ND ND 2570.14 ND ND ND 938.04 ND ND ND ND 1869.22 PBDE-154 1096.34 5115.36 434.95 1215.47 1481.84 1809.63 957.84 830.30 151.13 2342.61 125.17 158.44 6156.07 1311.23 1235.21 891.20 6398.79 1005.09 1493.78 717.96 1506.27 2346.96 809.71 3694.14 1783.52 522.14 1123.45 2668.31 315.39 750.52 2049.19 1931.00 4652.17 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name Class mz time quantitated m/z measured retention time measured Mass error (ppm) Time error (s) PCB-52 Analyte 219.9841 672 219.9841 671.8082 -0.1214 -0.1918 PBDE-28 Analyte 245.9673 930 245.9676 929.7869 1.2002 -0.2131 p'p-DDE Analyte 246.0000 862 245.9999 862.0917 -0.4720 0.4917 o.p-DDE Analyte 246.0000 806 245.9997 806.1966 -1.0553 -0.2034 PCB-28 Analyte 255.9608 619 255.9609 619.0833 0.2297 -0.1167 Hexachlorobenzene Analyte 283.8097 494 283.8098 494.4939 0.2100 0.0939 PCB-77 Analyte 291.9188 877 291.9189 877.3900 0.2210 0.1900 2,3,7,8-TCDF Analyte 305.8978 986 305.8980 986.1893 0.8097 -0.2107 2,3,7,8-TCDD Analyte 321.8927 1007 321.8928 1007.3544 0.4296 -0.0456 PBDE-47 Analyte 325.8757 1130 325.8759 1130.4329 0.6739 0.0329 PCB-101 Analyte 325.8797 814 325.8798 813.2907 0.1590 -0.3093 PCB-153 Analyte 359.8405 957 359.8407 956.6924 0.5780 -0.3076 PCB-138 Analyte 359.8405 1003 359.8406 1002.5033 0.3279 -0.0967 PCB-180 Analyte 393.8016 1118 393.8017 1118.7848 0.3197 0.3848 PBDE-100 Analyte 403.7860 1277 403.7864 1277.4486 0.9872 0.0486 PBDE-99 Analyte 403.7860 1325 403.7863 1324.4712 0.7056 -0.3288 Trans-Nonachlor Analyte 408.7833 830 408.7834 830.2733 0.1874 -0.1267 Cis-Nonachlor Analyte 408.7833 935 408.7834 934.5798 0.1267 -0.2202 PCB-170 Analyte 459.7339 1550 459.7339 1550.2848 0.0672 -0.1152 PBDE-154 Analyte 483.6941 1442 483.6945 1441.6214 0.8020 -0.1786 METABOLITES_END #END