#METABOLOMICS WORKBENCH murra668_20240801_090052 DATATRACK_ID:5071 STUDY_ID:ST003500 ANALYSIS_ID:AN005745 PROJECT_ID:PR002146 VERSION 1 CREATED_ON August 13, 2024, 7:38 am #PROJECT PR:PROJECT_TITLE A UHPLC-MS/MS Method for Profiling of Urinary Mercapturic Acids using Positive PR:PROJECT_TITLE Ion Mode PR:PROJECT_SUMMARY We describe an analytical global profiling approach with machine learning PR:PROJECT_SUMMARY predicted structural annotations for the characterization of mercapturic acids, PR:PROJECT_SUMMARY a detoxification product of chemical environmental exposure. PR:INSTITUTE University of Minnesota PR:DEPARTMENT School of Public Health, Division of Environmental Health Sciences PR:LABORATORY Balbo Research Group PR:LAST_NAME Murray PR:FIRST_NAME Kevin PR:ADDRESS 2-210 CCRB, 2231 6th St SE, Minneapolis, MN 55455 PR:EMAIL murra668@umn.edu PR:PHONE 612-626-2182 PR:FUNDING_SOURCE National Cancer Institute (5R01CA222005-05) #STUDY ST:STUDY_TITLE A UHPLC-MS/MS Method for Profiling of Urinary Mercapturic Acids using Positive ST:STUDY_TITLE Ion Mode ST:STUDY_SUMMARY We report the first application of a UHPLC-MS/MS method using positive ion mode ST:STUDY_SUMMARY detection for the unbiased characterization of mercapturic acids. The proposed ST:STUDY_SUMMARY method utilizes a neutral loss monitoring paradigm to monitor for two diagnostic ST:STUDY_SUMMARY fragmentation pathways for this class of compound. Using a cohort of 20 ST:STUDY_SUMMARY nonsmokers and 20 smokers, we detected 180 putative mercapturic acid signatures ST:STUDY_SUMMARY that exhibited a high degree of reproducibility from the complex urine ST:STUDY_SUMMARY metabolome background. Following a combination of multivariate and univariate ST:STUDY_SUMMARY statistics, we found 33 putative mercapturic acids associated with smoking ST:STUDY_SUMMARY status. ST:INSTITUTE University of Minnesota ST:LAST_NAME Murray ST:FIRST_NAME Kevin ST:ADDRESS 2-210 CCRB, 2231 6th St SE, Minneapolis, MN 55455 ST:EMAIL murra668@umn.edu ST:PHONE 612-626-2182 ST:NUM_GROUPS 2 ST:TOTAL_SUBJECTS 20 #SUBJECT SU:SUBJECT_TYPE Human SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 SU:AGE_OR_AGE_RANGE 21-61 SU:GENDER Male and female SU:HUMAN_SMOKING_STATUS Smoking and Nonsmoking #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 79 N079 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_1; Injection order=3; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_N079 SUBJECT_SAMPLE_FACTORS 82 N082 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_2; Injection order=3; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_N082 SUBJECT_SAMPLE_FACTORS 87 N087 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_1; Injection order=8; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_N087 SUBJECT_SAMPLE_FACTORS 88 N088 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_2; Injection order=8; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_N088 SUBJECT_SAMPLE_FACTORS 94 N094 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_1; Injection order=12; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_N094 SUBJECT_SAMPLE_FACTORS 96 N096 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_2; Injection order=12; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_N096 SUBJECT_SAMPLE_FACTORS 128 N128 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_1; Injection order=17; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_N128 SUBJECT_SAMPLE_FACTORS 133 N133 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_2; Injection order=17; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_N133 SUBJECT_SAMPLE_FACTORS 134 N134 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_1; Injection order=22; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_N134 SUBJECT_SAMPLE_FACTORS 140 N140 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_2; Injection order=22; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_N140 SUBJECT_SAMPLE_FACTORS 141 N141 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_1; Injection order=24; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_N141 SUBJECT_SAMPLE_FACTORS 142 N142 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_2; Injection order=24; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_N142 SUBJECT_SAMPLE_FACTORS 149 N149 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_1; Injection order=20; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_N149 SUBJECT_SAMPLE_FACTORS 161 N161 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_2; Injection order=20; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_N161 SUBJECT_SAMPLE_FACTORS 162 N162 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_1; Injection order=15; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_N162 SUBJECT_SAMPLE_FACTORS 167 N167 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_2; Injection order=10; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_N167 SUBJECT_SAMPLE_FACTORS 189 N189 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_2; Injection order=15; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_N189 SUBJECT_SAMPLE_FACTORS 194 N194 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_1; Injection order=10; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_N194 SUBJECT_SAMPLE_FACTORS 207 N207 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_1; Injection order=5; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_N207 SUBJECT_SAMPLE_FACTORS 215 N215 Sample source:Urine | Smoking Status:Non-Smoker Batch=Batch_2; Injection order=5; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_N215 SUBJECT_SAMPLE_FACTORS pool_01 pool_01 Sample source:Urine | Smoking Status:QC Batch=Batch_1; Injection order=1; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_pool_01 SUBJECT_SAMPLE_FACTORS pool_02 pool_02 Sample source:Urine | Smoking Status:QC Batch=Batch_1; Injection order=7; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_pool_02 SUBJECT_SAMPLE_FACTORS pool_03 pool_03 Sample source:Urine | Smoking Status:QC Batch=Batch_1; Injection order=13; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_pool_03 SUBJECT_SAMPLE_FACTORS pool_04 pool_04 Sample source:Urine | Smoking Status:QC Batch=Batch_1; Injection order=19; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_pool_04 SUBJECT_SAMPLE_FACTORS pool_05 pool_05 Sample source:Urine | Smoking Status:QC Batch=Batch_1; Injection order=25; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_pool_05 SUBJECT_SAMPLE_FACTORS pool_06 pool_06 Sample source:Urine | Smoking Status:QC Batch=Batch_2; Injection order=1; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_pool_06 SUBJECT_SAMPLE_FACTORS pool_07 pool_07 Sample source:Urine | Smoking Status:QC Batch=Batch_2; Injection order=7; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_pool_07 SUBJECT_SAMPLE_FACTORS pool_08 pool_08 Sample source:Urine | Smoking Status:QC Batch=Batch_2; Injection order=13; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_pool_08 SUBJECT_SAMPLE_FACTORS pool_09 pool_09 Sample source:Urine | Smoking Status:QC Batch=Batch_2; Injection order=19; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_pool_09 SUBJECT_SAMPLE_FACTORS pool_10 pool_10 Sample source:Urine | Smoking Status:QC Batch=Batch_2; Injection order=25; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_pool_10 SUBJECT_SAMPLE_FACTORS 102 S102 Sample source:Urine | Smoking Status:Smoker Batch=Batch_1; Injection order=2; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_S102 SUBJECT_SAMPLE_FACTORS 110 S110 Sample source:Urine | Smoking Status:Smoker Batch=Batch_2; Injection order=2; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_S110 SUBJECT_SAMPLE_FACTORS 117 S117 Sample source:Urine | Smoking Status:Smoker Batch=Batch_1; Injection order=6; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_S117 SUBJECT_SAMPLE_FACTORS 131 S131 Sample source:Urine | Smoking Status:Smoker Batch=Batch_2; Injection order=6; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_S131 SUBJECT_SAMPLE_FACTORS 132 S132 Sample source:Urine | Smoking Status:Smoker Batch=Batch_1; Injection order=11; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_S132 SUBJECT_SAMPLE_FACTORS 135 S135 Sample source:Urine | Smoking Status:Smoker Batch=Batch_2; Injection order=11; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_S135 SUBJECT_SAMPLE_FACTORS 136 S136 Sample source:Urine | Smoking Status:Smoker Batch=Batch_1; Injection order=16; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_S136 SUBJECT_SAMPLE_FACTORS 138 S138 Sample source:Urine | Smoking Status:Smoker Batch=Batch_2; Injection order=16; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_S138 SUBJECT_SAMPLE_FACTORS 144 S144 Sample source:Urine | Smoking Status:Smoker Batch=Batch_1; Injection order=21; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_S144 SUBJECT_SAMPLE_FACTORS 145 S145 Sample source:Urine | Smoking Status:Smoker Batch=Batch_2; Injection order=21; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_S145 SUBJECT_SAMPLE_FACTORS 146 S146 Sample source:Urine | Smoking Status:Smoker Batch=Batch_1; Injection order=23; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_S146 SUBJECT_SAMPLE_FACTORS 147 S147 Sample source:Urine | Smoking Status:Smoker Batch=Batch_2; Injection order=23; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_S147 SUBJECT_SAMPLE_FACTORS 150 S150 Sample source:Urine | Smoking Status:Smoker Batch=Batch_1; Injection order=18; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_S150 SUBJECT_SAMPLE_FACTORS 151 S151 Sample source:Urine | Smoking Status:Smoker Batch=Batch_2; Injection order=18; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_S151 SUBJECT_SAMPLE_FACTORS 152 S152 Sample source:Urine | Smoking Status:Smoker Batch=Batch_1; Injection order=14; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_S152 SUBJECT_SAMPLE_FACTORS 156 S156 Sample source:Urine | Smoking Status:Smoker Batch=Batch_2; Injection order=14; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_S156 SUBJECT_SAMPLE_FACTORS 157 S157 Sample source:Urine | Smoking Status:Smoker Batch=Batch_1; Injection order=9; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_S157 SUBJECT_SAMPLE_FACTORS 163 S163 Sample source:Urine | Smoking Status:Smoker Batch=Batch_2; Injection order=9; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_S163 SUBJECT_SAMPLE_FACTORS 171 S171 Sample source:Urine | Smoking Status:Smoker Batch=Batch_2; Injection order=4; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230512_S171 SUBJECT_SAMPLE_FACTORS 190 S190 Sample source:Urine | Smoking Status:Smoker Batch=Batch_1; Injection order=4; RAW_FILE_NAME(Raw file name)=balbo006_mckeo093_19145_20230510_S190 #COLLECTION CO:COLLECTION_SUMMARY Adult participants were recruited as part of an ongoing study conducted at the CO:COLLECTION_SUMMARY University of Minnesota. Demographic information including gender, race, age, CO:COLLECTION_SUMMARY and tobacco usage was obtained through a questionnaire. Urine samples were CO:COLLECTION_SUMMARY collected throughout a 24-hour period and stored at -80 °C until the LC-MS/MS CO:COLLECTION_SUMMARY analysis. Urine samples were normalized by total 24-hour collection volume prior CO:COLLECTION_SUMMARY to sample preparation. Urinary creatinine levels measured using the RayBio CO:COLLECTION_SUMMARY Creatinine Assay kit (RayBiotech, Norcross, GA). This study was approved by the CO:COLLECTION_SUMMARY University of Minnesota Institutional Review Board. CO:SAMPLE_TYPE Urine CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY Urine samples were collected from non smokers and smokers. Urine samples were TR:TREATMENT_SUMMARY collected from patients over a 24-hour period. Sample were snap frozen after TR:TREATMENT_SUMMARY collection. No additional sample treatment was applied. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Urine samples (200 μL) were acidified using 40 μL of a 30% aqueous HCl SP:SAMPLEPREP_SUMMARY solution and vortexed gently. A mixture of 10 ng D5-PhMA, 20 ng D3-2CaEMA, 20 ng SP:SAMPLEPREP_SUMMARY D3-2CyEMA, 10 ng D3-2HPMA, 50 ng D3-3HPMA, and 50 ng D3-3HMPMA was added to each SP:SAMPLEPREP_SUMMARY sample. Oasis MAX mixed mode reverse phase anion exchange solid phase extraction SP:SAMPLEPREP_SUMMARY cartridges (60 mg, 60 u, 2 mL reservoir size) were obtained from Waters Corp. SP:SAMPLEPREP_SUMMARY (Milford, MA, USA). Before sample introduction, the plate was preconditioned SP:SAMPLEPREP_SUMMARY with MeOH, water, and 2% aqueous NH4OH solutions. The samples were applied and SP:SAMPLEPREP_SUMMARY washed with 0.7 mL 2% aqueous NH4OH and 0.7 mL MeOH, then dried with nitrogen SP:SAMPLEPREP_SUMMARY gas for 20 min. The plate was washed with 0.7 mL 2% aqueous formic acid before SP:SAMPLEPREP_SUMMARY sample collection. Finally, the unfractionated urine extracts were collected SP:SAMPLEPREP_SUMMARY using 0.7 mL 90% MeOH in 2% formic acid wash. The samples were transferred from SP:SAMPLEPREP_SUMMARY the 96-well collection plate to fresh 1.2 mL silanized vials and dried to SP:SAMPLEPREP_SUMMARY completeness in a SpeedVac without heat. The dried samples were stored at -80 SP:SAMPLEPREP_SUMMARY °C until ready for LC-MS/MS analysis. SP:PROCESSING_STORAGE_CONDITIONS Room temperature SP:EXTRACT_STORAGE -80℃ #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Thermo Dionex Ultimate 3000 RS CH:COLUMN_NAME Self-packed C18 column Dr. Maisch GmbH ReproSil-PUR (450 mm x 100 µm, 1.9 µm, CH:COLUMN_NAME 120 Å C18aq) CH:SOLVENT_A 99.9% water; 0.1% formic acid CH:SOLVENT_B 99.9% acetonitrile; 0.1% formic acid CH:FLOW_GRADIENT Chromatographic separation was performed using a linear gradient starting at 0% CH:FLOW_GRADIENT B and increased to 50% B at 40 min, 90% B at 60 min, and held for 8 min followed CH:FLOW_GRADIENT by a return to starting conditions. CH:FLOW_RATE 325 nL/min CH:COLUMN_TEMPERATURE 55 °C CH:METHODS_FILENAME MA_18786_hcdOT_30K_CNL-MS3-50CE_77min_20230510 CH:SAMPLE_INJECTION 1% sample equivalent load in 1 μL reconstituted using a load solvent mixture of CH:SAMPLE_INJECTION 0.1% aqueous formic acid. CH:ANALYTICAL_TIME 68 min CH:SAMPLE_LOOP_SIZE 10 µL injection loop #ANALYSIS AN:ANALYSIS_TYPE MS AN:LABORATORY_NAME Center for Metabolomics and Proteomics AN:OPERATOR_NAME Kevin Murray AN:DETECTOR_TYPE Orbitrap AN:ACQUISITION_DATE 2023-05-12 AN:DATA_FORMAT .RAW #MS MS:INSTRUMENT_NAME Thermo Orbitrap Fusion Tribrid MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS Discovery DDA-CNL-MS3 analyses of experimental samples were performed using MS:MS_COMMENTS full-scan detection followed by data-dependent MS2 acquisition and a conditional MS:MS_COMMENTS neutral loss-triggered MS3 acquisition. All analyses were performed in positive MS:MS_COMMENTS ion mode. Full-scan detection was performed using Orbitrap detection at a MS:MS_COMMENTS resolution of 120,000, AGC targeted setting of 4 × 105, a maximum ion injection MS:MS_COMMENTS time of 50 ms, and an S-Lens RF setting of 40%. Scan ranges of 170 m/z – 600 MS:MS_COMMENTS m/z were used for full-scan detection. MS2 spectra were collected using a DDA MS:MS_COMMENTS design with a 2 sec cycle time in centroid mode with a 15 sec dynamic exclusion MS:MS_COMMENTS list. Fragment spectra were acquired with quadrupole isolation of 0.8 m/z, MS:MS_COMMENTS Orbitrap detection at a resolution of 30,000, an AGC setting of 1 × 105, and a MS:MS_COMMENTS 100 ms maximum injection time. The analysis utilized HCD fragmentation at a MS:MS_COMMENTS fixed collision energy of 30%. MS3 spectra were collected using a conditional MS:MS_COMMENTS neutral loss design that acquired an additional fragmentation scan on ions MS:MS_COMMENTS exhibiting a neutral loss of 131.0582 Da or 105.0426 Da from the selected MS:MS_COMMENTS precursor. MS3 spectra were acquired with a full-scan MS isolation of 1.5 m/z MS:MS_COMMENTS and MS2 isolation of 2.0 m/z, Orbitrap detection at a resolution of 30,000, AGC MS:MS_COMMENTS setting of 2 × 105, and a 200 ms maximum injection time. The MS3 analysis MS:MS_COMMENTS utilized HCD fragmentation at a fixed collision energy of 50%. Data processing: MS:MS_COMMENTS Fragmentation filtering was performed using the DFBuilder module incorporated in MS:MS_COMMENTS the open-source software MZmine to monitor diagnostic neutral loss and fragment MS:MS_COMMENTS ions characteristic of mercapturic acid conjugates. Putative mercapturic acid MS:MS_COMMENTS detection required the observation of a neutral loss of 131.0582 Da (C5H9NO3) or MS:MS_COMMENTS 105.0426 Da (C3H7NO3) in acquired MS/MS spectrum. A complete list of the MS:MS_COMMENTS processing parameters is summarized in Table S2. Fragmentation filtering using MS:MS_COMMENTS the DFBuilder module was performed for the specified neutral loss using a 5 ppm MS:MS_COMMENTS mass tolerance and minimum fragment ion intensity of 1 x 104 with a 1% base peak MS:MS_COMMENTS threshold. A targeted chromatogram was drawn around detected precursor masses MS:MS_COMMENTS using a 1.0 min retention time window. Masses of interest were detected using MS:MS_COMMENTS the centroid detector and noise threshold of 1 x 105. Chromatogram traces were MS:MS_COMMENTS computed using the ADAP chromatogram builder module with a mass tolerance of 5 MS:MS_COMMENTS ppm. Chromatographic peaks were detected using the Local Minimum search MS:MS_COMMENTS algorithm using a 30% chromatographic threshold, 0.1 min retention time minimum MS:MS_COMMENTS search range, a minimum peak height of 1 x 105, a minimum peak top/edge ratio of MS:MS_COMMENTS 10, and minimum peak width tolerance of 0.1 min. Duplicate peaks were removed MS:MS_COMMENTS using a 0.1 min retention time tolerance. Feature lists were deisotoped using MS:MS_COMMENTS the 13C isotope filter module. All samples and pooled replicate feature lists MS:MS_COMMENTS were aligned using the Join Aligner module with a mass tolerance of 5 ppm and MS:MS_COMMENTS retention time tolerance of 0.4 min. Commonly observed in-source fragments, MS:MS_COMMENTS adducts, and multimer complexes were detected and removed from consideration. MS:MS_COMMENTS Missing peak annotations were replaced using the Gap-Filling algorithm. Acquired MS:MS_COMMENTS MS/MS spectra were matched against the NIST mass spectral library (version 2020) MS:MS_COMMENTS to remove unrelated metabolic signatures. Finally, the remaining features were MS:MS_COMMENTS exported as CSV files for downstream analysis. MS:COLLISION_ENERGY NCE 30% MS:FRAGMENTATION_METHOD HCD MS:ION_SPRAY_VOLTAGE 2.1 kV MS:IONIZATION ESI MS:MASS_ACCURACY 5 ppm MS:PRECURSOR_TYPE [M+H]+ MS:MS_RESULTS_FILE ST003500_AN005745_Results.txt UNITS:Peak heights Has m/z:Yes Has RT:Yes RT units:Minutes #END