#METABOLOMICS WORKBENCH sbhattacharya_20191114_130654 DATATRACK_ID:1846 STUDY_ID:ST001277 ANALYSIS_ID:AN002118 PROJECT_ID:PR000862 VERSION 1 CREATED_ON November 18, 2019, 2:47 pm #PROJECT PR:PROJECT_TITLE Pseudoexfoliation Syndrome Humor Metabolites from Veterans Affairs Patients PR:PROJECT_TYPE untargeted LC-MS/MS IROA metabolomics PR:PROJECT_SUMMARY We performed high-performance liquid chromatography-tandem mass spectrometry PR:PROJECT_SUMMARY (LC-MS/MS) Isotopic Ratio Outlier Analysis of aqueous humor samples from PR:PROJECT_SUMMARY patients with pseudoexfoliation syndrome (PEX). PR:INSTITUTE University of Miami PR:DEPARTMENT Ophthalmology, Bascom Palmer Eye Institute PR:LAST_NAME Bhattacharya PR:FIRST_NAME Sanjoy PR:ADDRESS 1638 NW 10th Avenue, Miami, Florida, 33136, USA PR:EMAIL sbhattacharya@med.miami.edu PR:PHONE 3054824103 PR:CONTRIBUTORS Leila Abdelrahman #STUDY ST:STUDY_TITLE Pseudoexfoliation Aqueous Humor Metabolites from Veterans Affairs Patients ST:STUDY_SUMMARY We performed high-performance liquid chromatography-tandem mass spectrometry ST:STUDY_SUMMARY (LC-MS/MS) Isotopic Ratio Outlier Analysis of aqueous humor samples from ST:STUDY_SUMMARY patients from patients with pseudoefoliation syndrome (PEX). ST:INSTITUTE University of Miami ST:DEPARTMENT Ophthalmology, Bascom Palmer Eye Institute ST:LABORATORY Sanjoy K. Bhattacharya ST:LAST_NAME Bhattacharya ST:FIRST_NAME Sanjoy ST:ADDRESS 1638 NW 10th Avenue, Miami, FL 33136 ST:EMAIL SBhattacharya@med.miami.edu ST:PHONE 305-482-4103 ST:NUM_GROUPS 1 ST:TOTAL_SUBJECTS 31 #SUBJECT SU:SUBJECT_TYPE Human SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 SU:GENDER Male and female #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS - S001_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X145_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S002_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X154_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S003_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X167_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S004_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X187_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S005_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X188_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S006_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X197_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S007_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X200_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S008_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X219_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S009_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X336_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0010_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X352_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0011_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X359_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0012_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X362_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0013_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X379_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0014_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X406_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0015_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X413_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0016_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X418_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0017_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X423_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0018_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X425_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0019_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X449_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0020_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X459_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0021_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X464_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0022_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X483_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0023_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X485_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0024_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X559_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0025_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X561_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0026_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X562_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0027_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X579_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0028_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X586_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0029_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X598_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0030_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X653_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S0031_NEG Treatment1:PXE Acquisition method=PXE_NEG; Data file=X655_neg_IS.raw SUBJECT_SAMPLE_FACTORS - S001_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X145_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S002_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X154_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S003_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X167_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S004_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X187_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S005_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X188_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S006_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X197_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S007_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X200_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S008_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X219_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S009_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X336_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0010_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X352_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0011_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X359_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0012_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X362_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0013_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X379_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0014_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X406_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0015_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X413_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0016_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X418_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0017_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X423_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0018_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X425_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0019_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X449_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0020_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X459_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0021_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X464_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0022_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X483_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0023_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X485_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0024_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X559_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0025_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X561_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0026_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X562_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0027_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X579_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0028_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X586_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0029_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X598_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0030_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X653_pos_IS.raw SUBJECT_SAMPLE_FACTORS - S0031_POS Treatment1:PXE Acquisition method=PXE_POS; Data file=X655_pos_IS.raw #COLLECTION CO:COLLECTION_SUMMARY All materials were collected from human donors without identifiers under CO:COLLECTION_SUMMARY institutional review board exemption/approval. We acquired 31 aqueous humor (AH) CO:COLLECTION_SUMMARY samples from participants at the Veterans Administration (VA) Medical Center CO:COLLECTION_SUMMARY (Miami, FL). Patient samples used for Isotopic Ratio Outlier Analysis (IROA) CO:COLLECTION_SUMMARY consisted of individuals with PEX (n=31). CO:SAMPLE_TYPE Eye tissue CO:COLLECTION_LOCATION 1201 NW 16th St, Miami, FL 33125 #TREATMENT TR:TREATMENT_SUMMARY No Treatment #SAMPLEPREP SP:SAMPLEPREP_SUMMARY The 31 samples were prepared for isotopic ratio outlier analysis (IROA). SP:SAMPLEPREP_SUMMARY Approximately 800 µL of precipitate solution (8:1:1 Acetonitrile: Methanol: SP:SAMPLEPREP_SUMMARY Acetone) was added to the 50 µL of aqueous humor. The metabolites were vortexed SP:SAMPLEPREP_SUMMARY and incubated at 4 °C for 30 minutes. This was followed by another incubation SP:SAMPLEPREP_SUMMARY period at -20 °C for 1 hour. Each sample was centrifuged at 20,000 x g for 10 SP:SAMPLEPREP_SUMMARY minutes at 4 °C (Beckman Microfuge 18) to form a pellet and 375 µL of SP:SAMPLEPREP_SUMMARY supernatant was collected. The supernatant was dried in a speed vacuum for SP:SAMPLEPREP_SUMMARY approximately 20 minutes or until the sample was fully dry and stored at -20 SP:SAMPLEPREP_SUMMARY °C. The IROA internal standard (IROA-IS, IROA technologies) (U-95% 13C) was SP:SAMPLEPREP_SUMMARY reconstituted in 1.2 mL of LC-MS grade water. The samples were reconstituted in SP:SAMPLEPREP_SUMMARY 25 µL of LC-MS grade water and 10 µL was combined with 20 µL of IROA-IS and SP:SAMPLEPREP_SUMMARY subjected to LC-MS/MS analysis. A long-term reference standard (LTRS) was SP:SAMPLEPREP_SUMMARY reconstituted in 50 µL of LC-MS grade water and subjected to LC-MS/MS as the SP:SAMPLEPREP_SUMMARY first, last, and every 10 samples. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Thermo Accela 600 CH:COLUMN_NAME ACE Excel 2 C18-PFP (100 x 2.1mm, 2um) CH:FLOW_RATE 350 ul/min CH:COLUMN_TEMPERATURE 40 C CH:SOLVENT_A Water with 0.1% formic acid (FA) CH:SOLVENT_B Acetonitrile with 0.1 % FA #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Thermo Q Exactive Orbitrap MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE ESI MS:ION_MODE UNSPECIFIED MS:MS_COMMENTS Samples were ran in positive and negative ion mode Data dependent MS/MS MS:MS_RESULTS_FILE ST001277_AN002118_Results.txt UNITS:C12:C13 Ratio Has m/z:Yes Has RT:Yes RT units:Minutes #END