#METABOLOMICS WORKBENCH borjalanzon_20210819_092939 DATATRACK_ID:2805 STUDY_ID:ST001943 ANALYSIS_ID:AN003187 PROJECT_ID:000000 VERSION 1 CREATED_ON September 2, 2021, 4:48 pm #PROJECT PR:PROJECT_TITLE Urinary signature of chronic kidney disease in patients with severe obesity by PR:PROJECT_TITLE CE-MS PR:PROJECT_TYPE Human nephropathy in CKD obese patients PR:PROJECT_SUMMARY Urine metabolomic characterization of severely obese patients with and without PR:PROJECT_SUMMARY chronic kidney disease (CKD) by CE-MS. Analysis was performed in patients before PR:PROJECT_SUMMARY and after bariatric surgery. In the present studio, samples obtained from CKD PR:PROJECT_SUMMARY patients with severe obesity before bariatric surgery will be referred as OD PR:PROJECT_SUMMARY (obese disease). Samples obtained from CKD patients with severe obesity after PR:PROJECT_SUMMARY bariatric surgery will be referred as ODBS (Obese disease bariatric surgery). PR:PROJECT_SUMMARY Patients with severe obesity without CKD will be referenced as O (obese) and PR:PROJECT_SUMMARY after BS, OBS (obese bariatric surgery). In healthy group, when results refer to PR:PROJECT_SUMMARY the first void urine samples, the acronym will be Healthy1V, and the acronym for PR:PROJECT_SUMMARY urine samples collected at 24-hour will be Healthy24h. PR:INSTITUTE University Rey Juan Carlos PR:DEPARTMENT Basics Science of Health PR:LABORATORY LAFEMEX PR:LAST_NAME Lanzon PR:FIRST_NAME Borja PR:ADDRESS Avenida de Atenas S/N, Alcorcón, Madrid, 28922, Spain PR:EMAIL borja.lanzon@urjc.es PR:PHONE 663692554 #STUDY ST:STUDY_TITLE Urinary signature of chronic kidney disease in patients with severe obesity by ST:STUDY_TITLE CE-MS ST:STUDY_TYPE Human nephropathy in CKD obese patients ST:STUDY_SUMMARY Urine metabolomic characterization of severe obese patients with and without ST:STUDY_SUMMARY chronic kidney disease (CKD) by CE-MS. Analysis was performed in patients before ST:STUDY_SUMMARY and after bariatric surgery. In the present studio, samples obtained from CKD ST:STUDY_SUMMARY patients with severe obesity before bariatric surgery will be referred as OD ST:STUDY_SUMMARY (obese disease). Samples obtained from CKD patients with severe obesity after ST:STUDY_SUMMARY bariatric surgery will be referred as ODBS (Obese disease bariatric surgery). ST:STUDY_SUMMARY Patients with severe obesity without CKD will be referenced as O (obese) and ST:STUDY_SUMMARY after BS, OBS (obese bariatric surgery). In healthy group, when results refer to ST:STUDY_SUMMARY the first void urine samples, the acronym will be Healthy1V, and the acronym for ST:STUDY_SUMMARY urine samples collected at 24-hour will be Healthy24h. ST:INSTITUTE University Rey Juan Carlos ST:DEPARTMENT Basics Science of Health ST:LABORATORY LAFEMEX ST:LAST_NAME Lanzon ST:FIRST_NAME Borja ST:ADDRESS Avenida de Atenas S/N ST:EMAIL borja.lanzon@urjc.es ST:PHONE 663692554 ST:NUM_GROUPS 6 ST:TOTAL_SUBJECTS 27 #SUBJECT SU:SUBJECT_TYPE Human SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 SU:AGE_OR_AGE_RANGE 53 ± 15 SU:WEIGHT_OR_WEIGHT_RANGE 116 ± 25 SU:GENDER Male and female SU:HUMAN_INCLUSION_CRITERIA (i) body mass index (BMI)>35kg/m2 plus GFR 30–60ml/min and proteinuria >1 SU:HUMAN_INCLUSION_CRITERIA g/24h or GFR>60ml/min and proteinuria >2.5 g/24h despite receiving maximally SU:HUMAN_INCLUSION_CRITERIA tolerated doses of renin-angiotensin-aldosterone system (RAAS) blocker and (ii) SU:HUMAN_INCLUSION_CRITERIA BMI>40kg/m2 with a GFR>30ml/min and proteinuria >0.5 g/24h despite receiving SU:HUMAN_INCLUSION_CRITERIA maximally tolerated doses of RAAS blocker. For the renal function criterion, SU:HUMAN_INCLUSION_CRITERIA eGFR was used. The follow-up time was 24 months. SU:HUMAN_EXCLUSION_CRITERIA 1) Patients who had participated or were participating in another clinical trial SU:HUMAN_EXCLUSION_CRITERIA or had taken an experimental drug in the last 28 days. 2) Patients with renal SU:HUMAN_EXCLUSION_CRITERIA transplantation and/or chronic replacement therapy (hemodialysis and/or SU:HUMAN_EXCLUSION_CRITERIA peritoneal dialysis). 3) Subjects with poorly controlled blood pressure (SBP > SU:HUMAN_EXCLUSION_CRITERIA 170 mmHg or DBP > 110 mmHg). 4) Patients with a history of cardiovascular events SU:HUMAN_EXCLUSION_CRITERIA in the past six months. 5) Patients treated with immunosuppressants. 6) Subjects SU:HUMAN_EXCLUSION_CRITERIA with a history of renovascular disease, autoimmune diseases, cancer, drug use, SU:HUMAN_EXCLUSION_CRITERIA or obstructive uropathy. 6) Patients who did not sign the informed consent. 7) SU:HUMAN_EXCLUSION_CRITERIA Patients who were pregnant or lactating. eGFR was used to establish the renal SU:HUMAN_EXCLUSION_CRITERIA function criteria. Patients were monitored for 24 months. #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 - OD_1 Genotype:O RAW_FILE_NAME=OD_1; Acronym explanation=CKD patients with severe obesity SUBJECT_SAMPLE_FACTORS - OD_2 Genotype:O RAW_FILE_NAME=OD_2; Acronym explanation=CKD patients with severe obesity SUBJECT_SAMPLE_FACTORS - OD_3 Genotype:O RAW_FILE_NAME=OD_3; Acronym explanation=CKD patients with severe obesity SUBJECT_SAMPLE_FACTORS - OD_4 Genotype:O RAW_FILE_NAME=OD_4; Acronym explanation=CKD patients with severe obesity SUBJECT_SAMPLE_FACTORS - OD_5 Genotype:O RAW_FILE_NAME=OD_5; Acronym explanation=CKD patients with severe obesity SUBJECT_SAMPLE_FACTORS - OD_6 Genotype:O RAW_FILE_NAME=OD_6; Acronym explanation=CKD patients with severe obesity SUBJECT_SAMPLE_FACTORS - OD_7 Genotype:O RAW_FILE_NAME=OD_7; Acronym explanation=CKD patients with severe obesity SUBJECT_SAMPLE_FACTORS - OD_8 Genotype:O RAW_FILE_NAME=OD_8; Acronym explanation=CKD patients with severe obesity SUBJECT_SAMPLE_FACTORS - ODBS_1 Genotype:ODBS RAW_FILE_NAME=ODBS_1; Acronym explanation=CKD patients with severe obesity after bariatric surgery SUBJECT_SAMPLE_FACTORS - ODBS_2 Genotype:ODBS RAW_FILE_NAME=ODBS_2; Acronym explanation=CKD patients with severe obesity after bariatric surgery SUBJECT_SAMPLE_FACTORS - ODBS_3 Genotype:ODBS RAW_FILE_NAME=ODBS_3; Acronym explanation=CKD patients with severe obesity after bariatric surgery SUBJECT_SAMPLE_FACTORS - ODBS_4 Genotype:ODBS RAW_FILE_NAME=ODBS_4; Acronym explanation=CKD patients with severe obesity after bariatric surgery SUBJECT_SAMPLE_FACTORS - ODBS_5 Genotype:ODBS RAW_FILE_NAME=ODBS_5; Acronym explanation=CKD patients with severe obesity after bariatric surgery SUBJECT_SAMPLE_FACTORS - ODBS_6 Genotype:ODBS RAW_FILE_NAME=ODBS_6; Acronym explanation=CKD patients with severe obesity after bariatric surgery SUBJECT_SAMPLE_FACTORS - ODBS_7 Genotype:ODBS RAW_FILE_NAME=ODBS_7; Acronym explanation=CKD patients with severe obesity after bariatric surgery SUBJECT_SAMPLE_FACTORS - ODBS_8 Genotype:ODBS RAW_FILE_NAME=ODBS_8; Acronym explanation=CKD patients with severe obesity after bariatric surgery SUBJECT_SAMPLE_FACTORS - O_1 Genotype:O RAW_FILE_NAME=O_1; Acronym explanation=Patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - O_2 Genotype:O RAW_FILE_NAME=O_2; Acronym explanation=Patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - O_3 Genotype:O RAW_FILE_NAME=O_3; Acronym explanation=Patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - O_4 Genotype:O RAW_FILE_NAME=O_4; Acronym explanation=Patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - O_5 Genotype:O RAW_FILE_NAME=O_5; Acronym explanation=Patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - O_6 Genotype:O RAW_FILE_NAME=O_6; Acronym explanation=Patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - O_7 Genotype:O RAW_FILE_NAME=O_7; Acronym explanation=Patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - O_8 Genotype:O RAW_FILE_NAME=O_8; Acronym explanation=Patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - O_9 Genotype:O RAW_FILE_NAME=O_9; Acronym explanation=Patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - OBS_1 Genotype:OBS RAW_FILE_NAME=OBS_1; Acronym explanation=Patients with severe obesity without CKD after bariatric surgery SUBJECT_SAMPLE_FACTORS - OBS_2 Genotype:OBS RAW_FILE_NAME=OBS_2; Acronym explanation=Patients with severe obesity without CKD after bariatric surgery SUBJECT_SAMPLE_FACTORS - OBS_3 Genotype:OBS RAW_FILE_NAME=OBS_3; Acronym explanation=Patients with severe obesity without CKD after bariatric surgery SUBJECT_SAMPLE_FACTORS - OBS_4 Genotype:OBS RAW_FILE_NAME=OBS_4; Acronym explanation=Patients with severe obesity without CKD after bariatric surgery SUBJECT_SAMPLE_FACTORS - OBS_5 Genotype:OBS RAW_FILE_NAME=OBS_5; Acronym explanation=Patients with severe obesity without CKD after bariatric surgery SUBJECT_SAMPLE_FACTORS - OBS_6 Genotype:OBS RAW_FILE_NAME=OBS_6; Acronym explanation=Patients with severe obesity without CKD after bariatric surgery SUBJECT_SAMPLE_FACTORS - OBS_7 Genotype:OBS RAW_FILE_NAME=OBS_7; Acronym explanation=Patients with severe obesity without CKD after bariatric surgery SUBJECT_SAMPLE_FACTORS - OBS_8 Genotype:OBS RAW_FILE_NAME=OBS_8; Acronym explanation=Patients with severe obesity without CKD after bariatric surgery SUBJECT_SAMPLE_FACTORS - OBS_9 Genotype:OBS RAW_FILE_NAME=OBS_9; Acronym explanation=Patients with severe obesity without CKD after bariatric surgery SUBJECT_SAMPLE_FACTORS - Healthy24h_1 Genotype:Healthy24h RAW_FILE_NAME=Healthy24h_1; Acronym explanation=Healthy subjects 24h urine SUBJECT_SAMPLE_FACTORS - Healthy24h_2 Genotype:Healthy24h RAW_FILE_NAME=Healthy24h_2; Acronym explanation=Healthy subjects 24h urine SUBJECT_SAMPLE_FACTORS - Healthy24h_3 Genotype:Healthy24h RAW_FILE_NAME=Healthy24h_3; Acronym explanation=Healthy subjects 24h urine SUBJECT_SAMPLE_FACTORS - Healthy24h_4 Genotype:Healthy24h RAW_FILE_NAME=Healthy24h_4; Acronym explanation=Healthy subjects 24h urine SUBJECT_SAMPLE_FACTORS - Healthy24h_5 Genotype:Healthy24h RAW_FILE_NAME=Healthy24h_5; Acronym explanation=Healthy subjects 24h urine SUBJECT_SAMPLE_FACTORS - Healthy24h_6 Genotype:Healthy24h RAW_FILE_NAME=Healthy24h_6; Acronym explanation=Healthy subjects 24h urine SUBJECT_SAMPLE_FACTORS - Healthy24h_7 Genotype:Healthy24h RAW_FILE_NAME=Healthy24h_7; Acronym explanation=Healthy subjects 24h urine SUBJECT_SAMPLE_FACTORS - Healthy24h_8 Genotype:Healthy24h RAW_FILE_NAME=Healthy24h_8; Acronym explanation=Healthy subjects 24h urine SUBJECT_SAMPLE_FACTORS - Healthy24h_9 Genotype:Healthy24h RAW_FILE_NAME=Healthy24h_9; Acronym explanation=Healthy subjects 24h urine SUBJECT_SAMPLE_FACTORS - Healthy24h_10 Genotype:Healthy24h RAW_FILE_NAME=Healthy24h_10; Acronym explanation=Healthy subjects 24h urine SUBJECT_SAMPLE_FACTORS - Healthy1V_1 Genotype:Healthy1V RAW_FILE_NAME=Healthy1V_1; Acronym explanation=Healthy subjects first morning void urine SUBJECT_SAMPLE_FACTORS - Healthy1V_2 Genotype:Healthy1V RAW_FILE_NAME=Healthy1V_2; Acronym explanation=Healthy subjects first morning void urine SUBJECT_SAMPLE_FACTORS - Healthy1V_3 Genotype:Healthy1V RAW_FILE_NAME=Healthy1V_3; Acronym explanation=Healthy subjects first morning void urine SUBJECT_SAMPLE_FACTORS - Healthy1V_4 Genotype:Healthy1V RAW_FILE_NAME=Healthy1V_4; Acronym explanation=Healthy subjects first morning void urine SUBJECT_SAMPLE_FACTORS - Healthy1V_5 Genotype:Healthy1V RAW_FILE_NAME=Healthy1V_5; Acronym explanation=Healthy subjects first morning void urine SUBJECT_SAMPLE_FACTORS - Healthy1V_6 Genotype:Healthy1V RAW_FILE_NAME=Healthy1V_6; Acronym explanation=Healthy subjects first morning void urine SUBJECT_SAMPLE_FACTORS - Healthy1V_7 Genotype:Healthy1V RAW_FILE_NAME=Healthy1V_7; Acronym explanation=Healthy subjects first morning void urine SUBJECT_SAMPLE_FACTORS - Healthy1V_8 Genotype:Healthy1V RAW_FILE_NAME=Healthy1V_8; Acronym explanation=Healthy subjects first morning void urine SUBJECT_SAMPLE_FACTORS - Healthy1V_9 Genotype:Healthy1V RAW_FILE_NAME=Healthy1V_9; Acronym explanation=Healthy subjects first morning void urine SUBJECT_SAMPLE_FACTORS - Healthy1V_10 Genotype:Healthy1V RAW_FILE_NAME=Healthy1V_10; Acronym explanation=Healthy subjects first morning void urine SUBJECT_SAMPLE_FACTORS - QC_CKD_1 Genotype:QC_CKD RAW_FILE_NAME=QC_CKD_1; Acronym explanation=Quality controls CKD patients SUBJECT_SAMPLE_FACTORS - QC_CKD_2 Genotype:QC_CKD RAW_FILE_NAME=QC_CKD_2; Acronym explanation=Quality controls CKD patients SUBJECT_SAMPLE_FACTORS - QC_CKD_3 Genotype:QC_CKD RAW_FILE_NAME=QC_CKD_3; Acronym explanation=Quality controls CKD patients SUBJECT_SAMPLE_FACTORS - QC_CKD_4 Genotype:QC_CKD RAW_FILE_NAME=QC_CKD_4; Acronym explanation=Quality controls CKD patients SUBJECT_SAMPLE_FACTORS - QC_CKD_5 Genotype:QC_CKD RAW_FILE_NAME=QC_CKD_5; Acronym explanation=Quality controls CKD patients SUBJECT_SAMPLE_FACTORS - QC_CKD_6 Genotype:QC_CKD RAW_FILE_NAME=QC_CKD_6; Acronym explanation=Quality controls CKD patients SUBJECT_SAMPLE_FACTORS - QC_CKD_7 Genotype:QC_CKD RAW_FILE_NAME=QC_CKD_7; Acronym explanation=Quality controls CKD patients SUBJECT_SAMPLE_FACTORS - QC_CKD_8 Genotype:QC_CKD RAW_FILE_NAME=QC_CKD_8; Acronym explanation=Quality controls CKD patients SUBJECT_SAMPLE_FACTORS - QC_Control_1 Genotype:QC_Control RAW_FILE_NAME=QC_Control_1; Acronym explanation=Quality controls patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - QC_Control_2 Genotype:QC_Control RAW_FILE_NAME=QC_Control_2; Acronym explanation=Quality controls patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - QC_Control_3 Genotype:QC_Control RAW_FILE_NAME=QC_Control_3; Acronym explanation=Quality controls patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - QC_Control_4 Genotype:QC_Control RAW_FILE_NAME=QC_Control_4; Acronym explanation=Quality controls patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - QC_Control_5 Genotype:QC_Control RAW_FILE_NAME=QC_Control_5; Acronym explanation=Quality controls patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - QC_Control_6 Genotype:QC_Control RAW_FILE_NAME=QC_Control_6; Acronym explanation=Quality controls patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - QC_Control_7 Genotype:QC_Control RAW_FILE_NAME=QC_Control_7; Acronym explanation=Quality controls patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - QC_Control_8 Genotype:QC_Control RAW_FILE_NAME=QC_Control_8; Acronym explanation=Quality controls patients with severe obesity without CKD SUBJECT_SAMPLE_FACTORS - QC_CC_1 Genotype:QC_CC RAW_FILE_NAME=QC_CC_1; Acronym explanation=Quality controls patients with CKD and without CKD SUBJECT_SAMPLE_FACTORS - QC_CC_2 Genotype:QC_CC RAW_FILE_NAME=QC_CC_2; Acronym explanation=Quality controls patients with CKD and without CKD SUBJECT_SAMPLE_FACTORS - QC_CC_3 Genotype:QC_CC RAW_FILE_NAME=QC_CC_3; Acronym explanation=Quality controls patients with CKD and without CKD SUBJECT_SAMPLE_FACTORS - QC_CC_4 Genotype:QC_CC RAW_FILE_NAME=QC_CC_4; Acronym explanation=Quality controls patients with CKD and without CKD SUBJECT_SAMPLE_FACTORS - QC_CC_5 Genotype:QC_CC RAW_FILE_NAME=QC_CC_5; Acronym explanation=Quality controls patients with CKD and without CKD SUBJECT_SAMPLE_FACTORS - QC_CC_6 Genotype:QC_CC RAW_FILE_NAME=QC_CC_6; Acronym explanation=Quality controls patients with CKD and without CKD SUBJECT_SAMPLE_FACTORS - QC_CC_7 Genotype:QC_CC RAW_FILE_NAME=QC_CC_7; Acronym explanation=Quality controls patients with CKD and without CKD SUBJECT_SAMPLE_FACTORS - QC_CC_8 Genotype:QC_CC RAW_FILE_NAME=QC_CC_8; Acronym explanation=Quality controls patients with CKD and without CKD SUBJECT_SAMPLE_FACTORS - QC_Healthy_1 Genotype:QC_Healthy RAW_FILE_NAME=QC_Healthy_1; Acronym explanation=Quality controls healthy individuals SUBJECT_SAMPLE_FACTORS - QC_Healthy_2 Genotype:QC_Healthy RAW_FILE_NAME=QC_Healthy_2; Acronym explanation=Quality controls healthy individuals SUBJECT_SAMPLE_FACTORS - QC_Healthy_3 Genotype:QC_Healthy RAW_FILE_NAME=QC_Healthy_3; Acronym explanation=Quality controls healthy individuals SUBJECT_SAMPLE_FACTORS - QC_Healthy_4 Genotype:QC_Healthy RAW_FILE_NAME=QC_Healthy_4; Acronym explanation=Quality controls healthy individuals SUBJECT_SAMPLE_FACTORS - QC_Healthy_5 Genotype:QC_Healthy RAW_FILE_NAME=QC_Healthy_5; Acronym explanation=Quality controls healthy individuals SUBJECT_SAMPLE_FACTORS - QC_Healthy_6 Genotype:QC_Healthy RAW_FILE_NAME=QC_Healthy_6; Acronym explanation=Quality controls healthy individuals SUBJECT_SAMPLE_FACTORS - QC_Healthy_7 Genotype:QC_Healthy RAW_FILE_NAME=QC_Healthy_7; Acronym explanation=Quality controls healthy individuals SUBJECT_SAMPLE_FACTORS - QC_Healthy_8 Genotype:QC_Healthy RAW_FILE_NAME=QC_Healthy_8; Acronym explanation=Quality controls healthy individuals #COLLECTION CO:COLLECTION_SUMMARY First morning void and 24-hour urine samples, were collected, aliquoted and CO:COLLECTION_SUMMARY stored at -80°C until extraction. 24-h urine samples were prepared collecting CO:COLLECTION_SUMMARY all the urine after the first morning void until the first urine in the next CO:COLLECTION_SUMMARY day. CO:SAMPLE_TYPE Urine CO:COLLECTION_FREQUENCY First void and 24h urines CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY CKD patients with severe obesity and non-CKD patients with severe obesity TR:TREATMENT_SUMMARY underwent bariatric surgery. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY 100 µL of each urine sample were mixed in a 100 µL IS (internal standard) SP:SAMPLEPREP_SUMMARY mixture with 0.4 mM of methionine sulfone and 2 mM of paracetamol containing 5% SP:SAMPLEPREP_SUMMARY of acetonitrile, then, samples were vortexed. Some CKD patients showed an SP:SAMPLEPREP_SUMMARY elevated proteinuria and in order to avoid capillary collapse, a Merck Milipore SP:SAMPLEPREP_SUMMARY filters (Ultrafiltration Device with Ultracel YM-T membrane, product 4104) were SP:SAMPLEPREP_SUMMARY used. Samples mixed with IS mixture were added to filters and centrifuged (2000 SP:SAMPLEPREP_SUMMARY g, 60 min, 4 °C). Then, 90 µl of each sample was transferred to a glass vial. SP:SAMPLEPREP_SUMMARY Samples were stored at -20°C until analysis. Stability and reproducibility of SP:SAMPLEPREP_SUMMARY the system were checked with quality control (QC) samples. Four types of QC SP:SAMPLEPREP_SUMMARY samples were prepared combining 60 µl of each sample after filtration: 1. QC SP:SAMPLEPREP_SUMMARY from OD and OD BS samples (QC CKD). 2. QC from O and O BS samples (QC Control). SP:SAMPLEPREP_SUMMARY 3. QC from OD, OD BS and O and OD BS samples (QC CC). 4. QC from Healthy 1H and SP:SAMPLEPREP_SUMMARY Healthy 24H samples (QC Healthy). QC samples were vortex-mixed, transferred to a SP:SAMPLEPREP_SUMMARY vial and stored at -20°C until analysis. SP:PROCESSING_STORAGE_CONDITIONS -80℃ SP:EXTRACT_STORAGE On ice #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Analysis were performed in a capillary electrophoresis (CE) system coupled to a CH:CHROMATOGRAPHY_SUMMARY TOF-MS analyser (TOF: time-of-flight) purchased from Agilent Technologies (CE: CH:CHROMATOGRAPHY_SUMMARY 7100, MS: 6224). In coupling process, steath liquid was supplied by an ISO pump CH:CHROMATOGRAPHY_SUMMARY (Agilent 1200) to increase the volatility and compensate the volume for MS CH:CHROMATOGRAPHY_SUMMARY through an electrospray source. A fused silica capillary was used to perform CH:CHROMATOGRAPHY_SUMMARY metabolite separation (100 cm total length x 50 μm i.d. x 360 μm o.d., CH:CHROMATOGRAPHY_SUMMARY provided by Agilent Technologies). Capillary was conditioned with water, BGE and CH:CHROMATOGRAPHY_SUMMARY NaOH. Before analysis, capillary was rinsed for 5 min at 950 mbar with BGE and CH:CHROMATOGRAPHY_SUMMARY was applied a voltage of 30kV for 10 s to displace BGE ions. After conditioning, CH:CHROMATOGRAPHY_SUMMARY sample was loaded applying 50 mbar for 50 s, then, BGE was applied at 100 mbar CH:CHROMATOGRAPHY_SUMMARY for 10 s. In the capillary was applied a pressure of 25 mbar and a voltage of 30 CH:CHROMATOGRAPHY_SUMMARY kV to achieve metabolite separation. A positively charged spray is formed by a CH:CHROMATOGRAPHY_SUMMARY flow of 0.6 mL min−1 (1:100 split) when separated compounds leave the CH:CHROMATOGRAPHY_SUMMARY capillary from the auxiliary liquid system with a nebulization pressure of 10 CH:CHROMATOGRAPHY_SUMMARY psig with nitrogen and a capillary voltage of 3500 V. A hot nitrogen flow of 10 CH:CHROMATOGRAPHY_SUMMARY mL min−1 at 200 °C was applied to dry the spray. CH:CHROMATOGRAPHY_TYPE None (Direct infusion) CH:INSTRUMENT_NAME none CH:COLUMN_NAME none #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Agilent 6224 TOF MS:INSTRUMENT_TYPE CE-TOF-MS MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:MS_COMMENTS Ions in gaseous state were directed to the TOF using two different voltages for MS:MS_COMMENTS the fragmentor (125 V and 200 V) and voltages of 65 V and 750 V for the skimmer MS:MS_COMMENTS and the octopole, respectively. Mass range was set from 70 to 1050 Da at a MS:MS_COMMENTS scanning rate of 1.02 scans per second. Data was collected in ESI positive ion MS:MS_COMMENTS mode. MassHunter Workstation version B.06.01 (Agilent Technologies) was used to MS:MS_COMMENTS monitor CE-MS analysis. Voltage of 125 V in the fragmentor was applied to MS:MS_COMMENTS ordinary samples and QC control samples. Data acquired at 125 V, equivalent to MS:MS_COMMENTS MS1, provided information about adducts, isotopes, multimers and fragments ions MS:MS_COMMENTS with low intensity. Voltage of 200 V in the fragmentor was applied to four QC MS:MS_COMMENTS (covering the four types of QC prepared). Data acquired at 200 V is similar to MS:MS_COMMENTS data obtained in 10 eV MS/MS, although has been previously reported that some MS:MS_COMMENTS fragment ions could differ. An equilibration QC was prepared to adapt and MS:MS_COMMENTS prepare the equipment for the conditions of the samples to be analysed. MS:MS_COMMENTS Equilibration QC was prepared as a pool of the four types of QC used to cover MS:MS_COMMENTS completely the biological diversity of the samples. Equilibration QC was MS:MS_COMMENTS analysed before samples. QC samples were disposed throughout the instrumental MS:MS_COMMENTS analysis. MS:MS_RESULTS_FILE ST001943_AN003187_Results.txt UNITS:Area Has m/z:Neutral masses Has RT:Yes RT units:Minutes #END