#METABOLOMICS WORKBENCH borjalanzon_20210402_110628 DATATRACK_ID:2559 STUDY_ID:ST001825 ANALYSIS_ID:AN002961 PROJECT_ID:000000 VERSION 1 CREATED_ON May 11, 2021, 11:53 am #PROJECT PR:PROJECT_TYPE Human nephropathy in CKD obese patients PR:PROJECT_SUMMARY Obesity is a global pandemic with an increase prevalence over the years. This PR:PROJECT_SUMMARY condition elevates the risk of developing cardiovascular diseases, hypertension PR:PROJECT_SUMMARY and renal pathologies, like chronic kidney disease (CKD). In the present study, PR:PROJECT_SUMMARY the metabolomic and the lipidomic profiles of CKD obese patients were analyzed PR:PROJECT_SUMMARY comparing with obese subjects without CKD. Subsequently, CKD obese patients PR:PROJECT_SUMMARY underwent bariatric surgery and the effect of surgery in the CKD progression of PR:PROJECT_SUMMARY these subjects was evaluated. Serum and urine were measured by LC-MS and GC-HRAM PR:PROJECT_SUMMARY equipment. PR:INSTITUTE University Rey Juan Carlos PR:FIRST_NAME Borja PR:ADDRESS Avenida de Atenas S/N, Alcorcón, Madrid, 28922, Spain PR:EMAIL borja.lanzon@urjc.es PR:PHONE 663692554 PR:PROJECT_TITLE Metabolomic and lipidomic profiles of CKD in obese patients in serum and urine PR:PROJECT_TITLE (part 2 of 3) PR:LAST_NAME Lanzon #STUDY ST:STUDY_TITLE Metabolomic and lipidomic profiles of CKD in obese patients in serum and urine ST:STUDY_TITLE (part 2 of 3) ST:STUDY_SUMMARY Obesity is a global pandemic with an increase prevalence over the years. This ST:STUDY_SUMMARY condition elevates the risk of developing cardiovascular diseases, hypertension ST:STUDY_SUMMARY and renal pathologies, like chronic kidney disease (CKD). In the present study, ST:STUDY_SUMMARY the metabolomic and the lipidomic profiles of CKD obese patients were analyzed ST:STUDY_SUMMARY comparing with obese subjects without CKD. Subsequently, CKD obese patients ST:STUDY_SUMMARY underwent bariatric surgery and the effect of surgery in the CKD progression of ST:STUDY_SUMMARY these subjects was evaluated. Serum and urine were measured by LC-MS and GC-HRAM ST:STUDY_SUMMARY equipment. ST:INSTITUTE University Rey Juan Carlos ST:DEPARTMENT Basics Science of Health ST:LAST_NAME Lanzon ST:FIRST_NAME Borja ST:ADDRESS Avenida de Atenas S/N, Alcorcón, Madrid, 28922, Spain ST:EMAIL borja.lanzon@urjc.es ST:NUM_GROUPS 3 ST:TOTAL_SUBJECTS 35 ST:STUDY_COMMENTS Serum GC-HRAM data: part 2 of 3. ST:STUDY_TYPE Human nephropathy in CKD obese patients ST:PHONE 663692554 #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 #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 - O1 Genotype:O RAW_FILE_NAME=O1 SUBJECT_SAMPLE_FACTORS - O2 Genotype:O RAW_FILE_NAME=O2 SUBJECT_SAMPLE_FACTORS - O3 Genotype:O RAW_FILE_NAME=O3 SUBJECT_SAMPLE_FACTORS - O4 Genotype:O RAW_FILE_NAME=O4 SUBJECT_SAMPLE_FACTORS - O5 Genotype:O RAW_FILE_NAME=O5 SUBJECT_SAMPLE_FACTORS - O6 Genotype:O RAW_FILE_NAME=O6 SUBJECT_SAMPLE_FACTORS - O7 Genotype:O RAW_FILE_NAME=O7 SUBJECT_SAMPLE_FACTORS - O8 Genotype:O RAW_FILE_NAME=O8 SUBJECT_SAMPLE_FACTORS - O9 Genotype:O RAW_FILE_NAME=O9 SUBJECT_SAMPLE_FACTORS - O10 Genotype:O RAW_FILE_NAME=O10 SUBJECT_SAMPLE_FACTORS - O11 Genotype:O RAW_FILE_NAME=O11 SUBJECT_SAMPLE_FACTORS - O12 Genotype:O RAW_FILE_NAME=O12 SUBJECT_SAMPLE_FACTORS - O13 Genotype:O RAW_FILE_NAME=O13 SUBJECT_SAMPLE_FACTORS - OD1 Genotype:OD RAW_FILE_NAME=OD1 SUBJECT_SAMPLE_FACTORS - OD2 Genotype:OD RAW_FILE_NAME=OD2 SUBJECT_SAMPLE_FACTORS - OD3 Genotype:OD RAW_FILE_NAME=OD3 SUBJECT_SAMPLE_FACTORS - OD4 Genotype:OD RAW_FILE_NAME=OD4 SUBJECT_SAMPLE_FACTORS - OD5 Genotype:OD RAW_FILE_NAME=OD5 SUBJECT_SAMPLE_FACTORS - OD6 Genotype:OD RAW_FILE_NAME=OD6 SUBJECT_SAMPLE_FACTORS - OD7 Genotype:OD RAW_FILE_NAME=OD7 SUBJECT_SAMPLE_FACTORS - OD8 Genotype:OD RAW_FILE_NAME=OD8 SUBJECT_SAMPLE_FACTORS - OD9 Genotype:OD RAW_FILE_NAME=OD9 SUBJECT_SAMPLE_FACTORS - OD10 Genotype:OD RAW_FILE_NAME=OD10 SUBJECT_SAMPLE_FACTORS - OD11 Genotype:OD RAW_FILE_NAME=OD11 SUBJECT_SAMPLE_FACTORS - ODBS1 Genotype:ODBS RAW_FILE_NAME=ODBS1 SUBJECT_SAMPLE_FACTORS - ODBS2 Genotype:ODBS RAW_FILE_NAME=ODBS2 SUBJECT_SAMPLE_FACTORS - ODBS3 Genotype:ODBS RAW_FILE_NAME=ODBS3 SUBJECT_SAMPLE_FACTORS - ODBS4 Genotype:ODBS RAW_FILE_NAME=ODBS4 SUBJECT_SAMPLE_FACTORS - ODBS5 Genotype:ODBS RAW_FILE_NAME=ODBS5 SUBJECT_SAMPLE_FACTORS - ODBS6 Genotype:ODBS RAW_FILE_NAME=ODBS6 SUBJECT_SAMPLE_FACTORS - ODBS7 Genotype:ODBS RAW_FILE_NAME=ODBS7 SUBJECT_SAMPLE_FACTORS - ODBS8 Genotype:ODBS RAW_FILE_NAME=ODBS8 SUBJECT_SAMPLE_FACTORS - ODBS9 Genotype:ODBS RAW_FILE_NAME=ODBS9 SUBJECT_SAMPLE_FACTORS - ODBS10 Genotype:ODBS RAW_FILE_NAME=ODBS10 SUBJECT_SAMPLE_FACTORS - ODBS11 Genotype:ODBS RAW_FILE_NAME=ODBS11 SUBJECT_SAMPLE_FACTORS - QC1 Genotype:QC RAW_FILE_NAME=QC1 SUBJECT_SAMPLE_FACTORS - QC2 Genotype:QC RAW_FILE_NAME=QC2 SUBJECT_SAMPLE_FACTORS - QC3 Genotype:QC RAW_FILE_NAME=QC3 SUBJECT_SAMPLE_FACTORS - QC4 Genotype:QC RAW_FILE_NAME=QC4 SUBJECT_SAMPLE_FACTORS - QC5 Genotype:QC RAW_FILE_NAME=QC5 #COLLECTION CO:COLLECTION_SUMMARY Serum samples were collected for CKD obese patients (OD), obese patients without CO:COLLECTION_SUMMARY CKD (O) and CKD obese patients who underwent bariatric surgery (OD BS). Samples CO:COLLECTION_SUMMARY were centrifuged (3500 rpm, 15 min at 4 °C), aliquoted and stored at -80 °C CO:COLLECTION_SUMMARY until extraction. CO:SAMPLE_TYPE Blood (serum) CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY 30 µl of serum of each sample were randomized and vortex-mixed with 400 µl of TR:TREATMENT_SUMMARY MeOH at -20 °C containing 1 ppm of a mix of internal standards. Samples were TR:TREATMENT_SUMMARY oximated and silylated. Samples were analyzed in a Orbitrap system. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY 30 µl of serum of each sample were randomized and vortex-mixed with 400 µl of SP:SAMPLEPREP_SUMMARY MeOH at -20 °C containing 1 ppm of the following internal standards: SP:SAMPLEPREP_SUMMARY heptadecanoic acid, valine-d8, succinic acid-d4 and glutamic acid-d5 SP:SAMPLEPREP_SUMMARY (Sigma-Aldrich). Samples were incubated on ice for 30 min and centrifuged (9600 SP:SAMPLEPREP_SUMMARY rpm, 3 min). After that, 350 µl (400 µl for urine) of the supernatant of each SP:SAMPLEPREP_SUMMARY serum sample were transferred to a V-shaped GC-vial. Stability and SP:SAMPLEPREP_SUMMARY reproducibility of the system were checked with pooled samples prepared SP:SAMPLEPREP_SUMMARY colleting from all the extracts the same quantity of the remained supernatant. SP:SAMPLEPREP_SUMMARY Afterwards, pooled samples were vortex-mixed, centrifuged and 350 µl (400 µl SP:SAMPLEPREP_SUMMARY for urine) of the supernatant of each aliquot were transferred to a V-shaped SP:SAMPLEPREP_SUMMARY GC-vial. Derivatization. Supernatants were evaporate to dryness in a nitrogen SP:SAMPLEPREP_SUMMARY flow. Then, samples were converted to trimethylsilyl (TMS) and methoxime (MEOX) SP:SAMPLEPREP_SUMMARY derivate(s). Consequently, 25 µl of MOX reagent in pyridine (20 mg/ml) were SP:SAMPLEPREP_SUMMARY added, samples were vortex-mixed and incubated for 60 min at 45 °C. After SP:SAMPLEPREP_SUMMARY oximation, silylation was performed adding 25 µl of MSTFA, samples were SP:SAMPLEPREP_SUMMARY vortex-mixed and incubated for 60 min at 45 °C. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY GC-HRAM analysis were performed in a Q Exactive GC Orbitrap system (Thermo CH:CHROMATOGRAPHY_SUMMARY Scientific), mounted with a Rxi Guard column purchased at Restek (10 m X 0.37 CH:CHROMATOGRAPHY_SUMMARY mm, 0.25 µm i.d.) and a capillary column provided by Agilent Technologies (30 CH:CHROMATOGRAPHY_SUMMARY m, 0.25 mm, 0.25 µm i.d.). Injection (1µ) was done in splitless mode with a CH:CHROMATOGRAPHY_SUMMARY TriPlus RSH autosampler system provided by Thermo. Oven temperature was keep at CH:CHROMATOGRAPHY_SUMMARY 70 °C for the first 5 minutes. Then, temperature was increased to 260 °C (10 CH:CHROMATOGRAPHY_SUMMARY °C/min) to reach in the final step 300 °C (40 °C/min) for 5 min. The carrier CH:CHROMATOGRAPHY_SUMMARY gas used was Helium with a flow of 2.0 ml/min. Scan range and resolution were CH:CHROMATOGRAPHY_SUMMARY adjusted to 50 – 500 m/z and 60,000 respectively. MS Detector was operated in CH:CHROMATOGRAPHY_SUMMARY EI positive mode. CH:CHROMATOGRAPHY_TYPE Normal phase CH:INSTRUMENT_NAME Q Exactive GC orbitrap CH:COLUMN_NAME Agilent Technologies 30 m, 0.25 mm, 0.25 µm #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Thermo Q Exactive Orbitrap MS:INSTRUMENT_TYPE Triple quadrupole MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:MS_COMMENTS GC-HRAM analysis were performed in a Q Exactive GC Orbitrap system (Thermo MS:MS_COMMENTS Scientific), mounted with a Rxi Guard column purchased at Restek (10 m X 0.37 MS:MS_COMMENTS mm, 0.25 µm i.d.) and a capillary column provided by Agilent Technologies (30 MS:MS_COMMENTS m, 0.25 mm, 0.25 µm i.d.). Injection (1µ) was done in splitless mode with a MS:MS_COMMENTS TriPlus RSH autosampler system provided by Thermo. Oven temperature was keep at MS:MS_COMMENTS 70 °C for the first 5 minutes. Then, temperature was increased to 260 °C (10 MS:MS_COMMENTS °C/min) to reach in the final step 300 °C (40 °C/min) for 5 min. The carrier MS:MS_COMMENTS gas used was Helium with a flow of 2.0 ml/min. Scan range and resolution were MS:MS_COMMENTS adjusted to 50 – 500 m/z and 60,000 respectively. MS Detector was operated in MS:MS_COMMENTS EI positive mode. The ion source and the transfer line were kept at 280 °C. MS:MS_RESULTS_FILE ST001825_AN002961_Results.txt UNITS:Area Has m/z:Yes Has RT:Yes RT units:Minutes #END