#METABOLOMICS WORKBENCH Carol_Glez_20210601_004645 DATATRACK_ID:2661 STUDY_ID:ST001814 ANALYSIS_ID:AN002941 PROJECT_ID:PR001147 VERSION 1 CREATED_ON June 3, 2021, 12:38 am #PROJECT PR:PROJECT_TITLE Searching for prognostic biomarkers of Parkinson´s Disease development in the PR:PROJECT_TITLE Spanish EPIC cohort through a multiplatform metabolomics approach PR:PROJECT_SUMMARY The lack of knowledge about the onset and progression of Parkinson’s disease PR:PROJECT_SUMMARY (PD) hampers its early diagnosis and treatment. We used a multiplatform PR:PROJECT_SUMMARY untargeted metabolomics-based approach to uncover the biochemical remodeling PR:PROJECT_SUMMARY induced by PD in a really early and pre-symptomatic stage and unveiling early PR:PROJECT_SUMMARY potential diagnostic biomarkers. Baseline pre-clinical plasma samples (Pre-PD PR:PROJECT_SUMMARY n=39; control group n=39) were obtained from the European Prospective Study on PR:PROJECT_SUMMARY Nutrition and Cancer (EPIC) cohort, which healthy volunteers were followed for PR:PROJECT_SUMMARY around 15 years to ascertain incident PD. Our finding revealed alterations in PR:PROJECT_SUMMARY fatty acids metabolism, mitochondrial dysfunction, oxidative stress, and PR:PROJECT_SUMMARY gut-brain axis dysregulation. This study is of inestimable value since this is PR:PROJECT_SUMMARY the first study conducted with samples collected many years before the disease PR:PROJECT_SUMMARY development. PR:INSTITUTE Universidad CEU San Pablo PR:DEPARTMENT Center of Metabolomics and Bioanalysis PR:LAST_NAME Gonzalez-Riano PR:FIRST_NAME Carolina PR:ADDRESS Facultad de Farmacia, Universidad CEU San Pablo, Campus Monteprincipe, Boadilla PR:ADDRESS del Monte, Boadilla del Monte, Madrid, 28668, Spain PR:EMAIL carolina.gonzalezriano@ceu.es PR:PHONE 00 34 91 3724753 #STUDY ST:STUDY_TITLE Searching for prognostic biomarkers of Parkinson´s Disease development in the ST:STUDY_TITLE Spanish EPIC cohort through a multiplatform metabolomics approach ST:STUDY_SUMMARY The lack of knowledge about the onset and progression of Parkinson’s disease ST:STUDY_SUMMARY (PD) hampers its early diagnosis and treatment. We used a multiplatform ST:STUDY_SUMMARY untargeted metabolomics-based approach to uncover the biochemical remodeling ST:STUDY_SUMMARY induced by PD in a really early and pre-symptomatic stage and unveiling early ST:STUDY_SUMMARY potential diagnostic biomarkers. Baseline pre-clinical plasma samples (Pre-PD ST:STUDY_SUMMARY n=39; control group n=39) were obtained from the European Prospective Study on ST:STUDY_SUMMARY Nutrition and Cancer (EPIC) cohort, which healthy volunteers were followed for ST:STUDY_SUMMARY around 15 years to ascertain incident PD. Our finding revealed alterations in ST:STUDY_SUMMARY fatty acids metabolism, mitochondrial dysfunction, oxidative stress, and ST:STUDY_SUMMARY gut-brain axis dysregulation. This study is of inestimable value since this is ST:STUDY_SUMMARY the first study conducted with samples collected many years before the disease ST:STUDY_SUMMARY development. ST:INSTITUTE Universidad CEU San Pablo ST:LAST_NAME Gonzalez-Riano ST:FIRST_NAME Carolina ST:ADDRESS km 0, Universidad CEU-San Pablo Urbanización Montepríncipe. M-501, 28925 ST:ADDRESS Alcorcón ST:EMAIL carolina.gonzalezriano@ceu.es ST:PHONE 646251045 #SUBJECT SU:SUBJECT_TYPE Human SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 #FACTORS #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 Local Sample ID Sample name Factor1:Classification RAW_FILE_NAME=Raw Data file SUBJECT_SAMPLE_FACTORS 2 2 Factor1:CASE RAW_FILE_NAME=2 SUBJECT_SAMPLE_FACTORS 5 5 Factor1:CASE RAW_FILE_NAME=5 SUBJECT_SAMPLE_FACTORS 8 8 Factor1:CASE RAW_FILE_NAME=8 SUBJECT_SAMPLE_FACTORS 12 12 Factor1:CASE RAW_FILE_NAME=12 SUBJECT_SAMPLE_FACTORS 15 15 Factor1:CASE RAW_FILE_NAME=15 SUBJECT_SAMPLE_FACTORS 19 19 Factor1:CASE RAW_FILE_NAME=19 SUBJECT_SAMPLE_FACTORS 21 21 Factor1:CASE RAW_FILE_NAME=21 SUBJECT_SAMPLE_FACTORS 23 23 Factor1:CASE RAW_FILE_NAME=23 SUBJECT_SAMPLE_FACTORS 25 25 Factor1:CASE RAW_FILE_NAME=25 SUBJECT_SAMPLE_FACTORS 27 27 Factor1:CASE RAW_FILE_NAME=27 SUBJECT_SAMPLE_FACTORS 28 28 Factor1:CASE RAW_FILE_NAME=28 SUBJECT_SAMPLE_FACTORS 30 30 Factor1:CASE RAW_FILE_NAME=30 SUBJECT_SAMPLE_FACTORS 32 32 Factor1:CASE RAW_FILE_NAME=32 SUBJECT_SAMPLE_FACTORS 33 33 Factor1:CASE RAW_FILE_NAME=33 SUBJECT_SAMPLE_FACTORS 34 34 Factor1:CASE RAW_FILE_NAME=34 SUBJECT_SAMPLE_FACTORS 35 35 Factor1:CASE RAW_FILE_NAME=35 SUBJECT_SAMPLE_FACTORS 36 36 Factor1:CASE RAW_FILE_NAME=36 SUBJECT_SAMPLE_FACTORS 39 39 Factor1:CASE RAW_FILE_NAME=39 SUBJECT_SAMPLE_FACTORS 40 40 Factor1:CASE RAW_FILE_NAME=40 SUBJECT_SAMPLE_FACTORS 42 42 Factor1:CASE RAW_FILE_NAME=42 SUBJECT_SAMPLE_FACTORS 43 43 Factor1:CASE RAW_FILE_NAME=43 SUBJECT_SAMPLE_FACTORS 44 44 Factor1:CASE RAW_FILE_NAME=44 SUBJECT_SAMPLE_FACTORS 46 46 Factor1:CASE RAW_FILE_NAME=46 SUBJECT_SAMPLE_FACTORS 47 47 Factor1:CASE RAW_FILE_NAME=47 SUBJECT_SAMPLE_FACTORS 48 48 Factor1:CASE RAW_FILE_NAME=48 SUBJECT_SAMPLE_FACTORS 49 49 Factor1:CASE RAW_FILE_NAME=49 SUBJECT_SAMPLE_FACTORS 50 50 Factor1:CASE RAW_FILE_NAME=50 SUBJECT_SAMPLE_FACTORS 53 53 Factor1:CASE RAW_FILE_NAME=53 SUBJECT_SAMPLE_FACTORS 54 54 Factor1:CASE RAW_FILE_NAME=54 SUBJECT_SAMPLE_FACTORS 56 56 Factor1:CASE RAW_FILE_NAME=56 SUBJECT_SAMPLE_FACTORS 57 57 Factor1:CASE RAW_FILE_NAME=57 SUBJECT_SAMPLE_FACTORS 60 60 Factor1:CASE RAW_FILE_NAME=60 SUBJECT_SAMPLE_FACTORS 62 62 Factor1:CASE RAW_FILE_NAME=62 SUBJECT_SAMPLE_FACTORS 64 64 Factor1:CASE RAW_FILE_NAME=64 SUBJECT_SAMPLE_FACTORS 65 65 Factor1:CASE RAW_FILE_NAME=65 SUBJECT_SAMPLE_FACTORS 67 67 Factor1:CASE RAW_FILE_NAME=67 SUBJECT_SAMPLE_FACTORS 68 68 Factor1:CASE RAW_FILE_NAME=68 SUBJECT_SAMPLE_FACTORS 70 70 Factor1:CASE RAW_FILE_NAME=70 SUBJECT_SAMPLE_FACTORS 71 71 Factor1:CASE RAW_FILE_NAME=71 SUBJECT_SAMPLE_FACTORS C12A C12A Factor1:CONTROL RAW_FILE_NAME=C12A SUBJECT_SAMPLE_FACTORS C15B C15B Factor1:CONTROL RAW_FILE_NAME=C15B SUBJECT_SAMPLE_FACTORS C19B C19B Factor1:CONTROL RAW_FILE_NAME=C19B SUBJECT_SAMPLE_FACTORS C21B C21B Factor1:CONTROL RAW_FILE_NAME=C21B SUBJECT_SAMPLE_FACTORS C23A C23A Factor1:CONTROL RAW_FILE_NAME=C23A SUBJECT_SAMPLE_FACTORS C25B C25B Factor1:CONTROL RAW_FILE_NAME=C25B SUBJECT_SAMPLE_FACTORS C27A C27A Factor1:CONTROL RAW_FILE_NAME=C27A SUBJECT_SAMPLE_FACTORS C28A C28A Factor1:CONTROL RAW_FILE_NAME=C28A SUBJECT_SAMPLE_FACTORS C2A C2A Factor1:CONTROL RAW_FILE_NAME=C2A SUBJECT_SAMPLE_FACTORS C30B C30B Factor1:CONTROL RAW_FILE_NAME=C30B SUBJECT_SAMPLE_FACTORS C32B C32B Factor1:CONTROL RAW_FILE_NAME=C32B SUBJECT_SAMPLE_FACTORS C33A C33A Factor1:CONTROL RAW_FILE_NAME=C33A SUBJECT_SAMPLE_FACTORS C34B C34B Factor1:CONTROL RAW_FILE_NAME=C34B SUBJECT_SAMPLE_FACTORS C35B C35B Factor1:CONTROL RAW_FILE_NAME=C35B SUBJECT_SAMPLE_FACTORS C36A C36A Factor1:CONTROL RAW_FILE_NAME=C36A SUBJECT_SAMPLE_FACTORS C39A C39A Factor1:CONTROL RAW_FILE_NAME=C39A SUBJECT_SAMPLE_FACTORS C40B C40B Factor1:CONTROL RAW_FILE_NAME=C40B SUBJECT_SAMPLE_FACTORS C42A C42A Factor1:CONTROL RAW_FILE_NAME=C42A SUBJECT_SAMPLE_FACTORS C43A C43A Factor1:CONTROL RAW_FILE_NAME=C43A SUBJECT_SAMPLE_FACTORS C44B C44B Factor1:CONTROL RAW_FILE_NAME=C44B SUBJECT_SAMPLE_FACTORS C46B C46B Factor1:CONTROL RAW_FILE_NAME=C46B SUBJECT_SAMPLE_FACTORS C47A C47A Factor1:CONTROL RAW_FILE_NAME=C47A SUBJECT_SAMPLE_FACTORS C48A C48A Factor1:CONTROL RAW_FILE_NAME=C48A SUBJECT_SAMPLE_FACTORS C49A C49A Factor1:CONTROL RAW_FILE_NAME=C49A SUBJECT_SAMPLE_FACTORS C50B C50B Factor1:CONTROL RAW_FILE_NAME=C50B SUBJECT_SAMPLE_FACTORS C53A C53A Factor1:CONTROL RAW_FILE_NAME=C53A SUBJECT_SAMPLE_FACTORS C54A C54A Factor1:CONTROL RAW_FILE_NAME=C54A SUBJECT_SAMPLE_FACTORS C56B C56B Factor1:CONTROL RAW_FILE_NAME=C56B SUBJECT_SAMPLE_FACTORS C57A C57A Factor1:CONTROL RAW_FILE_NAME=C57A SUBJECT_SAMPLE_FACTORS C5A C5A Factor1:CONTROL RAW_FILE_NAME=C5A SUBJECT_SAMPLE_FACTORS C60A C60A Factor1:CONTROL RAW_FILE_NAME=C60A SUBJECT_SAMPLE_FACTORS C62B C62B Factor1:CONTROL RAW_FILE_NAME=C62B SUBJECT_SAMPLE_FACTORS C64B C64B Factor1:CONTROL RAW_FILE_NAME=C64B SUBJECT_SAMPLE_FACTORS C65A C65A Factor1:CONTROL RAW_FILE_NAME=C65A SUBJECT_SAMPLE_FACTORS C67B C67B Factor1:CONTROL RAW_FILE_NAME=C67B SUBJECT_SAMPLE_FACTORS C68A C68A Factor1:CONTROL RAW_FILE_NAME=C68A SUBJECT_SAMPLE_FACTORS C70A C70A Factor1:CONTROL RAW_FILE_NAME=C70A SUBJECT_SAMPLE_FACTORS C71B C71B Factor1:CONTROL RAW_FILE_NAME=C71B SUBJECT_SAMPLE_FACTORS C8A C8A Factor1:CONTROL RAW_FILE_NAME=C8A #COLLECTION CO:COLLECTION_SUMMARY On the day of the interview subjects were appointed for blood sample analysis CO:COLLECTION_SUMMARY within the next week at the primary care centre, where the interview took place, CO:COLLECTION_SUMMARY and a 30 ml blood sample were obtained at baseline. Fasting venous blood samples CO:COLLECTION_SUMMARY were drawn and immediately processed and were divided into 0.5 ml aliquots of CO:COLLECTION_SUMMARY plasma, serum, concentrated red blood cells and buffy coat, and stored in liquid CO:COLLECTION_SUMMARY nitrogen tanks at –190ºC until analysis. CO:SAMPLE_TYPE Blood (plasma) CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY Briefly, for GC-MS analysis, protein precipitation was achieved by mixing 1 TR:TREATMENT_SUMMARY volume of plasma with 3 volumes of cold (-20°C) acetonitrile, followed by TR:TREATMENT_SUMMARY methoximation with O-methoxyamine hydrochloride (15 mg/mL) in pyridine, and TR:TREATMENT_SUMMARY silylation with BSTFA:TMSC (99:1). Finally, 20 ppm of tricosane in heptane was TR:TREATMENT_SUMMARY added as internal standard (IS). For CE-MS analysis, 100 µL of plasma was mixed TR:TREATMENT_SUMMARY with 100 µL of 0.2 M formic acid containing 5% acetonitrile and 0.4 mM TR:TREATMENT_SUMMARY methionine sulfone as IS. The sample was transferred to a centrifree TR:TREATMENT_SUMMARY ultracentrifugation device (Millipore Ireland Ltd.,Carrigtohill, Ireland) with a TR:TREATMENT_SUMMARY 30 kDa protein cutoff for deproteinization through centrifugation (2000 g, 4 TR:TREATMENT_SUMMARY °C, 70 min). For LC-MS analysis, 100 µL of plasma was mixed with 300 µL of a TR:TREATMENT_SUMMARY cold mixture (-20°C) of methanol:ethanol (1:1, v/v) for deproteinization. TR:TREATMENT_SUMMARY Samples were centrifuged (13,000 g, 4°C, 20 min). After centrifugation, 100 µL TR:TREATMENT_SUMMARY of the supernatant was directly injected into the system. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY for GC-MS analysis, protein precipitation was achieved by mixing 1 volume of SP:SAMPLEPREP_SUMMARY plasma with 3 volumes of cold (-20°C) acetonitrile, followed by methoximation SP:SAMPLEPREP_SUMMARY with O-methoxyamine hydrochloride (15 mg/mL) in pyridine, and silylation with SP:SAMPLEPREP_SUMMARY BSTFA:TMSC (99:1). Finally, 20 ppm of tricosane in heptane was added as internal SP:SAMPLEPREP_SUMMARY standard (IS). For CE-MS analysis, 100 µL of plasma was mixed with 100 µL of SP:SAMPLEPREP_SUMMARY 0.2 M formic acid containing 5% acetonitrile and 0.4 mM methionine sulfone as SP:SAMPLEPREP_SUMMARY IS. The sample was transferred to a centrifree ultracentrifugation device SP:SAMPLEPREP_SUMMARY (Millipore Ireland Ltd.,Carrigtohill, Ireland) with a 30 kDa protein cutoff for SP:SAMPLEPREP_SUMMARY deproteinization through centrifugation (2000 g, 4 °C, 70 min). For LC-MS SP:SAMPLEPREP_SUMMARY analysis, 100 µL of plasma was mixed with 300 µL of a cold mixture (-20°C) of SP:SAMPLEPREP_SUMMARY methanol:ethanol (1:1, v/v) for deproteinization. Samples were centrifuged SP:SAMPLEPREP_SUMMARY (13,000 g, 4°C, 20 min). After centrifugation, 100 µL of the supernatant was SP:SAMPLEPREP_SUMMARY directly injected into the system. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY The analysis of the samples was accomplished using an UHPLC system (1200 CH:CHROMATOGRAPHY_SUMMARY Infinity system, Agilent Technologies, Waldbronn, Germany), coupled to a 6520 CH:CHROMATOGRAPHY_SUMMARY QTOF MS (Agilent Technologies) with an ESI ion source. The sample injection CH:CHROMATOGRAPHY_SUMMARY volume was set up to 10 μL. The separation was achieved using a Discovery® HS CH:CHROMATOGRAPHY_SUMMARY C18 15cm x 2.1 mm, 3 µm (Supelco analytical) reverse phase column at CH:CHROMATOGRAPHY_SUMMARY thermostated 40 °C. CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Agilent 1200 CH:COLUMN_NAME Phenomenex Kinetex C18 (150 x 2.1mm, 2.6 um) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Agilent 6520 QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE NEGATIVE MS:MS_COMMENTS Agilent Technologies MassHunter WorkStation Data Acquisition B.08.00 software; MS:MS_COMMENTS Agilent Technologies MassHunter Profinder B.08.00 SP1 software (Waldbronn, MS:MS_COMMENTS Germany) MS:MS_RESULTS_FILE ST001814_AN002941_Results.txt UNITS:AREA Has m/z:Neutral masses Has RT:Yes RT units:Minutes #END