#METABOLOMICS WORKBENCH yorch40_20210601_033941 DATATRACK_ID:2662 STUDY_ID:ST001810 ANALYSIS_ID:AN002934 PROJECT_ID:PR001144 VERSION 1 CREATED_ON June 1, 2021, 7:48 am #PROJECT PR:PROJECT_TITLE Metabolomics-driven identification of biochemical mechanisms underlying the PR:PROJECT_TITLE neuroprotective effects of pleiotrophin in a mouse model of Parkinson’s PR:PROJECT_TITLE disease PR:PROJECT_SUMMARY Pleiotrophin (PTN) is a cytokine involved in nerve tissue repair processes, PR:PROJECT_SUMMARY neuroinflammation and neuronal survival. PTN expression levels are upregulated PR:PROJECT_SUMMARY in the nigrostriatal pathway of Parkinson’s Disease (PD) patients. We aimed to PR:PROJECT_SUMMARY characterize the dopaminergic injury and glial activation in the nigrostriatal PR:PROJECT_SUMMARY pathway of mice with transgenic Ptn overexpression in the brain (Ptn-Tg) after PR:PROJECT_SUMMARY intrastriatal injection of the Parkinsonian toxin 6-hydroxydopamine (6-OHDA). PR:PROJECT_SUMMARY The injection of 6-OHDA induced a significant decrease of the number of tyrosine PR:PROJECT_SUMMARY hydroxylase (TH)-positive neurons in the substantia nigra and of the striatal TH PR:PROJECT_SUMMARY contents in Wild type (Wt) mice. In contrast, these effects of 6-OHDA were PR:PROJECT_SUMMARY blocked in Ptn-Tg mice. 6-OHDA injection did not cause robust changes in PR:PROJECT_SUMMARY microglia but induced an exacerbated astrocytic response in Wt mice compared PR:PROJECT_SUMMARY with Ptn-Tg mice. In metabolomics studies, we detected interesting metabolites PR:PROJECT_SUMMARY that significantly discriminate the more injured 6-OHDA-injected Wt striatum and PR:PROJECT_SUMMARY the more protected 6-OHDA-injected Ptn-Tg striatum. Particularly, we detected PR:PROJECT_SUMMARY groups of metabolites, mostly corresponding to phospholipids, whose trends were PR:PROJECT_SUMMARY opposite in both groups. In summary, the data confirm the neuroprotective effect PR:PROJECT_SUMMARY of brain PTN overexpression in this mouse model of PD. New lipid-related PD drug PR:PROJECT_SUMMARY candidates emerge from this study and the data presented here support the PR:PROJECT_SUMMARY increasingly recognized “lipid cascade” in PD. PR:INSTITUTE CEU San Pablo University PR:LAST_NAME Sáiz PR:FIRST_NAME Jorge PR:ADDRESS JULIÁN ROMEA 23, Madrid, Madrid, 28003, Spain PR:EMAIL jorge.saizgalindo@ceu.es PR:PHONE 913 72 47 11 #STUDY ST:STUDY_TITLE Metabolomics-driven identification of biochemical mechanisms underlying the ST:STUDY_TITLE neuroprotective effects of pleiotrophin in a mouse model of Parkinson’s ST:STUDY_TITLE disease ST:STUDY_SUMMARY Pleiotrophin (PTN) is a cytokine involved in nerve tissue repair processes, ST:STUDY_SUMMARY neuroinflammation and neuronal survival. PTN expression levels are upregulated ST:STUDY_SUMMARY in the nigrostriatal pathway of Parkinson’s Disease (PD) patients. We aimed to ST:STUDY_SUMMARY characterize the dopaminergic injury and glial activation in the nigrostriatal ST:STUDY_SUMMARY pathway of mice with transgenic Ptn overexpression in the brain (Ptn-Tg) after ST:STUDY_SUMMARY intrastriatal injection of the Parkinsonian toxin 6-hydroxydopamine (6-OHDA). ST:STUDY_SUMMARY The injection of 6-OHDA induced a significant decrease of the number of tyrosine ST:STUDY_SUMMARY hydroxylase (TH)-positive neurons in the substantia nigra and of the striatal TH ST:STUDY_SUMMARY contents in Wild type (Wt) mice. In contrast, these effects of 6-OHDA were ST:STUDY_SUMMARY blocked in Ptn-Tg mice. 6-OHDA injection did not cause robust changes in ST:STUDY_SUMMARY microglia but induced an exacerbated astrocytic response in Wt mice compared ST:STUDY_SUMMARY with Ptn-Tg mice. In metabolomics studies, we detected interesting metabolites ST:STUDY_SUMMARY that significantly discriminate the more injured 6-OHDA-injected Wt striatum and ST:STUDY_SUMMARY the more protected 6-OHDA-injected Ptn-Tg striatum. Particularly, we detected ST:STUDY_SUMMARY groups of metabolites, mostly corresponding to phospholipids, whose trends were ST:STUDY_SUMMARY opposite in both groups. In summary, the data confirm the neuroprotective effect ST:STUDY_SUMMARY of brain PTN overexpression in this mouse model of PD. New lipid-related PD drug ST:STUDY_SUMMARY candidates emerge from this study and the data presented here support the ST:STUDY_SUMMARY increasingly recognized “lipid cascade” in PD. ST:INSTITUTE CEMBIO ST:LAST_NAME Sáiz ST:FIRST_NAME Jorge ST:ADDRESS JULIÁN ROMEA 23, Madrid, Madrid, 28003, Spain ST:EMAIL jorge.saizgalindo@ceu.es ST:PHONE none #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 #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 - WT1-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT1-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT2-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT2-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT3-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT3-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT4-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT4-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT5-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT5-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT6-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT6-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT7-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT7-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT8-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT8-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT9-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT9-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT10-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT10-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT11-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT11-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT12-HD-POS.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT12-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T1-HD-POS.d Subject type:Transg | Treatment:OH-Dopamine RAW_FILE_NAME=T1-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T2-HD-POS.d Subject type:Transg | Treatment:OH-Dopamine RAW_FILE_NAME=T2-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T3-HD-POS.d Subject type:Transg | Treatment:OH-Dopamine RAW_FILE_NAME=T3-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T4-HD-POS.d Subject type:Transg | Treatment:OH-Dopamine RAW_FILE_NAME=T4-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T5-HD-POS.d Subject type:Transg | Treatment:OH-Dopamine RAW_FILE_NAME=T5-HD-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT1-VEH-POS.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT1-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT2-VEH-POS.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT2-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT3-VEH-POS.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT3-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT4-VEH-POS.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT4-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT6-VEH-POS.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT6-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT7-VEH-POS.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT7-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT8-VEH-POS.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT8-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT9-VEH-POS.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT9-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT10-VEH-POS.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT10-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T1-VEH-POS.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T1-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T2-VEH-POS.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T2-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T4-VEH-POS.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T4-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T5-VEH-POS.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T5-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T6-VEH-POS.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T6-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - T7-VEH-POS.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T7-VEH-POS.d; Polarity=POS SUBJECT_SAMPLE_FACTORS - QC1-POS Subject type:QC | Treatment:QC RAW_FILE_NAME=QC1-POS; Polarity=POS SUBJECT_SAMPLE_FACTORS - QC2-POS Subject type:QC | Treatment:QC RAW_FILE_NAME=QC2-POS; Polarity=POS SUBJECT_SAMPLE_FACTORS - QC3-POS Subject type:QC | Treatment:QC RAW_FILE_NAME=QC3-POS; Polarity=POS SUBJECT_SAMPLE_FACTORS - QC4-POS Subject type:QC | Treatment:QC RAW_FILE_NAME=QC4-POS; Polarity=POS SUBJECT_SAMPLE_FACTORS - QC5-POS Subject type:QC | Treatment:QC RAW_FILE_NAME=QC5-POS; Polarity=POS SUBJECT_SAMPLE_FACTORS - QC6-POS Subject type:QC | Treatment:QC RAW_FILE_NAME=QC6-POS; Polarity=POS SUBJECT_SAMPLE_FACTORS - QC7-POS Subject type:QC | Treatment:QC RAW_FILE_NAME=QC7-POS; Polarity=POS SUBJECT_SAMPLE_FACTORS - WT1-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT1-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT2-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT2-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT3-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT3-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT4-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT4-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT5-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT5-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT6-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT6-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT7-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT7-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT8-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT8-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT9-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT9-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT10-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT10-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT11-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT11-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT12-HD-NEG.d Subject type:WT | Treatment:OH-Dopamine RAW_FILE_NAME=WT12-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T1-HD-NEG.d Subject type:Transg | Treatment:OH-Dopamine RAW_FILE_NAME=T1-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T2-HD-NEG.d Subject type:Transg | Treatment:OH-Dopamine RAW_FILE_NAME=T2-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T3-HD-NEG.d Subject type:Transg | Treatment:OH-Dopamine RAW_FILE_NAME=T3-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T4-HD-NEG.d Subject type:Transg | Treatment:OH-Dopamine RAW_FILE_NAME=T4-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T5-HD-NEG.d Subject type:Transg | Treatment:OH-Dopamine RAW_FILE_NAME=T5-HD-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT1-VEH-NEG.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT1-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT2-VEH-NEG.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT2-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT3-VEH-NEG.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT3-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT4-VEH-NEG.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT4-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT6-VEH-NEG.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT6-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT7-VEH-NEG.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT7-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT8-VEH-NEG.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT8-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT9-VEH-NEG.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT9-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - WT10-VEH-NEG.d Subject type:WT | Treatment:Vehicle RAW_FILE_NAME=WT10-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T1-VEH-NEG.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T1-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T2-VEH-NEG.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T2-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T4-VEH-NEG.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T4-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T5-VEH-NEG.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T5-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T6-VEH-NEG.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T6-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - T7-VEH-NEG.d Subject type:Transg | Treatment:Vehicle RAW_FILE_NAME=T7-VEH-NEG.d; Polarity=NEG SUBJECT_SAMPLE_FACTORS - QC1-NEG Subject type:QC | Treatment:QC RAW_FILE_NAME=QC1-NEG; Polarity=NEG SUBJECT_SAMPLE_FACTORS - QC2-NEG Subject type:QC | Treatment:QC RAW_FILE_NAME=QC2-NEG; Polarity=NEG SUBJECT_SAMPLE_FACTORS - QC3-NEG Subject type:QC | Treatment:QC RAW_FILE_NAME=QC3-NEG; Polarity=NEG SUBJECT_SAMPLE_FACTORS - QC4-NEG Subject type:QC | Treatment:QC RAW_FILE_NAME=QC4-NEG; Polarity=NEG SUBJECT_SAMPLE_FACTORS - QC5-NEG Subject type:QC | Treatment:QC RAW_FILE_NAME=QC5-NEG; Polarity=NEG SUBJECT_SAMPLE_FACTORS - QC6-NEG Subject type:QC | Treatment:QC RAW_FILE_NAME=QC6-NEG; Polarity=NEG SUBJECT_SAMPLE_FACTORS - QC7-NEG Subject type:QC | Treatment:QC RAW_FILE_NAME=QC7-NEG; Polarity=NEG #COLLECTION CO:COLLECTION_SUMMARY Samples were collected and frozen at -80ºC CO:SAMPLE_TYPE Brain #TREATMENT TR:TREATMENT_SUMMARY Striatum resections were added to 300 µL MeOH:water (50:50) in 2 mL Eppendorf TR:TREATMENT_SUMMARY tubes and were first homogenized with glass beads in a TissueLyser LT (QIAGEN) TR:TREATMENT_SUMMARY for 2 min. The tubes were immersed in liquid N2 and homogenized again in the TR:TREATMENT_SUMMARY TissueLyser LT for another 2 min. A volume 100 µL of the homogenate was TR:TREATMENT_SUMMARY transferred into Eppendorf tubes of 1.5 mL and added with 320 µL of methanol TR:TREATMENT_SUMMARY and 80 µL of MTBE and the mixture was vortexed for 1 h. Afterwards, the vials TR:TREATMENT_SUMMARY were centrifuged at 4000 g for 20 min at 20 ºC and 300 µL of the supernatants TR:TREATMENT_SUMMARY were transferred into new tubes, which were evaporated to dryness in a vacuum TR:TREATMENT_SUMMARY concentrator. Finally, the residues were reconstituted in 50 µL of TR:TREATMENT_SUMMARY MeOH:water:MTBE (37:5:4), being the samples ready for their analysis. Blank TR:TREATMENT_SUMMARY samples were prepared following the sample procedure without the addition of any TR:TREATMENT_SUMMARY biological tissue. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Striatum resections were added to 300 µL MeOH:water (50:50) in 2 mL Eppendorf SP:SAMPLEPREP_SUMMARY tubes and were first homogenized with glass beads in a TissueLyser LT (QIAGEN) SP:SAMPLEPREP_SUMMARY for 2 min. The tubes were immersed in liquid N2 and homogenized again in the SP:SAMPLEPREP_SUMMARY TissueLyser LT for another 2 min. A volume 100 µL of the homogenate was SP:SAMPLEPREP_SUMMARY transferred into Eppendorf tubes of 1.5 mL and added with 320 µL of methanol SP:SAMPLEPREP_SUMMARY and 80 µL of MTBE and the mixture was vortexed for 1 h. Afterwards, the vials SP:SAMPLEPREP_SUMMARY were centrifuged at 4000 g for 20 min at 20 ºC and 300 µL of the supernatants SP:SAMPLEPREP_SUMMARY were transferred into new tubes, which were evaporated to dryness in a vacuum SP:SAMPLEPREP_SUMMARY concentrator. Finally, the residues were reconstituted in 50 µL of SP:SAMPLEPREP_SUMMARY MeOH:water:MTBE (37:5:4), being the samples ready for their analysis. Blank SP:SAMPLEPREP_SUMMARY samples were prepared following the sample procedure without the addition of any SP:SAMPLEPREP_SUMMARY biological tissue. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Agilent 1200 CH:COLUMN_NAME RP C8 column Agilent Poroshell (150 mm 2.1 mm, 2.7 μm) #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 All data were controlled and acquired using Mass Hunter Qualitative Analysis MS:MS_COMMENTS B.07.00 (Agilent Technologies). Data obtained from LC-MS were cleaned of MS:MS_COMMENTS background noise and unrelated ions. Peak detection, deconvolution and alignment MS:MS_COMMENTS were performed by the recursive feature extraction (RFE) using Profinder MS:MS_COMMENTS Software B.08.00 (Agilent Technologies). Blank subtraction and filtering by MS:MS_COMMENTS frequency of at least 50% of the QC and 60% of each group and relative standard MS:MS_COMMENTS deviation (RSD) less than 30% in QC were performed, to keep only the relevant MS:MS_COMMENTS features. Missing values were substituted by KNN algorithm. MS:MS_RESULTS_FILE ST001810_AN002934_Results.txt UNITS:area Has m/z:Neutral masses Has RT:Yes RT units:Minutes #END