#METABOLOMICS WORKBENCH michaelsa93_20170427_170716_mwtab.txt DATATRACK_ID:900 STUDY_ID:ST000604 ANALYSIS_ID:AN000924 PROJECT_ID:PR000328 VERSION 1 CREATED_ON May 11, 2017, 10:44 am #PROJECT PR:PROJECT_TITLE Impact Of High Sugar Diet On L-Arginine Metabolism In The Lung PR:PROJECT_SUMMARY Asthma is a progressive inflammatory airways disease that leads to structural PR:PROJECT_SUMMARY airway changes and debilitating symptoms in many severely affected adults. We PR:PROJECT_SUMMARY need novel therapeutic agents that are affordable, can decrease the reliance on PR:PROJECT_SUMMARY steroids, and can improve quality of life. This clinical and mechanistic study PR:PROJECT_SUMMARY has the potential to impact treatment of a subset of adult severe asthmatics and PR:PROJECT_SUMMARY to further our understanding of the mechanisms of L-arginine metabolism and NO PR:PROJECT_SUMMARY biology in the airways of asthmatics. We will pursue a clinical trial in PR:PROJECT_SUMMARY subjects not well controlled on standard drug therapy; this strategy will PR:PROJECT_SUMMARY address whether L-arginine is efficacious in patients receiving standard of care PR:PROJECT_SUMMARY medications. In studies using animal models, we and others have shown that PR:PROJECT_SUMMARY interventions that augment NO levels, through either supplementation of PR:PROJECT_SUMMARY L-arginine or inhibition of arginase, decrease allergic airway inflammation and PR:PROJECT_SUMMARY hyperresponsiveness-the two hallmarks of asthma. Overall, we hypothesize that a PR:PROJECT_SUMMARY responder subset of adult severe asthma patients will derive clinical benefit PR:PROJECT_SUMMARY from supplemental L-arginine therapy and that these patients will have a lower PR:PROJECT_SUMMARY exhaled NO concentrations (<20 ppb) and a higher NOS2/Arg1 mRNA and protein PR:PROJECT_SUMMARY ratio in their airway epithelial cells than non-responders. We aim to: 1) test PR:PROJECT_SUMMARY the hypothesis that uncontrolled, adult severe asthma patients with exhaled PR:PROJECT_SUMMARY breath NO concentrations <20 ppb will have fewer asthma exacerbations over 3 PR:PROJECT_SUMMARY months when treated with L-arginine compared to patients with FeNO > 25, 2) PR:PROJECT_SUMMARY determine the mechanisms by which L-arginine affects the regulation of NOS and PR:PROJECT_SUMMARY arginase enzymes in primary airway epithelial cell cultures from severe PR:PROJECT_SUMMARY asthmatic subjects, and 3) test the hypothesis that inhaled nanoparticle carrier PR:PROJECT_SUMMARY formulations of L-arginine will decrease airway inflammation, airway PR:PROJECT_SUMMARY hyperresponsiveness, and airway fibrosis at lower doses than systemically PR:PROJECT_SUMMARY administered L-arginine. The major impact of our study will be to identify the PR:PROJECT_SUMMARY adult severe asthma cohort that will benefit from supplemental L-arginine PR:PROJECT_SUMMARY therapy. Our ultimate goal is to develop novel therapeutic agents to treat adult PR:PROJECT_SUMMARY severe asthma patients better. PUBLIC HEALTH RELEVANCE: Asthma is a progressive PR:PROJECT_SUMMARY inflammatory airways disease that leads to structural airway changes and PR:PROJECT_SUMMARY debilitating symptoms in many severely affected adults. This clinical study has PR:PROJECT_SUMMARY the potential to improve the care of adult severe asthmatics and to further our PR:PROJECT_SUMMARY understanding of the mechanisms of L-arginine metabolism and nitric oxide PR:PROJECT_SUMMARY biology in the lung. If we demonstrate that L-arginine supplementation can PR:PROJECT_SUMMARY decrease asthma attacks in a subset of severe asthmatics, it will have great PR:PROJECT_SUMMARY implications for future research as well as for the daily lives of patients with PR:PROJECT_SUMMARY asthma. PR:INSTITUTE University of California, Davis PR:DEPARTMENT Genome and Biomedical Sciences Facility PR:LABORATORY WCMC Metabolomics Core PR:LAST_NAME Fiehn PR:FIRST_NAME Oliver PR:ADDRESS 1315 Genome and Biomedical Sciences Facility, 451 Health Sciences Drive, Davis, PR:ADDRESS CA 95616 PR:EMAIL ofiehn@ucdavis.edu PR:PHONE (530) 754-8258 PR:FUNDING_SOURCE NIH U24DK097154 #STUDY ST:STUDY_TITLE Impact Of High Sugar Diet On L-Arginine Metabolism In The Lung ST:STUDY_SUMMARY Asthma is a progressive inflammatory airways disease that leads to structural ST:STUDY_SUMMARY airway changes and debilitating symptoms in many severely affected adults. We ST:STUDY_SUMMARY need novel therapeutic agents that are affordable, can decrease the reliance on ST:STUDY_SUMMARY steroids, and can improve quality of life. This clinical and mechanistic study ST:STUDY_SUMMARY has the potential to impact treatment of a subset of adult severe asthmatics and ST:STUDY_SUMMARY to further our understanding of the mechanisms of L-arginine metabolism and NO ST:STUDY_SUMMARY biology in the airways of asthmatics. We will pursue a clinical trial in ST:STUDY_SUMMARY subjects not well controlled on standard drug therapy; this strategy will ST:STUDY_SUMMARY address whether L-arginine is efficacious in patients receiving standard of care ST:STUDY_SUMMARY medications. In studies using animal models, we and others have shown that ST:STUDY_SUMMARY interventions that augment NO levels, through either supplementation of ST:STUDY_SUMMARY L-arginine or inhibition of arginase, decrease allergic airway inflammation and ST:STUDY_SUMMARY hyperresponsiveness-the two hallmarks of asthma. Overall, we hypothesize that a ST:STUDY_SUMMARY responder subset of adult severe asthma patients will derive clinical benefit ST:STUDY_SUMMARY from supplemental L-arginine therapy and that these patients will have a lower ST:STUDY_SUMMARY exhaled NO concentrations (<20 ppb) and a higher NOS2/Arg1 mRNA and protein ST:STUDY_SUMMARY ratio in their airway epithelial cells than non-responders. We aim to: 1) test ST:STUDY_SUMMARY the hypothesis that uncontrolled, adult severe asthma patients with exhaled ST:STUDY_SUMMARY breath NO concentrations <20 ppb will have fewer asthma exacerbations over 3 ST:STUDY_SUMMARY months when treated with L-arginine compared to patients with FeNO > 25, 2) ST:STUDY_SUMMARY determine the mechanisms by which L-arginine affects the regulation of NOS and ST:STUDY_SUMMARY arginase enzymes in primary airway epithelial cell cultures from severe ST:STUDY_SUMMARY asthmatic subjects, and 3) test the hypothesis that inhaled nanoparticle carrier ST:STUDY_SUMMARY formulations of L-arginine will decrease airway inflammation, airway ST:STUDY_SUMMARY hyperresponsiveness, and airway fibrosis at lower doses than systemically ST:STUDY_SUMMARY administered L-arginine. The major impact of our study will be to identify the ST:STUDY_SUMMARY adult severe asthma cohort that will benefit from supplemental L-arginine ST:STUDY_SUMMARY therapy. Our ultimate goal is to develop novel therapeutic agents to treat adult ST:STUDY_SUMMARY severe asthma patients better. PUBLIC HEALTH RELEVANCE: Asthma is a progressive ST:STUDY_SUMMARY inflammatory airways disease that leads to structural airway changes and ST:STUDY_SUMMARY debilitating symptoms in many severely affected adults. This clinical study has ST:STUDY_SUMMARY the potential to improve the care of adult severe asthmatics and to further our ST:STUDY_SUMMARY understanding of the mechanisms of L-arginine metabolism and nitric oxide ST:STUDY_SUMMARY biology in the lung. If we demonstrate that L-arginine supplementation can ST:STUDY_SUMMARY decrease asthma attacks in a subset of severe asthmatics, it will have great ST:STUDY_SUMMARY implications for future research as well as for the daily lives of patients with ST:STUDY_SUMMARY asthma. ST:INSTITUTE University of California, Davis ST:DEPARTMENT Genome and Biomedical Sciences Facility ST:LABORATORY WCMC Metabolomics Core ST:LAST_NAME Fiehn ST:FIRST_NAME Oliver ST:ADDRESS 1315 Genome and Biomedical Sciences Facility, 451 Health Sciences Drive, Davis, ST:ADDRESS CA 95616 ST:EMAIL ofiehn@ucdavis.edu ST:PHONE (530) 754-8258 #SUBJECT SU:SUBJECT_TYPE Animal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 9606 SU:GENDER Male #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS QC Inj011_CSH_QC_01.d Source:Plasma | Species:Human | Treatment:None SUBJECT_SAMPLE_FACTORS QC Inj022_CSH_QC_02.d Source:Plasma | Species:Human | Treatment:None SUBJECT_SAMPLE_FACTORS QC Inj033_CSH_QC_03.d Source:Plasma | Species:Human | Treatment:None SUBJECT_SAMPLE_FACTORS QC Inj039_CSH_QC_04.d Source:Plasma | Species:Human | Treatment:None SUBJECT_SAMPLE_FACTORS QC Inj050_CSH_QC_05.d Source:Plasma | Species:Human | Treatment:None SUBJECT_SAMPLE_FACTORS QC Inj061_CSH_QC_06.d Source:Plasma | Species:Human | Treatment:None SUBJECT_SAMPLE_FACTORS QC Inj067_CSH_QC_07.d Source:Plasma | Species:Human | Treatment:None SUBJECT_SAMPLE_FACTORS QC Inj078_CSH_QC_08.d Source:Plasma | Species:Human | Treatment:None SUBJECT_SAMPLE_FACTORS QC Inj089_CSH_QC_09.d Source:Plasma | Species:Human | Treatment:None SUBJECT_SAMPLE_FACTORS QC Inj096_CSH_QC_10.d Source:Plasma | Species:Human | Treatment:None SUBJECT_SAMPLE_FACTORS HFS59 Inj012_CSH_Kid_HFS59.d Source:Kidney | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS HFS70 Inj013_CSH_Kid_HFS70.d Source:Kidney | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS CF90 Inj014_CSH_Kid_CF90.d Source:Kidney | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS VHF97 Inj015_CSH_Kid_VHF97.d Source:Kidney | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS CF54 Inj016_CSH_Kid_CF54.d Source:Kidney | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS CF78 Inj017_CSH_Kid_CF78.d Source:Kidney | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS CF49 Inj018_CSH_Kid_CF49.d Source:Kidney | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS HFS82 Inj019_CSH_Kid_HFS82.d Source:Kidney | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS HFS1 Inj020_CSH_Kid_HFS1.d Source:Kidney | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS VHF45 Inj021_CSH_Kid_VHF45.d Source:Kidney | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF92 Inj023_CSH_Kid_VHF92.d Source:Kidney | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF2 Inj024_CSH_Kid_VHF2.d Source:Kidney | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF80 Inj025_CSH_Kid_VHF80.d Source:Kidney | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS CF57 Inj026_CSH_Kid_CF57.d Source:Kidney | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS VHF64 Inj027_CSH_Kid_VHF64.d Source:Kidney | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS CF82 Inj028_CSH_Kid_CF82.d Source:Kidney | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS CF70 Inj029_CSH_Kid_CF70.d Source:Kidney | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS VHF86 Inj030_CSH_Kid_VHF86.d Source:Kidney | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS CF26 Inj031_CSH_Kid_CF26.d Source:Kidney | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS HFS69 Inj032_CSH_Kid_HFS69.d Source:Kidney | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS HFS87 Inj034_CSH_Kid_HFS87.d Source:Kidney | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS VHF31 Inj035_CSH_Kid_VHF31.d Source:Kidney | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS HFS55 Inj036_CSH_Kid_HFS55.d Source:Kidney | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS VHF70 Inj037_CSH_Kid_VHF70.d Source:Kidney | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS CF11 Inj038_CSH_Kid_CF11.d Source:Kidney | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS HFS55 Inj068_CSH_Liv_HFS55.d Source:Liver | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS HFS1 Inj069_CSH_Liv_HFS1.d Source:Liver | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS VHF64 Inj070_CSH_Liv_VHF64.d Source:Liver | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS HFS82 Inj071_CSH_Liv_HFS82.d Source:Liver | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS CF78 Inj072_CSH_Liv_CF78.d Source:Liver | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS VHF2 Inj073_CSH_Liv_VHF2.d Source:Liver | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF86 Inj074_CSH_Liv_VHF86.d Source:Liver | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF31 Inj075_CSH_Liv_VHF31.d Source:Liver | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF45 Inj076_CSH_Liv_VHF45.d Source:Liver | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS HFS59 Inj077_CSH_Liv_HFS59.d Source:Liver | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS CF57 Inj079_CSH_Liv_CF57.d Source:Liver | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS VHF92 Inj080_CSH_Liv_VHF92.d Source:Liver | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF97 Inj081_CSH_Liv_VHF97.d Source:Liver | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS HFS69 Inj082_CSH_Liv_HFS69.d Source:Liver | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS HFS87 Inj083_CSH_Liv_HFS87.d Source:Liver | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS CF49 Inj084_CSH_Liv_CF49.d Source:Liver | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS CF70 Inj085_CSH_Liv_CF70.d Source:Liver | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS VHF70 Inj086_CSH_Liv_VHF70.d Source:Liver | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF80 Inj087_CSH_Liv_VHF80.d Source:Liver | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS CF54 Inj088_CSH_Liv_CF54.d Source:Liver | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS CF11 Inj090_CSH_Liv_CF11.d Source:Liver | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS CF82 Inj091_CSH_Liv_CF82.d Source:Liver | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS CF90 Inj092_CSH_Liv_CF90.d Source:Liver | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS HFS92 Inj093_CSH_Liv_HFS92.d Source:Liver | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS HFS55 Inj040_CSH_Lung_HFS55.d Source:Lung | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS CF54 Inj041_CSH_Lung_CF54.d Source:Lung | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS HFS82 Inj042_CSH_Lung_HFS82.d Source:Lung | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS VHF2 Inj043_CSH_Lung_VHF2.d Source:Lung | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS CF90 Inj044_CSH_Lung_CF90.d Source:Lung | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS CF82 Inj045_CSH_Lung_CF82.d Source:Lung | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS VHF80 Inj046_CSH_Lung_VHF80.d Source:Lung | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF31 Inj047_CSH_Lung_VHF31.d Source:Lung | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS HFS87 Inj048_CSH_Lung_HFS87.d Source:Lung | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS VHF45 Inj049_CSH_Lung_VHF45.d Source:Lung | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF70 Inj051_CSH_Lung_VHF70.d Source:Lung | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS VHF86 Inj052_CSH_Lung_VHF86.d Source:Lung | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS CF70 Inj053_CSH_Lung_CF70.d Source:Lung | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS CF49 Inj054_CSH_Lung_CF49.d Source:Lung | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS VHF92 Inj055_CSH_Lung_VHF92.d Source:Lung | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS HFS59 Inj056_CSH_Lung_HFS59.d Source:Lung | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS HFS92 Inj057_CSH_Lung_HFS92.d Source:Lung | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS CF78 Inj058_CSH_Lung_CF78.d Source:Lung | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS HFS69 Inj059_CSH_Lung_HFS69.d Source:Lung | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS CF57 Inj060_CSH_Lung_CF57.d Source:Lung | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS CF11 Inj062_CSH_Lung_CF11.d Source:Lung | Species:Mouse | Treatment:Control Diet SUBJECT_SAMPLE_FACTORS VHF64 Inj063_CSH_Lung_VHF64.d Source:Lung | Species:Mouse | Treatment:Very High Fat Diet SUBJECT_SAMPLE_FACTORS HFS70 Inj064_CSH_Lung_HFS70.d Source:Lung | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS HFS1 Inj065_CSH_Lung_HFS1.d Source:Lung | Species:Mouse | Treatment:High Fat Diet SUBJECT_SAMPLE_FACTORS CF26 Inj066_CSH_Lung_CF26.d Source:Lung | Species:Mouse | Treatment:Control Diet #COLLECTION CO:COLLECTION_SUMMARY C57BL/6 mice, 6-7 weeks of age upon diet assignment were fed for 150 days and CO:COLLECTION_SUMMARY weighed daily. Mice were euthanized with an overdose of pentobarbital IP and CO:COLLECTION_SUMMARY lungs were flash frozen. 6 mg of lung tissue was extracted for GC-TOFMS and CO:COLLECTION_SUMMARY HILIC-QTOFMS analysis. CO:SAMPLE_TYPE Tissue #TREATMENT TR:TREATMENT_SUMMARY Male C57BL/6N mice (6-7 weeks of age) were provided ad libitum access to one of TR:TREATMENT_SUMMARY three diets for 150 days: low fat (10% kcals) control (CTRL) chow, high fat (45% TR:TREATMENT_SUMMARY kcals) with sugar (HFS) chow or very high (60% kcals) fat (VHF) chow. Body TR:TREATMENT_SUMMARY weight and food intake were measured daily. TR:TREATMENT_DOSEDURATION 150 Days #SAMPLEPREP SP:SAMPLEPREP_SUMMARY 1. Weigh 50 mg tissue sample in to a 25 ml conical polypropylene centrifuge SP:SAMPLEPREP_SUMMARY tube. 2. Add 2.5mL extraction solvent to the tissue sample and homogenize for 45 SP:SAMPLEPREP_SUMMARY seconds ensuring that sample resembles a powder. In between samples, clean the SP:SAMPLEPREP_SUMMARY homogenizer in solutions of methanol, acetone, water, and the extraction SP:SAMPLEPREP_SUMMARY solvent. 3. Centrifuge the samples at 2500 rpm. for 5 minutes. Aliquot 2 X SP:SAMPLEPREP_SUMMARY 500?l supernatant, one for analysis and one for a backup sample. Store backup SP:SAMPLEPREP_SUMMARY aliquot in the -20°C freezer. 4. Evaporate one 500?l aliquot of the sample in SP:SAMPLEPREP_SUMMARY the Labconco Centrivap cold trap concentrator to complete dryness 5. The dried SP:SAMPLEPREP_SUMMARY aliquot is then re-suspended with 500l 50% acetonitrile (degassed as given) 6. SP:SAMPLEPREP_SUMMARY Centrifuge for 2 min at 14000 rcf using the centrifuge Eppendorf 5415. 7. Remove SP:SAMPLEPREP_SUMMARY supernatant to a new Eppendorff tube. 8. Evaporate the supernatant to dryness in SP:SAMPLEPREP_SUMMARY the the Labconco Centrivap cold trap concentrator. 9. Submit to derivatization. SP:SAMPLEPREP_PROTOCOL_FILENAME SOP_Extraction_of_Mammalian_Tissue_Samples.pdf SP:SAMPLEPREP_PROTOCOL_COMMENTS This study combined 2 different injection volumes for certain samples when SP:SAMPLEPREP_PROTOCOL_COMMENTS processing positive mode data. The reason for this was because in certain SP:SAMPLEPREP_PROTOCOL_COMMENTS samples the TG's were overloaded with the 1uL injections, but everything else SP:SAMPLEPREP_PROTOCOL_COMMENTS was at a normal peak height. Therefore samples were run at a 0.1uL injection SP:SAMPLEPREP_PROTOCOL_COMMENTS volume and took the data from 9-12.5 minutes because that's when TG's elute. The SP:SAMPLEPREP_PROTOCOL_COMMENTS samples that were injected at 0.1uL were used for times 9 min-12.5 min and the SP:SAMPLEPREP_PROTOCOL_COMMENTS 1uL samples were used for times 0-8.99min. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Data Dictionary Fiehn laboratory_CSH QTOF lipidomics_05-29-2014.pdf CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Agilent 6530 CH:COLUMN_NAME Waters Acquity CSH C18 (100 x 2.1mm, 1.7um) CH:COLUMN_NAME 1.7um Pre-Column CH:FLOW_GRADIENT 15% B to 99% B CH:FLOW_RATE 0.6 mL/min CH:COLUMN_TEMPERATURE 65 C CH:SOLVENT_A 60:40 Acetonitrile:Water +10mM Ammonium Formate +10mM Formic Acid CH:SOLVENT_B 9:1 Isopropanol:Acetonitrile +10mM Ammonium Formate +10mM Formic Acid CH:COLUMN_PRESSURE 450-850 bar CH:INJECTION_TEMPERATURE 4 C CH:INTERNAL_STANDARD See data dictionary CH:RETENTION_TIME See data dictionary CH:SAMPLE_INJECTION 1.67 uL CH:ANALYTICAL_TIME 13 min CH:CAPILLARY_VOLTAGE 3500 CH:TIME_PROGRAM 15 min CH:WEAK_WASH_SOLVENT_NAME Isopropanol CH:STRONG_WASH_SOLVENT_NAME Isopropanol CH:TARGET_SAMPLE_TEMPERATURE Autosampler temp 4 C CH:RANDOMIZATION_ORDER Excel #ANALYSIS AN:ANALYSIS_TYPE MS AN:LABORATORY_NAME WCMC Metabolomics Core AN:DETECTOR_TYPE TOF MCP AN:SOFTWARE_VERSION Masshunter AN:DATA_FORMAT .d #MS MS:MS_COMMENTS - MS:INSTRUMENT_NAME Agilent 6530 QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:CAPILLARY_VOLTAGE 3500 MS:COLLISION_ENERGY 25 eV MS:COLLISION_GAS Nitrogen MS:DRY_GAS_FLOW 8L/min MS:DRY_GAS_TEMP 325 C MS:FRAGMENT_VOLTAGE 120 MS:FRAGMENTATION_METHOD Auto MSMS MS:ION_SOURCE_TEMPERATURE 325 C MS:ION_SPRAY_VOLTAGE 1000 MS:IONIZATION Pos MS:PRECURSOR_TYPE Intact Molecule MS:REAGENT_GAS Nitrogen MS:SOURCE_TEMPERATURE 325 C MS:DATAFORMAT .d MS:DESOLVATION_GAS_FLOW 11 L/min MS:DESOLVATION_TEMPERATURE 350 C MS:NEBULIZER 35 psig MS:OCTPOLE_VOLTAGE 750 MS:RESOLUTION_SETTING Exteded Dyamic Range MS:SCAN_RANGE_MOVERZ 60-1700 Da MS:SCANNING_CYCLE 2 Hz MS:SCANNING_RANGE 60-1700 Da MS:SKIMMER_VOLTAGE 65 MS:MS_RESULTS_FILE ST000604_AN000924_Results.txt UNITS:Counts #END