{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST002747","ANALYSIS_ID":"AN004455","VERSION":"1","CREATED_ON":"June 26, 2023, 5:29 pm"},

"PROJECT":{"PROJECT_TITLE":"Evolutionary genomics identifies host-directed therapeutics to treat intracellular bacterial infections","PROJECT_SUMMARY":"Obligate intracellular bacteria from the Rickettsiaceae family have shed essential biosynthetic pathways during their evolution towards host dependency. By systematically comparing this cytosolic family of bacteria to the related vacuolar Anaplasmataceae family using a novel computational pipeline called PoMeLo, we identified 20 metabolic pathways that may have been lost since the divergence of Anaplasmataceae and Rickettsiaceae, corresponding to the latter’s change to a cytosolic niche. We hypothesized that drug inhibition of these host metabolic pathways would reduce the levels of metabolic products available to the bacteria, thereby inhibiting bacterial growth. We tested 22 commercially available inhibitors for 14 of the identified pathways and found that 59% of the inhibitors reduced bacterial growth at concentrations that did not contribute to host cell cytotoxicity. Of these, 5 inhibitors with an IC50 under 5 µM were tested to determine whether their mode of inhibition was bactericidal or bacteriostatic. Both mycophenolate mofetil, an inhibitor of inosine-5'-monophosphate dehydrogenase in the purine biosynthesis pathway, and roseoflavin, an analog of riboflavin, displayed bactericidal activity. We then took an unbiased metabolomics approach to Rickettsia-infected cells to determine whether there was any overlap between our predicted host pathways and depletion of metabolite levels in infected cells, as measured by mass spectrometry. Our results show that 13 pathways were identified as metabolic gaps in both our computational predictions and our metabolomics analysis. These in vitro validation studies support the feasibility of a novel evolutionary genomics-guided approach for antibiotic drug development against obligate pathogens.","INSTITUTE":"CZ Biohub","LAST_NAME":"DeFelice","FIRST_NAME":"Brian","ADDRESS":"1291 Welch Rd., Rm. G0821 (SIM1), Stanford CA, California, 94305, USA","EMAIL":"bcdefelice@ucdavis.edu","PHONE":"5303564485"},

"STUDY":{"STUDY_TITLE":"Evolutionary genomics identifies host-directed therapeutics to treat intracellular bacterial infections","STUDY_SUMMARY":"Obligate intracellular bacteria from the Rickettsiaceae family have shed essential biosynthetic pathways during their evolution towards host dependency. By systematically comparing this cytosolic family of bacteria to the related vacuolar Anaplasmataceae family using a novel computational pipeline called PoMeLo, we identified 20 metabolic pathways that may have been lost since the divergence of Anaplasmataceae and Rickettsiaceae, corresponding to the latter’s change to a cytosolic niche. We hypothesized that drug inhibition of these host metabolic pathways would reduce the levels of metabolic products available to the bacteria, thereby inhibiting bacterial growth. We tested 22 commercially available inhibitors for 14 of the identified pathways and found that 59% of the inhibitors reduced bacterial growth at concentrations that did not contribute to host cell cytotoxicity. Of these, 5 inhibitors with an IC50 under 5 µM were tested to determine whether their mode of inhibition was bactericidal or bacteriostatic. Both mycophenolate mofetil, an inhibitor of inosine-5'-monophosphate dehydrogenase in the purine biosynthesis pathway, and roseoflavin, an analog of riboflavin, displayed bactericidal activity. We then took an unbiased metabolomics approach to Rickettsia-infected cells to determine whether there was any overlap between our predicted host pathways and depletion of metabolite levels in infected cells, as measured by mass spectrometry. Our results show that 13 pathways were identified as metabolic gaps in both our computational predictions and our metabolomics analysis. These in vitro validation studies support the feasibility of a novel evolutionary genomics-guided approach for antibiotic drug development against obligate pathogens.","INSTITUTE":"CZ Biohub","LAST_NAME":"DeFelice","FIRST_NAME":"Brian","ADDRESS":"1291 Welch Rd., Rm. G0821 (SIM1), Stanford CA, California, 94305, USA","EMAIL":"bcdefelice@ucdavis.edu","PHONE":"5303564485"},

"SUBJECT":{"SUBJECT_TYPE":"Cultured cells","SUBJECT_SPECIES":"Rickettsia parkeri; Homo sapiens","TAXONOMY_ID":"35792; 9606"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Pos_M1_QE2_HILIC_008",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Pos_M1_QE2_HILIC_008"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Neg_M1_QE2_HILIC_009",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Neg_M1_QE2_HILIC_009"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Pos_M2_QE2_HILIC_010",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Pos_M2_QE2_HILIC_010"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Neg_M2_QE2_HILIC_011",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Neg_M2_QE2_HILIC_011"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Pos_Inf5_QE2_HILIC_012",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Pos_Inf5_QE2_HILIC_012"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Neg_Inf5_QE2_HILIC_013",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Neg_Inf5_QE2_HILIC_013"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Pos_M5_QE2_HILIC_014",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Pos_M5_QE2_HILIC_014"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Neg_M5_QE2_HILIC_015",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Neg_M5_QE2_HILIC_015"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Pos_Inf3_QE2_HILIC_016",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Pos_Inf3_QE2_HILIC_016"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Neg_Inf3_QE2_HILIC_017",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Neg_Inf3_QE2_HILIC_017"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Pos_M3_QE2_HILIC_020",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Pos_M3_QE2_HILIC_020"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Neg_M3_QE2_HILIC_021",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Neg_M3_QE2_HILIC_021"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Pos_Inf1_QE2_HILIC_022",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Pos_Inf1_QE2_HILIC_022"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Neg_Inf1_QE2_HILIC_023",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Neg_Inf1_QE2_HILIC_023"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Pos_Inf4_QE2_HILIC_024",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Pos_Inf4_QE2_HILIC_024"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Neg_Inf4_QE2_HILIC_025",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Neg_Inf4_QE2_HILIC_025"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Pos_Inf2_QE2_HILIC_026",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Pos_Inf2_QE2_HILIC_026"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Neg_Inf2_QE2_HILIC_027",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Neg_Inf2_QE2_HILIC_027"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Pos_M4_QE2_HILIC_028",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Pos_M4_QE2_HILIC_028"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_Neg_M4_QE2_HILIC_029",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_Neg_M4_QE2_HILIC_029"}
},
{
"Subject ID":"-",
"Sample ID":"M2_uninfected_NA_2__008",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"M2_uninfected_NA_2__008"}
},
{
"Subject ID":"-",
"Sample ID":"M2_uninfected_NA_2__009",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"M2_uninfected_NA_2__009"}
},
{
"Subject ID":"-",
"Sample ID":"M3_uninfected_NA_3__010",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"M3_uninfected_NA_3__010"}
},
{
"Subject ID":"-",
"Sample ID":"M3_uninfected_NA_3__011",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"M3_uninfected_NA_3__011"}
},
{
"Subject ID":"-",
"Sample ID":"M5_uninfected_NA_5__012",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"M5_uninfected_NA_5__012"}
},
{
"Subject ID":"-",
"Sample ID":"M5_uninfected_NA_5__013",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"M5_uninfected_NA_5__013"}
},
{
"Subject ID":"-",
"Sample ID":"Inf5_infected_NA_5__014",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"Inf5_infected_NA_5__014"}
},
{
"Subject ID":"-",
"Sample ID":"Inf5_infected_NA_5__015",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"Inf5_infected_NA_5__015"}
},
{
"Subject ID":"-",
"Sample ID":"M1_uninfected_NA_1__016",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"M1_uninfected_NA_1__016"}
},
{
"Subject ID":"-",
"Sample ID":"M1_uninfected_NA_1__017",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"M1_uninfected_NA_1__017"}
},
{
"Subject ID":"-",
"Sample ID":"Inf2_infected_NA_2__020",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"Inf2_infected_NA_2__020"}
},
{
"Subject ID":"-",
"Sample ID":"Inf2_infected_NA_2__021",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"Inf2_infected_NA_2__021"}
},
{
"Subject ID":"-",
"Sample ID":"Inf4_infected_NA_4__022",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"Inf4_infected_NA_4__022"}
},
{
"Subject ID":"-",
"Sample ID":"Inf4_infected_NA_4__023",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"Inf4_infected_NA_4__023"}
},
{
"Subject ID":"-",
"Sample ID":"Inf1_infected_NA_1__024",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"Inf1_infected_NA_1__024"}
},
{
"Subject ID":"-",
"Sample ID":"Inf1_infected_NA_1__025",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"Inf1_infected_NA_1__025"}
},
{
"Subject ID":"-",
"Sample ID":"Inf3_infected_NA_3__026",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"Inf3_infected_NA_3__026"}
},
{
"Subject ID":"-",
"Sample ID":"Inf3_infected_NA_3__027",
"Factors":{"Genotype":"Rickettsia parkeri & human","Treatment":"PostInfection"},
"Additional sample data":{"RAW_FILE_NAME":"Inf3_infected_NA_3__027"}
},
{
"Subject ID":"-",
"Sample ID":"M4_uninfected_NA_4__028",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"M4_uninfected_NA_4__028"}
},
{
"Subject ID":"-",
"Sample ID":"M4_uninfected_NA_4__029",
"Factors":{"Genotype":"Human","Treatment":"control"},
"Additional sample data":{"RAW_FILE_NAME":"M4_uninfected_NA_4__029"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_BK_1_Pos_QE2_HILIC_006",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_BK_1_Pos_QE2_HILIC_006"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_BK_2_Pos_QE2_HILIC_018",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_BK_2_Pos_QE2_HILIC_018"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_BK_3_Pos_QE2_HILIC_030",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_BK_3_Pos_QE2_HILIC_030"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_BK_1_Neg_QE2_HILIC_007",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_BK_1_Neg_QE2_HILIC_007"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_BK_2_Neg_QE2_HILIC_019",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_BK_2_Neg_QE2_HILIC_019"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002H_BK_3_Neg_QE2_HILIC_031",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002H_BK_3_Neg_QE2_HILIC_031"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002_BK_1_Pos_QE1_Lipids_006",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002_BK_1_Pos_QE1_Lipids_006"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002_BK_2_Pos_QE1_Lipids_018",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002_BK_2_Pos_QE1_Lipids_018"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002_BK_3_Pos_QE1_Lipids_030",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002_BK_3_Pos_QE1_Lipids_030"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002_BK_1_Neg_QE1_Lipids_007",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002_BK_1_Neg_QE1_Lipids_007"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002_BK_2_Neg_QE1_Lipids_019",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002_BK_2_Neg_QE1_Lipids_019"}
},
{
"Subject ID":"-",
"Sample ID":"VIAH002_BK_3_Neg_QE1_Lipids_031",
"Factors":{"Genotype":"blank","Treatment":"blank"},
"Additional sample data":{"RAW_FILE_NAME":"VIAH002_BK_3_Neg_QE1_Lipids_031"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"A549 cells were seeded into 10 x 6-well tissue culture treated plates the day before infection. On the day of infection, cells were either infected with an MOI of 0.05 in 1 mL of DMEM (5 replicates) or mock-infected with DMEM only (5 replicates). Plates were centrifuged at 300 × g for 5 minutes and placed in a 33°C in 5% CO2 for 4 days. After 4 days, cells from each well of a 6-well plate were washed with 2 mL of 1x PBS, scraped, and collected into an 1.5 mL centrifuge tube. Samples were washed twice with 500 µl of 1x PBS before resuspending in 225 µl ice-cold methanol with 1.5% iSTD-SPLASH and freezing on dry ice. Samples were processed within 48 h. Sample preparation and analysis have been detailed previously (DOI: dx.doi.org/10.17504/protocols.io.e6nvwj86dlmk/v1 (Private link for reviewers: https://www.protocols.io/private/8962C900975C11ED8E800A58A9FEAC02 to be removed before publication.) Analysis of metabolomics data was performed on MetaboAnalyst 5.0 and GraphPad Prism (Version 9.5.1 (528); Chong et al., 2018).","SAMPLE_TYPE":"A549 cells"},

"TREATMENT":{"TREATMENT_SUMMARY":"A549 cells were seeded into 10 x 6-well tissue culture treated plates the day before infection. On the day of infection, cells were either infected with an MOI of 0.05 in 1 mL of DMEM (5 replicates) or mock-infected with DMEM only (5 replicates). Plates were centrifuged at 300 × g for 5 minutes and placed in a 33°C in 5% CO2 for 4 days. After 4 days, cells from each well of a 6-well plate were washed with 2 mL of 1x PBS, scraped, and collected into an 1.5 mL centrifuge tube. Samples were washed twice with 500 µl of 1x PBS before resuspending in 225 µl ice-cold methanol with 1.5% iSTD-SPLASH and freezing on dry ice. Samples were processed within 48 h. Sample preparation and analysis have been detailed previously (DOI: dx.doi.org/10.17504/protocols.io.e6nvwj86dlmk/v1 (Private link for reviewers: https://www.protocols.io/private/8962C900975C11ED8E800A58A9FEAC02 to be removed before publication.) Analysis of metabolomics data was performed on MetaboAnalyst 5.0 and GraphPad Prism (Version 9.5.1 (528); Chong et al., 2018)."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"A549 cells were seeded into 10 x 6-well tissue culture treated plates the day before infection. On the day of infection, cells were either infected with an MOI of 0.05 in 1 mL of DMEM (5 replicates) or mock-infected with DMEM only (5 replicates). Plates were centrifuged at 300 × g for 5 minutes and placed in a 33°C in 5% CO2 for 4 days. After 4 days, cells from each well of a 6-well plate were washed with 2 mL of 1x PBS, scraped, and collected into an 1.5 mL centrifuge tube. Samples were washed twice with 500 µl of 1x PBS before resuspending in 225 µl ice-cold methanol with 1.5% iSTD-SPLASH and freezing on dry ice. Samples were processed within 48 h. Sample preparation and analysis have been detailed previously (DOI: dx.doi.org/10.17504/protocols.io.e6nvwj86dlmk/v1 (Private link for reviewers: https://www.protocols.io/private/8962C900975C11ED8E800A58A9FEAC02 to be removed before publication.) Analysis of metabolomics data was performed on MetaboAnalyst 5.0 and GraphPad Prism (Version 9.5.1 (528); Chong et al., 2018)."},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_SUMMARY":"HILIC","CHROMATOGRAPHY_TYPE":"HILIC","INSTRUMENT_NAME":"Thermo Vanquish","COLUMN_NAME":"Waters ACQUITY UPLC BEH Amide (150 x 2.1mm,1.7um)","SOLVENT_A":"100% water; 10mM ammonium formate; 0.125% formic acid","SOLVENT_B":"95% acetonitrile; 10mM ammonium formate; 0.125% formic acid","FLOW_GRADIENT":"Gradient elution was performed from 100% (B) at 0–2 min to 70% (B) at 7.7 min, 40% (B) at 9.5 min, 30% (B) at 10.25 min, 100% (B) at 12.75 min, isocratic until 16.75 min with a column flow of","FLOW_RATE":"0.4 mL/min.","COLUMN_TEMPERATURE":"45C"},

"ANALYSIS":{"ANALYSIS_TYPE":"MS"},

"MS":{"INSTRUMENT_NAME":"Thermo Q Exactive HF hybrid Orbitrap","INSTRUMENT_TYPE":"Orbitrap","MS_TYPE":"ESI","ION_MODE":"NEGATIVE","MS_COMMENTS":"Spectra were collected using a Thermo Q Exactive HF Hybrid Quadrupole-Orbitrap mass spectrometer in both positive and negative mode ionization (separate injections). Full MS-ddMS2 data was collected, an inclusion list was used to prioritize MS2 selection of metabolites from the in-house ‘local’ library, when additional scan bandwidth was available MS2 was collected in a data-dependent manner. Mass range was 60-900 mz, resolution was 60k (MS1) and 15k (MS2), centroid data was collected, loop count was 4, isolation window was 1.0 Da. Collision energy was NCE 20, 50, 100 eV."},

"MS_METABOLITE_DATA":{
"Units":"counts, height",

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