#METABOLOMICS WORKBENCH Karin_20210630_021601_mwtab.txt DATATRACK_ID:2726 STUDY_ID:ST001883 ANALYSIS_ID:AN003047 PROJECT_ID:PR001187 VERSION 1 CREATED_ON July 24, 2021, 2:31 pm #PROJECT PR:PROJECT_TITLE Ndufs4 KO mouse urine metabolomics PR:PROJECT_TYPE Multi-platform metabolomics analysis of Ndufs4 KO mouse urine PR:PROJECT_SUMMARY Multi-platform metabolomics analysis of urine from the Ndufs4 knockout PR:PROJECT_SUMMARY (Ndufs4-/-) mouse model of human Leigh syndrome PR:INSTITUTE North-West University PR:LAST_NAME Louw PR:FIRST_NAME Roan PR:ADDRESS Hofman Street PR:EMAIL Roan.Louw@nwu.ac.za PR:PHONE +27 18 299 4074 #STUDY ST:STUDY_TITLE Metabolomics of Ndufs4 KO urine (part - I) ST:STUDY_SUMMARY Untargeted GC-TOF-MS analysis of Ndufs4 KO and WT mouse urine ST:INSTITUTE North-West University ST:LAST_NAME Louw ST:FIRST_NAME Roan ST:ADDRESS Hofman Street ST:EMAIL Roan.Louw@nwu.ac.za ST:PHONE +27 18 299 4074 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN Ndufs4, https://www.jax.org/strai n/02705 8 SU:AGE_OR_AGE_RANGE 45-50 days SU:GENDER Male SU:ANIMAL_LIGHT_CYCLE 12:12 h SU:ANIMAL_FEED Rodent Breeder, Cat. #RM1845, LabChef, Nutritionhub SU:ANIMAL_WATER ad libitum SU:ANIMAL_ANIMAL_SUPPLIER Jackson Laboratory (ME, USA) #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 - 57_3l Genotype:WT RAW_FILE_NAME=netCDF57.3l.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 9_3d Genotype:WT RAW_FILE_NAME=netCDF9.3d.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 82_2g Genotype:WT RAW_FILE_NAME=netCDF82.2g.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 88_3q Genotype:WT RAW_FILE_NAME=netCDF88.3q .WT_1.cdf SUBJECT_SAMPLE_FACTORS - 9_4j Genotype:WT RAW_FILE_NAME=netCDF9.4j.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 49_2a Genotype:WT RAW_FILE_NAME=netCDF49.2a.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 37_4p Genotype:WT RAW_FILE_NAME=netCDF37.4p.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 27_4n Genotype:WT RAW_FILE_NAME=netCDF27.4n.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 91_2i Genotype:WT RAW_FILE_NAME=netCDF91.2i.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 16_4m Genotype:WT RAW_FILE_NAME=netCDF16.4m.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 62_4b Genotype:WT RAW_FILE_NAME=netCDF62.4b.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 83_3p Genotype:WT RAW_FILE_NAME=netCDF83.3p.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 8_1v Genotype:WT RAW_FILE_NAME=netCDF8.1v.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 42_3k Genotype:WT RAW_FILE_NAME=netCDF42.3k.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 68_3n Genotype:WT RAW_FILE_NAME=netCDF68.3n.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 97_4h Genotype:WT RAW_FILE_NAME=netCDF97.4h.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 98_4h Genotype:WT RAW_FILE_NAME=netCDF98.4h.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 54_2c Genotype:WT RAW_FILE_NAME=netCDF54.2c.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 26_3g Genotype:WT RAW_FILE_NAME=netCDF26.3g.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 87_3q Genotype:WT RAW_FILE_NAME=netCDF87.3q.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 63_4b Genotype:WT RAW_FILE_NAME=netCDF63.4b.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 81_3p Genotype:WT RAW_FILE_NAME=netCDF81.3p.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 65_2u Genotype:WT RAW_FILE_NAME=netCDF65.2u.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 53_2c Genotype:WT RAW_FILE_NAME=netCDF53.2c.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 29_4n Genotype:WT RAW_FILE_NAME=netCDF29.4n.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 61_2e Genotype:WT RAW_FILE_NAME=netCDF61.2e.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 11_3d Genotype:WT RAW_FILE_NAME=netCDF11.3d.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 65_3n Genotype:WT RAW_FILE_NAME=netCDF65.3n.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 62_2t Genotype:WT RAW_FILE_NAME=netCDF62.2t.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 2_1u Genotype:WT RAW_FILE_NAME=netCDF2.1u.WT_1.cdf SUBJECT_SAMPLE_FACTORS - 67_2f Genotype:KO RAW_FILE_NAME=netCDF67.2f.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 18_2l Genotype:KO RAW_FILE_NAME=netCDF18.2l.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 60_2e Genotype:KO RAW_FILE_NAME=netCDF60.2e.KO_2.cdf SUBJECT_SAMPLE_FACTORS - 68_1o Genotype:KO RAW_FILE_NAME=netCDF68.1o.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 44_3k Genotype:KO RAW_FILE_NAME=netCDF44.3k.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 2_3s Genotype:KO RAW_FILE_NAME=netCDF2.3s.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 86_3q Genotype:KO RAW_FILE_NAME=netCDF86.3q.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 96_2i Genotype:KO RAW_FILE_NAME=netCDF96.2i.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 42_4q Genotype:KO RAW_FILE_NAME=netCDF42.4q.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 20_4m Genotype:KO RAW_FILE_NAME=netCDF20.4m.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 8_4j Genotype:KO RAW_FILE_NAME=netCDF8.4j.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 69_2f Genotype:KO RAW_FILE_NAME=netCDF69.2f.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 37_3j Genotype:KO RAW_FILE_NAME=netCDF37.3j.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 10_1v Genotype:KO RAW_FILE_NAME=netCDF10.1v.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 33_2o Genotype:KO RAW_FILE_NAME=netCDF33.2o.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 45_3k Genotype:KO RAW_FILE_NAME=netCDF45.3k.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 54_4a Genotype:KO RAW_FILE_NAME=netCDF54.4a.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 89_1r Genotype:KO RAW_FILE_NAME=netCDF89.1r.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 56_2d Genotype:KO RAW_FILE_NAME=netCDF56.2d.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 28_3g Genotype:KO RAW_FILE_NAME=netCDF28.3g.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 34_4o Genotype:KO RAW_FILE_NAME=netCDF34.4o.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 50_4s Genotype:KO RAW_FILE_NAME=netCDF50.4s.KO_1.cdf SUBJECT_SAMPLE_FACTORS - 2_4i Genotype:KO RAW_FILE_NAME=netCDF2.4i.KO_1.cdf #COLLECTION CO:COLLECTION_SUMMARY Urine was collected from individual mice in metabolic cages after overnight (12 CO:COLLECTION_SUMMARY hours) fasting with ad libitum access to water. Since Ndufs4 KO mice yielded CO:COLLECTION_SUMMARY limited urine volumes, urine droplets that failed to reach the collection tube CO:COLLECTION_SUMMARY were also collected. Hence, the separating cone, collection funnel and urine CO:COLLECTION_SUMMARY collection tubes were all treated with sodium azide (NaN3; 0.1% w/v; WT: 100 μL CO:COLLECTION_SUMMARY and KO: 50 μL) to prevent bacterial growth. Samples were snap-frozen in liquid CO:COLLECTION_SUMMARY nitrogen upon collection and stored at −80 °C until used. CO:SAMPLE_TYPE Urine CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY The animals did not receive any treatment #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Pre-determined volumes of urine, containing 0.0625 μmole creatinine, were SP:SAMPLEPREP_SUMMARY supplemented with internal standards and deproteinized using methanol. Urine SP:SAMPLEPREP_SUMMARY samples were then dried under nitrogen and derivatized (methoxymated and SP:SAMPLEPREP_SUMMARY silylated) prior to analysis. SP:SAMPLEPREP_PROTOCOL_FILENAME GC_sample_prep_protocol.pdf #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Agilent 7890A CH:COLUMN_NAME Restek Rtx-5Sil MS (30 x 0.25mm, 0.25um) CH:METHODS_FILENAME GC_data_acquisition_method.pdf #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Leco GC-TOF MS:INSTRUMENT_TYPE GC-TOF MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:MS_COMMENTS The LECO Corporation ChromaTOF® software (v 4.5x) was used for data acquisition MS:MS_COMMENTS and extraction. This included automatic baseline removal via the “spanning” MS:MS_COMMENTS tracking method (offset of 1; just above the noise) and auto smoothing, with the MS:MS_COMMENTS software’s Statistical Compare feature used to align peaks. Spectral matching MS:MS_COMMENTS was done using the NIST11 commercial library and an in-house mass spectral MS:MS_COMMENTS library in order to identify important analytes. MS:MS_RESULTS_FILE ST001883_AN003047_Results.txt UNITS:mg/g creatinine Has m/z:Yes Has RT:Yes RT units:Seconds #END