#METABOLOMICS WORKBENCH wenchaowei_20230317_231840 DATATRACK_ID:3795 STUDY_ID:ST002520 ANALYSIS_ID:AN004152 PROJECT_ID:PR001622 VERSION 1 CREATED_ON March 23, 2023, 10:08 am #PROJECT PR:PROJECT_TITLE The investigation of the role of dietary inulin in Non-alcoholic Steatohepatitis PR:PROJECT_TITLE (NASH) progression PR:PROJECT_TYPE non-targeted LC-MS analysis PR:PROJECT_SUMMARY To investigate the role of dietary fiber in non-alcoholic steatohepatitis (NASH) PR:PROJECT_SUMMARY progression, male C57 mice was randomly assigned into four groups that received PR:PROJECT_SUMMARY normal chow diet (NCD), choline deficient high fat diet (CDHFD), CDHFD + 10% PR:PROJECT_SUMMARY inulin (CDHFD-I), CDHFD + 10% Cellulose (CDHFD-C). Mice stool and portal vein PR:PROJECT_SUMMARY serum was collected. Additionally, some mice received inulin or cellulose was PR:PROJECT_SUMMARY treated with 13C labelled fiber for 36 hours. Metabolites were analyzed by PR:PROJECT_SUMMARY non-targeted metabolomics. PR:INSTITUTE The Chinese University of Hong Kong PR:LAST_NAME Wei PR:FIRST_NAME Wenchao PR:ADDRESS Prince of Wales Hospital PR:EMAIL 1155118538@link.cuhk.edu.hk PR:PHONE 56001913 #STUDY ST:STUDY_TITLE The investigation of the role of dietary inulin in Non-alcoholic Steatohepatitis ST:STUDY_TITLE (NASH) progression with portal vein serum metabolites ST:STUDY_SUMMARY To investigate the role of dietary fiber in non-alcoholic steatohepatitis (NASH) ST:STUDY_SUMMARY progression, male C57 mice was randomly assigned into four groups that received ST:STUDY_SUMMARY normal chow diet (NCD), choline deficient high fat diet (CDHFD), CDHFD + 10% ST:STUDY_SUMMARY inulin (CDHFD-I), CDHFD + 10% Cellulose (CDHFD-C). Mice portal vein serum was ST:STUDY_SUMMARY collected.Results shown that the fecal metabolites from Inulin treatment group ST:STUDY_SUMMARY signficantly distinguished from that from CDHFD only group, while only ST:STUDY_SUMMARY negligible alterations was induced by cellulose treatment, indicating that ST:STUDY_SUMMARY inulin not cellulose could be well fermented by gut microbiota. By overlaping ST:STUDY_SUMMARY with fecal metabolites, the inulin derived pentadecaonic acid was consistently ST:STUDY_SUMMARY enriched in portal vein serum, which was reported as a beneficial metabolite.The ST:STUDY_SUMMARY results indicate that inulin suppresses NASH may through direct modulating gut ST:STUDY_SUMMARY metabolites. ST:INSTITUTE The Chinese University of Hong Kong ST:LAST_NAME Wei ST:FIRST_NAME Wenchao ST:ADDRESS Prince of Wales Hospital ST:EMAIL 1155118538@link.cuhk.edu.hk ST:STUDY_TYPE non-targeted LC-Mass analysis ST:PHONE 56001913 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN C57 BL/6 SU:AGE_OR_AGE_RANGE 23-24 weeks SU:WEIGHT_OR_WEIGHT_RANGE 30-60g SU:HEIGHT_OR_HEIGHT_RANGE N/A SU:GENDER Male SU:ANIMAL_ANIMAL_SUPPLIER The Laboratory Animal Services Centre in the Chinese univeristy of Hong Kong SU:ANIMAL_HOUSING The Laboratory Animal Services Centre in the Chinese univeristy of Hong Kong SU:ANIMAL_LIGHT_CYCLE 12/12 SU:ANIMAL_FEED libitum SU:ANIMAL_WATER libitum #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 - X3771neg Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3771neg.mzML SUBJECT_SAMPLE_FACTORS - X3772neg Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3772neg.mzML SUBJECT_SAMPLE_FACTORS - X3773neg Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3773neg.mzML SUBJECT_SAMPLE_FACTORS - X3774neg Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3774neg.mzML SUBJECT_SAMPLE_FACTORS - X3775neg Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3775neg.mzML SUBJECT_SAMPLE_FACTORS - X3777neg Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3777neg.mzML SUBJECT_SAMPLE_FACTORS - X3780neg Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3780neg.mzML SUBJECT_SAMPLE_FACTORS - X3781neg Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3781neg.mzML SUBJECT_SAMPLE_FACTORS - X3784neg Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3784neg.mzML SUBJECT_SAMPLE_FACTORS - X3785neg Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3785neg.mzML SUBJECT_SAMPLE_FACTORS - X3786neg Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3786neg.mzML SUBJECT_SAMPLE_FACTORS - X3787neg Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3787neg.mzML SUBJECT_SAMPLE_FACTORS - X3788neg Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3788neg.mzML SUBJECT_SAMPLE_FACTORS - X3790neg Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3790neg.mzML SUBJECT_SAMPLE_FACTORS - X3791neg Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3791neg.mzML SUBJECT_SAMPLE_FACTORS - X3792neg Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3792neg.mzML SUBJECT_SAMPLE_FACTORS - X3793neg Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3793neg.mzML SUBJECT_SAMPLE_FACTORS - X3794neg Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3794neg.mzML SUBJECT_SAMPLE_FACTORS - X3796neg Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3796neg.mzML SUBJECT_SAMPLE_FACTORS - X3800neg Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3800neg.mzML SUBJECT_SAMPLE_FACTORS - X3717neg Treatment:NCD Sample type=Mice serum; RAW_FILE_NAME=X3717neg.mzML SUBJECT_SAMPLE_FACTORS - X3718neg Treatment:NCD Sample type=Mice serum; RAW_FILE_NAME=X3718neg.mzML SUBJECT_SAMPLE_FACTORS - X3725neg Treatment:NCD Sample type=Mice serum; RAW_FILE_NAME=X3725neg.mzML SUBJECT_SAMPLE_FACTORS - X3727neg Treatment:NCD Sample type=Mice serum; RAW_FILE_NAME=X3727neg.mzML SUBJECT_SAMPLE_FACTORS - X3729neg Treatment:NCD Sample type=Mice serum; RAW_FILE_NAME=X3729neg.mzML SUBJECT_SAMPLE_FACTORS - QC01neg Treatment:QC Sample type=Mice serum; RAW_FILE_NAME=QC01neg.mzML SUBJECT_SAMPLE_FACTORS - QC02neg Treatment:QC Sample type=Mice serum; RAW_FILE_NAME=QC02neg.mzML SUBJECT_SAMPLE_FACTORS - QC03neg Treatment:QC Sample type=Mice serum; RAW_FILE_NAME=QC03neg.mzML SUBJECT_SAMPLE_FACTORS - QC04neg Treatment:QC Sample type=Mice serum; RAW_FILE_NAME=QC04neg.mzML SUBJECT_SAMPLE_FACTORS - QC05neg Treatment:QC Sample type=Mice serum; RAW_FILE_NAME=QC05neg.mzML SUBJECT_SAMPLE_FACTORS - X3771pos Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3771pos.mzML SUBJECT_SAMPLE_FACTORS - X3772pos Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3772pos.mzML SUBJECT_SAMPLE_FACTORS - X3773pos Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3773pos.mzML SUBJECT_SAMPLE_FACTORS - X3774pos Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3774pos.mzML SUBJECT_SAMPLE_FACTORS - X3775pos Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3775pos.mzML SUBJECT_SAMPLE_FACTORS - X3777pos Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3777pos.mzML SUBJECT_SAMPLE_FACTORS - X3780pos Treatment:CDHFD Sample type=Mice serum; RAW_FILE_NAME=X3780pos.mzML SUBJECT_SAMPLE_FACTORS - X3781pos Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3781pos.mzML SUBJECT_SAMPLE_FACTORS - X3784pos Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3784pos.mzML SUBJECT_SAMPLE_FACTORS - X3785pos Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3785pos.mzML SUBJECT_SAMPLE_FACTORS - X3786pos Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3786pos.mzML SUBJECT_SAMPLE_FACTORS - X3787pos Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3787pos.mzML SUBJECT_SAMPLE_FACTORS - X3788pos Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3788pos.mzML SUBJECT_SAMPLE_FACTORS - X3790pos Treatment:CDHFD-I Sample type=Mice serum; RAW_FILE_NAME=X3790pos.mzML SUBJECT_SAMPLE_FACTORS - X3791pos Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3791pos.mzML SUBJECT_SAMPLE_FACTORS - X3792pos Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3792pos.mzML SUBJECT_SAMPLE_FACTORS - X3793pos Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3793pos.mzML SUBJECT_SAMPLE_FACTORS - X3794pos Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3794pos.mzML SUBJECT_SAMPLE_FACTORS - X3796pos Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3796pos.mzML SUBJECT_SAMPLE_FACTORS - X3800pos Treatment:CDHFD-C Sample type=Mice serum; RAW_FILE_NAME=X3800pos.mzML SUBJECT_SAMPLE_FACTORS - X3717pos Treatment:NCD Sample type=Mice serum; RAW_FILE_NAME=X3717pos.mzML SUBJECT_SAMPLE_FACTORS - X3718pos Treatment:NCD Sample type=Mice serum; RAW_FILE_NAME=X3718pos.mzML SUBJECT_SAMPLE_FACTORS - X3725pos Treatment:NCD Sample type=Mice serum; RAW_FILE_NAME=X3725pos.mzML SUBJECT_SAMPLE_FACTORS - X3727pos Treatment:NCD Sample type=Mice serum; RAW_FILE_NAME=X3727pos.mzML SUBJECT_SAMPLE_FACTORS - X3729pos Treatment:NCD Sample type=Mice serum; RAW_FILE_NAME=X3729pos.mzML SUBJECT_SAMPLE_FACTORS - QC01pos Treatment:QC Sample type=Mice serum; RAW_FILE_NAME=QC01pos.mzML SUBJECT_SAMPLE_FACTORS - QC02pos Treatment:QC Sample type=Mice serum; RAW_FILE_NAME=QC02pos.mzML SUBJECT_SAMPLE_FACTORS - QC03pos Treatment:QC Sample type=Mice serum; RAW_FILE_NAME=QC03pos.mzML SUBJECT_SAMPLE_FACTORS - QC04pos Treatment:QC Sample type=Mice serum; RAW_FILE_NAME=QC04pos.mzML SUBJECT_SAMPLE_FACTORS - QC05pos Treatment:QC Sample type=Mice serum; RAW_FILE_NAME=QC05pos.mzML #COLLECTION CO:COLLECTION_SUMMARY Mice portal vein serum were collected under anaesthesia. Serum was processed by CO:COLLECTION_SUMMARY centrifugation with 3000 rpm for 15 min after coagulation. Processed serum was CO:COLLECTION_SUMMARY stored in liquid nitrogen immediately and stored in -80°C fridge. CO:COLLECTION_PROTOCOL_FILENAME Protocol_of_the_role_of_dietary_inulin_in_NASH_progression_portal_vein_serum.docx CO:SAMPLE_TYPE Blood (serum) CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY Male C57 mice was randomly assigned into 4 groups that received Normal chow diet TR:TREATMENT_SUMMARY (NCD), Choline deficient high fat diet (CDHFD), CDHFD + 10% inulin (CDHFD-I), TR:TREATMENT_SUMMARY CDHFD + 10% Cellulose (CDHFD-C). Mice portal vein serum were collected. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY 50 μL of sample was transferred to an EP tube, and 200 μL extract solution SP:SAMPLEPREP_SUMMARY (acetonitrile: methanol = 1: 1) containing internal standard SP:SAMPLEPREP_SUMMARY (L-2-Chlorophenylalanine, 2 μg/mL) was added. After 1 min vortex, the samples SP:SAMPLEPREP_SUMMARY were sonicated for 10 min in ice-water bath. Then the samples were incubated at SP:SAMPLEPREP_SUMMARY -40 ℃ for 1 h and centrifuged at 12000 rpm for 15 min at 4 ℃. 200 μL of SP:SAMPLEPREP_SUMMARY supernatant was transferred to a fresh tube and dried in a vacuum concentrator SP:SAMPLEPREP_SUMMARY at 37 ℃. Then, the dried samples were reconstituted in 200 μL of 50% SP:SAMPLEPREP_SUMMARY acetonitrile by sonication on ice for 10 min. The constitution was then SP:SAMPLEPREP_SUMMARY centrifuged at 13000 rpm for 15 min at 4 ℃, and 75 μL of supernatant was SP:SAMPLEPREP_SUMMARY transferred to a fresh glass vial for LC/MS analysis. The quality control (QC) SP:SAMPLEPREP_SUMMARY sample was prepared by mixing an equal aliquot of the supernatants from all of SP:SAMPLEPREP_SUMMARY the samples. SP:SAMPLEPREP_PROTOCOL_FILENAME Protocol_of_the_role_of_dietary_inulin_in_NASH_progression_portal_vein_serum.docx #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Non-targeted metabolomic analysis was performed on a 1290 Infinity series UHPLC CH:CHROMATOGRAPHY_SUMMARY System (Agilent Technologies) coupled to a Triple TOF 6600 mass spectrometer (AB CH:CHROMATOGRAPHY_SUMMARY Sciex ) equipped with an electrospray ionization (ESI) source by Biotree company CH:CHROMATOGRAPHY_SUMMARY (Shanghai, China). Briefly, chromatographic separation was achieved by using a CH:CHROMATOGRAPHY_SUMMARY Waters BEH Amide column (2.1 * 100 mm, 1.7 µm) under 25 ℃. The mobile phase CH:CHROMATOGRAPHY_SUMMARY consisted of water with 25mM ammonium acetate and 25 mM ammonium hydroxide (A) CH:CHROMATOGRAPHY_SUMMARY and acetonitrile (B). The flow rate was 0.5 ml/min. CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME Agilent 1290 Infinity CH:COLUMN_NAME Waters ACQUITY UPLC BEH Amide (100 x 2.1mm,1.7um) CH:SOLVENT_A 100% water; 25mM ammonium acetate; 25 mM ammonium hydroxide CH:SOLVENT_B 100% acetonitrile CH:FLOW_GRADIENT 0~0.5 min, 95% B; 0.5~7 min, 95%~65% B; 7~8 min, 65%~40% B; 8~9 min, 40% B; CH:FLOW_GRADIENT 9~9.1 min, 40%~95% B; 9.1~12 min, 95% B CH:FLOW_RATE 0.5 ml/min. CH:COLUMN_TEMPERATURE 25 ℃ CH:METHODS_FILENAME Protocol_of_the_role_of_dietary_inulin_in_NASH_progression_portal_vein_serum.docx #ANALYSIS AN:ANALYSIS_TYPE MS AN:ANALYSIS_PROTOCOL_FILE Protocol_of_the_role_of_dietary_inulin_in_NASH_progression_portal_vein_serum.docx #MS MS:INSTRUMENT_NAME ABI Sciex 6600 TripleTOF MS:INSTRUMENT_TYPE Triple TOF MS:MS_TYPE ESI MS:ION_MODE NEGATIVE MS:MS_COMMENTS MS analysis was operated in information-dependent basis (IDA) mode under MS:MS_COMMENTS negative modes. The 12 most abundant ions were selected, fragmented and sent to MS:MS_COMMENTS TOF mass analyzer to acquire the MS2 information. MS:MS_RESULTS_FILE ST002520_AN004152_Results.txt UNITS:Peak ion intensity Has m/z:Yes Has RT:Yes RT units:Seconds #END