#METABOLOMICS WORKBENCH Liyuan_115_20250929_000638 DATATRACK_ID:6501 STUDY_ID:ST004259 ANALYSIS_ID:AN007088 PROJECT_ID:PR002687 VERSION 1 CREATED_ON October 4, 2025, 3:29 am #PROJECT PR:PROJECT_TITLE Phytate enhances gut Parasutterella colonization to alleviate radiation injury PR:PROJECT_TITLE via anti-inflammatory and antioxidant effects PR:PROJECT_SUMMARY Food as medicine shows promise for disease intervention or treatment. Here, we PR:PROJECT_SUMMARY found phytate, an active ingredient of plant-based diets, exhibits properties in PR:PROJECT_SUMMARY mitigating radiotherapy-related complications. Oral gavage of phytate restored PR:PROJECT_SUMMARY hematogenic organ atrophy, elevated peripheral blood neutrophils and white blood PR:PROJECT_SUMMARY cells, reduced inflammation, and improved gastrointestinal (GI) integrity in PR:PROJECT_SUMMARY irradiated mice. Phytate intake modulated the gut microbiota, facilitating the PR:PROJECT_SUMMARY colonization of symbiotic Parasutterella in GI tract, thus combating intestinal PR:PROJECT_SUMMARY radiation toxicity. In vitro assays and untargeted metabolomics identified PR:PROJECT_SUMMARY 3-Phenyllactic acid (PLA) and N-Acetyl-L-leucine (NL) as functional metabolites PR:PROJECT_SUMMARY produced by Parasutterella. In vitro, ex vivo, and in vivo models showed that PR:PROJECT_SUMMARY PLA induces M2-like polarization in macrophages, while NL reduced oxidative PR:PROJECT_SUMMARY stress, both counteracting radiation toxicity and working synergistically. Our PR:PROJECT_SUMMARY findings offer mechanistic insights into phytate for alleviating PR:PROJECT_SUMMARY radiation-associated complications and suggest that Parasutterella and its PR:PROJECT_SUMMARY metabolites might be employed as promising probiotics or postbiotics for cancer PR:PROJECT_SUMMARY patients with radiotherapy. PR:INSTITUTE Chinese Academy of Medical Sciences PR:LAST_NAME Li PR:FIRST_NAME Yuan PR:ADDRESS Baidi Road, Tianjin, Tianjin, 300192, China PR:EMAIL liyuan@irm-cams.ac.cn PR:PHONE +8615900383739 #STUDY ST:STUDY_TITLE Phytate enhances gut Parasutterella colonization to alleviate radiation injury ST:STUDY_TITLE via anti-inflammatory and antioxidant effects ST:STUDY_SUMMARY Food as medicine shows promise for disease intervention or treatment. Here, we ST:STUDY_SUMMARY found phytate, an active ingredient of plant-based diets, exhibits properties in ST:STUDY_SUMMARY mitigating radiotherapy-related complications. Oral gavage of phytate restored ST:STUDY_SUMMARY hematogenic organ atrophy, elevated peripheral blood neutrophils and white blood ST:STUDY_SUMMARY cells, reduced inflammation, and improved gastrointestinal (GI) integrity in ST:STUDY_SUMMARY irradiated mice. Phytate intake modulated the gut microbiota, facilitating the ST:STUDY_SUMMARY colonization of symbiotic Parasutterella in GI tract, thus combating intestinal ST:STUDY_SUMMARY radiation toxicity. In vitro assays and untargeted metabolomics identified ST:STUDY_SUMMARY 3-Phenyllactic acid (PLA) and N-Acetyl-L-leucine (NL) as functional metabolites ST:STUDY_SUMMARY produced by Parasutterella. In vitro, ex vivo, and in vivo models showed that ST:STUDY_SUMMARY PLA induces M2-like polarization in macrophages, while NL reduced oxidative ST:STUDY_SUMMARY stress, both counteracting radiation toxicity and working synergistically. Our ST:STUDY_SUMMARY findings offer mechanistic insights into phytate for alleviating ST:STUDY_SUMMARY radiation-associated complications and suggest that Parasutterella and its ST:STUDY_SUMMARY metabolites might be employed as promising probiotics or postbiotics for cancer ST:STUDY_SUMMARY patients with radiotherapy. ST:INSTITUTE Chinese Academy of Medical Sciences ST:LAST_NAME Li ST:FIRST_NAME Yuan ST:ADDRESS Baidi Road, Tianjin, Tianjin, 300192, China ST:EMAIL liyuan@irm-cams.ac.cn ST:PHONE +8615900383739 #SUBJECT SU:SUBJECT_TYPE Bacteria SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 #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 - PE0h-1 Treatment:Sample collection after 0h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP1_FZTM220089101-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN1_FZTM220089101-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE0h-2 Treatment:Sample collection after 0h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP1_FZTM220089102-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN1_FZTM220089102-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE0h-3 Treatment:Sample collection after 0h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP1_FZTM220089103-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN1_FZTM220089103-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE0h-4 Treatment:Sample collection after 0h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP1_FZTM220089104-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN1_FZTM220089104-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE0h-5 Treatment:Sample collection after 0h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP1_FZTM220089105-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN1_FZTM220089105-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE0h-6 Treatment:Sample collection after 0h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP1_FZTM220089106-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN1_FZTM220089106-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE6h-1 Treatment:Sample collection after 6h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP2_FZTM220089107-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN2_FZTM220089107-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE6h-2 Treatment:Sample collection after 6h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP2_FZTM220089108-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN2_FZTM220089108-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE6h-3 Treatment:Sample collection after 6h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP2_FZTM220089109-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN2_FZTM220089109-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE6h-4 Treatment:Sample collection after 6h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP2_FZTM220089110-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN2_FZTM220089110-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE6h-5 Treatment:Sample collection after 6h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP2_FZTM220089111-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN2_FZTM220089111-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE6h-6 Treatment:Sample collection after 6h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP2_FZTM220089112-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN2_FZTM220089112-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE24h-1 Treatment:Sample collection after 24h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP3_FZTM220089113-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN3_FZTM220089113-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE24h-2 Treatment:Sample collection after 24h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP3_FZTM220089114-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN3_FZTM220089114-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE24h-3 Treatment:Sample collection after 24h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP3_FZTM220089115-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN3_FZTM220089115-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE24h-4 Treatment:Sample collection after 24h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP3_FZTM220089116-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN3_FZTM220089116-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE24h-5 Treatment:Sample collection after 24h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP3_FZTM220089117-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN3_FZTM220089117-1A; Raw data file name=- SUBJECT_SAMPLE_FACTORS - PE24h-6 Treatment:Sample collection after 24h of cultivation | Sample source:Culture supernatant RAW_FILE_NAME()=HFX8_CP3_FZTM220089118-1A; RAW_FILE_NAME(Raw data file name)=HFX8_CN3_FZTM220089118-1A; Raw data file name=- #COLLECTION CO:COLLECTION_SUMMARY Culture supernatants of Parasutterella excrementihominis (DSM 21040) were CO:COLLECTION_SUMMARY collected at 0, 6, and 24 hours, followed by centrifugation at 3000 rpm for 10 CO:COLLECTION_SUMMARY minutes at 4ºC. The samples were then rapidly frozen in liquid nitrogen for 15 CO:COLLECTION_SUMMARY minutes and stored at -80ºC until further use. CO:SAMPLE_TYPE Media #TREATMENT TR:TREATMENT_SUMMARY Not applicable #SAMPLEPREP SP:SAMPLEPREP_SUMMARY The samples (1 mL) were freeze-dried and resuspended with prechilled 80% SP:SAMPLEPREP_SUMMARY methanol by well vortex. Then the samples were incubated on ice for 5 min and SP:SAMPLEPREP_SUMMARY centrifuged at 15,000 g, 4°C for 15 min. Some of supernatant was diluted to SP:SAMPLEPREP_SUMMARY final concentration containing 53% methanol by LC-MS grade water. The samples SP:SAMPLEPREP_SUMMARY were subsequently transferred to a fresh Eppendorf tube and thenwere centrifuged SP:SAMPLEPREP_SUMMARY at 15000 g, 4°C for 15 min. Finally, the supernatant was injected into the SP:SAMPLEPREP_SUMMARY LC-MS/MS system analysis. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Thermo Vanquish CH:COLUMN_NAME Thermo Hypersil GOLD aQ (100 x 2.1mm,1.9um) CH:SOLVENT_A 5 mM ammonium acetate, pH9.0 CH:SOLVENT_B Methanol CH:FLOW_GRADIENT 2%B, 1.5 min;2-85% B, 3 min; 85-100% B, 10 min；100-2% B, 10.1 min；2%B, 12 CH:FLOW_GRADIENT min. CH:FLOW_RATE 0.2 mL/min CH:COLUMN_TEMPERATURE 40°C #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Thermo Q Exactive HF hybrid Orbitrap MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE ESI MS:ION_MODE NEGATIVE MS:MS_COMMENTS The raw data files generated by UHPLC-MS/MS were processed using the Compound MS:MS_COMMENTS Discoverer 3.1 (CD3.1, ThermoFisher) to perform peak alignment, peak picking, MS:MS_COMMENTS and quantitation for each metabolite. The main parameters were set as follows: MS:MS_COMMENTS retention time tolerance, 0.2 minutes; actual mass tolerance, 5ppm; signal MS:MS_COMMENTS intensity tolerance, 30%; signal/noise ratio, 3; and minimum intensity, et al. MS:MS_COMMENTS After that, peak intensities were normalized to the total spectral intensity. MS:MS_COMMENTS The normalized data was used to predict the molecular formula based on additive MS:MS_COMMENTS ions, molecular ion peaks and fragment ions. And then peaks were matched with MS:MS_COMMENTS the mzCloud(https://www.mzcloud.org/)，mzVault and Mass List database to obtain MS:MS_COMMENTS the accurate qualitative and relative quantitative results. Statistical analyses MS:MS_COMMENTS were performed using the statistical software R (R version R-3.4.3), Python MS:MS_COMMENTS (Python 2.7.6 version) and CentOS (CentOS release 6.6), When data were not MS:MS_COMMENTS normally distributed, standardize according to the formula: sample raw MS:MS_COMMENTS quantitation value / (The sum of sample metabolite quantitation value / The sum MS:MS_COMMENTS of QC1 sample metabolite quantitation value)to obtain relative peak areas; And MS:MS_COMMENTS compounds whose CVs of relative peak areas in QC samples were greater than 30% MS:MS_COMMENTS were removed, and finally the metabolites' identification and relative MS:MS_COMMENTS quantification results were obtained. MS:MS_RESULTS_FILE ST004259_AN007088_Results.txt UNITS:Peak area Has m/z:Yes Has RT:Yes RT units:Minutes #END