{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST001667","ANALYSIS_ID":"AN002719","VERSION":"1","CREATED_ON":"12-15-2022"},

"PROJECT":{"PROJECT_TITLE":"NMR and MS Metabolomics Reveal a Distinct Metabolic State of Ricin-induced Hypoglycemia","PROJECT_TYPE":"Targeted NMR and Untargeted MS","PROJECT_SUMMARY":"Ricin toxin is a ribosome inactivating protein. Due to its toxic and chemical properties, ricin is a potential agent of bioterrorism but has also been studied for therapeutic use in immunotoxins. Previous research by our group has demonstrated lethal hypoglycemia associated with ricin toxicity. Research efforts have focused on better understanding this ricin-induced metabolic state of hypoglycemia to aid in better understanding the systemic effects of hypoglycemia, and the use of ricin in immunotherapy. Here, we have used a mouse model to characterize liver metabolome changes associated with hypoglycemia induced by two conditions. Mice were challenged with intraperitoneal injections of ricin at high and low doses for 2 hrs, 8 hrs, and overnight. To understand how ricin-induced hypoglycemia differs from that of fasting, another group of mice had food withheld for 8-hours and overnight. 1H NMR-based metabolomics was performed on polar molecules extracted from mouse livers, with metabolites annotated using Chenomx software. MetaboAnalyst was employed for multivariate statistical analysis. In this study, similar decreases in blood glucose in mice were observed following injection of a lethal dose of ricin and with overnight fasting. NMR analyses identified 59 polar metabolites present in mice livers from all treatments. Multivariate statistical analyses were used to evaluate global metabolic state differences. Results from these analyses indicated that the profiled liver metabolomes for mice subjected to the two conditions differ significantly at both 8 hours and overnight. Additionally, ricin treatment with a lethal dose reveal a progression of metabolic changes over time, from 2 to 22 hours. Additionally, mass spectrometry supported these findings, and NMR analyses revealed key metabolites that contribute to these differences. While both ricin and fasting induce hypoglycemia, the metabolic states resulting from these two conditions are different. Further analyses may give insights into mechanisms of ricin toxicity, specific metabolic pathways that are altered, and potential treatments for hypoglycemia. We propose to extend these studies to insulin-induced hypoglycemia.","INSTITUTE":"Montana State University","DEPARTMENT":"Chemistry & Biochemistry","LABORATORY":"Copié","LAST_NAME":"Kempa","FIRST_NAME":"Jake","ADDRESS":"103 Chemistry and Biochemistry Building, Bozeman, Montana, 59717, USA","EMAIL":"jkempa97@gmail.com","PHONE":"4067997200","DOI":"http://dx.doi.org/10.21228/M8NM4H"},

"STUDY":{"STUDY_TITLE":"LC-MS metabolomics analysis of ricin-induced and fasting hypoglycemia (part-II)","STUDY_SUMMARY":"Mice were subjected to ricin exposure or fasting conditions for 2 hours, 8 hours, or an overnight period. Following treatment, livers were removed and metabolites were extracted and analyzed by LC-MS.","INSTITUTE":"Montana State University","LAST_NAME":"Kempa","FIRST_NAME":"Jake","ADDRESS":"103 Chemistry and Biochemistry Building, Bozeman, Montana, 59717, USA","EMAIL":"jkempa97@gmail.com","PHONE":"4067997200","SUBMIT_DATE":"2021-01-19"},

"SUBJECT":{"SUBJECT_TYPE":"Mammal","SUBJECT_SPECIES":"Mus musculus","TAXONOMY_ID":"10090"},
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{
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{
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{
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{
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{
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},
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{
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{
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{
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{
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{
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{
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{
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},
{
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{
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{
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{
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{
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"Sample ID":"8--7",
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"Additional sample data":{"RAW_FILE_NAME":"8--7"}
},
{
"Subject ID":"-",
"Sample ID":"8--8",
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"Additional sample data":{"RAW_FILE_NAME":"8--8"}
},
{
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"Sample ID":"8--9",
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{
"Subject ID":"-",
"Sample ID":"4--1",
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"Additional sample data":{"RAW_FILE_NAME":"4--1"}
},
{
"Subject ID":"-",
"Sample ID":"4--10",
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"Additional sample data":{"RAW_FILE_NAME":"4--10"}
},
{
"Subject ID":"-",
"Sample ID":"4--2",
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"Additional sample data":{"RAW_FILE_NAME":"4--2"}
},
{
"Subject ID":"-",
"Sample ID":"4--3",
"Factors":{"Treatment":"Low Dose Ricin","Timepoint":"Overnight"},
"Additional sample data":{"RAW_FILE_NAME":"4--3"}
},
{
"Subject ID":"-",
"Sample ID":"4--4",
"Factors":{"Treatment":"Low Dose Ricin","Timepoint":"Overnight"},
"Additional sample data":{"RAW_FILE_NAME":"4--4"}
},
{
"Subject ID":"-",
"Sample ID":"4--5",
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"Additional sample data":{"RAW_FILE_NAME":"4--5"}
},
{
"Subject ID":"-",
"Sample ID":"4--6",
"Factors":{"Treatment":"Low Dose Ricin","Timepoint":"Overnight"},
"Additional sample data":{"RAW_FILE_NAME":"4--6"}
},
{
"Subject ID":"-",
"Sample ID":"4--7",
"Factors":{"Treatment":"Low Dose Ricin","Timepoint":"Overnight"},
"Additional sample data":{"RAW_FILE_NAME":"4--7"}
},
{
"Subject ID":"-",
"Sample ID":"4--8",
"Factors":{"Treatment":"Low Dose Ricin","Timepoint":"Overnight"},
"Additional sample data":{"RAW_FILE_NAME":"4--8"}
},
{
"Subject ID":"-",
"Sample ID":"4--9",
"Factors":{"Treatment":"Low Dose Ricin","Timepoint":"Overnight"},
"Additional sample data":{"RAW_FILE_NAME":"4--9"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"Livers were removed from mice immediately following sacrifice and stored at -80 C. Before metabolite extraction, livers were homogenized into powder with mortar and pestle over liquid nitrogen.","SAMPLE_TYPE":"Liver"},

"TREATMENT":{"TREATMENT_SUMMARY":"Mice were injected via intraperitoneal (IP) route with either ricin or saline in 10 microliter per g body weight. Mice in all but the fasted group were given food and water ad libitum. Fasted mice were transferred to new cages with fresh bedding where they had food withheld while given water ad libitum. Mice were sacrificed at 2h (ricin), 8h (ricin and fasted), and overnight (16-22h, control, ricin, fasted)."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"Microcentrifuge tubes with 30 mg of frozen liver tissue had 500 uL of ice-cold acetone and three Zirconia beads added to them. Tissues were lysed and homogenized, then sonicated and centrifuged. The supernatant was transferred to a new tube and 500 uL of ice-cold MeOH/H2O was added to the pellet. The pellet was sonicated and centrifuged again. The supernatant was combined with the acetone fraction and vacuum dried. Dried pellets were resuspended in acetonitrile/H2O with 1% Formic Acid, vortexed, sonicated, and centrifuged. 180 uL of eluate was used for LCMS analysis."},

"CHROMATOGRAPHY":{"INSTRUMENT_NAME":"Agilent 1290","COLUMN_NAME":"Waters Acquity BEH HILIC (150 x 2.1mm,1.7um)","COLUMN_PRESSURE":"600 bar","COLUMN_TEMPERATURE":"40","FLOW_GRADIENT":"100% B, 55% B, 20% B, 100%B","FLOW_RATE":"0.400 mL/min","RETENTION_TIME":"25 min","SOLVENT_A":"100% water; 0.1% formic acid","SOLVENT_B":"100% acetonitrile; 0.1% formic acid","CHROMATOGRAPHY_TYPE":"HILIC"},

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

"MS":{"INSTRUMENT_NAME":"Agilent 6530 QTOF","INSTRUMENT_TYPE":"QTOF","MS_TYPE":"ESI","MS_COMMENTS":"mzML formatting acquired in ProteoWizard msconvert, Feature analysis performed in XCMS Online v3.7.1","ION_MODE":"POSITIVE","MS_RESULTS_FILE":"ST001667_AN002719_Results.txt UNITS:Intensity Has m/z:Yes Has RT:Yes RT units:Minutes"}

}