{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST000365","ANALYSIS_ID":"AN000598","VERSION":"1","CREATED_ON":"08-09-2023"},

"PROJECT":{"PROJECT_TITLE":"Zebrafish Metabolomics: Model for Environmental Metal Toxicity","PROJECT_SUMMARY":"This metabolomics seed project will test the hypothesis that zebrafish can provide mechanistic insights into the human health effects of developmental exposure to Cd and Pb. We will use broad spectrum metabolomics of zebrafish larvae after exposure to Cd, Pb, and Cd and Pb compared to controls. Activity observed at 5 days post fertilization is will be used to determine if there is a correlation between biological pathways implicated by these metabolic profiles and cardiovascular, metabolic (obesity), and neurological phenotypes. The behavioral phenotypes have been quantified in zebrafish previously and have been measured in the Newborn Epigenetic STudy (NEST), a NIH-funded project that is investigating how environmental exposures and nutrition, in the womb and during childhood, affect how genes work and how these exposures developed into obesity and other diseases, disorders, and conditions. The results from this study will demonstrate the power of using zebrafish as a model for mechanism discovery in exposure using metabolomics to advance understanding of early life exposure to Cd and Pb and serve as preliminary data for opportunities.","INSTITUTE":"North Carolina State University","LAST_NAME":"Mattingly","FIRST_NAME":"Carolyn","ADDRESS":"142 David Clark Labs, Raleigh, NC 27695-7617","EMAIL":"cjmattin@ncsu.edu","PHONE":"919-515-2024","DOI":"http://dx.doi.org/10.21228/M8JW2K"},

"STUDY":{"STUDY_TITLE":"Zebrafish Metabolomics: Model for Environmental Metal Toxicity","STUDY_SUMMARY":"This metabolomics seed project will test the hypothesis that zebrafish can provide mechanistic insights into the human health effects of developmental exposure to Cd and Pb. We will use broad spectrum metabolomics of zebrafish larvae after exposure to Cd, Pb, and Cd and Pb compared to controls. Activity observed at 5 days post fertilization is will be used to determine if there is a correlation between biological pathways implicated by these metabolic profiles and cardiovascular, metabolic (obesity), and neurological phenotypes. The behavioral phenotypes have been quantified in zebrafish previously and have been measured in the Newborn Epigenetic STudy (NEST), a NIH-funded project that is investigating how environmental exposures and nutrition, in the womb and during childhood, affect how genes work and how these exposures developed into obesity and other diseases, disorders, and conditions. The results from this study will demonstrate the power of using zebrafish as a model for mechanism discovery in exposure using metabolomics to advance understanding of early life exposure to Cd and Pb and serve as preliminary data for opportunities.","INSTITUTE":"University of North Carolina","DEPARTMENT":"Discovery Sciences","LABORATORY":"Sumner Lab","LAST_NAME":"Sumner","FIRST_NAME":"Susan","ADDRESS":"Eastern Regional Comprehensive Metabolomics Resource Core, UNC Nutrition Research Institute, 500 Laureate Way, Kannapolis, NC, 28081","EMAIL":"susan_sumner @unc.edu","PHONE":"704-250-5066","SUBMIT_DATE":"2016-03-09"},

"SUBJECT":{"SUBJECT_TYPE":"Zebrafish","SUBJECT_SPECIES":"Danio rerio","TAXONOMY_ID":"7955","SPECIES_GROUP":"Fish"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"-",
"Sample ID":"CM_CdPool_1",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPool_2",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPool_3",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Cdhigh_1",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"high (1000 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Cdhigh_2",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"high (1000 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Cdhigh_3",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"high (1000 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Cdlow_1",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"low (10 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Cdlow_2",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"low (10 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Cdlow_3",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"low (10 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Cdmid_1",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"mid (100 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Cdmid_2",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"mid (100 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Cdmid_3",
"Factors":{"Metal Exposure":"Cd","Exposure Level":"mid (100 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPbPool_1",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPbPool_2",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPbPool_3-R",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_TotalPool_1",
"Factors":{"Metal Exposure":"CD+Pb","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_TotalPool_2",
"Factors":{"Metal Exposure":"CD+Pb","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_TotalPool_3",
"Factors":{"Metal Exposure":"CD+Pb","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPbhigh_1",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"high (1000 ppb Cd and 1000 ppb Pb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPbhigh_2",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"high (1000 ppb Cd and 1000 ppb Pb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPbhigh_3",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"high (1000 ppb Cd and 1000 ppb Pb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPblow_1",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"low (10 ppb Cd and 10 ppb Pb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPblow_2",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"low (10 ppb Cd and 10 ppb Pb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPblow_3",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"low (10 ppb Cd and 10 ppb Pb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPBmid_1",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"mid (100 ppb Cd and 100 ppb Pb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPBmid_2",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"mid (100 ppb Cd and 100 ppb Pb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_CdPBmid_3",
"Factors":{"Metal Exposure":"Cd+Pb","Exposure Level":"mid (100 ppb Cd and 100 ppb Pb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_NoExpPool_1",
"Factors":{"Metal Exposure":"control","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_NoExpPool_2",
"Factors":{"Metal Exposure":"control","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_NoExpPool_3",
"Factors":{"Metal Exposure":"control","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_NoExp_1",
"Factors":{"Metal Exposure":"control","Exposure Level":"No"}
},
{
"Subject ID":"-",
"Sample ID":"CM_NoExp_2",
"Factors":{"Metal Exposure":"control","Exposure Level":"No"}
},
{
"Subject ID":"-",
"Sample ID":"CM_NoExp_3",
"Factors":{"Metal Exposure":"control","Exposure Level":"No"}
},
{
"Subject ID":"-",
"Sample ID":"CM_PbPool_1",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_PbPool_2",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_PbPool_3-R",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"-"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Pbhigh_1",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"high (1000 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Pbhigh_2",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"high (1000 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Pbhigh_3",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"high (1000 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Pblow_1",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"low (10 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Pblow_2",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"low (10 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Pblow_3",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"low (10 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Pbmid_1",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"mid (100 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Pbmid_2",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"mid (100 ppb)"}
},
{
"Subject ID":"-",
"Sample ID":"CM_Pbmid_3",
"Factors":{"Metal Exposure":"Pb","Exposure Level":"mid (100 ppb)"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"The larvae were collected at 48 hours post fertilization (hpf).","SAMPLE_TYPE":"Fish larvae"},

"TREATMENT":{"TREATMENT_SUMMARY":"Zebrafish larvae were exposured to Cd (10, 100, and 1000 ppb), Pb (10, 100, and 1000 ppb), and Cd and Pb (10 ppb Cd and 10 ppb Pb, 100 ppb Cd and 100 ppb Pb, and 1000 ppb Cd and 1000 ppb Pb) and compared to controls. The exposure period was between 6-48 hours post fertilization (hpf). The larvae were collected at 48 hpf."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"Pools with 40 larvae each were washed and snap frozen before being shipped to the NIH RTI-RCMRC on dry ice and immediately stored at -80 °C after being logged in for metabolomics analysis. A total of 30 study samples were lyophilized overnight for sample preparation. Each sample was mixed with 750 μl of methanol, vortexed, and centrifuged. 300 μl of the supernatant were used for the study samples, phenotypic pools were created using 350 μl of each phenotype and a total study pool was created by mixing 35 μl from each sample. Three aliquots were created from each pool for a total of 15 pooled samples. All samples were dried in vacuum overnight. Each sample was reconstituted with 250 μl of 0.5 mM phosphate buffer (pH 7.5) and 25 μl of d-DSS/Chenomx as an internal standard. The tubes were vortexed for 2 min on a multi-tube vortexer and centrifuged at 16,000 rcf for 10 min. A 200 µl aliquot of the supernatant was transferred into pre-labeled 3 mm (4”) NMR tubes for data acquisition on a 700 MHz spectrometer. 1H NMR spectra of zebrafish extraction samples were acquired on a Bruker 700 MHz NMR spectrometer (located at David H. Murdock Research Institute, Kannapolis, NC, USA) using a 5 mm cryogenically cooled ATMA inverse probe and ambient temperature of 25 ℃. A 1D NOESY presaturation pulse sequence (noesypr1d, [recycle delay (RD)-90°-t1-90°-tm-90°-acquire free induction decay (FID) was used for data acquisition. For each sample 32 transients were collected into 65k data points using a spectral width of 17.17 ppm, 2 s relaxation delay, 100 ms mixing time, and an acquisition time of 3.9 s per FID. The water resonance was suppressed using resonance irradiation during the relaxation delay and mixing time. NMR spectra were processed using TopSpin 3.2 software (Bruker-Biospin, Germany). Spectra were zero filled, and Fourier transformed after exponential multiplication with line broadening factor of 0.5. Phase and baseline of the spectra were manually corrected for each spectrum. Spectra were referenced internally to the DSS-d6 signal. The quality of each NMR spectrum was assessed for the level of noise and alignment of identified markers. Spectra were assessed for missing data and underwent quality checks."},

"CHROMATOGRAPHY":[],

"ANALYSIS":{"ANALYSIS_TYPE":"NMR"},

"NM":{"INSTRUMENT_NAME":"Bruker Avance III","INSTRUMENT_TYPE":"FT-NMR","NMR_EXPERIMENT_TYPE":"1D 1H","SPECTROMETER_FREQUENCY":"700 MHz"},

"Data":[{"Bin range(ppm)":"NMR_BINNED_DATA_END","CM_CdPool_1":"","CM_CdPool_2":"","CM_CdPool_3":"","CM_Cdhigh_1":"","CM_Cdhigh_2":"","CM_Cdhigh_3":"","CM_Cdlow_1":"","CM_Cdlow_2":"","CM_Cdlow_3":"","CM_Cdmid_1":"","CM_Cdmid_2":"","CM_Cdmid_3":"","CM_CdPbPool_1":"","CM_CdPbPool_2":"","CM_CdPbPool_3-R":"","CM_TotalPool_1":"","CM_TotalPool_2":"","CM_TotalPool_3":"","CM_CdPbhigh_1":"","CM_CdPbhigh_2":"","CM_CdPbhigh_3":"","CM_CdPblow_1":"","CM_CdPblow_2":"","CM_CdPblow_3":"","CM_CdPBmid_1":"","CM_CdPBmid_2":"","CM_CdPBmid_3":"","CM_NoExpPool_1":"","CM_NoExpPool_2":"","CM_NoExpPool_3":"","CM_NoExp_1":"","CM_NoExp_2":"","CM_NoExp_3":"","CM_PbPool_1":"","CM_PbPool_2":"","CM_PbPool_3-R":"","CM_Pbhigh_1":"","CM_Pbhigh_2":"","CM_Pbhigh_3":"","CM_Pblow_1":"","CM_Pblow_2":"","CM_Pblow_3":"","CM_Pbmid_1":"","CM_Pbmid_2":"","CM_Pbmid_3":""},{"Bin range(ppm)":"#END","CM_CdPool_1":"","CM_CdPool_2":"","CM_CdPool_3":"","CM_Cdhigh_1":"","CM_Cdhigh_2":"","CM_Cdhigh_3":"","CM_Cdlow_1":"","CM_Cdlow_2":"","CM_Cdlow_3":"","CM_Cdmid_1":"","CM_Cdmid_2":"","CM_Cdmid_3":"","CM_CdPbPool_1":"","CM_CdPbPool_2":"","CM_CdPbPool_3-R":"","CM_TotalPool_1":"","CM_TotalPool_2":"","CM_TotalPool_3":"","CM_CdPbhigh_1":"","CM_CdPbhigh_2":"","CM_CdPbhigh_3":"","CM_CdPblow_1":"","CM_CdPblow_2":"","CM_CdPblow_3":"","CM_CdPBmid_1":"","CM_CdPBmid_2":"","CM_CdPBmid_3":"","CM_NoExpPool_1":"","CM_NoExpPool_2":"","CM_NoExpPool_3":"","CM_NoExp_1":"","CM_NoExp_2":"","CM_NoExp_3":"","CM_PbPool_1":"","CM_PbPool_2":"","CM_PbPool_3-R":"","CM_Pbhigh_1":"","CM_Pbhigh_2":"","CM_Pbhigh_3":"","CM_Pblow_1":"","CM_Pblow_2":"","CM_Pblow_3":"","CM_Pbmid_1":"","CM_Pbmid_2":"","CM_Pbmid_3":""}]

}