{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST000608","ANALYSIS_ID":"AN000929","VERSION":"1","CREATED_ON":"May 26, 2017, 11:40 am"},

"PROJECT":{"PROJECT_TITLE":"Comparing identified and statistically significant lipids and polar metabolites in 15-year old serum and dried blood spot samples for longitudinal studies","PROJECT_SUMMARY":"The use of dried blood spots (DBS) has many advantages over traditional plasma and serum samples such as the smaller blood volume required, storage at room temperature, and ability to sample in remote locations. However, understanding the robustness of different analytes in DBS samples is essential, especially in older samples collected for longitudinal studies. Here we analyzed the stability of polar metabolites and lipids in DBS samples collected in 2000-2001 and stored at room temperature. The identified and statistically significant molecules were then compared to matched serum samples stored at –80°C to determine if the DBS samples could be effectively used in a longitudinal study following metabolic disease. A total of 400 polar metabolites and lipids were identified in the serum and DBS samples using gas chromatograph/mass spectrometry (GC/MS), liquid chromatography (LC)/MS, and LC/ion mobility spectrometry-MS (LC/IMS-MS). The identified polar metabolites overlapped well between the sample types, though only one statistically significant metabolite was conserved in a case-control study of older diabetic males with low amounts of high-density lipoproteins and high body mass indices, triacylglycerides and glucose levels when compared to non-diabetic patients with normal levels, indicating that degradation in the DBS samples affects polar metabolite quantitation. Differences in the lipid identifications indicated that some oxidation occurs in the DBS samples. However, 36 statistically significant lipids correlated in both sample types. The difference in the number of statistically significant polar metabolites and lipids indicated that the lipids did not degrade to as great of a degree as the polar metabolites in the DBS samples and lipid quantitation was still possible.","INSTITUTE":"Pacific Northwest National Laboratory","LAST_NAME":"Casey","FIRST_NAME":"Cameron","ADDRESS":"902 Battelle Boulevard, Richland, WA, 99354, USA","EMAIL":"cameron.casey@pnnl.gov","PHONE":"509-371-6612","FUNDING_SOURCE":"Portions of this research were supported by grants from the National Institute of Environmental Health Sciences of the NIH (R01 ES022190), NIH Eunice Kennedy Shriver National Institute of Child Health and Human Development (R21 HD084788), National Institute of General Medical Sciences (P41 GM103493), and the Laboratory Directed Research and Development Program at Pacific Northwest National Laboratory. This research utilized capabilities developed by the Pan-omics program (funded by the U.S. Department of Energy Office of Biological and Environmental Research Genome Sciences Program) and by the National Institute of Allergy and Infectious Diseases under grant U19 AI106772. The Osteoporotic Fractures in Men (MrOS) Study in the US was supported by the National Institute on Aging (NIA), the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS), the National Center for Advancing Translational Sciences (NCATS), and NIH Roadmap for Medical Research under the following grant numbers: U01 AG027810, U01 AG042124, U01 AG042139, U01 AG042140, U01 AG042143, U01 AG042145, U01 AG042168, U01 AR066160, and UL1 TR000128. This work was performed in the W. R. Wiley Environmental Molecular Sciences Laboratory (EMSL), a DOE national scientific user facility at the Pacific Northwest National Laboratory (PNNL). PNNL is operated by Battelle for the DOE under contract DE-AC05-76RL0 1830.","PUBLICATIONS":"Comparing identified and statistically significant lipids and polar metabolites in 15-year old serum and dried blood spot samples for longitudinal studies","DOI":"10.1002/rcm.7808"},

"STUDY":{"STUDY_TITLE":"Comparing identified and statistically significant lipids and polar metabolites in 15-year old serum and dried blood spot samples for longitudinal studies","STUDY_SUMMARY":"The use of dried blood spots (DBS) has many advantages over traditional plasma and serum samples such as the smaller blood volume required, storage at room temperature, and ability to sample in remote locations. However, understanding the robustness of different analytes in DBS samples is essential, especially in older samples collected for longitudinal studies. Here we analyzed the stability of polar metabolites and lipids in DBS samples collected in 2000-2001 and stored at room temperature. The identified and statistically significant molecules were then compared to matched serum samples stored at –80°C to determine if the DBS samples could be effectively used in a longitudinal study following metabolic disease. A total of 400 polar metabolites and lipids were identified in the serum and DBS samples using gas chromatograph/mass spectrometry (GC/MS), liquid chromatography (LC)/MS, and LC/ion mobility spectrometry-MS (LC/IMS-MS). The identified polar metabolites overlapped well between the sample types, though only one statistically significant metabolite was conserved in a case-control study of older diabetic males with low amounts of high-density lipoproteins and high body mass indices, triacylglycerides and glucose levels when compared to non-diabetic patients with normal levels, indicating that degradation in the DBS samples affects polar metabolite quantitation. Differences in the lipid identifications indicated that some oxidation occurs in the DBS samples. However, 36 statistically significant lipids correlated in both sample types. The difference in the number of statistically significant polar metabolites and lipids indicated that the lipids did not degrade to as great of a degree as the polar metabolites in the DBS samples and lipid quantitation was still possible.","INSTITUTE":"Pacific Northwest National Laboratory","LAST_NAME":"Casey","FIRST_NAME":"Cameron","ADDRESS":"902 Battelle Boulevard, Richland, WA, 99354, USA","EMAIL":"cameron.casey@pnnl.gov","PHONE":"509-371-6612"},

"SUBJECT":{"SUBJECT_TYPE":"Human","SUBJECT_SPECIES":"Homo sapiens","TAXONOMY_ID":"9606"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"PO6573",
"Sample ID":"OHSUblotter_PO6573_1",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6573",
"Sample ID":"OHSUblotter_PO6573_2",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6573",
"Sample ID":"OHSUblotter_PO6573_3",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6716",
"Sample ID":"OHSUblotter_PO6716_1",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6716",
"Sample ID":"OHSUblotter_PO6716_2",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6716",
"Sample ID":"OHSUblotter_PO6716_3",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6764",
"Sample ID":"OHSUblotter_PO6764_1",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6764",
"Sample ID":"OHSUblotter_PO6764_2",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6764",
"Sample ID":"OHSUblotter_PO6764_3",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6882",
"Sample ID":"OHSUblotter_PO6882_1",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6882",
"Sample ID":"OHSUblotter_PO6882_2",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6882",
"Sample ID":"OHSUblotter_PO6882_3",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6927",
"Sample ID":"OHSUblotter_PO6927_1",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6927",
"Sample ID":"OHSUblotter_PO6927_2",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6927",
"Sample ID":"OHSUblotter_PO6927_3",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6935",
"Sample ID":"OHSUblotter_PO6935_1",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6935",
"Sample ID":"OHSUblotter_PO6935_2",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6935",
"Sample ID":"OHSUblotter_PO6935_3",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6976",
"Sample ID":"OHSUblotter_PO6976_1",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6976",
"Sample ID":"OHSUblotter_PO6976_2",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6976",
"Sample ID":"OHSUblotter_PO6976_3",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO7117",
"Sample ID":"OHSUblotter_PO7117_1",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO7117",
"Sample ID":"OHSUblotter_PO7117_2",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO7117",
"Sample ID":"OHSUblotter_PO7117_3",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO7324",
"Sample ID":"OHSUblotter_PO7324_1",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO7324",
"Sample ID":"OHSUblotter_PO7324_2",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO7324",
"Sample ID":"OHSUblotter_PO7324_3",
"Factors":{"Sample_Type":"blotter","Factor":"case"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO7355",
"Sample ID":"OHSUblotter_PO7355_1",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO7355",
"Sample ID":"OHSUblotter_PO7355_2",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO7355",
"Sample ID":"OHSUblotter_PO7355_3",
"Factors":{"Sample_Type":"blotter","Factor":"control"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6573",
"Sample ID":"OHSUserum_PO6573_1",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6573",
"Sample ID":"OHSUserum_PO6573_2",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6573",
"Sample ID":"OHSUserum_PO6573_3",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6716",
"Sample ID":"OHSUserum_PO6716_1",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6716",
"Sample ID":"OHSUserum_PO6716_2",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6716",
"Sample ID":"OHSUserum_PO6716_3",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6764",
"Sample ID":"OHSUserum_PO6764_1",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6764",
"Sample ID":"OHSUserum_PO6764_2",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6764",
"Sample ID":"OHSUserum_PO6764_3",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6882",
"Sample ID":"OHSUserum_PO6882_1",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6882",
"Sample ID":"OHSUserum_PO6882_2",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6882",
"Sample ID":"OHSUserum_PO6882_3",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6927",
"Sample ID":"OHSUserum_PO6927_1",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6927",
"Sample ID":"OHSUserum_PO6927_2",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6927",
"Sample ID":"OHSUserum_PO6927_3",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6935",
"Sample ID":"OHSUserum_PO6935_1",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6935",
"Sample ID":"OHSUserum_PO6935_2",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6935",
"Sample ID":"OHSUserum_PO6935_3",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO6976",
"Sample ID":"OHSUserum_PO6976_1",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO6976",
"Sample ID":"OHSUserum_PO6976_2",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO6976",
"Sample ID":"OHSUserum_PO6976_3",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO7117",
"Sample ID":"OHSUserum_PO7117_1",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO7117",
"Sample ID":"OHSUserum_PO7117_2",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO7117",
"Sample ID":"OHSUserum_PO7117_3",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO7324",
"Sample ID":"OHSUserum_PO7324_1",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO7324",
"Sample ID":"OHSUserum_PO7324_2",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO7324",
"Sample ID":"OHSUserum_PO7324_3",
"Factors":{"Sample_Type":"serum","Factor":"case"},
"Additional sample data":{"Replicate":"3"}
},
{
"Subject ID":"PO7355",
"Sample ID":"OHSUserum_PO7355_1",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"1"}
},
{
"Subject ID":"PO7355",
"Sample ID":"OHSUserum_PO7355_2",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"2"}
},
{
"Subject ID":"PO7355",
"Sample ID":"OHSUserum_PO7355_3",
"Factors":{"Sample_Type":"serum","Factor":"control"},
"Additional sample data":{"Replicate":"3"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"Matched DBS and serum samples were simultaneously collected in 2000–2001 from 10 overnight fasted male participants having an average age of 75.5 ± 6.15 years. The institutional review boards at the participating institutions approved the protocol and the study participants provided informed consent. Case participants included 5 diabetic men with high BMIs, TGs and glucose levels, and low amounts of HDL. The 5 control participants had normal metabolic profiles (normal BMIs, TGs, HDL and glucose levels).","STORAGE_CONDITIONS":"The DBS were stored at room temperature in the dark until analysis, while the serum was stored at –80°C."},

"TREATMENT":{"TREATMENT_SUMMARY":"Samples collected in 2000–2001 were stored until June 2015."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"For the serum samples, 10 mL of whole blood was drawn and centrifuged so the serum could be extracted. For the DBS samples, 500 μL of blood was collected and 125 μL was blotted in four areas on a Whatman FTA Classic Card. Polar metabolites and lipid extracts were derived from the same sample type (DBS or serum) using a modified Folch extraction in June 2015 and analyzed with MS shortly after. The DBS punch from each patient was transferred into to a 2.0 mL tube where 50 μL of water and then 1200 μL of –20°C 2:1 chloroform/methanol were added. Each sample was vortexed for 30 s then transferred into a shaker at 22°C for 60 min at 600 rpm. The samples were vortexed again for 30 s and then 250 μL of water was added to induce a phase separation. The sample was gently inverted several times, placed at room temperature for 5 min and then centrifuged at 10,000 g for 5 min at 4°C and put on ice to maintain the clear phase separation. Finally, 400 μL of the top polar layer was removed, dried in a Speedvac, and stored at –80°C for analysis of polar metabolites, while 700 μL of the bottom nonpolar layer was removed, dried in a Speedvac, and stored at –20°C in 250 μL of 2:1 chloroform/methanol for lipid analyses. Serum lipids were extracted using a similar procedure except 25 μL of serum was used and then 600 μL of –20°C 2:1 chloroform/methanol was added. After vortexing and shaking, 125 μL of water was added to induce a phase separation and 200 μL of the top polar layer and 350 μL of the bottom nonpolar lipid layer were removed and stored as outlined above. Prior to analysis, the total lipid extracts were dried down and then reconstituted in 70 μL and 100 μL of MeOH for the DBS and serum samples, respectively. To generate pooled case and control samples for LC/MS/MS and LC/IMS-MS analyses, 5 μL aliquots from each reconstituted DBS and serum sample were removed and combined."},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_TYPE":"Reversed phase","INSTRUMENT_NAME":"Waters NanoAcquity","COLUMN_NAME":"Waters Acquity HSS T3 (150 x 1mm, 1.8um)"},

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

"MS":{"MS_COMMENTS":"-","INSTRUMENT_NAME":"Thermo Velos Pro Orbitrap","INSTRUMENT_TYPE":"Orbitrap","MS_TYPE":"ESI","ION_MODE":"POSITIVE"},

"MS_METABOLITE_DATA":{
"Units":"Peak Height",

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