Stability of lipids in plasma and serum: Effects of temperature-related storage conditions on the human lipidome.
Author(s): Reis GB, Rees JC, Ivanova AA, Kuklenyik Z, Drew NM, Pirkle JL, Barr JR
Publication: J Mass Spectrom Adv Clin Lab, 2021, Vol. 22, Page 34-42
PubMed ID: 34939053 PubMed Review Paper? No
Purpose of Paper
The purpose of this paper was to investigate the potential impact on the lipidome of storing thawed plasma and serum in the refrigerator, at room temperature, or in a 37°C incubator. Additionally, the authors investigated potential differences in the plasma lipidome among blood specimens collected from fasting individuals and those that had eaten (post-prandial), as well as subsequent storage at 4°C or -80°C.
Conclusion of Paper
Cholesteryl ester (CE), phosphatidylcholine (PC), and triacylglycerol (TAG) accounted for 74% of the lipids detectable in thawed plasma specimens. Levels of CEs and PCs remained within the expected range when thawed plasma was stored in the refrigerator or on the bench top for up to 28 days. However, CE levels were higher than the expected range when thawed serum (but not thawed plasma) was stored in a 37°C incubator for 7, 21 or 28 days. PC levels were lower than the expected range after storage of thawed serum or plasma in a 37°C incubator for 7 or more days. TAGS remained within the expected range at all timepoints and storage temperatures, but the authors attribute this to the low abundance of many TAG species which may have prevented the detection of changes due to a low signal to noise ratio. Changes in phosphatidylethanolamine (PE), diacylglycerol (DAGs), free fatty acids (FFAs) and lysophospholipids were also observed after storage of thawed serum and plasma, but sphingomyelin (SM) and cholesteryl ester (CE) levels largely remained within the expected range at all temperatures. When individual species within a class were examined, changes in minor species (<10%) were not necessarily reflective of the lipid class as a whole. With the exception of FFAs and a few lysophospholipids, all changes associated with storage of thawed plasma/serum at 4°C were within the range of inter-individual variation.
A total of 4 TAGs were elevated in specimens collected from individuals immediately after consumption of a high fat emulsion compared to specimens collected from the same individuals before the fast was broken. One of these TAGS was also affected by storage of thawed plasma/serum? (decreased after meal consumption, increased with storage). In a second experiment, 4 FFAs, 3 lysophosphatidylcholine (LPCs) and one lysophosphatidylethanolamine (LPE) were affected by storage of fresh plasma.
Studies
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Study Purpose
The purpose of this study was to investigate the potential impact of storing thawed plasma and serum in the refrigerator, at room temperature, or in a 37°C incubator on the lipidome. Additionally, the authors investigated potential differences in the plasma lipidome among blood specimens collected from fasting individuals and those that had eaten (post-prandial), as well as subsequent storage at 4°C or -80°C. Ten plasma and ten serum specimens from healthy individuals were purchased, shipped on dry ice and stored at -80°C until use. Plasma and serum were thawed at 4°C and aliquoted for storage in a 37.5°C incubator, at room temperature (20±2°C), and in a refrigerator (4°C) for 0, 3, 7, 14, 21 and 28 days before analysis. To test the effects of storage and meal consumption on fresh plasma, plasma was collected (details not specified) from five healthy volunteers before and 2 h after consumption of a fatty drink (100 g long chain triglyceride emulsion). Blood specimens were transported on ice, aliquoted and stored at -80°C and 4°C for 0 and 28 days before analysis. Lipids were extracted using a modification of the Bligh and Dyer extraction method and analyzed using the Liquid chromatography Differential Mobility Spectrometry (LC-DMS-MS/MS)-based Lipidizer platform.
Summary of Findings:
CEs, PCs and TAGs accounted for 74% of detectable plasma lipids. Levels of CEs and PCs remained within the expected range when thawed plasma was stored in the refrigerator or bench top for up to 28 days. However, CE levels were higher than the expected range when thawed serum (but not thawed plasma) was stored in a 37°C incubator for 7, 21 or 28 days. Nevertheless, changes in PCs remained within the range of method variability. Levels of TAGS remained within the expected range at all storage timepoints and temperatures, but the authors attributed this to the low abundance of many TAG species that may have prevented the detection of changes due to a low signal to noise ratio. In thawed plasma specimens and serum PEs decreased over time while DAGs and FFAs increased when stored at room temperature or in a 37°C incubator; these changes were greater than the level of method variability. SM and CER levels remained mostly within the expected range at all storage temperatures. Lysophospholipids displayed an expected initial increase followed by a decrease with post-thaw storage of plasma and serum, which the authors attributed to production of PCs and PEs via hydrolysis followed by further hydrolysis to PFAs. When individual species within a class were looked at changes in minor species (<10%) were not always reflective of the lipid class as a whole. In general, lipids with a 16:0 or 18:0 fatty acid at one position and a PUFA at the other showed the largest changes. Plasma and serum levels of PCs and FFAs were inversely correlated with one another during post-thaw storage at room temperature or in a 37°C incubator, with stronger correlations observed when specimens were stored in the 37°C incubator. Although storage induced changes in PC and LPCS levels were correlated with one another, the direction and magnitude of the changes were dependent upon sample type (plasma versus serum) and the temperature of post-thaw storage. With the exception of FFAs and a few lysophospholipids, all changes associated with post-thaw storage at 4°C fell within the range of inter-individual variation.
A total of 4 TAGs were elevated in specimens collected from individuals immediately after consumption of a high fat emulsion compared to specimens collected from the same individuals before the fast was broken. One of these TAGS was also affected by storage of thawed plasma/serum? (decreased after meal consumption, increased with storage). In a second experiment, 4 FFAs, 3 lysophosphatidylcholine (LPCs) and one lysophosphatidylethanolamine (LPE) were affected by storage of fresh plasma.
Biospecimens
Preservative Types
- None (Fresh)
- Frozen
Diagnoses:
- Normal
Platform:
Analyte Technology Platform Lipid LC-MS or LC-MS/MS Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Acquisition Time of biospecimen collection Before eating
2 h post-consumption of high fat drink
Biospecimen Aliquots and Components Blood and blood products Plasma
Serum
Storage Storage temperature -80°C
4°C
Room temperature
37°C
Storage Storage duration 0 days
3 days
7 days
14 days
21 days
28 days