Stability of targeted metabolite profiles of urine samples under different storage conditions.
Author(s): Rotter M, Brandmaier S, Prehn C, Adam J, Rabstein S, Gawrych K, Brüning T, Illig T, Lickert H, Adamski J, Wang-Sattler R
Publication: Metabolomics, 2017, Vol. 13, Page 4
PubMed ID: 27980503 PubMed Review Paper? No
Purpose of Paper
This paper investigated the effects of storage duration and temperature and freeze-thaw cycling on levels of 63 metabolites in urine.
Conclusion of Paper
Although 56 of the 63 (89%) metabolites were not significantly affected by storage of urine at any of the temperatures examined, significant declines from baseline were noted in the levels of arginine (Arg), methionine (Met), serine (Ser), valine (Val), leucine/Isoleucine (xLeu), and hexose H1 (H1) after 24 h at 20˚C and in Arg, Val, and xLeu after 24 h at 9˚C while an increase occurred in hexenoylcarnitine (C6:1) after 24 h at 9˚C. All analytes were stable for up to 24 h of storage at 4 or -20˚C. Overall, a significant interaction between storage duration and temperature was found for levels of Arg, glutamic acid (Glu), Met, phenylalanine (Phe), proline (Pro), Ser, Threonine (Thr), Val, xLeu, and H1. One to two freeze-thaw cycles had no effect on any of the metabolites, but levels of H1and propionylcarnitine (C3) were significantly higher than baseline after three freeze-thaw cycles and there was a significant association between the number of freeze-thaw cycles and levels of C3, iso-/butyrylcarnitine (C4), octenoylcarnitine (C8:1), 3-OH-hexadecenoylcarnitine (C16:1-OH), and H1.
Studies
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Study Purpose
This study investigated the effects of storage duration and temperature and freeze-thaw cycling on levels of 63 metabolites in urine. Urine from six healthy fasting females was collected into sterile containers, pooled, and then aliquoted. Four aliquots from each donor were stored at -80, -20, 4, ~9 (cool pack), and ~20˚C (room temperature) for 2, 8, and 24 h. To investigate the effects of freeze-thaw cycling, twelve aliquots were subjected to three cycles of freezing at -80˚C for 24 h and thawing at 20˚C for 2 h. One hundred sixty-two targeted metabolites were measured using the AbsoluteIDQTM p150 kit on a flow injection-electrospray ionization-triple quadruple mass spectrometry (FIA-ESI-MS/MS). Sixty-three of the metabolites passed quality control parameters and were included in this study. Based on multiple corrections, statistical significance was defined as P<0.00079.
Summary of Findings:
Although 56 of the 63 (89%) metabolites were not significantly affected by storage of urine at any of the temperatures examined for up to 24 h, significant declines from baseline were observed in the levels of Arg (-40.9%, P=0.00031), Met (-43.7%, P=0.00013), Ser (36.2%, P=0.00004), Val (-59.0%, P=0.00001), xLeu (-39.6%, P=0.000088), and hexose H1 (-30.3%, P=0.0000095) after 24 h at 20˚C and in Arg (-40.2%, P=0.000072), Val (-42.5%, P=0.000048), and xLeu (-35.3% , P=0.00011) after 24 h at 9˚C. A 16.2% increase in C6:1 occurred after 24 h at 9˚C (P=0.000081). All analytes were stable during 24 h of storage at 4 or -20˚C. Overall, a significant interaction between storage duration and temperature was found for levels of Arg (P=0.000022), Glu (P=0.000025), Met (P=0.00000012), Phe (P=0.000078), Pro (P=0.000058), Ser (P=0.00000000098), Thr (P=0.000077), Val (P=0.000028), Leu/Ile (P=0.0000012), and H1 (P=0.00000045).
One to two freeze-thaw cycles had no effect on any of the metabolites, but levels of H1 and C3 were significantly higher than baseline after three freeze-thaw cycles (P=0.00077 and P=0.00031, respectively). There was a significant association between the number of freeze-thaw cycles and levels of C3 (P=0.00025), C4 (P=0.00018), C8:1 (P=0.00029), C16:1-OH (P=0.00025), and H1 (P=0.00015).
Biospecimens
Preservative Types
- Frozen
- None (Fresh)
Diagnoses:
- Normal
Platform:
Analyte Technology Platform Small molecule ESI MS Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Storage Storage temperature -80˚C
-20˚C
4˚C
9˚C (cool pack)
20˚C (room temperature)
Storage Freeze/thaw cycling 0 cycles
1 cycle
2 cycles
3 cycles
Storage Storage duration 0 h
2 h
8 h
24 h