Abstract
BackgroundInformation regarding the variability of metabolite levels over time in an individual is required to estimate the reproducibility of metabolite measurements. In intervention studies, it is critical to appropriately judge changes that are elicited by any kind of intervention. The pre-analytic phase (collection, transport and sample processing) is a particularly important component of data quality in multi-center studies.MethodsReliability of metabolites (within-and between-person variance, intraclass correlation coefficient) and stability (shipment simulation at different temperatures, use of gel-barrier collection tubes, freeze-thaw cycles) were analyzed in fasting serum and plasma samples of 22 healthy human subjects using a targeted LC-MS approach.ResultsReliability of metabolite measurements was higher in serum compared to plasma samples and was good in most saturated short-and medium-chain acylcarnitines, amino acids, biogenic amines, glycerophospholipids, sphingolipids and hexose. The majority of metabolites were stable for 24 h on cool packs and at room temperature in non-centrifuged tubes. Plasma and serum metabolite stability showed good coherence. Serum metabolite concentrations were mostly unaffected by tube type and one or two freeze-thaw cycles.ConclusionA single time point measurement is assumed to be sufficient for a targeted metabolomics analysis of most metabolites. For shipment, samples should ideally be separated and frozen immediately after collection, as some amino acids and biogenic amines become unstable within 3 h on cool packs. Serum gel-barrier tubes can be used safely for this process as they have no effect on concentration in most metabolites. Shipment of non-centrifuged samples on cool packs is a cost-efficient alternative for most metabolites.
Highlights
The inclusion of the serum or plasma metabolome analysis in clinical trials is an appealing approach for several reasons
Studies investigating sample stability during shipment focus on a small metabolite panel including cholesterol [4,5,6], vitamins [6] lipids [6,7], amino acids [8], glucose [9] or acylcarnitines [10], or they are limited by a small sample size [11].In this study, we address questions regarding the reproducibility of targeted metabolomics measurements in the same individual at three different time points and of pre-analytic stability of metabolites
Samples taken from 20 healthy human subjects on three different days were used to calculate the intraclass correlation coefficient (ICC), the within-person coefficient of variance (CV) (WCV) and the betweenperson CV (BCV) for 159 metabolites in serum and plasma
Summary
The inclusion of the serum or plasma metabolome analysis in clinical trials is an appealing approach for several reasons. Observed changes in the metabolome could be linked to the clinical response to a study medication or any other kind of intervention. Studies investigating sample stability during shipment focus on a small metabolite panel including cholesterol [4,5,6], vitamins [6] lipids [6,7], amino acids [8], glucose [9] or acylcarnitines [10], or they are limited by a small sample size [11].In this study, we address questions regarding the reproducibility of targeted metabolomics measurements in the same individual at three different time points and of pre-analytic stability of metabolites. The pre-analytic phase (collection, transport and sample processing) is a important component of data quality in multi-center studies
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