Most blood biomarkers related to vitamin status, one-carbon metabolism, and the kynurenine pathway show adequate preanalytical stability and within-person reproducibility to allow assessment of exposure or nutritional status in healthy women and cardiovascular patients

Abstract

Knowledge of stability during sample transportation and changes in biomarker concentrations within person over time are paramount for proper design and interpretation of epidemiologic studies based on a single measurement of biomarker status. Therefore, we investigated stability and intraindividual vs. interindividual variation in blood concentrations of biomarkers related to vitamin status, one-carbon metabolism, and the kynurenine pathway. Whole blood (EDTA and heparin, n = 12) was stored with an icepack for 24 or 48 h, and plasma concentrations of 38 biomarkers were determined. Stability was calculated as change per hour, intraclass correlation coefficient (ICC), and simple Spearman correlation. Within-person reproducibility of biomarkers was expressed as ICC in samples collected 1-2 y apart from 40 postmenopausal women and in samples collected up to 3 y apart from 551 patients with stable angina pectoris. Biomarker stability was similar in EDTA and heparin blood. Most biomarkers were essentially stable, except for choline and total homocysteine (tHcy), which increased markedly. Within-person reproducibility in postmenopausal women was excellent (ICC > 0.75) for cotinine, all-trans retinol, cobalamin, riboflavin, α-tocopherol, Gly, pyridoxal, methylmalonic acid, creatinine, pyridoxal 5'-phosphate, and Ser; was good to fair (ICC of 0.74-0.40) for pyridoxic acid, kynurenine, tHcy, cholecalciferol, flavin mononucleotide, kynurenic acid, xanthurenic acid, 3-hydroxykynurenine, sarcosine, anthranilic acid, cystathionine, homoarginine, 3-hydroxyanthranilic acid, betaine, Arg, folate, total cysteine, dimethylglycine, asymmetric dimethylarginine, neopterin, symmetric dimethylarginine, and Trp; and poor (ICC of 0.39-0.15) for methionine sulfoxide, Met, choline, and trimethyllysine. Similar reproducibilities were observed in patients with coronary heart disease. Thus, most biomarkers investigated were essentially stable in cooled whole blood for up to 48 h and had a sufficient within-person reproducibility to allow one-exposure assessment of biomarker status in epidemiologic studies. The Western Norway B Vitamin Intervention Trial was registered at clinicaltrials.gov as NTC00354081.

Trial registration: ClinicalTrials.gov NCT00354081.

Conflict of interest statement

Author disclosures: M. K. Townsend, O. Nygård, S. S. Tworoger, P. Brennan, and M. Johansson, no conflicts of interest. Ø. Midttun and P. M. Ueland are members of the steering board of the nonprofit Foundation to Promote Research Into Functional Vitamin B12 Deficiency.

Figures

FIGURE 1

Stability of biomarkers in EDTA blood stored in a Styrofoam container with an icepack for 24 or 48 h. Biomarkers (pyridoxal, choline, total homocysteine, Arg, and hydroxyanthranilic acid) that changed substantially over time (left panel) had a low intraclass correlation coefficient ( 0.4) correlation with values at time 0 (right panel). This reflects a parallel increase in concentrations over time and implies that the ranking of the values at fixed time points was maintained. ADMA decreased moderately during storage, but both intraclass correlation coefficient and ranking were low, which is explained by the low between-person variability for ADMA. n = 12. ADMA, asymmetric dimethylarginine; FMN, flavin mononucleotide; SDMA, symmetric dimethylarginine.