Biological Variability Dominates and Influences Analytical Variance in HPLC‐ECD Studies of the Human Plasma Metabolome

Abstract

Background: Biomarker-based assessments of biological samples are widespread in clinical, preclinical, and epidemiological investigations. We previously developed serum metabolomic profiles assessed by HPLC-separations coupled with coulometric array detection that can accurately identify ad libitum fed and caloric-restricted rats. These profiles are being adapted for human epidemiology studies, given the importance of energy balance in human disease.Methods: Human plasma samples were biochemically analyzed using HPLC separations coupled with coulometric electrode array detection. Results:We identified these markers/metabolites in human plasma, and then used them to determine which human samples represent blinded duplicates with 100% accuracy (N = 30 of 30). At least 47 of 61 metabolites tested were sufficiently stable for use even after 48 hours of exposure to shipping conditions. Stability of some metabolites differed between individuals (N = 10 at 0, 24, and 48 hours), suggesting the influence of some biological factors on parameters normally considered as analytical. Conclusion:Overall analytical precision (mean median CV, ~9%) and total between-person variation (median CV, ~50-70%) appear well suited to enable use of metabolomics markers in human clinical trials and epidemiological studies, including studies of the effect of caloric intake and balance on long-term cancer risk. BackgroundAfter tobacco, over-nutrition is, arguably, the major cause of excess morbidity in developed countries, affecting a broad spectrum of diseases including cancer, cardio-/cere-brovascular disease, and type II diabetes. This association may be seen in both broad demographic groups, such as the American Cancer Society study group (900,000 U.S. adults) [1] and in more narrowly defined demographic