High-throughput 1H NMR-based metabolic analysis of human serum and urine for large-scale epidemiological studies:: validation study

Background Metabolic profiling of biofluid specimens is an established method for investigating disease states in clinical studies but is only recently being applied to large-scale human population studies. As part of protocol development for the UK Biobank study, a H-1 nuclear magnetic resonance (NMR)-based metabonomic analysis of specimen storage effects and analytical reproducibility was carried out using urine and serum specimens from 40 volunteers. Methods Aliquots of each specimen were stored for t=0 and t=24 h at 4 degrees C prior to freezing, and in the case of serum samples for a further 12 h (t = 36), to determine whether the storage times affected specimen composition and quality. A blinded split-specimen matching exercise was implemented to assign candidate spectral pairs stored for different times using multivariate statistical analysis of the NMR data. Results Using a chemometric strategy, split specimens at time t=0 and t=24 or 36 h after storage at 4 degrees C were easily paired and the split-specimen matching task was reduced to a. workable size. H-1 NMR profiling established that the t=24 h urine and serum groups showed no systematic metabolite changes, indicating biochemical stability. Some small differences in serum specimens stored for t=36 h at 4 degrees C were detectable only by multivariate analysis, and were attributed to generalized alterations in proteins and protein fragments, and possibly trimethylamine-N-oxide. No other specific metabolite was implicated. Conclusions For the purposes of NMR-based analysis, storage of urine and serum for up to t=24 h at 4 degrees C does not detectably affect the metabolic profile and the methodology is robust. Future application of multivariate methods to data-rich studies should substantially enhance information recovery from epidemiological studies.