The

BACKGROUND

Metabolomics represents a powerful tool for exploring modulation of the human metabolome in response to food intake. However, the choice of multivariate statistical approach is not always evident, especially for complex experimental designs with repeated measurements per individual. Here we have investigated the serum metabolic responses to two breakfast meals: an egg and ham based breakfast and a cereal based breakfast using three different multivariate approaches based on the Projections to Latent Structures framework.

METHODS

In a cross over design, 24 healthy volunteers ate the egg and ham breakfast and cereal breakfast on four occasions each. Postprandial serum samples were subjected to metabolite profiling using

RESULTS

The Orthogonal Projections to Latent Structures with Discriminant Analysis model correctly classified 92 and 90% of the samples from the cereal breakfast and egg and ham breakfast, respectively, but confounded dietary effects with inter-personal variability. Orthogonal Projections to Latent Structures with Effect Projections removed inter-personal variability and performed perfect classification between breakfasts, however at the expense of comparing means of respective breakfasts instead of all samples. ANOVA-decomposed Projections to Latent Structures managed to remove inter-personal variability and predicted 99% of all individual samples correctly. Proline, tyrosine, and N-acetylated amino acids were found in higher concentration after consumption of the cereal breakfast while creatine, methanol, and isoleucine were found in higher concentration after the egg and ham breakfast.

CONCLUSIONS

Our results demonstrate that the choice of statistical method will influence the results and adequate methods need to be employed to manage sample dependency and repeated measurements in cross-over studies. In addition,

TRIAL REGISTRATION

Registered with ClinicalTrials.gov, identifier: NCT02039596 . Date of registration: January 17, 2014.