The present study aims to investigate the dose dependent effects of consuming diets enriched in flavonoid-rich and flavonoid-poor fruits and vegetables on the urine metabolome of adults who had a ≥1.5 fold increased risk of cardiovascular diseases. A single-blind, dose-dependent, parallel randomized controlled dietary intervention was conducted where volunteers (n = 126) were randomly assigned to one of three diets: high flavonoid diet, low flavonoid diet or habitual diet as a control for 18 weeks. High resolution LC–MS untargeted metabolomics with minimal sample cleanup was performed using an Orbitrap mass spectrometer. Putative biomarkers which characterize diets with high and low flavonoid content were selected by state-of-the-art data analysis strategies and identified by HR-MS and HR-MS/MS assays. Discrimination between diets was observed by application of two linear mixed models: one including a diet-time interaction effect and the second containing only a time effect. Valerolactones, phenolic acids and their derivatives were among sixteen biomarkers related to the high flavonoid dietary exposure. Four biomarkers related to the low flavonoid diet belonged to the family of phenolic acids. For the first time abscisic acid glucuronide was reported as a biomarker after a dietary intake, however its origins have to be examined by future hypothesis driven experiments using a more targeted approach. This metabolomic analysis has identified a number of dose dependent urinary biomarkers (i.e. proline betaine or iberin-N-acetyl cysteine), which can be used in future observation and intervention studies to assess flavonoids and non-flavonoid phenolic intakes and compliance to fruit and vegetable intervention.
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FLAVURS study group: Julie A Lovegrove (PI); Anne-Marie Minihane; Michael H Gordon, Jeremy PE Spencer, Kieran M. Tuohy, Orla B Kennedy, Trevor W George (Study Co-ordinator), Mary Foong-Fong Chong (Study Co-ordinator), Dauren Alimbetov, Yin Jin and Anna Macready. The authors would like to express their sincere thanks to all those involved in the FLAVURS project and in particular, to Orla B. Kennedy, Jeremy P.E. Spencer and Mike Gordon, for their critical reading of this article.
This metabolomic study was financially supported by the ADP2014 project, funded by the Autonomous Province of Trento, Italy. The work by Jan Stanstrup was supported by a research grant (VKR023371) from VILLUM FONDEN.
MU conceived and conducted the metabolomics analyses with KT, performed metabolite annotation, and wrote the body of the paper with FM, JS, KMT and JAL; JS conceived and conducted the LC–MS data analysis with PF and MU. KT helped with the data analysis; JAL conceived, obtained funding and conducted the intervention study, of which she was coordinator, with KMT, AMM, TG, MCFF and the FLAVURS study group. FM designed and coordinated the metabolomic research and obtained funding, together with KMT. All authors reviewed, edited and approved the final version of the manuscript.
Conflict of interest
The authors declare that they have no conflict of interest.
FLAVURS was registered as a randomized clinical trial (ISRCTN47748735) and conducted according to the guidelines laid down in the Declaration of Helsinki. Ethical approval for the study was obtained from the Local Research Ethics Committee of the Isle of Wight, Portsmouth and South East Hampshire (REC: 07/H0501/81), and the University of Reading’s Research (REC: 07/22) Ethics Committee. Informed consent was obtained before participation.
For the FLAVURS study group listed below Acknowledgment section.
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Ulaszewska, M.M., Trost, K., Stanstrup, J. et al. Urinary metabolomic profiling to identify biomarkers of a flavonoid-rich and flavonoid-poor fruits and vegetables diet in adults: the FLAVURS trial. Metabolomics 12, 32 (2016). https://doi.org/10.1007/s11306-015-0935-z
- Untargeted metabolomics
- Cardiovascular health
- Dietary intervention
- Fruit and vegetables