Two apples a day modulate human:microbiome co-metabolic processing of polyphenols, tyrosine and tryptophan



Validated biomarkers of food intake (BFIs) have recently been suggested as a useful tool to assess adherence to dietary guidelines or compliance in human dietary interventions. Although many new candidate biomarkers have emerged in the last decades for different foods from metabolic profiling studies, the number of comprehensively validated biomarkers of food intake is limited. Apples are among the most frequently consumed fruits and a rich source of polyphenols and fibers, an important mediator for their health-protective properties.


Using an untargeted metabolomics approach, we aimed to identify biomarkers of long-term apple intake and explore how apples impact on the human plasma and urine metabolite profiles. Forty mildly hypercholesterolemic volunteers consumed two whole apples or a sugar and energy-matched control beverage, daily for 8 weeks in a randomized, controlled, crossover intervention study. The metabolome in plasma and urine samples was analyzed via untargeted metabolomics.


We found 61 urine and 9 plasma metabolites being statistically significant after the whole apple intake compared to the control beverage, including several polyphenol metabolites that could be used as BFIs. Furthermore, we identified several endogenous indole and phenylacetyl-glutamine microbial metabolites significantly increasing in urine after apple consumption. The multiomic dataset allowed exploration of the correlations between metabolites modulated significantly by the dietary intervention and fecal microbiota species at genus level, showing interesting interactions between Granulicatella genus and phenyl-acetic acid metabolites. Phloretin glucuronide and phloretin glucuronide sulfate appeared promising biomarkers of apple intake; however, robustness, reliability and stability data are needed for full BFI validation.


The identified apple BFIs can be used in future studies to assess compliance and to explore their health effects after apple intake. Moreover, the identification of polyphenol microbial metabolites suggests that apple consumption mediates significant gut microbial metabolic activity which should be further explored.

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We thank Massimo Pindo and the FEM Sequencing Platform for performing the DNA sequencing


This project was funded in part by the AGER project “Apple fruit quality in the post-genomic era, from breeding new genotypes to post-harvest: nutrition and health”, funded by the AGER (Agribusiness and research) with Grant no. 2010–2119, internal funding of Fondazione Edmund Mach, and the European Union’s Horizon2020 research and innovation grant agreement No 696295—ERA-Net Cofund ERA-HDHL “Biomarkers for Nutrition and Health implementing the JPI HDHL objectives” (, projects FOODBALL ( and CABALA_DIET&HEALTH ( The Renetta apples were kindly provided by Consorzio Melinda S.C.A., Cles, Trentino, Italy.

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Correspondence to Maria M. Ulaszewska or Julie Lovegrove or Fulvio Mattivi.

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Ulaszewska, M.M., Koutsos, A., Trošt, K. et al. Two apples a day modulate human:microbiome co-metabolic processing of polyphenols, tyrosine and tryptophan. Eur J Nutr 59, 3691–3714 (2020).

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  • Untargeted metabolomics
  • Apples
  • Polyphenols
  • Tryptophan
  • Tyrosine
  • Orbitrap
  • Biomarker of food intake
  • Validation