Characterization of Blueberry (Poly)phenolics in Foods, their Metabolites in Human Biological Samples and Anti-Inflammatory Effects of Phytochemicals In Vitro

Blueberries’ popularity is in part supported by the health benefits associated with their consumption. Research has strongly focused on their polyphenol and anthocyanin concentration, one of the highest amongst commonly consumed foods. Polyphenols have demonstrated antioxidant effects in vitro, and consuming polyphenol-rich foods is associated with protective effects towards non-communicable diseases, but their direct bioactivity in the human body remains to be fully understood. It is estimated that about 90 to 95% of the initial polyphenol intake reaches the colon without being absorbed. Instead, phenolic compounds go through extensive biotransformations, consisting of catabolism into smaller compounds mostly carried out by the gut microbiota and enabling their absorption, and/or conjugation through phase II metabolism. Hence, the compounds resulting from dietary consumption and circulating in the blood are mostly phenolic-derived metabolites while the parent compounds found in blueberries are present at very low concentrations. It is crucial to understand the metabolic fate of blueberry polyphenols to be able to characterize the bioactive compounds to which health benefits are attributed. The main objectives of this dissertation were to determine the metabolites resulting from blueberry consumption and evaluate the anti-inflammatory effects of blueberry phytochemicals in vitro. Several food products were prepared with wild blueberry powder, and the stability of the polyphenols in the products was assessed during storage at different temperatures, for up to 8 weeks. The most stable products were then used in a clinical trial to supplement children with wild blueberry powder (or a placebo) for 5 days. Plasma and urine samples were collected hourly on the 6th day for 5 hours. Phenolic-derived metabolites were characterized in the samples using an ultra-high performance liquid chromatography method coupled with mass-spectrometry/mass spectrometry multiple reactions monitoring, which suitability to identify and quantify the targeted metabolites was previously assessed. The literature surrounding blueberries and their health benefits is extensive, but the variety of models and the large range of polyphenols concentrations used, some of them not being representative of their low bioavailability, makes it challenging to interpret observed effects. To address some of these considerations, blueberry polyphenol and volatile extracts were developed and tested in two models of inflammation to determine the potential anti-inflammatory effects of blueberry native phytochemicals at low concentrations. The results obtained in this dissertation provide a phenolic-derived metabolite profile in the plasma and urine of children after blueberry or placebo consumption, and show a clear difference between the two groups. The blueberry polyphenol and volatile extracts significantly decreased the production of the pro-inflammatory interleukin-6 in whole blood, but only at high concentrations that are unlikely to be recovered in the blood, for phenolic compounds, after consumption of one serving of blueberries. No clear modulation of markers of inflammation were observed in the human macrophage cell culture model. These results strengthened the initial hypotheses that blueberry native compounds may not exert strong bioactive effects in vivo due to the molecules undergoing extensive biotransformations once consumed, and support the need to include metabolite compounds in further research assessing health benefits of blueberry consumption

The Effects of Blueberry Phytochemicals on Cell Models of Inflammation and Oxidative Stress

Blueberries have been extensively studied for the health benefits associated with their high phenolic content. The positive impact of blueberry consumption on human health is associated in part with modulation of proinflammatory molecular pathways and oxidative stress. Here, we review in vitro studies examining the anti-inflammatory and antioxidant effects of blueberry phytochemicals, discuss the results in terms of relevance to disease and health, and consider how different blueberry components modulate cellular mechanisms. The dampening effects of blueberry-derived molecules on inflammation and oxidative stress in cell models have been demonstrated through downregulation of the NF-κB pathway and reduction of reactive oxygen species (ROS) and lipid peroxidation. The modulatory effects of blueberry phytochemicals on the mitogen-activated protein kinase (MAPK) pathway and antioxidant system are not as well described, with inconsistent observations reported on immune cells and between models of endothelial, dermal, and ocular inflammation. Although anthocyanins are often reported as being the main bioactive compound in blueberries, no individual phytochemical has emerged as the primary compound when different fractions are compared; rather, an effect of whole blueberry extracts or synergy between different phenolic and nonphenolic extracts seems apparent. The major molecular mechanisms of blueberry phytochemicals are increasingly defined in cell models, but their relevance in more complex human systems needs further investigation using well-controlled clinical trials, in which systemic exposures to blueberry-associated molecules are measured concurrently with physiologic indices of inflammation and oxidative stress