Plant Polyphenols: Characterisation, Metabolism and Beneficial Activity in Model Systems

The pursuit of efficient and safe alternatives to improve health and treat physical/mental illness has found in plant extracts a potential source of bioactive compounds such as polyphenols. The work presented within this thesis aimed to investigate the changes in the plant extract after ingestion, identify the remaining polyphenols and phase II conjugates, and to finally test their possible biological activity in in vitro assays. German chamomile (GC) was used as experimental case which was treated with a multiple-enzyme hydrolysis to mimic the process most likely to occur in the gastrointestinal tract. This methodology was previously used in five different fruit matrices and it was successfully applied to plant-based materials. It was possible to identify and quantify a series of compounds before and after treatment by LC-ESI/MS, and these results were compared with data found in the literature. In addition, it was found that differences in sample preparation and a suitable organic solvent for liquid-liquid extraction had an important impact in the quantification of these compounds. The transformation of polyphenols after the phase II metabolism was also investigated. For this purpose, hesperetin aglycone was used in the laboratory to produce the main monosulfate commonly found in urine samples. For the first time, p-nitropheyl sulfate (PNS) was used as co-cofactor in the biological sulfonation of polyphenols which enhanced the transformation to hesperetin-3'-O-sulfate (Hp3'SO4). Indeed, it was the first time that Hp3'SO4 was quantified in urine samples after orange juice consumption using a true standard. Finally, the anti-inflammatory activity of tested plant extracts and their main polyphenols was assessed by measuring their inhibitory activity against recombinant human COX-2. An enzyme immunoassay (EIA) and in vitro-LC-MS methodologies were used for this purpose. Consistent COX-2 inhibition was found for green tea (GT) and its two major polyphenols, epigallocatechin gallate (EGCG) and epigallocatechin (EGC) which showed inhibition in a concentration dependence manner. On the other hand, various polyphenols and plant extracts also presented COX-2 inhibition, but did not show concentration dependence