Investigating Polyphenol Exposure at the Omic-scale in Biological Matrices by Liquid Chromatography Coupled to Mass Spectrometry

Polyphenols, bioactive molecules present in plants and fungi, have been known for centuries to positively influence human wellbeing. Besides impacting human health directly, polyphenols can have an indirect effect either through modulating the microbiome by promoting or inhibiting the growth of certain microbes, or through combinatory effects, both synergistic and antagonistic, with other exogenous compounds that humans are exposed to. Studying polyphenols poses a challenge as they are an immensely diverse chemical class, containing a variety of isomers and biotransformation products originating from both human and microbial metabolism. Therefore, innovative workflows based on liquid chromatography coupled with mass spectrometry (LC-MS) were developed and benchmarked to better investigate polyphenols in biological matrices and gain insight into their impact on human health. Firstly, a sensitive targeted LC-MS method and a high-throughput sample preparation for 90 distinct polyphenols, that represent all the major chemical sub-classes, was developed. The method was then validated in-house for three different human matrices (urine, serum, and plasma). Secondly, using the targeted method as reference, an untargeted LC-MS workflow was developed and benchmarked. This workflow showed the potential and suitability of untargeted approaches to investigate polyphenols present in humans beyond those with readily available reference standards. Thirdly, the applicability of the developed workflows was demonstrated in two separate studies. The interchangeability of the workflows between different biological matrices was demonstrated by applying them to nine different plant and mushroom species. This allowed the comprehensive profiling of polyphenols present in these samples in order to better understand the selectivity of polyphenol oxidases found naturally in the samples. The workflows were then applied in a pilot study involving mother-infant pairs to investigate changes in infant exposure to dietary xenobiotics when complementary foods are introduced to their diet. Additionally, correlations between xenobiotics and the infant gut microbiome were explored. The results of the developed workflows demonstrated their potential, especially the untargeted platform, to gain a better understanding of the high variety of polyphenols present in different biological matrices, and their potential link to human health.