Session 12: Fundamental and mechanistic food research

Reaction kinetics at elevated pressures is of importance for the development of pressure-based technologies and especially for hyperbaric storage (HS), a potential alternative for the energy-consuming refrigeration. While the effect of pressure on reactions in equilibrium is well studied, the effect on reaction kinetics in foods is mostly unreported. Degradation of polyphenolic compounds is a set of reactions resulting and numerous degradation products. The effect of pressure on degradation kinetics of a polyphenol, EGCG (Epigallocatechin-gallate), was explored at pressures up to 200 MPa (HS levels) for several hours, with and without fructose. In a baro-resistant buffer, pressure enhanced EGCG degradation, due to a negative activation volume (ΔV^≠=-6.87 ± 1.54 ,), while in phosphate buffer the pH decreased (as is also expected to occur in many foods) resulting in a superposition of accelerating and protective effects. A previously undescribed protective, pressure-level dependent, effect of fructose was revealed. Novel in-situ spectroscopy and HPLC analysis revealed that in addition to the effect on EGCG degradation rate, pressure also modifies the ratios between the numerous degradation products, due to a varying effect on the steps involved in the degradation pathway. High-pressure enhanced the degradation rate of EGCG compared to storage at an identical temperature at atmospheric conditions, possibly negatively affecting quality under HS, yet due to other co-occurring effects, such as pH changes and the presence of co-solutes, a different outcome may prevail in foods. The effect of pressure was found to go beyond the influence on the degradation of the original compound, as likely further steps in the degradation pathway (that are not fully described even at atmospheric pressure) are differently influenced, resulting, at the end of the storage, in different ratios between the degradation products, with possibly varying combined (bio)functionality.

In these dynamic times, there is an evident need to keep reinforcing the safety of foods with sound scientific evidence. To this end, the food additive carrageenan (CGN) is at the heart of considerable debate^1-3. In fact, a 2018 EFSA panel suggested the instalment of an acceptable daily intake (ADI) for CGN to be set at 75 mg/kg body weight per day². This work will provide attendees with an overview of a series of studies interrogating the anti-nutritional effects of carrageenan on digestive proteolysis in toddlers, adults, and seniors. 

First, commercial κ-,ι- and λ-CGN preparations characterized by SEC-MALS and zeta-potential measurements indicate differences in MW, MW distribution, electrophoretic mobility and the existence of low MW CGN. Second, proteomic analyses of CGN-protein mixtures fed into a semi-dynamic in vitro digestion model highlight CGN ability to differentially modulate proteolysis and the bioaccessibility of milk-derived bioactive peptides in toddlers, adults and seniors⁵. Noting high CGN exposure in children^2,4, an effort was done to underpin CGN macromolecular characteristics governing its ability to attenuate the digestive proteolysis of milk proteins α-lactalbumin, β-lactoglobulin and lactoferrin in the gut of children. Further, PCA multivariate analyses (explaining 83.41% of variance) of flow behaviour, zeta-potential and particle size distributions (d4,3) at 3<pH<7 establish electrostatic biopolymer interactions as the strongest determinants of the digestive fate of a chocolate milk drink model system. Thus, this work highlights the need to better our understanding of CGN digestive fate in children and other liable populations.

The labelling of food ingredients with 14C atoms (isotopic labelling) is an exciting approach to monitor their fate during food processing, storage and adulteration. Most foodstuffs represent matrices with enormous complexity and permanently ongoing chemical dynamics. This complexity makes it difficult to decipher the fate of individual precursors or the generation of final or intermediate metabolites. Using 14C labelled compounds, their fate can be monitored along their full molecular life. and even of sub-molecular units, if labelling positions are selected respectively.

Food processing controls numerous chemical reaction cascades leading to desired and undesired reaction products with high importance for product quality and safety. Only by knowing these reactions in full detail enables the targeted optimisation of processes in order to boost or supress the formation of specific reaction products. Controlling the formation of neurological active metabolites of aromatic amino acids or supressing the formation of acrylamide are two examples.

Using the unique radioactivity of 14C-labelled precursors, their remains, metabolites, and fragments can easily be identified and monitored along food processing chains in complex matrices and metabolic events by radioactivity detection combined with high-resolution mass spectrometry. As seen in the case of Acrylamide, the fate of any ingredient may have a significant impact on food safety and quality. Comprehensive knowledge about specific reactions will help to preserve or generate desired compounds and help to inhibit process contaminants.

The presentation will address two examples of our recent research: the fate of pesticides during food processing and the fate of nutrients during food processing and digestion.

These results demonstrate that further investigations on the fate of chemicals in food processing have to be conducted and that realistic processing steps need to be considered e.g. for regulatory processes. Thereby it is possible to elucidate and finally assess potential hazards caused by unknown process metabolites. 

Many bitter compounds have important health benefits in mammals but their consumption can be incompatible with consumer acceptance. Perception of these bitter molecules in humans is mediated by 25 bitter taste receptors (TAS2Rs) and identification of molecules able to modulate the activation of these receptors can help in increasing the food palatability. The sophisticated TAS2Rs combinatorial coding scheme (one TAS2R binds many bitter compounds, the same compound can bind more than one TAS2R) and the lack of experimental 3D structures make our goal a challenging task. An accurate combination of computational methodologies (homology modelling, docking and virtual screening) for prediction of new TAS2Rs activators has been successfully applied by our lab using a structure-based approach [1]. This results were made possible despite the low sequence identity (~15%) with available templates, usually resulting in low resolution homology models that need to be furtherly refined.

Here, we focused our attention on one bitter taste receptor, TAS2R14, using a novel approach to predict not only activators but also inhibitors. Upon generation of TAS2R14 homology model [1], induced fit docking or normal mode analysis were used to increase the conformational space exploration of the receptor binding site. Evaluation of these receptor model’s ability to discriminate between active compounds and a list of decoys has been performed using docking followed by calculation of enrichment factors and ROC-AUC. The two homology models showing best performances in agonists or antagonists identification were selected and used to successfully suggest new compounds targeting the receptor through virtual screening of a large library of compounds. The promising results of this ongoing study suggest that this strategy, while already common in drug design, can be employed for discovery and design of new food compounds. [1] Di Pizio A, et al. Cellular and Molecular Life Sciences, 2020, 77, 531-542.