## 3 Results and Discussion
### Absorption Evaluation Using the In Vitro Dynamic Intestinal Model  
All the dietary polyphenols are characterized by an intestinal metabolization and a low intestinal absorption, which represent the main reason for the decrease in their bioactivities registered in the literature. Conversely, they seem to be stable at the gastric level in the presence of acidic conditions. In addition, polyphenol stability and bioaccessibility are strictly dependent on their chemical class and the composition and structure of the food matrix. Another important issue affecting the absorption data reported in the literature is the applied digestion method; in fact, the bioaccessibility values can vary widely in relation to the approach used to measure them [37; 38; 39]. In this study, a dynamic system was used to set-up the digestion key-step (i.e., the intestinal chamber (Figure 2)) by using Caco-2 cells, which are a well-known traditional model to study epithelial barrier function and transepithelial transporters expressed in these cells, such as P-glycoprotein (Pgp) and multidrug-resistant proteins (MRPs); these proteins clearly participate in the intestinal transport and absorption mechanisms of compounds [2]. To assess the functionality of the Caco-2 epithelial barrier plated in the millifluidic chambers, Transepithelial Electrical Resistance (TEER) [40] was measured at the starting (t0) and end (24 h) points of the experiment to monitor the intestinal barrier intercellular junctional integrity and the possible damage of the cellular monolayer over time. TEER values were measured by a voltmeter Millicell(r) ERS-2 Voltohnmmeter (Merck Millipore). The Caco-2 monolayer generates a stable TEER measurement of 250 \(\Omega\).cm\({}^{2}\), which is a value commonly reported in the literature for Caco-2 cells to confirm the corrected tight junction (TJ) integrity and monolayer permeability [40]. The effect of in vitro dynamic conditions on the tested molecule absorption was then investigated by collecting and analyzing the medium circulating in the system after the addition of the tested compounds, at a prefixed time. The analyses were performed by an RP-HPLC-DAD method, adapting the binary mobile phase to the different compounds with slight modifications to the existent literature [41; 42; 43; 44]. The percentage of each molecule in the extracellular fluid was considered as the non-absorbed fraction by Caco-2 cells and it was calculated at different times (from 1 to 24 h), considering the initial concentration (t0 = 0.1 mg/mL) as 100% of not-absorbed compound. Figure 3: Caco-2 cell viability (%) when exposed to 0.1 mg/mL of tested compounds for 6 and 24 h.  
The results obtained for each tested compound at different monitoring times are reported in Table 1.  
The chromatographic profiles registered during the monitoring period indicated that no metabolites were present, and therefore, the quantification was performed considering only the peak related to the tested molecule (Figures S1-S4). The most important monitoring time was 6 h, as it represents the time required for food to pass from the small intestine to the colon (normal range: 5-6 h), while 24 h can be considered the time required for a complete digestion [45]. Quinic acid (QA) (Figure 1a) was completely absorbed after 1 h as evident from the chromatographic profile where no peak was detected. In fact, 0.1 mg/mL QA was not toxic (Figure 3), and therefore, no loss of Caco-2 cell membrane integrity that could affect the absorption mechanism can be speculated, and apparently the obtained result could be attributed to a total absorption. However, the analysis of t\({}_{0}\) sample also revealed no peak, and therefore, QA behavior does not seem to depend on the contact between the compound and cell culture. In the literature, a high permeability has been attributed to QA, but with a resulting low intestinal absorption.