## 3 Results and Discussion
### Anti-Radical Activity of Free and Included Phloroglucinol along the Gastrointestinal Tract  
The antioxidant activity of a compound, expressed as its ability to interact and neutralise free radicals, can be affected by a broad range of factors. When considering oral intake, simulating the gastrointestinal conditions helps understand how factors, such as the pH of the different digestive fluids and interactions with digestive enzymes, may influence the antioxidant activity. In the case of an inclusion compound, the activity of an antioxidant guest molecule is further modulated by its affinity to the cyclodextrin host: guests with high affinity to their host are released more slowly to the medium, in what is deemed as a sustained release system. In this section, the effect of gastrointestinal fluids on the ability of phloroglucinol and its inclusion compound, \(\gamma\)-CD\(\cdot\)PG, to scavene two radical species, O\({}^{\bullet-}\) and NO\({}^{\bullet}\), was studied. For that, the compounds were immersed in a sequence of different media and times of exposure that mimic the digestive path: 2 min in an \(\alpha\)-amylase solution to simulate the oral cavity, followed by 1 h in presence of pepsin, to simulate the stomach, and finally the mixture was added with pancreatin and bile salts for 3 h to simulate the intestine. As expected, the scavenging activity of PG on both O\({}_{2}^{\bullet}\) - and NO\({}^{\bullet}\) scavenging assays decreased throughout the simulated digestive process, as perceived by the increase in the IC\({}_{50}\) values (Figure 7). This observation agrees with our previous studies on phlorotannins extracted from _Fucus vesiculosus_, which were found to be susceptible to the extreme conditions of the gastrointestinal fluids, consequently losing their antioxidant activity throughout the different compartments of the digestive tract [30]. In fact, the loss of integrity and bioactivity, such as antioxidant, is extremely common among phenolic compounds in general, which are well-known to be degraded and metabolised during their passage through the gastrointestinal tract [42; 43]. Figure 6: DTA traces of phloroglucinol (PG), \(\gamma\)-CD, their 1:1 physical mixture (1:1 mix) and inclusion compound \(\gamma\)-CD\(\cdot\)PG. Molecular encapsulation of PG with \(\gamma\)-CD appears as a possible approach to prevent this compound from being exposed to the gastrointestinal conditions, protecting it from degradation and ensuring its sustained release in the intestinal lumen, where it will be available for absorption. In fact, in both assays, the inclusion compound was shown to exert increased scavenging activity as it progressed from the oral to the gastric conditions, maintaining stable IC\({}_{50}\) values throughout the rest of the digestive process. These observations could be explained by the sustained-release effect of the \(\gamma\)-CD since exposure to the oral cavity fluid lasts only 2 min, which is a rather low contact time comparing with that from the following compartments (at least 1 h). It also indicates that time is a key factor for the encapsulated phloroglucinol to produce desirable effects. Interestingly, inclusion of PG into \(\gamma\)-CD resulted in a strong improvement of its scavenging activity against O\({}_{2}\)\({}^{\bullet}-\). This may be attributed not only to the protective effect of \(\gamma\)-CD against PG degradation, but also to a promoting effect of \(\gamma\)-CD on this reaction, in what is attributed to an enzyme mimicry activity of cyclodext [44,45]. This effect is not, however, universal. Indeed, this did not occur with the scavenging activity of NO\({}^{\bullet}\). This can possibly be explained by the fact that the scavenging activity of PG itself against NO\({}^{\bullet}\) is far more effective (in the order of \(\mathrm{\SIUnitSymbolMicro M}\)) than against O\({}_{2}\)\({}^{\bullet}-\) (in the order of mM).