## 3 Results and Discussion
### Antioxidant Activity of Phenolic Compounds on Phosphatidylcholine Membranes  
In order to check the antioxidant activity of phenolic compounds (AP, RT, RS, FA), we monitored the peroxidation of phosphatidylcholine membranes induced by hydrophilic free radical generator AAPH, employing a DPH probe. DPH operates as a probe to reveal the oxidative events within the membrane, both due to lipid oxidation as well as its own oxidation. In both cases it is expected that the DPH fluorescence intensity decreases, due to a variation of the membrane structure (fluidity) as a result of lipid peroxidation and disruption of DPH molecules. The antioxidant activity of phenolic compounds in liposomes  
Figure 7: Zeta potential of PC liposomes in the presence of oleanolic acid (**a**), and oleanolic acid mixed with apigenin (**b**), rutin (**c**), resveratrol (**d**) and ferulic acid (**e**). was tested at 25 \({}^{\circ}\)C in the presence of AAPH after 10 min of incubation. The initial probe fluorescence intensity (Fo) was divided by the intensity after incubation with radicals at specified antioxidant concentrations. The lower the DPH Fo/F value is, the higher the antioxidant activity of the compound was recorded. As a result of free radical action on phosphatidylcholine liposomes in the absence of OA, DPH fluorescence decreased to about 62% of the initial value (Figure 8). After the addition of 5-100 \(\upmu\)mol OA, a reduction of fluorescence by about half was observed. This demonstrates the pro-oxidant effect of oleanolic acid in phosphatidylcholine membranes. In subsequent experiments, the impact of phenolic compounds on the peroxidation of liposomes containing 20 \(\upmu\)mol OA was analyzed. For apigenin and rutin (100 \(\upmu\)mol), a decrease of Fo/F value to 1.31 and 1.23, respectively, versus 1.85 and 1.92 in control samples, was observed. This indicates a pronounced antioxidant effect that was dependent on the flavonoid concentration. A weaker antioxidant activity was observed for ferulic acid: in this case, at 100 \(\upmu\)mol a reduction of Fo/F value to 1.44 was noted, compared to 2.16 in the control. On the contrary, for resveratrol, there was a pro-oxidant effect that manifested itself most clearly at concentrations of 50 \(\upmu\)mol and 100 \(\upmu\)mol, causing an increase of Fo/F to 1.94 and 2.13, respectively, compared to 1.69 in the absence of RS (Figure 8). The antioxidant activity of phenolic compounds is widely recognized. [PERSON] and co-workers [40] investigated the antioxidant activity of polyphenols added during preparation of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) liposomes and incubated with membranes after their hydration. As the agent that induces peroxidation, AAPH was used. Lipid peroxidation inhibition capacity (LPIC), expressed as Trolox equivalent, was equal to 0.36, 3.62 and 2.23 after the addition of, respectively, apigenin, rutin and ferulic acid to DOPC before membrane hydration. When polyphenols were added after liposome hydration, LPIC values changed and amounted to 2.38, 4.68 and 4.28, respectively [40]. The observed differences are due to the presence of phenolic compounds in the aqueous phase in the second variant of the experiment. This allowed them to directly scavene free radicals generated from water-soluble AAPH outside the membrane. Resveratrol (10-20 \(\upmu\)mol) is able to reduce the peroxidation rate of phosphatidylcholine liposomes in the presence of AAPH [41]. In this range of concentrations in our experiment, the anti- or pro-oxidant effect of RS was not pronounced. However, a clear pro-oxidation impact was noted at 50-100 \(\upmu\)mol. In an ORAC (Oxygen Radical Absorption Capacity) assay, resveratrol showed antioxidant activity up to 35 \(\upmu\)mol (8 \(\upmu\)g/mL), and in DPPH (2,2-diphenyl-l-picrylhydrazyl) radical-scavenging activity tests up to 280 \(\upmu\)mol (64 \(\upmu\)g/mL) [42].