# Study of the Phenolic Compound Profile of _Arbutus unedo_ L.
## 3 Results and Discussion
### Optimization of the Extraction Procedure of Phenolic Compounds
As can be seen in Figure 3d, the best results were found for the 1:12 (\(w/v\)) sample/solvent ratio, reaching a compromise between the TPC, the RSA, and the TAC. To the best of our knowledge, the extraction procedure of PC from strawberry tree berries has not been optimized in other previous published works and, in all of them, a very high volume of organic solvents was used, which implies a greater environmental impact and a more expensive extraction methodology. For example, [PERSON] et al. used 2 \(\times\) 50 mL of MeOH for the extraction of 1 g of sample during 12 h [5]. [PERSON] et al. extracted 1.5 g of sample with 25 mL of 96% EtOH for 30 min [25], and [PERSON] et al. employed 20 mL of acidic MeOH:water (50:50, \(v/v\); pH 2) and 20 mL of acetone/water (70:30, \(v/v\)) in a second extraction cycle for 0.5 g of sample [26]. More recently, [PERSON] et al. used 3 \(\times\) 30 mL of MeOH:water (80:20, \(v/v\)) to extract 1 g of sample during 1 h, followed by a purification step by solid-phase extraction (SPE) with C-18 SepPakVac 3 cc cartridges prior to HPLC analysis to remove sugars and other polar substances [23]. Therefore, the extraction procedure proposed in this work provides an improvement over other previous protocols, since a lower volume of organic solvent (5 mL) and less extraction time (1 min)are needed to obtain the extraction of PC, leading to a reduction of the environmental impact, the cost, and the time consumption of the process. TPC, RSA and TAC of Strauberry Tree Berries at Different Ripening Stages, Harvested in Different Years and at Different Geographical Locations  
The strawberry berries collected at three ripening stages in different years and in different geographical locations (Figure 1) were analyzed to determine their TPC, RSA, and TAC. Firstly, the moisture percentage was determined in all samples (Table 1). As it can be observed, the average water content was between 67.2-74.3% and 72.3-78.1% (\(w/w\)) for unripe and ripe fruits, respectively. This suggests the moisture content slightly increases with the maturation stage. The small differences observed in the moisture content during the three years evaluated are probably due to different annual environmental conditions, such as rain or temperatures, which contribute to fruit drying. In the ripe strawberry tree fruits, the moisture content was similar or slightly higher than those previously reported for samples collected in Portugal and Spain by other authors [5,27]. These variations are probably due to the different environmental and climatic conditions of the different geographical locations, such as water availability, rain, temperatures, sunlight, or wind exposure. In addition, these results confirmed that, similarly to other wild fruits, the moisture content of the strawberry tree berries is lower than for most conventional fruits (in general, between 80-95%). As shown in Table 1, the highest values of TPC and RSA were observed in the unripe berries in all the years evaluated. Conversely, the lowest TPC and RSA values were provided by the strawberry tree fruits collected at an intermediate ripening stage, whereas a slight increase was observed when the berries reached their ripe stage. Good correlation between the TPC and the RSA in the three ripening stages was observed, suggesting that the high antioxidant activity of _A. unedo_ L berries is related to its PC content. These results agree with the ones reported by [PERSON] et al., who showed lower TPC values in  
Figure 3: Evaluation of (**a**) solvent, (**b**) cycles of extraction, (**c**) type and time of agitation, and (**d**) sample/solvent ratio to optimize the strawberry tree berry extraction conditions. The TPC is expressed as mg GAE/100 g FW, the RSA is expressed as mg TE/100 g FW, and the TAC is expressed as mg cyn-3-glu/100 g FW. ripe berries than in unripe berries [26].