## 4 Discussion
The similar patterns of the contour plots reinforced the idea that observations on the specific activity plots (Figure 7) were caused indirectly, in which the activity was \"diluted\" by non-active co-extractives, rather than directly, through differences in the amount of extracted UV-absorbing molecules. Thus, ethanol, LSR, and time might be good parameters to adjust in order to target higher-purity/potency extractions, especially for applications where purification is not meant to be performed. The specific ORAC extract activity (Figure 4) was significantly impacted by all three variables under study. As expected, the more ethanol in the solvent, the higher the antioxidant activity. Alcohols were the preferred solvent for recovering antioxidant activity, as polar metabolites soluble in alcohols are often responsible for crude extract antioxidant activity [40]. In _G. turuturu_, antioxidant polyphenols, chlorophylls, and other hydroxyl-containing molecules occurred, such as polar carotenoids and tocopherols, all of which became more readily solubilized by the presence of ethanol in the solvent [19; 33; 41]. However, ORAC activity in water extracts from _G. turuturu_ is worth studying, as sulphated carrageenan can present this activity [22; 23] and their presence in a higher quantity might result in worthy application studies of these molecules. In the case of water extracts, both high volumes of solvent and long extraction times benefitted antioxidant recovery from _G. turuturu_. Actually, the 40 mL-g\({}^{-1}\) and 100 min used in this study as upper limits of these variables did not decrease and rather increased. However, 10 mL-g\({}^{-1}\) of solvent extracted for 100 min led to maximum ORAC activity, even higher than using 40 mL-g\({}^{-1}\) for 100 min. If enough extraction time was employed, then increasing volume worsened the extract's specific antioxidant activity, likely because it allowed the extraction of non-active components that \"diluted\" the active ones. For 50% (_v/v_) ethanol, the same conditions (low volume and high time) resulted in maximum ORAC activity as well. However, for this solvent, higher volumes invariably resulted in same-or-poorer results, even at low times of extraction, which indicates that the recovery of the ethanol-soluble antioxidants was limited by the rate of mass transfer, and not by solubility. Total ORAC activity (Figure 5) recovered per gram of biomass followed similar trends, in the case of 25% and 50% (_v/v_) ethanol. For water extracts, however, extraction under sub-optimal conditions regarding specific-activity, namely higher solvent volumes, regardless of time, because of an increase in yield, maximized antioxidant recovery to the point of obtaining maximum ORAC activity using 40 mL-g\({}^{-1}\) for 20 min. If the extraction was made under optimal specific-ORAC activity, i.e., 10 mL-g\({}^{-1}\) for 100 min, only 66% of total capacity available was recovered, despite being purer (higher per gram of extract). In any case, specific- and total-ORAC activities of _G. turuturu_ hydroethanolic extracts had maximums in the limits of time and LSR tested, meaning that higher values of ORAC might be available in _G. turuturu_ biomass if further optimization of these variables is performed. The overall analysis of LSR, time, and ethanol percentage effects on the three parameters (yield, ORAC, and UV-absorption) of _G. turuturu_ extracts suggested that ethanol percentage resulted in extracts fundamentally different at the chemical composition level, to a point where it should still be a variable under optimization in the study of temperature and pH effects. Thus, this variable was maintained in the second Box-Behnken design. For this second set of extractions, a LSR and time value was chosen by considering the overall effects observed in the first Box-Behnken, attempting to maximize yield, ORAC, and UV-absorption: 60 min of extraction and 40 mL-g\({}^{-1}\). Under these conditions, temperature and pH effects on different hydroethanolic extractions were characterized (Figures 6-10).