## 3 Results and Discussion
### Identification and Quantification of Phenolic Compounds in Brine  
Oleuropein and other \(\beta\)-glucosidase are the major fermentative substrates from the olive drupe; they are enzymatically hydrolyzed by \(\beta\)-glucosidase, which releases glucose and aglycones. The latter are then quantitatively degraded by esterase to simple and non-bitter phenolics, such as hydroxytyrosol and tyrosol. Some strains of LAB can accelerate the process, by hydrolyzing such glycosides as oleuropein and ligstroside--mainly responsible for the bitter taste of olives. The breakdown of oleuropein and ligstroside takes place in two phases: (i) hydrolysis of the glycosidic bond by \(\beta\)-glucosidase, with formation of oleuropein or ligstroside-aglycons and (ii) hydrolysis of the aglycons by esterase, with formation of elenolic acid and hydroxytyrosol (from oleuropein-aglycon) or tyrosol (from ligstroside-aglycon) [33; 44; 45]. The hydrolysis of oleuropein during fermentation is also attributed to the acid pH, coupled with the activity of \(\beta\)-glucosidase produced by oleuropeinolytic yeasts [40]. The concentrations of oleuropein, hydroxytyrosol, verbascoside, and tyrosol were monitored throughout the fermentation process (Figure 4). Oleuropein attained its maximum value, ca. 1200 mg/L, by 7 months. Within the same period, hydroxytyrosol increased steadily up to 1700 mg/L and went on increasing up to 2000 mg/L by the end of the process, although at a much lower rate. Verbascoside reached 1500 mg/L within just 2 months, and then increased slowly up to 1700 mg/L by the end of the process. Tyrosine was found to much lower levels (about one third) than the other phenolic compounds and exhibited a linear pattern throughout the process. Figure 3: Changes in (**a**) lactic, (**b**) acetic, (**c**) citric, and (**d**) succinic organic acids in brine during spontaneous fermentation of _Cobranosa_ table olives. As for oleuropein, the importance of the sweetening stage (with brine changes) in removing this compound is apparent, as it leaches off the pulp into the surrounding water. In the salting stage, LAB and yeasts able to degrade oleuropein (associated to a bitter taste) into hydroxytyrosol (associated to a non-bitter taste) accordingly attain a great importance. Hydroxytyrosol is considered the main marker to estimate oleuropein degradation, as well as the diffusion of phenols from fruit to brine [6, 13, 33]. Our results corroborate the continuous increase in hydroxytyrosol throughout fermentation _versus_ the oleuropein profile, which attains a maximum at 5-6 months (or end of sweetening stage); oleuropein reaches a maximum because it is released into the brine but simultaneously undergoes conversion to hydroxytyrosol by LAB and yeasts present therein, while hydroxytyrosol grows monotonically for being a final product. This growth is accompanied by significant growth of LAB--their population by ca. 6 months became slightly higher in number than the yeast population (ca. 1 log CFU/mL). Another explanation for the higher rate of production of hydroxytyrosol in the first 3 months (typically November to January) than in the later 3 months may be temperature (typically July to August, Table 1). [PERSON] et al. [46] indeed reported that inactivation of \(\beta\)-glucosidase during olive brining increases with increasing room temperature. Considering that the drums are kept inside warehouses not designed to maintain a constant temperature, the hydroxytyrosol profile developed during fermentation might also be influenced by environmental temperature. The evolution of the concentration of verbascoside agrees with [PERSON] et al. [47], who found verbascoside in all debittering methods of fermentation. Concerning table olive cultivars, it was different--as expected from the results reported by [PERSON] et al. [48] and [PERSON] et al. [49].