## 3 Results and Discussion
### Optimization of MAE Extraction Conditions
#### 3.2.1 Optimization of Extraction Conditions for the Content of Total Compounds
\begin{table}
\begin{tabular}{c c c c c c} \hline \hline  & \multicolumn{4}{c}{**Total Compounds**} \\ \hline
**Source** & **SS** & **DF** & **MS** & **F-Value** & **\(p\)-Value** \\ \hline X\({}_{1}\) & 1723.74 & 1 & 1723.74 & 230.05 & 0.0043 \({}^{\ast}\) \\ X\({}_{2}\) & 1071.32 & 1 & 1071.32 & 142.98 & 0.0069 \({}^{\ast}\) \\ X\({}_{3}\) & 24.51 & 1 & 24.51 & 3.27 & 0.2122 \\ X\({}_{11}\) & 898.99 & 1 & 898.99 & 119.98 & 0.0082 \({}^{\ast}\) \\ X\({}_{12}\) & 20.26 & 1 & 20.26 & 2.70 & 0.2418 \\ X\({}_{13}\) & 59.98 & 1 & 59.98 & 8.01 & 0.1055 \\ X\({}_{22}\) & 15.05 & 1 & 15.05 & 2.01 & 0.2921 \\ X\({}_{23}\) & 12.60 & 1 & 12.60 & 1.68 & 0.3242 \\ X\({}_{33}\) & 367.86 & 1 & 367.86 & 49.09 & 0.0198 \({}^{\ast}\) \\ Lack of fit & 112.07 & 3 & 37.36 & 4.99 & 0.1716 \\ Pure error & 14.99 & 2 & 7.49 & & \\ Total (corr.) & 4280.1 & 14 & & & \\ R\({}^{2}\) & 97.03 & & & & \\ Adj R\({}^{2}\) & 91.69 & & & & \\ \hline \hline \end{tabular} \(X_{1}\): temperature, X\({}_{2}\): % EtOH, X\({}_{3}\): Time, SS: Sum of squares, DF: Degree of freedom, MS: Mean square, R\({}^{2}\): Quadratic correlation coefficient; Adj R\({}^{2}\): Quadratic correlation coefficient adjusted, \({}^{\ast}\) Significant (\(p\) < 0.05).  
\end{table} 
Table 2: ANOVA test for the response variable of total compounds. Optimal conditions for the MAE of olive leaves have been reported previously in other research. [PERSON] et al. [35] reported the highest phenolic content, expressed as mg TAE/g power (dw), in 'Koroneiki', 'Roghani' and 'Mission' cultivar at 50% EtOH and 15 min. This study used a constant composition of different solvents (water, 80% methanol, 50% ethanol and acetone). [PERSON] et al. [36] found that the optimum conditions of 200 W, 8 min and 80% ethanol obtained the maximum value of eleuporein and flavonoids: 2.32% of oleuporein, verbascoside 631 mg Kg\({}^{-1}\) d.w., apigenin-7-glucoside 1076 mg Kg\({}^{-1}\) d.w., luteolin-7-glucose 1016 mg Kg\({}^{-1}\) d.w. in olive leaves. The optimum conditions found in these previous studies are different to ours because these studies did not evaluate temperature as the third MAE parameter and nor is it indicated even though it significantly affects phenolic recovery from olive leaves in MAE. At high temperatures, the solvent power increases because of a decrease in viscosity and surface tension, which helps the solvent to solubilize solutes and improves matrix wetting and penetration. In addition, in a closed vessel microwave extraction, the temperature exceeds the boiling point of solvent, leading to improved extraction efficiency [37]. Moreover, the pressure of the system increases the boiling temperature of the solvent [38]. There are few studies that have been carried out on the optimization of three MAE parameters, including the extraction temperature. [PERSON] et al. [28] reported the optimal MAE parameters of 80 \({}^{\circ}\)C, 80% methanol and 6 min to obtain a yield of 16.70% of fresh leafweight. [PERSON] et al. [39] optimized the MAE conditions for the highest content of total compounds in olive leaves by using deep eutectic solvents with optimum conditions of 79.98 \({}^{\circ}\)C, 15.28 min and 48.63% of water, to obtain 28.52 mg g\({}^{-1}\) d.w. of total compounds from olive leaves by HPLC-ESI-TOF-MS. Thus, there is a great difference between these MAE optimum conditions compared to those obtained in the present study. This could be because these previous studies used methanol and water as an extraction solvent. Nevertheless, ethanol has been shown to be a good solvent for phenolic extraction and also it is safe for human consumption [40]. Therefore, this study is the first time that the optimization of these MAE parameters has been carried out using a mixture of ethanol/water as an extraction solvent in order to obtain the maximum total compounds from olive leaves samples by HPLC-ESI-TOF-MS.