## 2 Results and Discussion
### Physicochemical Analysis
The effect of the PEF process is shown in Figure 1. Juice cloud at T0 resulted to be not only preserved but significantly (\(p\leq 0.01\)) improved, switching from 2.731 \(\pm\) 0.0007 to 2.749 \(\pm\) 0.0007, indicating that the PEF treatment and the correlated electroporation and dielectric breakdown of the juice cells did not produce detrimental effects on juice turbidity, which, in turn, was well maintained and enhanced. In line with these findings, the % residual PME activity measured in PEFJ immediately after the process (T0) was significantly (\(p\leq 0.01\)) lower with respect to UJ. Indeed, remaining PME activity equal to 22.03 \(\pm\) 0.006% was recorded in PEFJ at T0, thus showing that PEF treatment inactivated \(\sim\)80% of the PME activity recorded in UJ. PME is a heat-sensitive enzyme; therefore, it is currently inactivated in citrus juice by thermal pasteurization [30]. Our results are in line with most of the previous studies carried out on PEF inactivation of PME in orange juice. [PERSON] et al. [13] reported that a PEF treatment (35 kV/cm, 59 us) achieved 90% PME inactivation in \"Valencia\" orange juice, while [PERSON] et al. [25] achieved 80% inactivation in \"Navelina\" orange juice applying 35 kV/cm of electric field strength and 1500 us treatment time. Even if most of the available literature on this topic is in accordance with our findings, it must be highlighted out that [PERSON] et al. [31] reported that the PEF treatment (23 kV/cm monopolar pulses of 2 us) achieved only 34% PME inactivation. It  
Figure 1: Cloud values (**A**) and pectinethylesterase activity (**B**) in UJ (untreated juice) and PEFJ (PEF-treated juice) immediately after the process (T0). Data are expressed as means of three analytical replicates \(\pm\) standard errors. Means significant differences are expressed by different letters: \(p\leq 0.01\); \(n=3\). must be pointed out that PEF process parameters are very likely to influence the effect on the overall quality of the treated juice; therefore, different operative conditions, such as batch or continuous systems, pulse shape, frequency, energy density, pulse width, treatment time, geometric properties of the chamber, and maximum temperature reached during the process, could putatively lead to different PME inactivation results. During 20 days of refrigerated storage, both cloudiness values and PME activity showed a constant trend with no significant changes recorded at the different timepoints. Cloud values ranged from 2.749 \(\pm\) 0.0007 at T0 to 2.7495 \(\pm\) 0.0010 a T20, while PME activity, expressed as umol carboxyl released/min \(\times\) mL, ranged from 0.3550 \(\pm\) 0.019 at T0 to 0.3102 \(\pm\) 0.007 at T20. Thus, it can be concluded that PME activity was not restored and PEF treatment was effective in causing an irreversible deactivation of PME. Our results are in accordance with what was previously reported by other authors [13; 17; 25; 32], who stated that PME activity significantly decreased after PEF treatment and remained stable during refrigerated storage. Other authors [23] concluded, in their recent review, that the mechanism under which the enzymatic inactivation is achieved after PEF treatment is mainly attributable to changes in structure and conformation of the enzyme itself, which are responsible for the modification of the enzymatic activity. Moreover, these modifications are mainly dependent on the applied PEF system and PEF process operative parameters, especially the energy density.