## 3 Non-Thermal Technologies in Winemaking Process
### Pulsed Electric Field (PEF) Effects on Microbial Populations, Phenolic Extraction, and Wine Quality  
Pulsed electric field is relatively novel non-thermal and emerging technology for food preservation that involves the application of short microsecond pulses of high electric field strength (5-50 kV/cm) to liquid or semi-solid foods placed between two electrodes [82]. In wine production, PEF technology has great potential due to its continuous operating mode [5]. Since PEF has the ability to inactivate microorganisms and enhance mass transfer by increasing cell membrane permeability via an electroporation mechanism (Figure 6), it presents a promising method for the reduction in wine spoilage microorganisms, extraction improvement of valuable compounds during winemaking, and acceleration of wine aging [11]. The main factors that can impact PEF efficiency in grape/wine processing can be divided into process parameters, properties of treated medium, and the microorganisms' characteristics [83]. The effects of PEF on grapes, must, and wine are shown in Table 4.  
Recently, [PERSON] et al. [47] investigated the effects of PEF on the microbial population in Tempranillo red wines aged in oak barrels without using additional preservatives. A PEF continuous flow system was used, operating at an electric field strength of 23 kV/cm with a pulse width of 8 \(\upmu\)s, specific energy of 95 kJ/kg, and a frequency of 330 Hz. The results showed significant inactivation of must yeasts (_Brettanomyces_, _Dekera_, _Pichia_) and lactic acid bacteria (_Lactobacillus_, _Oenococcus_, _Pediococcus_) in aged wines right after PEF treatment, which was also accompanied by a reduction in volatile phenols (4-ethylphenol and 4-ethylguaiacol) in wine with the highest population of _Brettanomyces_/_Dekera_. However, during the sampling period (5-25 months later), the recovery of some microorganisms (_Brettanomyces_/_Dekera_ and _Pediococcus_) in PEF-treated wines was observed. Similar findings were reported in another study by [PERSON] et al. [48], where PEF treatments (PEF alone and PEF with 15 mg/L SO\({}_{2}\)) applied to red wine just after maloalactic fermentation resulted in higher inactivation of yeasts, acetic acid, and lactic acid bacteria compared to one achieved only with SO\({}_{2}\) (30 mg/L) four days after treatments. After 6 months, the results showed that all treatments were effective for controlling most of microbial population, with the exception of lactic acid bacteria which was still viable in some of the wines. Moreover, PEF maintained wine sensory quality and provided wines with lower volatile acidity and greater color intensity despite reduced anthocyanin content. Figure 6: Pulsed electric field (PEF) mechanism of microbial inactivation in wine. T: temperature. This Figure was published in Preservatives and preservation approaches in beverages, Vol 15, [PERSON] and [PERSON]. [PERSON], Nonthermal preservation of wine, 203–235, Copyright Elsevier (2019) [75]. More recently, [PERSON] et al. [46] reported on the lethal effects (up to 4.0 log reductions) of both individual PEF treatments (15-25 kV/cm) and those that are moderate (15 kV/cm) combined with SO\({}_{2}\) (0-30 mg/L) on _S. cerevisiae_ and _O. Oeni_ in treated red wine after fermentation. After 4 months, PEF-treated wine after alcoholic fermentation had no viable yeasts, while lower than 1 \(\times\) 10\({}^{2}\) CFU/mL of _O. Oeni_ viable cells were detected in PEF-treated wine with 20 mg/L of SO\({}_{2}\) after malolactic fermentation. Moreover, applied PEF treatments did not affect oenological as well as wine sensory characteristics after the storage period. Similar conclusions were drawn by [PERSON] et al. [44], who reported that PEF treatments did not influence the physicochemical and aroma composition of Chardonnay white wine during 4 months of storage.