# Accumulation of Vitamin C in Yeast under Pulsed Electric Field (PEF) Conditions
## 3 Results and Discussion
### Optimal PEF Conditions for Vitamin C Accumulation in Yeast
C1—control culture without vitamin C added to the medium and PEF treatment, C2—control culture with vitamin C added to the medium (5 mg/mL medium) and without PEF treatment, orange bars—cultures treated with PEF (vitamin C, 5 mg/mL medium; pulse width of 10 μs; treatment time, 10 min; number of pulses, 600; after 20 h of cultivation). Each value is the mean \(\pm\) standard deviation (\(n=3\)). Bars with the same letter (a–d) are not significantly different (\(p<0.05\)). Figure 4: Effect of treatment time on vitamin C accumulation in yeast cells (**a**). Effect of treatment time on the number of dead yeast cells and on biomass (**b**). C1—control culture without vitamin C added to the medium and PEF treatment, C2—control culture with vitamin C added to the medium (5 mg/mL medium) and without PEF treatment, orange bars—cultures treated with PEF (vitamin C, 5 mg/mL medium; voltage, 1000 V; pulse width, 10 μs; after 20 h of cultivation). Each value is the mean \(\pm\) standard deviation (\(n=3\)). Bars with the same letter (a–c) are not significantly different (\(p<0.05\)). Figure 5: Effect of cultivation time on vitamin C accumulation in yeast cells (**a**). Effect of cultivation time on the number of dead yeast cells and on biomass (**b**). C1—control culture without vitamin C added to the medium and PEF treatment, C2—control culture with vitamin C added to the medium and PEF treatment, C2—control culture with vitamin C added to the medium and PEF treatment, orange bars—cultures treated with PEF (vitamin C, 5 mg/mL medium; voltage, 1000 V; pulse width, 10 μs; treatment time, 20 min; number of pulses, 1200). Each value is the mean \(\pm\) standard deviation (\(n=3\)). Bars with the same letter (a–d) are not significantly different (\(p<0.05\)). Figure 3: Effect of pulse width on vitamin C accumulation in yeast cells (**a**). Effect of pulse width on the number of dead yeast cells and on biomass (**b**). C1—control culture without vitamin C added to the medium and PEF treatment, C2—control culture with vitamin C added to the medium and PEF treatment, C2—control culture with vitamin C added to the medium (5 mg/mL medium) and without PEF treatment, orange bars—cultures treated with PEF (vitamin C, 5 mg/mL medium; voltage, 1000 V; treatment time, 10 min; number of pulses, 600; after 20 h of cultivation). Each value is the mean \(\pm\) standard deviation (\(n=3\)). Bars with the same letter (a–d) are not significantly different (\(p<0.05\)). During the optimization experiment, we also studied the influence of PEF parameters on cell viability and biomass production. Figure 2b shows the effect of voltage on both tested features. The application of 400 V caused a decrease in biomass compared to sample C2, whereas for a voltage in the range 500-1000 V, no significant effect on biomass and cell viability was observed. [PERSON] et al. [27] reported that the intensity of the pulsed electric field did not significantly affect the viability of _S. cerevisiae_ cells. Moreover, other authors confirm that an electric field greater than 10 kV/cm is required to inactivate various types of yeast [28; 29]. In some cases, even higher viability of the yeast _S. cerevisiae_ was noted with increasing electric field strength and treatment time [30]. In the case of pulse width (Figure 3b), increasing the parameter value did not cause a significant decrease in cell biomass. The highest fraction of dead cells (4.5%) was observed at 50 \(\upmu\)s. [PERSON] et al. [31] showed that yeast cell death is dependent on the parameters of the electric field impulse and increases with increasing electric field intensity and the number of pulses. The duration of the process (Figure 4b) slightly influenced the number of dead cells. We noted the highest share of dead cells when the treatment time was 20 min, but it was only 5%. The biomass production for the same treatment time was only 9% lower than in the control not exposed to PEF. Longer PEF treatment of yeast cultures resulted in a significant reduction in biomass and an increase in the fraction of dead cells.