Pulsed electric fields (PEF) have emerged as a promising technology to modulate microbial activity during fermentation processes. In this study, low-intensity PEF processing was applied to kefir grains (KG) before fermentation to stimulate biomass growth. A Box¿Behnken experimental design was employed to evaluate the effects of PEF parameters (electric field strength: 1¿5 kV/cm, pulse width: 8¿16 ¿s, and frequency: 5¿15 Hz) on KG biomass growth after 24 h of fermentation and on physicochemical characteristics of the obtained kefir beverage (KB), having untreated KG and KB as references. Response surface methodology was used to determine optimal processing conditions. Although most PEF treatments significantly influenced KG biomass growth, the treatment at 3 kV/cm, 133 pulses of 12 ¿s, and 10 Hz produced the greatest effect, increasing KG biomass by 16.59 ± 2.67% and significantly reducing KB pH. Scanning electron microscopy revealed no major structural damage in PEF-treated KG, although localized changes in protein extractability were observed. KB from PEF-treated KG exhibited higher titratable acidity than control KB, while no significant differences were observed in total phenolic content and antioxidant capacity. Finally, microbial populations evaluated through qPCR showed no significant differences in gene copy numbers of microorganisms between kefir mixes prepared with untreated and PEF-treated KG and their corresponding KB. These findings suggest that low-intensity PEF application in KG can stimulate biomass growth without compromising grain structural integrity or key physicochemical properties of KB, providing new insights into PEF application on KG and its potential to modulate complex symbiotic fermentation systems.
