ORIGINAL ARTICLE Biosynthetic requirements for the repair of sublethally injured Saccharomyces cerevisiae cells after pulsed electric fields M. Somolinos, D. Garcı´a, S. Condo ´ n, P. Man ˜ as and R. Paga ´n Departamento de Produccio ´ n Animal y Ciencia de los Alimentos, Facultad de Veterinaria, Universidad de Zaragoza, Zaragoza, Spain Introduction Pulsed electric field (PEF) treatment is a promising food preservation technique aiming to inactivate pathogenic and spoiling micro-organisms without temperature increase (Barbosa-Ca ´novas et al. 1999). Thus, PEF tech- nology is able to lessen the disadvantages of thermal treatments and, at the same time, to offer fresh-like qual- ity products. The achievement of this purpose requires, among other studies, to describe the mechanism of microbial inactivation adequately (Man ˜as and Paga ´n 2005). Microbial inactivation by PEF is believed to be due to the effects of PEF on cell envelopes (Hamilton and Sale 1967; Man ˜as and Paga ´n 2005; Garcı´a et al. 2007). Recent studies of our research group have demonstrated that PEF causes sublethal injury in bacteria (Garcı´a et al. 2003, 2005a) and yeasts (Somolinos et al. 2007) as a function of the treatment conditions. Whereas the occurrence of sub- lethal injury in bacteria depended on the type of bacteria and the pH of the treatment medium, PEF-injured cells of Saccharomyces cerevisiae and Dekkera bruxellensis were detected at any pH investigated in the range of 3Æ0–7Æ0. The use of the selective-medium plating technique con- firmed that PEF affects both the integrity and functional- ity of the membrane. The occurrence of sublethal damages in PEF-treated cells is a very relevant aspect to be taken into account, as injured cells have shown sensitivity to stress conditions after PEF. For instance, PEF-injured bacterial cells were inactivated under acid conditions in apple juice (Garcı´a et al. 2005b), and PEF-injured yeast cells died in the pres- ence of antimicrobial compounds such as sorbic acid (Somolinos et al. 2007). Thus, PEF has demonstrated to Keywords membrane damage, pulsed electric fields, repair, Saccharomyces cerevisiae, sublethal injury. Correspondence Rafael Paga ´ n Toma ´ s, Departamento de Produccio ´ n Animal y Ciencia de los Alimentos, Facultad de Veterinaria, Universidad de Zaragoza, C ⁄ Miguel Servet, 177, 50013 Zaragoza, Spain. E-mail: pagan@unizar.es 2007 ⁄ 1204: received 29 July 2007, revised 12 December 2007 and accepted 14 December 2007 doi:10.1111/j.1365-2672.2008.03726.x Abstract Aims: The aim was to evaluate the biosynthetic requirements for the repair of sublethal membrane damages in Saccharomyces cerevisiae cells after exposure to pulsed electric fields (PEF). Methods and Results: The partial loss of the integrity and functionality of the cytoplasmic membrane was assessed by adding sodium chloride to the recovery medium. More than 2 log 10 cycles of survivors were sublethally injured after PEF. Repair of sublethal membrane damages occurred when survivors to PEF were incubated in Sabouraud Broth for 4 h at room temperature. The addition of inhibitors, such as chloramphenicol, rifampicin, 5-fluorocytosine, nalidixic acid, cycloheximide, cerulenin, miconazol and sodium azide to the liquid repair medium showed that the repair of PEF-injured cells required energy and pro- tein synthesis. The extent of the sublethal damages was greater in PEF-treated cells at pH 4Æ0 than at pH 7Æ0. Conclusions: This work confirms that membrane damage is an important event in the PEF-inactivation of yeast. The mechanism of yeast inactivation by PEF seems to differ from that of bacteria, as the repair of sublethal damages requires protein synthesis. Significance and Impact of the study: Knowledge about the damages inflicted by PEF leads to a better description of the mechanism of yeast inactivation. Journal of Applied Microbiology ISSN 1364-5072 166 Journal compilation ª 2008 The Society for Applied Microbiology, Journal of Applied Microbiology 105 (2008) 166–174 ª 2008 The Authors