Short communication Recovery of Saccharomyces cerevisiae sublethally injured cells after Pulsed Electric Fields M. Somolinos, P. Mañas, S. Condón, R. Pagán, D. García ⁎ Tecnología de los Alimentos, Facultad de Veterinaria, C/ Miguel Servet,177, 50013, Zaragoza, Spain ABSTRACT ARTICLE INFO Article history: Received 6 June 2007 Received in revised form 23 April 2008 Accepted 25 April 2008 Keywords: Saccharomyces cerevisiae Repair Recovery medium Pulsed Electric Fields pH Sublethal injury The objective was to investigate the influence of the recovery liquid medium on the repair of Saccharomyces cerevisiae sublethally injured cells after Pulsed Electric Fields (PEF) in media of different pH. Sublethal injury was detected in the yeast S. cerevisiae after 50 pulses at 12.0 kV cm - 1 in both pH 4.0 and 7.0, by using a selective medium plating technique. PEF treatments cause a repairable sublethal injury in S. cerevisiae. Injured cells showed their maximum repair capacity when suspended in Sabouraud Broth compared to Peptone Water or citrate–phosphate buffer of pH 4.0. The extent to which cells repair their injuries depended on the treatment medium pH, and on the nature of the storage medium. No repair was detected when the recovery liquid medium was citrate–phosphate buffer of pH 7.0. Acid conditions favour repair and survival of PEF-treated S. cerevisiae cells. This work contributes to the description of the mechanisms of PEF injury and inactivation, which would be useful in defining adequate PEF treatments, alone or in combination with additional hurdles, to assure food stability. © 2008 Elsevier B.V. All rights reserved. 1. Introduction Thermal processing of food allows obtaining safe products with a prolonged shelf-life by inactivating micro-organisms, among other effects (Knorr et al., 1994). However, the collateral effects of heat on sensorial and nutritional properties of food have increased the interest in the development of novel inactivation technologies, such as High Hydrostatic Pressure (HHP), Pulsed Electric Fields (PEF), amongst others (Mañas and Pagan, 2005). PEF technology has demonstrated its potential to inactivate spoiling and pathogenic micro-organisms without temperature increase (Barbosa-Canovas and Altunakar, 2006). However, there is a need to understand the way micro- organisms are inactivated. The description of the mechanisms involved would be useful in defining adequate and economic PEF treatments to assure food safety and stability. The most accepted theory to explain the death of micro-organisms under external electric field strengths was proposed by Zimmermann (1986). According to this theory, which is based on model systems such as liposomes, protoplasts or large eukaryote cells, such as erythrocytes and chloroplasts (Tsong, 1991; Ho and Mittal, 1996), PEF treatments would cause the accumulation of free charges at both membrane surfaces, which compresses the cytoplasmic cell mem- brane. At high electric field strengths, such compression would result in mechanical instability of the cell membrane and cell permeabiliza- tion, thereby accounting for cell death. Since in most micro-organisms the cytoplasmic membrane is surrounded by other external struc- tures, such as the outer membrane or cell walls, it is expected that these will interfere in the mechanism of inactivation. In fact, studies on the mechanism of bacterial inactivation by PEF have revealed a different and characteristic behaviour in Gram- positive and Gram-negative bacteria, in aspects such as the occurrence of sublethal injury after PEF or the influence of treatment medium pH on the PEF-resistance (García et al., 2005a). The occurrence of sublethal injury, measured by plating survivors in selective and non- selective media, was detected when Listeria monocytogenes and Ba- cillus subtilis were treated at pH 7.0 but not at pH 4.0. The opposite was true for Escherichia coli and Salmonella typhimurium. These results along with bacterial permeabilization studies (García et al., 2007), suggested that differences in structure and composition between the cell envelopes of Gram-positive and Gram-negative bacteria could effectively be responsible for the different behaviour observed under PEF. The study of these aspects on several distinct micro-organisms, such as yeasts, that show variations in cell organization and envelopes, would allow a better understanding of the mechanism of microbial inactivation by PEF. In a recent study carried out in our laboratory, we have demonstrated for the first time the occurrence of sublethal injury also in yeasts (Somolinos et al., 2007), and also the efficacy of sorbic acid in the inactivation of such injured cells. To obtain the maximal profit from this new technology and its combinations, it is needed a better understanding of the events linked to yeast cell injury and inactivation by PEF. The aim of this work was to investigate the influence of the recovery medium on the repair of Saccharomyces cerevisiae sublethally injured cells after a PEF treatment at pH 4.0 and 7.0. International Journal of Food Microbiology 125 (2008) 352–356 ⁎ Corresponding author. Tel.: +34 976 761581; fax: +34 976 761590. E-mail address: dgarcia@unizar.es (D. García). 0168-1605/$ – see front matter © 2008 Elsevier B.V. All rights reserved. doi:10.1016/j.ijfoodmicro.2008.04.023 Contents lists available at ScienceDirect International Journal of Food Microbiology journal homepage: www.elsevier.com/locate/ijfoodmicro