APPLIED MICROBIAL AND CELL PHYSIOLOGY Cyclopropanation of unsaturated fatty acids and membrane rigidification improve the freeze-drying resistance of Lactococcus lactis subsp. lactis TOMSC161 H. Velly & M. Bouix & S. Passot & C. Penicaud & H. Beinsteiner & S. Ghorbal & P. Lieben & F. Fonseca Received: 17 September 2014 /Accepted: 9 October 2014 # Springer-Verlag Berlin Heidelberg 2014 Abstract This work aimed at characterizing the biochemical and biophysical properties of the membrane of Lactococcus lactis TOMSC161 cells during fermentation at different tem- peratures, in relation to their freeze-drying and storage resis- tance. Cells were cultivated at two different temperatures (22 and 30 °C) and were harvested at different growth phases (from the middle exponential phase to the late stationary phase). Bacterial membranes were characterized by determin- ing the fatty acid composition, the lipid phase transition, and the membrane fluidity. Cultivability and acidification activity losses of L. lactis were quantified after freezing, drying, and 3 months of storage. The direct measurement of membrane fluidity by fluorescence anisotropy was linked to lipid com- position, and it was established that the cyclopropanation of unsaturated fatty acids with concomitant membrane rigidifi- cation during growth led to an increase in the freeze-drying and storage resistance of L. lactis. As expected, cultivating cells at a lower fermentation temperature than the optimum growth temperature induced a homeoviscous adaptation that was demonstrated by a lowered lipid phase transition temper- ature but that was not related to any improvement in freeze- drying resistance. L. lactis TOMSC161 was therefore able to develop a combined biochemical and biophysical response at the membrane level during fermentation. The ratio of cyclic fatty acids to unsaturated fatty acids (CFA/UFA) appeared to be the most relevant parameter associated with membrane rigidification and cell resistance to freeze-drying and storage. This study increased our knowledge about the physiological mechanisms that explain the resistance of lactic acid bacteria (LAB) to freeze-drying and storage stresses and demonstrated the relevance of complementary methods of membrane characterization. Keywords Lactic acid bacteria . Fermentation . Fatty acid composition . Fluorescence anisotropy . Lipid phase transition . Acidification activity . Lyophilization Introduction Lactic acid bacteria (LAB) are of great importance to the food and dairy industries because of both their central role as starters for manufacturing a diversity of fermented products and their potential health benefit (Daly et al. 1998; Leroy and de Vuyst 2004). Freeze-drying is widely used to provide ready to use highly concentrated starter cultures. This process makes it possible to minimize the degradation reactions and to maintain adequate physical, chemical, and biological stability of the product during long-term storage, even at ambient temperature (Franks 1998; Coulibaly et al. 2011). However, during this low-temperature desiccation process, bacteria are exposed to several environmental stresses (thermal, osmotic, water re- moval, and oxidative stresses), which could induce major cellular damages, leading to loss of viability and functional activities (acidification, production of aroma compounds and texturizing agents, etc.) (Coulibaly et al. 2010; Velly et al. 2014). Damages to cellular systems resulting from freeze- H. Velly CNIEL, 42 rue de Châteaudun, 75014 Paris, France H. Velly : M. Bouix : S. Passot : C. Penicaud : H. Beinsteiner : S. Ghorbal : P. Lieben : F. Fonseca INRA, UMR 782 Génie et Microbiologie des Procédés Alimentaires (GMPA), Avenue Lucien Brétignières, 78850 Thiverval-Grignon, France H. Velly : M. Bouix : S. Passot : C. Penicaud : S. Ghorbal : P. Lieben : F. Fonseca (*) AgroParisTech, UMR 782 Génie et Microbiologie des Procédés Alimentaires (GMPA), Avenue Lucien Brétignières, 78850 Thiverval-Grignon, France e-mail: fonseca@grignon.inra.fr Appl Microbiol Biotechnol DOI 10.1007/s00253-014-6152-2