Sugar esters, substances made from bonding fatty acid tails to a sugar head, can play a number of key roles in food systems from antimicrobial agents to emulsifiers. These unique and very useful properties result from their water-loving and water-avoiding ends. Lactose, a sugar found in milk, based esters are important, as they are environmentally friendly and inexpensive, however, they are not very well understood. I created four different types of lactose esters: lactose monooctanoate (LMO), lactose monodecanoate (LMD), lactose monolaurate (LML) and lactose monomyristate (LMM), and then compared them to each other to see which would be the best emulsifier and which would be the best antimicrobial. My previous studies showed that LML was inhibitory against Listeria monocytogenes a common food pathogen. This encouraged us to evaluated the microbial inhibitory (bacterial killing) properties of LMO, LMD, LMM, along with LML, specifically, the influence of the fatty acid chain length in each ester and how that influenced my results. The esters, in order of highest microbial inhibitory properties, were LML, LMD, LMM followed by LMO. LML was inhibitory against all the Gram- positive bacteria tested including Bacillus cereus, Mycobacterium KMS, Streptococcussuis, L. monocytogenes, Enterococcus faecalis, and Streptococcus mutans but not Gram-negative bacteria (Escherichia coli O157:H7). Sugar esters are a large class of emulsifiers used in the food industry, and so my second research objective was to evaluate the influence of the fatty acid chain length on the emulsification properties of LMO, LMD, LML and LMM and compare them to each other and controls (Tween-20 and Ryoto L-1695) in a standard oil-in-water (O/W) emulsion. I did this by observing how long my emulsions lasted after mixing before they would start to separate. I also looked at the actual size of the oil drops in each of my emulsions, the smaller the oil drops remained, the better they stay in the emulsion and thus the more stable the emulsion. My results showed that the best emulsifier was LML, followed by LMD, LMM, and LMO, respectively. Therefore, my lactose esters contained both microbial inhibitory and emulsification activities. L. monocytogenes is an infamous food pathogen and one of the largest sources of food-borne illness from dairy foods in the United States. Addition of LMD and LML previously were shown to have microbial inhibitory effects in my lab so I wanted to see how well they would work in a food: milk. In general, bacterial deaths in the LMD milk samples were great and many times greater than the LML samples. However, both were greatly affected by milk fat content and how warm each of the samples were kept. LMD may play a useful role in increasing the safety of some foods.

Parmi les différentes méthodes de lutte contre les microorganismes pathogènes et/ou altérants en agroalimentaire, l’utilisation de bactéries lactiques (LAB) bioprotectrices s'avère être un outil prometteur pour la préservation des aliments. Ce travail de thèse collaboratif, entre l'équipe PAPC (AgroSup Dijon, Université de Bourgogne) et le laboratoire BioDyMIA (Université Lyon1-Isara Lyon), concerne l'étude de systèmes bioprotecteurs immobilisant des cellules entières de LAB dans une matrice polymérique d'alginate de sodium et de caséinate de sodium pour une activité ciblée contre Listeria spp. Dans un premier temps, la méthodologie mise en œuvre a consisté à sélectionner des souches de LAB bioprotectrices sur la base de leur activité antimicrobienne évaluée par la méthode de diffusion en milieu gélosé contre trois souches de Listeria spp. Quatre souches sur 19 ont ainsi été sélectionnées. Une caractérisation partielle des métabolites antimicrobiens produits par ces 4 souches a ensuite été réalisée en appliquant des traitements thermiques et enzymatiques aux surnageants de culture correspondants pour évaluer si ces traitements altéraient l’activité des métabolites antimicrobiens présents. Une purification et une identification partielle des actifs antimicrobiens de nature peptidique ont été réalisées uniquement pour la souche d'intérêt principale : Lactococcus lactis LAB3. Dans un second temps, une formulation de la matrice polymérique d’immobilisation des LAB sélectionnées a été choisie en réalisant le diagramme de phases du système aqueux alginate de sodium/caséinate de sodium : 1,5 % (m/m) d'alginate de sodium / 4 % (m/m) de caséinate de sodium / 20 % (m/m) bouillon MRS. Cette formulation a permis d'obtenir une matrice composée d’une phase continue riche en alginate et d’une phase dispersée riche en caséinate dans laquelle les cellules de LAB se localisent préférentiellement d’après les observations en microscopie de fluorescence confocale à balayage laser. Suite à l'inclusion des cellules de LAB dans ces matrices liquides et gélifiées d'alginate seul et d'alginate/caséinate, leur cultivabilité et leur activité anti-Listeria ont été suivies à 30°C pendant 12 jours. Ceci a révélé que la cultivabilité et l’activité antimicrobienne des cellules de LAB se maintiennent à des niveaux plus élevés dans les matrices d'alginate/caséinate que dans celles uniquement à base d’alginate. Ces matrices à base d’alginate et de caséinate apparaissent donc comme un système prometteur pour l'immobilisation de LAB bioprotectrices. Leur intérêt pour l’inclusion de LAB a pu être corrélé à leur viabilité et à la structure composite de cette matrice à base de protéines qui favoriserait la production et la libération des métabolites antimicrobiens / Among the various methods to control foodborne pathogenic and/or food spoilage microorganisms in food chain, bioprotective lactic acid bacteria (LAB) appear to be promising tools for food biopreservation. This collaborative study, between PAPC (Agrosup Dijon, University of Burgundy) and BioDyMIA (University Lyon1-Lyon Isara) laboratories, concerned the development of sodium alginate/sodium caseinate polymeric matrices intended to entrap LAB cells selected for their anti-Listeria spp. activity. First, 4 LAB strains from 19 LAB strains were selected for their anti-Listeria spp. activity: this screening was performed by the method of agar diffusion against three Listeria spp strains. Then, antimicrobial metabolites produced by the selected LAB strains were partially characterized by assessing the effect of various thermal and enzymatic treatments on the anti-Listeria spp. activity of their culture supernatants. A partial purification and identification of antimicrobial active peptides produced by the main strain of interest (Lactococcus lactis LAB3) was also performed. A composition of the polymer matrix has been selected by performing the phase diagram of sodium alginate/sodium caseinate system: 1.5% (w/w) sodium alginate / 4% (w/w) of caseinate sodium / 20% (w/w) MRS broth. This formulation provides a rich alginate continuous phase and a rich caseinate dispersed phase in which LAB cells localize according to the study by confocal microscopy. LAB cells were immobilized in liquid and gelled matrices of alginate and alginate/caseinate. Culturability and anti-Listeria activities were measured during a storage at 30°C for 12 days. The alginate/caseinate matrices were more effective in better maintaining LAB cells cultivability and their antimicrobial activity than alginate matrix. This effectiveness seemed correlated with cell viability and the dispersion-like structure of the protein-based system which enhance production and release of antimicrobial metabolites. Thus, this type of polymeric matrix appeared as a promising immobilization system of bioprotective LAB

Seafood permits the transmission of many bacterial pathogens. In order to reconcile consumer demands with important safety standards, traditional means of regulating microbial spoilage and safety hazards in foods are combined with novel technologies. These include biological antimicrobial systems, such as the use of lactic acid bacteria (LAB) and/or their bacteriocins, such as Carnobacterium maltaromaticum CS526 and its bacteriocin piscicocin CS526. The aims of this study were to investigate the presence of Listeria monocytogenes in temperate seafood, namely fresh and smoked salmon, fresh and smoked haddock, and fresh mussels and oysters. Additionally, there was an aim to recover, characterise and use bacteriocin-like-substance to control Listeria monocytogenes in cold smoked haddock. Vibrio spp., Enterobacteriaceae representatives, total aerobic heterotrophic counts and Listeria monocytogenes were isolated from commercially prepared smoked and fresh Atlantic salmon, smoked and fresh haddock, live mussels and oysters using selective media and tryptone soya agar (TSA). Vibrio spp. occurred in high densities (>106 CFU gˉ1) in mussels and Enterobacteriaceae representatives were recorded at >106 CFU gˉ1 in fresh salmon. Total aerobic heterotrophic counts in fresh salmon, live mussels and oysters reached 107, > 107, and > 106 CFU gˉ1, respectively. Listeria monocytogenes was recorded at 5.0 x 104 CFU gˉ1 in mussels. In total sixty one bacterial isolates were recovered from the seafood examined. The results revealed 19 genera of bacteria, i.e. Acinetobacter, Aerococcus, Aeromonas, Bacillus, Brochothrix, Carnobacterium, Citrobacter, Corynebacterium, Enterobacter, Escherichia coli, Moraxella, Micrococcus, Pseudomonas, Psychrobacter, Serratia, Shewanella, Staphylococcus, Vibrio and Listeria. The prominent characteristics of fish spoilage isolates were demonstrated by the ability of the isolates to reduce trimethylamine oxide (TMAO) to trimethylamine, and to produce H₂S. Sh. baltica OS185, Aeromonas spp. HB-6, Sh. baltica, Sh. putrefaciens, A. hydrophila HX201006-3, A. salmonicida subsp. achromogenes, A. hydrophila, C. freundii, Enterobacter cloacae were strong producers of TMA and H₂S. The spoilage microorganisms were tested for potential pathogenicity. The result revealed that 6/15 of the spoilage microorganisms produced proteolytic, lecithinase, blood (β and α haemolysin) and elastinase activity, respectively, whereas 7/15 of the spoilage microorganisms showed lipolytic activity. Cell free supernatants, ammonium sulphate precipitated supernatants and semi-purified bacteriocin-like substances of Carnobacterium maltaromaticum MMF-32 and KOPRI 25789 producing strains isolated from commercially prepared smoked salmon were investigated for their potential antimicrobial activity against potentially pathogenic and food spoilage microorganisms. Generally, a broad spectrum of activity was revealed against potentially pathogenic and food spoilage microorganisms in vitro. Cold-smoked haddock treated with bacteriocin producing C. maltaromaticum MMF-32, C. piscicola A9b bacˉ phenotype nonbacteriocin producing strain a mutant of C. piscicola A9b bac+, cell free supernatants, ammonium sulphate precipitated supernatants and semi-purified bacteriocin-like substances was challenged with L. monocytogenes ATCC 19114 up to 103 CFU gˉ1, respectively. Samples were stored at 4 °C for 10 days. L. monocytogenes and total bacterial counts were determined along with changes in total volatile base nitrogen (TVBN) and biogenic amines production as well as texture, colour and odour. Although the study on anti-listerial effects of C. maltaromaticum MMF-32 was not successful, this organism did have a positive effect on retention of firmness and sensory perception in cold smoked haddock.