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This study investigated the feasibility of using hyperspectral imaging (HSI) to characterize the diffusion of acid and water within food structures during gastric digestion. Two different sweet potatoes (steamed and fried) and egg white gel (pH5 and pH9 EWGs) structures were exposed to in vitro gastric digestion before scanning by HSI. Afterward, the moisture or acid present in the digested sample was analyzed for calibration purposes. Calibration models were subsequently built using partial least-squares (PLS). The PLS models indicated that the full-wavelength spectral range (550–1700 nm) had a good ability to predict the spatial distribution of acid (Rcal2 > 0.82) and moisture (Rcal2 > 0.88). The spatiotemporal distributions of moisture and acid were mapped across the digested food, and they were shown to depend on the food composition and structure. The kinetic data revealed that the acid and moisture uptakes are governed by Fickian diffusion or by both diffusion and erosion-controlled mechanisms.

Salmonella Enteritidis is the most prevalent food-borne pathogen associated with egg-related outbreaks in the European Union. In order to colonise eggs, S. Enteritidis must resist the powerful anti-bacterial activities of egg white (EW). Possibly, the major EW antibacterial property is iron restriction, which results from the presence of the Fe3+-binding protein, ovotransferrin. To circumvent iron restriction, S. Enteritidis synthesise two types of catecholate siderophores, enterobactin and salmochelin, that can chelate iron from host iron-binding proteins. However, EW contains a lipocalin (Ex-FABP) that is known to bind enterobactin. Two other lipocalins, Cal-¿ and a1-ovoglycoprotein, are found in EW but their siderophore-binding potential was yet to be exploredThe aim of this project was to study the antimicrobial activity of three EW lipocalins through sequestration of bacterial siderophores synthesised by S. Enteritidis. Among those lipocalins, Cal-¿ and a1-ovoglycoprotein were shown to bind neither enterobactin nor salmochelin. Further, it was confirmed that Ex-FABP binds only enterobactin and not salmochelin. In standard growth media, S. Enteritidis escaped Ex-FABP-mediated growth inhibition thanks to salmochelin synthesis. However, no clear antibacterial activity was observed for Ex-FABP in EW. This surprising observation might be correlated with the weak growth of S. Enteritidis in EW and a lack of requirement for siderophores in persistence., Salmonella Enteritidis est le pathogène le plus fréquemment identifié dans les œufs et ovoproduits au sein de l’Union Européene. Pour coloniser les œufs, S. Enteritidis doit surmonter de nombreux obstacles liés à l’activité antibactérienne du blanc d'œuf (BO). Un des mécanismes antibactériens majeurs du BO est la restriction en fer imposée par l'ovotransferrine (protéine liant le fer ferrique, Fe3+). Pour pallier cette carence, S. Enteritidis est capable de synthétiser deux types de sidérophores, l’entérobactine et la salmochéline, capables d’acquérir le fer des protéines de l'hôte. Cependant, le BO contient également une protéine de type lipocaline (Ex-FABP) connue pour séquestrer l’entérobactine. Deux autres lipocalines, Cal-¿ et l’a1-ovoglycoprotéine ont été identifiées dans le BO, mais leur capacité à séquestrer des sidérophores restait à explorer.L'objectif de ce projet était d'étudier l’activité antimicrobienne de trois lipocalines du BO par leur capacité à séquestrer les deux sidérophores de S. Enteritidis. Les résultats montrent que Cal-¿ et l’a1-ovoglycoprotéine ne sequestrent ni l’entérobactine ni la salmochéline. Nos observations confirment qu’Ex-FABP lie uniquement l’entérobactine et non la salmochéline. En milieux de culture, S. Enteritidis échappe à l'inhibition de croissance induite par Ex-FABP grâce à la synthèse de salmochéline. Néanmoins, le rôle antimicrobien d’Ex-FABP via la séquestration de sidérophores n’a pas été observé dans le BO. Ceci pourrait être lié à la faible croissance de S. Enteritidis dans le BO et au fait que les sidérophores ne semblent pas être nécessaires à sa survie dans ce milieu.

Salmonella Enteritidis is the most prevalent food-borne pathogen associated with egg-related outbreaks in the European Union. During egg colonization, S. Enteritidis must resist the powerful anti-bacterial activities of egg white (EW) and overcome ovotransferrin-imposed iron-restriction (the most important anti-bacterial mechanism of EW). Many pathogens respond to iron restriction by secreting iron-chelating chemicals called siderophores but EW contains a siderophore-sequestering "lipocalin" protein (Ex-FABP) that is predicted to limit the usefulness of siderophores in EW. S. Enteritidis produces two siderophores: enterobactin, which is strongly bound by Ex-FABP; and the di-glucosylated enterobactin-derivative, salmochelin (a so-called "stealth" siderophore), which is not recognized by Ex-FABP. Thus, production of salmochelin may allow S. Enteritidis to escape Ex-FABP-mediated growth inhibition under iron restriction although it is unclear whether its EW concentration is sufficient to inhibit pathogens. Further, two other lipocalins (Cal-γ and α-1-ovoglycoprotein) are found in EW but their siderophore sequestration potential remains unexplored. In addition, the effect of EW lipocalins on the major EW pathogen, S. Enteritidis, has yet to be reported. We overexpressed and purified the three lipocalins of EW and investigated their ability to interact with the siderophores of S. Enteritidis, as well as their EW concentrations. The results show that Ex-FABP is present in EW at concentrations (5.1 μM) sufficient to inhibit growth of a salmochelin-deficient S. Enteritidis mutant under iron restriction but has little impact on the salmochelin-producing wildtype. Neither Cal-γ nor α-1-ovoglycoprotein bind salmochelin or enterobactin, nor do they inhibit iron-restricted growth of S. Enteritidis. However, both are present in EW at significant concentrations (5.6 and 233 μM, respectively) indicating that α-1-ovoglycoprotein is the 4th most abundant protein in EW, with Cal-γ and Ex-FABP at 11th and 12th most abundant. Further, we confirm the preference (16-fold) of Ex-FABP for the ferrated form (K d of 5.3 nM) of enterobactin over the iron-free form (K d of 86.2 nM), and its lack of affinity for salmochelin. In conclusion, our findings show that salmochelin production by S. Enteritidis enables this key egg-associated pathogen to overcome the enterobactin-sequestration activity of Ex-FABP when this lipocalin is provided at levels found in EW.

Introduction et but de l’étude: La pepsine, sécrétée dans l’estomac, est la première protéase rencontrée par les protéines alimentaires au cours de la digestion. L’estomac acidifie lentement les bols alimentaires et il a été montré que le pH gastrique a un impact majeur sur les cinétiques de protéolyse 1.En effet, il est généralement admis que la pepsine n’est active qu’à pH acide, pour pH ≤ 5, avec un maximum d’activité autour de pH 2 pour beaucoup de substrats 2.Elle est également représentée comme un diacide 3, ce qui semble vrai lorsqu’elle agit sur l’hémoglobine, substrat de référence pour mesurer son activité. Ce comportement en fonction du pH n’a pas été validé sur des mélanges de protéines alimentaires telles que les caséines, principales protéines du lait, ou les protéines du blanc d’œuf. L’objectif principal de cette étude est de comparer les profils d’activité de la pepsine en fonction du pH sur les caséines du lait de vache (CA) et les protéines du blanc d’œuf (BO).Matériel et méthodes: Les cinétiques d’hydrolyse de protéines de BO entier et de micelles de CA par la pepsine, à différents pH compris entre 1 et 8, ont été suivies au pH-stat pendant 2h, avec quantification du degré d’hydrolyse final (DH) par la méthode OPA. Les vitesses initiales ont été calculées à partir de ces cinétiques entre 0 et 5 minutes. Les concentrations en substrat et en pepsine étaient les mêmes pour chaque expérience, et l’environnement salin était celui du protocole INFOGEST 4. Les CA se présentent en structures supramoléculaires, les micelles, qui précipitent à pH acide. Une étape d’acidification à pH 4 des CA a permis d’étudier des suspensions d’agrégats tous formés dans les mêmes conditions. L’aspect et la taille de ces agrégats ont été observés par microscopie confocale et par morpho-granulométrie.Résultats et Analyse statistique: Les cinétiques d’hydrolyse obtenues suivent, dans leur majorité, des profils de type loi de puissance en fonction du temps. Pour les protéines de BO, le DH final est maximal à pH1 (14,4%) et diminue de manière linéaire en fonction du pH,jusqu’à atteindre des valeurs négligeables pour les pH ≥6. A l’inverse, les valeurs de DH finaux sur les CA ne sont pas significativement différentes sur la gamme de pH 1 à 5 (7,5 ± 1,4 %) puis diminuent à partir de pH 6 et atteignent des valeurs négligeables à pH 8.Le rapport entre la vitesse d’hydrolyse initiale et le DH final des CA est supérieur à celui des protéines du BO quel que soit le pH, suggérant une cinétique d’hydrolyse plus rapide pour les CA.Ces résultats montrent que les caséines du lait,dont le pH se situe un peu au-dessus de 6, n’ont pas besoin d’être longuement acidifiées pour que l’hydrolyse par la pepsine puisse commencer dans l’estomac, atteignant une activité quasi optimale de la pepsine dès pH 5.Conclusion: Les cinétiques d’hydrolyse par la pepsine en fonction du pH diffèrent donc entre un substrat de type protéines globulaires comme l’œuf, et les caséines du lait. Le profil d’activité de la pepsine en fonction du pH est également différent de celui du standard hémoglobine. En outre, contrairement à ce qui est généralement admis, la pepsine peut hydrolyser certains substrats à des pH plus hauts que pH 5.

National audience; Changing the structure of foods can modulate their nutritional quality by modifying the digestion process, a complex process that is still imperfectly understood. In particular, digestion dynamics have been poorly studied, although it can has significant metabolic consequences. The objective of this study is to model the proteolytic cascade by pepsin, and to highlight a potential "structural effect". Egg white is an interesting model to meet this dual objective, as it offers the possibility of obtaining gels with different structures. In a probabilistic modeling approach of the peptide cleavage dynamics, we sought to identify leverages for the cleavage of a peptide after a given digestion time. For this purpose, peptides are identified and quantified at different time points in an in vitro digestion experiment. A Generalized Additive Modeling of the probability of a cleavage based on the physicochemical profile of peptides is proposed, considering either all kinds of cleavages made by the pepsin or only those made on preferential peptide bonds. The most significant variables are related to the length of the peptide and its location on the ovalbumin sequence. These results suggest that the action of pepsin depends more on structural criteria than on the presence of specific cleavage sites.

Food microstructure and gastric digestion: the egg white gel model. STLOpendays

Conference: 11th International Symposium on Biometals (Biometals) Location: Ottawa, CANADA Date: 2018; Salmonella enterica serovar Enteritidis (SE) is the most frequently-detected Salmonella in foodborne outbreaks in the European Union. Among such outbreaks, egg and egg products were identifiedas the most common vehicles of infection. Possibly, the major antibacterial property of egg white is ironrestriction, which results from the presence of the ironbinding protein, ovotransferrin. To circumvent ironrestriction, SE synthesise catecholate siderophores (i.e. enterobactin and salmochelin) that can chelateiron from host iron-binding proteins. Here, we highlight the role of lipocalin-like proteins found in eggwhite that could enhance egg-white iron restriction through sequestration of certain siderophores, includingenterobactin. Indeed, it is now apparent that the egg-white lipocalin, Ex-FABP, can inhibit bacterialgrowth via its siderophore-binding capacity in vitro. However, it remains unclear whether Ex-FABP performssuch a function in egg white or during bird infection. Regarding the two other lipocalins of eggwhite (Cal-c and a-1-glycoprotein), there is currently no evidence to indicate that they sequestersiderophores.

In situ disintegration of protein gels (egg white) by pepsin based on time-lapse confocal microscopy . Première réunion plénière du GDR SLAMM (Solliciter LA Matière Molle)

The present study investigated proteolysis combined with heat treatment to make hen Egg White (EW) an efficient emulsifier. EW was hydrolyzed by protease (Thermoase®) at various Enzyme Concentrations (EC) (w/w, 0.1%, 0.2%, 0.4%, 0.8%), followed by heating at 90°C for 8 min. Results showed that optimal emulsifying ability and stability, determined by measurement of emulsion turbidity, were obtained when EC was 0.4%, followed by heat treatment at 90°C. The hydrolysate thus prepared had higher emulsifying ability and stability than either native egg white (nEW) or small molecular weight EW peptides (Runpep®), close to the properties of Egg Yolk (EY) which was a reference as a food emulsifier. Surface hydrophobicity (H0) was found to be linearly related to the emulsifying activity and stability of hydrolyzed egg white proteins.

Chicken egg white protects the embryo from bacterial invaders by presenting an assortment of antagonistic activities that combine together to both kill and inhibit growth. The key features of the egg-white anti-bacterial system are iron restriction, high pH, antibacterial peptides and proteins, and viscosity. Salmonella enterica serovar Enteritidis is the major pathogen responsible for egg-borne infection in humans, which is partly explained by its exceptional capacity for survival under the harsh conditions encountered within egg white. However, at temperatures up to 42 ˚C, egg white exerts a much stronger bactericidal effect on S. Enteritidis than at lower tempertaures, although the mechanism of egg-white-induced killing is only partly understood. Here, for the first time, the impact of exposure of S. Enteritidis to egg white under bactericidal conditions (45 ˚C) is explored by global-expression analysis. A large-scale (18.7% of genome) shift in transcription is revealed suggesting major changes in specific aspects of S. Enteritidis physiology: induction of egg-white related stress-responses (envelope damage, exposure to heat and alkalinity, and translation shutdown); shift in energy metabolism from respiration to fermentation; and enhanced micronutrient provision (due to iron and biotin restriction). Little evidence of DNA damage or redox stress was obtained. Instead, data are consistent with envelope damage resulting in cell death by lysis. A surprise was the high degree of induction of hexonate/hexuronate utilisation genes, despite no evidence indicating the presence of these substrates in egg white.

Background: Ovalbumin-related protein X (OVAX) is an uncharacterized ovalbumin-serpin. Results: This egg white-specific serpin lacks protease inhibitory activity, but unlike its ovalbumin homolog, OVAX exhibits antibacterial properties, partly through its heparin-binding site(s). Conclusion: OVAX, a non-inhibitory serpin is a heparin-binding molecule with antibacterial activity. Significance: OVAX participates in egg defense and constitutes a natural agent against Listeria and Salmonella., Ovalbumin family contains three proteins with high sequence similarity: ovalbumin, ovalbumin-related protein Y (OVAY), and ovalbumin-related protein X (OVAX). Ovalbumin is the major egg white protein with still undefined function, whereas the biological activity of OVAX and OVAY has not yet been explored. Similar to ovalbumin and OVAY, OVAX belongs to the ovalbumin serine protease inhibitor family (ov-serpin). We show that OVAX is specifically expressed by the magnum tissue, which is responsible for egg white formation. OVAX is also the main heparin-binding protein of egg white. This glycoprotein with a predicted reactive site at Lys367-His368 is not able to inhibit trypsin, plasmin, or cathepsin G with or without heparin as a cofactor. Secondary structure of OVAX is similar to that of ovalbumin, but the three-dimensional model of OVAX reveals the presence of a cluster of exposed positive charges, which potentially explains the affinity of this ov-serpin for heparin, as opposed to ovalbumin. Interestingly, OVAX, unlike ovalbumin, displays antibacterial activities against both Listeria monocytogenes and Salmonella enterica sv. Enteritidis. These properties partly involve heparin-binding site(s) of the molecule as the presence of heparin reverses its anti-Salmonella but not its anti-Listeria potential. Altogether, these results suggest that OVAX and ovalbumin, although highly similar in sequence, have peculiar sequential and/or structural features that are likely to impact their respective biological functions.

International audience; Egg white is an important contributor to the protection of eggs against bacterial contaminations during the first half of incubation (day zero to 12), prior to the egg white transfer into the amniotic fluid to be orally absorbed by the embryo. This protective system relies on an arsenal of antimicrobial proteins and on intrinsic physicochemical properties that are generally unfavorable for bacterial multiplication and dissemination. Some changes in these parameters can be observed in egg white during egg storage and incubation. The aim of this work was to characterize changes in the antibacterial potential of egg white in embryonated eggs (FE) during the first half of incubation using unfertilized eggs (UF) as controls. Egg white samples were collected at day zero, 4, 8, and 12 and analyzed for pH, protein concentration, and protein profile. Antibacterial properties of egg white proteins were evaluated against Listeria monocytogenes, Streptococcus uberis, Staphylococcus aureus, Escherichia coli, and Salmonella Enteritidis. During incubation, differential variations of egg white pH and protein concentrations were observed between UF and FE. At equal protein concentrations, similar activities against L. monocytogenes and S. uberis were observed for FE and UF egg white proteins. A progressive decline in these activities, however, was observed over incubation time, regardless of the egg group (UF or FE). SDS-PAGE analysis of egg white proteins during incubation revealed discrete changes in the profile of major proteins, whereas the stability of some less abundant antimicrobial proteins seemed more affected. To conclude, the antibacterial activity of egg white proteins progressively decreased during the first half of egg incubation, possibly resulting from the alteration of specific antimicrobial proteins. This apparent decline may be partly counterbalanced in embryonated eggs by the increase in egg white protein concentration. The antibacterial potential of egg white is very effective during early stages of embryonic development but its alteration during incubation suggests that extra-embryonic structures could then progressively ensure protective functions.

International audience; Proteins started being used as pharmaceuticals in the 1920s with insulin extracted from pig pancreas. In the early 1980s, human insulin was prepared in recombinant bacteria and it is now used by all patients suffering from diabetes. Several other proteins and particularly human growth hormone are also prepared from bacteria. This success was limited by the fact that bacteria cannot synthesize complex proteins such as monoclonal antibodies or coagulation blood factors which must be matured by post-translational modifications to be active or stable in vivo. These modifications include mainly folding, cleavage, subunit association, gamma-carboxylation and glycosylation. They can be fully achieved only in mammalian cells which can be cultured in fermentors at an industrial scale or used in living animals. Several transgenic animal species can produce recombinant proteins but presently two systems started being implemented. The first is milk from farm transgenic mammals which has been studied for 20 years and which allowed a protein, human antithrombin III, to receive the agreement from EMEA (European Agency for the Evaluation of Medicinal Products) to be put on the market in 2006. The second system is chicken egg white which recently became more attractive after essential improvement of the methods used to generate transgenic birds. Two monoclonal antibodies and human interferon-beta 1a could be recovered from chicken egg white. A broad variety of recombinant proteins were produced experimentally by these systems and a few others. This includes monoclonal antibodies, vaccines, blood factors, hormones, growth factors, cytokines, enzymes, milk proteins, collagen, fibrinogen and others. Although these tools have not yet been optimized and are still being improved, a new era in the production of recombinant pharmaceutical proteins was initiated in 1987 and became a reality in 2006. In the present review, the efficiency of the different animal systems to produce pharmaceutical proteins are described and compared to others including plants and micro-organisms.

Salmonella enterica serovar Enteritidis is the prevalent egg-product-related food-borne pathogen. The egg-contamination capacity of S. Enteritidis includes its exceptional survival capability within the harsh conditions provided by egg white. Egg white proteins, such as lysozyme and ovotransferrin, are well known to play important roles in defence against bacterial invaders. Indeed, several additional minor proteins and peptides have recently been found to play known or potential roles in protection against bacterial contamination. However, although such antibacterial proteins are well studied, little is known about their efficacy under the environmental conditions prevalent in egg white. Thus, the influence of factors such as temperature, alkalinity, nutrient restriction, viscosity and cooperative interactions on the activities of antibacterial proteins in egg white remains unclear. This review critically assesses the available evidence on the antimicrobial components of egg white. In addition, mechanisms employed by S. Enteritidis to resist egg white exposure are also considered along with various genetic studies that have shed light upon egg white resistance systems. We also consider how multiple, antibacterial proteins operate in association with specific environmental factors within egg white to generate a lethal protective cocktail that preserves sterility.

This work has been supported by the grant Reducing Egg Susceptibility to Contamination in Avian Production in Europe (RESCAPE) from the European Commission (FOOD-CT-2006-036018).; Bacillus cereus group bacteria are opportunistically pathogenic spore-forming microorganisms well known in the sector of pasteurized food products because of their involvement in spoilage events. In the sector of egg product processing, these bacteria may lead to important economic losses. It seemed then relevant to study their behavior in egg white, a widely used egg product usually recognized as developing different levels of antimicrobial activities depending on the environmental conditions. A strong bactericidal effect (decrease in the bacterial population of 6.1 ± 0.2 log CFU/ml) was observed for 68 B. cereus group isolates, independently incubated at 30°C in egg white at pH 9.3 (natural egg white pH). To determine which components could explain such a strong bactericidal effect, an experimental strategy was carried out, based on egg white fractionation by ultrafiltration and by anion-exchange liquid chromatography. The role of the protein fraction was thus demonstrated, and subsequent nano-liquid chromatography-tandem mass spectrometry analyses allowed identification of ovotransferrin as a major protein involved. The strong bactericidal effect was confirmed in the presence of commercial ovotransferrin. Such a bactericidal effect (i.e., a decrease in the bacterial population through cell death) had never been described because ovotransferrin is known for its bacteriostatic effect (i.e., inhibition of growth) due to its ability to chelate iron. Surprisingly, the addition of iron did not reverse the bactericidal effect of ovotransferrin under alkaline conditions (pH 9.3), whereas it completely reversed this effect at pH 7.3. Ovotransferrin was shown to provoke a perturbation of the electrochemical potential of the cytoplasmic membrane. A membrane disturbance mechanism could, hence, be involved, leading to the lysis of B. cereus group bacteria incubated in egg white.

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International audience; Bacterial membranes are often thought to be the main targets of the antimicrobial activity of egg white. In order to test this hypothesis, the state of the membranes of Escherichia coli K-12 cells during either bactericidal (45°C) or bacteriostatic (30°C) incubation in egg white at natural alkaline pH was studied by biochemical methods. Namely, the permeability of the outer membrane was evaluated through its ability to incorporate a hydrophobic fluorescent probe (1-N-phenylnaphthylamine), and the permeability of the cytoplasmic membrane was evaluated through the release of a specific intracellular enzyme (β-galactosidase). The bacteria were observed by atomic force microscopy in order to support the biochemical results. At 45°C, the outer membrane of E. coli K-12 incorporated the hydrophobic probe, suggesting that it was disrupted. In addition, the cytoplasmic β-galactosidase was released at this temperature. The atomic force microscopy analysis revealed the formation of spheroplasts, which provided further evidence of the cell wall disruption and a progressive release of cellular contents. At 30°C, biochemical and micrographic experiments confirmed that membrane integrity was preserved. These techniques provide a useful approach for studying the mechanisms of bacterial cell death in egg white.

Ce travail porte sur l’analyse de 63 échantillons de blanc d’oeuf dont 30 de blanc cru et 33 de blanc pasteurisé. Les analyses réalisées sont le dénombrement de la flore total à 30°C, l’étude des capacités d’altération des isolats après inoculation (de 3 à 5 log UFC/mL) dans de la crème anglaise et incubation à 10 °C pendant 21 jours. Les isolats altérants la crème anglaise sont ensuite identifiés par séquençage de l’ADN 16S puis caractériser selon leurs capacités à produire des protéases, lipases et amylases sur des milieux gélosés à base de crème anglaise. La révélation de ces activités enzymatiques sont basés sur la présence et la mesure d’un halo d’éclaircissement, la fluorescence due à la rhodamine B, ou la présence d’un halo après coloration au lugol pour les activités protase, lipase et amylase respectivement. Les résultats montrent que près de 80% des échantillons de blanc analysés étaient contaminés avec des niveaux de populations autour de 2±1,6 et 0,94±0,8 log UFC/mL du blanc cru et pasteurisé respectivement. L’ensemble des isolats étaient capables d’altérer la crème anglaise à 10 °C. Cette altération se caractérise par des niveaux de populations autour de 9,3 log UFC/ml, une diminution du pH, un épaississement, une apparition d’un déphasage, et une présence de bulles ou trous ainsi que des bruits à l’ouverture du tube dus à la production de gaz. La présence d’un de ces critères d’altération, variant selon les genres identifiés, a permis de distinguer 17 profils d’altération. L’identification de bactéries d’altération, a montré une prépondérance des genres Enterobacter et Escherichia dans le blanc d’oeuf cru et d’Enterococcus dans le blanc d’oeuf pasteurisé. Les autres genres identifiés sont Yersinia, Rahnella, Salmonella, Pantoae, Pseudomonas et Bacillus dans le cru et des genres tels que Enterococcus, Enterobacter, Escherichia, Serratia, Shigella et Klebsiella communs aux 2 types de blanc d’oeuf. L’ensemble des isolats testés est capable de produire des protéases mais à des niveaux variables (épaisseurs des halos de 1 à 6,2 mm). Soixante pourcent, cent pourcent et quatre-vingt pourcent des isolats présentent des activités lipase, protéase et amylase respectivement. Ce travail souligne l’importance de la maitrise des flores bactériennes issues des ovoproduits qui sont capable d’altérer des produits finis tels que des desserts réfrigérés., This work focuses on the analysis of 63 liquid egg white samples, including 30 raw and 33 pasteurized samples. The performed analysis are the total enumeration flora at 30 °C, the spoilage abilities of the isolates after inoculation (3 to 5 log CFU/mL) and incubation in the custard cream at 10 °C for 21 days. The spoilage isolates were then identified by DNA 16S sequencing and characterized according to their ability to produce proteases, lipases and amylases on agar custard based media. The revelation of these enzymatic activities are based on the presence and translucent halo extent, fluorescence due to rhodamine B, or the presence of a halo after Lugol staining for the protease, lipase and amylase activities respectively. The results show that about 80 % of egg white samples were contaminated with population levels around 2±1,6 and 0,94±0,8 log CFU/mL in raw and pasteurized white respectively. All isolates were able to spoil custard cream at 10 °C. The custard cream spoilage is characterized by a bacterial growth reaching level as high as 9,3 log CFU/mL, a decrease in the pH, thickening, an appearance of a phase shift and the presence of bubbles or holes as well as noises opening tube due to gas production. The presence of these alteration criteria, varying genera identified, allowed to distinguish 17 alteration profiles. The identification of spoilage bacteria showed a preponderance of Escherichia and Enterobacter in raw egg white and Enterococcus in the pasteurized one. Other identified types are Yersinia, Rahnella, Salmonella, Pantoae, Pseudomonas and Bacillus exclusively found in raw egg white and Enterococcus, Enterobacter, Escherichia, Serratia, Shigella and Klebsiella common to the two types of egg white. All isolates tested can produce proteases but at varying levels (thicknesses halos 1 to 6,2 mm). Sixty percent, one hundred percent and eighty percent of the isolates have lipase, protease and amylase activities, respectively. This work highlights the importance of controlling the bacterial flora from egg products which are able to spoil finished products such as chilled desserts.

National audience

La présente invention concerne l'utilisation d'un coproduit issu d'un procédé d'extraction du lysozyme à partir de blanc d'oeuf, pour l'obtention d'au moins une protéine basique de blanc d'oeuf. Ce coproduit est une solution obtenue à l'issue d'une étape d'extraction du lysozyme préalablement cristallisé à pH basique à partir d'un éluat contenant la fraction protéique de blanc d'oeuf retenue sur un support de chromatographie échangeur de cations. La présente invention concerne également un procédé pour la purification d'au moins une protéine basique de blanc d'oeuf, comprenant une opération d'extraction et de purification de ladite protéine basique à partir dudit coproduit issu d'un procédé d'extraction du lysozyme à partir de blanc d'oeuf., The present invention concerne to the use of a co-product from a method for extracting lysozyme from egg whites, in order to at least one basic Obtain egg white protein. Said co-product is a solution Obtained from a step of extracting pre-crystallized lysozyme Having a basic pH Containing the eluate from an egg white protein fraction was kept cation exchange chromatography substrate. The present invention concerne aussi to a method for purifying at least one basic egg white protein, Including an operation of extracting and purifying said using said basic protein co-product from a method for extracting lysozyme from egg white

Les protéines du blanc d'oeuf sont souvent utilisées dans l'industrie agro-alimentaire et pharmaceutique comme agents texturants. Elles ont, en particulier, d'excellentes propriétés interfaciales. Après l’adsorption des protéines, des concentrations très élevées, de l'ordre de plusieurs centaines de grammes par litre, ont été mises en évidence à l’interface air-solution. Afin de mieux comprendre la contribution de cette surconcentration aux propriétés interfaciales, nous avons compressé par dialyse à l’équilibre des solutions d’ovalbumine et de lysozyme, choisies pour leurs comportements différents à l’interface, sur une gamme de pressions osmotiques allant jusqu’à 50 bar pour établir l'équation d'état liant la concentration à la pression osmotique. Des différences entre ces protéines sont mises en évidence. Elles vont dans le même sens que celles observées à l’interface, mais sont inattendues par rapport à la biochimie. Il y a donc des vraies différences au niveau de la nature des protéines, et leur confinement, indépendamment du contact avec l’interface hydrophobe, joue sur leurs propriétés interfaciales.

L‟industrie des ovoproduits comporte un secteur dit « produits élaborés » parmi lesquels on retrouve les omelettes, les oeufs dur, les oeufs pochés… destinés à la restauration hors domicile. Certain de ces produits sont cuits directement dans l‟emballage pour de nombreuses raisons : augmentation de la durée de vie, meilleure qualité, meilleur rendement …Il s‟avère que, lors de la cuisson, des liaisons entre l‟emballage et le blanc d‟oeuf se créent et posent des problèmes lors de l‟ouverture des conditionnements. Par empirisme, la société a développé une formulation (un mélange d‟huile) qu‟elle pulvérise sur les films plastiques pour éviter ces problèmes d‟adhérence. Aujourd‟hui, elle souhaite éliminer cette formulation qui impose des contraintes qualité. En effet, ces huiles ne doivent pas être obligatoirement signalées sur l‟emballage, mais avec la nouvelle législation concernant l‟étiquetage, plus exigeante vis à vis des OGM et des allergènes, ces huiles vont perdre leur statut d‟agent technologique. Considérées comme additifs, elles devront apparaître dans la composition du produit sur l‟étiquette de l‟emballage. Pour cela, il est essentiel de comprendre les liaisons qui se mettent en place afin d‟envisager un moyen de les éviter. L‟un des objectifs de ce stage est de compléter la bibliographie concernant les interactions entres les protéines du blanc d‟oeuf et les films plastiques. Il est également important de référencer les techniques qui permettent de caractériser la présence ou non de protéines sur les films plastiques ainsi que les forces d‟adhésion présentes. Un autre objectif est de mettre en place des expérimentations simples qui permettront de définir les raisons de l‟adhésion afin d‟envisager un emballage plus adéquat.

Hen egg is a polyfunctional ingredient as it can bring concurrently different functionalities in formulated foods. Its emulsifying, foaming, gelling, thickening, colouring and aromatising properties are largely researched. At industrial scale, egg is transformed in egg products of first (whole egg, yolk, albumen) or second (classical recipes) transformation. This industry is rather recent and is knowing a continuous growth. During the last fifteen years, laboratory researches have brought important knowledge concerning constituents and different levels of egg structure, understanding of functional mechanisms and thermomechanical treatment impact. These knowledges serve now to better control functionalities of egg products in food applications. Furthermore, advances realised on extraction of minor constituents could serve for new applications in biological and pharmaceutical industry. The aim of this paper is to describe the different steps to obtain egg products, with a brief presentation of structure, constituents and functionalities of egg., L’œuf de poule est un ingrédient polyfonctionnel, c’est-à-dire qu’il peut remplir simultanément plusieurs fonctions technologiques dans un produit alimentaire formulé. Ses propriétés émulsifiantes, foisonnantes, gélifiantes, épaississantes, colorantes et aromatiques en font un ingrédient de base de la cuisine domestique et de l’agroalimentaire. Plus spécifiquement, le jaune d’œuf est l’agent émulsifiant par excellence, alors que le blanc d’œuf est une référence en termes de foisonnement. A l’échelle industrielle, l’œuf est transformé en ovoproduits dits de première transformation (œuf entier, jaune ou blanc, à l’état liquide, congelé ou poudre), ou dans certains cas cuit et/ou cuisiné, de manière à reproduire certaines recettes ménagères classiques ; on parle alors d’ovoproduits de deuxième transformation. Cette industrie relativement récente connaît depuis plusieurs années une croissance importante. Les connaissances sur les constituants, la structure de l’œuf et la compréhension des mécanismes mis en jeu au cours des traitements thermomécaniques, issues de la recherche développée depuis plus d’une vingtaine d’années, ont permis d’une part de mieux maîtriser les fonctionnalités des constituants de l’œuf et d’autre part d’adapter les procédés de transformation utilisés dans le secteur des ovoproduits. De même, les progrès réalisés dans l’extraction et la connaissance des constituants mineurs de l’œuf et de leurs propriétés biologiques ouvrent des perspectives de développement pour des applications hors du secteur de l’agroalimentaire (biologie, médecine, pharmacie). Cet article, a pour objectif de décrire les différentes étapes de fabrication des ovoproduits, après avoir abordé succinctement la structure, les constituants, et les principales applications de l’œuf et de certains de ses constituants.

L'oeuf renferme de nombreuses molécules contribuant à la protection de l'embryon, qui limitent en outre les toxi-infections alimentaires liées à sa consommation (Salmonelloses). Certaines sont connues dans le blanc d'oeuf (lysozyme, ovotransferrine) mais il existe d'autres protéines, telles que les antiprotéases, qui pourraient participer à la défense de l'oeuf. La régulation de leurs activités dans le blanc d'oeuf reste méconnue. Dans ce contexte, nous avons analysé l'activité bactériostatique et antiprotéasique de différents blancs d'oeufs non embryonnés (NE-E) conservés à 4°C, 20°C ou 37°C pendant 30 jours ou de blancs d'oeufs embryonnés (EE) incubés pendant 10 jours. Nous constatons que les blancs des oeufs conservés à 37°C perdent leur activité bactériostatique contre Salmonella et leur potentiel antiprotéasique, dès 14 jours de conservation. Aucune différence d'activité n'est observée dans les oeufs (NE-E) stockés à 4°C ou 20°C. Par ailleurs, on observe une diminution du potentiel antibactérien des blancs des oeufs embryonnés au cours de l'incubation alors que les activités antiprotéasiques restent identiques. Ces résultats suggèrent que la perte du potentiel antibactérien (E-E) n'est pas due à l'activité des antiprotéases. Les analyses du blanc d'oeuf en SDS-PAGE ont révélé l'apparition de bandes nouvelles au cours de l'incubation des oeufs à 37°C après 14 jours de conservation (NE-E) ou dès 4 jours d'incubation (E-E) mais l'analyse en spectrométrie de masse de ces bandes n'a pas permis d'identifier des molécules antimicrobiennes connues pouvant expliquer l'altération de l'activité bactériostatique observée.

High-intensity electric fields have been successfully applied to the destruction of Salmonella Enteritidis in diaultrafiltered egg white. The effects of electric field strength (from 20 to 35 kV x cm(-1)), pulse frequency (from 100 to 900 Hz), pulse number (from 2 to 8), temperature (from 4 to 30 degrees C), pH (from 7 to 9), and inoculum size (from 10(3) to 10(7) CFU x ml(-1)) were tested through a multifactorial experimental design. Experimental results indicate that, for Salmonella inactivation, the electric field intensity is the dominant factor with a strongly positive effect, strengthened by its positive interaction with pulse number. Pulse number, temperature, and pH have also significant positive effects but to a lesser extent. In the most efficient conditions, the pulsed electric field (PEF) treatment is capable of 3.5 log10 reduction in viable salmonellae. Simultaneously, the measure of surface hydrophobicity does not indicate any increase after PEF treatment. These results suggest that no protein denaturation occurs, unlike what is observed after comparable heat treatment in terms of Salmonella inactivation (55 degrees C for 15 min).

Background Egg white must provide nutrients and protection to the developing avian embryo. One way in which this is achieved is an arsenal of antimicrobial proteins and peptides which are essentially extensions of the innate immune system. Gallin is a recently identified member of a family of peptides that are found in egg white. The function of this peptide family has not been identified and they are potentially antimicrobial. Results We have confirmed that there are at least 3 forms of the gallin gene in the chicken genome in 3 separate lines of chicken, all the forms are expressed in the tubular cells of the magnum region of the oviduct, consistent with its presence in egg white. mRNA expression levels are in the order 10,000 times greater in the magnum than the shell gland. The conservation between the multiple forms of gallin in the chicken genome compared with the conservation between gallin and other avian gallin like peptides, suggests that the gene duplication has occurred relatively recently in the chicken lineage. The gallin peptide family contains a six cysteine motif (C-X5-C-X3-C-X11-C-X3-C-C) found in all defensins, and is most closely related to avian beta-defensins, although the cysteine spacing differs. Further support for the classification comes from the presence of a glycine at position 10 in the 41 amino acid peptide. Recombinant gallin inhibited the growth of Escherischia coli (E. coli) at a concentration of 0.25 μM confirming it as part of the antimicrobial innate immune system in avian species. Conclusions The relatively recent evolution of multiple forms of a member of a new defensin related group of peptides that we have termed ovodefensins, may be an adaptation to increase expression or the first steps in divergent evolution of the gene in chickens. The potent antimicrobial activity of the peptide against E. coli increases our understanding of the antimicrobial strategies of the avian innate immune system particularly those of the egg white and the evolution of the defensin family. The potential of this peptide and others in the family can now be investigated in a number of novel antimicrobial roles.