Lysozyme Modification Using Proteolytic Enzymes.

The lysozyme in the chicken egg white consists of various bioactive amino acids. However, these compounds are inactive when they are in the sequence of parent proteins. They become active only when isolated from these proteins. The aim of this study was to modify lysozyme with proteolytic enzymes under specific conditions of the reaction environment so as to obtain active biopeptides. The physicochemical properties of the resulting preparations were also assessed. Our study showed that the modification of lysozyme with hydrolytic enzymes (pepsin and trypsin) under strictly specified conditions resulted in obtaining biopeptide preparations with new and valuable properties, as compared with native lysozyme. After the enzymatic modification of lysozyme, two structural fractions were distinguished in the composition of the resulting preparations-the monomeric fraction and the peptide fraction. The modified lysozyme exhibited high surface hydrophobicity and high total antibacterial activity despite the decrease in the hydrolytic activity. Modification of lysozyme with hydrolytic enzymes, especially pepsin, resulted in preparations with very good antioxidative properties.

The Evaluation of Activity of Selected Lactic Acid Bacteria for Bioconversion of Milk and Whey from Goat Milk to Release Biomolecules with Antibacterial Activity.

The aim of the study was to assess the antibacterial features of functional macromolecules released during the fermentation of goat milk and whey from goat milk by selected lactic acid bacteria strains that are components of kefir grain microflora. Two milk sources were used: goat milk and whey from goat milk. The lactic acid bacteria (LAB) and indicator microorganisms used were Lactobacillus plantarum PCM 1386, Lactobacillus fermentum PCM 491, Lactobacillus rhamnosus PCM 2677, Lactobacillus acidophilus PCM 2499, Escherichia coli PCM 2793, Salmonella enteritidis PCM 2548, Micrococcus luteus PCM 525, and Proteus mirabilis PCM 1361. The metabolic activity of LAB was described by the Gompertz model, and the parameters proposed for this experiment were the maximum rate of change of electrical impedance and potential biodegradability. Antibacterial activity was examined using the culture method in a liquid medium, determination of the reduction in indicator microorganisms, and optical density changes. Results show that the selective LAB produced certain active biomolecules with antibacterial activity from whey, a by-product that is sometimes troublesome for goat milk processors to manage. Lactobacillus acidophilus is a microorganism that is characterized by the highest metabolic activity in goat milk and whey from goat milk. It has the possibility to produce macromolecules with antibacterial activity.

Comprehensive Assessment of Environmental Pollution in a Poultry Farm Depending on the Season and the Laying Hen Breeding System.

The odors and dust emitted from hen houses affect human health and the condition of crops. The source of fumes is an element of the poultry house environment that affects the level of dust (litter and feed), the concentration of volatile compounds and the composition of the microflora (litter, dust and fodder). The research carried out as part of this study is a comprehensive assessment of the microbiological contamination (Pseudomonas, Enterobacteriaceae, and microscopic fungi) of all the elements that make up the environment of the poultry house (feed, litter, dust pollution and the atmosphere of the poultry house) in an annual cycle. The air from both types of farms is tested in terms of the quantity and quality of volatile compounds. Two types of laying hens reared on litter were compared: commercial and backyard farms. It was found that the seasons of the year and the system of keeping hens have a significant impact on the microbiological contamination with volatile compounds of the environment and the atmosphere of the hen houses. The obtained results of chemical, microbiological and questionnaire tests show that commercial farms carry a lower microbiological risk to the environment than backyard farm.

Assessment of Antimicrobial Properties of Phenolic Acid Extracts from Grain Infected with Fungi from the Genus Fusarium.

Problems related with biological contamination of plant origin raw materials have a considerable effect on prevention systems at each stage of food production. Concerning the antimicrobial action of phenolic acids, studies were undertaken to investigate antibacterial properties against bacterial strains of Escherichia coli (EC), Pseudomonas fluorescence (PF), Micrococcus luteus (ML) and Proteus mirabilis (PM), as well as antifungal properties targeting microscopic fungi Fusarium spp., extracts of phenolic compounds coming from inoculated grain from various genotypes of cereals. This study evaluated the antimicrobial action of phenolic acids extracts obtained from both naturally infested and inoculated with microorganisms. For this purpose a total of 24 cereal cultivars were selected, including 9 winter and 15 spring cultivars. The analyses showed a bactericidal effect in the case of 4 extracts against Micrococcus luteus (ML), 14 extracts against Pseudomonas fluorescence (PF), 17 extracts against Escherichia coli (EC) as well as 16 extracts against Proteus mirabilis (PM). It was found that 3 out of the 24 extracts showed no antibacterial activity. In turn, fungicidal action was observed in the case of 17 extracts against Fusarium culmorum (FC) (NIV), 16 extracts against FC (3AcDON), 12 extracts against Fusarium graminearum (FG) (3AcDON), while 12 other extracts showed antifungal action against FG (NIV) and 19 extracts against Fusarium langsethiae (FL). Based on the conducted analyses it was found that grain of small-grained cereals exposed to fungal infection is a source of bioactive compounds exhibiting antimicrobial properties. It was observed that the qualitative and quantitative profiles of polyphenols vary depending on the cereal cultivar. This extracts may be used to develop an antimicrobial preparation applicable in organic farming.

Antimicrobial Activities Evaluation and Phytochemical Screening of Some Selected Plant Materials Used in Traditional Medicine.

Plant extracts are a source of valuable ingredients that can be used in many industries. This paper presents research on the content of selected bioactive compounds in extracts obtained from various plant materials. Raw materials have a documented use in traditional medicine not only in Poland. The tested plants were: bitter melon (fruit), elderberry (flowers, fruit, leaves), wild rose (fruit, flesh, seeds), mountain ash (fruit), guelder rose (fruit), and sea buckthorn (fruit, leaves, pomace). The main goal of these tests is to indicate the potential raw materials that may constitute an alternative source of bioactive compounds with antimicrobial activity. The plant material was tested for the content of bioactive antioxidant compounds and possible antimicrobial activity. The content of polyphenols (phenolic acids and flavonoids) was analyzed using UPLC/PDA, sterols, organic acids, and other bioactive compounds. The minimum inhibitory concentration (MIC) was determined. The total free phenolic acids (TPC) and total free flavonoids (TFC) of all plant raw materials was varied and ranged from 0.21 (mg RUTE/1 g of extract) to 38.30 mg RUTE/1 g of extract) for TFC. The concentration of sterols was, on average, about 10 mg/1 g of extract. The value of approx. 20 mg/1 g of the extract was recorded for bitter melon and beach rose. The content of organic acids was about 1.5 mg/1 g of the extract to even 13 mg/1 g of the extract for sea buckthorn berries. The most sensitive to the extracts' activity were the following bacteria: M. luteus, P. mirabilis, P. fragii, S. enteritidis, and E. coli. The tested plant materials can be used in various industries as a source of bioactive compounds of an antibacterial nature.

Sambucus Nigra Extracts-Natural Antioxidants and Antimicrobial Compounds.

Due to the health-promoting properties of elderberry fruits, which result from their rich chemical composition, this raw material is widely used in herbal medicine and the food industry. The aim of the study was to demonstrate the antibacterial activity of the elderberry fruit extracts. The research showed that the content of phenolic acids and flavonoids in the extracts determined their antibacterial activity. The research showed that the content of phenolic acids and flavonoids in the extracts determined their antibacterial activity. The following phenolic acids were predominant: chlorogenic acid, sinapic acid, and t-cinnamic acid. Their average content was, respectively, 139.09, 72.84, 51.29 mg/g extract. Rutin and quercetin (their average content was 1105.39 and 306.6 mg/g extract, respectively) were the dominant flavonoids. The research showed that the elderberry polyphenol extracts exhibited activity against selected strains of bacteria within the concentration range of 0.5-0.05%. The following bacteria were the most sensitive to the extracts: Micrococcus luteus, Proteus mirabilis, Pseudomonas fragii, and Escherichia coli. Of the compounds under analysis, apigenin, kaempferol and ferulic, protocatechuic, and p-coumarin acids had the greatest influence on the high antibacterial activity of elderberry extracts. The results of the microbiological and chemical analyses of the composition of the extracts were analyzed statistically to indicate the bioactive compounds of the greatest antimicrobial significance.

Unconventional effects of long-term storage of microwave-modified chicken egg white lysozyme preparations.

Thermal modification is an effective method that induces significant expansion of the antimicrobial properties and other valuable properties of chicken egg white lysozyme. In our latest research, a new innovative method of enzyme modification was developed, in which microwave radiation was used as an energy source to process liquid lysozyme concentrate (LLC). After modification, high-quality preparations were obtained. However, long-term storage in a concentrated form initiated various processes that caused darkening over time and could also lead to other significant changes to their structure and, consequently, to their functional properties. This necessitated multidirectional research to explain this phenomenon. This paper presents the results of research aimed at assessing the physicochemical changes in the properties of microwave-modified lysozyme in the form of a liquid concentrate after long-term storage under refrigeration conditions. The assessment also considered the conditions under the acidity of the modifying medium and the duration of the microwave modification. The analysis showed that the values of the basic parameters determining the quality and usefulness of the modified enzyme significantly improved during long-term storage of the preparations. The greatest changes were observed in the preparations modified for the longest time and in the most acidic environment (process time 260 s, pH 2.0), the number of oligomers under these conditions increased by 18% after 12 months of holding, and the surface hydrophobicity increased by as much as 31%. In addition, microbiological tests showed that the preparations of microwave-modified lysozyme had an effect on gram-positive bacteria as well as on gram-negative, and this effect was significantly enhanced after 12 months. The results confirm that LLC modification with microwave radiation is a highly efficient method to prepare high-quality and high utility potential lysozyme. Notably, an interesting and important phenomenon was the observation of the unconventional behaviour of the preparations during their long-term storage, which increased their utility potential significantly.

The Influence of Packing Methods and Storage Time of Poultry Sausages with Liquid and Microencapsulated Fish Oil Additives on Their Physicochemical, Microbial and Sensory Properties.

The aim of the study was to evaluate the influence of refined fish oil additives in liquid and microencapsulated forms, packing method (VP-vacuum packing, MAP-modified atmosphere packing) and storage time (1, 7, 14, 21 days) on selected physicochemical, microbiological and sensory characteristics of minced poultry sausage. Principle component analysis (PCA) showed that the fish oil additive, packing method and storage time significantly influenced some of the physicochemical characteristics of the sausages. The pH value was negatively correlated with the type of sample and packing method. The water activity decreased along with the storage time. The sausages with microcapsules had distinguishable hardness, gumminess and chewiness than the other samples. This tendency increased in the subsequent storage periods. The packing method and storage time of the samples had a statistically significant influence on the growth of the total colony count and count of lactic acid bacteria (p < 0.05). The most aerobic bacteria were found in the control sample, and the least in the sample with microcapsules, regardless of the packing method. The use of MAP and the addition of microcapsules resulted in the lowest microbiological contamination of the sausages. The sensory analysis made by a trained panel did not show any significant differences between the samples. After 21-day storage of the sausages there was a slight decrease in some of the sensory parameters, e.g., color, smell, taste. The liquid oil and microencapsulated oil additives in the meat filling did not negatively affect the taste or any physicochemical characteristics of the meat products. From the microbiological perspective, there were better effects from the MAP method.

A Chemometric Approach to Oxidative Stability and Physicochemical Quality of Raw Ground Chicken Meat Affected by Black Seed and Other Spice Extracts.

The effects of black seed (Nigella sativa), allspice, bay leaf, caraway, cardamom, clove, and nutmeg extracts on the quality of raw ground chicken legs stored at 4 °C were investigated. During 12 days of storage, conjugated diene (CD) content, thiobarbituric acid reactive substances (TBARS), oxidation induction time (IP) by DSC (differential scanning calorimetry), hexanal content by GC-SPME-MS, thiol group (SH) content were determined. Moreover, microbial growth, pH and color of the samples were investigated. Sensory analysis was also realized. All extracts increased oxidative stability and safety of meat, significantly changed the color of the samples, stabilized the pH and increased their sensory scores (except color of samples with bay leaf and black seed) when comparing to control. Black seed, allspice and clove extracts showed high antioxidant capacity in lipid (CD = 0.23%, 0.28%, and 0.37%, respectively; TBARS = 0.55, 0.50, and 0.48 mg/kg, respectively) and protein fraction (SH content = 47.9, 52.1 and 52.7 nmol/g, respectively), although the ABTS•+ radical scavenging activity of black seed (33.1 µM/g) was significantly lower than the cloves (2496 µM/g) and allspice (815 µM/g). In the sensory analysis the highest scores were ascribed to the sample with cardamom followed by cloves. Principal component analysis (PCA) indicated complex and inseparable interrelationship among lipid and protein oxidation processes and the relationship of the protein oxidation on the lightness of meat. The results enabled to discriminate the meat samples, showing a great impact of the extracts on the final quality of raw chicken meat with black seed being potent antioxidant active additive.

The influence of the conditions of acquisition and storage of table eggs on changes in their quality and the presence of mycobiota and Fusarium mycotoxins.

The study showed that the hen housing system and egg storage conditions had significant influence on differences in the qualitative traits of eggs. The most rapid qualitative traits changes were observed in the eggs laid by the hens kept in the free-range and litter systems. After the first week of storage fungi were found on the eggshell surface but significant growth of fungi on the eggshell surface was noted after 3 wk, regardless of the storage conditions. Higher humidity during egg storage causes the dynamic development of fungi in the albumen. The principal component analysis proved the dependency between ergosterol and the trichothecene concentration in the albumen. Mycotoxins in the albumen indicate the presence of pathogenic species of microscopic fungi. The albumen of the eggs laid by the hens kept in cages had the lowest concentration of mycotoxins. The results of the study showed that the conditions of the henhouse environment significantly influenced the initial contamination of eggshells. Egg storage conditions, i.e., high air humidity, favor the development of fungi on the eggshell surface, affect the dynamics of fungal growth into the albumen and the production of mycotoxins.

Characterisation of the Mycobiota on the Shell Surface of Table Eggs Acquired from Different Egg-Laying Hen Breeding Systems.

Microbial safety is an important factor contributing to the egg quality. During egg acquisition, there is significant risk of contamination of the eggshell surface with microscopic fungi. Mycelial hyphae may grow on the eggshell surface and penetrate into the egg content. However, there is no information on the populations of microscopic fungi on the eggshell surface and, consequently, on possible production of mycotoxins. Therefore, the aim of the study was to identify the species of microscopic fungi present on the eggshell surface acquired from different breeding systems and to measure the number of selected mycotoxins. The qualitative analysis resulted in the identification of 41 isolates on the surface of eggs. There were 7 isolates from the organic production system, 11 from the free-range production system, 14 from the deep litter indoor housing system and 9 from the cage farming production system. The research proved that the diversification in the population of mycobiota on the eggshells depended on the egg-laying hen breeding system. The microscopic fungi isolated from the eggshells included toxigenic and pathogenic species such as Fusarium culmorum and F. equiseti. As the egg storage time increased, fungi, including the pathogenic species, penetrated through the eggshells. In consequence, mycotoxins were identified in the egg whites. Type-A and type-B trichothecenes were found in the eggshell samples containing F. culmorum.

Microbiological response of Japanese quail eggs to disinfection and location in the setter during incubation.

The aim of this study was to evaluate the effectiveness of ethyl alcohol (75%) disinfection of Japanese quail (Coturnix coturnix japonica) hatching eggs and analysis of microbial contamination of eggs during incubation, depending on their location in the setter. Disinfected eggshells were found to have lower total bacteria (TBC) and fungi (TFC) count. Concerning the vertical location of eggs (top, middle, bottom), disinfected eggs were characterized by similar values of the TBC (x = 1.54 log CFU/shell surface). For eggs without disinfection, it was found that those from middle and bottom levels of the setter had similar and lower TBC (by about 1.22 log CFU/shell surface) as compared to eggs from the top level. No statistically significant differences between levels were found in the case of TFC. Hatch breakouts (dead-in-shell embryos) from non-disinfected eggs were characterized by higher TBC (on average 0.37 log CFU/g). Disinfected eggs, located at the middle and bottom levels of the incubator, had similar and lower TBC in comparison with eggs from the top level. There were no microscopic fungi inside disinfected eggs of hatch breakouts. On the other hand, the non-disinfected eggs, placed on trays from the middle level of the incubator, had greater TFC (by about 0.9 log CFU/g) than those from top and bottom levels. Regardless of whether the eggs were disinfected or not, the largest group of microscopic fungi included Aspergillus and Penicillium.

Microflora and mycotoxin contamination in poultry feed mixtures from western Poland.

OBJECTIVE: Contamination of feeds with pathogenic microflora and mycotoxins constitutes a serious threat both for animals and humans. The aim of the study was to determine the degree of risk of the occurrence of microscopic fungi, selected bacteria and mycotoxins from the trichothecene group in poultry feeds in western Poland. RESULTS: In feed mixtures, the concentration of ergosterol (ERG), being a specific quantitative biomarker for the content of microscopic fungi, was determined. Grower and finisher feeds were characterized by a higher count of bacteria and fungi in comparison to starter feeds. A considerable variation was found in the amount of ergosterol in analyzed feeds. Mean ergosterol content in feeds amounted to 19.34 mg/kg. The most common genera of fungi detected in the tested feeds included Aspergillus, Rhizopus and Mucor. Irrespective of the type of feed, the proportion of trichothecenes group B was five times higher than that of trichothecenes group A in relation to the total content of these mycotoxins in samples. In terms of the analyzed mycotoxins, feeds contained the highest concentration of deoxynivalenol (DON). A statistically significant correlation was shown between DON and ERG and between total trichothecenes and ERG. CONCLUSION: Recorded results indicate that the level of microbiological contamination in feeds for broiler chickens produced in western Poland is within the requirements of the binding standards.

Effect of modified lysozyme on the microflora and sensory attributes of ground pork.

The effects of lysozyme monomer and thermochemically modified lysozyme on the microflora and sensory attributes of heated and unheated ground pork were investigated in this study. The dimer and trimer fractions of the modified lysozyme were 36.1 and 33.5%, respectively. The modified lysozyme exhibited higher hydrophobicity (40,600 U/mg of protein) and lower enzymatic activity (1,020 U/mg of protein) than the hydrophobicity (890 U/mg of protein) and activity (17,950 U/mg of protein) of the lysozyme monomer. Portions of ground pork (150 g) without lysozyme or supplemented with 5 mg/g lysozyme or modified lysozyme and either not heated or heated at 60°C for 10 min were stored at 4 ± 1°C and sampled at various times between 1 and 144 h. Meat color was not affected by either additive. After storage for 72 h, the mean odor score for meat supplemented with modified lysozyme and heated decreased from 5.0 at 1 h to 4.1, while the scores for all the other preparations were ≤3.2. After 144 h, the counts of Pseudomonas and Enterobacteriaceae in meat that was supplemented with modified lysozyme and not heated were, respectively, 1.1 and 0.9 log less than in the controls, and the numbers in such meat that was heated were, respectively, 1.2 and 2.4 log less than the numbers in the controls. The counts in meat supplemented with lysozyme and the controls were comparable. Heat treatment increased the bacteriostatic effect of modified lysozyme on gram-negative bacteria.

Potential possibilities of production, modification and practical application of lysozyme.

INTRODUCTION: Lysozyme, taking a stand in many biological fluids and tissues of a large number of living organisms, is a strongly basic protein. Hen egg white is its rich source and from this source enzyme can be obtained on a commercial scale as a preparation of biological activity. Monomer of lysozyme is known as hydrolase cutting the b-1-4 glycosidic bond, but its dimeric form received after modification of monomer form, exerts different and new valuable properties. In this study we indicated ways of production and modification of lysozyme and possibilities of its practical application. MATERIAL AND METHODS: The material for producing of lysozyme was fresh egg white. Enzyme was isolated by direct crystallization, ion-exchange chromatographic and utrafiltration methods. Lysozyme received by ion-exchange method has been used for modification. Modification of enzyme was carried out by chemical, chemical-thermal, thermal and membrane methods. RESULTS: The presented methods of lysozyme isolation from hen egg white yielded good results. Depending on employed procedure it was recovered from 20 to 85 % of enzyme. However, after modification approximately the quantity of 50-70% of polymerized enzyme was received, which contained from 30-40% of dimer. CONCLUSION: The method of isolating and modifying lysozyme can be successfully used to produce high active preparation of enzyme. Lysozyme monomer, and especially its modified form, shows the possibility of wide use not only in food industry, but also in medicine, pharmacology and veterinary medicine.

Sodium lactate addition on the quality and shelf life of refrigerated sliced poultry sausage packaged in air or nitrogen atmosphere.

The aim of this study was to determine the effect of sodium lactate addition on shelf-life extension of sliced poultry sausage packaged both in air and nitrogen atmospheres and stored in refrigerated conditions. Basic chemical composition, pH, and malonaldehyde content were assayed and color measurement using the reflection method was carried out. Microbiological examination consisted of determination of total number of aerobic psychrotrophic bacteria and number of lactic acid bacteria. Sensory evaluation of products was performed. Microbiological and sensory quality of sliced poultry meat sausage was dependent on the addition during production of sodium lactate and the composition of gases (air or nitrogen) used in packaging. Slices of poultry sausage with 1% as well as 2% of sodium lactate maintained their initial quality of evaluated sensory attributes longer, irrespective of the applied gases. Sodium lactate inhibited growth of aerobic psychrotrophic bacteria and lactic acid bacteria during refrigerated storage. Sodium lactate also inhibited the formation of malonaldehyde in sliced poultry sausage during refrigerated storage. The effectiveness of this process depended on the concentration of sodium lactate addition. It was concluded that 1% as well as 2% addition of sodium lactate could extend the shelf life of sliced poultry sausage packaged in air atmosphere and stored at 5 to 7 degrees C by 3 or 4 times, respectively. Sliced poultry sausage treated with 2% sodium lactate packed in nitrogen had the longest (35-day) shelf life. This was a sevenfold increase in the shelf life of sliced poultry sausage compared with the control.