Mixture Of Lysozyme Research Articles

Temperature-dependent dynamics at protein-solvent interfaces.

We perform differential scanning calorimetry, broadband dielectric spectroscopy (BDS), and nuclear magnetic resonance (NMR) studies to understand the molecular dynamics in mixtures of ethylene glycol with elastin or lysozyme over broad temperature ranges. To focus on the protein-solvent interface, we use mixtures with about equal numbers of amino acids and solvent molecules. The elastin and lysozyme mixtures show similar glass transition steps, which extend over a broad temperature range of 157-185K. The BDS and NMR studies yield fully consistent results for the fastest process P1, which is caused by the structural relaxation of ethylene glycol between the protein molecules and follows an Arrhenius law with an activation energy of Ea = 0.63eV. It involves quasi-isotropic reorientation and is very similar in the elastin and lysozyme matrices but different from the α and β relaxations of bulk ethylene glycol. Two slower BDS processes, viz., P2 and P3, have protein-dependent time scales, but they exhibit a similar Arrhenius-like temperature dependence with an activation energy of Ea ∼ 0.81eV. However, P2 and P3 do not have a clear NMR signature. In particular, the NMR results for the lysozyme mixture reveal that the protein backbone does not show isotropic α-like motion on the P2 and P3 time scales but only restricted β-like reorientation. The different activation energies of the P1 and P2/P3 processes do not support an intimate coupling of protein and ethylene glycol dynamics. The present results are compared with previous findings for mixtures of proteins with water or glycerol, implying qualitatively different dynamical couplings at various protein-solvent interfaces.

Fabrication of food-grade Pickering high internal phase emulsions (HIPEs) stabilized by a dihydromyricetin and lysozyme mixture

This study evaluated the feasibility of fabricating food-grade HIPEs using a dihydromyricetin and lysozyme mixture. The effects of the oil phase volume fraction (φ), composition (lysozyme:dihydromyricetin, k), and addition amount (w) of the mixture on the formation and properties of the HIPEs were analyzed. Then, the interactions of dihydromyricetin and lysozyme were investigated. The results indicated that when w was 0.4%, HIPEs with φ value of 90% could be obtained. Furthermore, the k also affected the microstructure, mechanical properties, oil oxidation, and lutein protection ability of the HIPEs. However, the presence of dihydromyricetin did not affect lysozyme activity. Both isothermal titration calorimetry and molecular simulations proved that they did not form a typical host–guest complex. But, dihydromyricetin could absorb on the lysozyme surface. Therefore, we speculated that lysozyme and dihydromyricetin particles could overlap and form a 3D network structure to stabilize the HIPEs, which was consistent with the microstructure observations.

Eggshell derived hydroxyapatite microspheres for chromatographic applications by a novel dissolution - precipitation method

Hydroxyapatite ((Ca10(PO4)6(OH)2, HAp) based chromatography matrix has attracted great interest in the field of protein separation. However, researchers have been trying to combat the growing costs associated with the HAp matrix. In the present investigation, we utilized a cheap biological waste material, viz. eggshells, for the development of hydroxyapatite (HAp) resins and evaluated them for protein purification. Initially, the calcite of the eggshell carbonate was converted into metastable vaterite microspheres. The HAp microspheres (ECHAp) were then prepared from eggshell carbonate microspheres using a novel dissolution-precipitation process. Synthetic source calcium carbonate was also used to prepare HAp microspheres (CHAp) for comparison. The purity and morphology of the apatite microspheres were characterized using X-ray diffraction (XRD) method, Fourier Transform Infrared spectroscopy (FTIR), Scanning Electron Microscopy (SEM), and laser diffraction particle analysis. Although both the apatites have similar morphology, the ECHAp has a larger surface area of 33.8 m2 g−1 compared to CHAp of 17.27 m2 g−1 by surface area analysis method. A commercial HAp matrix (CHT) with similar properties was also studied for comparison. All the apatite microspheres were found to have a similar protein binding capacity for bovine serum albumin (BSA). But ECHAp showed better protein separation for BSA and lysozyme mixture compared to CHAp and CHT matrices. The ECHAp matrix was also found to be highly stable over 20 purification cycles. Hence, the eggshell waste seems to have the potential for HAp matrix by a novel carbonate route with ease of preparation and also an economical packing material for chromatographic purification of biomolecules.

Ultra-Small Lysozyme-Protected Gold Nanoclusters as Nanomedicines Inducing Osteogenic Differentiation.

PurposeUltra-small gold nanoclusters (AuNCs), as emerging fluorescent nanomaterials with excellent biocompatibility, have been widely investigated for in vivo biomedical applications. However, their effects in guiding osteogenic differentiation have not been investigated, which are important for osteoporosis therapy and bone regeneration. Herein, for the first time, lysozyme-protected AuNCs (Lys-AuNCs) are used to stimulate osteogenic differentiation, which have the potential for the treatment of bone disease.MethodsProliferation of MC3T3E-1 is important for osteogenic differentiation. First, the proliferation rate of MC3T3E-1 was studied by Cell Counting Kit-8 (CCK8) assays. Signaling pathways of PI3K/Akt play central roles in controlling proliferation throughout the body. The expression of PI3K/Akt was investigated in the presence of lysozyme, and lysozyme-protected AuNCs (Lys-AuNCs) by Western blot (WB) and intracellular cell imaging to evacuate the osteogenic differentiation mechanisms. Moreover, the formation of osteoclasts (OC) plays a negative role in the differentiation of osteoblasts. Nuclear factor κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) signaling pathways are used to understand the negative influence of the osteogenic differentiation by the investigation of Raw 264.7 cell line. Raw 264.7 (murine macrophage-like) cells and NIH/3T3 (mouse fibroblast) cells were treated with tyloxapol, and the cell viability was assessed. Raw 264.7 cells have also been used for in vitro studies, on understanding the osteoclast formation and function. The induced osteoclasts were identified by TRAP confocal fluorescence imaging. These key factors in osteoclast formation, such as (NFATc-1, c-Fos, V-ATPase-2 and CTSK), were investigated by Western blot.ResultsBased on the above investigation, Lys-AuNCs were found to promote osteogenic differentiation and decrease osteoclast activity. It is noteworthy that the lysozyme (protected template), AuNPs, or the mixture of Lysozyme and AuNPs have negligible effects on osteoblastic differentiation compared to Lys-AuNCs.ConclusionThis study opens up a novel avenue to develop a new gold nanomaterial for promoting osteogenic differentiation. The possibility of using AuNCs as nanomedicines for the treatment of osteoporosis can be expected.

The Effect of Dimethyl Sulfoxide on the Lysozyme Unfolding Kinetics, Thermodynamics, and Mechanism.

The thermal stability of proteins in the presence of organic solvents and the search for ways to increase this stability are important topics in industrial biocatalysis and protein engineering. The denaturation of hen egg-white lysozyme in mixtures of water with dimethyl sulfoxide (DMSO) with a broad range of compositions was studied using a combination of differential scanning calorimetry (DSC), circular dichroism (CD), and spectrofluorimetry techniques. In this study, for the first time, the kinetics of unfolding of lysozyme in DMSO–water mixtures was characterized. In the presence of DMSO, a sharp decrease in near-UV CD and an increase in the fluorescence signal were observed at lower temperatures than the DSC denaturation peak. It was found that differences in the temperatures of the CD and DSC signal changes increase as the content of DMSO increases. Changes in CD and fluorescence are triggered by a break of the tertiary contacts, leading to an intermediate state, while the DSC peak corresponds to a subsequent complete loss of the native structure. In this way, the commonly used two-state model was proven to be unsuitable to describe the unfolding of lysozyme in the presence of DMSO. In kinetic studies, it was found that even high concentrations of DMSO do not drastically change the activation energy of the initial stage of unfolding associated with a disruption of the tertiary structure, while the enthalpy of denaturation shows a significant dependence on DMSO content. This observation suggests that the structure of the transition state upon unfolding remains similar to the structure of the native state.

Mixed Polymer Brushes for the Selective Capture and Release of Proteins

Selective protein adsorption is a key challenge for the development of biosensors, separation technologies, and smart materials for medicine and biotechnologies. In this work, a strategy was developed for selective protein adsorption, based on the use of mixed polymer brushes composed of poly(ethylene oxide) (PEO), a protein-repellent polymer, and poly(acrylic acid) (PAA), a weak polyacid whose conformation changes according to the pH and ionic strength of the surrounding medium. A mixture of lysozyme (Lyz), human serum albumin (HSA), and human fibrinogen (Fb) was used to demonstrate the success of this strategy. Polymer brush formation and protein adsorption were monitored by quartz crystal microbalance, whereas protein identification after adsorption from the mixture was performed by time-of-flight secondary ion mass spectrometry (ToF-SIMS) with principal component analysis and gel electrophoresis with silver staining. For the ToF-SIMS measurements, adsorption was first performed from single-protein solutions in order to identify characteristic peaks of each protein. Next, adsorption was performed from the mixture of the three proteins. Proteins were also desorbed from the brushes and analyzed by gel electrophoresis with silver staining for further identification. Selective adsorption of Lyz from a mixture of Lyz/HSA/Fb was successfully achieved at pH 9.0 and ionic strength of 10-3 M, while Lyz and HSA, but not Fb, were adsorbed at ionic strength 10-2 M and pH 9.0. The results demonstrate that by controlling the ionic strength, selective adsorption can be achieved from protein mixtures on PEO/PAA mixed brushes, predominantly because of the resulting control on electrostatic interactions. In well-chosen conditions, the selectively adsorbed proteins can also be fully recovered from the brushes by a simple ionic strength stimulus. The developed systems will find applications as responsive biointerfaces in the fields of separation technologies, biosensing, drug delivery, and nanomedicine.

Direct Analysis of Proteins from Solutions with High Salt Concentration Using Laser Electrospray Mass Spectrometry.

The detection of lysozyme, or a mixture of lysozyme, cytochrome c, and myoglobin, from solutions with varying salt concentrations (0.1 to 250mM NaCl) is compared using laser electrospray mass spectrometry (LEMS) and electrospray ionization-mass spectrometry (ESI-MS). Protonated protein peaks were observed up to a concentration of 250mM NaCl in the case of LEMS. In the case of ESI-MS, a protein solution with salt concentration > 0.5mM resulted in predominantly salt-adducted features, with suppression of the protonated protein ions. The constituents in the mixture of proteins were assignable up to 250mM NaCl for LEMS and were not assignable above a NaCl concentration of 0.5mM for ESI. The average sodium adducts (<n>) bound to the 7+ charge state of lysozyme for LEMS measurements from salt concentrations of 2.5, 25, 50, and 100mM NaCl are 1.71, 5.23, 5.26, and 5.11, respectively. The conventional electrospray measurements for lysozyme solution containing salt concentrations of 0.1, 1, 2, and 5mM NaCl resulted in <n> of 2.65, 6.44, 7.57, and 8.48, respectively. LEMS displays an approximately two orders of magnitude higher salt tolerance in comparison with conventional ESI-MS. The non-equilibrium partitioning of proteins on the surface of the charged droplets is proposed as the mechanism for the high salt tolerance phenomena observed in the LEMS measurements. Graphical Abstract ᅟ.

Simulation of mono‐PEGylated lysozyme separation in heparin affinity chromatography using a general rate model

AbstractBACKGROUNDThe bioavailability of therapeutic proteins is improved through PEGylation. This chemical modification involves the production of isomers with different numbers and sites of attached PEG chains, which are difficult to separate efficiently. Their purification with chromatography requires an understanding of the operation and the evaluation of different operational conditions. The General Rate Model (GRM) was applied for modelling the linear salt gradient elution of mono‐PEGylated and native lysozyme in Heparin Affinity Chromatography (HAC) considering mass balance equations for proteins in the bulk‐fluid phase, in the particle phase and the kinetic adsorption.RESULTSThe model was able to simulate the individual proteins and the separation of these in a PEGylation reaction using as proof‐of‐concept a mono‐PEGylated and native lysozyme mixture under changes of operational parameters such as the gradient length (5, 13, 25 column volumes) and flow (0.8 and 1.2 mL min‐1) with a relative error in retention times of less than 6% and correlation coefficients greater than 0.78.CONCLUSIONSimulation of the elution curves of PEGylated lysozyme in HAC was performed in this work and the diverse information generated by the model is explained through the physicochemical protein properties. This simulation represents a tool for optimization, prediction and future scale‐up of PEGylated proteins purification, which would reduce the investment in time and resources to test several operating conditions. © 2017 Society of Chemical Industry

Effects of Seeding on Lysozyme Amyloid Fibrillation in the Presence of Epigallocatechin and Polyethylene Glycol.

Preformed amyloid fibrils can act as seeds for accelerating protein fibrillation. In the present study, we examined the effects of preformed seeds on lysozyme amyloid fibrillation in the presence of two distinct inhibitors - epigallocatechin (EGC) and polyethylene glycol 2000 (PEG). The results demonstrated that the effects of fibrillar seeds on the acceleration of lysozyme fibrillation depended on the aggregation pathway directed by an inhibitor. EGC inhibited lysozyme fibrillation and modified the peptide chains with quinone moieties in a concentration-dependent manner. The resulting aggregates showed amorphous off-pathway morphology. Preformed fibril seeds did not promote lysozyme fibrillation in the presence of EGC. PEG also inhibited lysozyme fibrillation, and the resulting aggregates showed on-pathway protofibrillar morphology. In contrast, the addition of fibril seeds into the mixture of lysozyme and PEG significantly stimulated fibril growth. Assays of cell viability showed that both EGC and PEG inhibited the formation of cytotoxic species. In accordance with thioflavine T data, the seeds failed to alter the cell-damaging potency of the EGC-directed off-pathway aggregates, but increased the cytotoxicity of the PEG-directed on-pathway fibrils. We suggest that the pattern of interaction between lysozyme and an inhibitor determines the pathway of aggregation and therefore the effects of seeding on amyloid formation. EGC covalently modified lysozyme chains with quinones, directing the aggregation to proceed through an off-pathway, whereas PEG affected the protein in a noncovalent manner, and fibril growth could be stimulated under seeding through an on-pathway.

Synergistic Antilisterial Effects of Mixtures of Lysozyme and Organic Acids

We investigated the synergistic effects of lysozyme combined with organic acids to inhibit the growth of Listeria monocytogenes . The antilisterial effects of the combination of lysozyme and acetic acid, citric acid, lactic acid, malic acid, or succinic acid were evaluated using the checkerboard method and time-kill assay. The MIC was 25,000 mg/liter for lysozyme, 625 mg/liter for acetic acid, and 1,250 mg/liter for the other acids. The MBC was 10,000 mg/liter for all of the tested organic acids. The combination of lysozyme and each organic acid showed synergistic effects via the checkerboard method; however, the time-kill assay showed synergistic effects for only three combinations of 1,250 mg/liter lysozyme with succinic acid (312 and 625 mg/liter) or malic acid (625 mg/liter). The results of this study indicate that the combination of lysozyme and malic acid or succinic acid can be effectively used as a food preservative to control L. monocytogenes .

Crystallization of lysozyme from lysozyme – ovalbumin mixtures: Separation potential and crystal growth kinetics

Lysozyme was successfully separated from mixtures of lysozyme and ovalbumin by crystallization. The purity of the lysozyme product is more than 98%, the remaining activity is greater than 97%, and the yields of the crystal products were greater than 80%. The experimental conditions used were varied to study the effect of the operating parameters on the growth kinetics of lysozyme crystal and the separation ability of the process. The growth rates of lysozyme are second order with respect to the relative supersaturation. Therefore the growth kinetics of the crystallization process is controlled by the surface integration mechanism. The calculated growth rate constants were 5.4×10−6cm/h and 2.5×10−6cm/h for the crystallization process at 20°C and 10°C, respectively. There is no significant effect of the ovalbumin impurity up to the concentration of 67.5% ovalbumin (based on total protein) on the growth kinetics of lysozyme. Changing the NaCl concentration from 4% to 3% had no effect on the growth kinetics of lysozyme, although this does change the solubility and therefore the yield. The calculated activation energy was 53.08kJ/mol which supports the hypothesis that the crystallization process is controlled by the surface integration mechanism.

Evaluation of Lysis Methods for the Extraction of Bacterial DNA for Analysis of the Vaginal Microbiota.

BackgroundRecent studies on the vaginal microbiota have employed molecular techniques such as 16S rRNA gene sequencing to describe the bacterial community as a whole. These techniques require the lysis of bacterial cells to release DNA before purification and PCR amplification of the 16S rRNA gene. Currently, methods for the lysis of bacterial cells are not standardised and there is potential for introducing bias into the results if some bacterial species are lysed less efficiently than others. This study aimed to compare the results of vaginal microbiota profiling using four different pretreatment methods for the lysis of bacterial samples (30 min of lysis with lysozyme, 16 hours of lysis with lysozyme, 60 min of lysis with a mixture of lysozyme, mutanolysin and lysostaphin and 30 min of lysis with lysozyme followed by bead beating) prior to chemical and enzyme-based DNA extraction with a commercial kit.ResultsAfter extraction, DNA yield did not significantly differ between methods with the exception of lysis with lysozyme combined with bead beating which produced significantly lower yields when compared to lysis with the enzyme cocktail or 30 min lysis with lysozyme only. However, this did not result in a statistically significant difference in the observed alpha diversity of samples. The beta diversity (Bray-Curtis dissimilarity) between different lysis methods was statistically significantly different, but this difference was small compared to differences between samples, and did not affect the grouping of samples with similar vaginal bacterial community structure by hierarchical clustering.ConclusionsAn understanding of how laboratory methods affect the results of microbiota studies is vital in order to accurately interpret the results and make valid comparisons between studies. Our results indicate that the choice of lysis method does not prevent the detection of effects relating to the type of vaginal bacterial community one of the main outcome measures of epidemiological studies. However, we recommend that the same method is used on all samples within a particular study.

Rapid Assessment of Resistance to Antibiotic Inhibitors of Protein Synthesis in the Gram-Positive Pathogens, Enterococcus faecalis and Streptococcus pneumoniae, Based on Evaluation of the Lytic Response.

A novel assay for rapid determination of resistance to antibiotic inhibitors of protein synthesis was developed for the gram-positive pathogens, Enterococcus faecalis and Streptococcus pneumoniae. To this purpose, a lytic response was obtained by a brief incubation with lysozyme or a mixture of lysozyme, Triton X-100, and EDTA for E. faecalis (n = 82) and S. pneumoniae (n = 51), respectively. Lysis was quantified by visualizing the released nucleoids. Antibiotic-susceptible bacteria treated with Clinical and Laboratory Standards Institute (CLSI) breakpoint doses of erythromycin, azithromycin, or doxycycline that inhibited protein synthesis demonstrated a large reduction of lysed cells with respect to the control, that is, without antibiotics. However, cell lysis prevention was much lower in nonsusceptible strains, with unsuccessful inhibition of protein synthesis. ROC analysis showed that a reduction value of ≥35.6% and ≥40.4% discriminates susceptible and nonsusceptible strains for erythromycin and for doxycycline, respectively, in E. faecalis, whereas ≥20.0% is adequate for both macrolides and doxycycline in S. pneumoniae. Resistant stains were identified in 90-120 min with sensitivity and specificity between 91.7% and 100%. This is a proof of concept that evaluation of the lytic response may be a rapid and efficient test for determination of resistance to antibiotic inhibitors of protein synthesis.

Titanium Surface Priming with Phase-Transited Lysozyme to Establish a Silver Nanoparticle-Loaded Chitosan/Hyaluronic Acid Antibacterial Multilayer via Layer-by-Layer Self-Assembly.

ObjectivesThe formation of biofilm around implants, which is induced by immediate bacterial colonization after installation, is the primary cause of post-operation infection. Initial surface modification is usually required to incorporate antibacterial agents on titanium (Ti) surfaces to inhibit biofilm formation. However, simple and effective priming methods are still lacking for the development of an initial functional layer as a base for subsequent coatings on titanium surfaces. The purpose of our work was to establish a novel initial layer on Ti surfaces using phase-transited lysozyme (PTL), on which multilayer coatings can incorporate silver nanoparticles (AgNP) using chitosan (CS) and hyaluronic acid (HA) via a layer-by-layer (LbL) self-assembly technique.MethodsIn this study, the surfaces of Ti substrates were primed by dipping into a mixture of lysozyme and tris(2-carboxyethyl)phosphine (TCEP) to obtain PTL-functionalized Ti substrates. The subsequent alternating coatings of HA and chitosan loaded with AgNP onto the precursor layer of PTL were carried out via LbL self-assembly to construct multilayer coatings on Ti substrates.ResultsThe results of SEM and XPS indicated that the necklace-like PTL and self-assembled multilayer were successfully immobilized on the Ti substrates. The multilayer coatings loaded with AgNP can kill planktonic and adherent bacteria to 100% during the first 4 days. The antibacterial efficacy of the samples against planktonic and adherent bacteria achieved 65%-90% after 14 days. The sustained release of Ag over 14 days can prevent bacterial invasion until mucosa healing. Although the AgNP-containing structure showed some cytotoxicity, the toxicity can be reduced by controlling the Ag release rate and concentration.ConclusionsThe PTL priming method provides a promising strategy for fabricating long-term antibacterial multilayer coatings on titanium surfaces via the LbL self-assembly technique, which is effective in preventing implant-associated infections in the early stage.

A method developed to fractionate intact proteins based on capillary electrophoresis.

Reduction in the sample complexity enables more thorough intact protein analysis using MS-based proteomics. A capillary electrophoresis method, namely the velocity gap mode of capillary electrophoresis (VGCE), is proposed to separate protein mixtures with high resolution. Although the separation mechanism of VGCE is also based on the difference of the mass-to-charge ratios of the proteins, it fractionates the sample zone into small pieces of subunits. In this way, the resolution can be dramatically improved due to less longitudinal dispersion of the sample. The effect of the new approach is evaluated by separation of three groups of reference protein mixtures, i.e. a mixture of lysozyme and BSA; a mixture of lysozyme, β-lactoglobulin, and ribonuclease A; and a mixture of cytochrome C, lysozyme, BSA, β-lactoglobulin, ribonuclease A, conalbumin, carbonic anhydrase, and hemoglobin. The results indicate that the new approach shows great potential to couple with MS for top-down analysis of complex mixtures.

Improvement of wine aromatic quality using mixtures of lysozyme and dimethyl dicarbonate, with low SO2 concentration

The use of sulphur dioxide (SO2) in the treatment of foodstuffs presents some problems as it could lead to pseudo-allergies in some people. The aim of this research work was to study the addition of different preservative mixtures and their influence on the concentration of volatile compounds and sensorial quality in wine. To do so, vinifications were carried out using Garnacha must to which lysozyme, dimethyl dicarbonate (DMDC) and mixtures of these with SO2 were added at different doses (25 and 50 mg l−1). The results were compared with a control sample to which only SO2 had been added (50 mg l−1). In general, mixtures of SO2 with lysozyme and DMDC favoured the formation of volatile compounds in the wines. Wines obtained from the mixtures of lysozyme and DMDC with 25 mg l−1 of SO2 had better sensorial quality than the wines obtained with 50 mg l−1 as the only preservative used.

Separation of protein mixtures by an integrated electro-ultrafiltration–electrodialysis process

An integrated process for improving selectivity and permeate flux in ultrafiltration based protein fractionation was developed by combining electrodialysis (ED) with electro-ultrafiltration (EUF). The performance of such a process was first investigated using individual proteins (bovine serum albumin and lysozyme) and then with a mixture of the two. The experimental results showed that as with EUF on its own, the build-up of lysozyme concentration polarization near the cation exchange membrane in the permeate compartment significantly affected ion migration and led to change in pH and conductivity of the feed solution during an EUF–ED process with lysozyme, or bovine serum albumin and lysozyme mixture. As compared with EUF, EUF–ED of protein mixture resulted in a 20% increase in permeate flux with the lysozyme transmission remaining the same. The demineralization that occurred during EUF–ED made this process suitable for protein separation from the feed solutions with high conductivity.

A microporous walled micro-capillary film module for cation-exchange protein chromatography

Opportunities exist in preparative chromatography for alternative chromatography media that possess high binding capacity and throughput, but are also economically feasible for single use disposability and avoid column packing. An ion-exchange functionalised, microporous walled micro-capillary film (MMCF), has been developed as a module for cation-exchange separation of proteins. A MMCF module has been operated on a standard AKTA chromatography system at pressures up to 1.5MPa and superficial flow velocities up to 54,000cmh−1. The dynamic binding capacity of the MMCF module at 10% breakthrough was 13.8mglysozyme/ml adsorbent volume, which is comparable to the capacity of current commercial adsorbents. Frontal analysis studies using a mixture of lysozyme and bovine serum albumin (BSA) have shown that lysozyme can be isolated free of BSA to the limit of detection of the SDS gel assay used. 98.8% of the total sample eluted was the target protein lysozyme with only 1.2% BSA impurity. MMCF may thus be a viable chromatographic medium for preparative protein chromatography.

Do low surfactants concentrations change lysozyme colloid properties?

Multi-component aqueous/octane systems, which contain low-molecular-weight ionic surfactants (sodium dodecyl sulfate (SDS) or dodecyltrimethylammonium bromide (DTAB)) and lysozyme, were investigated using the scintillation phase method and the pendant drop technique. Experiments were carried out in the solutions of high ionic strength. Interfacial tension measurements reveal the appearance of local extremums for the SDS–lysozyme mixture that were attributed to changes in the nature of the SDS–lysozyme complex, which was confirmed by a radiochemical approach (determination of the distribution ratios and values of the surface excess of each component on the background of the other component). The formation of lysozyme–ionic surfactant complexes was observed for both cationic and anionic surfactants. Low surfactant concentrations of both SDS and DTAB result in the same changes of the colloidal behavior of protein. We proposed two possible mechanisms of interaction of lysozyme with cationic and anionic surfactants at low concentrations. The interactive process associated with the formation of lysozyme–ionic surfactant complexes by means of protein binding sites following by co-adsorption with SDS and by step-by-step hydrophilization and displacement from the adsorption layer by DTAB. The increase in the detergent concentration results in the usual substitution of protein from the adsorption layer that was observed by interfacial tension measurements and the radiochemical method.

Application of active zein-based films with controlled release properties to control Listeria monocytogenes growth and lipid oxidation in fresh Kashar cheese

Abstract The antimicrobial and antioxidant potential of zein and zein–wax composite films having different release profiles for lysozyme and mixture of lysozyme, catechin and gallic acid were tested on cold-stored fresh Kashar cheese inoculated with Listeria monocytogenes ATCC 7644. All lysozyme containing films prevented the increase of L. monocytogenes counts in Kashar cheese for 8 weeks at 4 °C, but it was only the zein–wax composite films with sustained lysozyme-release rates which caused a significant reduction (− 0.4 decimals) in initial microbial load of inoculated cheese samples. The mixture of catechin and gallic acid improved the in vitro antimicrobial effect of films against L. monocytogenes, but showed no considerable antimicrobial effect in cheese. However, the films containing catechin and gallic acid were effective in preventing oxidative changes in cheese. This study showed the possibility of increasing safety and quality of fresh cheeses by use of active packaging employing natural antimicrobial compounds and controlled release technology. Industrial relevance There has been a growing interest to develop the functional properties of packaging materials and to obtain packed foods with better shelf-life and quality. Active packaging incorporating antimicrobials is one of the most promising areas since application of this method can improve safety of foods by inhibiting pathogenic bacteria or controlling spoilage flora by using minimum amounts of active compounds. Recently, health concerns of consumers and environmental problems related to plastics increased the popularity of using natural antimicrobial substances in edible films and coatings. This work employs a GRAS status natural antimicrobial lysozyme and zein a major by-product of rapidly growing oil and bioethanol industries which can form self-standing films, coatings or casings. The results of this study showed the possibility of using flexible antimicrobial and antioxidant films with controlled release properties in diary industry.