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Whey protein-enriched cheese can be produced by means of a high-temperature treatment of a part of the cheese milk. In this way, the nutritional quality of the resulting cheeses can be increased while resources are conserved. High-performance thin-layer chromatography-immunostaining (HPTLC-IS) using specific β-lactoglobulin (β-LG) antibodies was applied to study the implementation and stability of β-LG in two different sample sets of whey protein-enriched Edam model cheeses, including industrial-scale ones. Two methods were compared for the extraction of the proteins/peptides from the cheese samples. By applying tryptic hydrolysis directly from a suspended cheese sample instead of a supernatant of a centrifuged suspension, a better yield was obtained for the extraction of β-LG. When applying this method, it was found that selected epitopes in the tryptic β-LG peptides remain stable over the ripening period of the cheese. For four of the tryptic β-LG peptides detected by immunostaining, the amino acid sequence was identified using MALDI-TOF-MS/MS. One of the peptides identified was the semi-tryptic peptide VYVEELKPTP. A linear relationship was found between the content of this peptide in cheese and the proportion of high-heated milk in the cheese milk. β-LG enrichment factors of 1.72 (

In order to apply emulsion-based delivery systems to food, they have to be stable in a protein rich environment. This study investigated the stability of solid lipid nanoparticles (SLN) during heat treatment in the presence or absence of β-lactoglobulin (BLG). SLN were stabilized either by Tween 20 (TS) or by the protein itself (BS) and were enriched to a total BLG content of 56 mg/mL. The sizes of both types of SLN were initially in the range of 170 nm. The amount of free protein was determined before and after enrichment with BLG. As revealed by particle size and zeta potential measurements, a protein layer of BLG (hard corona) adsorbed on BS but not on TS. By contrast, a soft corona was formed around both BS and TS. SLN were heat treated in the presence and absence of protein and were characterized regarding size and zeta potential. According to transmission electron microscopy imaging, heating did not affect the shape of TS and BS: TS were platelets, whereas BS exhibited a spherical or platelet like shape. Upon heat treatment, the particle size of TS increased to about 3.5 fold of the initial size (to appr. 600 nm) in the presence and in the absence of excess protein. The cloudy protein layer (soft corona) around TS could thus not prevent coalescence of TS. By contrast, BS did not experience a change in particle size. Hence, by the choice of emulsifier, an encapsulation system that is stable against heat treatment can be obtained.

A nanoparticlulate delivery system was prepared for developing a food grade carrier for the major bioactive constituent in green tea; (─)-epigallocatechingallate (EGCG), in order to protect it against degradation during storage and digestion under simulated gastrointestinal pH conditions. EGCG-loaded solid lipid nanoparticles (EGCG-SLNs) were produced by the hot homogenization method. The lipid matrix used in the production consisted of pure cocoa butter. A combination of sodium stearoyl-2-lactylate (SSL) and mono- and diglycerides (MDG) was applied as a surfactant blend. The nanoparticles loaded with different concentrations of EGCG had an average particle size in the range of 108–122 nm. A maximal encapsulation efficiency of 68.5% was obtained. The produced food grade SLNs successfully protected the encapsulated EGCG along the storage period as well as under the adverse conditions at neutral pH values. The developed system offers good potential for enriching food products with EGCG.

Selected influencing factors in processed cheese making (protein and fat content, fat globule size, and rework addition) affecting the physical changes known as “creaming” were investigated for their effect on this multistage structure formation reaction. The creaming curve (viscosity vs. time) shows four typical stages: an initiation phase, a first exponential stage, a plateau, and a second exponential phase. Increasing the protein content from 10 to 17% (w/w) accelerated the reaction. Light microscopy showed that the fat content (0–20%) affected the shape of the creaming curve as well and it was shown that a fat level of 15–20% is required for the characteristic creaming curve to occur. Moreover, modifications in the initial milkfat globule size (3.7 μm down to 1.1 μm) by means of upstream homogenization (0–250/50 bar) accelerated the exponential phase and modified the shape of the creaming curve, shortening the initiation and plateau phases. The reaction started earlier with decreasing incoming fat globule size, and the slope was steeper. When fat was present in the system, it was not only the content, but the milkfat globule size which dictates the viscosity change and shape of the curve. The addition of rework dramatically affects the structure formation process, rework probably acting as a catalyst accelerating the reaction. However, protein polymerization was found to be constant during the entire course of the reaction suggesting that weaker physical bonds are responsible for the structuring of the matrix.

Edible solid lipid nanoparticles (SLN) stabilized by a mixture of food grade emulsifiers (soy bean lecithin, Tween 20, sucrose stearate) were added to o/w emulsions previously stabilized by anionic SDS, non-ionic Tween 20 or cationic CTAB. The aim of the study was to understand the fate of both SLN and oil droplets in these mixtures focusing on the impact of the surfactant used to stabilize the emulsion. The presence of SLN in emulsions led to increased emulsion stability as reflected by droplet size measurements and accelerated creaming experiments. This could be attributed to an increase in the viscosity of the sample, but also to changed properties of the o/w interface. Zeta potential measurements revealed that the surfactant composition at the o/w interface had changed in SDS and CTAB stabilized but not in Tween 20 stabilized emulsions. SLN remained detectable in the continuous phase of each emulsion system over three weeks of storage but were not detected at the o/w interface of oil droplets. The particle size of the SLN remained unchanged whereas their zeta potential increased in SDS and CTAB stabilized emulsions to a similar magnitude (+/− 68 mV) but opposite signs. The melting temperature and melting enthalpy of SLN decreased in emulsions indicating that part of the lipid matrix was dissolved by oil from the emulsion. Accordingly, a time-dependent transfer of crystalline triglycerides originating from the SLN into the oil phase of separated model emulsion systems was verified.

The lactobacilli associated with a fermented goat milk product from Tajikistan were isolated to characterize their technological properties and antibiotic resistances in order to assess their suitability for development as starter cultures. In this study, twenty three strains were identified by 16S rRNA sequencing as typical dairy-associated lactic acid bacterial strains, i.e. L. plantarum, L. pentosus, L. delbrueckii, L. helveticus and L. paracasei strains. These strains were generally susceptible to most antibiotics tested in this study and this allowed a selection of strains as safe starters. The draft genomes of four representative strains were sequenced and the number of contigs of the four assembled genomes ranged from 51 to 245 and the genome sizes ranged from 1.75 to 3.24 Mbp. These representative strains showed differences in their growth behavior and pH reducing abilities in in vitro studies. The co-inoculation of these Lactobacillus spp. strains together with a yeast Kluyveromyces marxianus MBT-5698, or together with the yeast and an additional Streptococcus thermophilus MBT-2, led to a pH reduction to 3.4 after 48 h. Only in the case of fermentation inoculated with the co-culture, the viscosity of the milk increased noticeably. In contrast, fermentations with single strains did not lead to gelation of the milk and to decrease the pH after 24h. The results of this study provide a comprehensive understanding of the predominant lactobacilli related to Tajikistan fermented milk products.

Casein micelles (CM) loaded with Vitamin D2 (Vit. D2) were spray- and freeze-dried at pilot scale. The powders showed constant Vit. D2 contents over four months of storage. The CM structure was retained after processing and drying. Low fat yoghurt was enriched with the spray-dried powder or with free Vit. D2. An in vitro proteolysis showed that 90% of the Vit. D2 added as an encapsulated product, e.g. dried CM, remained active compared to only 67% when Vit. D2 was supplemented as a free substance. Since all proteins were hydrolysed after in vitro proteolysis, it can be assumed that the Vit. D2 contents determined would be ultimately available in the lumen.

Summary To apply native casein micelles (CM) as nanocarriers for lipophilic substances in non- or low-fat food products, they have to be conditioned before loading. In this study, an experimental set-up for the production and loading of CM was developed. Microfiltration was used to separate CM from skimmed milk. To identify optimal loading conditions temperatures (2, 20, 40 °C), pH values (6.8 and 5.5) and holding times (5, 15, 30, 60 min) were varied. The release of calcium, phosphate and protein from the micellar phase as well as static light scattering and transmission electron microscopy showed that CM were optimally primed at 2 °C and a pH of 5.5 for 5 min. Therefore, loading with β-carotene was performed under those conditions. After the back-extraction of β-carotene, the photometrical analyses revealed total recovery rates of >79% whereby 94% of it was associated with the native CM.

Nono, an important traditional fermented dairy food produced from cow’s milk in Nigeria, was studied for microbial diversity and for starter culture development for industrial production. On the basis of a polyphasic approach, including phenotypic and genotypic methods such as 16S rRNA gene sequencing, repetitive element PCR (rep-PCR) fingerprinting metagenomics, and whole genome sequencing, we identified Lactobacillus (Lb.) helveticus, Limosilactobacillus (L.) fermentum, Lb. delbrueckii, and Streptococcus (S.) thermophilus as predominant bacterial species involved with milk fermentation during traditional nono production in Nigeria, while the predominant yeast species in nono was identified as Saccharomyces cerevisiae. Using metagenomics, Shigella and potential pathogens such as enterobacteria were detected at low levels of abundance. Strains of the predominant lactic acid bacteria (LAB) were selected for starter cultures combination on the basis of their capacities for rapid growth in milk and reduction of pH below 4.5 and their gelling characteristic, which was demonstrated noticeably only by the S. thermophilus strains. Whole genome sequence analysis of selected bacterial strains showed the largest assembled genome size to be 2,169,635 bp in Lb. helveticus 314, while the smallest genome size was 1,785,639 bp in Lb. delbrueckii 328M. Genes encoding bacteriocins were not detected in all the strains, but all the LAB possessed genes potentially involved in diacetyl production and citrate metabolism. These bacteria isolated from nono can thus be used to improve the microbial safety quality of nono in Nigeria, in addition to improving technological parameters such as gelling viscosity, palatability, and product consistency.

The aim of the present study is the investigation of lipid oxidation in protein based oleogels. For this purpose, oleogels are prepared by suspension of freeze dried whey protein isolate aggregates and small amounts of water (

Although the connection between protein oxidation, amyloid aggregation and diseases such as Alzheimer’s is well known there is no information on such effects during preparation of beta-lactoglobulin fibrils. Different morphologies of amyloid aggregates of beta-lactoglobulin were prepared by incubation at pH 2 or pH 3.5 for up to 72 h. After 5 h, amyloid aggregates at pH 2 formed typical fibrils, which consisted of peptides. At pH 3.5, the amyloid aggregates were worm-like and consisted of intact protein. After 72 h, the building blocks at both pH values changed towards smaller peptides. The apparent tyrosine oxidation reached a maximum after 5 h at both pH values, whereas N-formylkynurenine and carbonyls increased continuously during 72 h. In case amyloid structures are used as edible material, the health related effects caused by protein oxidation needs to be considered.

Clostridium tyrobutyricum is the main spoilage agent of late blowing defect (LBD) in Grana Padano and Parmigiano-Reggiano cheeses; LBD is characterized by openings and holes and is sometimes accompanied by cracks and an undesirable flavor. Even a very few spores remaining in the cheese curd may cause LBD; thus, it is essential to eradicate them during milk natural creaming. By this process, most of the bacteria, somatic cells, and spores rise to the top of the milk, together with the fat globules, and are removed with the cream. Previous studies suggested that milk immunoglobulins mediate the interactions between fat globules and bacteria that occur upon creaming but no direct evidence for this has been found. Moreover, other physical chemical interactions could be involved; for example, physical entrapment of spores among globule clusters. To maximize the efficiency of the natural creaming step in removing Cl. tyrobutyricum, it is essential to understand the nature of spore–globule interactions. With this aim, raw milk was contaminated with spores of Cl. tyrobutyricum before going to creaming overnight at 8°C, after which spore and bacteria removal was >90%. The obtained cream was analyzed by light interference contrast and fluorescence microscopy and by transmission electron microscopy (TEM). Results showed that most of the vegetative cells and spores, which were stained with malachite green before addition to milk, adhered tightly to the surface of single fat globules, the membranes of which appeared heterogeneous when stained with the fluorescent dye DilC18(3)-DS. Using the same dye, we observed transient and persistent interactions among globules, with formation of clusters of different sizes and partial coalescence of adhering membranes. Transmission electron microscopy examination of replicates of freeze-fractured cream allowed us to observe tight adhesion of spores to fat globules. Ultrathin sections revealed that this adhesion is mediated by an amorphous, slightly electron-opaque material, sometimes granular in appearance. Bacteria also adhered to different fat globules, linking them together, which suggests that adhesion was strong enough to maintain a stable contact. Although we cannot exclude physical entrapment of bacteria among fat globule clusters, we show for the first time that most of the bacteria are adhered to fat globules by an electron-opaque material whose nature has yet to be determined. Immunoglobulins are certainly the best candidates for adhesion but other compounds may be involved.

Summary Photocentrifugation at 1200 g and 60 °C was used to monitor emulsion stability of spreadable processed cheese (PC) with a low content (0.7 and 1.0 g 100 g−1) of typical emulsifying salts (ES) after different manufacturing conditions. This method of dispersion analysis has been used more frequently for nonfood materials rather than for foodstuffs. The PC contained 36.5 g 100 g−1 dry matter, 40 g 100 g−1 fat in dry matter and was manufactured in a laboratory cooker at 82 or 92 °C and 1000–3000 rpm cutter speed for 5–19 min. Insufficient fat emulsification and a heterogeneous casein matrix of a PC with 0.7% was well illustrated by photocentrifugation. At 1 g 100 g−1 ES and 3000 rpm cutter speed, the effect of manufacturing time on the behaviour of the colloidal dispersion was demonstrated by the different evolutions of the corresponding transmission profiles. Sedimentation velocities were measured at a level of 20% transmission. These results showed that dispersion analysis of spreadable PC by photocentrifugation at 60 °C is a sensitive method and applicable as a new approach for characterisation of emulsion stability resulting from physicochemical changes during manufacture.

Titanium dioxide (TiO2) is one of the most applied nanomaterials and widely used in food and non-food industries as an additive or coating material (E171). It has been shown that E171 contains up to 37% particles which are smaller than 100 nm and that TiO2 nanoparticles (NPs) induce cytotoxicity and inflammation. Using a nuclear factor Kappa-light-chain enhancer of activated B cells (NF-κB) reporter cell line (Caco-2nfkb-RE), Real time polymerase chain reaction (PCR), and inhibition of dynamin and clathrin, it was shown that cellular responses induced by 5 nm and 10 nm TiO2 NPs (nominal size) depends on endocytic processes. As endocytosis is often dependent on the epithelial growth factor receptor (EGFR), further investigations focused on the involvement of EGFR in the uptake of TiO2 NPs: (1) inhibition of EGFR reduced inflammatory markers of the cell (i.e., nuclear factor (NF)-κB activity, mRNA of IL8, CCL20, and CXCL10); and (2) exposure of Caco-2 cells to TiO2 NPs activated the intracellular EGFR cascade beginning with EGFR-mediated extracellular signal-regulated kinases (ERK)1/2, and including transcription factor ELK1. This was followed by the expression of ERK1/2 target genes CCL2 and CXCL3. We concluded that TiO2 NPs enter the cell via EGFR-associated endocytosis, followed by activation of the EGFR/ERK/ELK signaling pathway, which finally induces NF-κB. No changes in inflammatory response are observed in Caco-2 cells exposed to 32 nm and 490 nm TiO2 particles.

Substituting sodium with potassium in emulsifying salts of pasteurized block-type dairy cheese analogues was investigated. Rennet casein and butter were used as protein and fat sources for the manufacture of a product with 51% dry matter and 50% fat in dry matter. Different emulsifying salt blends contained sodium or potassium citrate and sodium polyphosphate. The cheese analogues were tested for their melting properties by oscillation rheometry and by empirical tests for meltability and fat release after 2 days, 4 and 12 weeks of cold storage. A recipe containing about 60% less sodium and a typical standard recipe without potassium showed very similar results during 4 weeks of cold storage.

Differently homogenised and HTST-heated milk (3.5% fat) was foamed at temperatures between 4 and 60 °C. Foaming was achieved by air injection through fritted glass. Initial foam density, drainage and corresponding bubble size were analysed. Transmission electron microscopic (TEM) images completed the study. The studies showed that whole milk was better foamable between 50 and 60 °C than at lower temperatures. This was mainly due to the completely liquid milk fat and the increased protein adsorption at the air–serum interface. The resulting bubbles of these two foams maintained their spherical shape also for 20 min of draining. However, the average bubble diameter and the drainage mass in relation to the initial foam mass increased from about 20 g/100 g after 1 min to about 80 g/100 g after 20 min. It was surprising to learn that milk homogenisation and corresponding fat globule size had only marginal effect on foam formation and stability. TEM images suggested that the air–serum interface consisted mainly of protein monomers and oligomers, while casein micelles were not directly adsorbed. The membrane of the homogenised fat globules was destroyed near the interface and coalesced liquid fat formed a restricted film on the bubble that was obviously of minor importance for the foam properties.

The objective of this study was to investigate the effect of foaming temperature and impact of heat on the foaming properties of skimmed milk. Increasing foaming temperatures from 4 to 60 °C led to a decreased average foam bubble diameter ( d 10 ) from 0.26 to 0.18 mm. Temperature increase also led to increased foam density values (0.14–0.22 g cm −3 ). Overall, the skimmed milk foam that was prepared at 50–60 °C was the most stable, while minimal drainage was detected in the range of 30–40 °C. Pre-heating of skimmed milk was performed by high-temperature-short-time (HTST), high temperature (HT)- or ultra-high-temperature (UHT)-heating. The ( d 10 )-values of the milk foam increased from non-heated to HT-heated milk. Foam density decreased in a continuous fashion from unheated to UHT-heated milk. The composition of non-denatured whey proteins in milk samples and the respective foams differed only marginally. Foam from unheated milk was the most stable. Transmission electron microscope-images revealed differences in foam microstructure in relation to the age of the foam and the heat treatment applied. The air–serum interface in the foam of skimmed milk consists of casein and whey protein monomers to oligomers. Casein micelles do not seem to be an integral part of the air–serum interface in skimmed milk foam.

The influence of low molecular weight emulsifiers (SDS, CTAB, lecithin, mono-diglycerides) on rheological properties and the microstructure of model processed cheese, made using rennet casein, at three different pH values was investigated. Interactions between the low molecular weight emulsifiers and rennet caseins in model processed cheese are consistent with those found in model food emulsions of liquid continuous phase. Compared with the control, the addition of CTAB (cationic) resulted in the hardest and most elastic processed cheese, while the incorporation of SDS (anionic) produced the softest and least elastic cheese. Processed cheese with added lecithin or mono-diglycerides behaved much like the controls, but with an increase in syneresis level. No syneresis was observed with the SDS cheeses. In general, low pH cheeses (pH 5.45) were harder than high pH cheeses (pH 6.05). Rheologically, all the processed cheese samples can be described as [weak gels]. The protein matrix of the high pH processed cheese containing CTAB consisted mainly of protein particles linked into a chain-like form, while those containing lecithin and SDS showed a mixture of individual as well as short chained protein aggregates, and the control had protein aggregates of intermediate size. The added low molecular weight emulsifiers resulted in a finer dispersion of the fat in the protein matrix. Protein-emulsifier charge interactions seem to be the prime determinant of the rheological properties of these model processed cheeses.

The properties of casein micelles in milk concentrates are of interest for the use of ultrafiltered (UF) skim milk concentrates in dairy products, and for the general understanding of colloidal stability and behavior of the casein micelle. The rheological behavior of UF skim milk concentrate with a casein concentration of 19.5% (wt/wt) was investigated at different pH and NaCl concentrations by analyzing flow viscometry and small amplitude oscillatory shear measurements. Viscometric flow curves were fitted to the Carreau-Yasuda model with the aim of determining values for the viscosity at infinite high shear rates and thereby estimate the voluminosity of the casein micelles (nu(casein)) in the UF concentrate. The voluminosity of the casein micelles increased with addition of NaCl and decreased when pH was decreased from 6.5 to 5.5. At pH 5.2, nu(casein) increased because of acid-induced aggregation of the casein micelles. The changes in nu(casein) could be interpreted from transmission electron microscopy of freeze-fractured samples of the UF concentrate and partly from dynamic light scattering measurements. Altered interactions between casein micelles due to different pH and NaCl concentrations are proposed to occur due to collapse of the kappa-casein layer, changed ionic strength, and altered distance between casein micelles.