Karsten Bruun Qvist

Emulsifying Properties of Milk Proteins Cross-linked with Microbial Transglutaminase

Oil-in-water emulsions containing 20% oil were prepared with n-tetradecane and sodium caseinate or β-lactoglobulin in the aqueous phase. The proteins had been cross-linked with microbial transglutaminase prior to emulsification or were cross-linked in the emulsions after emulsification. Extensive cross-linking reduced the stability of the emulsions to coalescence or strong flocculation, as determined by droplet size measurements, whereas limited cross-linking improved coalescence stability. The creaming stability of milk protein stabilized emulsions was improved even with relatively extensive cross-linking and this was ascribed to changes in the adsorbed layer or increased viscosity of the continuous phase. Cross-linking greatly reduced Ostwald ripening or aggregation of the milk protein stabilized emulsions in the presence of ethanol, probably due to a reduction of the diffusion of oil from the emulsion droplets because of increased viscosity/elasticity of the adsorbed layer or due to hindering of the aggregation of protein, that in the non-cross-linked state probably results in a collapse of the surface protein layer.

Relation between sensory texture analysis and rheological properties of stirred yogurt

The sensory and rheological characteristics of stirred yogurts varying in fermentation temperature, heat treatment of milk, dry matter content and composition of bacterial cultures were investigated. Two independent sensory properties, non-oral viscosity and oral viscosity, were evaluated. To reflect these properties precisely it was necessary to perform rheological measurements at both small and large deformations. Non-oral and oral viscosities were successfully modelled from a set of rheological variables by partial least squares regression, resulting in a three component model explaining 83·8% of the variation in non-oral viscosity and a two component model explaining 82·0% of the variation in oral viscosity. Non-oral viscosity was highly correlated with the complex modulus ( G *) from the dynamic oscillatory measurements ( r =0·823) and the viscosity obtained from a Brookfield viscometer (with Helipath stand) operating at 5 rev./min ( r =0·862). Similarly, there were relationships between oral viscosity and the hysteresis loop area from the viscometry test ( r =0·867) and between oral viscosity and the viscosity obtained from the Posthumus funnel ( r =0·834).

Influence of pH on surface properties of aqueous egg albumen solutions in relation to foaming behaviour

The surface properties of aqueous egg albumen protein solutions (0.1 g litre−1) were studied at pH values of 4.8, 7.0, 9.2 and 10.7 and related to foaming behaviour such as bubble size distribution, overrun and drainage. By measurements far from equilibrium of dynamic steady state surface dilation using the overflowing cylinder technique, egg albumen showed ability to slow down surface expansion and to lower the dynamic surface tension. The pH-effect was small, but at pH 4.8 the film length, at which a motionless surface was created, was longer than at higher pH indicating a somewhat more rigid surface at low pH. Near equilibrium sinusoidal surface area deformation resulted in relatively high moduli of egg albumen, with a significant effect of pH. The surface modulus E showed at pH 4.8 an increase in the course of time, but at higher pH it was constant. Large deformation of egg albumen surface was not destructive, and for all pH values the surface behaved viscoelastic, with highest loss modulus E″ and tan θ values at pH 4.8. Surface deformation frequency sweeps revealed the relaxation processes to be relatively slow at pH 4.8 and faster at pH 7.0–10.7. Foamability measured as overrun of foam as a result of shaking and stirring was highest at pH 4.8 and lowest at pH 10.7. Foam stability against drainage was best at pH 7.0 after 30 min, but at a long-term scale foam at pH 4.8 was most resistant to drainage. Foam samples were subjected to microscopy and image analysis. The smallest bubbles were found at pH 4.8 (mean diameter 142 µm) and the largest at pH 7.0 (mean diameter 328 µm). In conclusion, the foaming behaviour of an aqueous egg albumen solution at pH 4.8 can be related to dynamic surface properties as follows: the more rigid behaviour of the surface at this pH favours a small bubble size and slow drainage of liquid from the foam. The high overrun at this pH can be explained by a lower dynamic surface tension, but also here film stability during foam making can be promoted by a more rigid liquid surface.

Molecular self-assembly of partially hydrolysed α-lactalbumin resulting in strong gels with a novel microstructure

Gelation of alpha-lactalbumin (alpha-la) incubated with a protease from Bacillus licheniformis (BLP) at 50 degrees C for 4 h was monitored using small oscillatory shear and the large deformation properties of final gels were characterized by uniaxial compression. Transmission electron microscopy was used to visualize the microstructure. Gels made from alpha-la (10 g/l) using BLP were almost transparent, although somewhat whitish, and they were more than 20 times stiffer (measured as complex modulus) than equivalent gels made from beta-lactoglobulin (beta-lg) at the same concentration. The microstructure of the gels consisted of non-branching, apparently hollow strands with a uniform diameter close to 20 nm, similar in overall structure to microtubules. Adding Ca2+ in amounts of 50 or 100 mM changed the spatial distribution of the strands and resulted in a reduction in the failure stress recorded in uniaxial compression. Apart from affecting the microstructure, Ca2+ was shown to be essential for the formation of the gels. It is proposed. that the mechanism behind the self-assembly of the partially hydrolysed alpha-la into long tubes is a spatially restricted creation of ionic bonds between Ca2+ and carboxyl acid groups on peptide fragments resulting from the action of BLP on alpha-la. Proteolysis of alpha-la with BLP in the presence of Ca2+ thus results in formation of a strong gel with a microstructure not previously observed in food protein systems.

Effect of limited hydrolysis on the interfacial rheology and foaming properties of β-lactoglobulin A

Hydrolysis of beta-lactoglobulin (beta-Lg), genetic variant A, using a serine protease specific for glutamic and aspartic acid residues from Bacillus licheniformis (BLP), resulted in improved foam overrun and foam stability. Limited hydrolysis (19-26% hydrolysed beta-Lg) led to a more rapid increase in the viscoelastic properties of air/water interfacial films and a concomitant increase in foam overrun compared with intact beta-Lg, presumably due to increased exposure of hydrophobic areas. The increased exposure did not, however, cause formation of an interfacial layer with increased viscoelastic properties. More extended hydrolysis (86% hydrolysed beta-Lg) resulted in a higher initial overrun than the unhydrolysed sample and the best foam stability. The interfacial elasticity and viscosity, though, was the lowest observed. Thus, high maximum values of these interfacial properties are not necessary prerequisites for formation of a voluminous and stable foam.

Enzyme-induced gelation of whey proteins: Effect of protein denaturation

Abstract The enzyme-induced gelation properties of unheated and denatured whey proteins (WP) were investigated. Solutions (9%, w v ) of whey protein isolate (WPI) were denatured by heating at 80 °C for 2–30 min, and gelation was induced by incubation with a Bacillus licheniformis protease (40 °C, pH 7.0). The gelation characteristics, as well as gel strength and microstructure, were examined. The heat treatments resulted in irreversible denaturation of up to 98% of the WP. Bacillus licheniformis protease (BLP) was able to induce gelation of both unheated and pre-heated WP, but the gelation process was strongly dependent on the extent of denaturation (%D). Higher %D resulted in earlier gelation and a faster increase in gel firmness. The relation between %D and the rate of gel firming was almost linear. The strength of the enzyme-induced gels increased continuously during 30 h of incubation. After 9 h, the gel with 98%D was 3.2 times stronger than that with 47%D, and 18 times stronger than the gel from unheated WPI (≈8%D). The latter gel had a particulate microstructure, whereas the gel from the highly denatured WPI (98%D) had a fine-stranded structure. This enzyme-induced gelation of partly denatured WP may be of interest for food applications.

Hydrolysis of β-lactoglobulin by four different proteinases monitored by capillary electrophoresis and high performance liquid chromatography

Abstract The hydrolysis of β-lactoglobulin (β-lg) by four enzymes [porcine trypsin (PT), Fusarium oxysporum trypsin (FOT), Bacillus licheniformis proteinase (BLP) and Bacillus subtilis proteinase (Neutrase®)] was characterised by size exclusion—and reversed phase high performance liquid chromatography (SE-HPLC and RP-HPLC) and by capillary electrophoresis (CE). After 24 h of hydrolysis, all β-lg had been degraded by PT and BLP, but a large part of the protein was still intact after hydrolysis by FOT or Neutrase®. The main fraction of peptides was found to have MWs ranging from 1.0 to 3.0kDa. The hydrolysis catalysed by each enzyme resulted in different peptide profiles by RP-HPLC and CE. Hydrolysates produced by PT or FOT are resolved into 18 peaks, while BLP hydrolysates were resolved into 25 peaks, corresponding well to the numbers of possible cleavage sites in β-lg. Neutrase®, with a broad specificity, produced the largest number of peptides. Seven peptides from PT hydrolysis, eight from FOT hydrolysis and one from BLP hydrolysis were identified by mass spectrometry (MS) and Edman degradation.

Characteristics of oil water emulsions stabilised by an industrial α-lactalbumin concentrate, cross-linked before and after emulsification, by a microbial transglutaminase

Abstract Industrial α-lactalbumin concentrate, cross-linked with a microbial transglutaminase, showed lower dilational surface viscosity at a planer oil–water interface than a non-cross-linked α-lactalbumin concentrate. Properties of emulsions containing cross-linked α-lactalbumin were influenced by the sequence of cross-linking and emulsification. Emulsions stabilised by α-lactalbumin concentrate (even without crosslinking) were generally unstable. While cross-linking before emulsification decreased the stability further, the emulsion stability was improved when cross-linking was carried out after emulsification. Results from the sodium dodecyl sulphate (SDS) gel electrophoresis of adsorbed protein suggested that, irrespective of the sequence of cross-linking and emulsification, the adsorbed protein was polymerised too extensively to be resolvable on the gel matrix. Results from the reverse-phase HPLC suggested that the amount of adsorbed protein in emulsions containing protein cross-linking before emulsification was lower than that containing cross-linking after emulsification.

Camel and Bovine Chymosin: The Relationship between Their Structures and Cheese-Making Properties.

Analysis of the crystal structures of the two milk-clotting enzymes bovine and camel chymosin has revealed that the better milk-clotting activity towards bovine milk of camel chymosin compared with bovine chymosin is related to variations in their surface charges and their substrate-binding clefts.

Bovine Chymosin: A Computational Study of Recognition and Binding of Bovine κ-Casein

Bovine chymosin is an aspartic protease that selectively cleaves the milk protein kappa-casein. The enzyme is widely used to promote milk clotting in cheese manufacturing. We have developed models of residues 97-112 of bovine kappa-casein complexed with bovine chymosin, using ligand docking, conformational search algorithms, and molecular dynamics simulations. In agreement with limited experimental evidence, the model suggests that the substrate binds in an extended conformation with charged residues on either side of the scissile bond playing an important role in stabilizing the binding pose. Lys111 and Lys112 are observed to bind to the N-terminal domain of chymosin displacing a conserved water molecule. A cluster of histidine and proline residues (His98-Pro99-His100-Pro101-His102) in kappa-casein binds to the C-terminal domain of the protein, where a neighboring conserved arginine residue (Arg97) is found to be important for stabilizing the binding pose. The catalytic site (including the catalytic water molecule) is stable in the starting conformation of the previously proposed general acid/base catalytic mechanism for 18 ns of molecular dynamics simulations.