Effects of various whey protein hydrolysates on the emulsifying and surface properties of hydrolysed lecithin

Oil-in-water emulsions (30 wt% sunflower oil) containing various concentrations of commercial whey protein hydrolysates (0-4 wt%) and hydrolysed lecithin (0.4-1.8 wt%) were prepared by means of a high pressure homogeniser. The degrees of hydrolysis used ranged from 10 to 27%. The individual and interactive effects of these factors on the particle size distribution, emulsion stability, consistency and interfacial tension were investigated using a three-level factorial design according to the principle of response surface methodology. The properties of the emulsions containing both hydrolysed lecithin and whey protein hydrolysate (WPH) were significantly influenced by the degree of hydrolysis of WPH, the protein content and the second-order interaction between both. Addition of WPH, with a 10-20% degree of hydrolysis, improved the stability of lecithin-stabilised emulsions and slightly decreased the average droplet size, compared to those emulsions with only protein or hydrolysed lecithin. However, when extensively hydrolysed WPH (DH=27%) was mixed with hydrolysed lecithin, rapid coalescence and oiling-off of the emulsion droplets resulted, suggesting competition between the surface active components of this WPH and the hydrolysed lecithin. High amounts of such an extensively hydrolysed WPH, together with low lecithin concentrations, were found to be especially detrimental. The different behaviour of partially and extensively hydrolysed WPH in oil-in-water emulsions containing hydrolysed lecithin, was in good agreement with their interfacial activity, as measured by the drop volume method.     

Behaviour of protein-stabilised emulsions under various physiological conditions

Emulsion forms a major part of many processed food formulations. During the past few decades, the physico-chemical properties of oil-in-water emulsions under various food processing conditions have been extensively studied. However, over the recent years, interest has turned to understanding the behaviour of emulsions during consumption, i.e. physiological processing. In general, on ingestion, an emulsion is exposed to a relatively narrow range of physical (e.g. shear and temperature) and biochemical (e.g. dilution, pH, pepsin, pancreatin, mucins and bile salts) environments as it passes through the mouth into the stomach and then the intestines. There is currently limited knowledge of the physico-chemical and structural changes, which an emulsion may undergo when it passes through the physiologically active regime. A better understanding of the gastro-intestinal processing of emulsions would allow manipulation of physico-chemical and interfacial properties to modulate lipid ingestion, improve bioavailability of lipid soluble nutrients and reduce absorption of saturated fats, cholesterol and trans fats.Food emulsions are commonly stabilised by proteins, as they are not only excellent emulsifiers but also provide nutritional benefits to the product. The effects of digestion conditions on interfacial protein structures are complicated because of potential breakdown of these structures by proteolytic enzymes of the gastrointestinal tract. Studies dealing directly with the behaviour of protein-based emulsions under digestion conditions are very limited. This paper provides an overview of the behaviour of oil-in-water emulsions stabilised with globular proteins, namely lactoferrin and β-lactoglobulin. Recent advances in understanding the interactions between interfacial proteins on oil droplets and various physiological materials (e.g. enzymes and bile salts) in in vitro digestion systems are considered. Major emphasis is placed on the recent work carried out in our laboratory at Massey University on the behaviour of milk protein based emulsions (lactoferrin or β-lactoglobulin) during their passage through the gastro-intestinal tract.     

Flocculation and coalescence of droplets in oil-in-water emulsions formed with highly hydrolysed whey proteins as influenced by starch

The effects of added unmodified amylopectin starch, modified amylopectin starch and amylose starch on the formation and properties of emulsions (4 wt.% corn oil) made with an extensively hydrolysed commercial whey protein (WPH) product under a range of conditions were examined. The rate of coalescence was calculated based on the changes in the droplet size of the emulsions during storage at 20 degrees C. The rates of creaming and coalescence in emulsions containing amylopectin starches were enhanced with increasing concentration of the starches during storage for up to 7 days. At a given starch concentration, the rate of coalescence was higher in the emulsions containing modified amylopectin starch than in those containing unmodified amylopectin starch, whereas it was lowest in the emulsions containing amylose starch. All emulsions containing unmodified and modified amylopectin starches showed flocculation of oil droplets by a depletion mechanism. However, flocculation was not observed in the emulsions containing amylose starch. The extent of flocculation was considered to correlate with the rate of coalescence of oil droplets. The different rates of coalescence could be explained on the basis of the strength of the depletion potential, which was dependent on the molecular weight and the radius of gyration of the starches. At high levels of starch addition (>1.5%), the rate of coalescence decreased gradually, apparently because of the high viscosity of the aqueous phase caused by the starch.     

A study of oil-in-water emulsions stabilized by whey proteins

Like many other emulsifiers, whey protein concentrates stabilize oil-in-water emulsions. However, the emulsifying capacity of whey proteins is affected by several factors, e. g., type of homogenizer, degree of homogenization, protein concentration, oil volume fraction, pH and ionic strength of the aqueous phase. For the present study, oilin-water emulsions were made by homogenizing known amounts of whey protein concentrate with a vegetable oil (i. e. grapeseed oil) at different pH. The emulsifying properties of whey proteins are expressed as a function of the particle size and size distribution of oil droplets as measured by light scattering, and of the surface charge density derived from the electrophoretic mobility.The whey protein concentrate was shown to have an isoelectric point at pH 4.4. Near this pH value, the oil-in-water emulsions exhibited poor stability as expected from the low surface coverage.     

Effects of heat on physicochemical properties of whey protein-stabilised emulsions

The effect of heating has been studied for whey protein-stabilised oil-in-water emulsions (25.0% (w/w) soybean oil, 3.0% (w/w) whey protein isolate, pH 7.0). These emulsions were heated between 55 and 95 °C as a function of time and the effect on particle size distribution, adsorbed protein amount, protein conformation and rheological properties was determined. Heating the emulsions as a function of temperature for 25 min resulted in an increase of the mean diameter (d₃₂) and shear viscosity with a maximum at 75 °C. Heating of the emulsions at different temperatures as a function of time in all cases resulted in a curve with a maximum for d₃₂. A maximum increase of d₃₂ was observed after about 45 min at 75 °C and after 6–8 min at 90 °C. Similar trends were observed with viscosity measurements. Confocal scanning laser micrographs showed that after 8 min of heating at 90 °C large, loose aggregates of oil droplets were formed, while after 20 min of heating compact aggregates of two or three emulsion droplets remained. An increase of the adsorbed amount of protein was found with increasing heating temperature. Plateau values were reached after 10 min of heating at 75 °C and after 5 min of heating at 90 °C. Based on these results we concluded that in the whole process of aggregation of whey protein-stabilised emulsions an essential role is played by the non-adsorbed protein fraction, that the kinetics of the aggregation of whey protein-stabilised emulsions follow similar trends as those for heated whey protein solutions and that upon prolonged heating rearrangements take place leading to deaggregation of initially formed large, loose aggregates of emulsion droplets into smaller, more compact ones.     

Physico-chemical characteristics of oil-in-water emulsions based on whey protein-phospholipid mixtures

Emulsions prepared with whey proteins, phospholipids and 10% of vegetable oil were used for a model typifying dressings, coffee whitener and balanced diets. For the present study, two whey proteins (partial heat-denatured whey protein concentrate (WPC) and undenatured whey protein isolate (WPI)) in combination with different phospholipids (hydrolysed and unmodified deoiled lecithin) were chosen to investigate the interactions between proteins, phospholipids and salt (sodium chloride) in such emulsion systems. Oil-in-water (o/w) emulsions (10 wt.% sunflower oil) containing various concentrations of commercial whey proteins (1-2%), phospholipids (0.39-0.78%) and salt (0.5-1.5%) were prepared using a laboratory high pressure homogeniser under various preparation conditions. Each emulsion was characterised by droplet size, creaming rate, flow behaviour and protein load. The dynamic surface activity of the whey proteins and lecithins at the oil-water interface was determined using the drop volume method. The properties of emulsions were significantly influenced by the content of whey protein. Higher protein levels improved the emulsion behaviour (smaller oil droplets and increased stability) independent of the protein or lecithin samples used. An increase of the protein content resulted in a lower tendency for oil droplet aggregation of emulsions with WPC to occur and emulsions tending towards a Newtonian flow behaviour. The emulsification temperature was especially important using the partial heat-denatured WPC in combination with the deoiled lecithin. A higher emulsification temperature (60 degrees C) promoted oil droplet aggregation, as well as an increased emulsion consistency. Emulsions with the WPC were significantly influenced by the NaCl content, as well as the protein-salt ratio. Increasing the NaCl content led to an increase of the droplet size, higher oil droplet aggregation, as well as to a higher creaming rate of the emulsions. An increase of the lecithin content from 0.39 to 0.78% in the emulsion system resulted in a small reduction of the single droplet size. This effect was more pronounced when using the hydrolysed lecithins.     

Oil-in-water emulsions stabilized by whey protein aggregates: Effect of aggregate size,pH of aggregation and emulsion pH

Oil-in-water emulsions (60% oil (w/w)) were prepared using whey protein aggregates as the sole emulsifying agent. The effects of whey protein aggregate size (the diameter between 0.92 and 10.9?µm), the pH of emulsions (4–8.6) and storage time on physical properties, droplet size, and stability of emulsions were investigated. The results indicate that increment of whey protein aggregate size caused an increase in the firmness, droplet size, and viscosity of emulsions, and also a decrease in the emulsion creaming. The emulsion viscosity, firmness, and droplet size were reduced by increasing the emulsion pH; however, the creaming process was accelerated. Viscosity, creaming, and droplet size of emulsions were increased slightly during 21 days storage at 40°C.     

Microstructure of beta-lactoglobulin-stabilized emulsions containing non-ionic surfactant and excess free protein: influence of heating

The influence of the non-ionic surfactant Tween 20 on the microstructure of beta-lactoglobulin-stabilized emulsions with substantial excess free protein present was investigated via confocal microscopy. The separate distributions of oil droplets and protein were determined using two different fluorescent dyes. In the emulsion at ambient temperature the excess protein and protein-coated oil droplets were associated together in a reversibly flocculated state. The pore-size distribution of the initial flocculated emulsion was found to depend on the surfactant/protein ratio R, and at higher values of R the system became more inhomogeneous due to areas of local phase separation. Evidence for competitive displacement of protein from the oil-water interface by surfactant was obtained only on heating (from 25 to 85 degrees C) during the process of formation of a heat-set emulsion gel. By measuring fluorescence intensities of the protein dye inside and outside of the oil-droplet-rich areas, we have been able to quantify the evolving protein distribution during the thermal processing. The results are discussed in relation to previous work on the competitive adsorption of proteins and surfactants in emulsions and the effect of emulsion droplets on the rheology of heat-set protein gels.     

A differential microcalorimetric study of whey proteins and their behaviour in oil-in-water emulsions

Oil-in-water emulsions (20% soya oil, 1% protein) have been prepared containing lysozyme or isolates of -lactalbumin and β-lactoglobulin from whey protein. The structural characteristics of these proteins adsorbed at an oil/water interface were determined by following their thermal transitions using differential scanning microcalorimetry. Thermograms of the proteins in the adsorbed state differed markedly from the corresponding transitions seen for the proteins in solution. This suggests that the proteins underwent substantial changes in secondary and tertiary structure upon adsorption. In general, for all the proteins studied, a net decrease in the total energy absorbed during denaturation was found when the proteins were in an adsorbed state. Both lysozyme and -lactalbumin were in part “surface denatured”, and they showed a certain degree of reversibility between solution and the adsorbed state. β-Lactoglobulin showed the highest degree of denaturation upon adsorption and the conformational change was irreversible.     

Preparation and characteristics of multiple emulsions containing liquid crystals

In this paper, multiple emulsions containing liquid crystals were prepared successfully and the influence of formulation parameters on the formation mechanism was studied. Moreover, differential scanning calorimetry (DSC), small-angle X-ray scattering (SAXS) spectra analysis and stability analysis were used to characterise the property of them. The results showed that the chemical structure of water-in-oil (W/O) emulsifiers directly impacted on the formation of multiple structure, but the effect on the formation of liquid crystal structure was negligible. With the gap of the polarity between inner and outer liquid oils decreased, both multiple structure and liquid crystal structure were harder to form. The content of sodium chloride in internal aqueous phase, which should be neither too high nor too low, has great impact on the formulation of multiple structure. It was easier to form two structures simultaneously when the carbon chain length of fatty alcohols was closer to that of emulsifier C₂₂ alkyl polyglucoside (202). DSC elucidated the phase transitions of water in the liquid crystal layer and the W/O emulsions. SAXS indicated that the liquid crystal orientation was lamellar. The stability analysis showed that the presence of liquid crystal structure had a significant contribution to the stability of the multiple emulsions.     

Egg yolk protein gels and emulsions

Egg yolk remains a key ingredient of a number of food products. Yet, its main functional properties, e.g. emulsifying ability and gel structure formation, upon heating, have not attracted the attention of too many researchers specializing in the area of food colloids. It is not surprising then that there have been only very few major advances in the field over the period of the last few years. These are discussed in the present review and include recent research findings on competitive adsorption between yolk protein constituents at emulsion oil–water interfaces, and also on the relationship between yolk particle supermolecular structure disorganization and the rheological properties of yolk-based emulsions and gel-network structures.     

Influence of sodium dodecyl sulfate on the physicochemical properties of whey protein-stabilized emulsions

The influence of sodium dodecyl sulfate (SDS) on the flocculation of droplets in 20 wt.% soybean oil-in-water emulsions stabilized by whey protein isolate (WPI) was investigated by light scattering, rheology and creaming measurements. The SDS concentrations used were low enough to prevent depletion flocculation by surfactant micelles and extensive protein displacement. In the absence of SDS, emulsions were prone to droplet flocculation near the isoelectric point of the proteins (4<pH<6), but were stable at a higher and lower pH. Flocculation led to an increase in emulsion viscosity, pronounced shear thinning behavior and accelerated creaming. When the surfactant-to-protein molar ratio was increased from 0 to 10, the emulsion instability range shifted to lower pH values due to binding of the negatively charged SDS molecules to the droplets. Our results indicate that the physicochemical properties of protein-stabilized emulsions can be modified by utilizing surfactant–protein interactions.     

Release of electrolytes from W/O/W double emulsions stabilized by a soluble complex of modified pectin and whey protein isolate

W/O/W double emulsions (DEs) stabilized by charged soluble complexes of whey protein isolate (WPI) and modified pectins were investigated in relation to their stability and the release of two types of electrolytes, NaCl and sodium ascorbate.WPI alone cannot properly stabilize the DEs. The droplet size is relatively large (100 μm) and increases with time. However, addition of modified pectin to form a soluble complex with WPI significantly improved the stability.DEs prepared with two types of oils (medium chain triglycerides (MCT) and R(+)-limonene) were studied by measuring droplet size, creaming, viscosity, and electrolyte release. Irrespective of their very different oil phase nature, both emulsions were stable against coalescence, but R(+)-limonene formed smaller droplets (25 μm) than MCT (35 μm). The electrolyte release rate was significantly higher from the R(+)-limonene that formed DEs with much lower viscosity. R(+)-limonene-DE released 75% of the NaCl after 28 days, while MCT-DE released only 50%. NaCl was released more slowly than sodium ascorbate.Apparently, the release mechanism from R(+)-limonene-DE was found to be “thinning the outer interface and release of the entire inner droplets” while it seems that the release from MCT-DE was slower and “diffusion controlled”.DEs stabilized by WPI/C63 released 12% of the sodium ascorbate after 1 day in milk and remained stable for at least 8 days. However, DEs stabilized with only WPI released about 50% of the sodium ascorbate after 1 day, and phase separated after 8 days.     

Development of the emulsions containing modified fats formed via enzymatic interesterification catalyzed by specific lipase with various amount of water

The aim of this work was to obtain and evaluate the stability of new emulsion systems, in which diacylglycerols derived from enzymatic interesterification of mutton tallow with hemp oil were used as emulsifiers. In order to achieve a higher content of the polar fat fraction in the final fat blend, a different amount of water was added to the reaction mixtures. The modified fats with a mixture of mono and diacylglycerols served as a fat base for emulsions. Based on the results of Turbiscan test, the droplet size, emulsion texture, and studies of rheological properties, it was found that addition of water to the reaction mixture in the range of 1.00–1.25%, wt./wt., caused the formation of a sufficient amount of emulsifiers stabilizing the dispersion system.

The novelty of this work was to determine the optimal amount of water added to the interesterification of mutton tallow with vegetable oil, ensuring the synthesis of a high-efficiency emulsifying system. Another new aspect of this work was to show that the diacylglycerols obtained during such fat modification constitute effective emulsifiers for new stable emulsion systems that may find potential use as food emulsions (dressings) or cosmetic products dedicated to sensitive skin.     

Determination of free L- and D-alanine in hydrolysed protein fertilisers by capillary electrophoresis

of racemisation of hydrolysed protein fertilisers (HPFs) using an The objective of this study was to determine the degree inexpensive and easy to handle analytical method for qualitative control of the products. Using a polyacrylamide coated capillary and a run buffer containing 0.1 M Tris-borate+2.5 mM EDTA-Na2+0.1% sodium dodecylsulfate+10 mM beta-cyclodextrin a quantitative separation of D- and L-alanine (Ala) was made from an not treated HPF sample derivatised with dansyl chlorine by capillary electrophoresis. The D-Ala:[D-Ala+L-Ala] ratio, called degree of racemisation (RD), was calculated. The analysis of ten commercial HPFs has shown that more than 60% of HPFs have an RD > or = 40%. while only one product has shown an RD <5%. These results showed that most of the HPFs on the market are obtained with strong hydrolytic processes and high contents of D-amino acids are probably less effective as plant nutrients or even potentially dangerous to plants.     

Stability of emulsions of water in mineral oils containing asphaltenes

Emulsions of water in mineral oils are stable if the oil phase contains asphaltenes which are near the condition of incipient flocculation. This condition is determined by the composition of the oil phase and by the nature of the asphaltenes. High aromaticity of the oil phase and the presence of deflocculants prevent flocculation of asphaltenes; the deflocculants may be interfacially active agents or asphaltene-like compounds with better solubility in the oil phase. Conditions of incipient flocculation of asphaltenes correlate very well with a considerable increase of rheological resistance of the interface between the oil phase and distilled water, determined according to the torsion oscillation method. Stabilization of the water-in-oil emulsions is therefore caused by the build-up of a coherent layer of asphaltenes in the water-oil interface in these cases. Deflocculants of asphaltenes in the oil phase destroy their stabilizing effect; however, the deflocculants themselves may stabilize the water-in-oil emulsions by adsorption on the water-oil interface and then the correlation between the condition of asphaltenes and emulsion stability does not hold, nor is the interfacial viscosity perceptibly increased. Under borderline conditions of emulsion stability a few percent of sodium chloride in the water phase counteracts the build-up of a stabilizing layer of asphaltenes in the water-oil interface and so do higher p_H values of a buffered water phase. At low p_H-values emulsion stability does not correlate with interfacial resistance. It can be concluded that asphaltenes stabilize water-in-oil emulsions if they accumulate on the water-oil interface. This interfacial layer may show a coherence, which is an indication of the presence of asphaltenes rather than a condition for stability of the emulsions.     

Thermoanalytical and microscopical investigation of the microstructure of emulsions containing polymeric emulsifier

Polymeric emulsifiers provide exceptional stability to oil-in-water, water-in-oil or multiple emulsions by their steric stabilization. Pemulens as polymeric emulsifiers are able to stabilize o/w type emulsions because their short lipophilic part integrates into the oil droplets while their long hydrophilic part forms a micro gel around the droplet. In our present study the microstructure and integration of the polymeric emulsifier at the water-oil interface was investigated with thermogravimetric and microscopical methods. It was established that depending on the amount of both of the polymeric emulsifier and added coemulsifier the microstructure of the system changes.     

Chromatographic amino acid analysis of protein hydrolysed in polyacrylamide after removal of ammonia

Summary A simple method for the removal of NH₃ from amino acids is presented. The method is based on a cation-exchange resin from which amino acids are eluted with NH₄OH. The eluent is then removed under reduced pressure. The method allows the ninhydrin-based detection of amino acids after hydrolysis of stained protein bands in polyacrylamide gels. This was previously not possible since NH₃ produced by the hydrolysis of polyacrylamide interferes with the ninhydrin-detection of basic amino acids. The method should also be applicable to the detection of amino acids with o-phthalaldehyde.     

Potential of various archae- and eubacterial strains as industrial polyhydroxyalkanoate producers from whey

Three different microbial wild-type strains are compared with respect to their potential as industrial scale polyhydroxyalkanoate (PHA) producers from the feed stock whey lactose. The halophilic archaeon Haloferax mediterranei as well as two eubacterial strains (Pseudomonas hydrogenovora and Hydrogenophaga pseudoflava) are investigated. H. mediterranei accumulated 50 wt.-% of poly-3-(hydroxybutyrate-co-8%-hydroxyvalerate) from hydrolyzed whey without addition of 3-hydroxyvalerate (3HV) precursors (specific productivity q(p): 9.1 mg x g(-1) x h(-1)). Using P. hydrogenovora, the final percentage of poly-3-hydroxybutyrate (PHB) amounted to 12 wt.-% (q(p): 2.9 mg x g(-1) x h(-1)). With H. pseudoflava, it was possible to reach 40 wt.-% P-3(HB-co-5%-HV) on non-hydrolyzed whey lactose plus addition of valeric acid as 3HV precursor (q(p): 12.5 mg x g(-1) x h(-1)). A detailed characterization of the isolated biopolyesters and an evaluation with regard to the economic feasibility completes the study.