Heat stability and freeze–thaw stability of oil-in-water emulsions stabilised by sodium caseinate–maltodextrin conjugates

A sodium caseinate (NaCN)–maltodextrin (Md100) conjugate was prepared by a Maillard-type reaction by dry heat treatment of a NaCN–Md100 mixture at 60 °C and 79% relative humidity for 4 days. Conjugation resulted in a 35.7% loss of available amino groups in the NaCN and a 25.9% loss of available reducing groups in the Md100. The crude conjugate was purified by batch anion exchange chromatography to remove non-conjugated Md100. Purification reduced the available reducing groups in the conjugate from 74.1% to 23.7% and increased the protein content from 45.6% to 83.9%. The emulsifying properties of the conjugates were assessed in oil-in-water (o/w) emulsions; crude and purified conjugate stabilised emulsions had improved storage stability and freeze–thaw stability when compared to NaCN stabilised emulsions. Purified conjugate stabilised emulsions had better thermal stability than NaCN, NaCN–Md mixture and non-purified conjugate stabilised emulsions. These results indicate a potential for these NaCN–Md conjugates as speciality functional food ingredients.     

Factors influencing the production of o/w emulsions stabilized by β-lactoglobulin–pectin membranes

A primary emulsion was prepared by homogenizing 10 wt% corn oil with 90 wt% aqueous β-lactoglobulin solution (0.5 wt% β-lg, pH 3 or 7) using a two-stage high-pressure valve homogenizer. This emulsion was mixed with aqueous pectin (citrus, 59% DE) stock solution (2 wt%, pH 3 or 7) and NaCl solution to yield secondary emulsions with 5 wt% corn oil, 0.225 wt% β-lactoglobulin, 0.2 wt% pectin and 0 or 100 mM NaCl. The final pH of the emulsions was then adjusted (3–8). Primary and secondary emulsions were ultrasonically treated (30 s, 20 kHz, 40% amplitude) to disrupt any flocculated droplets. Secondary emulsions were more stable than primary emulsions at intermediate pHs. Secondary emulsions prepared at pH 7 had smaller particle diameters (0.35 to 6 μm) than those prepared at pH 3 (0.42 to 18 μm) across the whole pH range studied, and also had smaller diameters than the primary emulsions (0.35 to 14 μm). Ultrasound treatment reduced the particle diameter of both primary and secondary emulsions and lowered the rate of creaming. The presence of NaCl screened the charges and thus the electrostatic interaction between biopolymer molecules and primary emulsion droplets. Secondary emulsions were more stable to the presence of 100 mM NaCl at low pHs (3–4) than primary emulsions. This study shows that stable emulsions can be prepared by engineering their interfacial membranes using the electrostatic interaction of natural biopolymers, especially at intermediate pHs where proteins normally fail to function.     

Stability and rheology of egg-yolk-stabilized concentrated emulsions containing cereal β-glucans of varying molecular size

The effects of barley and oat β-glucans on rheological and creaming behaviour of concentrated egg-yolk-stabilized model emulsions were investigated. Four polysaccharide preparations were used, two from each cereal; one sample with high and one with low molecular weight, i.e. the molecular weights were alike in pairs (110×10³ and 40×10³, respectively). In order to elucidate the mechanism of action of β-glucans in emulsions, Tween 20-stabilized emulsions were also examined. Tween 20 enhances neither the continuous phase viscosity nor the interactions between the droplets, so the changes could be easily attributed to β-glucans. It appeared that the low Mw β-glucan samples stabilize emulsions against creaming by means of network formation in the continuous phase while their high molecular weight counterparts enhance the viscosity of the continuous phase. Comparison of dynamic rheological tests between a reference emulsion without β-glucans and emulsions containing β-glucans showed that the polysaccharides largely affects the viscoelastic behaviour of the emulsion. Ageing of β-glucan-containing emulsions did not affect significantly the viscoelastic properties except for the emulsions containing low Mw β-glucans extracted from oat. Interestingly, all emulsions containing β-glucans creamed approximately the same after 30 days of storage regardless which preparation was used. The egg yolk constituents seemed to play a dominant role on the viscoelastic and the creaming behaviour of the emulsions, i.e. the viscoelastic behaviour was further enhanced and this could not only be attributed to the presence of the β-glucans but also to the stronger interactions between the oil droplets. Ageing did not affect the viscoelastic properties of β-glucan-containing emulsions while the reference emulsion, prepared only with egg yolk, showed a decrease in the value of storage modulus. The former could be interpreted as a steady consistency of the product during storage independent of the creaming behaviour. The creaming behaviour varied among the samples with the high molecular weight β-glucans from oat showing the highest stability.     

The molecular characterisation and antimicrobial properties of amidated bovine β-lactoglobulin

Amidation of bovine β-lactoglobulin (β-Lg) imparted antimicrobial properties to this protein. Amidated β-Lg was strongly bactericidal against resting cells of Pseudomonas fluorescens, Pseudomonas fragi and Bacillus subtilis, but had a much weaker effect against Escherichia coli, Enterococcus faecalis, Salmonella typhimurium and Listeria monocytogenes. Neither native nor amidated β-Lg was effective against the yeast Saccharomyces cerevisiae and the mould Penicillium candidum. Mass spectrometric analysis demonstrated that amidation of β-Lg converted aspartyl and glutamyl residues to asparaginyl and glutaminyl residues, respectively, and that the amidation reaction did not occur to the same extent on every β-Lg molecule. The charge change was also confirmed by SDS–PAGE, ion exchange chromatography and the change in isoionic point of β-Lg. Reverse-phase chromatography showed that amidation also led to alterations in hydrophobicity of the β-Lg molecule. The antibacterial properties of the amidated β-Lg appear to be dependent on the net positive charge and charge distribution on the molecule.     

Impact of cosolvents on formation and properties of biopolymer nanoparticles formed by heat treatment of β-lactoglobulin–Pectin complexes

The purpose of this study was to determine the influence of neutral cosolvents on the formation and properties of biopolymer nanoparticles formed by thermal treatment of protein–polysaccharide electrostatic complexes. Biopolymer particles were formed by heating (85 °C, 20 min) an aqueous solution containing a globular protein (β-lactoglobulin) and an anionic polysaccharide (beet pectin) above the thermal denaturation temperature (T_m) of the protein under pH conditions where the biopolymers formed electrostatic complexes (pH 5). The impact of two neutral cosolvents (glycerol and sorbitol) on the self-association of β-lactoglobulin and on the formation of β-lactoglobulin–pectin complexes was examined as a function of solution pH (3–7) and temperature (30–95 °C). Glycerol had little impact on the pH-induced self-association or aggregation of the biopolymers, but it did increase the thermal aggregation temperature (T_a) of the protein–polysaccharide complexes, which was attributed to its ability to increase aqueous phase viscosity. Sorbitol decreased the pH where insoluble protein–polysaccharide complexes were formed, and greatly increased their T_a, which was attributed to its ability to increase T_m, alter biopolymer–biopolymer interactions, and increase aqueous phase viscosity. This study shows that neutral cosolvents can be used to modulate the properties of biopolymer nanoparticles prepared by thermal treatment of protein–polysaccharide electrostatic complexes.     

Heat-induced phase behavior of β-lactoglobulin/polysaccharide mixtures

The influence of polysaccharides on the thermal stability of β-lactoglobulin at pH 6.8 was investigated regarding polysaccharide type, concentration and size. Two kinds of polysaccharides, sulfate-containing polysaccharides (carrageenans and dextran sulfate with different molecular mass) and neutral polysaccharides (dextran with different molecular mass), were investigated. At low ratios of sulfate-containing polysaccharide to β-lactoglobulin, heat-induced aggregation was decreased as shown by lower turbidity. Increasing the ratio induced a significant increase in turbidity, leading to segregative phase separation. Phase diagrams were established by centrifugation, chemical assays and visual observation for β-lactoglobulin/kappa-carrageenan and β-lactoglobulin/dextran sulfates. Significant phases (stable, separated and gel) were found, indicating the varieties of phase behavior and a strong competition between phase separation and gelation caused by thermal treatment. Moreover, gelation was reversible in β-lactoglobulin/dextran sulfate systems depending on the polysaccharide concentration.     

Microencapsulation of β‐carotene using native pinhão starch,modified pinhão starch and gelatin by freeze‐drying

Beta‐carotene was microencapsulated by freeze‐drying using native pinhão starch, hydrolysed pinhão starch 6 dextrose equivalent (DE), hydrolysed pinhão starch 12 DE and the mixture of these materials with gelatin as coating material. The purpose of this research was to produce and characterize these microcapsules. The capsules’ efficiency, surface content, moisture, morphology, solubility, particle size and glass transition temperature were analysed. The hydrolysed pinhão starch 12 DE showed the highest total β‐carotene content and the lowest surface β‐carotene content, unlike the native starch. Using scanning electron microscopy, it was observed that all microcapsules presented undefined shapes. The samples with gelatin had wider particle size distribution, higher diameters, lower solubility and higher glass transition temperature when compared with other the samples. Results obtained suggest that the modified pinhão starch can be considered as potential wall material for encapsulation of β‐carotene.     

Cold-set thickening mechanism of β-lactoglobulin at low pH: Concentration effects

There is an interest in developing protein based thickening agents for nutritional considerations. A procedure to convert whey protein concentrates or isolates into a pH modified cold-thickening ingredient was developed. Concentration effects on thickening mechanism of this whey protein ingredient were studied with a β-lactoglobulin model system at the pH of the modification procedure, 3.35. In this study, concentration effects on thermal aggregation of β-lactoglobulin were studied at low pH using capillary and rotational viscometry, transmission electron microscopy (TEM), and high performance liquid chromatography coupled with multi-angle laser light scattering (HPLC-MALS). From the results of capillary viscometry, a critical concentration (C_c  6.9% w/w) was identified below which no significant thickening functionality could be achieved. Microscopy revealed formation of flexible fibrillar network at pH 3.35 during heating at all concentrations. These flexible fibrils had a diameter of about 5 nm and persistence length of about 35 nm as compared to more linear and stiff fibrils formed at pH 2 and low ionic strength conditions. Under similar heating conditions at concentration above C_c, larger aggregates similar to microgels were observed compared to the concentration below C_c, where isolated fibrils with an average contour length of about 130 nm were observed. These microgels and apparently stronger interactions between aggregates at concentrations above C_c were seemingly responsible for thickening functionality of heated β-lactoglobulin solutions and subsequently modified powders. Further investigation of β-lactoglobulin aggregation at this pH may provide capability to mechanistically tailor the functional attributes of modified ingredients.     

Effect of gamma irradiation parameters on the in-vitro digestibility of β-lactoglobulin

Radiation level and protein conformation during irradiation may modify a protein's response to ionising energy and change its nutritional value. Effects of gamma irradiation on protein in-vitro digestibility and amino acid release were investigated using β-lactoglobulin as a model protein. Irradiation with ⁶⁰Co was performed at a range of 1–50 kGy on liquid or frozen samples atA_w 0.93, with pH adjusted to 5.2 or 7.0, and on dry samples atA_w 0.13. Amino acid composition, in-vitro protein digestibility and amino acid release were not impaired by irradiation at the highest dose of 50 kGy. Thus the nutritional quality of β-lactoglobulin was not affected by irradiation in a dose range likely to be used in food processing.     

Stability of whipped dairy creams containing locust bean gum/λ-carrageenan mixtures during freezing–thawing processes

The role of locust bean gum (LBG)-λ-carrageenan mixtures on the stability of whipped dairy creams in freezing-thawing processes was analysed as a function of the gum concentration (between 0 and 0.1% w/w). DSC analysis of ice crystallisation in the cream aqueous phase was carried out in order to know the influence of the hydrocolloid ratio on freezable water content. From this analysis, a potential cryostabiliser effect of whey proteins could be deduced since they provoked an increase in the T_m′ value. Changes in overrun, viscoelastic behaviour (creep compliance parameters) and stiffness (extrusion test) due to freezing–thawing and frozen storage (1 month at −18 °C) of the whipped cream were analysed in samples. Freezing provoked collapse of the foam structure, but samples containing λ-carrageenan at concentration greater than 0.085% showed a well preserved firmness, as deduced from the small changes observed in their viscoelastic parameters. The λ-carrageenan cryoprotection mechanism is not based on the freezable water content reduction since this value was similar for all gum mixtures.     

The role of pulsed electric fields treatment in enhancing the stability of amino acid – sugar complexes:‐ interactions between L‐Phenylalanine and β‐Cyclodextrin

In this work, the effect of pulsed electric fields (PEF) treatment on the interactions between amino acids (using L‐Phenylalanine: L‐Phe) and sugar (using β‐Cyclodextrin: β‐CD) complex was analysed by fluorescence spectroscopy, Raman spectroscopy and simultaneous thermal analyzer. Moreover, the molecular dynamics of β‐CD–L‐Phe inclusion complex treated by PEF was calculated by molecular modelling. The results indicated that β‐CD–L‐Phe complexes are formed by a molar ratio of 1:1, and the stability constant of such complexes increased from 147 to 614 M^?1 by PEF treatment. Thermal characterisations of β‐CD–L‐Phe complexes indicated that the PEF treatment could increase the yield of complexes. The PEF treatment resulted in an increase in the reaction enthalpy of β‐CD–L‐Phe inclusion complexes by DSC curve. These results show that PEF treatment has the potential to promote the chemical processing, especially the small organic molecules participate in inclusion or cross‐linking reaction.     

Modified stainless steel surfaces targeted to reduce fouling – Evaluation of fouling by milk components

Several stainless steel based surfaces with different properties were evaluated according to their fouling behaviour for different dairy products under different conditions. Surface properties were obtained by the following modification techniques: , and TiC ion implantation; diamond-like carbon (DLC) sputtering; DLC, DLC–Si–O and SiO_x, plasma enhanced chemical vapor Deposition (PECVD); autocatalytic Ni–P–PTFE and silica coating. Aqueous solutions that simulate milk (SMUF – simulated milk ultrafiltrate for the mineral components, β-lactoglobulin for the protein components and FMF – fouling model fluid for complex milk systems) were used to study the fouling behaviour during pasteurisation. Bacteriological deposition studies were also performed with two heat resistant strains of Bacillus. The experiments were carried out at laboratory scale for the evaluation of calcium phosphate and protein deposition, and at pilot scale for adhesion of bacteria and deposits from complex milk systems.

In all cases, the fouling behaviour was affected by the surface material, although in different ways for the deposition or the cleaning phases. For the non-microbiological deposits (calcium phosphate, whey protein and FMF milk-based product), the Ni–P–PTFE surface was the most promising one, since it generally promoted less deposit build up and, in all cases, was the easiest to clean. On the other hand, for bacterial adhesion, the most suitable surface was the ion implanted (TiC) surface, which also showed less spores after the cleaning process.     

SOME BARLEY GRAIN AND GREEN MALT PROPERTIES AND THEIR INFLUENCE ON MALT HOT-WATER EXTRACT: I. β-GLUCAN, β-GLUCAN SOLUBILASE AND ENDO-β-GLUCANASE

A study has been made of the variation between varieties in some properties of barley and malt and how this variation relates to malt hot water extract (HWE). The development of enzyme activity along the grain during germination was investigated. In this first paper we have examined β-glucan-related characters and found significant varietal variation in maximum enzyme activities and in the activities in different sections of grain during germination. Varietal variation was greater than environmental variation for each character. The fraction of β-glucan soluble in acid was the character most highly correlated with HWE.     

Statistical Modeling of β-galactosidase Inhibition During Lactose Hydrolysis

A statistical model approach called response surface methodology was used to describe the product and substrate inhibition effect on β-galactosidase enzyme during lactose hydrolysis. The effect of independent variables, namely the initial concentrations of lactose (73 - 146 mM), galactose (44 - 122 mM) and glucose (83 - 167 mM) on the reaction rate of β-galactosidase was evaluated. The enzymatic reaction rate was influenced by both combined and individual effects of all the substrate and products. Although, glucose acted as an activator at low lactose and low galactose concentrations, glucose caused the inhibition of β-galactosidase at higher concentrations of lactose and galactose. The effect of galactose concentration on β-galactosidase enzyme was in the direction of inhibition. At low lactose concentrations and high glucose concentrations, galactose concentration became more effective on the reaction rate.     

β-GLUCANASE: THE SUCCESSFUL APPLICATION OF GENETIC ENGINEERING

The now well established principles of genetic engineering are described in relation to the solution of problems associated with β-glucans in beer. The application of these techniques has enabled the isolation of a Bacillus subtilis endo-1, 3–1, 4-β-D-glucanase gene which expresses a biologically active enzyme in yeast.^15,16 Although this enzyme is capable of hydrolysing beer β-glucans during fermentation, thereby enhancing beer filtration, insufficient β-glucanase is produced in yeast to enable successful commercial implementation. The requirements for the efficient production of β-glucanase in genetically manipulated brewing yeast are described.     

Release of β-casomorphins 5 and 7 during simulated gastro-intestinal digestion of bovine β-casein variants and milk-based infant formulas

The release of β-casomorphin-5 (BCM5) and β-casomorphin-7 (BCM7) was investigated during simulated gastro-intestinal digestion (SGID) of bovine β-casein variants (n = 3), commercial milk-based infant formulas (n = 6) and experimental infant formulas (n = 3). SGID included pepsin digestion at pH 2.0, 3.0 and 4.0 and further hydrolysis with Corolase PP™. β-Casein (β-CN) variants were extracted from raw milks coming from cows of Holstein-Friesian and Jersey breeds. Genomic DNA was isolated from milk and the β-CN genotype was determined by a PCR-based method. Phenotype at protein level was determined by capillary zone electrophoresis in order to ascertain the level of gene expression. Recognition and quantification of BCMs involved HPLC coupled to tandem MS. Regardless of the pH, BCM7 generated from variants A1 and B of β-CN (5–176 mmol/mol casein) the highest amount being released during SGID of form B. As expected, the peptide was not released from variant A2 at any steps of SGID. BCM5 was not formed in hydrolysates irrespective of either the genetic variant or the pH value during SGID. Variants A1, A2 and B of β-CN were present in all the commercial infant formulae (IFs) submitted to SGID. Accordingly, 16–297 nmol BCM7 were released from 800 ml IF, i.e. the daily recommended intake for infant. Industrial indirect-UHT treatments (156 °C × 6–9 s) did not modify release of BCM7 and, during SGID, comparable peptide amounts formed in raw formulation and final heat-treated IFs.     

Effect of dextran glycation on nanofibril assembly of soya β‐conglycinin at pH 2.0 and the pH stability of nanofibrils

In this study, we investigated the effects of dextran glycation on soya β‐conglycinin self‐assembly into nanofibrils at 85 °C and pH 2.0, as well as their stability at pH 2.0–10.0. Although the hydrolysis rate of β‐conglycinin decreased, glycation significantly increased the structural change rate in the initial stage of nanofibril formation and the growth of nanofibril. It is suggested that the glycation of three subunits (α′, α, β) in β‐conglycinin may promote fibril assembly because the extension regions of α′ and α subunits play an important role in affecting the rate of structural changes in fibril formation. At neutral pH, conjugate nanofibrils are highly dispersible and transparent and remained greater structural stability compared with mixture. The improvement stability of conjugate nanofibrils may be contributed to dextran which provides some steric hindrance to prevent the aggregation of nanofibrils; this would also facilitate the application of soya β‐conglycinin nanofibrils in food industry.     

Effect of Mashing Temperatures and Endo-β-Glucanase on β-Glucan Content of Malt Worts

Similar basal levels of β-D-glucans were released into worts produced at 45°C from enzymically active or inactivated flours of milled malts. In contrast, significantly higher levels of β-D-glucans were found in worts derived from either enzymically active or inactivated malt flours mashed at 65°C. In general, mashing temperature may play a more important role in releasing β-D-glucans during mashing than enzymes described as β-glucan-releasing. In this context, the physical release of β-D-glucan during mashing should be separated from the enzymic release and degradation of β-D-glucan which occur during malting.     

Residues of β-lactam antibiotics in bovine milk: confirmatory analysis by liquid chromatography tandem mass spectrometry after microbial assay screening

A procedure for the simultaneous determination of the residues of seven β-lactam antibiotics (penicillin G, ampicillin, oxacillin, amoxicillin, dicloxacillin, cephalexin, cephapirin) in bovine raw milk using tandem liquid chromatography mass spectrometry (LC-MS/MS) is described. The antibiotics were extracted by an acetic acid solution after centrifugation and filtration. The β-lactams were separated using reversed-phase liquid chromatography. A triple quadrupole mass spectrometer was used in positive-ion mode as a detector via a Turbo Ionspray interface for electrospray ionization (ESI). The limits of detection and quantitation of the method were below the legal tolerances, except for ampicillin. The method was used to confirm 53 samples found positive by a microbial method (Delvotest SP®) at the Istituto Zooprofilattico Sperimentale per la Lombardia e l'Emilia Romagna of Brescia during 2001. Penicillin G was found in 26 samples at concentrations ranging from less than 4 to 6240 ±550 μg l^-1. Amoxicillin was found in three samples at concentrations ranging from 8.5 ±0.1 to 53.7 ±2.3 μg l^-1. Cephapirin was found in two samples at 5.7 ±0.1 and 6.4 ±0.3 μg l^-1.