SURFACE PROPERTIES OF PROTEIN LAYERS ADSORBED FROM MIXTURES OF GELATIN WITH VARIOUS CASEINS

We report surface tensions and surface shear viscosities of protein layers adsorbed from mixtures of gelatin +α_s11-casein, β-casein, k-casein or sodium caseinate. Under conditions where the two protein components carry opposite net charges, the combined experimental data at 25C and neutral pH are consistent with a two-layer model of the mixed protein film. The surface properties of sodium caseinate lie intermediate between those for the two major individual caseins (α_s1 and β).     

Factors Affecting Rheological Characteristics of Fibril Gels: The Case of β-Lactoglobulin and α-Lactalbumin

ABSTRACT:  Some of the factors that affect the rheological characteristics of fibril gels are discussed. Fibrils with nanoscale diameters from β-lactoglobulin (β-lg) and α-lactalbumin (α-la) have been used to create gels with different rheological characteristics. Values of the gelation time, t_c , the critical gel concentration, c ₀, and the equilibrium value of the storage modulus, G , such as     at long gelation times, derived from experimental rheological data, are discussed. Fibrils created from β-lg using solvent incubation and heating result in gels with different rheological properties, probably because of different microstructures and fibril densities. Partial hydrolysis of α-la with a serine proteinase from Bacillus licheniformis results in fibrils that are tubes about 20 nm in diameter. Such a fibril gel from a 10% (w/v) α-la solution has a higher modulus than a heat-set gel from a 10% (w/w) β-lg, pH 2.5 solution; it is suggested that one reason for the higher modulus might be the greater stiffness of α-la fibrils. However, the gelation times of α-la fibrils are longer than those of β-lg fibrils.     

Casein diversity in caprine milk and its relation to technological properties: heat stability

Caprine milk is characterized by a genetically linked diversity in casein content. Of particular importance is the genetic polymorphism of α_s1-casein, which is under the influence of seven known alleles controlling the level of α_s1-casein expression in milk. In this article, the effect of α_s1-casein genotype on milk composition and heat stability is considered. Two different types of caprine milk were used: milk high in α_s1-casein from animals heterozygous AE or BE with respect to α_s1-casein genotype and milk low in α_s1-casein from animals homozygous FF. Milk high in α_s1-casein was less stable upon heating than milk low in α_s1-casein. Unlike bovine milk there was no recovery in heat stability at pH values greater than 7.0. These differences were associated with factors in the serum rather than the casein fraction and could not be explained by the variation in the protein composition, /3–lactoglobulin to casein ratio or the small variation in Ca²⁺ and lactose levels between the two milks. On the other hand, milk low in α_s1-casein had a higher urea content, which had a significant stabilizing effect.     

Evaluation of Serpa cheese proteolysis by nitrogen content and capillary zone electrophoresis

Proteolysis of Serpa cheese produced traditionally (B) and semi-industrially (C) was evaluated for the first time by determination of nitrogen content and capillary zone electrophoresis (CZE). A citrate dispersion of cheese was fractionated to determine the nitrogen in pH 4.4, trichloroacetic and phosphotungstic acid soluble fractions (pH 4.4-SN, TCA-SN and PTA-SN, respectively). The pH 4.4-SN was significantly higher for B ( P  < 0.001), while TCA-SN was significantly higher for C ( P  < 0.001). PTA-SN was also higher for C but at 60 days ripening no significant difference was found between B and C. Degradation of α_s1- and β-caseins evaluated by CZE was in good agreement with the maturation index (pH 4.4-SN/TN).     

Production of hard cheese from caprine milk by the use of two types of probiotic cultures as adjuncts

Hard cheeses (Kefalotyri-like) were manufactured from caprine milk with yoghurt as a starter (A), and with its partial replacement with the probiotic adjuncts Lactobacillus rhamnosus LC 705 (B) and/or Lactobacillus paracasei ssp. paracasei DC 412 (C). Both adjuncts retarded the growth of enterococci, and the environment in cheese B did not favour the recovery of lactic acid bacteria (LAB) on Rogosa agar. However, better recovery of the LAB population on M17 agar from cheeses B and C made with adjuncts was recorded early in ripening, and this was accompanied by a greater decrease in pH. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) of whole-cell protein demonstrated that cheese C, made with Lb. paracasei ssp. paracasei as adjunct, is a better vehicle for delivery of live probiotic cells (10 ⁷   cfu/g) to the gastrointestinal tract than cheese B, made with Lb. rhamnosus ; the latter did not belong to the predominant microflora of one out of the two B cheeses. Urea-PAGE electrophoresis results indicated that adjunct lactobacilli enhanced the degradation of both α _S -casein (α _S -CN) and β-casein (β-CN). In the fresh cheese, hydrolysis of α _S -CN was more rapid than β-CN, and the free amino acid content of B and C was higher than in A. Lipolysis products were also higher in B and C than in A as ripening progressed, and the organoleptic characteristics of these cheeses resulted in higher scores, in the order C > B > A. Thus, making Kefalotyri-like cheese from caprine milk with probiotic lactobacilli, particularly Lb. paracasei ssp. paracasei, as adjunct can be considered an effective way of producing a cheese with a large number of probiotic cells.     

Evaluation of changes during storage of probiotic dahi at 7°C

Low-fat probiotic dahi (an Indian traditional fermented milk product like yogurt) prepared with lactococci starter and two adjunct probiotic cultures of Lactobacillus acidophilus and Lactobacillus casei was stored at 7°C. The survival of bacterial species was affected during storage of dahi, in which viable counts increased up to 2 days and gradually decreased from 2 days, up to 8 days. The quality of refrigerated dahi was assessed by a panel of trained judges. On storage, pH significantly decreased over storage time, indicating that the dahi samples did not develop much acidity under the storage conditions of the study. However, after 8 days of storage, the samples were disliked by the panel, which reported slight bitterness, which may have been due to proteolysis. Proteolysis increased during storage. More α_s1-casein was degraded than β-casein. On the basis of the results of this study we can conclude that dahi stored up to 8 days may be acceptable to consumers.     

Fractionating Soybean Storage Proteins Using Ca2+ and NaHSO3

ABSTRACT:  Individual soybean storage proteins have been identified as having nutraceutical properties, especially β-conglycinin. Several methods to fractionate soy proteins on industrial scales have been published, but there are no commercial products of fractionated soy proteins. The present study addresses this problem by using calcium salts to achieve glycinin-rich and β-conglycinin-rich fractions in high yields and purities. A well-known 3-step fractionation procedure that uses SO_2, NaCl, and pH adjustments was evaluated with CaCl₂ as a substitute for NaCl. Calcium was effective in precipitating residual glycinin, after precipitating a glycinin-rich fraction, into an intermediate fraction at 5 to 10 mM CaCl₂ and pH 6.4, eliminating the contaminant glycinin from the β-conglycinin-rich fraction. Purities of 100%β-conglycinin with unique subunit compositions were obtained after prior precipitation of the glycinin-rich and intermediate fractions. The use of 5 mM SO₂ in combination with 5 mM CaCl₂ in a 2-step fractionation procedure produced the highest purities in the glycinin-rich (85.2%) and β-conglycinin-rich (80.9%) fractions. The glycinin in the glycinin-rich fraction had a unique acidic (62.6%) to basic (37.4%) subunit distribution. The β-conglycinin-rich fraction was approximately evenly distributed among the β-conglycinin subunits (30.9%, 35.8%, and 33.3%, for α', α, and β subunits, respectively). Solids yields and protein yields, as well as purities and subunit compositions, were highly affected by pH and SO₂ and CaCl₂ concentrations.     

ADSORPTION BEHAVIOR OF β-LACTOGLOBULIN ONTO STAINLESS STEEL SURFACES

Nonporous stainless steel particles with an average diameter of 25.5 μm and an average surface area of 215 cm²/g were used to study the adsorption of β-lactoglobulin (β-lg). Scanning electron microscopy of stainless steel particle surfaces with adsorbed β-lg showed an obvious eradication of crevices as protein was adsorbed. The adsorption process for β-lg was very rapid in the first 5 min and essentially reached equilibrium within 10 min. Precipitation of calcium phosphate onto the stainless steel surface was very slow compared to monolayer deposition of β-lg. A monolayer of β-lg adsorbed very tightly to the stainless steel surface and a significant fraction of the protein was irreversibly adsorbed. The amount of adsorbed protein decreased with increasing ratio of surface area to reaction volume. pH in the range 6.0–6.8 did not significantly affect the amount of adsorbed protein.     

Influence of the Coagulant Level on Early Proteolysis in Ovine Cheese-like Systems Made with Sterilized Milk and Cynara cardunculus

ABSTRACT: The effect of coagulant level on the quality and quantity of protein breakdown during the first 24 h of ripening of cheese-like systems, manufactured with sterilized ovine milk using crude aqueous extracts of Cynara cardunculus as coagulant, was experimentally assessed. Urea-polyacrylamide gel electrophoresis was performed on both water-soluble and water-insoluble cheese extracts to monitor the casein degradation pattern; the ripening extension index and the ripening depth index were thus calculated. Peptides from the water-soluble fraction were isolated by reverse-phase, high-performance liquid chromatography and partially sequenced by Edman degradation. Higher residual coagulant levels in curdled milk led to earlier breakdown of caseins, as expected. The primary cleavage sites were Phe105-Met106 in k-casein, Phe23-Val24 in α_s1-casein, and Leu127-Thr128, Ser142-Trp143, Leu165-Ser166, and Leu190-Tyr191 in β-casein.     

Proteolysis in ultra-filtered and conventional Iranian white cheese during ripening

Conventional and ultra-filtered (UF) Iranian white cheeses were made with almost identical gross chemical composition and the extent and characteristics of proteolysis were studied during ripening. UF cheeses exhibited a lower rate of development of pH 4.6-soluble nitrogen than conventional cheeses. The rates of degradation of α_s1-casein and particularly β-casein were lower in UF cheeses than in conventional cheeses. Plasmin activity was lower in UF cheeses than that in conventional cheese, whereas coagulant activity was higher in the former. Noticeable qualitative and quantitative differences were observed in reverse-phase high performance liquid chromatography (RP-HPLC) peptide profiles between UF and conventional white cheeses and chemometric analysis of peak height data distributed the cheeses into two separate groups. The levels of free amino acids in UF cheeses were lower than in conventional cheeses. Lower peptide degradation and production of amino acids suggested slower ripening, which may have been associated with the weak aroma development characteristic of UF cheeses.     

EFFECT OF ADDITION OF α-, β-, AND κ-CASEIN, AND Na-CASEINATE ON VISCOELASTIC PROPERTIES OF SKIM MILK CURD

Creep compliance was used to determine the effects of the addition of α-, β-, and κ-casein, and Na-caseinate on the viscoelastic properties of skim milk curd. The results of all measurements can be represented by a six-element mechanical model. Addition of α- and β-casein, and Na-caseinate (1.80g/L) to raw skim milk reduced the instantaneous modulus of rigidity and final viscosity of the curd, while κ-casein addition at the same level increased both viscoelastic parameters. Shielding of κ-casein and depletion of serum Ca⁺⁺ ions by α- and β-casein is thought to have caused the reduction of curd rigidity and viscosity. Subsequent experiments indicated that the addition of β-casein before and after rennet hydrolysis produced different curd strength with the latter producing a stronger curd.     

PROTEIN DEGRADATION IN CHEDDAR CHEESE SLURRIES

SUMMARY— Preferential degradation of caseins was noted in the ripening of Cheddar curd slurries, with the degradation generally being in the order of para-K, β-, α_s-caseins. The addition of reduced glutathione to the slurry caused an immediate release of peptides from the protein mass and decreased the initial rate of β-casein degradation. The peptides appeared to be preferentially utilized during the first 4 days of ripening; thereafter rapid degradation of β-casein occurred. In quiescent slurries, α_s-casein degradation exceeded that of β-casein, whereas periodic agitation reversed the degradation rate of the two caseins. The rate and extent of characteristic flavor development appeared to be related directly to β-casein degradation, but not to changes in concentration of pare-K or α_s-caseins.     

Sodium caseinates — composition and properties of different preparations

A number of samples of commercially available spray-dried and roller-dried caseinates have been compared with freshly prepared caseinate from the laboratory. The compositions of the materials were measured using chromatography, and their calcium susceptibilities were determined using turbidity measurements. The chromatographic analysis showed that the commercial caseinates differed substantially from the fresh material, especially in the regions where α-casein and α_s2-caseins were eluted. The fresh caseinate was more affected by the presence of calcium ions than the others, and there were also differences between the individual spray-dried caseinates in this respect. The roller-dried caseinates were very insensitive to the presence of calcium ions.     

Characterisation of soybean glycinin and β-conglycinin fractionated by using MgCl2 instead of CaCl2

A simple two step precipitation method was used to investigate the effect of MgCl₂ instead of CaCl₂ on fractionation of soybean glycinin and β-conglycinin. Compositional and physicochemical properties of the resulting protein fractions were characterised. The optimised procedure, in terms of protein yield, purity, phytate content and physicochemical properties, was obtained when the addition of 5 m m MgCl₂ was used. After application of 5 m m MgCl₂, the phytate content of the glycinin-rich and β-conglycinin-rich fractions was about 0.4% and 1.3%, respectively, but the addition of 5 m m CaCl₂ increased the phytate content of the glycinin-rich fraction to 1.25% and decreased that of β-conglycinin-rich fraction to 0.67%. Low phytate protein product was suitable for use in infant formula and acidic food. The solubility of the glycinin-rich fractions with MgCl₂ was significantly higher than that with CaCl₂ at pH < 4.5. Application of MgCl₂ improved thermal stability of the β-conglycinin-rich fraction.     

Elucidation of structural changes in soy protein isolate upon heating by Raman spectroscopy

Changes in protein secondary and tertiary structure of soy protein isolate (SPI) upon heating process were investigated by Raman spectroscopy. SPI refrigerated and heated at 70, 100 and 200 °C was analysed. Non-significant differences in secondary structure (α-helix, β-sheet, unordered and turn) were observed in SPI upon heating at these temperatures. However, comparison of the Raman spectra of SPI refrigerated and heated at 70, 100 and 200 °C reveals changes in hydrophobic group environments. An intensity decreasing trend of the 1340 cm⁻¹ band attributable to tryptophan vibrations was observed, which involves an increase in solvent exposure of tryptophan residues. The spectral results also showed intensity increasing trends for the 1450 and 2935 cm⁻¹ bands of SPI upon heating, which can be ascribed to breaking of hydrophobic contacts and subsequent solvent exposure of the corresponding hydrophobic aliphatic groups. Finally, it was found tertiary structural transitions towards greater proportions of buried tyrosine residues upon heating of SPI.     

OPTIMIZING STABILITY OF ORANGE JUICE FORTIFIED WITH WHEY PROTEIN AT LOW pH VALUES

Abstract The influence of temperature, heating time and pH on the stability of whey protein-fortified Valencia orange juice was determined by uronic acid content, degree of esterification (DE), % transmission measurements (%T) and capillary electrophoretic analysis of the juice-protein supernatants. Uronic acid content and charge of pectins showed no significant change in heat-treated samples with added proteins. The %T decreased with decreasing pH and increasing temperature and heating time for α-lactalbumin (α-lac), β-lactoglobulin (β-lg) and whey protein isolate (WPI). The lowest transmission values were shown at pH 3.0 and 85C. Capillary electropherograms confirmed more extensive juice-protein interactions in WPI and β-lg added juices than in those containing α-lac, especially at low pH, resulting in more stable juice-protein mixtures.     

Probiotic Cheddar Cheese: Influence of Ripening Temperatures on Proteolysis and Sensory Characteristics of Cheddar Cheeses

ABSTRACT:  Bifidobacterium longum 1941, B. animalis subsp. lactis LAFTI^® B94, Lactobacillus casei 279, Lb. casei LAFTI L26, Lb. acidophilus 4962, or Lb. acidophilus LAFTI L10 were used as an adjunct in the production of Cheddar cheeses, which were ripened at 4 and 8 °C for 24 wk. Effects of ripening temperatures and probiotic adjuncts on proteolysis and sensory evaluation of the cheeses were examined. Higher ripening temperature increased the level of proteolysis in the cheeses. Product of proteolysis and organic acids released during ripening were shown to be important for the flavor of Cheddar cheeses. There were positive and significant correlations between the levels of soluble nitrogen, lactic, acetic, and butyric acids, percentage hydrolysis of α_s1-CN and β-CN to the scores of cheddary flavor ( P < 0.05). Scores for sour-acid and vinegary flavors were higher in cheeses with the addition of Bifidobacterium sp. or Lb. casei 279 ripened at 8 °C. The scores were positively and significantly correlated to the level of lactic, acetic, and free amino acids in the cheeses ( P < 0.05). The results show that both 4 and 8 °C have potential for use in the ripening of probiotic Cheddar cheeses.     

Effect of technological parameters on the characteristics of kasseri cheese made from raw or pasteurized ewes' milk

Two groups of kasseri cheese (pasta filata type) were manufactured from raw or pasteurized ewes' milk, without starter cultures. Cheeses of each group were divided into two subgroups: the first was ripened and stored at 4°C and packaged in plastic film; the second ripened and stored at 15°C and coated with paraffin wax. Milk pasteurization and technological parameters had a significant effect on the pH ( P  < 0.05), while only technological parameters had an effect on the total solids content. At day 120, the range of mean cfu/g counts for the mesophilic aerobic flora was 9.5 × 10⁷−1.4 × 10⁸; for the thermophilic streptococci, the range was 2.6 × 10⁷−7.6 × 10⁷; and for the thermophilic bacilli, 9.8 × 10⁶−1.7 × 10⁷. Changes in the N fractions became significant after 30 days of ripening. For mature 120-day-old cheeses, the percentage of total N soluble at pH 4.6 was 22.7%–22.9% in raw milk cheeses and 19.0%–21.7% in pasteurized milk cheeses. The percentage of total N soluble at 12% TCA was 10.1%–12.2% in raw milk cheeses and 7.3%–11.5% in pasteurized milk cheeses; the percentages of total N soluble at 5% PTA were 3.1%–4.0% and 2.6%–3.6%, respectively. The residual α_s-casein percentages at day 120 ranged between 63% and 78% of the respective area at day 1; the residual β-casein ranged between 67% and 75%. There were some characteristic differences in the reverse phase-HPLC peptide profiles of the four cheeses. In general, the effect of the different ripening conditions was more pronounced in cheeses made from pasteurized milk.     

Changes in the microbiological and chemical characteristics of an artisanal, low-fat cheese made from raw ovine milk during ripening

Changes in the microbial flora of batzos cheese made from raw ovine milk were studied during ripening. Lactic acid bacteria and Enterobacteriaceae were the predominant groups of micro-organisms. Cheeses manufactured in summer had higher microbial counts than those made in spring, with the exception of staphylococci. Nevertheless, Enterobacteriaceae and coliforms decreased more rapidly in cheese made in summer and counts at the end of storage were lower than those in spring cheese.
Enterococci predominated in the ripened curd of cheese made in spring, whereas lactobacilli were the most abundant lactic acid bacteria in cheese made in summer. Enterococcus faecium was the predominant species in spring, and Lactobacillus paracasei ssp. paracasei predominated in cheese made in summer. The pH of the cheeses was > 5.0 throughout ripening, and NaCl-in-moisture content (> 8.0%) permitted the growth and survival of salt-tolerant micro-organisms. α_s1-Casein degraded at a faster rate than β-casein; both caseins were hydrolysed more rapidly in spring than in summer. The free amino acid content became higher in summer cheese (566.24–3460.25 µg/g of glycine equivalent) than in spring cheese because of the progress of ripening. Moreover, the milk fat of the cheese was degraded more in the summer than in the spring. The results suggest that there could be advantages to using starter cultures and improving the level of hygiene during milk and cheese production in order to eliminate undesirable micro-organisms and standardize cheese quality.     

Natural preservative ingredient from marine alga Ulva lactuca L.

The contents of total chlorophyll (T-Chl), carotenoids and phenolics compounds were quantified in the biomasses of Ulva lactuca grown either in normal or artificial sea water under indoor conditions. The antioxidant and antibacterial activities of U. lactuca crude organic extracts ( Ulva- COEs) were determined. Thirty-four compounds in Ulva- COEs were characterised by thin layer chromatography and high-performance liquid chromatography. The major compounds were chlorophyll a (Chl a ) (15.60–30.90%) and b (Chl b ) (12.20–14.89%) , 9-cis β-carotene (13.12–14.47%), α-carotene (11.44–11.47%) and all-trans β-carotene (6.16–29.70%, of total carotenoids).The Ulva- COEs exhibited remarkable antioxidant activity, with an IC₅₀ (concentration which causes a 50% of DPPH radical scavenging activity) values ranged from 16.5 and 18.7 μg mL⁻¹, which could be compared with the synthetic antioxidants: α-tocopherol (14.4 μg mL⁻¹), butylated hydroxyanisol (13.1 μg mL⁻¹) and butylated hydroxyltoluene (13.1 μg mL⁻¹). Also, Ulva- COEs exhibited great potential antibacterial activities against six bacterial strains, with minimal inhibitory concentration values ranged from 0.40 to 0.35 mg mL⁻¹.