Use of Platelet Aggregometer to monitor the chymosin-initiated coagulation of casein micelles

1986 ◽  
Vol 53 (3) ◽  
pp. 359-370 ◽  
Author(s):  
Neal A. Bringe ◽  
John E. Kinsella
Keyword(s):  
Light Transmission ◽  
Average Rate ◽  
Enzyme Concentration ◽  
Casein Micelle ◽  
Clotting Time ◽  
Casein Micelles ◽  
Catalysed Reaction ◽  
High Enzyme ◽  
Lag Period ◽  
Limiting Value

SUMMARYThe chymosin-initiated coagulation of casein micelles was followed by monitoring light transmission using a Platelet Aggregometer. The release of macropeptide by chymosin was monitored using fluorescamine. The lag period in the clotting reaction was proportional to clotting time and the reciprocal of enzyme concentration. The average rate of coagulation, which was approximately equal to the reciprocal of clotting time (Tc), increased in proportion to enzyme concentration at low enzyme concentrations and reached a limiting value at high enzyme concentrations. The percentage hydrolysis at the Tc was 47 ± 5% in the presence of 20 mM-CaCl2 and it was calculated that a 5-fold decrease in the speed of the enzyme-catalysed reaction would decrease this value at the Tc to 43 ± 5%. The possible uses and limitations of the Platelet Aggregometer for determining the influence of the chemical environment on the velocity of the chymosin-catalysed reaction and para-casein micelle aggregatability are discussed.

Influence of calcium chloride on the chymosin-initiated coagulation of casein micelles

1986 ◽  
Vol 53 (3) ◽  
pp. 371-379 ◽  
Author(s):  
Neal A. Bringe ◽  
John E. Kinsella
Keyword(s):  
Calcium Chloride ◽  
Average Rate ◽  
Casein Micelle ◽  
Clotting Time ◽  
Casein Micelles ◽  
Casein Hydrolysis ◽  
Catalysed Reaction ◽  
Low Concentrations ◽  
Hydrolysis Of ◽  
Electrostatic Repulsions

SUMMARYCoagulation of para-casein micelles was monitored using a Platelet Aggregometer at 37·8 °C and pH 6·7. The macropeptide released by chymosin was determined quantitatively with fluorescamine. The aggregatability of para-casein micelles integrated over the complete hydrolysis of κ-casein was calculated from the amount of κ-casein hydrolysed during the clotting time (Tc). Low concentrations of CaCl2 enhanced the rate of κ-casein hydrolysis but increases in CaCl2 concentration above 8 mM caused marked decreases in this rate. Calcium chloride enhanced the ability of para-casein micelles to aggregate. Little aggregation of para-casein micelles occurred at 0·6 mM-CaCl2 even after all of the κ-casein was hydrolysed. As the concentration of CaCl2 was increased from 3 to 60 mM, aggregation of para-casein micelles took place at progressively lower levels of κ-casein hydrolysis and the percentage hydrolysis at the Tc decreased markedly from 71±7% to 26±8%. The combined influence of CaCl2 on the velocity of the chymosin-catalysed reaction and on para-casein micelle aggregatability accounted for its effect on the average rate of coagulation (calculated by the reciprocal of the Tc). Results are consistent with the hypothesis that electrostatic repulsions and ionic bonding are involved in the interaction between chymosin and κ-casein.

Derivation of a mathematical model for the mechanism of casein micelle coagulation by rennet

1981 ◽  
Vol 48 (2) ◽  
pp. 189-200 ◽  
Author(s):  
Donald F. Darling ◽  
Antoon C. M. Van Hooydonk
Keyword(s):  
Protein Concentration ◽  
Surface Protein ◽  
Enzyme Concentration ◽  
Lag Phase ◽  
Stability Factor ◽  
Casein Micelle ◽  
Clotting Time ◽  
Total Protein Concentration ◽  
Casein Micelles ◽  
The Stability

SummaryThe action of rennet on the stability of casein micelles has been described theoretically by combining Michaelis–Menten enzyme kinetics with von Smoluchow-ski's theory for slow coagulation. The casein micelle is considered to be stabilized by stcric interactions arising from a highly hydrated surface protein layer. The stability factor, in von Smoluchowski's theory, is then considered as a variable and determined by the concentration of unhydrolysed κ-casein.The theoretical approach describes semi-quantitatively experimental observations on clotting time (CT) as a function of temperature, rennet concentration, and total protein concentration. The model also predicts that, under certain circumstances, the CT is inversely proportional to the enzyme concentration. Furthermore, the model agrees with other workers' experimental observations on the effect of κ-cascin concentration, the presence of a lag phase, and that the lag phase is commonly about 60% of the CT.

Disruption and reassociation of casein micelles during high pressure treatment: influence of whey proteins

2007 ◽  
Vol 74 (2) ◽  
pp. 194-197 ◽  
Author(s):  
Thom Huppertz ◽  
Cornelis G de Kruif
Keyword(s):  
High Pressure ◽  
Treatment Time ◽  
Light Transmission ◽  
Whey Proteins ◽  
Casein Micelle ◽  
Casein Micelles ◽  
Β Lactoglobulin ◽  
Steric Stabilisation ◽  
Prolonged Holding

In the study presented in this article, the influence of added α-lactalbumin and β-lactoglobulin on the changes that occur in casein micelles at 250 and 300 MPa were investigated by in-situ measurement of light transmission. Light transmission of a serum protein-free casein micelle suspension initially increased with increasing treatment time, indicating disruption of micelles, but prolonged holding of micelles at high pressure partially reversed HP-induced increases in light transmission, suggesting reformation of micellar particles of colloidal dimensions. The presence of α-la and/or β-lg did not influence the rate and extent of micellar disruption and the rate and extent of reformation of casein particles. These data indicate that reformation of casein particles during prolonged HP treatment occurs as a result of a solvent-mediated association of the micellar fragments. During the final stages of reformation, κ-casein, with or without denatured whey proteins attached, associates on the surface of the reformed particle to provide steric stabilisation.

On the mechanism of milk clotting by rennin

1972 ◽  
Vol 39 (1) ◽  
pp. 55-63 ◽  
Author(s):  
Margaret L. Green
Keyword(s):  
Strong Evidence ◽  
Casein Micelle ◽  
Clotting Time ◽  
Casein Micelles ◽  
Milk Clotting ◽  
Milk Serum ◽  
Decisive Evidence ◽  
Subsequent Addition

SummaryTwo possible hypotheses for the mechanism of milk clotting were tested. The results obtained constituted strong evidence against one and suggested that the second is improbable. Milk was separated into a 5-fold-concentrated casein micelle suspension and milk serum. Pre-renneting of the serum did not reduce the rate of clotting on subsequent addition to the micelle suspension whether or not the conditions were such that the para-κ-casein became extensively aggregated. Washing of casein micelles up to 3 times with milk dialysate at 23°C extracted very little casein from the micelles and did not increase the clotting time of micelles resuspended to about the same concentration as in milk. The results appear to constitute decisive evidence against the hypothesis of milk clotting proposed by Parry & Carroll (1969). S-carboxymethyl-κ-casein, S-carboxymethyl-κ-casein containing 2·5 dimethylaminonaphthalene sulphonyl residues per mole, and rennin-treated dimethylaminonaphthalene sulphonated-S-carboxymethyl-κ-casein all bound Ca to the same extent at 30°C and pH 6·5, over the range 0·5–15·3 mM-CaCl2. This adds support to existing evidence that milk clotting does not involve formation of Ca bridges between casein micelles.

The effect of the chemical structure of additives on the coagulation of casein micelle suspensions by rennet

1980 ◽  
Vol 47 (3) ◽  
pp. 359-369 ◽  
Author(s):  
Richard J. Marshall ◽  
Margaret L. Green
Keyword(s):  
Molecular Weight ◽  
Chemical Structure ◽  
Hydrophobic Interactions ◽  
Sodium Dodecyl ◽  
Low Molecular Weight ◽  
Casein Micelle ◽  
Clotting Time ◽  
Casein Micelles ◽  
Hydrophobic Binding ◽  
Positively Charged

SummaryCasein micelles in milk-salts solution adsorbed charged detergents and highly-charged polypeptides strongly, neutral detergents less strongly and low molecular-weight amines weakly. A tetra-amine was adsorbed more strongly than a tri-amine. The extent of adsorption of proteins tended to rise as the molecular weight increased. Glycerol and lactate were adsorbed to a limited extent but dextran and α-ketoglutarate were not adsorbed at all. Proline was partly adsorbed, indicating that hydrophobic binding sites were available, and caused some disruption of the casein micelles. Additives were bound to approximately the same extent by casein micelles and rennet coagula. The proportions adsorbed were constant over at least 10-fold ranges of concentration. Additives which increased the rennet clotting time (RCT) acted by binding Ca2+. Most additives decreased the RCT, the extent increasing with the amount adsorbed and the positive charge on the additive. The greatest reduction in RCT was observed with those additives which had positively-charged and hydrophobic moieties and bound most strongly to casein micelles. Of the additives tested, only sodium dodecyl sulphate affected the enzymic action of rennet. The reduction in RCT may have resulted from the neutralization of the negative charge of the micelles or enhancement of their hydrophobicity, favouring hydrophobic interactions between the particles.

The acceleration by cationic materials of the coagulation of casein micelles by rennet

1977 ◽  
Vol 44 (3) ◽  
pp. 521-531 ◽  
Author(s):  
Margaret L. Green ◽  
R. J. Marshall
Keyword(s):  
Minimum Amount ◽  
Casein Micelle ◽  
Specific Interactions ◽  
Clotting Time ◽  
Coagulation Time ◽  
Casein Micelles ◽  
Milk Serum ◽  
Cationic Materials

SummaryThree cationic materials markedly reduced the rennet clotting time of casein micelle suspensions, the efficacy of each being primarily dependent on the charge and the amount absorbed by the micelles. The reduction in coagulation time was unaffected by components of the milk serum other than salts. No enzymic action by lysozyme on casein micelles was detected. All materials acted by the same mechanism, increasing the affinity of rennet for the micelles and accelerating the aggregation phase. Coagulation did not occur until a minimum amount of κ-casein had been hydrolysed to para-κ-casein. All additives increased the proportion of added rennet retained by the casein in the coagulum. The results indicated that coagulation occurs by specific interactions between micelles modified by rennet.

Hydration of casein micelles and caseinates: Implications for casein micelle structure

2017 ◽  
Vol 74 ◽  
pp. 1-11 ◽  
Author(s):  
Thom Huppertz ◽  
Inge Gazi ◽  
Hannemieke Luyten ◽  
Hans Nieuwenhuijse ◽  
Arno Alting ◽  
...  
Keyword(s):  
Casein Micelle ◽  
Casein Micelles ◽  
Micelle Structure

scholarly journals Association of casein micelle size and enzymatic curd strength and dry matter curd yield

2019 ◽  
Vol 49 (3) ◽  
Author(s):  
Denise Ribeiro de Freitas ◽  
Fernando Nogueira de Souza ◽  
Jamil Silvano de Oliveira ◽  
Diêgo dos Santos Ferreira ◽  
Cristiane Viana Guimarães Ladeira ◽  
...  
Keyword(s):  
Fixed Effects ◽  
Dry Matter ◽  
Linear Models ◽  
Favorable Effect ◽  
Casein Micelle ◽  
Casein Micelles ◽  
Micelle Size ◽  
Milk Protein Content ◽  
Bulk Tank ◽  
Cheese Production

ABSTRACT: The aim of the present study was to explore the association between milk protein content and casein micelle size and to examine the effects of casein micelle size on enzymatic curd strength and dry matter curd yield using reduced laboratory-scale cheese production. In this research, 140 bulk tank milk samples were collected at dairy farms. The traits were analyzed using two linear models, including only fixed effects. Smaller micelles were associated with higher κ-casein and lower αs-casein contents. The casein micellar size (in the absence of the αs-casein and κ-casein effects) did not affect the enzymatic curd strength; however, smaller casein micelles combined with higher fat, lactose, casein and κ-casein contents exhibited a favorable effect on the dry matter curd yield. Overall, results of the present study provide new insights into the importance of casein micelle size for optimizing cheese production.

scholarly journals Switches induced by quorum sensing in a model of enzyme-loaded microparticles

2018 ◽  
Vol 15 (140) ◽  
pp. 20170945 ◽  
Author(s):  
Tamás Bánsági ◽  
Annette F. Taylor
Keyword(s):  
Drug Delivery ◽  
Quorum Sensing ◽  
Cell Density ◽  
Group Size ◽  
Enzyme Concentration ◽  
Autocatalytic Reaction ◽  
Enzyme Loading ◽  
High Enzyme ◽  
Different Types ◽  
Hydrolysis Of

Quorum sensing refers to the ability of bacteria and other single-celled organisms to respond to changes in cell density or number with population-wide changes in behaviour. Here, simulations were performed to investigate quorum sensing in groups of diffusively coupled enzyme microparticles using a well-characterized autocatalytic reaction which raises the pH of the medium: hydrolysis of urea by urease. The enzyme urease is found in both plants and microorganisms, and has been widely exploited in engineering processes. We demonstrate how increases in group size can be used to achieve a sigmoidal switch in pH at high enzyme loading, oscillations in pH at intermediate enzyme loading and a bistable, hysteretic switch at low enzyme loading. Thus, quorum sensing can be exploited to obtain different types of response in the same system, depending on the enzyme concentration. The implications for microorganisms in colonies are discussed, and the results could help in the design of synthetic quorum sensing for biotechnology applications such as drug delivery.

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