Advances in Analysis of Milk Proteases Activity at Surfaces and in a Volume by Acoustic Methods

This review is focused on the application of surface and volume-sensitive acoustic methods for the detection of milk proteases such as trypsin and plasmin. While trypsin is an important protein of human milk, plasmin is a protease that plays an important role in the quality of bovine, sheep and goat milks. The increased activity of plasmin can cause an extensive cleavage of β-casein and, thus, affect the milk gelation and taste. The basic principles of surface-sensitive acoustic methods, as well as high-resolution ultrasonic spectroscopy (HR-US), are presented. The current state-of-the-art examples of the application of acoustic sensors for protease detection in real time are discussed. The application of the HR-US method for studying the kinetics of the enzyme reaction is demonstrated. The sensitivity of the acoustics biosensors and HR-US methods for protease detection are compared.

STUDY OF THE ENZYMATIC STAGE OF MILK GELATION: CHANGES IN VISCOSITY AND MICROSTRUCTURE

The article presents the results of experimental joint studies of changes in the viscosity and microstructure of milk at the enzymatic stage of gelation. Based on the statistical processing of the array of research results, it was determined that the viscosity change at this stage is not monotonic, as it is usually stated, but two-stage in the middle part and S-shaped, preceding the gel point, at its end. It was found that the S-shaped change in viscosity at the end of the enzymatic stage of milk coagulation coincides with changes in the microstructure of casein micelles and reflects the existence of a cooperative conformational phase transition in casein molecules of micelle clusters. A description of the possible mechanism of this phase transition is proposed. It was noted that the moment of the S-shaped change in the milk viscosity at the enzymatic gelation stage and the corresponding cooperative phase transition in casein micelles are a physical reflection of the gel point. The research results provide a better understanding of the mechanism of enzymatic coagulation of milk in a cheesemaking tank.

Measurement of milk clotting activity by rotational viscometry

Standard method for the determination of the activity of milk coagulants is the rotating bottle method, where clotting time is defined as the time when visually observable flocculation starts. Aim of this study was to verify whether it is possible to determine milk clotting time by rotational viscometry. Using three different coagulants and reconstituted milk of different pH and temperature, flocculation time and viscosity in steady shear was determined, and milk gelation was monitored by small amplitude oscillating shear rheometry. The results show that, independent of pH and temperature, milk clotting time is related to an apparent viscosity of 7·24±0·45 mPa.s, indicating that rotational viscometry can be used for the screening of flocculation time with an accuracy of approximately 6%.

Gelation of casein-whey mixtures: effects of heating whey proteins alone or in the presence of casein micelles

The aim of the present work was to investigate the role of whey protein denaturation on the acid induced gelation of casein. This was studied by determining the effect of whey protein denaturation both in the presence and absence of casein micelles. The study showed that milk gelation kinetics and gel properties are greatly influenced by the heat treatment sequence. When the whey proteins are denatured separately and subsequently added to casein micelles, acid-induced gelation occurs more rapidly and leads to gels with a more particulated microstructure than gels made from co-heated systems. The gels resulting from heat-treatment of a mixture of pre-denatured whey protein with casein micelles are heterogeneous in nature due to particulates formed from casein micelles which are complexed with denatured whey proteins and also from separate whey protein aggregates. Whey proteins thus offer an opportunity not only to control casein gelation but also to control the level of syneresis, which can occur.