CHEMISTRY OF MILK

Module 4. Enzymes in milk

Lesson 14
INTRODUCTION AND SIGNIFICANCE OF ENZYMES IN MILK

14.1 Introduction

Milk being a biological secretion of epithelial cells of the mammary gland has to undergo several biochemical reactions during its secretion. Consequently some of the enzymes will be entering the milk which have not been utilized during the biosynthesis of the milk. Similarly some enzymes get incorporated directly as a measure to protect the constituents after the production of milk. As such the enzymes naturally found in milk play a very significant role not only during the processing but also during their storage.

14.2 Enzymes in Milk

Approximately 50 enzymatic activities have been detected in bovine milk. Milk chemists are interested in quantitation of these enzymes, in deleterious or beneficial reactions that they catalyze in milk and dairy products, and in their inactivation. Catalysis of the enzyme is not similar for all enzyme some enzymes have a very fast activity Ex.Conversion up to 1,000,000 mols of reactant in one minute where as some enzymes catalyse only few hundred molecules of reactantper min in to the product. The rate of catalysis per molecule of enzyme is expressed as k _cat sometimes called the turnover number for the enzyme. The k _cat is a function of the efficiency of the enzyme and the chemistry of the reaction. Enzyme activity is frequently expressed in units. The Enzyme Commis­sion has defined a unit of enzyme as the quantity that will catalyze the transformation of one micromole of substrate to product(s) per minute under standard conditions. For comparative purposes and quantization, initial velocities are preferred. In order to employ the relation between reaction velocity and enzyme concentration to quan­titate the latter, three other parameters viz. substrate concentration, temper­ature, and pH-must be recognized and controlled. In general a plot of initial velocity against substrate concentration is a section of a rectangular hyper­bola. Such a relation is rationalized and explained on the basis of the well-known Michaelis-Menten equation

v_i =V _max[S]/(K_m+[S])

where vi =initial velocity of reaction, V_max = maximum velocity (or activity),K_m = Michael is constant, and [S] = substrate concentration.

In general, enzymes are active only over a limited range of pH, and usually a distinct pH optimum is observed. This may result from an effect of pH on the V max, on affinity of enzyme for substrate,or on the stability of the enzyme. The activity of many enzymes depends on particular ionized groups in the active site. Obviously, these will be influenced by even small variations in pH and Ionic strength. More extreme high or low pH may denature and hence inactivate the enzyme.

The overall effect of temperature on enzyme-catalyzed reactions is a resultant of the accelerating influence of temperature on the reaction itself and the thermal denaturation and inactivation of the enzyme. Thus, the rate of the catalyzed reaction passes through a maximum as it is examined at a series of increasing temperatures. The optimal temperature therefore, is generally higher when the time during which the reaction is measured is shorter. The optimum differs greatly among individual enzymes because they differ in susceptibility to thermal denaturation. It should be apparent from this brief Introduction: that measurement of enzyme activity must be performed under rigidly controlled conditions in which observed activity can be translated into activity with 100% of the enzyme in the ES form. Therefore, enzyme activity is a value equivalent to V max for the concentration of enzyme available, that is, [E] k_cat (The values given in represent such values.)

Enzymatic activities detected in bovine milk are listed in Table given below. They are named and classified according to the recommendations of the Enzyme Commission of the International Union of Biochemistry. All classes of enzymes except ligases have been detected in milk.

Some of the milk enzymes (e.g., catalase) are constituents of leukocytes, and some (e.g.,plasmin) may gain entrance to milk from blood. Most, how­ever, are constituents or products of the mammary cells that enter milk as rather benign accidental constituents during the secretory processes. Some (e.g. the galactosyl transferase component of lactose synthase) are con­stituents of Golgimembranes, others (e.g., alkaline phosphatase) of the cell membrane. Enzymes of microbial origin are not considered in this chapter. Individual milk enzymes are associated with casein micelles, fat globules, or leukocytes, or are dispersed in the serum. Locations are specified in the appendix, for cases in which they are known.

14.3 Activities of Some Enzymes in Bovine Milk

The enzymes in milk play a significant role by having an activity specific to that enzyme. The important enzymes along with their EC No, pH of optimum activity and temperature and the ratio of their activity in relation to the activity in bovine and human are presented in Table No. 14.1

Table 14.1 Activities of some enzymes in bovine milk

Concentrations of milk enzymes vary greatly among species, and within a species large variations occur among individual animals and during the course of lactation. Data on such variations for most of the milk enzymes are not very complete or definitive.

A few of the enzymes found in milk (e.g., lactose synthetase) have known functions in the biosynthetic processes of the mammary cells. Some milk enzymes act on substrates present as normal constituents of milk and may under suitable conditions play either beneficial or deleterious roles in dairy processes and products. Hydrolases, such as lipase and proteinase, may facilitate resorption of milk constituents if and when milking is stopped. There is no well-documented case of an enzyme in bovine milk beingof direct benefit to the primary consumer-the calf. It has been suggested,however, that a lipase in human milk facilitates digestion of fat by the human infant.