442 DAIRY CHEMISTRY AND BIOCHEMISTRY Ascorbic acid Reduction‖ Oxidation H,O Dehydroascorbic acid 2, 3.Diketogulonic acid HoH Hydrated hemiketal form Figure 11.2 Chemical structures of ascorbic acid and its derivatives. due mainly to the denaturation of B-lactoglobulin(and the consequent exposure of-SH groups)and loss of O2. Compounds formed by the Maillard reaction between lactose and proteins can also influence the eh of heated milk, particularly dried milk products Fermentation of lactose during the growth of micro-organisms in milk has a major effect on its redox potential. The decrease in the En of milk caused by the growth of lactic acid bacteria is shown in Figure 11. 3. A rapid decrease in Eh occurs after the available O, has been consumed by the bacteria. Therefore, the redox potential of cheese and fermented milk products is negative. Reduction of redox indicators (e.g. resazurin or
442 DAIRY CHEMISTRY AND BIOCHEMISTRY CHPOH I H-$-OH Ascorbic acid Rcduction Oxidation 11 CH2OH I H-C-OH 0 Hk2 Dehydroascorbic acid 11 Hzo CH20H I H-C-OH OH CHzOH %O I * H-c-OH I H-C-OH ,COOH 00 ‘c-c II II 2, SDiketogulonic acid Hydrated hemiketal form Figure 11.2 Chemical structures of ascorbic acid and its derivatives. due mainly to the denaturation of b-lactoglobulin (and the consequent exposure of -SH groups) and loss of 0,. Compounds formed by the Maillard reaction between lactose and proteins can also influence the E, of heated milk, particularly dried milk products. Fermentation of lactose during the growth of micro-organisms in milk has a major effect on its redox potential. The decrease in the E, of milk caused by the growth of lactic acid bacteria is shown in Figure 11.3. A rapid decrease in Eh occurs after the available 0, has been consumed by the bacteria. Therefore, the redox potential of cheese and fermented milk products is negative. Reduction of redox indicators (e.g. resazurin or
IYSICAL PROPERTIES OF MILK 443 Time(h) igure 11.3 Decrease in the redox potential of milk caused by the growth of Lactococcus lactis subsp. lactis at25°C. methylene blue)can be used as an index of the bacterial quality of milk by measuring the ' reduction time, at a suitable temperature, of milk containing Riboflavin absorbs light maximally at about 450 nm and in doing so be excited to a triplet state. This excited form of riboflavin can interact with triplet O2 to form a superoxide anion O2(or H2O2 at low pH). Excited iboflavin can also oxidize ascorbate, a number of amino acids and proteins nd orotic acid. Riboflavin- catalysed photo-oxidation results in the produc tion of a number of compounds, most notably methional(11. 1)which is the principal compound responsible for the off-favour in milk exposed to light Methional Photo-oxidation of milk constituents was discused in detail by Walstr and Jenness(1984) 11.4 Colligative properties of milk Colligative properties are those physical properties which are governed by the number, rather than the kind, of particles present in solution. The important colligative properties of milk are its freezing and boiling points (c.-0522 and 100.15.C, respectively) and its osmotic pressure( approxi
PHYSICAL PROPERTIES OF MILK 0.2 - 0.1 - 0.0 - -0.1 - -0.2 - 443 1 -0.31 . I ' I ' I ' I . I . I . I 0 1 2 3 4 5 6 7 Time (h) Figure 11.3 Decrease in the redox potential of milk caused by the growth of Lactococcus lactis subsp. lactis at 25°C. methylene blue) can be used as an index of the bacterial quality of milk by measuring the 'reduction time', at a suitable temperature, of milk containing the dye. Riboflavin absorbs light maximally at about 450nm and in doing so can be excited to a triplet state. This excited form of riboflavin can interact with triplet 0, to form a superoxide anion 0; (or H,O, at low pH). Excited riboflavin can also oxidize ascorbate, a number of amino acids and proteins and orotic acid. Riboflavin-catalysed photo-oxidation results in the production of a number of compounds, most notably methional(11.1) which is the principal compound responsible for the off-flavour in milk exposed to light. Methional Photo-oxidation of milk constituents was discused in detail by Walstra and Jenness (1984). 11.4 Colligative properties of milk Colligative properties are those physical properties which are governed by the number, rather than the kind, of particles present in solution. The important colligative properties of milk are its freezing and boiling points (c. -0.522 and 100.15"C, respectively) and its osmotic pressure (approxi-
DAIRY CHEMISTRY AND BIOCHEMISTRY mately 700kPa at 20C), all of which are interrelated. Since the osmotic pressure of milk remains essentially constant(because it is regulated by that of the cow's blood), the freezing point is also relatively constant The freezing point of an aqueous solution is governed by the concentra- tion of solutes in the solution. The relationship between the freezing point of a simple aqueous solution and concentration of solute is described by a relation based on Raoult s law: T=Ke (11.10) where T is the difference between the freezing point of the solution and that of the solvent, K is the molal depression constant( 1.86C for water) and m is the molal concentration of solute. However, this equation is valid only for dilute solutions containing undissociated solutes. Raoult's law is thus limited to approximating the freezing point of milk The freezing point of bovine milk is usually in the range -0.512 to 0.550C, with a mean value close to -0.522C( Sherbon, 1988)or 0.540C (Jenness and Patton, 1959). Despite variations in the conc tions of individual solutes, the freezing point depression of milk is quite constant since it is proportional to the osmotic pressure of milk(approxi mately 700kPa at 20 C), which is regulated by that of the cows blood. The freezing point of milk is more closely related to the osmotic pressure of mammary venous blood than to that of blood from the jugular vein. Owing to their large particle or molecular mass, fat globules, casein nicelles and whey proteins do not have a significant effect on the freezing point of milk, to which lactose makes the greatest contribution. The freezing nt depression in milk due to lactose alone has been calculated to be 0.296.C. Assuming a mean freezing point depression of 0.522 C, all other constituents in milk depress the freezing point by only 0. 226C. Chloride is also an important contributor to the colligative properties of milk. Assum ing a CI concentration of 0.032 M and that Cl- is accompanied by a monovalent cation (i.e. Na or K), the freezing point depression caused by CI" and its associated cation is 0. 119C. Therefore, lactose, chloride and its accompanying cations together account for about 80% of the freezing point depression in milk. Since the total osmotic pressure of milk is regulated by that of the cows blood there is a strong inverse correlation between lactose and chloride concentrations( Chapter 5) Natural variation in the osmotic pressure of milk(and hence freezing point)is limited by the physiology of the mammary gland. variations in the freezing point of milk have been attributed to seasonality, feed, stage of lactation, water intake, breed of cow, heat stress and time of day. These factors are often interrelated but have relatively little infuence on the freezing point of milk. Likewise, unit operations in dairy processing which do not influence the net number of osmotically active molecules/ ions in solution do not influence the freezing point, Cooling or heating milk causes
444 DAIRY CHEMISTRY AND BIOCHEMISTRY mately 700 kPa at 20"C), all of which are interrelated. Since the osmotic pressure of milk remains essentially constant (because it is regulated by that of the cow's blood), the freezing point is also relatively constant. The freezing point of an aqueous solution is governed by the concentration of solutes in the solution. The relationship between the freezing point of a simple aqueous solution and concentration of solute is described by a relation based on Raoult's law: Tf = K,m (11.10) where is the difference between the freezing point of the solution and that of the solvent, K, is the molal depression constant (136°C for water) and m is the molal concentration of solute. However, this equation is valid only for dilute solutions containing undissociated solutes. Raoult's law is thus limited to approximating the freezing point of milk. The freezing point of bovine milk is usually in the range -0.512 to -O.55O0C, with a mean value close to -02~22°C (Sherbon, 1988) or - 0.540"C (Jenness and Patton, 1959). Despite variations in the concentrations of individual solutes, the freezing point depression of milk is quite constant since it is proportional to the osmotic pressure of milk (approximately 700 kPa at 20"C), which is regulated by that of the cow's blood. The freezing point of milk is more closely related to the osmotic pressure of mammary venous blood than to that of blood from the jugular vein. Owing to their large particle or molecular mass, fat globules, casein micelles and whey proteins do not have a significant effect on the freezing point of milk, to which lactose makes the greatest contribution. The freezing point depression in milk due to lactose alone has been calculated to be 0.296"C. Assuming a mean freezing point depression of 0.522"C, all other constituents in milk depress the freezing point by only 0.226"C. Chloride is also an important contributor to the colligative properties of milk. Assuming a C1- concentration of 0.032M and that C1- is accompanied by a monovalent cation (i.e. Na' or K'), the freezing point depression caused by C1- and its associated cation is 0.119"C. Therefore, lactose, chloride and its accompanying cations together account for about 80% of the freezing point depression in milk. Since the total osmotic pressure of milk is regulated by that of the cow's blood, there is a strong inverse correlation between lactose and chloride concentrations (Chapter 5). Natural variation in the osmotic pressure of milk (and hence freezing point) is limited by the physiology of the mammary gland. Variations in the freezing point of milk have been attributed to seasonality, feed, stage of lactation, water intake, breed of cow, heat stress and time of day. These factors are often interrelated but have relatively little influence on the freezing point of milk. Likewise, unit operations in dairy processing which do not influence the net number of osmotically active molecules/ions in solution do not influence the freezing point. Cooling or heating milk causes
PHYSICAL PROPERTIES OF MILK transfer of salts to or from the colloidal state. However evidence for an effect of cooling or moderate heating(e.g. HTST pasteurization or minimum UhT processing)on the freezing point of milk is contradictory, perhaps ince such changes are slowly reversible over time. Direct UHT treatment involves the addition of water(through condensed steam). This additional water should be removed during fash cooling, which also removes gases, e.g. CO2, from the milk, causing a small increase in freezing point. Vacuum treatment of milk, i.e. vacreation(to remove taints), has been shown to increase its freezing point, presumably by degassing. However, if vacuum treatment is severe enough to cause a significant loss of water, the freezing CO2. Fermentation of milk has a large effect on its freezing point sina point will be reduced, thus compensating fully or partially for the loss fermentation of 1 mol lactose results in the formation of 4 mol lactic acid Likewise, fermentation of citrate influences the freezing point of milk Accurate measurement of the freezing point depression in milk requires great care. The principle used is to supercool the milk sample(by 1.0 to 1. C), to induce crystallization of ice, after which the temperature increases rapidly to the freezing point of the sample(Figure 11.4). For water, the temperature at the freezing point will remain constant until all the latent heat of fusion has been removed (i.e. until all the water is frozen ). However, for milk the temperature is stable at this maximum only momentarily and falls rapidly because ice crystallization causes concentration of solutes which eads to a further depression of freezing point. The observed freezing point of milk(maximum temperature after initiation of crystallization) is not the same as its true freezing point since some ice crystallization will have occurred before the maximum temperature is reached. Correction factors have been suggested to account for this but, in practice, it is usual to report T(°C) Observed freez point of milk sample 0.522 -1.5 Induction of crystallization Figure 11.4 Temperature-time curve for the freezing of milk
PHYSICAL PROPERTIES OF MILK 445 transfer of sdts to or from the colloidal state. However, evidence for an effect of cooling or moderate heating (e.g. HTST pasteurization or minimum UHT processing) on the freezing point of milk is contradictory, perhaps since such changes are slowly reversible over time. Direct UHT treatment involves the addition of water (through condensed steam). This additional water should be removed during flash cooling, which also removes gases, e.g. CO,, from the milk, causing a small increase in freezing point. Vacuum treatment of milk, i.e. vacreation (to remove taints), has been shown to increase its freezing point, presumably by degassing. However, if vacuum treatment is severe enough to cause a significant loss of water, the freezing point will be reduced, thus compensating fully or partially for the loss of CO,. Fermentation of milk has a large effect on its freezing point since fermentation of 1 mol lactose results in the formation of 4 mol lactic acid. Likewise, fermentation of citrate influences the freezing point of milk. Accurate measurement of the freezing point depression in milk requires great care. The principle used is to supercool the milk sample (by 1.0 to 1.2"C), to induce crystallization of ice, after which the temperature increases rapidly to the freezing point of the sample (Figure 11.4). For water, the temperature at the freezing point will remain constant until all the latent heat of fusion has been removed (i.e. until all the water is frozen). However, for milk the temperature is stable at this maximum only momentarily and falls rapidly because ice crystallization causes concentration of solutes which leads to a further depression of freezing point. The observed freezing point of milk (maximum temperature after initiation of crystallization) is not the same as its true freezing point since some ice crystallization will have occurred before the maximum temperature is reached. Correction factors have been suggested to account for this but, in practice, it is usual to report M v) -1.5 -. Induction of crystallization Time Figure 11.4 Temperature-time curve for the freezing of milk