1. Enzymes in Dairy Products

3. The concentration of proteins in milk grams/ liter % of total protein _______________________________________________________ Total Protein 33 100 Total Caseins 26 79.5 alpha s1 10 30.6 alpha s2 2.6 8.0 beta 9.3 28.3 kappa 3.3 10.1 Total Whey Proteins 6.3 19.3 alpha lactalbumin 1.2 3.7 beta lactoglobulin 3.2 9.8 BSA 0.4 1.2 Immunoglobulins 0.7 2.1 Proteose peptone 0.8 2.4 _______________________________________________________

4. Constituents in milk In solution, 87% water, dissolved is the milk sugar (lactose), four water - soluble vitamins (thiamine, riboflavin, niacin and ascorbic acid). Minerals including chlorides, citrates, potassium, magnesium and sodium ions as well as calcium phosphate are in solution.

5. In colloidal dispersion, ● Dispersed in the water, but not in true solution, are calcium phosphate and protein, both in colloidal form. These colloids are responsible for the opaqueness of milk. ● The proteins in milk – casein and whey (serum) proteins. ● Casein accounts for approximately 80 % of the proteins in milk. ● Casein separated into three major fractions: alpha-casein (slightly more than ½), β-casein (slightly less than 1/3), and κ-casein (approximately 15% of the total complex known as casein). ● All three types are organized in a complex with Ca++ ions to form aggregates known as “micelles.” ● Casein molecules are phosphoproteins – therefore, these colloidal particles have been referred to as calcium phosphocaseinate.

6. A colloid is a type of chemical mixture where one substance is dispersed evenly throughout another. The particles of the dispersed substance are only suspended in the mixture, unlike a solution, where they are completely dissolved within. This occurs because the particles in a colloid are larger than in a solution - small enough to be dispersed evenly and maintain a homogenous appearance, but large enough to scatter light and not dissolve. Because of this dispersal, some colloids have the appearance of solutions. A colloidal system consists of two separate phases: a dispersed phase (or internal phase) and a continuous phase (or dispersion medium). A colloidal system may be solid, liquid, or gaseous.mixturesolution homogenouslight solutionssolidliquid gaseous

7. MilkMilk is an emulsified colloid of liquid butterfat globules dispersed within a water-based fluid.emulsified butterfat water

8. In chemistry, a solution is a homogeneous mixture composed of two or more substances. In such a mixture, a solute is dissolved in another substance, known as a solvent. Gases may dissolve in liquids, for example, carbon dioxide or oxygen in water. Liquids may dissolve in other liquids. Gases can combine with other gases to form mixtures, rather than solutions. All solutions are characterized by interactions between the solvent phase and solute molecules or ions that result in a net decrease in free energy.chemistry homogeneousmixturesolutesolvent Gasesdissolvecarbon dioxideoxygen

9. An emulsion is a mixture of two or more immiscible (unblendable) liquids. One liquid (the dispersed phase) is dispersed in the other (the continuous phase). Many emulsions are oil/water emulsions, with dietary fats being one common type of oil encountered in everyday life. Examples of emulsions include butter and margarine, milk and cream, and vinaigrettes. In butter and margarine, fat surrounds droplets of water (a water-in-oil emulsion). In milk and cream, water surrounds droplets of fat (an oil-in- water emulsion).immisciblephasedispersedphasebuttermargarinemilk vinaigrettes

10. A. Two immiscible liquids, not yet emulsified; B. An emulsion of Phase II dispersed in Phase I; C. The unstable emulsion progressively separates; D. The surfactant (purple outline) positions itself on the interfaces between Phase II and Phase I, stabilizing the emulsion

11. ● The micelles of casein are dispersed throughout the milk and are the main source of its whiteness. ● Both α-casein and β-casein are sensitive to calcium and precipitated by the calcium ions in milk if κ-casein is not present in the complex. κ-casein renders the complex stable. ● Whey proteins roughly 20% of the total proteins in milk contain two major fractions: the globulins, the albumins. In emulsion, The fat in milk as small droplets or globules, 3 to 6 microns, may be up to 10, depending on the breed of cow. Fat droplets in milk are prevented from coalescing by a thin coating of emulsifier (a few millimicrons thick) around the fat globules at the liquid-fat interface. Diagram of the structure of the fat globule membrane in milk is on the next slide.

12. Triglyceride, fatty acid chains intermeshed with non-polar prongs of the phospholipid molecules. Molecules of vitamin A and of cholesterol are interspersed among the molecules of phospholipids. Hydrophilic group of phospholipid are oriented toward hydrophilic side chain of the first protein layer. Hydrophobic R groups oriented away from the fat globule. Hydrophobic side chain of second protein layer are pointed inward and the hydrophilic ones pointed outward.

13. Agglutinin promotes clustering. Rise top of the milk. Cluster of fat droplets, creaming. Fat droplets contribute viscosity to milk and cream: low viscosity of skim milk. In goat’s milk the fat globules are so small that they are unable to float to the top of the milk and cream.

14. To eliminate the creaming of milk, it is homogenized. Milk is pasteurized and forced, under pressure, through fine orifices. This reduces the fat globules to an average diameter of 1 micron. Increased surface of the fat globules in the homogenized milk and spreading of the casein on this surface make whiter and more viscous than unhomogenized milk with the same fat content. The sense of richness of the homogenized milk is associated with its greater viscosity.

15. I. Natural enzymes in milk Enzymes are a group of proteins that have the ability to catalyze chemical reactions and the speed of such reactions. The action of enzymes is very specific. Milk contains both indigenous and exogenous enzymes. Exogenous enzymes mainly consist of heat-stable enzymes produced by psychrotrophic bacteria: lipases, and proteinases. There are many indigenous enzymes that have been isolated from milk. The most significant group are the hydrolases: Indigenous enzymes lipoprotein lipase plasmin alkaline phosphatase

16. Lipoprotein lipase (LPL): A lipase enzyme splits fats into glycerol and free fatty acids. This enzyme is found mainly in the plasma in association with casein micelles. The milkfat is protected from its action by the FGM. If the FGM has been damaged, or if certain cofactors (blood serum lipoproteins) are present, the LPL is able to attack the lipoproteins of the FGM (fat globule membrane) Plasmin: Plasmin is a proteolytic enzyme; it splits proteins. Plasmin attacks both β-casein and α(s2)-casein. It is very heat stable and responsible for the development of bitterness in pasteurized milk and UHT processed milk. It may also play a role in the ripening and flavour development of certain cheeses, such as Swiss cheese.

17. Alkaline phosphatase: Phosphatase enzymes are able to split specific phosporic acid esters into phosphoric acid and the related alcohols. Unlike most milk enzymes, it has a pH and temperature optima differing from physiological values; pH of 9.8. The enzyme is destroyed by minimum pasteurization temperatures, therefore, a phosphatase test can be done to ensure proper pasteurization. Phosphoric acids and phosphate ester on the next slides.

18. Phosphoric acids

19. II. Milk Clotting A. Enzymatic Milk Coagulation ● In coagulation of milk to make cheese. ● Most proteolytic enzymes clot milk. ● Not all make acceptable cheese. ● The Food and Nutrition Board of the United States National Research Council describes “rennet” as all milk-clotting enzyme preparations (except porcine pepsin) used for cheese making. ● Rennet is defined as “aqueous extracts made from the fourth stomachs of calves, kids, or lambs.” (kid: young goat) ● Bovine rennet is defined as “aqueous extract made from the fourth stomach of bovine animals, sheep, or goats.” Bovine: any of ruminant (even-toed, hoofed, herbivorous having a stomach with four cavities, cattle, deer, sheep, goats, giraffes, camels)

20. ● Microbial rennet is defined as “followed by the name of the organism” is the approved nomenclature for milk-clotting preparations derived from microorganisms. ● US Standards of Identity for Cheddar cheese allow the use of “rennet and/or other clotting enzymes of animal, plant, or microbial origin.” ● Milk-clotting enzymes generally recognized as safe (GRAS) are rennet and bovine rennet. ● Microbial rennets derived from Cryphonectria parasitica (formerly Endothia parasitica), Bacillus cereus, Mucor pusillus var. Lindt, and Mucor miehei var. Cooney et Emerson may be used as secondary direct food additives.

21. ● Chymosin from the abomasa of suckling calves remains the enzyme of choice and the standard against which all others are evaluated. ● Genetic engineering has been employed to produce chymosin microbially in Escherichia coli and in Saccharomyces cerevisiae. ● No significant difference between the two cheese prepared with recombinant chymosin and calf rennet. ● Recombinant enzymes are now approved for commercial use. ● Milk-clotting activity is present as chymosin in the abomasum of a bovine fetus by the sixth month of development and increases in potency as the fetus approaches full term.

22. ● At birth, chymosin is present in gastric mucosa at 2-3 mg/g but its production declines after 1 week. ● Liquid rennet preparations have a pH between 5.6 and 5.8 to provide the most stable environment. Bacterial content can be reduced by filtration or centrifugation. ● Porcine pepsin is used as a substitute for part of the rennet in making varieties of cheese. This enzyme is secreted by hog stomach mucosa as pepsinogen (40,400 Daltons).

23. ● The pH of cheese milk is such that both the activity and stability of porcine pepsin are far from optimum (pH 2.0). Slow coagulation and a weak set resulting in excessive fat losses and reduced yield occur if insufficient enzyme is used. Porcine pepsin is inexpensive relative to chymosin. Mixtures of porcine pepsin and bovine rennet are used, typically containing 20-25% chymosin, 40-45% bovine pepsin, and 30-40% porcine pepsin activity. ● Fungal proteases are used extensively as substitute for chymosin in milk clotting. Mucor miehei rennet is the most common fungal milk- clotting preparation. It is also most heat stable of all the commonly used milk-clotting enzymes.

24. ● Mucor miehei rennet remains active in the whey and is concentrated in condensed whey products, causing problems when the whey products are mixed with casein. Therefore, Mucor miehei rennet used by cheese industry is now modified to decrease its heat stability. ● Mucor pusillus var. Lindt protease has been used as a chymosin substitute and not all strains are capable of producing acceptable cheese.

26. 1. Colloidal Dispersion ● Dispersed in the water, but not in true solution, are calcium phosphate and protein, both in colloidal form. These colloids are responsible for the opaqueness of milk. ● The proteins in milk – casein and whey (serum) proteins. ● Casein accounts for approximately 80 % of the proteins in milk. ● Casein separated into three major fractions: alpha-casein (slightly more than ½), β-casein (slightly less than 1/3), and κ-casein (approximately 15% of the total complex known as casein). B. Milk Coagulation Chemistry

27. ● All three types are organized in a complex with Ca ++ ions to form aggregates known as “micelles.” ● Casein molecules are phosphoproteins – therefore, these colloidal particles have been referred to as calcium phosphocaseinate. ● C asein proteins exist in a colloidal particle known as the casein micelle. Its biological function is to carry large amounts of highly insoluble CaP to mammalian young in liquid form and to form a clot in the stomach for more efficient nutrition. ● The micelles of casein are dispersed throughout the milk and are the main source of its whiteness. ● Both alpha-casein and β-casein are sensitive to calcium and precipitated by the calcium ions in milk if κ-casein is not present in the complex. κ-casein renders the complex stable.

30. Old model for casein micelle (20 years old) CMP: caseinomacropeptide

31. New model for casein micelle

32. ● Colloidal calcium phosphate (CCP): Acts as a cement between the hundreds or even thousands of submicelles that form the casein micelle. ● Role of Ca ++ : More than 90% of the calcium content of skim milk is associated in some way or another with the casein micelle. The removal of Ca ++ leads to reversible dissociation of β-casein without micellular disintegration. The addition of Ca ++ leads to aggregation. ● pH: Lowering the pH leads to dissoLution of calcium pHosphate until, At the isoelec4ric poiNt (pH 4.6), all phosphate is dissolv%d and the caseins precipitate.

33. ● Casein micelle aggregation Caseins are able to aggregate if the surface of the micelle is reactive The Schmidt model further illustrates this. ● Although the casein micelle is fairly stable, there are four major ways in which aggregation can be induced: 1. chymosin - rennet or other proteolytic enzymes as in Cheese manufacturingCheese 2. acid 3. heat 4. age gelation

34. Age Gelation Age gelation is an aggregation phenomenon that affects shelf-stable, sterilized dairy products, such as concentrated milk and UHT milk products. After weeks to months storage of these products, there is a sudden sharp increase in viscosity accompanied by visible gelation and irreversible aggregation of the micelles into long chains forming a three- dimensional network. The actual cause and mechanism is not yet clear, however, some theories exist: Proteolytic breakdown of the casein: bacterial or native plasmin enzymes that are resistant to heat treatment may lead to the formation of a gel Chemical reactions: polymerization of casein and whey proteins due to Maillard type or other chemical reactions Formation of kappa-casein-ß -lactoglobulin complexes

35. 2. Effect of rennet on casein ● Milk clotting begins with enzymatic cleavage of κ-casein, destroying the ability of κ-casein to stabilize casein micelles. Primary action of rennet on casein is specific, cleaving the peptide bond between phenylalanine and methionine in the k-casein. k-casein para-k-casein + macropeptide ● This cleavage is followed by non-enzymatic aggregation of the altered casein micelles. Ca -- acting as a bridge between micelles

36. ● Then aggregates of casein micelles form a firm gel structure. Under normal cheese making conditions, aggregation of micelles begins at the same time as enzymatic cleavage of k-casein. ● Curd structure tightens and syneresis occurs. ● Cheese starter cultures lower the pH before milk-clotting enzyme is added and continue to lower the pH during setting of the curd. ● Both pH and temperature affect milk coagulation. Each milk-clotting enzyme has an optimal pH that must be considered in balance with culture activity to attain the correct pH at setting and cutting of the curds. ● With an increase in temperature up to 38-40 ℃, rate of rennet reaction increases.

37. Cheese Manufacture The most popular cheese in the US, Canada and England is Cheddar cheese. All cheese types begin with curdmaking and then involve various manipulations of the curd or whey. When the casein coagulates, the curd traps much of the fat, some of the lactose, and some of the water and minerals in the coagulant. The remaining liquid and its dissolved lactose, proteins, minerals, and other minor constituents are the whey. Cheese curd can be made from raw or pasteurized milk. Raw milk cheese should be ripened for 60 days or more as a safeguard against pathogens. Storage conditions under the acid conditions of the cheese destroys the common disease-producing organisms that could be present in the milk. Most of cheddar cheese is made from pasteurized milk. Pasteurization destroys most spoilage organisms and undesirable enzymes and gives better control over subsequent fermentation of the curd.

38. Cheddar Cheese 1. Setting the Milk Whole milk is added to a vat and brought to about 31 ℃. A lactic acid –producing starter culture of Streptococcus lactis is added. Orange color (Annatto) may be added at this point to the stirring milk. After about 30 min, a mildly acidic condition of about 0.2% acidity develops, and then rennin in the form of diluted solution is added. The commercial rennin preparation is known as rennet. Rennin (also called chymosin) refers to the pure enzyme. Commercial rennet is obtained from the fourth stomach of the calf.

39. 2. Cutting the Curd Curd knives are made up of wires strung across a frame (a harplike wire knife). One knife has the wires going vertically and the other horizontally. By drawing the knives through the length of the vat and then back and forth with the width of the vat, the curd is cut into small cubes. In the case of Cheddar, these are 0.25-0.5 inches on a side. The curd for different types of cheese, therefore, is cut into different size cubes: the smaller the cubes, the greater the surface area, and so the quicker and more complete is the removal of whey from the cubes, which can lead to a drier cheese.

40. 3. Cooking The cubes are gently agitated and jacketed vat is heated with steam to raise the temperature of the curd and whey to about 38 ℃ over a 30-min period and held for about 45 min or longer. Heat increases the rate of acid production and makes the curd cubes shrink. Both help expel the whey and toughen the curd cubes. During the cooking the curds continue to be gently agitated. Rennet and lactic acid bacteria entrapped in the curds (enzyme activities from rennet and the peptidase from the bacteria continue). ● With an increase in temperature up to 38-40 ℃, rate of rennet reaction increases.

41. 4. Draining Whey and Matting Curd. Agitation of the curds is stopped and they are permitted to settle. The whey is drained from the cheese vat and curds are trenched along the sides of the vat to further facilitate whey drainage. After all of the whey has been drained, the curds are allowed to mat for about 15 min. During matting the individual curd pieces fuse together to form a continuous rubbery slab.

42. 5. Cheddaring This step involves cutting the matted curd into blocks, turning the blocks at 15-min intervals, and then piling the blocks on one another two or three deep. Purposes of cheddaring are to allow acid formation to continue and to squeeze whey from the curd. The weight of the blocks on one another is a mild form of pressure. During cheddaring the vat is maintained warm. Stacking and turning the blocks goes on for about 2 hr or until the whey coming from the blocks reaches 0.5-0.6% acid.

43. 6. Milling and Salting The rubbery slabs of cheddared curd are passed through a mill that cuts the blocks into small pieces. The milled pieces are spread out over the floor of the vat and sprinkled with salt (2.5% of curd weight). The salt and curd are stirred to uniformly distribute the salt. Purposes of salt: 1) to further draw whey out of the curd by osmosis, 2) to inhibit proteolytic and other types of spoilage organisms that might otherwise grow in later stages of the cheesemaking operation, 3) to add flavor to the final cheese.

44. 7. Pressing (Hooping) The milled and salted curd pieces are placed in hoops fitted with cheesecloth and the hoop are placed in a hydraulic press at about 1.4 x 10 5 Pa (20 psi) overnight. The more moisture or whey retained in the cheese from the press, the more acidity that can be fermented from it. This in turn affects the final texture of the cheese and what organisms can grow during the subsequent ripening period. 8. Curing or Ripening Cheese is removed from hoop and placed in a cool room at about 16 ℃ and 60% RH for 3 or 4 days. This causes mild surface drying and forms a slight rind. The cheese block is dipped in hot paraffin to prevent mold growth on the surface and from excessive drying out during the long ageing period.

45. The waxed cheese is boxed and placed in the curing room for ripening at about 4 ℃ and 85% RH at least 60 days whether the cheese milk was raw or pasteurized. Cheddar types: 4 - 10 ℃, 8-10 ℃ is the recommended range. It is desirable to initiate ripening for several weeks at 4-6 ℃ and then increase the temperature to 8 - 10 ℃. Low temperature initially, minimizes early growth of starter and non-starter bacteria and reduces the risk of too rapid ripening and off flavour development. It also minimizes the risk of the minimum pH reaching levels below 5.0. Most European varieties are stored at 10 - 15 ℃ for initial ripening and then 4 ℃ until consumed. For peak flavor, ripening may be continued for 12 months or longer.

46. During this period bacteria in the cheese and enzymes in the rennet preparation modify the cheese texture, flavor, and color by continuing to ferment residual lactose and other organic compounds into acids and aroma compounds, by partial hydrolysis of the milk fat and further breakdown of fatty acids, and mild proteolysis of the protein. Rennet and lactic acid bacteria entrapped in the curds (enzyme activities from rennet and the peptidase from the bacteria continue).

47. Advanced Processes Recent advances utilize reverse osmosis and ultrafiltration. These treatments concentrate milk solids for efficient further processing. Lactoglobulin and lactalbumin can be retained with the cheese solids rather than being lost to the whey, improving cheese yield and nutritional values. Lactose levels can be reduced when less acidity is desired.

48. The automated ultrafiltration system shows numerous membrane cartridges connected in series and provided with recirculation loops for progressive separation and concentration of milk solids.

49. Cottage Cheese A low-fat, soft cheese, generally coagulated with lactic acid bacteria rather than rennin. Curd is not pressed, not aged or ripened under long storage. 1. Pasteurized skim milk is warmed in a vat to 22 ℃. 2. A lactic starter (at about 1% level) is added to produce acid. In addition to S. lactis, the starter usually contains Leuconostoc citrovorum, a flavor-producing bacterium. 3. The vat is set and fermented for about 14 hr (long-set method). 4. The coagulated milk is cut into small cubes. 5. The curd cubes are cooked for about 90 min with stirring, and the temperature is gradually increased to 50 ℃.

50. 6. The whey is drained and the curd is washed with cold water to remove excess whey and limit acidity. 7. The curd is trenched to drain all water. 8. The curd may now mildly salted, as a mild preservative measure and for flavor. 9. The curd also may be blended with sweet or soured cream to give 4% fat. The product is then called creamed cottage cheese. 10. Packaged as loose curd particles and undergoes no further processing. Short-set method: a larger amount of lactic starter (about 6%) and a temperature of 32°C develop the proper degree of acidity for coagulation and curd cutting within 5 hr. Another variation employs low levels of rennet plus the starter for milk coagulation.

51. Swiss Cheese 1. To the raw milk in large kettle is added a multiple-organism starter, Lactobacillus bulgaricus, heat-tolerant Streptococcus thermophilus, and eye-forming Propionibacterium or this may come in with the raw milk. 2. Following an initial period of lactic acid fermentation, rennet is added to the kettle to coagulate the milk. 3. The curd is cut with a harplike wire knife into rice size particles. 4. The curds and whey are now heated and cooked at about 53 ℃ for about an hour.

52. 5. At this point, unlike Cheddar-making, the stirred heated curd is allowed to settle. 6. A cloth with a fitted steel strip edge is slid under the curd, and the entire curd mass is hoisted from the kettle to drain.

53. 7. The entire curd from a kettle is placed in a single large hoop in which it is pressed for 1 day to form a beginning rind. 8. The cheese is removed from the hoop and placed in a large brine tank at about 10 ℃ to float for about 3 days. Its top is periodically salted. 9. The cheese is now moved to a warm ripening room maintained at about 21 ℃ and 85% RH and remained here for about 5 weeks during which time eyes are formed by the fermentation of Propionibacterium.

54. Propionibacterium Lactic acid Propionic acid + CO 2 CH 3 CHOHCOOH (nutty flavor) (eye form) CH 3 CH 2 COOH Propionibacter produce CO 2, which is not able to escape, so it concentrates in various places in the cheese. The pressure produced by this gas results in the eyes in Swiss cheese. 10. After about weeks, the cheese is moved to a colder curing room at about 7 ℃ and remained from 4 to 12 months to develop the full sweet nutty flavor. Blue-Veined Cheese Four well-known varieties of blue-veined cheeses - Blue cheese: Denmark, USA, and other countries (cow’s milk) Stilton: England (cow’s milk) Gorgonzola: Italy (cow’s milk) Roquefort: France (sheep’s milk)

55. Blue-green mold Penicillium roqueforti is added prior to being hooped and pressed. Ripening period for 3 to 10 months at cool, moist, cavelike conditions of about 4 ℃ and 90% RH. To encourage the mold growth throughout the cheese mass during ripening period, the pressed cheese is pierced. This allows the air to penetrate the cheese and support mold growth throughout mass. Penicillium roqueforti not only produces the mottled blue color but is an active splitter of milk fat. This gives rise to fatty acids and ketones, which contribute to the sharp, peppery flavor of blue-veined cheese.

56. Camembert This cheese is characterized by a soft cream-colored curd and a white mold growth that covers its entire surface. The mold is Penicillium camemberti. The mold is inoculated onto the pressed cheese curd after removal from the hoop by spraying a mist of mold spores onto the cheese surfaces. Ripening is under damp conditions at about 7 ℃ and 95% RH for about 3 weeks.

57. P. camemberti is highly proteolytic and breaks down the curd protein, from the surface inward, to the texture of soft butter. If proteolysis goes too far, because of prolonged storage in the supermarket or the home, the cheese develop a strong ammonia odor. Limburger After milk is coagulated, the curd is packed into rectangular forms and residual whey is allowed to drain. When the cheese is firm enough to retain its shape, it is removed from the form and salted and turned frequently. Major ripening agent is a surface bacterium called Brevibacterium linens. Limburger is a semisoft cheese like Camembert, which is ripened from the surface inward with a characteristic decomposition. It is thought that proteolytic enzymes secreted by the organism diffuse into the cheese and cause the characteristic softening and flavor by proteolytic decomposition..

58. Processed Cheese Mixing or grinding different lots of natural cheeses together and then melting them into a uniform mass. Different lots of natural cheese varying in moisture, acidity, texture, flavor, age, and other characteristics: e.g. a highly acidic cheese blended with a bland cheese for a totally acceptable product. The best known process cheese is the popular American cheese made from blended and melted Cheddar. Labeling for the cheese may be “Process Cheese Food” or “Process Cheese Spread” if minimum federal standards are met for no less than 23% fat and no more than 44% moisture.

59. 1. The mixed lots are melted together by heating to about 71 ℃. This also pasteurizes the cheese. 2. Emulsifiers such as sodium citrate and disodium phosphate are added to prevent fat separation, and to add smoothness to the texture. 3. The hot melted cheese is then filled into cartons and allowed to cool and solidify.

60. American cheese being extruded and sliced.

61. Other Related Products Sweet Milk Milk is coagulated with rennet to a custard-like consistency plus flavor. Cultured Buttermilk Skim milk (or partially-skimmed milk) is mildly coagulated with lactic acid culture (containing Leuconostoc bacteria for flavor) and consumed as a beverage. Sour Buttermilk Liquid drained from the butter churn (or called cream) is pasteurized and mildly coagulated with lactic acid culture plus Leuconostoc flavor bacteria. It is higher in fat and heavier in consistency than cultured buttermilk.

62. Acidophilus Milk Whole or low-fat milk is pasteurized and inoculated with Lactobacillus acidophilus. It may provide health benefits by favorably altering the microflora of the intestinal tract. Yogurt Whole or low-fat milk is pasteurized and coagulated to a custard-like consistency with a mixed lactic acid culture containing Lactobacillus bulgaricus and Streptococcus thermophilus. It may be flavored or unflavored.

63. III. Cheese Ripening A. Proteolysis Milk clotting enzymes are added to milk to cleave k-casein and begin coagulation of the milk, but they also have general proteolysis capabilities that contribute to cheese aging. However, excessive general proteolysis leads to excessive loss of fat and cheese yield and adversely affects flavor and texture. In hard cheese, 25-35% of the insoluble protein of the curd may be converted into soluble protein, whereas in soft varieties such as, Brie, Camembert, or Limburger, over 80% of the insoluble protein can be converted to water-soluble compounds such as peptides, amino acids, and ammonia. Some rennet substitutes show high rates of proteolysis and frequently produce bitter taste in cheese. Bitter flavor results when the bitter peptides are formed faster than they can be broken down further by proteolytic enzymes of the starter organisms.

64. Peptide bitterness The bitter taste has long been known as a fault during cheese maturation. The bitter taste originates from the very products of proteolysis, the peptides. The problem of bitterness is, thus, inherent in any application of proteases for food. General consensus that the bitterness is caused by the presence of peptides of medium to short chain length, with an exceptionally high content of hydrophobic amino acid side chains. It is generally observed that the higher the hydrophobicity of a particular peptide, the more intense is its bitter taste. The majority of peptide material in protein hydrolysate is non-bitter and bitterness is caused solely by the presence of a small fraction of soluble highly hydrophobic

65. B. Lipolysis Lactobacilli and Micrococci play significant roles in lipolysis during cheese ripening. Most hard cheeses are ripened by bacterial action in the cheese, whereas the soft varieties of cheese are ripened largely by yeasts, molds, or bacteria growing on the cheese surface. Because of the fairly high pH range for milk lipase (6-9) and its sensitivity to heat, it does not seem to play an important role in the ripening of cheese made from pasteurized milk. Milk lipases may lead to undesirable rancidity if freshly drawn milk is cooled too rapidly. “Membrane lipase” is adsorbed on the fat globules on rapid cooling and initiates lipolysis. The lipase becomes active if raw milk is homogenized or agitated or if foaming or great temperature fluctuations occur.

66. In some of the mold cheese, such as Roquefort, Gorgonzola, blue, and Stilton, the lipases produced by Penicillium roqueforti play an important part in flavor formation. C. Accelerated Cheese Ripening Enzymes often are added as crude preparations made by heat treating or freeze shocking microorganisms to significantly accelerate cheese flavor development in the manufacture of cheese paste products. Flavor enhancing enzymes have been added as pastes, as solutions, through encapsulation techniques, and as soluble enzymes with salt. In order to supplement lipolytic activity of microorganisms during cheese ripening, efforts have been made through lipase addition.

67. Addition of pregastric esterases is practiced for the production of Italian type cheese. Traditionally, Italian cheeses have been prepared with rennet paste made by drying the entire stomachs of calves, kids (young goat), or lambs, including the milk contents of the stomach. The paste made from kids or lambs particularly gives the cheese a characteristic piquant flavor that cannot be obtained with rennet extracts or milk free pastes. The difference is created by enzymes secreted by glands at the base of the tongue of the animals. These enzymes are extracted from the excised glands of calves, lambs, or kids. They are the major source of the characteristic lipolytic activity associated with rennet paste. These pregastric esterase enzymes are sometimes called oral lipases or oral glandular lipases.

68. IV. Lactase A. Lactose Intolerance Lactose is the major carbohydrate of milk, and milk is the source of Lactose. Use of dairy foods as dietary sources of specific nutrients such as calcium suggested the need for lactose treatment of milk and milk products. The inability of some populations to catabolize lactose properly because of inadequate intestinal lactase (β-galactosidase) activity is a concern of for the dairy industry and for the nutritionists.

69. B. Lactase in Dairy Processing Lactose is in abundant supply, particularly in cheese whey, presenting serious disposal problems in the cheese industry. Lactose has poor solubility (about 15% in water at 20 ℃ ) and is less than one-sixth as sweet as sucrose. Lactose hydrolysis produces a more soluble, sweeter mixture of equal proportions of glucose and galactose and small amounts of diverse polysaccharides. The lactose fermenting yeast such as Kluyveromyces lactis ( formerly Saccharomyces lactis), and Kluyveromyces fragilis, or fungi such as Aspergillus oryzae are the most frequently used sources of the enzyme lactase.

70. V. Hydrogen Peroxide and Catalase Treatment Farms where heat pasteurization is not feasible, addition of H 2 O 2 can be added, 1ml 33% H 2 O 2 per liter of milk at the farm and another addition of 1 ml H 2 O 2 at the dairy plant, heating to 50 ℃ for 30 min, cooling to 35 ℃, and adding catalase. Pasteurization destroys not only milk pathogens but also acid forming microorganisms, and inactivates some of the natural enzymes of the milk. H 2 O 2 (total 0.05% or less) is effective in reducing the counts of pathogenic microorganisms and inactivates milk catalase and peroxidase, but it permits lactic acid-forming microorganisms to survive, and lipases, proteases, and phosphatases are not affected for cheese making.

71. The H 2 O 2 / catalase treatment cannot be substituted legally for pasteurization, since complete elimination of pathogens is not assured. Catalase is widely distributed among animals, plants, bacteria, and fungi. Catalase for commercial use is obtained from animal liver, bacteria (Micrococcus lysodeikticus), and fungi (Aspergillus niger). H 2 O 2 is extremely toxic to cells because they attack unsaturated fatty acid components of membrane lipids, thus damaging membrane structure. Certain enzyme system, DNA. Not completely known. 2 H 2 O 2 Catalase 2H 2 O + O 2

72. Other Commercial Applications of Hydrogen Peroxide In combination with glucose oxidase, for treatment of food packaging materials to prevent oxidative deterioration of food. For prevention of moisture accumulation in the package. Pasteurization of egg white. In combination of H 2 O 2 and heat, based on 0.05% addition to the albumen, a dose of 0.005% (wt/wt) catalase (fungal) is recommended as a starting dose. Impregnation of these two enzymes and the dianisidine on the paper strip to test the presence of glucose in urine.

73. Desugaring Traces of glucose in the egg white. On drying or during subsequent storage at temp. much above freezing, glucose combines with egg proteins and the Maillard browning reaction (discoloration) occurs. Removal of glucose through fermentation by yeasts or with commercial enzymes prior to the drying of all egg white.