Enzymes in Food Industries

1. Enzymes in Food Industries By Lydia ETCHEBEST Frédéric RIVAL

2. Immobilization of enzymes Why it is important to choose a method of attachment prevent loss of enzyme activity? - To avoid reaction - The structure is retained in the enzyme through hydrogen bonding or the formation of electron transition complexes: prevent vibration of the enzyme and increase thermal stability The different methods are: Carrier-Binding: the binding of enzymes to water-insoluble carriers Cross-linking: intermolecular cross-linking of enzymes by bi-functional or multi-functional reagents. Entrapping: incorporating enzymes into the lattices of a semi-permeable gel or enclosing the enzymes in a semi-permeable polymer membrane

3.  Carrier-Binding • The oldest immobilization technique for enzymes • Some of the most commonly used carriers for enzyme immobilization are polysaccharide derivatives such as cellulose, dextran, agarose, and polyacrylamide gel. • The selection of the carrier depends on the nature of the enzyme itself, as well as the: - Particle size - Surface area - Molar ratio of hydrophilic to hydrophobic groups - Chemical composition • the carrier-binding method can be further sub-classified into: • Physical Adsorption • Covalent Binding • Ionic Binding

4. 1-1 : Physical Adsorption Of enzyme protein on the surface of water-insoluble carriers. Advantages : no reagents and only a minimum of activation steps are required Disadvantages : the adsorbed enzyme may leak from the carrier during use due to a weak binding force between the enzyme and the carrier. Moreover, the adsorption is non-specific, further adsorption of other proteins or other substances

5. 1-2 : Covalent Binding Based on the binding of enzymes and water-insoluble carriers by covalent bonds The functional groups that may take part in this binding are Amino group, Carboxyl group, Sulfhydryl group, Hydroxyl group, Imidazole group, Phenolic group, Thiol group, Threonine group,Indole group Disadvantages : covalent binding may alter the conformational structure and active center of the enzyme, resulting in major loss of activity and/or changes of the substrate Advantages : the binding force between enzyme and carrier is so strong that no leakage of the enzymes occurs, even in the presence of substrate or solution of high ionic strength.

6. 1-3 : Ionic Binding Of the enzyme protein to water-insoluble carriers containing ion-exchange residues Polysaccharides and synthetic polymers having ion-exchange centers are usually used as carriers Advantages : the enzyme to carrier linkages is much stronger for ionic binding Disadvantages : the binding forces between enzyme proteins and carriers are weaker than those in covalent binding

7.  Cross-Linking Either to other protein molecules or to functional groups on an insoluble support matrix It is used mostly as a means of stabilizing adsorbed enzymes and also for preventing leakage from polyacrylamide gels The most common reagent used for cross-linking is glutaraldehyde Disadvantages : Cross-linking reactions are carried out under relatively severe conditions. These harsh conditions can change the conformation of active center of the enzyme; and so may lead to significant loss of activity.

8.  Entrapping Enzymes

9.  Entrapping Enzymes continuation Based on the localization of an enzyme within the lattice of a polymer matrix or membrane It can be classified into lattice and micro capsule types. This method differs from the covalent binding and cross linking in that the enzyme itself does not bind to the gel matrix or membrane. This results in a wide applicability Disadvantages : The conditions used in the chemical polymerization reaction are relatively severe and result in the loss of enzyme activity.

10. Enzymes That Aid Beverages • Enzymes perform many functions in beverages • They can help to form nutrients for the fermentation process, facilitate processing, and affect the color, flavor and clarity of the finished product • They are Biological catalysts based mainly on protein, they remain unchanged at the completion of the reaction • Enzymes are typically named for the reactions they catalyze. They fall into six major categories: oxidoreductases, hydrolases, lysases, transferases, ligases and isomerases. Of these types, hydrolases play the most important role in the beverage industry.

11. Enzymes That Aid Beverages • Most enzymes catalyze highly specific reactions (alpha-1-4-glucan glucanhydrolase) but not all enzymes show the same degree of specificity (papain) • A number of factors affect enzyme activity : T°C, pH, concentration, contact time with the substrate, trace metals, salt and salt ions, and oxidizing agents. • enzymes in beverages can occur naturally in the ingredients used to formulate the beverage. Most fruits contain low levels of pectinase, and the malting process produces significant levels of amylase. In other cases, a product designer can add an enzyme preparation to achieve a specific goal or to supplement or standardize naturally occurring enzymes.

12.  The brew crew • crucial role in the production of beer and other types of malted liquor, such as whiskey • three major functions: the formation of sugars to be used during fermentation; viscosity control; and, in beer, "chill-proofing." • The additional enzymes can help make up for the lack of amylases in the grains used and increase the level of fermentable sugars. • The beta-glucans and pentosans are left intact : they absorb high levels of water, increasing the viscosity. • Protein precipitation can cause the “Chill haze” in beer : addition of papain

13.  The brew crewcontinuation

14.  The Vine • Even if wine-making generally relies on the natural enzymes present in the grape or formed as a product of fermentation, added enzymes could help in several areas. • Added pectinase can aid in pressing and clarification. Particularly helpful during during the mash process, since higher temperatures mean increased levels of pectin in the juice

15.  Juices • Enzymes are used to extract juice from fruits and prepare a finished product. For non-citrus juices, such as apple, grape and berry, processors add enzymes at the beginning of the mash stage. • The cell walls of fruits consist of cellulose, hemicelluloses, pectin and proteins. To extract a larger amount of juice and to facilitate pressing, structures must be break down the viscosity of the juice decreases and the size of the particles is increased. The combination of those two elements causes a floc that settles out, and the clarified juice can be removed • The most prevalent enzymes used in juice processing are pectinases • Enzymes in fruit juice also affect the color and flavor of the juice

17. Dairy doses (milk and other fluid dairy products) • The enzymatic process of interest is the hydrolysis of lactose, for the lactose-intolerant • Lactase, catalyzes the hydrolysis of the beta-D-galactoside, converting it into glucose and galactose.

18. Starch & sugar Industries • Considerable quantities of the sweeteners used throughout the world are derived from starch as opposed to cane or beet sugar • The treatment of starch with enzymes results in a variety of sweet syrups

19. Starch & sugar Industries continuation • Three stages can be identified in starch modification: • amylases liberate "maltodextrin" by the liquefaction process; they are not very sweet as they contain dextrins and oligosaccharides. • The dextrins and oligosaccharides are further hydrolysed by enzymes such as pullulanase and glucoamylase in a process known as saccharification. Complete saccharification converts all the limit dextrans to glucose, maltose and isomaltose. The resulting syrups are moderately sweet and are frequently modified further. • Treatment of glucose/maltose syrups with glucose isomerase converts a large proportion of the glucose to fructose which is sweeter than glucose, process of isomerisation(50 % fructose and 50 % glucose: High Fructose Syrups )

20. Breadmaking • Important in human nutrition: half of their required carbohydrates and about one-third of their protein from bread • Process uses: • Endogenous enzymes • Exogenous enzymes (more efficience)

21. Breadmaking - Amylase • Purposes: • to improve or control dough-handling properties • to improve product quality • Activity: hydrolisation of starch • Origins: • Fungal • Bacterial: cheaper but thermostable (excess of activity)

22. Breadmaking - Proteinase • Purposes: • destruction of gluten protein cohesiveness • improve elasticity and handling properties of doughs (good volume) • Over - Activity: Decomposition of bread structure

23. Breadmaking - Pentosanase • Purposes (rye flour): • a less tough dough • improve volume • a softer crumb • better storage properties

24. Meat tenderization • Characteristics of good meat: • juiciness • good chewability • firm texture • color • taste

25. Meat tenderization - Enzymes • Main Enzymes: • Papain • Bromelain • calpain

26. Meat tenderization - Enzymes • Characterisics of calpain: • The protease must be endogenous to skeletal muscle cells • The protease must have the ability to reproduce post-mortem changes in myofibrils in vitro • The protease must have access to myofibrils in tissue

27. Meat tenderization - Enzymes • Mechanism of tenderization (calpain): • proteolysis of key myofibrillar proteins • resistance of myofibrillar proteins to calpains • regulation and stability of calpains in muscles

28. Meat tenderization - Enzymes • Calpains attack certain proteins of the Z-lineproteolysis of key myofibrillar proteins • cathepsins attack myosin and actin

29. Cheese making • a way of preserving the nutrients of milk • More than 1000 varieties of cheese

30. Cheese making Process: • clotting • enzymatic conversion of k-casein into para-k-casein • coagulation of the micelles of paracasein • aging

31. Cheese making • Inside the raw milk, there are endogenous enzymes • Pasteurized milk includes inactivated enzymes • Food industry must use exogenous enzymes

32. Cheese making • Bacteria convert lactose to lactic acid. This acid environment would eventually coagulate the protein. • Using of rennet • Split the k-casein • One part is extremly soluble and the other part remains in micelles • Micelles coagule

33. Cheese making • lipases • Fat hydrolysis • Flavour development • 13-galactosidase and lactase (dairy industry) • increase sweetness • prevent crystallization

34. Conclusion: • Endogenous enzymes are used traditionally • Exogenous enzymes are more and more used in food industry: • Control the process • Homogenization of the production • More efficient

35. That’s all ! Thank you!