Enzymes in Food Industries

By Lydia ETCHEBESTFrédéric RIVAL

Immobilization of enzymes

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

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

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

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

1-1 : Physical Adsorption

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.

1-2 : Covalent 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

1-3 : Ionic Binding

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.

Entrapping Enzymes

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.

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.

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.

Enzymes That Aid Beverages

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

Enzymes Uses

Amylases Starch breakdown

Β glucanases Improved filtering ability

Proteases Prevent “chill haze”

The brew crew

continuation

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

The Vine

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

Juices

Enzymes Uses

Amyloglucosidase Starch breakdown in early season fruit

Cellulase Liquefaction of fruit

Esterase Aroma development

Lipoxygenase Aroma development

Pectinesterase Clarification of juice

Polygaclacturonase Clarification of juice

Polyphenoloxidase Color and flavor

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.

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

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 )

Starch & sugar Industries continuation

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)

Breadmaking

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)

Breadmaking - Amylase

Purposes: destruction of gluten protein

cohesiveness improve elasticity and handling

properties of doughs (good volume) Over - Activity: Decomposition of

bread structure

Breadmaking - Proteinase

Purposes (rye flour): a less tough dough improve volume a softer crumb better storage properties

Breadmaking - Pentosanase

Characteristics of good meat: juiciness good chewability firm texture color taste

Meat tenderization

Main Enzymes: Papain Bromelain calpain

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

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

Meat tenderization - Enzymes

Calpains attack certain proteins of the Z-line proteolysis of key myofibrillar proteins

cathepsins attack myosin and actin

Meat tenderization - Enzymes

a way of preserving the nutrients of milk

More than 1000 varieties of cheese

Cheese making

Process: clotting enzymatic conversion of k-casein

into para-k-casein coagulation of the micelles of

paracasein aging

Cheese making

Inside the raw milk, there are endogenous enzymes

Pasteurized milk includes inactivated enzymes

Food industry must use exogenous enzymes

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

Cheese making

lipases Fat hydrolysis Flavour development 13-galactosidase and lactase

(dairy industry) increase sweetness prevent crystallization

Cheese making

Endogenous enzymes are used traditionally

Exogenous enzymes are more and more used in food industry:

Control the process Homogenization of the production More efficient

Conclusion:

Thank you!

That’s all !