Group Members: Nguyễn Thị Cẩm Nhi (BTFTIU17041)

Đặng Tống Trọng Nghĩa (BTFTIU17010)

Đinh yến Nhi (BTFTIU17080)

Lê Thanh Bảo Ngọc (BTFTIU17065)

Assignment 1: Enzymes in protein modification

(Outline)
A. Overview
Enzymes: a biological catalyst that promotes and speeds up a biochemical reaction
without itself being altered in the process.
Proteins: large biomolecules, or macromolecules, consisting of one or more long
chains of amino acid residues.
 Proteins are widely used as ingredients in the food industry and can be isolated
from many types of raw materials like milk (casein and whey), wheat (gluten), soya,
meat (gelatine and meat extracts), etc.
 The opportunities to use proteins as ingredients in food products are often limited by
the properties of the proteins.
=> One of the methods to modify these properties is the hydrolysis of the
proteins to smaller peptides.

B. Enzymes in protein modification

 Hydrolysis reaction
 Hydrolysis is the chemical breakdown of substances by water and depends on the
chemistry, solubility, pH, and the oxidation-reduction potential of compound.
 The properties of the hydrolyzed proteins are adjusted by the selection of the
protease, pretreatment of raw material, the parameters of hydrolysis as well as the
downstream processing of the hydrolyzate
 The protease catalyzed reaction cleaves a peptide bond in the protein.
For example,

Reaction scheme for protease catalyzed reaction

 The extent or degree of hydrolysis is defined by the percentage of peptide bonds
hydrolyzed:
 peptide bonds cleaved
Degree of h ydrolysis ( DH ) %= ∗100
total number of peptide bonds

=> There is one important detail about the hydrolysis reaction.

 Controlling the hydrolysis reaction
 Several aspects of this reaction are utilized in the control of the DH.
 Some are related to the formation of the amino groups, others to the acidity of the
peptides or to the change in other properties of the resulting peptides.
 The most accurate method for following the reaction is the pH-stat method.
 A good alternative for monitoring the reaction is the use of osmolality.
 Chemical methods used for detection of the amino groups are widely used.
 Some methods of analysis use the correlation between degree of hydrolysis and
properties of the protein itself.

 Proteases and properties of hydrolysis protein

Proteases: are classified according to the source of origin, the site they hydrolyze a
protein. It is very common to use combinations of proteases to optimize the properties
of the end product.
 Due to differences in specificity of the enzymes the resulting peptides also widely
differ.

 Properties of Hydrolysis Protein:

1. Taste
 One of the most important properties for food protein hydrolyzates is taste.
 When proteins are hydrolyzed there are two main contributions to the taste of the
peptides.
 the release of flavour components ‘hidden’ in the protein structure
 the formation of small peptides with a relative high content of hydrophobic amino
acids => create bitterness
 The tendency of bitterness formation is different from one source of protein to
another.
 Prevention of bitterness is achieved by selection of the type of protease(s) and
operating condition soft hehydrolysis process.
 The remaining bitterness fits better into the flavour of orange, lime and grapefruit
flavoured formulas than into strawberry flavoured drinks.
2. Solubility
 Protein hydrolyzates are used as ingredients in beverages,their solubility is
obviously very important.
 Native proteins have different solubilities, dependant, amongst other factors, on the
pH of the solution.
 For a specific enzyme and a specific substrate, the solubility will be higher at higher
DH.

3. Viscosity
 Normally the hydrolysis of proteins causes a decrease in the viscosity of the protein
solution.
 The high viscosity was caused by aggregation of the peptides causing a gelation.
 The concept of protein viscosity reduction is being applied industrially.
Ex: the use of protease in a fish meal stick water plant
 By adding protease the viscosity is reduced, which makes processing of the protein
solutions much easier.
 Low viscosity is desirable in protein-fortified drinks and clinical nutrition products.

4. Emulsification
 The emulsifying properties are changed, since both the size of the molecules, the
charge and the distribution of hydrophobic and hydrophilic regions are altered.
 Even when the emulsifying capacity increased as a result of the hydrolysis, there
was no improvement in the stability of emulsions compared with un-hydrolyzed
proteins.
 There is no direct relation between DH and emulsifying properties
 The emulsifying properties of extensively hydrolyzed proteins are of importance in
low allergenic baby food products
 A good emulsion in food products promotes the uptake of the nutrients besides
giving a better appearance.

5. Foaming
The use of hydrolysis to improve the foaming properties has been suggested for making
an egg white substitute from soya protein
The type of raw material used for making hydrolyzates is important.
There are big variations in the foaming properties, dependent upon the type of enzyme
used
6. Gelling
By specific proteolysis, the k- casein in milk is hydrolyzed at the pepetide bond Phe 105-
Met106, causing the charge of the casein micelle to change, thus enabling gelation.
Instead of producing cheese from the curd it can be used for production of rennet
casein. In this case, the curd is heated which contracts the gel and releases the whey,
followed by drying of the casein.
Trypsin, papain, Pronase and a protease from Streptomyces griseus all caused gelling
of whey protein
The most important parameters for plastein formation are type and size of peptides,
concentration of peptides and pH of the reaction mixture. Generally, the DH of the
protein hydrolyzate must be high to promote the plastein formation.
The plastein reaction can be used to reduce the bitterness of protein hydrolyzates,but
difficult to control reaction

7. Allergenicity
The use of protein hydrolysis to lower the allergenicity of baby formulations
They reduce whey protein or casein (ingredient which make human allergenic) due to
the slightly better amino acid composition but also to the inherent difficulties in
producing a low- bitter product from casein.

8. Bioactive peptides.
The scientists use peptide as bioactive components
The peptides originate from different parts of the caseins and whey proteins in the milk
Cow milk protein help us: Opioid antagonists, antihypertensive peptides, antithrombotic
peptides, antimicrobial peptides, immunostimulants, mineral carriers and anti-stress
peptides
In the body, these peptides can be formed by the enzyme activity in the digestion
system and/or as result of the lactic acid fermentation in the intestine.
Peptides can also have positive effects on the cardiovascular system
(hypocholesterolemic and antioxidative)
The peptides can be produced in three different ways:
 Enzymatic hydrolysis by digestive enzymes.
 Enzymatic hydrolysis by enzymes from microorganisms or plants.
 Fermentation of milk by proteolytic starter cultures.
9. Protein hydrolyzates on the markets

1. Processing issues
- Preparing the raw material
- Hydrolysis
- Inactivating the proteases
- Recovery of the protein/peptide product
- Concentration, formulation and drying
2. Conclusion
- The use of proteases for modification of protein properties to produce protein
ingredients with improved properties is well established in the market.
- Controlling the properties of the peptides is a very complex task and may require
new proteases.
3. References