Download FOOD BIOTECHNOLOGY BIT 313 3(3-0)

Dr. Zaffar Mehmood
Definitions:
 Food: means a raw, cooked, or processed edible substance, ice,
beverage, or ingredient used or intended for use or for sale in whole or
in part for human consumption, or chewing gum.
 Food Technology: is the application of food science & engineering
principles to the selection, preservation, processing, packaging,
distribution, and use of safe, nutritious, and wholesome food
 Food Security:
 Food security is achieved when all people, at all times, have physical, social
and economic access to sufficient, safe and nutritious food to meet their
dietary needs and food preferences for an active and healthy life. (800
million people suffer from hunger)
 Food Safety:
 A suitable product which when consumed orally either by a human or an
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animal does not cause health risk to consumer
Food Biotechnology: Application of technology to modify genes of
animals, plants, and microorganisms to create new species which have
desired production, marketing, or nutrition related properties. Called
genetically engineered (GE) foods, they are a source of an unresolved
controversy over the uncertainty of their long-term effects on humans and
food chains
Principles of Biochemistry
 Biochemical Building blocks
 Proteins
 Carbohydrates
 Lipids
 Nucleic acids
 Protein synthesis
 Enzymes
Proteins
 1. The 20 amino acids that cells use to make proteins have a
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common core structure.
a. Most amino acids have a central carbon atom to which is
attached a hydrogen atom, an amino group, NH3 +, and a
carboxyl group, COO–.
b. The side chain or R group distinguishes each amino acid
chemically.
2. Assembly of the amino acids to form peptides and
proteins occurs by stepwise fusion of the carboxyl group of
one amino acid with the amino group of another, with loss of
a molecule of water during the reaction to form a peptide
bond.
3. Proteins can have a broad diversity of structures depending
on their amino acid sequences and composition.
4. The central carbon and the atoms involved in end-to-end
linkage of the amino acids form the polypeptide backbone,
with the side chains protruding outwardly to interact with
other parts of the protein or with other molecules.
Proteins continued
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The 20 common amino acids can be classified into groups with similar side chain
chemistry.
1. The nonpolar or hydrophobic amino acids—glycine, alanine, valine, leucine,
and isoleucine–have alkyl side chains (or simply a hydrogen atom in the case of
glycine).
2. Serine and threonine are small, polar amino acids that have hydroxyl groups.
3. The sulfur-containing amino acids are cysteine and methionine.
4. The aromatic amino acids, phenylalanine, tyrosine, and tryptophan, have ring
structures and are nonpolar with the exception of the hydroxyl group of tyrosine.
5. The acidic amino acids, aspartic acid and glutamic acid, have carboxyl groups.
6. The amides of the carboxylic amino acids, asparagine and glutamine, are
uncharged and polar.
 7. Members of the basic group, histidine, lysine, and arginine, have weak-base
 side chains.
 8. Proline is unique; it is an amino acid because its side chain loops back to
form a five-membered ring with its amino group, which causes proline to
produce kinks in the polypeptide backbone.
 Essential amino acids: Histidien, Isolucine, lucine, lysin, methionine,
phenylalanine, threonine, tryptophan, valine.
Protein structure:
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A. Primary structure refers to the linear sequence of amino acids linked by peptide bonds to make up a protein.
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B. Secondary structure describes the twisting of the polypeptide backbone into regular structures that are stabilized by hydrogen
bonding.
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1. The α-helix is a coiled structure stabilized by intrastrand hydrogen bonds
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a. The structure is both extensible and springy, which contributes to the function of proteins that are primarily α-helix, such as keratins
of fingernails,hair, and wool.
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b. Amino acid side chains project outward, away from the axis of the α-helix and decorate its exterior surface.
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2. _-Sheet structures are made from highly extended polypeptide chains that link together by hydrogen bonds between the neighboring
strands and can be oriented in parallel or antiparallel arrays.
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a. Due to the very extended conformation of the polypeptide backbone, β-sheets resist stretching.
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b. The amino acid side chains project on either side of the plane of a β-sheet.
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c. Silk is composed of the protein fibroin, which is entirely β-sheet.
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C. Tertiary structure is formed by combinations of secondary structural elements into a three-dimensional organization that is mainly
stabilized by noncovalent interactions, such as hydrogen bonds.
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1. Protein folding is the complex process by which tertiary structures form within the cell.
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2. Regions of proteins that are capable of folding independently and that often have distinct functions are called domains.
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3. The side chains of highly polar amino acids tend to reside on the exterior of proteins, where they can form hydrogen bonds with water.
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4. The side chains of nonpolar amino acids are normally clustered in the interior of proteins to shield them from water.
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D. Quaternary structure occurs in proteins that have multiple polypeptide chains, called subunits.
CARBOHYDRATES
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Monosacharides:
Carbohydrates have a carbon backbone bearing hydroxyl groups with either an
aldehyde or ketone at one carbon.
B. Simple sugars may take on several types of structures in solution.
1. Simple sugars or monosaccharides are classified according to the number of
carbons in the backbone.
a. Pentoses have five carbons; examples include ribose and ribulose.
b. Hexoses have six carbons: examples include glucose, galactose, fructose, and
mannose.
Hexoses have six carbons: examples include glucose, galactose, fructose, and
mannose.
2. Most sugars are asymmetric and designated either D- or L- in stereochemistry.
3. Simple sugars in aqueous solution usually form cyclic structures, either hemiacetals or hemiketals
a. The rings may have five or six members.
b. Depending on how the cyclic structure was formed, the substituents at the connecting carbon may
be anomers—having either α or β configuration.
4. The hexoses are structurally distinguished by different configurations at one or more carbons.
a. Diastereomers are molecules differing in configuration at one or more carbons.
b. Epimers are molecules that differ in their configurations at only one carbon, thus glucose and
galactose are both epimers and diastereomers. Modifications of one or more groups convert simple
sugars into a variety of sugar derivatives.
a. Replacement of −OH by −H converts the sugar into a deoxymonosaccharide, such as
deoxyribose.
b. Replacement of −OH by −NH2 converts the sugar into an amino sugar designated as -osamine,
eg, glucosamine.
LIPIDS
 Lipids are formed from structural units with a pronounced hydrophobicity. This
solubility characteristic, rather than a common structural feature, is unique for this class
of compounds. Lipids are soluble in organic solvents but not in water. Water insolubility
is the analytical property used as the basis for their facile separation from proteins and
carbohydrates. Some lipids are surface-active since they are amphiphilic molecules
(contain both hydrophilic and hydrophobic moieties). Hence, they are polar and thus
distinctly different from neutral lipids.
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 The majority of lipids are derivatives of fatty acids. In these so-called acyl lipids the fatty
acids are present as esters and in some minor lipid groups in amide form (Table 3.1). The
acyl residue influences strongly the hydrophobicity and the reactivity of the acyl lipids.
Some lipids act as building blocks in the formation of biological membranes which
surround cells and subcellular particles. Such lipids occur in all foods, but their content is
often less than 2% . Nevertheless, even as minor food constituents they deserve
particular attention, since their high reactivity may strongly influence the organoleptic
quality of the food.
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 Primarily triacylglycerols (also called triglycerides) are deposited in some animal tissues
and organs of some plants. Lipid content in such storage tissues can rise to 15–20% or
higher and so serve as a commercial source for isolation of triacylglycerols. When this
lipid is refined, it is available to the consumer as an edible oil or fat. The
nutritive/physiological importance of lipids is based on their role as fuel molecules (37
kJ/g or 9 kcal/g triacylglycerols) and as a source of essential fatty acids and vitamins.
Apart from these roles, some other lipid properties are indispensable in food handling or
processing.
 c. Oxidation of the terminal −CH2OH to −COOH converts the sugar
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into a -uronic acid, such as glucuronic acid.
C. Sugars can be polymerized or interconnected to create chains
termed oligosaccharides
(≤ 8 sugars) or polysaccharides (> 8 sugars)
1. The linkage between sugars is formed by condensation of the
hemiacetal or hemiketal of one sugar with a hydroxyl of another sugar
with loss of water in the reaction.
2. The linkage is called a glycosidic bond and can either be classified
as α or β depending on the stereochemistry of the anomeric carbons at
the bridge points.
3. The important difference between α and β glycosidic bonds can be
seen in the digestibility of the major plant polysaccharides cellulose
and starch.
a. Cellulose, the primary component of plant cell walls, is made up of
_–1,4- linked glucose, which cannot be broken down by digestive
enzymes. So humans cannot use cellulose as a direct dietary source of
glucose.
b. Starch, the main form of stored sugar in plants, is made up of _–1,4linked glucose, which can be hydrolyzed by enzymes of the digestive
tract, eg,
α-amylase. Thus, starch is an important dietary source of glucose.
BENEFITS OF FERMENTATION:
the use of mild conditions of pH and temperature
which maintain (and often improve)
the nutritional properties and sensory
characteristics of the food
• the production of foods which have flavours or
textures that cannot be achieved by other methods
• low energy consumption due to the mild operating
conditions
• relatively low capital and operating costs
• relatively simple technologies.
Fermentation and enzyme technology
The main advantages of technical enzymes are:
• they cause highly specific and controlled changes to
foods
• there is minimal loss of nutritional quality at the
moderate temperatures employed
• lower energy consumption than corresponding
chemical reactions
• the production of new foods, not achievable by other
methods.
The main factors that control the growth and
activity of micro-organisms in food
fermentations are:
• availability of carbon and nitrogen sources,
and any specific nutrients required by
individual micro-organisms
• substrate pH
• moisture content
• incubation temperature
• redox potential
• stage of growth of micro-organisms
• presence of other competing microorganisms
APPLICATION OF ENZYME TECHNOLOGY:
micro-encapsulation in polymer membranes which retain
the enzyme but permit the passage of substrates and
products
• electrostatic attachment to ion exchange resins
• adsorption onto colloidal silica and/or cross linking with
glutaraldehyde
• covalent bonding to organic polymers
• entrapment in polymer fibres (for example cellulose
triactetate or starches)
• co-polymerisation with maleic anhydride
• adsorption onto charcoal, polyacrylamide, or glass
LIMITATIONS OF TECHNOLGY
the higher cost of carriers, equipment and
process control
• changes to the pH profiles and reaction
kinetics of enzymes
• loss of activity (25–60% loss)
• risk of microbial contamination
ENZYME CHARACTERISTICS:
• short residence times for a reaction
• stability to variations in temperature and
other operating conditions over a period of
time (for example glucose isomerase is
used for 1000 h at 60–65ºC)
• suitability for regeneration.
The requirements of commercial enzyme
production from micro-organisms are as follows:
• micro-organisms must grow well on an
inexpensive substrate
• substrates should be readily available in adequate
quantities, with a uniform quality
• micro-organisms should produce a constant high
yield of enzyme in a short time
• methods for enzyme recovery should be simple
and inexpensive
• the enzyme preparation should be stable
Bread Industry
 Basic ingredient, flour
 Types of wheat and wheat milling
 Composition (Germ 2.5%, Endosperm 82.5% and Bran 15%
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(epidermis, epicarp, endocorp, testa and aleauron layers)).
Milling 72 % extraction, 28% Bran and shorts.
Cake flour and Baking flour
Ideal flour: (colour, strength, tolerance, High absorption,
Uniformity).
Compressed baker’s yeast. 30% solids, 2c storage./ dried
yeast.
Leavening: Mechanical, CO2, Chemicals, Water vapours.
Baking
 Flour mixing stages: Pick up, drying, clean up, runny
 Fermentation: 5C, 25-30C, humidity 70-75%.
 Punching.
 Proofing: 35-40C and 80-85% humidity.
 Types of Fermentation:
 Straight Dough and Sponge Dough Method
 Straight dough: weighing sifting and blending flour
tempering of water, preliminary mixing of yeast dried milk
etc.
Dough mixed
dough placed in trough
dough allowed to rise, turned and folded
rounding
intermediate proof
moulding
Panning
pan
proof
baking
cooling
slicing .
 Sponge Dough Process:
 weighing sifting and blending flour tempering of
water, preliminary mixing of yeast dried milk etc.
Dough mixed
sponge mixed
sponge placed
in trough
sponge allowed to mature
sponge
placed in mixer
sponge broken up and mixed
with dough ingredients
final dough placed in
trough
allowed to rise sometimes turned and
folded
dough sent to bench or divider
Dividing and scaling
rounding
intermediate
proof
Moulding
panning
pan
proof
baking
cooling
slicing.
Beer Prodution:
Beer Fermentation ICT
Requirement: Continuous beer fermentation
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stirred vs. unstirred tanks
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single vessels vs. multiple vessels in a series
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open systems vs. closed systems
Major issue of contamination.
Choice of Carrier material:
 (i) attachment to the support surface, which can
be spontaneous or induced by linking agents;
(ii) Entrapment within a porous matrix;
(iii) containment behind or within a barrier;and
(iv) self-aggregation, naturally or artificially induced
Immobilized Cell technology
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High cell mass loading capacity
Easy access to nutrient media
Simple and gentle immobilization procedure
Immobilization compounds approved for food applications
High surface area-to-volume ratio
Optimum mass transfer distance from flowing media to
centre of support
Mechanical stability (compression, abrasion)
Chemical stability
Highly flexible: rapid start-up after shut-down
Sterilizable and reusable
Suitable for conventional reactor systems
Low shear experienced by cells
Easy separation of cells and carrier from media
Readily up-scalable
Economically feasible (low capital and operating costs)
Desired flavor profile and consistent product
Complete attenuation
Controlled oxygenation
Control of contamination
Controlled yeast growth
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Wide choice of yeast
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MILK FERMENTATION
 Cow's milk contains approximately 3.2% protein, 4.8% lactose,
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3.9% lipids, 0.9% minerals, traces of vitamins, and 87.2% water.
Among the proteins, casein in colloidal suspension as calcium
caseinate is present in higher amounts than the other two
soluble proteins, albumin and globulin.
Lactose is the main carbohydrate and is present in solution.
lipids are dispersed as globules of different sizes in emulsion (fat
in water).
Minerals are present in solution and as colloid with casein.
Water-soluble vitamins are present in aqueous phase, whereas
fat-soluble vitamins are present with the lipids. The solid
components (12.8%) are designated as total solids (TS), and TS
without lipids is designated as solid-not-fat (SNF; ca. 8.9%).
The whey contains principally the water-soluble components,
some fat, and water
Milk compositin (Fat)
 Fat globules, are surrounded by a polar milk fat globule
membrane (MFGM).
 Triacylglycerols are the predominant lipid fraction
(98%) of the total lipids alongwith 2% Diacylglycerols,
monoacylglycerols, fatty acids, phospholipids, and
sterols.
 The phospholipids are integral components of the
MFGM. 65% saturated (26% palmitic acid and 15%
stearic acid). Rest are short- and middle-chain fatty
acids, including 3.3% butyric acid.
 These fatty acids and the breakdown products of these
fatty acids are important contributors to the flavor of
many cultured dairy products.
Milk Proteins
 Caseins 80% of the total protein and are insoluble at a
pH of 4.6, but are heat stable. The casein micelles exist
as a colloidal dispersion, with a diameter ranging from
40 to 300 nm and containing approximately 10,000
casein molecules. The principal casein proteins, αs1,
αs2, β, and κ, present in the ratio 40:10:35:12,
 Whey proteins remain soluble at pH 4.6 and are heat
sensitive. Four major proteins, β- lactoglobulin (50%),
α-lactalbumin (20%), blood serum albumin (10%), and
immunoglobulins (10%). Cysteine and cysteine
residues in these proteins form disulfide linkages with
other proteins following heat treatment
The production of lactic acid by lactic acid bacteria
decreases the pH of the milk to cause coagulation of the
casein. A pH below 5.3, colloidal calcium phosphate is
solubilized from the casein micelle, causing the micelles
to dissociate and the casein proteins to aggregate and
precipitate at the isoelectric point of casein (pH 4.6).
The resulting gel, which is somewhat fragile in nature,
provides the structure for sour cream, yogurt, and acidprecipitated cheeses, such as cream cheese andcottage
cheese.
Lactic Acid Bacteria:
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Lactic Acid Bacteria
Streptococcus,
Lactococcus,
Leuconostoc, and
Lactobacillus genera.
These bacteria are gram-positive bacteria and belong to
either the Streptococcaceae or Lactobacillaceae families.
Optimal temperature for growth, with 20–30◦C the optimal
temperature for mesophilic bacteria and 35–45◦C. Although
the lactic acid bacteria are quite diverse in growth
requirements, morphology, and physiology, they all have
the ability to metabolize lactose to lactic acid and reduce
the pH of the milk to produce specific cultured dairy
products.
DVI
Many commercial dairy processors now use the direct vat inoculation
(DVI) process for frozen or freeze-dried cultures (up to 1012 bacteria per
gram of starter) in the processing of cultured dairy products.
Benefits:
 Direct addition of the cultures
 No on-site culture preparation.
 Increased phage resistance,
 minimum formation of mutants,
 enhanced the ability to characterize the composition of the cultures,
 improved the consistent quality of cultured dairy products.
Limitations:
 Additional cost of these cultures,
 the dependence of the cheese plants on the starter suppliers for the
selection and production of the starters,
 increased lag phase of these cultures in comparison to on-site culture
preparation.
Enzyme Coagulation:
Rennet: a mixture of chymosin and pepsin, obtained from calf
stomach, is most commonly recognized as the enzyme for
coagulation of casein.
Proteases: from microorganisms and produced through
recombinant DNA technologies have been successfully adapted as
alternatives to calf rennet.
Chymosin: in rennet, cleaves the peptide bond between Phe-105
and Met-106 of κ-casein, releasing the hydrophilic, charged casein
macropeptide, while the para-κ-casein remains associated with the
casein micelle. The loss of the charged macropeptide reduces the
surface charge of the casein micelle and results in the aggregation
of the casein micelles to form a gel network stabilized by
hydrophobic interactions.
Temperature influences both the rate of the enzymatic reaction
and the aggregation of the casein proteins, with 40–42◦C, the
optimal temperature for casein coagulation. The use of rennet to
hydrolyze the peptide bond and cause aggregation of the casein
micelles is used in the manufacture of most ripened cheeses .
Homogenization:
Milk fat globules have a tendency to coalesce and separate upon
standing. Homogenization reduces the diameter of the fat globules
from 1–10 μm to less than 2 μm and increases the total
fat globule surface area.
The physical change in the fat globule occurs through forcing the
milk through a small orifice under high pressure. The decrease in
the size of the milk fat globules reduces the tendency of the fat
globules to aggregate during the gelation period.
In addition, denaturation of the whey proteins and interactions of
the whey proteins with casein or the fat globules can alter the
physical and chemical properties of the milk proteins to result in a
firmer gel with reduced syneresis. Milk to be used to process
yogurt, cultured buttermilk, and unripened cheeses is commonly
homogenized to improve the quality of the final product.
Pasturization:
The heat process, which must be sufficient to inactivate alkaline
phosphatase, also destroys many pathogenic and spoilage
microorganisms, and enzymes that may have a negative impact on
the quality of the finished products.
The time-temperature treatments for the fluid milk pasteurization
have been adapted for the milk to be used in the processing of
cultured dairy products (62.8◦C for 30 minutes or 71.1◦C for
15 seconds).
More severe heat treatments than characteristic of pasteurization
causes denaturation of whey proteins and interactions between βlactoglobulin and κ-casein. In cheeses, this interaction decreases
the ability of chymosin to hydrolyze the casein molecule and
initiate curd precipitation and formation.
Pasteurization has a significant effect of the flavor
profile of the milk. Cultured dairy products produced
from pasteurized milks tend to have less intense flavor
characteristics due to the heat inactivation of the
naturally occurring microorganisms and enzymes in the
milk that contribute to flavor formation.
Lactones and heterocycles are also formed during the
heat treatment of raw milk to contribute cooked flavors.
Cooling
The processing of cultured dairy products relies on the
metabolic activity of the starter cultures to contribute to
acid formation and flavor and texture development.
Once the desired pH or titratable acidity is reached for
these products, the products are cooled to 5–10◦C to slow
the growth of the bacteria and limit further acid
production and other biological reactions.
Probiotics
Probiotics are live microorganisms which, when
administered in sufficient quantity, confer a health benefit to
the host. They must have (a). Viable cells (b). Enough
quantity.
 Improved carbohydrate digestion in the gastrointestinal
(GI) tract.
 reduction of the incidence of diarrhoea.
 immune system enhancement,
 blood cholesterol reduction .
Official recognition. In light of these potential health
benefits,digestive health is considered as one of the ten key
food trends for 2010. Market predicted to reach US$30 billion
by 2015 (Starling 2010).
Prebiotics
Prebiotics are nondigestible food ingredients that beneficially
affect the host by selectively stimulating the growth and/or activity
of one or a limited number of bacteria in the GI system, and
thereby confer health benefits to the host (Roberfroid 2007).
This definition overlaps with the definition of dietary fiber, with
the exception of its selectivity for certain bacterial species and
a wider range of health effects. Peptides, proteins, and lipids
contain prebiotics characteristics, but some carbohydrates have
received the most attention, including lactulose, inulin, and a
range of oligosaccharides that supply a source of fermentable
carbohydrate for the beneficial bacteria in the colon (Prado et al.
2008).
Cheese
400 types of cheese.
Natural vs. ripened Cheese :
Mild flavor, High moisture content (30-55%), pH 5-5.3,
Process:
Standardization of milk:
Casein: fat ratio, Cal. Chloride (0.1%) to facilitate coagulation with
rennet.
Coagulation of Proteins:
Aggregation of protein network (casein micelles) entraps fat and
fat/water . Curd and starter culture bind to this network and yield
no coagulation below 10C and above 65C. 40-45C ideal temp.
Cutting the coagulum: (scalding) draining of whey, 20-55
Pressing and shaping: 20-27C, 95% RH
Salting: 0.7-4% salt.
Ripening: Degradation of fats/proteins lipids to volatiles. 3 wks3years. 4-20C temperatures. Soft cheese at 4C.
Cheese (Fresh)
Yoghurt