further decomposition of hydroperoxides
TRANSCRIPT
Further decomposition of hydroperoxides
C-C-C=C-CHO
C-C-C=C-CHO O O H
R-CHO + OHC-CH2-CHO (malonaldehyde)
Formation of malonaldehyde is one of the major
products of lipid oxidation.
Malonaldehyde can cross-link with proteins,
enzymes and DNA and cause health problems.
Thiobarbituric acid (TBA) test
Measuring TBARS (Thiobarbituric acid reactive
substances) is a general test used to evaluate the
extent of lipid oxidation.
Oxidation products of unsaturated systems produce
a colour reaction with TBA.
Colour results from condensation of two molecules
of TBA and one molecule of malonaldehyde.
2 mol TBA + malonaldehyde red colour
The product can be measured quantitatively at 530
nm using a spectrophotometer.
Role of metal ions in lipid oxidation
Metal ions can catalyze the oxidation of lipids.
Metals possessing two or more valency states and a
suitable oxidation-reduction potential between them
are effective pro-oxidants. e.g. Fe, Cu, Mn, Co
Even at concentrations as low as 0.1 ppm, they can
decrease the induction period and thereby increase
the rate of oxidation.
Trace amounts of heavy metals are found in edible
oils originating from:
• the soil in which the plant was grown
• the animal
• metallic equipment used in processing or storage
Role of metal ions in lipid oxidation
They are also naturally occurring components of all
foods and are present in both free and bound forms.
Mechanisms for metal catalysis of oxidation are as
follows.
1. Activation of molecular oxygen to give singlet oxygen
and peroxy radical.
-e- 1O2
Mn+ + O2 M(n+1) + 3O2
+H+
HOO0
2. Direct reaction with the unoxidized substrate.
Mn+ + RH M(n-1)+ + H+ + Ro
Role of metal ions in lipid oxidation
3. Acceleration of hydroperoxide decomposition
Mn+ + ROOH M(n+1)+ + OH- + ROo
Mn+ + ROOH M(n-1)+ + H+ + ROOo
Lipoxydase (Lipoxygenase) catalysed oxidation:
This is an enzymatic reaction which needs O2.
1,4 pentadiene structure is also needed.
RH + Lipoxydase + O2 RH + O2
Lipoxydase
ROOH ROOH R + OOH
+ Lipoxydase Lipoxydase
Lipoxydase
Lipoxygenase catalysed oxidation ……
• Lipoxygenase is present in plants and animals.
• Activation energy for the above reaction (3-4 kcal/
mol) is low compared with autooxidation hence it can
take place at low temperature, even at refrigeration
temperature.
• Under frozen conditions aw is low and the mobility of
reactants is low. Hence the rate of reaction reduced.
• Enzymatic cleavage of ROOH yields a variety of
breakdown products which are responsible for the
characteristic flavor of natural products.
• This reaction can be inhibited by phenolic anti-
oxidants (tocopherol, hydroquinones etc.)
Hematin catalysed oxidation ……
• Hematin compounds present in many food tissues
are also important pro-oxidants. E.g. myoglobin,
haemoglobin, cytochrome
• Even in well bled tissues very low amount of hematin
is present. Fe3+ is involved in this reaction.
• This reaction is different to other 2 forms due to the
requirement of pre-formed hydroperoxides.
ROOH + hematin
RH
Ro + carboxyl compounds + hematin
The activation energy for reaction is 3.3 kcal /molecule.
ROOH + hematin
Antioxidants
• Substances that can delay the onset or slow down
the rate of oxidation
• Main lipid soluble antioxidants used in foods are
monohydric or polyhydric phenols
E.g. Tocopherol, BHT, BHA, PG, TBHQ
• For maximum efficiency, they are used in combination
with metal sequestering agents
Mechanism of action:
A substance delays autooxidation reaction, if;
• it inhibits formation of free radicals
• it interrupts propagation of free radicals
Antioxidants
• Antioxidants inhibit free radical formation by:
• Quenching singlet oxygen
• Chelating metal ions
• Decomposing hydroperoxides
• An antioxidant inhibits the chain reaction by acting as
hydrogen donor (free radical acceptor) for Ro and
ROOo radicals.
Ro + AH RH + Ao
ROOo + AH ROOH + Ao
• The resulting antioxidant radical will not initiate new
free radicals and they may undergo a variety of
reactions forming stable products.
Antioxidants…..
Ao + Ao AA
Ao+ ROOo ROOA
Dihydric phenols dismute to yield quinones with the
formation of original antioxidant.
Ao + Ao AH + quinone
Effectiveness of an antioxidant is influenced by its,
• Chemical potency
• Solubility in oil (accessibility to free radical)
• Volatility (stability during heating, storage)
Synergism of Antioxidants
• Synergism occurs when a mixture of antioxidants
produces a greater activity than the sum of the
activity of each antioxidant in the mixture, when
tested individually.
• Two types of synergism are recognized:
1. Action of mixed free radical acceptors:
ROOo + AH ROOH + Ao
Ao + BH AH + Bo
The presence of the second antioxidant (BH) will
have a sparing effect since it regenerates the primary
antioxidant (AH).
E.g. Phenolic antioxidant and ascorbic acid
Synergism of Antioxidants
• Phenolic antioxidant is the primary antioxidant (more
effective one) while ascorbic acid is the synergist.
• It is possible for two phenolic antioxidants to exhibit
synergism in a similar way.
2. Combined action of a free radical acceptor and a
metal chelating agent:
• Metal chelating agents are compounds which can
partly deactivate trace metals present.
• When antioxidant property of a free radical acceptor
is enhanced by the presence of a metal chelating
agent synergism occurs. E.g. citric acid, phosphoric
acid, polyphosphates.
Choice of the Antioxidant
• Antioxidants exhibit substantial differences in their
effectiveness when used with different types of fatty foods
and under different processing and handling conditions.
• Factors to be considered in selecting an anti-oxidant are;
• Chemical potency of the antioxidant
• Ease of incorporation into the food
• Carry-through characteristics
• Sensitivity to pH
• Hydrophilic-lipophilic properties
• Tendency to produce off-flavour or off-colour
• Availability
• Cost
Choice of the Antioxidant
• In bulk oils – TBHQ and PG are more effective.
• In oil-water emulsions, polar lipid membranes,
intracellular micelles of neutral lipids - more lipophilic
antioxidants, such as BHA, BHT and tocopherols are
the most effective.
Thermal Decomposition
• Heating of food produces various chemical changes,
some of which are important to flavor, appearance,
nutritive value and toxicity.
• Different nutrients in food undergo decomposition
reactions and also interact among themselves in
extremely complex ways to form a large number of
new compounds.
• Lipid oxidation at high temperatures is complicated:
thermolytic and oxidative reactions taking place
simultaneously.
• Both saturated and unsaturated fatty acids undergo
decomposition when exposed to heat.
Thermal decomposition
I. Thermal, non-oxidative reactions of SFA:
Heating of saturated triglycerides to >200 oC yields
detectable amounts of hydrocarbons, acids and
ketones due to thermolysis.
II. Thermal, oxidative reactions of SFA:
Even though SFA are more stable to heat than their
unsaturated analogs, above 150 oC they can also
undergo oxidation, giving rise to a complex decomposition
pattern.
Major oxidative products are, series of carboxylic acids
and hydrocarbons: 2-alkanones, n-alkanals, n-alkanes,
1-alkenes and lactones.
Thermal decomposition
III. Thermal non-oxidative reactions of USFA:
Unsatutared fatty acids form dimeric compounds and low
molecular weight compounds during high heat in the
absence of oxygen.
IV. Thermal oxidative reactions of USFA:
At elevated temperatures oxidative decomposition of
USFA takes place very rapidly.
Major compounds formed at high temperatures are
qualitatively the same as that of room temperature
autooxidation. But at elevated temperatures
hydroperoxide decomposition and secondary oxidation
are extremely rapid.
Fatty acids, esters
and triglycerides
Saturated Unsaturated
Thermolytic Oxidative Thermolytic Oxidative
reactions reactions reactions reactions
(α,β,γ attack)
acids alkanes acyclic and volatile and
hydrocarbons alkenes cyclic dimeric
propenediol alkanals dimers products of
acrolein lactones autooxidation
ketones carboxylic acids
Generalized scheme for thermal decomposition of lipids
Chemistry of Frying
• Foods fried in oil, contribute significantly to the
energy in the diet because 5-40% of the oil can be
absorbed to food.
• During deep-fat frying, foods contact oil at high
temperatures (around 180 oC) and is exposed to air
for a variable period of time.
• Thus frying has the greatest potential for causing
chemical changes in food.
Chemistry of Frying
Behaviour of the frying oil:
• The physical and chemical changes that can be
observed in the oil during frying include;
• Increase in viscosity
• Increase in free fatty acid content
• Development of a dark colour
• Decrease in iodine value
• Decrease in surface tension
• Changes in refractive index
• Increased tendency to foam
Above changes are due to following classes of
compounds produced from the oil, during frying.
Chemistry of Frying……
1. Volatiles:
• Oxidative reactions involving the formation and
decomposition of hydroperoxides, lead to the
formation of saturated and unsaturated aldehydes,
ketones, hydrocarbons, lactones, alcohols, acids
and esters.
• Volatiles produced vary widely depending on the type
of oil, type of food and the heat treatment.
• They reach a plateau value with time probably
because a balance is achieved between the formation
and loss of volatiles.
Chemistry of Frying
2. Non-polymeric polar compounds of moderate
volatility (E.g. hydroxyl and epoxy acids):
These compounds are produced due to various oxidative
pathways involving the alkoxy radical.
3. Dimeric- and polymeric acids, and dimeric- and
polymeric glycerides:
These compounds occur from thermal and oxidative
reaction combinations of free radicals. Polymerization
results in an increase of viscosity of the frying oil.
4. Free fatty acids:
These compounds arise from the hydrolysis of
triglycerides in the presence of heat and water.
Chemistry of Frying
Behaviour of the food during frying:
• Water is continuously released from the food into the
hot oil. This produces a steam distillation effect,
sweeping volatile oxidative products from the oil.
• Released moisture also agitates the oil and hastens
the hydrolysis making more FFA available.
• Blanket of steam produced above the surface of oil
tends to reduce the amount of oxygen available for
oxidation.
• Volatiles may develop in the food itself or from the
interaction between food and oil.
Chemistry of Frying
• Food absorbs varying amount of oil during deep fat
frying. E.g. in potato chips the final fat content is
about 35 %.
• Food itself may release some of its endogenous lipids
(e.g. fat from chicken) into the frying medium and
consequently the oxidative stability of the new mixture
may be different from that of the original oil.
• The oil may get darken at an accelerated rate due to
the presence of food.
• Extensive decomposition due to uncontrolled frying
operation can be a potential source of damage to
sensory properties, nutritive value and safety of food.
Browning Reactions in Foods
Browning reactions in foods are of 2 major types:
1. Enzymatic:
2. Non-enzymatic:
• Maillard browning
• Caramelization
• Ascorbic acid oxidation
Enzymatic browning (phenolase / oxidative browning):
• Enzymatic browning is the common form of browning
occurring in fruits and vegetables when they are
damaged or bruised. E.g. apple, banana, pear, guava,
avocado, mango, grapes, potato, brinjal, lettuce etc.
Also found in shrimp and lobsters.
• It is the reaction between oxygen and a phenolic
substrate catalyzed by the enzyme phenolase.
Enzymatic browning
• It is a major contributor to the desirable colour of tea,
apple juice, cocoa and cider.
• Responsible for the normal colour of raisins, prunes,
dates and figs.
• Phenolase activities in fruits and vegetables are not
desirable because the ensuing brown colour is not
pleasing. Enzymatic browning is detrimental to quality,
particularly in post-harvest storage of fresh fruits, juices
and some shellfish.
• In intact plant tissues phenolic substrates are separated
from phenolase and browning does not occur.
• In cut surfaces of light-coloured fruits and vegetables
enzymatic browning is prominent.
Enzymatic browning …..
• Exposure of cut surface (cutting, peeling, bruising) to
air results in rapid browning due to oxidation of
phenols to ortho-quinones, which then rapidly
polymerize into brown colour melanin pigments.
• Enzymes catalyzing oxidation of phenols are known
as “phenolase” and it includes phenolases,
polyphenol oxidases, tyrosinases or catecholases.
• Phenol enzymes have a copper prosthetic group.
• Phenolases catalyse two types of reactions.
Enzymatic browning ……
1. Hydroxylation reaction (known as phenol hydroxylase or
cresolase activity)
2. Oxidation reaction (known as polyphenol oxidase or
catecholase activity)
• First reaction results in ortho-hydroxylation of a phenol and
the second, oxidation of the diphenol to an ortho-quinone.
• Phenolases have a Cu prosthetic group.
Substrates:
1. Monophenols – e.g. tyrosine
2. Ortho-diphenols – e.g.
• catechol
Enzymatic browning ……
• Caffeic acid
• Protocatechuric acid
• Chlorogenic acid
3. Flavanoids in apple – e.g. rutin, quercetin
4. Tannins in peach, tea
5. Catechins in tea
Reaction:
• When tyrosine is the substrate, phenolase first catalyzes its
hydroxylation to DOPA (L-3,4 dihydroxy phenylalanine) and
subsequently catalyzes oxidation of DOPA to DOPA quinone.
Enzymatic browning …..
enzyme + H2O enzyme +O2
Tyrosine DOPA DOPA quinone fast
fast
Hallochrome (red) Leuco-compound
polymerization
Indole 5,6 quinone Melanin
• DOPA quinone formation is enzyme and O2 dependent.
• After the quinone step reactions will proceed without the
involvement of enzymes.
• From hallochrome onwards a gradual change of colour
occurs.
• Melanin, the final product, can interact with protein to form
complexes.
Enzymatic browning …..
• Hydroxylation of monophenol is the rate-limiting step.
• Some phenolase enzymes (e.g. in tobacco, tea and sweet
potato) do not hydroxylate monophenols but some (e.g. in
mushrooms and potato) perform both functions.
• Ortho-diphenols are readily attacked by catecholase
component of the phenolase.
• Meta-diphenols (e.g. rescorcinol) do not participate in the
oxidation reaction. It acts as an inhibitor.
Rescorcinol (benzene-1,3 diol)
• Para-diphenols (e.g. quinol) can act as a substrate but the
reaction rate is slower than ortho-diphenol since monophenol
hydroxylation reaction is required.
Quinol (benzene-1,4 diol)
• Although enzymatic browning is desirable in tea and apple
juice industries, in most fruits and vegetables and frozen
products, it should be prevented.
Methods to control enzymatic browning:
1. Blanching to destroy the enzyme, most commonly used method
2. HTST Pasteurization for fruit puree and not for fruit slices
(textural changes occur).
3. For potato, microwave heating causes more rapid inactivation
of phenolases than hot water treatment.
4. Dipping in water – limits O2 access to cut surfaces. E.g. potato
chips. Textural and flavour changes may occur.
5. Surface treatment of fruit slices (apple, pear, peach) with
excess ascorbic acid (antioxidant).
6. Vacuum packaging to exclude air.
7. Immersing in sucrose solution before freezing of fruits
(reduces dissolved oxygen).
8. Reduction of pH by adding acids. Citric, malic, phosphoric and
ascorbic acids are used to lower pH. They also serve as
chelators for Cu. Addition of lemon juice or vinegar is also
effective to lower pH.
Optimum pH for the reaction is pH 5-7. Below pH 3, the
enzyme is irreversibly inactivated. In apple juice production
malic acid is used to get pale brown colour (rate of reaction is
only reduced by malic acid).
9. Use of 1% NaCl solution: Cl- inhibits enzymic browning.
10. Addition of sulphites: Na2S, KMS, SO2 to inactivate enzyme.
11. Methylation of the substrate using SAM (methyl donor) and
ortho-methyl transferase enzyme. E.g.
Catechol + SAM + enzyme guaicol
Caffeic acid + SAM + enzyme ferulic acid
12. High Pressure Processing (HPP):
HPP is a technique of food processing where food is subjected
to high pressure (500-700 atmosphere) to achieve microbial
and enzyme inactivation.
High pressure processing causes minimal changes in foods.
Compared to thermal processing, HPP results in foods with
fresher taste and better appearance, texture and nutrition.
13. Ultrafiltration:
Ultrafiltration is a membrane separation process, driven by a
pressure gradient. The membrane separates liquid
components according to their size and structure. In the food
industry this technique is applied for white wine and fruit juices.
Ultrafiltration is able to remove larger molecules like enzyme
polyphenol oxidase, but not lower-molecular-weight
compounds like polyphenols.
Maillard Browning
• Maillard browning is a non-enzymic browning reaction
that takes place during the processing or storage of
protein foods containing reducing carbohydrates or
carbonyl compounds (e.g. aldehydes or ketones derived
from lipid oxidation).
• The minimum reactant requirements for Maillard browning
are the presence of an amino-bearing compound, usually
a protein, a reducing sugar and some water.
• The rate of Maillard reaction is markedly enhanced during
cooking, heat processing, evaporation and drying.
Maillard Browning
The Maillard reaction is responsible for many colors
and flavors in foodstuffs such as:
• caramel made from milk and sugar
• the browning of bread into toast
• the color of beer, chocolate, coffee, and maple
syrup
• the flavor of roast meat
• roasted nuts
• the color of dried or condensed milk
Maillard Browning
• Reaction rates are greatest in foods with an intermediate
moisture content such as roasted nuts, toasted breakfast
cereals and roller-dried milk powders.
• Maillard reaction begins with condensation of a non-
ionized amino group (terminal α-NH2 group of amino
acid) and a reducing sugar (aldose or ketose).
• During the first step, aldosylamines or ketosylamines are
formed by aldose and ketose sugars, respectively.
• Next, aldosylamines are transformed by „Amadori
rearrangement‟ into ketosamines while ketosylamines are
transformed by „Heyn‟s rearrangement‟ into aldosamines,
both of which are stable compounds.
Maillard Browning
Aldose sugar carbonyl Aldosyl Ketosamine
+ amino acid amine amine (Amadori
product)
Ketose sugar carbonyl Ketosyl Aldosamine
+ amino acid amine amine (Heyn‟s
product)
• During the second step, ketosamines and aldosamines
evolve into numerous carbonyl and polycarbonyl
unsaturated derivatives, such as reductones.
R – C = C – C - R‟
| | || - Reductone
OH OH O
Maillard Browning
• Some of these derivatives may react with amines and amino
acids leading to formation of ammonia and new carbonyl
compounds (Strecker degradation).
• Decarboxylation of free amino acids may also take place.
• Commercially the Strecker degradation is used to produce
distinctive flavours of chocolate, honey, maple syrup and
bread. Common Strecker aldehydes include ethanal (fruity,
sweet aroma), methylpropanal (malty) and 2-phenylethanal
(flowery/honey like aroma). Thus at times Maillard and
Strecker reactions are favourable.
• In the next step, polycarbonyl unsaturated derivatives undergo
both scission and polymerization reactions leading on one
hand to volatile compounds (aldehydes, pyrazines) and on the
other hand to brown or black pigments known as melanoidins,
with high molecular weight and complex structures.
Maillard Browning
• Such pigments are responsible for the colour of bread
and bakery products.
• It is important to control both Maillard and Strecker
reactions because of their adverse contribution to flavour
and odour and possible toxicity of degradation products.
• `Premelanoidin‟ products may also contribute to
nitrosamine formation, which is mutagenic.
Factors affecting the rate of browning
• Browning reaction occurs at a pH range of 7.8 - 9.2. In
less than pH 6, browning is reduced. In strong acidic
solutions browning does not occur.
• Browning is high at intermediate water levels.
• Copper and iron enhance browning. Fe3+ is more effective
than Fe2+
• Sugar type has an effect on the rate of Maillard reaction.
Pentoses > hexoses > disaccharides
• Among hexoses fructose is less reactive than aldoses.
Nutritional effects of Maillard reaction
1. Loss of biologically available lysine (in bread, milk). Other
basic amino acids such as L-arginine and L-histidine are
also susceptible.
2. Amadori products may inhibit intestinal absorption of
certain essential amino acids.
3. Formation of melanoidins destroys the digestibility of the
protein fraction of the molecules.
4. Melanoidins may be carcinogenic although they are
generally less absorbable (colloidal nature).
Methods to control Maillard browning
1. Decreasing moisture to very low levels or if
the food is a liquid, diluting it.
2. Lowering the pH
3. Lowering temperature
4. Removing one of the substrate, e.g. sugar
5. Addition of sulphur dioxide or sulphites.
Caramelization
• Direct heating of carbohydrates, particularly sugars and
sugar syrups, produces a complex group of reactions
called caramelization. It is used extensively in cooking for
the resulting nutty flavor and brown color. Caramelization
generally occurs at high temperatures (~150 °C), low
water content or high sugar content.
• Reactions are facilitated by small amount of acid and
certain salts.
• Thermolysis causes dehydration with formation of
anhydro rings or introduction of double bonds into sugar
rings.
• The latter produces intermediates to unsaturated rings,
such as furans. Conjugated double bonds absorb light
and produce colour.
Caramelization
• Maltol, isomaltol, hydroxyl methyl furans contribute to
the flavour and can be used to enhance various
flavours and sweeteners.
• In unsaturated ring systems, condensation will occur to
polymerize ring systems, yielding useful colours and
flavours.
• Catalysis speed up the reaction and often is used to
direct the reaction to specific types of caramel colours,
solubilities and acidities.
• Sucrose is commonly used to produce caramel colours
and flavours. It is heated in solution with acid or acidic
ammonium salts to produce a variety of products used
in food, candies and beverages (cola drinks, beer).
Caramelization
• Caramel pigments contain hydroxyl groups of varying
acidity, carbonyl, carboxyl, enolic and phenolic
hydroxyl groups.
• Rapid initial reaction is oxygen dependent and
proceeds until oxygen is completely exhausted.
• Increasing the temperature and increasing pH
increases the reaction rate. At pH 10 the reaction is 10
times faster than at pH 5.9.
• In the absence of buffering salts, humic substances
are formed which produce a bitter taste.
Ascorbic Acid Oxidation
• Decomposition of ascorbic acid is reported to be the major
deteriorative reaction occurring during the storage of orange
juice. Further, there exists a high correlation between the
percentage loss of ascorbic acid and an increase in browning
in grapefruit juices.
• Ascorbic acid breakdown in orange juice results in furfural
production and furfural buildup closely parallels quality loss in
citrus products.
• It has been demonstrated that amino acids accelerate ascorbic
acid breakdown, and in the presence of amine it is the
dehydroascorbic acid (DHA) that is the reactive intermediate in
the pathway to furfural and brown pigment production.
• If DHA has already been formed in juices brown color is
produced more intensely under non-oxidative conditions than
under oxidative conditions.
Ascorbic acid oxidation