Browning is a process that produces brown color in foods through enzymatic or non-enzymatic reactions. Enzymatic browning is caused by polyphenol oxidase (PPO) enzymes acting on phenolic compounds in fruits and vegetables, producing quinones that polymerize to form brown melanoidins. It can impact food quality and market value. PPO activity can be controlled by excluding oxygen, denaturing the enzyme through heat or pH reduction, chelating the copper cofactor, or preventing reactions with phenolic substrates. Common prevention methods in food industry include oxygen exclusion, heat treatment, acidification, and sulfur dioxide application.

1. BROWNING
As a food process

2. Browning is a process that
produces a brown color in
food.

3. Importance of browning reactions
in food systems
• Browning may increase the acceptability of food by
developing appropriate flavor and color in food.
For example, flavor development in tea, flavor and color
development in figs and raisins, toast production in bread.
• Browning may cause food quality deterioration and thus
a decline in the market value of food.
Since color is the first sensory quality by which food is judged,
browning after peeling/slicing fruit and vegetables, mushroom
discoloration, and blackspots in shrimps and lobsters have great
economic cost.

4. Two Types of Browning
• Enzymatic browning
• Non-enzymatic browning

5. Enzymatic Browning
phenolases
A chemical process involving and
other enzymes that produce benzoquinones
quinones
and melanins from natural phenols
phenols, resulting in
a brown color in fruit and vegetables.

6. - enzymes that act on the
• Phenolases
substrates to produce the brown products
- substrates acted upon by the
• Phenols
enzymes to produce the brown products
– the brown products produced
• Quinones
by the enzymes from the substrates

7. • EB is one of the most important color
reactions affecting fruit and vegetables, and
seafoods.

8. Desirable effects of EB
• Flavor and color development in tea, coffee,
cocoa, and dried fruits
• Production of melanoidins, which may exhibit
antibacterial, antifungal, anticancer, and
antioxidant properties.
• Wound healing and post-molting hardening of
the shell (sclerotization) in insects and
crustaceans such as shrimp and lobsters.
• Fermentation

9. Undesirable effects of EB
• Hinders ease of processing fruit slices and
juices
• Unwanted color changes in lettuce and other
green leafy vegetables; potatoes and other
starchy staples such as sweet potato,
breadfruit, and yam; mushrooms; fruits such
as apples, avocados, bananas, grapes, olives,
and peaches; and crustaceans

10. The Enzymes

11. Enzymes involved in EB are collectively called
PHENOLASE and, since their discovery, their
international nomenclatures have undergone
marked changes.

12. Phenolase
• An oxidoreductase enzyme that catalyzes
the oxidation of phenols and other related
substances.
• This enzyme is found in many plants, fungi and
microorganisms. It catalyzes the oxidation of
certain molecules such as tyrosine.
• The action of this enzyme is evident
in fruit browning when fruits such as apples
and bananas are cut or bruised.

13. PHENOLASES
• Polyphenol oxidase initiates browning
• Catechol oxidase syn. diphenol oxidase)
• Laccase

14. Polyphenol oxidase (PPO)
• Systematic name: 1,2-Benzenediol:oxygen
oxidoreductase (EC 1.10.3.2)
• Synonyms: phenoloxidase, phenolase,
monophenol oxidase, diphenol oxidase,
tyrosinase, catechol oxidase
• An oxidoreductase, with oxygen as the
hydrogen acceptor, that acts on phenols
• Requires the presence of BOTH a copper
prosthetic group and oxygen for its activity

15. Two basic reactions catalyzed
by PPO
• o-Hydroxylation of monophenols to o-diphenols
(cresolase or monophenol oxidase activity)
E.g., oxidation of catechol to o-benzoquinone
• Oxidation of diphenol to o-benzoquinones
(catecholase or diphenol oxidase activity)
E.g., oxidation of L-tyrosine to 3,4-
dihydroxyphenylalanine (occurs in potatoes)

16. PPO-Catalyzed Reactions

17. Catechol Oxidase (CO)
• Systematic name: 1,2-Benzenediol:oxygen
oxidoreductase (EC 1.10.3.1)
• Synonym: Diphenol oxidase
• Copper-containing enzyme with a similar activity
to tyrosinase (EC 1.14.18.1)
• Catalyzes, using dioxygen, the oxidation of a broad
range of o-diphenols such as catechol to the
corresponding o-quinones coupled with the
reduction of oxygen to water. The
yellow compound produced (benzoquinone) is)
then oxidized in air to form dark-brown melanin.
• Found in fruits (e.g., banana, apple, and pear

18. Laccase (LAC or DPO)
• p-Diphenol oxidase or urushiol oxidase (EC
1.10.3.2) (DPO or LAC)
• Copper-containing oxidase that catalyzes the
oxidation of p-diphenols and o-diphenols
• Also oxidizes 4-benzenediol to 4-benzosemiquinone
• Occur in many phytopathogenic fungi, higher
plants, peaches and apricots
• Is involved in lignin degradation, pigment
biosynthesis and detoxification of lignin-derived
products.

19. PPO and CO oxidize ONLY o-diphenols,
whereas LAC oxidizes BOTH o-phenols and p-phenols.

20. PEROXIDASE (POD)
A hemoprotein that catalyzes the
oxidation of phenolic substrates through
the associated reduction of hydrogen
peroxide in the peroxidative cycle that
produces reactive oxygen species such as
a superoxide anion (O2
-●) or a hydroxyl
radical (OH●)

21. Peroxidase
• Oxidoreductase acting on phenols in the
presence of H2O2

22. Reactions Catalyzed by PPO and POD

23. Common Phenolic Compounds
in Food Material
1. Simple phenols - include mono-, o-di- and tri-phenols
E.g., L-tyrosine, catechol, and gallic acid
2. Cinnamic acid derivatives
E.g., chlorogenic acid, p-coumaric acid, caffeic acid,
ferulic acid, sinapic acid
3. Flavonoids - structurally related to flavones
E.g., catechins, leucoanthocyanidins (food tannins),
anthocyanins, and flavonols

24. Structures of Phenolic Compounds
•

25. •

26. •

27. Food sources of Phenolic Compounds
Source Phenolic Compounds
Apple Chlorogenic acid (flesh), catechin
(peel), catechol, DOPA
Avocado 4-Methyl catechol, dopamine,
pyrogallol
Cacao Catechins, anthocyanins
Coffee beans Chlorogenic acid, caffeic acid
Lobster, shrimp Tyrosine
Tea Flavonols, tannins, cinnamic acid

28. EB in
Schematic
Summary
Phenolase
Phenolics
Quinones

29. How do you control EB?
Answer: Inhibit PPO activity

30. Inhibition of PPO Activity
1. Exclusion of reactants such as oxygen
2. Denaturation of enzyme
3. Interaction of copper prosthetic group
4. Interaction with phenolics or quinones

31. (1) Oxygen Exclusion
• Oxygen exposure prevention, the simplest method
of which is water immersion.
• The method is limited for fruit and vegetables as
they will brown upon air re-exposure or via oxygen
occurring naturally in plant tissue.
• May lead to anaerobiosis in case of extended
storage of fruits and vegetables, which in turn may
lead to tissue breakdown

32. Inhibition of PPO Activity
1. Exclusion of reactants such as oxygen
2. Denaturation of enzyme
3. Interaction of copper prosthetic group
4. Interaction with phenolics or quinones

33. (2) Enzyme Denaturation
• Heat treatment
PPOs work at room temperature.

34. Optimal Temperatures for Activity of
PPOs from Different Sources
Source Optimal Temp (oC)
Apricot 25
Banana 37
Apple 25-30
Grape 25-30
Potato 22

35. (2) Enzyme Denaturation
• Heat treatment
PPOs work at room temperature.
• pH reduction
Optimum pH range of most PPOs: 4-7
• Application of powerful PPO inhibitors

36. PPO Inhibitors
• Cinnamon acid, benzoic acid, ascorbic acid
in apple juice
• Carbon monoxide in mushrooms
• 4-Hexylresorcinol in shrimp
• Inorganic halides
• Sodium chloride
• Zinc chloride + calcium chloride, ascorbic
acid, citric acid

37. Inhibition of PPO Activity
1. Exclusion of reactants such as oxygen
2. Denaturation of enzyme
3. Interaction of copper prosthetic group
4. Interaction with phenolics or quinones

38. (3) Binding of Copper Prosthetic Group
• Addition of complexing agents that binds to
the copper prosthetic group
E.g., ethylenediaminetetraacetate (EDTA)
diethyldithiocarbamate (DIECA), sodium
azide, potassium ethylxanthate, sodium
acid pyrophosphate, and citric acid
• Cu2+ is essential for PPO activity. Thus,
chelating Cu2+ will inhibit PPO activity

39. Inhibition of PPO Activity
1. Exclusion of reactants such as oxygen
2. Denaturation of enzyme
3. Interaction of copper prosthetic group
4. Interaction with phenolics or quinones

40. (4) Prevention of Action on Polyphenol
• Addition of polyvinylpolypyrollidone (PVPP),
which bind polyphenols, thereby eliminating
substrate of PPO, preventing browning
• Addition of compounds with similar chemical
structures to o-diphenols but are not PPO
substrates
Guaiacol Resorcinol Phloroglucinol

41. Methods of EB prevention used in
the Food Industry?
• Oxygen exclusion
• Heat treatment
• Acid treatment
• Application of sulfur dioxide and sulfites

42. Oxygen Exclusion
1. Water immersion
2. Use of O2-impermeable packages
3. Use of edible films such as sulfated
polysaccharides, e.g., carrageenan, amylose
sulfate, and xylan sulfate as wrappers
4. Prevention of mechanical bruising during
shipping of fresh fruits to prevent O2
exposure of fruit flesh

43. 5. Use of N2 headspace in packaging
6. Reduced oxygen packaging (i.e., vacuum
packaging, modified atmosphere packaging,
controlled atmosphere packaging)
But not too low an O2 concentration:
• Off-flavor production by anaerobic
glycolysis
• Risk of Clostridium botulinum growth

44. Heat Treatment
1. Blanching
2. Water immersion at 93oC for 2 min.
3. Steaming and water immersion at 70-105oC
4. Pasteurization at 60-85oC
5. Refrigeration, e.g., freezing at -18oC
High-temperature inactivates PPO. Low
temperature retards PPO activity.
The heat inactivation of PPO is dependent on
time and pH.

45. Acid Treatment
1. Addition of acids occurring naturally in plants, e.g.,
citric, malic, phosphoric and ascorbic acids
[Ascorbic acid is a very effective PPO inhibitor. At its
level used in the industry, it has no detectable flavor
or corrosive action on metals.]
• Used extensively in the food industry
• Based on the fact that lowering plant tissue pH
will retard EB
• Sodium acid pyrophosphate has been suggested
as an alternative to organic acids. SSAP is less sour
than most organic acids and minimizes after-cooking
blackening in potatoes.

46. Sulfur Dioxide and Sulfite Application
• Powerful reducing agents and PPO inhibitors that can be
applied in gaseous or solution form.
• Applicable in cases where heating will cause undesirable
effects, e.g., textural changes and off-flavor
• Disadvantages: Off-flavor and off-odor development,
bleaching of natural food pigments, hastening of can
corrosion, degradation of Vitamin B1, and toxicity at
high levels
• Advantages: High effectivity, preservation of Vitamin C,
and low cost

47. •

48. Fill in the blanks:
Enzymatic Browning
A chemical process involving __________ and
other enzymes that produce ____________
and melanins from natural _______, resulting
in a brown color in fruit and vegetables.

49. NON-ENZYMATIC
BROWNIING

50. NON-ENZYMATIC BROWNING (NEB)
A chemical process that produces a
brown color in food without
involving
enzymes

51. TYPES OF NEB
1. Maillard reaction
2. Caramelization
3. Ascorbic acid oxidation
4. Lipid browning

52. MAILLARD REACTION
• A chemical reaction between a free amino
group from amino acids, peptides, or proteins
and the carbonyl group of a reducing sugar
• A NEB reaction, caused by the condensation of
an amino group and a reducing sugar,
resulting in complex changes in biological and
food systems

53. FEATURES OF
MAILLARD REACTION
• First described by Louis Maillard in 1912
• Generally requires heat addition, BUT also
occurs during storage
• Favored under alkaline condition

54. ADVANTAGES OF
MAILLARD REACTION
• Development of caramel aroma
• Development of golden brown color
• Formation of antioxidative Maillard
reaction products (e.g., in honey and
tomato puree manufacture)

55. DISADVANTAGES OF MAILLARD REACTION
• Dark pigmentation
• Off-flavor development
• Deterioration of proteins during food processing
and storage
• Reduction in protein digestibility
• Loss of nutritional quality
• Formation of mutagenic and carcinogenic
compounds during frying,
grilling, and baking of meat
(e.g., acrylamide)

56. ACRYLAMIDE
• IUPAC name is prop-2-enamide (C3H5NO).
• Considered a carcinogen
• Discovered accidentally in food in April 2002
by scientists in Sweden
• Forms at moderate levels (5–50 g/kg) during
heating of protein-rich food and at higher
levels (150–4000 g/kg) during heating of
carbohydrate-rich food

57. Mutagen from the Maillard Reaction
Asparagine + reducing sugar
Strecker aldehyde
Acrylamide

59. Aldose sugar + amino compound N-substituted glycosylamine + H2O
Amadori
rearrangement
1-amino-1-deoxy-2-ketose
Sugar dehydration
Sugar fragmentation
Strecker degradation
Volatile and nonvolatile monomers
Schiff base of HMF or furfural
Reductones
Fission products
Aldehydes
Others
MAILLARD
REACTION
Aldol condensation
Carbonyl amine
polymerization
Highly colored products
Aldols and N-free products
Melanoidins

60. THREE STAGES OF
THE MAILLARD REACTION
1. Early stage
2. Intermediate stage
3. Final stage

61. EARLY STAGE
• Condensation of primary amino groups of
amino acids, peptides, or proteins with the
carbonyl group of reducing sugars (aldoses) --
The carbonylamino reaction
• Formation of Amadori products via Schiff’s
base formation and Amadori rearrangement

62. EARLY STAGE
amino acid or protein + glucose
Schiff’s base b-pyranosyl
Amadori rearragement
Amadori products

64. AMADORI PRODUCTS
Alanine-fructose and leucine-fructose (precursors
of numerous compounds important in the
formation of characteristic flavors, aromas, and
brown polymers)
Hydroxyproline-fructose and tryptophan-fructose
Alanine-fructose Tryptophan-fructose
• These are formed before the occurrence of
sensory changes.

65. INTERMEDIATE STAGE
• Breakdown of Amadori compounds (or other
products related to Schiff’s base)
• Formation of degradation products, reactive
intermediates (3-deoxyglucosone), and
volatile compounds (flavor compounds)

67. HETEROCYCLIC COMPOUNDS
• Flavor compounds in processed foods, such
as beef products, soy products, processed
cheese, coffee, tea, potatoes
• Include pyrazines, pyrroles, oxazoles,
oxazolines, and thiazoles (formed from
sulfur amino acids)
• ALSO formed from Strecker degradation

68. STRECKER DEGRADATION
• Oxidative degradation of amino acids into
aldehydes in the presence of α-dicarbonyls or
other conjugated dibarbonyls formed from
Amadori compounds.
• NOT directly involved in pigment formation
• Aldehydes formed contribute to flavor
development

69. STRECKER DEGRADATION PATHWAY

70. FINAL STAGE
• Production of nitrogen-containing
brown polymers and copolymers known
as melanoidins
• Series of aldol condensation and
polymerization reactions

71. TWO MAJOR PATHWAYS FROM AMADORI
COMPOUNDS TO MELANOIDINS
Amadori
compounds

72. Aromas and volatile compounds produced from
L-amino acids in the Maillard reaction system
Amino acid Volatile compound Aroma
Alanine Acetaldehyde Roasted barley
Cysteine Thiol, H2S Meaty
Valine 2-Methylpropanal
Leucine 3-Methylbutanal Cheesy
Lysine Breadlike
Methionine Methional

73. FACTORS AFFECTING THE
MAILLARD REACTION
1. pH
2. Type of reducing sugar
3. Type of amino acid
4. Temperature
5. Concentration and ratio of
reducing sugar to amino acid
6. Water activity (Aw)
7. Metals

74. pH
• An increase in pH enhances Maillard reaction
Therefore, high acidity (that is, low pH)
makes food less susceptible to the reaction.
• Has a less dramatic effect on aroma than
temperature, time or water content
• The most desirable meaty and pot-roasted
aroma has been obtained at pH 4.7
• NOTE Buffer concentration also affects the
reaction: A higher buffer concentration leads to
a higher reaction rate.

76. TYPE OF REDUCING SUGAR
• Pentoses (e.g., ribose) react more readily than
hexoses (e.g., glucose)
• Hexoses are more reactive than disaccharides
(e.g., lactose)
• Glucose is more reactive than fructose
And therefore pentoses, hexoses and glucose
enhance Maillard reaction compared with their
counterparts.

77. TYPE OF AMINO ACID
•

78. TEMPERATURE
• An increase in temperature increases the
rate of browning.
• The temperature of a chemical reaction is
often expressed as the activation energy
(Ea), which is highly dependent on pH and
the participating reactants; thus, it is
difficult to isolate the effect of temperature
as a single variable.

79. CONCENTRATION AND RATIO OF
REDUCING SUGAR TO AMINO ACID
• Browning reaction rate increases with
increasing glycine:glucose ratio in the range
from 0.1:1 to 5:1 (Wolfrom et al., 1974).
• Using a model system of intermediate
moisture (aw, 0.52), Warmbier et al. (1976)
observed an increase in browning reaction
rate when the molar ratio of glucose to lysine
increased from 0.5:1 to 3.0:1.

80. WATER ACTIVITY (aw)
• The Maillard reaction occurs less readily in food
with high aw.
At a high aw, the reactants are diluted.
• BUT note that a low aw does not translate to a
high reaction rate. Note that, at a low aw, the
mobility of the reactants is limited, despite their
increased concentrations.
• The browning reaction rate is maximum in the
aw range from 0.5 to 0.8 in dried and
intermediate-moisture foods.

81. METALS
• Metals form metal complexes with amino
acids and for yet unexplained reasons such
complex formation hastens Maillard
reaction
• For instance, browning is accelerated by
Cu2+ and Fe3+.

82. TYPES OF NEB
1. Maillard reaction
2. Caramelization
3. Ascorbic acid oxidation
4. Lipid browning

83. CARAMELIZATION
• Pyrolysis of food carbohydrates by heat
treatment above the melting point of the sugar
under alkaline or acidic condition
• Involves only carbohydrates, not amines
• Occurs when surfaces are heated strongly, when
processing foods with high sugar content, or in
wine processing

84. DESIRABLE EFFECTS OF
CARAMELIZATION
• Pleasant caramel flavor
• Enticing brown color in some foods

85. UNDESIRABLE EFFECTS OF
CARAMELIZATION
• Formation of mutagenic compounds
• Excessive changes in sensory attributes

86. STEPS IN CARAMELIZATION OF
REDUCING SUGARS
1. Ring opening of the hemiacetal ring
2. Enolization via acid-base catalysis
3. Formation of isomers (aldose to ketose
interconversion; rate increases with
increasing pH)

87. ISOMER FORMATION STEP

88. STEPS IN CARAMELIZATION OF
REDUCING SUGARS
1. Ring opening of the hemiacetal ring
2. Enolization via acid-base catalysis
3. Formation of isomers (aldose to ketose
interconversion; rate increases with
increasing pH)
4. Dehydration - leads to the formation of
furaldehydes (e.g., hydroxymethylfurfural)

89. DEHYDRATION STEP

90. STEPS IN CARAMELIZATION OF
REDUCING SUGARS
5. Formation of fragmentation products such
as acetol, acetoin, and diacetylformic and
oxidation products such as acetic, and other
organic acids
6. Reaction of these products, forming brown
pigments and flavor compounds

91. FACTORS AFFECTING
CARAMELIZATION
1. pH
2. Temperature
3. Water activity
4. Type of sugar

92. 1. pH Reaction occurs faster under alkaline
condition than under neutral or acid condition.
The optimum pH for the reaction is 10.
2. Temperature Reaction is favored at >120oC.
Reaction rate increases from 80 to 110oC.
3. Aw Faster browning at Aw approaching 1 than at
Aw=0.75
4. Type of sugar Faster reaction with fructose than
with sucrose as well as with glucose than with
starch
Sugars with more reducing groups hastens
reaction.

93. TYPES OF NEB
1. Maillard reaction
2. Caramelization
3. Ascorbic acid oxidation
4. Lipid browning

94. ASCORBIC ACID OXIDATION (AAO)
• Ascorbic acid browning
• Spontaneous thermal decomposition of
ascorbic acid under both aerobic and
anaerobic conditions, by oxidative or
nonoxidative mechanisms, in either the
presence or absence of amino compounds
• Observed in citrus, asparagus, broccoli,
cauliflower, peas, potatoes, spinach, apples,
green beans, apricots, melons, strawberries,
corn, and dehydrated fruits

95. FACTORS AFFECTING AAO
1. Temperature
2. Salt and sugar concentration
3. pH
4. Oxygen
5. Enzyme (ascorbic acid oxidase)
6. Metal catalysts (Cu2+ and Fe2+)
7. Amino acids
8. Oxidants or reductants
9. Initial concentration of ascorbic acid
10. Ratio of ascorbic acid to dehydroascorbic acid

96. Anaerobic Aerobic

97. TYPES OF NEB
1. Maillard reaction
2. Caramelization
3. Ascorbic acid oxidation
4. Lipid browning

98. LIPID BROWNING
• Oxidative deterioration of unsaturated
glycerides followed by polymerization
accelerated by ammonia, amines or proteins
• Protein browning caused by reaction of
acetaldehyde (derived from unsaturated
lipids) with protein-free amino groups, by
repeated aldol condensations
• Protein-oxidized fatty acid reactions

100. LIPID BROWNING
• First observed in discoloration of white fish
muscle during frozen storage
• May be non-enzymatic or enzymatic
• Its first stage is lipid oxidation, which produces
hydroperoxides as the initial products
• Via polymerization, brown oxypolymers can
be produced subsequently from lipid
oxidation derivatives

101. LIPID BROWNING
• Oxidized products can also interact with free
amino groups of amino acids, peptides,
proteins
• Observed during storage and processing of
some fatty foods, salted sun-dried fish, boiled
and dried anchovy, smoked tuna, meat and
meat products, and rancid oils and fats with
amino acids or proteins

102. Comparison of Mechanisms of
Nonenzymatic Browning
Mechanism Requires O2 Requires amino group ph optimum
in initial reaction
Maillard reaction − + Alkaline
Caramelization − − Alkaline, acid
Ascorbic acid + − Slightly acid
oxidation

103. LIPID BROWNING
• Reaction of 4,5-epoxy-2-alkenals (formed during
lipid peroxidation) with the amino group of
amino acids or proteins
• Always accompanied by the production of N-substituted
pyrroles (II), which are stable
• N-Substituted 2-(1-hydroxyalkyl) pyrroles are
also formed, but are unstable; they polymerize
rapidly and spontaneously to produce brown
macromolecules with fluorescent melanoidin-like
characteristics

106. DESIRABLE EFFECT OF LIPID
BROWNING
• Produces lipid-amino acid reaction products
that exert antioxidant properties when
added to vegetable oils

107. UNDESIRABLE EFFECT OF LIPID
BROWNING
• Loss of nutritional quality due to the
destruction of essential amino acids such as
tryptophan, lysine, and methionine and of
essential fatty acids.
• Decrease in digestibility and inhibition of
proteolytic and glycolytic enzymes

108. CONTROL OF NONENZYMATIC
BROWNING
1. Addition of sulfites, thiol compounds,
maltitol, sugars, and sorbitol
2. Modified atmosphere packaging
3. Microwave heating
4. Ultrasound assisted thermal processing
5. Pulsed electric field processing
6. Carbon dioxide-assisted high-pressure
processing

109. END