Changes during ripening, mechanism behind the changes, Role of ethylene in ripening contact: dhota3@gmail.com

1. Mechanism of ripening in Fruit crops
Submitted to
Dr Prabhakar Singh
Professor and Head
Submitted by
Debashish Hota
Ph.D. 1st year

2. Course outline
1. What is ripening ?
2. Factors affecting ripening
3. Classification of fruit according to ripening
4. Role of ethylene in fruit ripening
5. Ethylene biosynthesis
6. Respiration
7. Fruit abscission
8. Changes during ripening
9. Regulation of ripening
10. Ethylene management
11. Case Study
12. Conclusion

3. • Fruit ripening is a highly co-ordinated,
genetically programmed, an irreversible
phenomenon and probably independent of one
another, involving a series of physiological,
biochemical, and organoleptic changes that
lead to the development of a soft and edible
ripe fruit with desirable quality attributes.

4. • Ripening is associated with change in
composition i.e.
• Conversion of starch to sugar.
• Change in colour
• Change in firmness
• Shape and size
• Odour /smell

6. Factors affecting ripening
1. Stage of development
2. Moisture content
3. Peel thickness
4. Type of tissue
5. Volatiles
6. Transpiration
7. Respiration
1. Temperature
2. Radiation
3. Air Humidity
4. Mechanical damage
5. O2 & CO2
concentration
6. Growth Substances
Internal factor External factor

8. Fruit classification according to ripening

9. Role of ethylene in fruit ripening
• Promotes its own biosynthesis
• External ethylene Hasten climacteric rise
(climacteric fruit)
• Accompanied by an increase in oxygen uptake
• This process is not reversed.
• External ethylene Increase Respiration
(Non-climacteric fruit)
• Ethylene production is largely pre-determined in both
time and amount by the genetics of the fruit
• During ripening both carbon dioxide and ethylene
increase significantly

12. Increase in rate of respiration
• There are two pathways in conversion of starch or sucrose to
glucose-6 P glycolysis
pentose phosphate pathway
Storedorganic materials Simple end product + Energy
(carbohydrates, proteins, fats)
Movement of fructose from the vacuole to the cytoplasm.
C₆H₁₂O₆+6O₂→6CO₂+6H ₂O+ENERGY
Respiration declines gradually throughout the season until
several weeks before it ripens
Just before ripening metabolic functions of the fruit are in a
near resting stage

13. FRUIT ABSCISSION
Degradation of
lipid bi-layers
Cell damage Senescence
Pedicel
Abscission
zone
Separation
fruit
Pedicel cells
Ethylene
signal
Enzyme
production
Pectinase
production
Weaken cell
wall
Fruit drop

14. Changes during ripening
1. Chlorophyll degradation
2. Hydrolysis of Starch to sugar
3. Textural Softening
4. Cell wall degradation
5. Production of volatiles compound
6. Changes in amino acids and protein
7. Involvement of different enzyme

15. Colour Changes
 The chlorophyll degradation is mainly due to: pH
changes, Oxidative systems, Chlorophyllase
 Formation of carotene in plastid
 Formation of anthocyanin in vacuoles and in epidermal
layer
 Masking effect of chlorophyll
 Degradation of chlorophyll into colourless product

16. • Replacement of Mg atom
in the chlorophyll by
hydrogen atom under
acidic conditions with the
formation of pheophytin.
• Pheophytin formation: a
colour change from bright
green to dull olive green
• Hydrolysis of chlorophyll
to chlorophyllin and phytol
catalyzed by chlorophyllase
followed by a replacement of
Mg atom with hydrogen
resulting in the formation of
pheophorbide

17. Chlorophyll Degradation

18. Fruits Pigment Colouring agent
Papaya
Yellow Pigment Caricaxanthin
Red Pigment Lycopene
Yellow orange Pigment Beta cryptoxanthin
Red fleshed of Solo variety Alpha carotene
Tamarind
Red Pigment Anthocyanin
Common Brown Pigment Leucoanthocyanin
Yellow Pigment Xanthophyll
Guava Red Colour Lycopene
Grape fruit Pink colour Lycopene
Mango Orange-yellow Caroteniod
Carambola Light Green Beta cryptoflavin and
mutatoxanthin
Phalsa Purple Delphinidine and Cyanidinide

19. Hydrolysis of starch to sugars
• Increase in the concentration of sugars, either by
hydrolysis of starch within the fruit (e.g banana,
mango, kiwi fruits etc.) or by continued import of
sugars from other part of the plant (e g. that ripen on
the vine, melon).
• In climacteric fruits, the starch content generally
increases during development but sucrose decreases
during storage due to its usage for metabolic
processes.

20. Starch solubilizing enzymes
Enzyme Mode of action
α-Amylase Hydrolysis α (1-4) linkage of amylase at
random to produce
mixture of glucose and maltose.
β-amylase Attacks penultimate linkages and
releases maltose
Starch phosphorylase Hydrolyses α (1-4) linkages to give
glucose-1, phosphate which can be
converted into glucose 6-phosphate by
the action of glucose phosphate mutase.
α (1-6) glucosidase Attacks α1-6 glucose linkage of
amylopectin

21. Textural Softening
Ripening probably depends on the species, changes in cell wall
composition, especially cell wall mechanical strength and cell-
to-cell adhesion, loss of turgor pressure, degradation of starch
etc.
Firm
ness
Crisp
ness
Meal
iness
Juici
ness
Hard
iness
Texture
Texture is controlled by the wall to wall adhesion of cells.
Breakdown
of starch
Breakdown
of pectic
substances
Reduced
cellular
rigidity
Softening
It is the result of enzymatic degradation of structural as well
as storage polysaccharides.

22. Cell wall degradation and softening of fruits
• Cell wall degradation involved:
– (i) Depolymerisation (shortening of chain length) and
– (ii) Deesterification
Complex
polysaccharides
Numerous
proteins
Plant cell
walls
Pectin(>50%) Cellulose Hemicellulose Polysaccharides
Increase in
soluble
pectin
Decrease in
insoluble
pectin
Decrease in
Firmness
Cell wall
degradation

23. • In ripening fruits, much attention was focused on
the depolymerization of acidic pectins by
polygalacturonase.
• De-esterification of polygalacturonic chains by
pectin methyl esterase (PME) make the chains
more susceptible to PG-degradation, facilitating
rapid loss of cell wall structure.
• PME activity were highest at green stage when
the fruit firmness was high and decreased as
ripening progressed, whereas both ethylene and
endo- (1 4) -D- mananase increased as ripening
proceed.

24. Production of aroma volatiles
• Esters of aliphatic alcohols and short chain fatty acids.
• In fruits, major volatile compounds are isoamyl acetate,
aldehydes and terpenoid compounds.
• Volatiles originate from proteins, carbohydrates, lipids, and
vitamins.
• Taste is provided by many non-volatile components,
including sugars and acids present in fruits.
• Short-chain unsaturated aldehydes and alcohols (C3-C6) and
esters are important contributors to the aroma.

25. Fruit Flavouring compound
Apple ( ripe) Ethyl 2- methylbutyrate
Apple ( green) Hexanal, 2- hexanal
Banana ( ripe) Eugenol
Banana ( green) 2- hexanal
Banana ( overripe) Isopentanol
Grapefruit Nootakatone
Lemon Citral
Orange Valencene
Pineapple Methyl Propionate Ester
Carambola Methyl Anthranilate
Durian Hydrogen Sulphide
Tamarind 2-acetyl furan
Raspberry 1- (α– hydroxyphenyl)- 3- butanone
Cherry Methyl Salicylate and Methyl Anthranilate

26. Changes in organic acids
Malate and citrate are more common among fruits. Also phenolic
compounds and tannins may affect the taste.
Onset of
Ripening
Increase
membrane
permeability
Reduction in
translocation
from the leaves
Conversion of
starch to sugar
Acids are used
in respiration
Increase in
volume of fruit
Leading to
dilution of
acids

27. Change in amino acids and proteins
• Amino acids generally decrease with ripening in
many fruits.
• They are reduced towards maturity because of
incorporation into proteins required for synthesis
of various enzymes.
• It is presumed that lowering of amino acids
indicates the advancement of maturity.
• Aspartic acid, glutamic acids, serine fractions
decreases with ripening in muskmelon.
• Similarly, in tomato, leucine and iso-leucine
decreases with fruit ripening.

28. Enzymes involved in ripening and
senescence of fruits
• The major enzymes implicated in softening of fruits are
pectineasteraes,polygalacturonase,β-(1-4)gluconase or
cellulase and β-galactosidase.
The mode of action of these enzymes are as follows
• Pectineasterase –Act to remove the methyl group from
the C-6 position of a galacturonic acid.
• Polygalacturonase-Hydrolyses the α(1-4) link between
adjacent dimethylated galacturonic acid residue.
• Cellulase-Hydrolyses the β-(1-4) link between adjacent
glucose residue
• β-galactosidase - In some cases attacks on native
galactan polymar.

29. • Lipase: Lipase activity is increase during ripening of
avocado with parallel increase in fatty acid.
• Malic enzyme: malic enzyme degrades malic acid in
apple , mango, grapes,etc.. increased during ripening.
• Citrate synthetase,: This enzyme is mainly found in citric
fruits and it synthesizes citric acid, citrate synthetase
found in citrus.
• Acid phosphatase.: It involved in carotenoid biosynthesis
increase during ripening of mango and banana.
• Phenylalanine ammonia lyase and flavone synthase are
key enzymes for the synthesis of anthocyanins.
• The important group of enzymes responsible for the
metabolism of phenolics is the polyphenol oxidase (PPO)
which are of two type catechol oxidase and laccase which
catalyse the oxidation of O-diphenols and P-diphenols,
respectively, and have been classified together under the
general nature of monophenol mono oxygenase.

30. Regulation in Ripening
1. Ethylene Regulation
2. Regulation of O2 & CO2
3. Calcium application
4. Using Ionizing radiation
5. Bioregulators

31. ETHYLENE MANAGEMENT
• Potassium permanganate
• Celite-KMnO4
• Silica gel-KMnO4
• Alumina silicates-KMnO4
• Activated charcoal
• Zeolites
• Sodium Permanganate
• AVG= Aminoethoxy-vinyl-
glycine
• AOA= Amino oxyacetic acid
• 1-MCP= 1-methyl cyclo
propane
• AgNO3= Silver Nitrate
• Ag(S2O3)2
3-=Silver
thiosulphate
• Co2+= Cobalt ion
Ethylene inhibitor Ethylene absorbant

32. Changes in pectic enzymes and cellulase activity
during guava fruit ripening
Abu-Bakr A. Abu-Goukh, Hind A. Bashir
•Changes in activities of the cell wall degrading enzymes,
pectinesterase (PE), polygalacturonase (PG) and cellulase,
were studied during the ripening of white- and pink-fleshed
guava fruit types.
•PE activity increased in both guava types up to the
climacteric peak of respiration (flesh firmness of 1.21
kg/cm2) and subsequently decreased.
•Activities of PG and cellulase increased progressively
during the ripening of both guava fruit types with a high
correlation between the increase in the activity of the two
enzymes and the loss of fruit flesh firmness.
Food Chemistry 83 (2003) 213–218

38. Conclusion
• Fruit ripening is a highly coordinated, genetically programmed, and
an irreversible phenomenon involving a series of physiological,
biochemical, and organoleptic changes, that finally lead to the
development of a soft, edible and ripe fruit with desirable quality
attributes.
• Starch, pectins, cellulose, and hemicelluloses are the major classes of
cell wall polysaccharides that undergo modifications during ripening.
• Increase in respiration occurs in climacteric fruits during ripening,
while no such rise in respiration is observed in non-climacteric ones.
• Various biochemical changes associated with fruit ripening involve
chlorophyll degradation, synthesis of anthocyanins, carotenoids etc.,
decreased acidity, polyphenols, development of volatiles etc.
• Ethylene a growth hormone has been found to regulate fruit ripening.
• Auxin and GA help in delaying the ripening while ABA and ethylene
accelerates the process.

39. Thank You