Cultivation, Collection, Processing of Medicinal Plants

UNIT II
CULTIVATION, COLLECTION, PROCESSING
AND STORAGE OF DRUGS OF
NATURAL ORIGIN
1
Prepared by: Ms. Divya Kanojiya
Assistant Professor in Pharmacognosy
Sumandeep Vidyapeeth Deemed to be University

CONTENT:
 CULTIVATION AND COLLECTION OF DRUGS OF NATURAL
ORIGIN
 FACTORS INFLUENCING CULTIVATION OF MEDICINAL
PLANTS
 PLANT HORMONES AND THEIR APPLICATIONS
 POLYPLOIDY, MUTATION AND HYBRIDIZATION WITH
REFERENCE TO MEDICINAL PLANTS
2

CULTIVATION AND COLLECTION OF
DRUGS OF NATURAL ORIGIN
3

CULTIVATION OF CRUDE DRUGS:
→ Cultivation of medicinal plants requires intensive care and management.
→ The conditions and duration of cultivation required vary depending on
the quality of medicinal plant materials required.
4

 ADVANTAGES OF CULTIVATION:
→ It ensures quality and purity of medicinal plants.
→ Collection of crude drugs from cultivated plants gives a better yield and
therapeutic quality.
→ Cultivation ensures regular supply of a crude drug.
→ Cultivation permits application of modern technological aspects such as
mutation, polyploidy and hybridisation.
 DISADVANTAGES OF CULTIVATION:
→ Cultivation drugs as compared to wild source and losses due to
ecological imbalance such as storms, earthquakes, droughts are major
disadvantages of cultivation.
5

 Methods of cultivation:
1. Asexual method
2. Sexual method (Propagation from seeds)
3. Micro propagation / Plant Tissue Culture
6

ASEXUAL METHOD
→ Vegetative propagation (Asexual propagation):
→ Vegetative propagation can be defined as regeneration or formation of a
new individual from any vegetative part of the plant body.
→ The method of vegetative propagation involves separation of a part of
plant body, which develops into a new plant.
7

 Other Vegetative propagation methods:
1. Cutting
2. Layering
3. Grafting
9

1. Cutting:
→ These are the parts of the plant (stem, root or leaf) which, if grown under
suitable conditions, develop new plants.
→ Stem cutting are generally used to obtained new plants. Examples:
Sugarcane and rose, etc.
10

2. Layering:
→ Roots are induced on the stem
while it is still attached to the
parent plant.
→ This part of stem is later detached
from the parent plant and grown
into a new plant. Examples:
Jasmine plant
11

3. Grafting:
→ New variety is produced by joining parts of two different plants.
→ The rooted shoot of one plant, called stock, is joined with a piece of shoot
of another plant known as scion.
→ Examples: Rose, citrus and rubber, etc.
12

 Advantages of Asexual Propagation:
→ As resultant species formed through asexual process are genetically identical,
useful traits can be preserved among them.
→ Asexual propagation allows propagation of crops that do not possess seeds or
those which are not possible to grow from seeds.
→ For e.g. Jasmine. sugarcane, potato, banana, rose etc.
→ Plants grown through vegetative propagation bear fruits early.
→ In this type, only a single parent is required and thus it eliminates the need for
propagation mechanisms such as pollination, cross pollination etc.
→ The process is faster than sexual propagation.
→ This helps in rapid generation of crops which in turn balances the loss.
13

 Disadvantages of Asexual Propagation
→ Due to over crowding of large number of plants near the parent plant,
there is a severe competition between the members of same species.
Thus many plants become stunted and weak.
→ The organs used in vegetative reproduction are very poor means of
propagation.
14

SEXUAL PROPAGATION
 The process of sexual propagation:
1. Pollination: This is the transfer of pollen grains from the anther to
the stigma.
2. Fertilization: Fusion of male and female gametes takes place,
resulting in the formation of zygote.
3. Seedling: Multiplication of plants by using seed is called as seed
propagation
15

 Dormancy:
→ It is term used to describe a seed that will not germinate because of any condition
associated either with the seed itself or with existing environmental factors such as
temperature and moisture.
 Rest Period:
→ Some seeds will not germinate immediately after harvest even if conditions are
favorable.
→ This failure to germinate is due to physiological condition.
→ This is said to be the seeds are in the rest period.
 Seed viability and longevity:
→ Viability means the presence of life in the seed. Longevity refers to the length of
time that seeds will retain their viability.
→ Some seeds are short lived. (Citrus).
16

 Advantages of Sexual Propagation:
→ Simplest, easiest and the most economical process
→ Some plants, trees, vegetables or fruits species can propagate only
through sexual propagation. E.g. -marigold, papaya, tomato.
→ This type of propagation leads to better crop species that are stronger,
disease- resistant and have longer life-span.
→ Viral transmission can be prevented in this type of propagation
→ Easy storage and transportation of seeds.
17

 Disadvantages of Sexual Propagation
→ Seeds take a long time to turn into mature plants i.e. time interval
between sowing and flowering is longer.
→ Seedlings propagated through sexual propagation are unlikely to have
same genetic characteristics as that of parent plants.
→ Some plant species do not produce viable seeds through sexual
propagation and hence are unsuitable to propagate for the same.
→ Plants that do not have seeds can't be propagated through this process.
→ There are many factors that can affect the viability of seeds, including
moisture, air, temperature, and light.
18

MICRO PROPAGATION
→ This method consists of growing cell, tissue and organ in culture.
→ Small pieces of plant organs or tissues are grown in a container with suitable
nutrient medium, under sterilized conditions.
→ The tissue grows into a mass of undifferentiated cells called callus which later
differentiates into plantlets.
→ These are then transferred into pots or nursery beds and allowed to grow into
full plants.
→ Plant tissue culture is widely used to produce clones of a plant in a method
known as micropropagation to conserve rare or endangered plant species.
→ Micro propagation is useful in raising disease free plants, homozygous
diploids, and those without viable seeds. 19

COLLECTION OF DRUGS
1. Medicinal plant materials should be collected during the appropriate season or time
period to ensure the best possible quality of both source materials and finished
products.
2. It is well known that the quantitative concentration of biologically active
constituents varies with the stage of plant growth and development.
3. The best time for collection (quality peak season or time of day) should be
determined according to the quality and quantity of biologically active constituents
rather than the total vegetative yield of the targeted medicinal plant parts.
4. In general, the collected raw medicinal plant materials should not come into direct
contact with the soil.
5. If underground parts (such as the roots) are used, any adhering soil should be
removed from the plants as soon as they are collected.
21

6. In general, the collected raw medicinal plant materials should not come into
direct contact with the soil.
7. If underground parts (such as the roots) are used, any adhering soil should be
removed from the plants as soon as they are collected.
8. Collected material should be placed in clean baskets, mesh bags, other well
aerated containers.
9. After collection, the raw medicinal plant materials may be subjected to
appropriate preliminary processing, including elimination of undesirable
materials and contaminants, washing (to remove excess soil), sorting and cutting.
10. The collected medicinal plant materials should be protected from insects, rodents,
birds and other pests, and from livestock and domestic animals.
22

11. If the collection site is located some distance from processing facilities, it may
be necessary to air or sun-dry the raw medicinal plant materials prior to
transport.
12. If more than one medicinal plant part is to be collected, the different plant
species or plant materials should be gathered separately and transported in
separate containers.
13. Cross-contamination should be avoided at all times.
14. Collecting implements, such as machetes, shears, saws and mechanical tools,
should be kept clean and maintained in proper condition.
15. Those parts that come into direct contact with the collected medicinal plant
materials should be free from excess oil and other contamination.
23

FACTORS AFFECTING CULTIVATION
24

→ Cultivation of medicinal plants offers wide range of advantages over the plants
obtained from wild sources.
→ There are few factors to concern which have a real effect on plant growth and
development, nature and quantity of secondary metabolites.
→ The factors affecting cultivation are altitude, temperature, rainfall, length of day,
day light, soil and soil fertility, fertilizers and pests.
→ The effects of these factors have been studied by growing particular plants in
different environmental conditions and observing variations.
→ Nutrients have the ability to enhance the production of secondary metabolites, at
the same time they may reduce the metabolites as well.
26

ALTITUDE
1. Altitude is a very important factor in cultivation of medicinal plants.
2. Tea, cinchona and eucalyptus are cultivated favorably at an altitude of
1,000-2,000 metres.
3. Cinnamon and cardamom are grown at a height of 500-1000 metres, while
Senna can be cultivated at sea level.
4. The following are the examples of medicinal and aromatic plants
indicating the altitude for their successful cultivation.
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28
Plant Altitude(meters)
Tea 1000-1500
Cinchona 1000-2000
Camphor 1500-2000
Cinnamon 250-1000
Coffee 1000-2000
Clove Up to 900
Saffron Up to 1250
Cardamom 600-1600

TEMPERATURE
1. Temperature is a crucial factor controlling the growth, metabolism and there
by the yield of secondary metabolites of plants.
2. Even though each species has become adapted to its own natural environment,
they are able to exist in a considerable range of temperature.
3. Many plants will grow better in temperate regions during summer, but they
lack in resistance to withstand frost in winter.
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Plant Optimum temperature (ºF)
Cinchona 60-75
Coffee 55-70
Tea 70-90
Cardamom 50-100

RAINFALL
1. For the proper development of plant, rainfall is required in proper
measurements.
2. Xerophytic plants(a plat that can survive with very little water) like aloes do
not require irrigation rainfall.
3. The effects of rainfall on plants must be considered in relation to the annual
rainfall throughout the year with the water holding properties of the soil.
4. Variable results have been reported for the production of constituents under
different conditions of rainfall.
5. Excessive rainfall could cause a reduction in the secondary metabolites due
to leaching of water soluble substances from the plants.
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DAY LENGTH AND DAY LIGHT
1. It has been proved that even the length of the day has an effect over the
metabolites production.
2. The plants that are kept in long day conditions may contain more or less
amount of constituents when compared to the plants kept in short day.
3. For example peppermint has produced menthone, menthol and traces of
menthofuran in long day conditions and only menthofuran in short day
condition.
4. The developments of plants vary much in both the amount and intensity of the
light they require.
31

5. The wild grown plants would meet the required conditions and so they
grow but during cultivation we have to fulfill the requirements of plants.
6. The day light was found to increase the amount of alkaloids in belladonna,
stramonium, cinchona, etc.
7. Even the type of radiation too has an effect over the development and
metabolites of plants.
32

SOIL
1. Each and every plant species have its own soil and nutritive
requirements.
2. The three important basic characteristics of soils are their physical,
chemical and microbiological properties.
3. Soil provides mechanical support, water and essential foods for the
development of plants.
4. Soil consists of air, water, mineral matters and organic matters.
5. Variations in particle size result in different soils ranging from clay,
sand and gravel.
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6. Particle size influences the water holding capacity of soil.
7. The type and amount of minerals plays a vital role in plant cultivation.
8. Calcium favours the growth of certain plants whereas with some plants it
does not produce any effects.
9. The plants are able to determine their own soil pH range for their growth;
microbes should be taken in to consideration which grows well in certain
pH.
10. Nitrogen containing soil has a great momentum in raising the production of
alkaloids in some plants.
34

SOIL FERTILITY
1. It is the capacity of soil to provide nutrients in adequate amounts and in
balanced proportion to plants.
2. If cropping is done without fortification of soil with plant nutrients, soil
fertility gets lost.
3. It is also diminished through leaching and erosion.
4. Soil fertility can be maintained by addition of animal manures, nitrogen-
fixing bacteria or by application of chemical fertilizers.
5. The latter is time saving and surest of all above techniques.
35

36
Particle size (diameter) Type of soil
Less than 0.002 mm Fine clay
0.002-0.02 mm Coarse clay
0.02 – 0.2 mm Fine sand
0.2- 2.0 mm Coarse sand

FERTILIZERS AND MANURES
→ Plant also needs food for their growth and development.
→ What plants need basically for their growth are the carbon sun-rays, water
and mineral matter from the soil.
→ It is seen that with limited number of chemical elements, plants build up
fruits, grains, fibres, etc. and synthesize fixed and volatile oils, glycosides,
alkaloids, sugar and many more chemicals.
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→ Chemical:
→ Animals are in need of vitamins, plants are in need of sixteen nutrient elements for
synthesizing various compounds.
→ Some of them are known as primary nutrients like nitrogen, phosphorus and
potassium.
→ Magnesium, calcium and sulphur are required in small quantities and hence, they
are known as secondary nutrients.
→ Trace elements like copper, manganese, iron, boron, molybdenum, zinc are also
necessary for plant growths are known as micronutrients.
→ Carbon, hydrogen, oxygen and chlorine are provided from water and Every element
has to perform some specific function in growth and development of plants.
→ Its deficiency is also characterized by certain symptoms.
38

MANURES
39
→ Farm yard manure (FYM/compost), castor seed cake, poultry manures,
neem and karanj seed cakes vermin compost, etc. are manures.
→ Oil-cake and compost normally consists of 3-6% of nitrogen, 2%
phosphates and 1-1.5% potash.
→ They are made easily available to plants.
→ Bone meal, fish meal, biogas slurry, blood meal and press mud are the other
forms of organic fertilizers.

BIOFERTILIZERS
→ Inadequate supply, high costs and undesirable effects if used successively are
the demerits of fertilizers or manures and hence the cultivator has to opt for
some other type of fertilizer.
→ Biofertilizers are the most suitable forms that can be tried.
→ These consist of different types of micro organisms or lower organisms which
fix the atmospheric nitrogen in soil and plant can use them for their day to day
use.
→ Thus they are symbiotic. Rhizobium, Azotobactor, Azosperillium, Bijericcia,
Blue- green algae, Azolla, etc. are the examples of biofertilizers.
40

PEST AND PEST CONTROL
→ Pests are undesired plant or animal species that causes a great damage to
the plants.
→ There are different types of pests; they are microbes, insects, non insect
pests and weeds.
41

MICROBES
→ They include fungi, bacteria and viruses. Armillaria Root Rot (Oak Root
Fungus) is a disease caused by fungi Armil-laria mellea (Marasmiaceae) and in
this the infected plant become nonproductive and very frequently dies within
two to four years.
→ Plants develop weak, shorter shoots as they are infected by the pathogen.
→ Dark, root-like structures (rhizomorphs), grow into the soil after symptoms
develop on plants.
42

GENERAL METHODS OF PEST CONTROLS
43
Controlling
technique
Methods involved
Cultural Changing the time of sowing and harvesting, maintenance of storage, special
cultivation methods, proper cleaning, Using trap crops
and resistant varieties
Physical Mechanical control Utilization of physical factors (temperature, less oxygen
concentration, humidity, passing CO2)
Biological Using predators, parasites, pathogens,
sterilization, genetic manipulation, pheromones
Chemical Use of pesticides, herbicides, antifeedants (a naturally occuring sunstance in
certain plants which adversely affects insects or other animals which eat them.)

OTHER FACTORS THAT AFFECT THE
CULTIVATED PLANTS
 Air Pollution
→ Chemical discharges into the atmosphere have increased dramatically
during this century, but the total effect on plants is virtually unknown.
→ It has been demonstrated that air pollutants can cause mortality and
losses in growth of plants.
→ Nearly all species of deciduous and coniferous trees are sensitive to
some pollutants.
44

 Herbicide
 Herbicides should be handled very carefully; misapplication of herbicides can
often damage nontarget plants.
 The total extent of such damage remains unclear, but localized, severe damage
occurs.
 Symptoms of herbicide injury are variable due to chemical mode of action,
dosage, duration of expo-sure, plant species, and environmental conditions.
 Some herbicides cause growth abnormalities such as cupping or twisting of
foliage while others cause foliage yellowing or browning, defoliation, or death.
45

PLANT HORMONES AND
THEIR APPLICATIONS
46

→ Plant hormones (phytohormones) are physiological intercellular messengers
that control the complete plant lifecycle, including germination, rooting,
growth, flowering, fruit ripening, foliage and death.
→ In addition, plant hormones are secreted in response to environmental factors
such as excess of nutrients, drought conditions, light, temperature and chemical
or physical stress.
→ So, levels of hormones will change over the lifespan of a plant and are
dependent upon season and environment.
→ The term ‘plant growth factor’ is usually employed for plant hormones or
substances of similar effect that are administered to plants.
→ Growth factors are widely used in industrialized agriculture to improve
productivity.
47

 Five major classes of plant hormones are mentioned: auxins, cytokinins,
gibbereilins, abscisic acid and ethylene.
→ However as research progresses, more active molecules are being found and
new families of regulators are emerging; one example being polyamines
(putrescine or spermidine).
→ Plant growth regulators have made the way for plant tissue culture
techniques, which were a real boon for mankind in obtaining therapeutically
valuable secondary metabolites
48

 General plant hormones:
⸙ Auxins: cell elongation
⸙ Gibberellins: cell elongation + cell division – translated into growth
⸙ Cytokinins: cell division + inhibits senescence
⸙ Abscisic acid: abscission of leaves and fruits + dormancy induction of buds
and seeds
⸙ Ethylene: promotes senescence, epinasty and fruit ripening
49

→ The term auxin is derived from the Greek word auxein which means to
grow.
→ Generally compounds are considered as auxins if they are able to induce cell
elongation in stems and otherwise resemble indoleacetic acid (the first auxin
isolated) in physiological activity.
→ Auxins usually affect other processes in addition to cell elongation of stem
cells but this characteristic is considered critical of all auxins and thus
‘helps’ define the hormone.
→ Auxins were the first plant hormones discovered.
52

→ Indole acetic acid (IAA) is the principle natural auxin and other natural
auxins are indole-3-acetonitrile (IAN), phenyl acetic acid and 4-
chloroindole-3-acetic acid.
→ The exogenous or synthetic auxins are indole-3-butyric acid (IBA), α-
napthyl acetic acid (NAA), 2-napthyloxyacetic acid (NOA), 1-napthyl
acetamide (NAD), 5-carboxymethyl-N, N-dimethyl dithiocarbamate,
2,4-dichlorophenoxy acetic acid (2,4-D), etc.
53

 FUNCTIONS OF AUXIN:
1. Stimulates cell elongation.
2. The auxin supply from the apical bud suppresses growth of lateral buds.
3. Apical dominance is the inhibiting influence of the shoot apex on the
growth of axillary buds.
4. Removal of the apical bud results in growth of the axillary buds.
5. Replacing the apical bud with a lanolin paste containing IAA restores the
apical dominance.
6. The mechanism involves another hormone - ethylene. Auxin (IAA) causes
lateral buds to make ethylene, which inhibits growth of the lateral buds.
55

7. Differentiation of vascular tissue (xylem and phloem) is stimulated by IAA.
8. Auxin stimulates root initiation on stem cuttings and lateral root
development in tissue culture (adventitious rooting).
9. Auxin mediates the tropistic response of bending in response to gravity and
light (this is how auxin was first discovered).
10. Auxin has various effects on leaf and fruit abscission, fruit set, development,
and ripening, and flowering, depending on the circumstances.
56

 PRODUCTION AND OCCURRENCE
1. Produced in shoot and root meristematic tissue, in young leaves, mature root
cells and small amounts in mature leaves.
2. Transported throughout the plant parts and the production of IAA will be more
in day time.
3. It is released by all cells when they are experiencing conditions which would
normally cause a shoot meristematic cell to produce auxin.
4. Ethylene has direct or indirect action over to enhance the synthesis auxin.
5. IAA is chemically similar to the amino acid tryptophan which is generally
accepted to be the molecule from which IAA is derived.
6. Three mechanisms have been suggested to explain this conversion:
58

7. Tryptophan is converted to indolepyruvic acid through a transamination
reaction. Indolepyruvic acid is then converted to indoleacetaldehyde by a
decarboxylation reaction.
8. The final step involves oxidation of indoleacetaldehyde resulting in
indoteacetic acid.
9. Tryptophan undergoes decarboxylation resulting in tryptamine.
10. Tryptamine is then oxidized and deaminated to produce indoleacetaldehyde.
This molecule is further oxidized to produce indoleacetic acid.
11. IAA can be produced via a tryptophan-independent mechanism. This
mechanism is poorly understood, but has been proven using tip (-) mutants.
59

12. Other experiments have shown that, in some plants, this mechanism is
actually the preferred mechanism of IAA biosynthesis.
13. The enzymes responsible for the biosynthesis of IAA are most active in
young tissues such as shoot apical meristems and growing leaves and fruits.
These are the same tissues where the highest concentrations of IAA are
found.
14. Another control mechanism involves the production of conjugates which
are, in simple terms, molecules which resemble the hormone but are
inactive.
15. The formation of conjugates may be a mechanism of storing and
transporting the active hormone. Conjugates can be formed from IAA via
hydrolase enzymes. 60

16. Conjugates can be rapidly activated by environmental stimuli signaling a
quick hormonal response. Degradation of auxin is the final method of
controlling auxin levels.
17. This process also has two proposed mechanisms outlined below:
18. The oxidation of IAA by oxygen resulting in the loss of the carboxyl group
and 3-methyleneoxindole as the major breakdown product. IAA oxidase is
the enzyme which catalyses this activity.
19. Conjugates of IAA and synthetic auxins such as 2,4-D can not be destroyed
by this activity.
20. Conjugates of IAA and synthetic auxins such as 2,4-D can not be destroyed
by this activity.
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21. C-2 of the heterocyclic ring may be oxidized resulting in oxindole-3-acetic
acid. C-3 may be oxidized in addition to C-2 resulting in dioxindole-3-
acetic acid.
22. The mechanisms by which biosynthesis and degradation of auxin molecules
occur are important to future agricultural applications.
23. Information regarding auxin metabolism will most likely lead to genetic and
chemical manipulation of endogenous hormone levels resulting in desirable
growth and differentiation of important plant species.
62

63
Figure: Pathways of IAA biosynthesis

65
→ Cytokinins are compounds with a structure resembling adenine which promote cell
division and have other similar functions to kinetin.
→ They also regulate the pattern and frequency of organ production as well as
position and shape.
→ They have an inhibitory effect on senescence.
→ Kinetin was the first cytokinin identified and so named because of the compounds
ability to promote cytokinesis (cell division).
→ Though it is a natural compound, it is not made in plants, and is therefore usually
considered a ‘synthetic’ cytokinin.
→ The common naturally occurring cytokinin in plants today is called zeatin which
was isolated from corn.

DISCOVERY OF CYTOKININ
1. In the 1950s, Folke Skoog and Carlos Miller studying the influence of auxin
on the growth of tobacco in tissue culture.
2. When auxin was added to artificial medium, the cells enlarged but did not
divide. Miller took herring- sperm DNA.
3. Miller knew of Overbeek's work, and decided to add this to the culture
medium, the tobacco cells started dividing.
4. He repeated this experiment with fresh herring-sperm DNA, but the results
were not repeated. Only old DNA seemed to work.
5. Miller later discovered that adding the purine base of DNA (adenine) would
cause the cells to divide.
66

6. Adenine or adenine-like compounds induce cell division in plant tissue
culture. Miller, Skoog and their coworkers isolated the growth factor
responsible for cellular division from a DNA preparation calling it kinetin
which belongs to a class of compounds called cytokinins.
7. In 1964, the first naturally occurring cytokinin was isolated from com
called zeatin.
8. Zeatin and zeatin riboside are found in coconut milk.
9. All cytokinins (artificial or natural) are chemically similar to adenine.
67

FUNCTIONS OF CYTOKININ
1. Stimulate cell division (cytokinesis).
2. Stimulate morphogenesis (shoot initiation/bud formation) in tissue culture.
3. Stimulate the growth of lateral (or adventitious) buds release of apical
dominance.
4. Stimulate leaf expansion resulting from cell enlargement.
5. May enhance stomatal opening in some species. Promotes the conversion of
etioplasts into chloroplasts via stimulation of chlorophyll synthesis.
6. Stimulate the dark-germination of light-dependent seeds.
7. Delays senescence.
8. Promotes some stages of root development. 68

PRODUCTION AND OCCURRENCE
1. Produced in root and shoot meristematic tissue, in mature shoot cells and in
mature roots in small amounts.
2. If is rapidly transported in xylem stream.
3. Peak production occurs in day time and their activity is reduced in plants
suffering drought.
4. It is directly or indirectly induced by high levels of Gibberlic acid.
5. Cytokinin is generally found in meristematic regions and growing tissues. They
are believed to be synthesized in the roots and translocated via the xylem to
shoots.
6. Cytokinin biosynthesis happens through the biochemical modification of
adenine. They are synthesized by following pathway: 69

A product of the mevalonate pathway called isopentyl pyrophosphate is isomerized.
This isomer can then react with adenosine monophosphate with the aid of an enzyme called
isopentenyl AMP synthase.
The result is isopentenyl adenosine-5’-phosphate (isopentenyl AMP).
This product can then be converted to isopentenyl adenosine by removal of the phosphate
by a phosphatase and further converted to isopentenyl adenine by removal of the ribose
group.
70

Isopentenyl adenine can be converted to the three major forms of naturally
occurring cytokinins.
→ Degradation of cytokinins occurs largely due to the enzyme cytokinin
oxidase.
→ This enzyme removes the side chain and releases adenine.
→ Derivatives can also be made but the difficulties are with pathways, which
are more complex and poorly understood.
71

 PRODUCTION AND OCCURRENCE:
1. Produced in the roots, embryo and germinating seeds.
2. The level of gibberellins goes up in the dark when sugar cannot be
manufactured and will be reduced in the light.
3. It is released in mature cells (particularly root) when they do not have enough
sugar and oxygen to support both themselves and released by all cells when
they are experiencing conditions which would normally cause a mature root
cell to produce GA.
4. Gibberellins are diterpenes synthesized from acetyl CoA via the mevalonic
acid pathway.
5. They all have either 19 or 20 carbon units grouped into either four or five ring
systems.
75

6. The fifth ring is a lactone ring as shown in the structures above attached to ring
A.
7. Gibberellins are believed to be synthesized in young tissues of the shoot and
also the developing seed.
8. It is not clear whether young root tissues also produce gibberellins.
9. There is also some evidence that leaves may also contain them.
10. The gibberellins are formed through the pathway, three acetyl CoA molecules
are oxidized by two NADPH molecules to produce three CoA molecules as a
side product and mevalonic acid.
11. Mevalonic acid is then Phosphorylated by ATP and decarboxylated to form
isopentyl pyrophosphate. Four of these molecules form geranylgeranyl
pyrophosphate which serves as the donor for all GA carbon atoms.
76

12. This compound is then converted to copalylpyrophosphate which has 2 ring
systems. Copalylpyrophosphate is then converted to kaurene which has 4-
ring systems. Subsequent oxidations reveal kaurenol (alcohol form),
kaurenal (aldehyde form), and kaurenoic acid respectively.
13. Kaurenoic acid is converted to the aldehyde form of GA12 by
decarboxylation. GA12 is the first true gibberellane ring system with 20
carbons.
14. From the aldehyde form of GA12 arise both 20 and 19 carbon gibberellins
but there are many mechanisms by which these other compounds arise.
15. During active growth, the plant will metabolize most gibberellins by
hydroxylation to inactive conjugates quickly with, the exception of GA3.
77

16. GA3 is degraded much slower which helps to explain why the symptoms
initially associated with the hormone in the disease bakanae are present.
17. Inactive conjugates might be stored or translocated via the phloem and
xylem before their release (activation) at the proper time and in the proper
tissue.
78

 HISTORY
1. Ethylene has been used in practice since the ancient times, where people would
use gas figs in order to stimulate ripening, burn incense in closed rooms to
enhance the ripening of pears.
2. It was in 1864, that leaks of gas from street lights showed stunting of growth,
twisting of plants, and abnormal thickening of stems.
3. In 1901, a Russian scientist named Dimitry Neljubow showed that the active
component was ethylene. Doubt 1917, discovered that ethylene stimulated
abscission. In 1932 it was demonstrated that the ethylene evolved from stored
apple inhibited the growth of potato shoots enclosed with them. In 1934 Gane
reported that plants synthesize ethylene.
4. In 1935, Crocker proposed that ethylene was the plant hormone responsible for
fruit ripening as well as inhibition of vegetative tissues.
5. Ethylene is now known to have many other functions as well.
80

 Functions of ethylene:
1. Production stimulated during ripening, flooding, stress, senescence,
mechanical damage, infection.
2. Regulator of cell death programs in plants (apoptosis).
3. Stimulates the release of dormancy.
4. Stimulates shoot and root growth and differentiation (triple response).
5. Regulates ripening of climacteric fruits.
6. May have a role in adventitious root formation.
7. Stimulates leaf and fruit abscission.
8. Flowering in most plants is inhibited by ethylene.
9. Mangos, pineapples and some ornamentals are stimulated by ethylene
81

10. Induction of femaleness in dioecious flowers.
11. Stimulates flower opening.
12. Stimulates flower and leaf senescence.
82

 Production and occurrence:
1. Production is directly induced by high levels of Auxin, root flooding and
drought.
2. It is found in germinating seeds and produced in nodes of stems, tissues of
ripening fruits, response to shoot environmental, pest, or disease stress and in
senescent leaves and flowers.
3. Light minimizes the production of ethylene.
4. It is released by all cells when they are experiencing conditions which would
normally cause a mature shoot cell to produce ethylene.
5. Ethylene is produced in all higher plants and is produced from methionine in
essentially all tissues.
6. Production of ethylene varies with the type of tissue, the plant species, and
also the stage of development. The mechanism by which ethylene is
produced from methionine is a three step process. 83

7. ATP is an essential component in the synthesis of ethylene from methionine.
8. ATP and water are added to methionine resulting in loss of the three phosphates
and S-adenosyl methionine (SAM). 1-amino-cyclopropanel-carboxylic acid
synthase (ACC-synthase) facilitates the production of ACC from SAM.
9. Oxygen is then needed in order to oxidize ACC and produce ethylene.
10. This reaction is catalysed by an oxidative enzyme called ethylene forming
enzyme.
11. The control of ethylene production has received considerable study.
12. Study of ethylene has focused around the synthesis promoting effects of auxin,
wounding, and drought as well as aspects of fruit-ripening.
13. ACC synthase is the rate limiting step for ethylene production and it is this
enzyme that is manipulated in biotechnology to delay fruit ripening in the
‘flavor saver’ tomatoes.
84

 HISTORY:
1. Natural growth inhibiting substances are present in plants and affect the
normal physiological process of them. One such compound is abscisic
acid, a single compound unlike the auxins, gibberellins, and cytokinins.
2. It was called ‘abscisin II’ originally because it was thought to play a major
role in abscission of fruits.
3. At about the same time another group was calling it ‘dormin’ because they
thought it had a major role in bud dormancy. Though abscisic acid
generally is thought to play mostly inhibitory roles, it has many promoting
functions as well.
86

4. In 1963, when Frederick Addicott and his associates were the one to
identify abscisic acid. Two compounds were isolated and named as abscisin
I and abscisin II.
5. Abscisin II is presently called abscisic acid (ABA). At the same time Philip
Wareing, who was studying bud dormancy in woody plants and Van
Steveninck, who was studying abscission of flowers and fruits discovered
the same compound.
87

 Functions of abscisic acid
1. The abscisic acid stimulates the closure of stomata (water stress brings about an increase
in ABA synthesis).
2. Involved in abscission of buds, leaves, petals, flowers, and fruits in many, if not all,
instances, as well as in dehiscence of fruits.
3. Production is accentuated by stresses such as water loss and freezing temperatures.
4. Involved in bud dormancy.
5. Prolongs seed dormancy and delays germination.
6. Inhibits elongation.
7. ABA is implicated in the control of elongation, lateral root development, and
geotropism, as well as in water uptake and ion transport by roots.
8. ABA coming from the plastids promotes the metabolism of ripening.
9. Promotes senescence.
10. Can reverse the effects of growth stimulating hormones.
88

 PRODUCTION AND OCCURRENCE:
1. ABA is a naturally occurring sesquiterpenoid (15-carbon) compound in plants,
which is partially produced via the mevalonic pathway in chloroplasts and
other plastids.
2. Because it is synthesized partially in the chloroplasts, it makes sense that
biosynthesis primarily occurs in the leaves. The production of ABA is by
stresses such as water loss and freezing temperatures.
3. The biosynthesis occurs indirectly through the production of carotenoids.
4. Breakdown of these carotenoids occurs by the following
mechanism:Violaxanthin (forty carbons) is isomerized and then splitted via an
isomerase reaction followed by an oxidation reaction. 89

5. One molecule of xanthonin is produced from one molecule of violaxanthonin
and it is not clear what happens to the remaining by products.
6. The one molecule of xanthonin produced is unstable and spontaneously
changed to ABA aldehyde.
7. Further oxidation results in ABA. Activation of the molecule can occur by two
methods.
8. In the first, method, an ABA-glucose ester can form by attachment of glucose
to ABA.
9. In the second method, oxidation of ABA can occur to form phaseic acid and
dihyhdrophaseic acid. Both xylem and phloem tissues carries ABA.
10. It can also be translocated through parenchyma cells. Unlike auxins, ABA is
capable of moving both up and down the stem.
90

POLYPLOIDY, MUTATION AND
HYBRIDIZATION WITH
REFERENCE TO MEDICINAL
PLANTS
91

 W HAT I S PLOYPLOIDS ?
→ Polyploids are organisms with multiple sets of chromosomes in excess of the
diploid number.
→ Polyploids is common in nature and provide a major mechanism for adaptation
and speciation.
→ Approximately 50-70% of angiosperms, which include many crops plants,
have undergone polyploidy during their evolutionary process.
→ Classification of Polyploids:
1. Based on their chromosomal composition:
2. Euploids
3. Aneuploids
4. Euploids constitute the major of polyploids.
93

 EUPLOIDY:
 Euploidy are polyploids with multiples of the complete set of
chromosomes specific to a species.
 Depending on the composition of the genome, euploids can be further
classified into autopolyploids and allopolyploids
94

 AUTOPOLYPLOIDY
→ Autopolyploidy Contains of multiple copies of the basic set (x) of
chromosomes of the same genome.
95

→ It occurs in nature through union reduced gametes.
→ Natural autoploids include tetraploid crops such as alfafa, peanut, potato,
coffee and triploid bananas.
96

 ALLOPOLYPLOIDY
→ A combination of genomes from different species.
→ They result from hybridization of two or more genomes followed by
chromosome doubling or by the fusion of unreduced gametes between
species.
→ This mechanism is called non-disjunction.
→ These meiotic aberrances result in plants with reduced vigor.
→ Economically important natural alloploid crops include strawberry, wheat,
oat, upland cotton, oilseed rape, blueberry and mustard.
97

 ANEUPLOIDY
→ This are polyploids that contain either an addition or subtraction of one or
more specific chromosome(s) to the total number of chromosomes that usually
make up the ploidy of a species.
99

→ Aneuploids result from the formation of univalentsand multivalents during
meiosis of euploids.
→ With no mechanism of dividing univalents equally among daughter cells during
anaphase I, some cells inherit more genetic material than others.
→ Similarly, multivalents such as homologous chromosomes may fail to separate
during meiosis leading to unequal migration of chromosomes to opposite poles.
100
 CLASSIFICATION:
Term Chromosome number
Monosomy 2n-1
Nullisomy 2n-2
Trisomy 2n+2
Tetrasomy 2n+2
Pentasomy 2n+3

 INDUCING POLYPLOIDS:
1. They occur spontaneously through the process of chromosome doubling.
2. For example, induced autotetraploids in the watermelon crop are used for the
production of seedless triploid by hybrids fruits.
3. Such polyploids are induced through the treatment of diploids with mitotic
inhibitors such as dinitroaniles and colchicine.
4. The increase in nuclear ploidy affects the structuraland anatomical
characteristics of the plant.
5. Polyploidy results in increased leaf and flower size, stomatal density, cell
size and chloroplast count.
101

 INTRODUCTION:
→ Sudden heritable change in genetic material or character of an organism is
known as mutation.
→ Individuals showing these changes are known as mutants.
→ An individual showing an altered phenotype due to mutation are known as
variant.
→ Factor or agents causing mutation are known as mutagens.
→ Mutation which causes changes in base sequence of a gene are known as gene
mutation or point mutation.
103

 CHARACTERISTICS OF MUTATION:
1. Generally mutant alleles are recessive to their wild type or normal alleles
2. Most mutations have harmful effect, but some mutations are beneficial
3. Spontaneous mutations occurs at very low rate
4. Some genes shows high rate of mutation such genes are called as
mutablegene
5. Highly mutable sites within a gene are known as hot spots.
6. Mutation can occur in any tissue/cell (somatic or germinal) of an organism.
104

 CLASSIFICATION OF MUTATION
 Based on the survival of an individual:
1. Lethal mutation - when mutation causes death of all individuals undergoing
mutation are known as lethal
2. Sub lethal mutation - causes death of 90% individuals
3. Sub vital mutation- such mutation kills less than 90% individuals
 Vital mutation
1. When mutation don't affect the survival of an individual are known as vital
2. Supervital mutation - This kind of mutation enhances the survival of
individual
105

 Based on causes of mutation:
1. Spontaneous mutation-Spontaneous mutation occurs naturally without any cause.
2. The rate of spontaneous mutation is very slow
3. eg- Methylation followed by deamination of cytosine.
4. Rate of spontaneous mutation is higher in eukaryotes than prokaryotes.
5. Eg, UV light of sunlight causing mutation in bacteria
 Induced Mutation:
1. Mutations produced due to treatment with either a chemical or physicalare called
induced mutation.
2. The agents capable of inducing such mutations are known as mutagen.
3. Use of induced mutation for crop improvement program is known as mutation
breeding.
4. Eg. X-rays causing mutation in cerealscereals
106

 BASED ON TISSUE OF ORIGIN:
1. Somatic mutation-A mutation occurring in somatic cell is called somatic
mutation.
2. Germinal Mutation-When mutation occur in gametic cells or reproductive cells
are known as germinal mutation.
3. In reproductive species only germinal mutation are transmitted to the next
generation.
 BASED ON DIRECTION OF MUTATION:
1. Forward mutation- When mutation occurs from the normal/wild type allele to
mutant allele are known as forward mutation.
2. Reverse mutation- When mutation occurs in reverse direction that is from
mutant allele to the normal/wild type allele are known as reverse mutation.
107

 TYPES OF GENE MUTATIONS:
1. Point Mutations
2. Substitutions
3. Insertions
4. Deletions
5. Frameshift
108

HYBRIDIZATION WITH REFERENCE TO
MEDICINAL PLANTS
109

 MEANING OF HYBRIDIZATION
→ Individual produced as a result of cross between two genetically different
parents is known as hybrid.
→ The natural or artificial process that results in the formation of hybrid is known
as hybridization.
→ Hybridization is an important method of combining characters of different
plants.
→ Hybridization does not change genetic contents of organisms but it produces
new combination of genes.
110

 Objective and aim of hybridization:
1. The chief objective of hybridization is to create genetic variation.
2. The aim of hybridization may be transfer of one or few qualitative
characters, the improvement in one or more quantitative character or the
use of F1 as a hybrid variety.
3. To artificially create a variable population for the selection of types with
desired combination of characters.
4. To combine the desired characters into a single individual.
5. To exploit and utilize the hybrid varieties.
111

 TYPES OF HYBRIDIZATION:
1. Intervarietal Hybridization:
→ The parents involved in hybridization belong to the same species.
→ In crop improvement programme this type of hybridization is commonly used
e. g crossing of two varieties of wheat or crops.
2. Intraspecific hybridization:
→ Gene flow between genetically distinct populations
→ Increases heterozygosity
→ Natural hybrids generally show intermediate phenotypes
→ Artificial hybrids may show transgressive phenotypes (e.g. maize)
112

3. Interspecific hybridization:
→ Gene flow between diverged species
→ Increases heterozygosity and can generate new polymorphisms
→ Hybrids may show intermediate, transgressive, or novel phenotypes
113

 Procedure of Hybridization:
 It involves the following steps:
1. Selection of parents.
2. Selfing of parents or artificial self-pollinat
3. Emasculation
4. BaggingTagging
5. Crossing
6. Harvesting and storing the F, seeds
7. Raising the F₁ generation.
114

 STEPS IN HYBRIDIZATION EXPERIMENT:
 SELECTION OF PURE BREEDING PARENTS:
1. Bagging
2. Seed setting
3. Collection of seeds
4. Raising of F, generation plants
5. F1, plants self pollinated
6. Seed setting of F, plants
7. Collection of seeds from F1, plants
8. Raising F2, generation plants
9. Raising F3 Plants-F4, plants-F5, plants--F7 plants. 115

 PROCESS OF HYBRIDISATION
 EMASCULATION:
1. Removal of anthers from bisexual flowers before they shed their pollen is known as
emasculation.
2. It is done in order to prevent self fertilisation.
 BAGGING:
1. After emasculation, flower buds are kept enclose in bags made up of cloth, plastic or
polythene etc.
2. It is done to prevent pollination through unknown pollen.
 TAGGING:
1. The female parents are then labelled properly.
2. The labelling should bear the following information.
3. Serial number
4. Details of male parents and female parents.
5. Date of emasculation and crossing
116

 CHARACTERISTICS OF HYBRIDISED PLANTS:
1. Hybridised plants have high resistance.
2. Hybridised plants have high yielding capacity.
3. Hybridised plants have high productivity.
4. Hybridised plants have life-longness.
5. Example: Brinjal, Ladyfinger, Chilli, Tomato etc.
117

THANK YOU!
118
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