Extraction, isolation and identification lect 1

1. Extraction and
Phytochemical Analysis of
Medicinal
Plants

2. Medicinal
plants
• The Medicinal plants constitute an
effective source of both traditional and
modern medicines.
• Herbal medicine has been shown to have
genuine utility and about 80% of rural
population dependson it as primary
health care. [WHO,(2005)]

3. Medicinal plants are the
richest bio-resource
drugs of traditional systems of medicine,
modern medicines,
nutraceuticals,
food supplements,
folk medicines,
pharmaceutical intermediates,
chemical entities for synthetic drugs.

4. Evidence for Ethnopharmacology
• De Materia Medica,
• Historia Plantarum,
• Species Plantarum.
have been variously published in attempt
to provide scientific information on the
medicinal uses of plants.

5. Methods in natural product
chemistry

6. Bioassay guided isolation
 Is the physical process used to isolate
biologically active chemicals from a natural
source.
 Biomass: any sample of biological origin
 It does aim to find new drug leads

7. Extraction of crude
natural material
Bioassay measurements
of the total extract
Chromatographic separation and isolation
of the constituents
Characterization of the
isolated compounds
Investigation of the
biosynthetic pathway
Bioactivity of pure
known compounds
Determination of
3D structure of
compound
Lead compound
Specific and selective
Chemical synthesis
Or modification

8. Extraction
• Is the separation of medicinally active
portions of plant (and animal) tissues using
selective solvents through standard
procedures.
• The products so obtained from plants are
relatively complex mixtures of metabolites,
in liquid or semisolid state or in dry
powder form (after removing the solvent),
& are intended for oral or external use.

9. Parameters for Selecting an Appropriate
Extraction Method
I. Authentication of plant material by botanist.
II. Use the right plant part +the age of plant +the time,
season &place of collection.
III. The nature of its chemical constituents.
IV. Grinding methods &powdering techniques.
V. Nature of constituents (polar/nonpolar(
VI. The quality of water / menstruum.
VII.The design &material of fabrication of the extractor.
VIII.Analytical parameters of the final extract,
(TLC/HPLC).

11. Selection of plant
• Plant based natural constituents can be derived
from any part of the plant like bark, leaves,
flowers, roots, fruits, seeds, etc.
• Plants are usually air dried to a constant weight
before extraction.
• oven drying: every part were cut into pieces
dried in an oven @ 60°C for 9 hrs.&
pulverized.
• Other method for drying the plants is the oven
drying at about 40°C for 72 h.

12. parameters influencing the
quality of an extract
• Plant part used as starting material
• Solvent used for extraction
• Extraction procedure

13. Steps Involved in the
Extraction of
Medicinal Plants
1. Size reduction
2. Extraction
3. Filtration
4. Concentration
5. Drying

14. 1. Size Reduction
Objective:
• Torupture plant organ, tissue &cell structures so
that its medicinal ingredients are exposed to the
extraction solvent.
• Size reduction maximizes the surface area, which in
turn enhances the mass transfer of active principle
from plant material to the solvent.
The 30-40 mesh size is optimal.
Hammer mill or a disc pulverizer which has built in
sieves controlled by varying the speed of the rotor
clearance b/w the hammers & the lining of the
grinder.

15. 2. Extraction
Medicinalplants
1.Cold aqueous
percolation
2.Hot aqueous extraction
(decoction(
3.Solvent extraction
(cold / hot)
Aromatic plants
1. Essential oils
2. Concretes
3. Bsolutes
4. Pomades
5. Esinoids

16. Choice of solvents
• Successful determination of biologically
active compounds depends on the type of
solvent used in the extraction procedure.
• The choice of solvent is influenced by what is
intended with the extract.

17. Properties of a good
solvent in plant extractions
 low toxicity,
 ease of evaporation at low heat,
 promotion of rapid physiologic absorption of
the extract,
 preservative action,
 inability to cause the extract to complex or
dissociate.

18. Water
• Water is universal solvent.
• used to extract plant products with antimicrobial
activity.
• Traditional healers use primarily water &
consistent antimicrobial activity is obtained.
Plant extracts: organic solvents >>> water extract.
• Water soluble flavonoids (mostly anthocyanins)
have no antimicrobial significance.
• only water soluble phenolics are important as
antioxidant compound.

19. Acetone
• Dissolves many hydrophilic and lipophilic
components.
• a very useful extractant, especially for
antimicrobial studies (phenolic group extract).
•extraction of tannins +phenolics:
aqueous acetone >>> aqueousmethanol
• Both acetone and methanol were found to extract
saponins  antimicrobial activity.

20. Alcohol
• The identified components from plants
(antimicrobial) =aromatic or saturated organic
compounds  most often obtained through initial
ethanol or methanol extraction.
• Ethanol, found easier to penetrate the cellular
membrane to extract the intracellular
ingredients(polyphenols) from the plant material.
• Methanol is more polar than ethanol but due toits
cytotoxic nature.
• The higher concentrations of more bioactive
flavonoid compounds were detected with
aqua/ethanol,70% due to its higher polarity than
pure ethanol.

21. Chloroform
• Used to obtain tannins and terpenoids.
• Terpenoid lactones successive
extractions of dried barks with
chloroform.

22. Ether
• Commonly used selectively for the
extraction of coumarins and fatty acids.

23. Dichloromethanol
• Specially used for the selective extraction of
only terpenoids.

24. Flow chart of general extraction
procedures

25. 3. Filtration

26. 4. Concentration
• The enriched extract from percolators or extractors, known
as miscella, is fed into a rota-evaporator where it is
concentrated under vacuum to produce a thickconcentrated
extract.
• The concentrated extract is further fed into a vacuum
chamber dryer to produce a solid mass free fromsolvent.
• The solvent recovered from the evaporator and vacuum
chamber dryer is recycled back to the percolatoror
extractor for the next batch of plant material.
• The solid mass thus obtained is
pulverized and used directly for
the desired pharmaceutical
formulations or further processed for
isolation of its phytoconstituents.

28. 5. Drying
• The filtered extract is subjected to spray drying
with a high pressure pump at a controlled feed
rate and temperature  to get dry powder.
• The desired particle size of the product is obtained
by controlling the inside temperature of the
chamber and by varying the pressure of the pump.
• The dry powder is mixed with suitable diluents or
excipients and blended in a double cone mixer to
obtain a homogeneous powder that can bestraight
away used (for example, for filling in capsules or
making tablets(.

30. Variation in extraction methods
• Length of the extraction period,
• Solvent used,
• pH of the solvent,
• Temperature,
• Particle size of the plant tissues,
• The solvent-to-sample ratio.

31. The general techniques of
medicinal plant extraction
 maceration,
 infusion,
 percolation,
 digestion,
 decoction,
 hot continuous extraction (Soxhlet),
 aqueous-alcoholic extraction byfermentation,
 counter-current extraction,
 microwave-assisted extraction,
 ultrasound extraction (sonication),
 supercritical fluid extraction,
 phytonic extraction (with hydrofluorocarbon
solvents).

32. Maceration
• The whole / coarsely powdered crude drug is
placed in a stoppered container with the solvent.
• Allow to stand at room temperature for a period of
at least 3 days with frequent agitation until the
soluble matter gets dissolved.
• The mixture then is strained, the marc (the damp
solid material) is pressed,
• The combined liquids are clarified by filtration or
decantation after standing.
• This method is best suitable
for use in case of the
thermolabile drugs.

33. Infusion
• Fresh infusions are prepared by macerating
the crude drug for a short period of time
with cold or boiling water.
• These are dilute solutions of the readily
soluble constituents of crude drugs.

34. Digestion
• This is a form of maceration in which gentle heat
is used during the process of extraction.
• It is used when moderately elevated
temperature is not objectionable.
• The solvent efficiency of the menstruum is
thereby increased.

35. Decoction
• In this process, the crude drug is boiled in a
specified volume of water (1:4) for a defined time,
• Volume is reduced to 1/4th the original
• It is then cooled and strained / filtered.
• This procedure is suitable for extracting  water-
soluble, heat-stable constituents.
• Typically used in preparation of Ayurvedic extracts
=“quath” / “kawath”

36. Percolation
• Used most frequently to
extract active ingredients in
the preparation of tinctures
and fluid extracts.
• The solid ingredients are
moistened with an
appropriate amount of
the specified
menstruum,
• Allowed to stand for
approximately 4 hours in a
well closed container.
• After stand time, the tap is
opened and allow to drain
slowly.

37. Hot Continuous Extraction
(Soxhlet(
• The finely ground crude drug is placed in a
porous bag or “thimble” made of strong filter
paper, which is placed in chamber of the
Soxhlet apparatus.
• The extracting solvent in flask is heated, and its
vapors condense in condenser.
• The condensed extractant drips into the
thimble containing the crude drug &extracts it
by contact.

38. Soxhlet
apparatus• When the level of liquid in chamber rises to the
top of siphon tube, the liquid contents of chamber
siphon into flask
• This process is continuous and is carried out until
a drop of solvent from the siphon tube does not
leave residue when evaporated.

39. Aqueous Alcoholic Extraction
by Fermentation
• Some medicinal preparations of Ayurveda (asava &
arista) adopt the technique of fermentation for
extracting the active principles.
• The extraction procedure involves soaking the
crude drug, [powder / a decoction (kasaya)], for a
specified period of time
• Undergoes fermentation &generates alcohol in situ.
• This facilitates the extraction of the active
constituents contained in the plant material.
• The alcohol thus generated also serves as a
preservative.

40. Counter-current Extraction
• Countercurrent distribution is a separation process that is
founded on the principles of liquid–liquid extraction where a
chemical compound is distributed (partitioned) between two
immiscible liquid phases (oil and water for example) according
to its relative solubility in the two phases.
• Wet raw material is pulverized using toothed disc
disintegrators to produce a fine slurry.
• Material to be extracted is moved in one direction generally
in the form of a fine slurry within a cylindrical extractor
where it comes in contact with extraction solvent.

41.  The further the starting material moves, the more
concentrated the extract becomes.
 Complete extraction is thus possible when the quantities of
solvent more than the material. Their flow rates should be
optimized.
sufficiently
concentrated
extract comes out
at one end ofthe
extractor while
the marc,
practically free of
visible solvent
falls out from the
other end

42. Ultrasound
Extraction
(Sonication(• The procedure involves the use of ultrasound with
frequencies ranging from 20 kHz to 2000 kHz.
•This increases the permeability of
cell walls &produces cavitation.
Eg:extraction of rauwolfia root.
• Deleterious effect: Ultrasound energy (>20 kHz) on
the active constituents of medicinal plants through
formation of free radicals and consequently
undesirable changes in the drug molecules.

43. Supercritical Fluid Extraction
• Cylindrical extraction vessels are used.
• The collection of the extracted analyte following SFEis
another important step: significant analyte loss can
occur during this step.
• CO2 as the extracting fluid.
• Organic solvents are frequently added to the CO2
extracting fluid to alleviate the polarity limitations
• The component recovery rates generally increase with
increasing pressure/temperature.

45. MICROWAVE EXTRACTION
•Microwave-assisted extraction is a process that removes solutes from
a solid matrix into a solvent.
•Phenomena such as electromagnetic transfer, heat transfer, mass
transfer, and momentum transfer make the process complex.
•Accurate and controlled heat is possible because of the capacity of
microwave radiation to penetrate and combine with a substrate.
•Therefore, the microwave procedure can be designed to transport
electromagnetic energy with specific power to the location of the
compounds of interest in the substrate.
•The energy-saving factors and short processing times lead to a
reduction in manufacturing costs, and improvement of product
uniformity and yields, resulting in products with high quality compared
with other extraction techniques.

47. Aromatic
Plant Extracts
• essential oils,
• concretes,
• absolutes,
• pomades
• resinoids.

49. Essential
oils
• Used in a wide variety of consumer goods viz.,
detergents, soaps, toilet products, cosmetics,
pharmaceuticals, perfumes, confectionery food
products, soft drinks, distilled alcoholic
beverages (hard drinks) and insecticides.
• Production technology is an essential element
to improve the overall yield &quality of
essential oil.

50. Presence of Essential Oils in plant parts.

51. Hetreogenous chemical group present in
essential oils

52. Method
s
1. Distillation:
• water distillation.
• water and steam distillation.
• direct steam distillation.
2. Hydrolytic maceration distillation.
3. Expression.
4. Cold fat extraction / Enfleurage.

53. Water and steam distillation

54. • Distillation methods are good for powdered
almonds, rose petals and rose blossoms.
• Maceration is adaptable when oil yield from
distillation is poor.
• Solvent extraction is suitable for expensive,
delicate &thermally unstable materials like
jasmine, tuberose, and hyacinth.

55. Concrete
• This is an extract of fresh flowers, herbs, leaves and the
flowering tops of plants obtained by the use of a hydrocarbon
solvent such as butane, pentane, hexane and petroleumether.
• Concrete is rich in hydrocarbon soluble material &devoidof
water-soluble components.
• It is generally a waxy, semisolid, dark-colored material free
from the original solvent.
• concretes are produced in
static extractors.
• It is a normal practice to circulate fresh
solvent through a battery ofextractors.

56. Absolutes
• Tomake an absolute, the concrete is mixed with absolute alcohol
&agitated thoroughly in a vessel with an agitator.
• During agitation, the temperature is kept at 40°-60° Cand the
concrete is immersed in the solution.
• The solution is cooled down to -5° to -10° Cto precipitate outthe
wax, since waxes are normally insoluble in alcohol below -1° C.
• The precipitated wax is removed by passing the solution through
a rotary filter.
• The filtrate from the rotary filter is pumped into a primary
evaporator, where it is concentrated to about 10% alcohol
content.

57. Resinoids
• Resinoid is an extract of naturally resinous material,
made with a hydrocarbon solvent.
• Resinoids are usually obtained from dry materials.
• The extraction process is same as that of concrete
production, except that perforated discs are not used
for stacking the material;
•instead powder from dry plant
material is fed into the extractor.

58. Pomades
•Pomades are obtained by a process known as
enfleurage, which is a cold fat extractionmethod.
• The fat is spread out on glass plates contained in wooden
frames, leaving a clear margin near theedges.
• The absorptive surface of the fat is increased by surface
grooves made with a wooden spatula.
• Fresh flowers are spread out on the surface of the fat andthe
frames are stacked in piles.
• After the perfume oils have been absorbed from theflowers,
the spent flowers are removed byhand.
• Fresh flowers are again spread on the fatsurface.
• This is repeated until the fat surface is completely enriched
with perfume oils.
• The pomade so obtained is ready for cold alcoholic extraction

59. Standardized
Extraction
The purpose of standardized extraction
procedures for crude drugs
(medicinal &aromatic plant parts)
 Toattain the therapeutically desired portions
 Toeliminate unwanted material by treatment
with a selective solvent known as “menstrum”

60. The extract thus obtained, after standardization,
may be used as medicinal agent
• as such in the form oftinctures
• fluid extracts
• further processed to be incorporated in any
dosage form such as tablets and capsules

61. STANDARIZATION
Requirement to have a minimum amount of one or
several compounds or groups of compounds in the
extract
Applies only to extracts
Standardization guarantees the content of one or
more active constituents and marker compounds.
The plant environment and genetic factors could
significantly affect the biochemical components of the
plant extract.

62.  Standardization involves adjusting the herbal drug
preparation to a defined content of a constituent or a
group of substances with known therapeutic activity
Botanical extracts made directly from crude plant
material show substantial (considerable) variation in
composition, quality, and therapeutic effects.
 Standardized extracts are high-quality extracts
containing consistent levels of specified compounds,
and they are subjected to accurate quality controls
during all phases of the growing, harvesting, and
manufacturing processes.

63.  No regulatory definition exists for standardization of dietary
supplements. As a result, the term “standardization” may
mean many different things. Some manufacturers use the
term standardization incorrectly to refer to uniform
manufacturing practices; following a recipe (formula) is not
sufficient for a product to be called standardized.
Therefore, the presence of the word “standardized” on a
supplement label does not necessarily indicate product
quality. When the active principles are unknown, marker
substance(s) should be established for analytical purposes
and standardization.
 Marker substances are chemically defined
constituents of a herbal drug that are important for the
quality of the finished product. Ideally, the chemical
markers chosen would also be the compounds that are
responsible for the botanical’s effects in the body.

64.  Special extract: a process of defining a
range for certain compounds or classes of
compounds in order to enrich desired
compounds and reducing the amount of
undesired ones.

65. Why is standardization necessary and
important?
 Reproducible composition and higher quality of
the product
 Provided that the product is registered, it thus
becomes a medicine that should comply with
the basic standards required for all drugs
 Standardization allows comparison of the
clinical effectiveness, pharmacological
effects and side effects of a series of products
 Such products give patients greater security and
increase their trust
 Ensuring the quality of the products sold is a key
responsibility of the pharmacist

66. There are two types of standardization
I. In the first category, “truly” standardized
extract (Type A):
 Extracts standardized to active constituents
(single or groups)
Example: belladonna leaf dry extract, standardized to
0.95-1.05% of alkaloids calculated as hyoscyamine

67. II. In the second category, “quantified ”
extract (Type B1):
 Standardized to constituents that contribute to
the activity, by blinding (randomize) different
batches of herbal drug before extraction or by
mixing different lots of herbal drugs
preparations. Adjustment using excipients is not
accepted
 Includes the special extracts

68. Example: Ginkgo with its 26% ginkgo flavones and
6% terpenes is a classic example.
 These products are highly concentrated and no
longer represent the whole herb, and are now
considered as phytopharmaceuticals. In many
cases they are vastly (greatly) more effective than
the whole herb.

69. III. In the third category, “other ” extracts
(Type B2):
 The other type of standardization is based on
manufacturers guaranteeing the presence of a
certain percentage of lead compounds of
unknown pharmacological relevance serve as
quality marker compounds ; these are not
indicators of therapeutic activity or quality of the
herb. Nevertheless, the extract is
pharmacologically active.
 Give information on the over all quality of the
phytomedicine
Example: Echinaceae radix

70. Phytochemical
screening methods

71. Quantitative Analysis
Total Phenolic Content
Determined by Folin-Ciocalteau
assay method (Singleton and
Rossi, 1965)
Instrument: UV-Vis
Spectrophotometer, absorbance
measured at 765 nm
Expressed as Gallic acid
equivalent (GAE) in milligrams
per gram of fresh leaf
Total Flavonoid Content
Determined by Colourimetric
method (Yun et al., 2009)
Instrument: UV-Vis
Spectrophotomer, absorbance
measured at λ415 nm
Expressed as mg rutin equivalent
(mg RE) per gram of fresh leaf

72. Yadav and Agarwala, 2011
Assam, India
Phytochemical Analysis of Some Medicinal Plants
Objective
To carry out qualitative and quantitative phytochemical analysis of
selected medicinal plants

73. Methodology
Plant sources
Bryophyllum pinnatum (Leaves)
Ipomea aquatica (Leaves)
Oldenlandia corymbosa
(Whole plant)
Ricinus communis (roots)
Terminalia bellerica (Leaves)
Tinospora cordifolia
(Leaves/Stem)
Xanthium strumarium (Leaves)

74. Hot water extraction
5gm of dried finely powdered plant
material mixed with200ml of distilled
water
Heated on a hot plate with continuous
stirring at 30º-40ºC for 20 minutes
Solvent extraction
20gm powdered plant material packed
into a thimble and extracted with 250ml
of solvents
Extraction continues for 24 hours or till
the solvent in siphon tube of an extractor
become colourless
filtered through filter paper
kept on hot plate and heated at 30-40ºC
Preparation of extracts

75. Table 1. Phytochemical constituents of medicinal plants
Results and discussion
L = leaves; S = stem

76. Fig 1. Total
phenolic
content
Fig 2. Total flavonoid
content

77. Conclusion
 Results revealed that extracts from these plants can be
used as a good source for drugs
 Further work should be carried out to isolate, purify and
characterize the active constituents responsible for the
activity of these plants

78. Pranoothi et. al., 2014
Andhra Pradesh, India
Studies on Qualitative, Phytochemical Analysis and Screening of In
Vitro Biological Activities of Leucas indica (L)
Objective
To carry out qualitative and quantitative phytochemical analysis of
aerial parts of Leucas indica (L)

79. Methodology
Cleaned, shade dried,
mechanically
grinded & coarsely
powdered
Aerial parts of Leucas indica
Preparation of extracts
Powdered material
Subjected to
Solvent extraction with hexane, acetone, methanol & water
Extracts were concentrated using Rotary Evaporator
Phytochemical screening

80. Table 6. Physico-chemical evaluation
Solvent
Initial weight of
the powder (g)
Final weight of
the powder (g)
Weight of the
crude extract
(g)
Hexane 50 44.563 5.437 10.874 Dark brown
Acetone 50 40.415 9.585 19.17 Dark green
Methanol 50 35.552 14.448 28.896 Dark green
Water 50 38.621 11.379 22.758 Dark red
Crude
extract (%)
Colour of the
extract
Figure 3. Yield of extracts
Results and discussion

81. Table 7. Phytochemical analysis of whole aerial part extracts of Leucas indica (L)
Sl. No.Tests
Extracts
1 Alkaloids
Hexane Acetone Methanol Water
2
Mayers
Dragon
Wagners
Hagers
Phenolics
FeCl2 test
-
-
-
-
-
+
+
+
+
+
+
-
+
-
+
+
+
+
+
+
3 Flavonoids
Lead acetate test - + + +
4
5
NaOH test
Ethyl acetate test
Anthraquinones
Borntrager’s test
Steroids
Salkowski’s test
-
-
-
+
+
-
-
+
+
-
-
+
-
-
-
+

82. 6 Tannins
Hexane Acetone Methanol Water
FeCl2 test - + + -
Lead acetate test - + + -
Potassium dichromate test - + + -
7 Saponins
Vigorous shaking test + + + +
8 Anthocyanins
Ammonia-HCL test - - - -
9 Leuco-Anthocyanin
Iso amyl alcohol test - - - -
10 Coumarins
NaOH test - - - -
11 Reducing sugars
Keller-Kiliani test + + + +
Sl. No. Tests
Extracts
Contd...

83. Table 8. Total phenol content
)µg/ml( extract extract extract
100 15.69 18.5 25.6 22.8
200 22.54 26.9 45.86 41.8
300 30.41 38.9 72.8 68.4
400 42.89 54.78 88.96 76.5
500 54.58 65.8 105.68 95.8
% of phenol content µg GAE/µg
Concentration Hexane Acetone Methanol Water
extract
Figure 4. Total phenol content

84. Table 9. Total flavonoid content
% of phenol content µg GAE/µg
Hexane Methanol
100 00 10.28 17.25 00
200 00 15.56 26.35 00
300 00 32.47 29.12 13.20
400 00 41.58 43.52 18.25
500 00 50.24 62.34 24.69
Concentration Acetone Water
Figure5.dionovalf latoTtnetnoc

85. Conclusion
 The presence of most general phytochemicals in Leucas
indica might be responsible for their therapeutic effects
 It further reflects a hope for the development of many more
novel chemotherapeutic agents from plants which in future
may serve for the production of synthetically improved
therapeutic agents

86. Pandey et. al., 2014
Uttarakhand, India
Phytochemical Screening of Selected Medicinal Plant
Cinnamon zeylanicum Bark Extract
Objective
To screen the phytochemicals present in Cinnamon zeylanicum
aqueous bark extract

87. Washed, shade dried and powdered
Barks of Cinnamon zeylanicum
Preparation of extracts
Maceration technique
Powdered material mixed with 150ml distilled water for 1 hour
in rotary shaker
Extract was filtered using muslin cloth & Whatman filter paper
Concentrated by evaporation on water bath
The extract was dried & used as powder
Methodology

88. Phytoconstituents water water water
Carbohydrates
15ºC
+
70ºC
+
45ºC
+ - - -
Steroids + + + + + +
Proteins - - - - - -
Glycosides - - - - - -
Alkaloids + + + + + +
Flavonoids + - - + - +
Saponins + + + + + +
Tannins & phenol - - - + + +
Table 16. Phytochemical screening of secondary metabolites of plant extracts
Extracts
Cold Hot Warm Ethanol Methanol Acetone
Results and discussion

89. Conclusion
Your home work

90. Singh and Bag, 2014
Manipur, India
Phytochemical Analysis and Determination of Total
Phenolics Content in Water Extracts of Three
Species of Hedychium
Objective
To identify and compare the bioactive constituents
present in Hedychium species and determine total
phenolic content

91. Hedychium rubrum
Cleaned,shadedried,mechanically
grinded & coarsely powdered
Preparation of extracts
Extracts were concentrated using Rotary Evaporator
Phytochemical screening
Powdered material
Subjected to
Solvent extraction with hexane, acetone, methanol & water
Hedychium spicatum
Hedychium coronarium
Methodology

92. Table 17. Comparative analysis of phytochemical constituents of three different species of
Genus Hedychium
Phytochemical Chemical tests
constituents H. spicatum
Water extract
H. coronarium H. rubrum
Alkaloids
Carbohydrates
(reducing sugar) Fehling’s test + + +
Proteins Xanthoproteic test + + +
Flavonoids Alkaline reagent test + + +
Phenolic compounds Lead acetate test + + +
Tannins Lead acetate test
Ferric chloride test
+
-
+
+
+
+
Steroids & terpenoids Salkowski’s test + + +
Saponins Froth test + + +
Cardiac glycosides Keller-killiani test + + +
Oil + + +
Phlobatannin - - +
Hager’s test
Benedict’s test
-
-
--
-+
Results and discussion

93. Table 18. Total phenolic content in the water extracts of H. Spicatum, H.
Coronarium and H. rubrum
Water extracts Concentration
)mg/ml(
mg of gallic acid/g of extract
Mean ± (Standard Deviation(
H. Spicatum 1 29.39 ± 0.01
H. Coronarium 1 34.93 ± 0.01
H. rubrum 1 66.48 ± 0.01
Figure 8. Callibaration curve of gallic acid

94. Conclusion
Your home work

95. Vastrad et. al., 2015
Karnataka, India
Identification of Bio-active Components in Leaf Extracts
of Aloe vera, Ocimum tenuiflorum (Tulasi) and
Tinospora cordifolia (Amrutballi)
Objective
To screen various bio-active compounds present in the leaf
extracts of A. vera, O. tenuiflorum and T. cordifolia and evaluate total
phenolic content & total flavonoid content

96. Tinospora
cordifolia
Cleaned,shadedried,mechanically
grinded & coarsely powdered
Preparation of extracts
Extracts were concentrated using Rotary Evaporator
Phytochemical screening
Powdered material
Subjected to
Solvent extraction with hexane, acetone, methanol & water
Aloe vera
Ocimum tenuiflorum
Methodology

97. Figure 9 . Yield of extracts
Results and discussion

98. Table 20. Total phenolic content (TPC) of the plant leaf extracts
Total phenolic content (GAE* mg/g)
Extraction
solvent
Aqueous
Ethanol
A. vera
94.42 ± 4.92
138.13 ± 6.63
O. tenuiflorum
80.82 ± 8.63
113.07 ± 9.81
114.34 ± 11.86
T. cordifolia
465.82 ± 23.04
264.06 ± 18.41
301.42 ± 29.69Methanol 95.20 ± 3.23
GAE = Gallic acid equivalent
Fig 10. Total phenolic content (TPC): Calibration curve

99. Table 21. Total flavonoid content (TFC) of the plant leaf extracts
Figure 11. Total flavonoid content (TFC): Calibration curve
Total flavonoid content (RE* mg/g)
Extraction
solvent
Aqueous
Ethanol
Methanol
RE = Rutin equivalent
A. vera
72.28 ± 8.70
76.50 ± 8.57
88.59 ± 8.38
O. tenuiflorum
61.84 ± 7.25
95.46 ± 4.12
96.34 ± 5.85
T. cordifolia
178.43 ± 6.61
208.36 ± 2.86
132.59 ± 7.59

100. Conclusion
Your home work