Extraction and phytochemical analysis of medicinal plants
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Transcript of Extraction and phytochemical analysis of medicinal plants
2
Seminar on
Extraction and Phytochemical Analysis
of Medicinal Plantsby
Shameembanu A. ByadgiShameembanu A. Byadgi
Medicinal plants
Medicinal plants constitute an effective source of both
traditional and modern medicines
Herbal medicine has been shown to have genuine utility
About 80% of rural population depends on it as primary
health care. [WHO, (2005)]
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
Natural bioactive compounds found in different parts of plant
(fruit, flower, stem, leaf, root) Provide definite physiological action on the human body Bioactive substances include tannins, alkaloids, carbohydrates,
terpenoids, steroids and flavonoids Widely used in the human therapy, veterinary, agriculture,
scientific research and countless other areas Have inhibitory effects on all types of microorganisms in vitro
Phytochemicals
Extraction
……… is the separation of medicinally active portions of
plant tissues using selective solvents through standard procedures
The basic parameters influencing the quality of an extract
•Plant part used as starting material
•Solvent used for extraction
•Extraction procedure
Choice of solvents
Successful determination of biologically active compounds depends on the type of solvent used in the extraction procedure
Property of a good solvent in plant extraction
•Low toxicity
•Ease of evaporation at low heat
•Promotion of rapid physiologic absorption of the extract
•Preservative action
The factors affecting the choice of solvent
•Quantity of phytochemicals to be extracted
•Rate of extraction
•Diversity of different inhibitory compounds extracted
•Ease of subsequent handling of the extracts
•Toxicity of the solvent in the bioassay process
•Potential health hazard of the extractants
Solvents used for active component extraction
Water Ethanol Methanol Chloroform Ether AcetoneAnthocyaninsStarchesTanninsSaponinsTerpenoidsPolypeptides
Lectins
TanninsPolyphenolsPolyacetylenesFlavonolsTerpenoidsSterols
Alkaloids
AnthocyaninsTerpenoidsSaponinsTanninsXanthoxyllinesTotarolQuassinoidsLactonesFlavonesPhenones
Polyphenols
Terpenoids
Flavonoids
AlkaloidsTerpenoidsCoumarins
Fatty acids
Phenol
Flavonols
General techniques of medicinal plant extraction
Plant tissue homogenization
Maceration
Infusion
Percolation
Digestion
Decoction
Soxhlet extraction (Hot continuous extraction)
Sonication (Ultrasound extraction)
Plant tissue homogenization
Maceration
Whole / coarsely powdered crude drug is placed in a stoppered container with the solvent
Allow to stand @ 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
Infusion
Digestion
• A form of maceration in which gentle heat is used during the process of extraction
• Used when moderately elevated temperature is not objectionable
• The solvent efficiency of the menstruum is thereby increased
Microwave digestion system
Decoction
Suitable for extracting water-soluble, heat-stable constituents
Typically used in preparation of Ayurvedic extracts
Percolation
• Used most frequently to extract active ingredients in the preparation of 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 mass is packed & the top of the percolator is closed
• The mixture is allowed to macerate in the closed percolator for 24 h
,• Additional menstruum is added as required, until the
percolate measures about three-quarters of the required
volume of the finished product
• The marc is then pressed and the expressed liquid is
added to the percolate
• Sufficient menstruum is added to produce the required
volume
• The mixed liquid is clarified by filtration or by standing
followed by decanting
Soxhlet Extraction (Hot Continuous Extraction)
Sonication (Ultrasound Extraction)
• Involves the use of ultrasound with frequencies ranging from 20 kHz to 2000 kHz
• Increases the permeability of cell walls & produces cavitation
Disadvantage Deleterious effect of ultrasound energy
(>20 kHz) on the active constituents of medicinal plants through formation of free radicals and consequently undesirable changes in the drug molecules
Effect of extracted plant phytochemicals depends on
• The nature & origin of the plant material
• Degree of processing
• Moisture content
• Particle size
Variation in extraction methods
• Length of the extraction period • Solvent used• pH of the solvent • Temperature• Particle size of the plant tissues• Solvent-to-sample ratio
Phytochemical screening methods
Phytochemicals Tests Reagents Positive resultsAlkaloids Dragendorff test Dragendorff’s reagent Prominent yellow ppt
Wagner test Wagner’s reagent Reddish brown pptMayer test 1% HCl, Mayer’s
reagentTurbid extract is obtained
Flavonoids Ammonia test 1% NH3 Yellow colourSodium hydroxide test
20% NaOH, HCl Yellow colour turns to colourless
Tannins Ferric chloride test 5% FeCl3 Blue-black or blue-green colouration
Phenolic compounds
Gelatine test 1% gelatine solution containing 10% NaCl
White ppt
Lead acetate test 10% lead acetate Bulky white pptSaponins Foam test 20 mL distilled water
(mixed vigorously for 15 minutes)
Presence of froth
Qualitative Analysis
Phytochemicals Tests Reagents Positive resultsTerpenoids Salkowski test 0.5 mL chloroform,
1 mL conc. H2SO4
Reddish brown colouration at the interface
Carbohydrates Molish test Conc. HCl Violet ringFehling test Conc. HCl & Mg
turningsYellow & brick red ppt
Proteins Biuret test 4% NaOH, 1% CuSO4 Violet or pink colourGlycosides Legal’s test Pyridine, sodium
nitroprussidePink to red colour
Kellar killani test Glacial acetic acid, 5% FeCl3
Reddish brown & bluish green colour
Contd....
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
Yadav and Agarwala, 2011Assam, India
To carry out qualitative and quantitative phytochemical analysis of selected medicinal plants
Phytochemical Analysis of Some Medicinal Plants
Objective
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)
Hot water extraction
5gm of dried finely powdered plant material mixed with 200ml of distilled
water
Heated on a hot plate with continuous stirring at 30º-40ºC for 20 minutes
filtered through filter paper
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
kept on hot plate and heated at 30-40ºC
Preparation of extracts
Phytochemical Analysis
Qualitative Quantitative
Total phenolic content FolinCiocalteu reagent expressed as
gallic acid equivalent, mg/g of extracted compound
Total flavonoid content Aluminium chloride colourimetric
method expressed as quercetin equivalent, mg/g of extracted compound
Proteins (Millon’s & Ninhydrin test) Carbohydrates (Fehling’s,
Benedict’s, Molisch’s & Iodine test) Phenols & tannins (Ferric chloride
test) Flavonoids (Shinoda & alkaline
reagent test) Saponins (Foam test) Glycosides (Liebermann’s,
Salkowski’s & Keller-kilani test) Steroids Terpenoids Alkaloids (Mayer’s & Wagner’s test)
Table 1. Phytochemical constituents of medicinal plants
Results and discussion
L = leaves; S = stem
Fig 1. Total phenolic content
Fig 2. Total flavonoid content
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
Badugu, 2012
To screen the presence of phytochemicals and estimate the amount of total phenols and flavonoids in Cyamopsis tetragonoloba
Phytochemical Screening, Quantitative Estimation Total Phenolics and Total Flavonoids, Antimicrobial Evaluation of Cyamopsis tetragonoloba
Objective
Methodology
Seeds of Cyamopsis tetragonoloba
Washed, shade dried & powdered mechanically
Preparation of extractsPowdered material was weighed
Soxhlet extraction using methanol, acetone, chloroform & hexane
Solvent was recovered using Rotary Vacuum Evaporator
Phytochemical screening
Subjected to
48 hrs
Phytochemical Analysis
Qualitative Quantitative
Total phenolic content Folin-Ciocalteu reagent expressed in
terms of milligrams of catechol per gram of dry weight
Total flavonoid content Aluminium chloride colourimetric
method expressed as catechin equivalent, mg catechin/g dried extract
Carbohydrates, reducing sugars, monosaccharides, alkaloids, saponins & tannins (Evans, 1996)
Flavonoids (Shinoda’s test) Terpenes/ steroids (Liebermann-
Burchard’s test) Anthraquinones (Borntrager’s test) Cardiac glucosides (Sodium nitro
proside method) Proteins (Copper sulphate & Folin-
Ciocalteau solution) Amino acids (Ninhydrin test)
Table 2. Phytochemical screening of Cyamopsis tetragonoloba
Sl. No.
Secondary metabolites
TestsSolvents
Methanol Acetone Chloroform Hexane
1 Carbohydrates Molisch’s test +ve +ve +ve +ve2 Reducing sugars Fehling test +ve +ve -ve -ve3 Monosaccharide Barfoed’s test -ve -ve -ve -ve4 Tannins Ferric chloride test
Lead sub acetate test-ve -ve -ve +ve
5 Saponins Frothing test +ve -ve -ve -ve6 Flavonoids Shinoda’s test +ve +ve -ve -ve7 Terpenes/ steroids Liebermann Burchard’s
test+ve -ve +ve -ve
8 Alkaloids Mayer’s testWagner’s test
+ve +ve -ve -ve
9 Cardiac glucosides
Sodium nitroproside +ve +ve -ve +ve
10 Proteins Copper sulphate Folin Ciocalteau solution
-ve -ve -ve -ve
11 Amino acids Ninhydrin -ve +ve -ve +ve12 Anthraquinones Borntrager’s test -ve -ve -ve -ve
Results and discussion
Table 3. Total phenolic and flavonoids content of different extracts of Cyamopsis tetragonoloba
Extract Total phenolic contentmg of Catechin equivalents/
200mg dried extract
Total flavonoids contentmg of Catechol equivalents/
200mg dried extractMethanol 14.51 17.34Acetone 9.17 8.75Chloroform 11.32 14.05Hexane 5.53 6.41
Conclusion
The study indicated that methanol is used as best solvent for extraction of phytochemicals
Other solvents like chloroform, acetone and hexane are less commonly used for the extraction of phytochemicals
The herbal extract can be used for curing diseases without any side effects
Hasanuzzaman et. al., 2013Bangladesh
To investigate total phenolic content of methanolic extract of Averrhoa bilimbi fruits
Evaluation of Total Phenolic Content, Free Radical Scavenging Activity and Phytochemical Screening of Different Extracts of Averrhoa
bilimbi
Objective
Methodology
Fruits of Averrhoa bilimbi
Washed, sliced & dried under sun
Preparation of extracts
Powdered fruits (500g)
2500 ml methanol for 15 days at room temperature
Extract filtered using filter cloth & Whatman’s filter paper
Filtrate evaporated under ceiling fan & in a water bath below 40°C until dried
Soaked in
Cold maceration technique
Phytochemical Analysis
Qualitative Quantitative
Total phenolic content Folin-Ciocalteu reagent (oxidizing
agent) & gallic acid (standard) expressed as mg of GAE (gallic acid equivalent) / gm of the extractive
Alkaloids (Mayer’s & Dragendoff’s test) Tannins (FeCl3 test) Saponins (Foam test) Flavonoids (chip of magnesium & HCl) Glycosides (NaCl & Fehling’s solutions
A & B) Steroids & triterpenes (Ethylic, H2SO4
& anhydride acetic) Phenols (FeCl3 & K3Fe(CN6)) Cardiac glycosides (Acetic, FeCl3 &
conc. H2SO4) Carbohydrates (Alcoholic α-naphthol,
Benedict’s reagent)
Table 4. Phytochemical screening of different extractives of Averrhoa bilimbi fruits
TestsExtracts
MEF PTSF CTSF CSF AQSF
Alkaloids + + - + +
Tannins + + + - -
Saponins + + - - +
Flavonoids + + + + +
Cardiac glycosides + + - + +
Glycosides + + + + +
Phytosterols - - - - -
Triterpenes + + - + -
Phenols + + + + +
Carbohydrates + - + + +MEF = methanolic extract; PTSF = pet-ether soluble fraction; CTSF = carbon tetrachloride soluble fraction; CSF = chloroform soluble fraction; AQSF = aqueous soluble fraction
Results and discussion
Table 5. Total phenolic content of different extractives of Averrhoa bilimbi fruits
Extractives Total phenol content (mg of GAE/g of extractive)
Methanolic extract 65.16 ± 0.52
Pet-ether soluble fraction 55.31 ± 1.01
Carbon tetrachloride soluble fraction 52.00 ± 0.90
Chloroform soluble fraction 68.67 ± 0.94
Aqueous soluble fraction 50.23 ± 0.56
GAE = Gallic acid equivalent
Conclusion
It may be concluded that Averrhoa bilimbi fruits is a good source of phytochemicals
Different extractives showed significant total phenolic content
Pranoothi et. al., 2014Andhra Pradesh, India
To carry out qualitative and quantitative phytochemical analysis of aerial parts of Leucas indica (L)
Studies on Qualitative, Phytochemical Analysis and Screening of In Vitro Biological Activities of Leucas indica (L)
Objective
Methodology
Aerial parts of Leucas indica
Cleaned, shade dried, mechanically grinded & coarsely powdered
Preparation of extracts
Powdered material
Solvent extraction with hexane, acetone, methanol & water
Extracts were concentrated using Rotary Evaporator
Phytochemical screening
Subjected to
Phytochemical Analysis
Qualitative Quantitative
Total phenolic content FolinCiocalteu reagent expressed as
µg of gallic acid equivalents per gram dry mass of extract (µg GAE/gDM)
Total flavonoid content Aluminium chloride colourimetric
assay expressed with the rutin equivalents per g of dried fraction)
Alkaloids Steroidal compounds Phenolic compounds Flavonoids Saponins Tannins Coumarins Cardiac glycosides
Table 6. Physico-chemical evaluation
SolventInitial weight of the powder (g)
Final weight of the powder (g)
Weight of the crude extract
(g)
Crude extract (%)
Colour of the extract
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
Figure 3. Yield of extracts
Results and discussion
Table 7. Phytochemical analysis of whole aerial part extracts of Leucas indica (L)
Sl. No. TestsExtracts
Hexane Acetone Methanol Water
1 Alkaloids
Mayers - + + +
Dragon - + - +
Wagners - + + +
Hagers - + - +
2 Phenolics
FeCl2 test - + + +
3 Flavonoids
Lead acetate test - + + +
NaOH test - + + -
Ethyl acetate test - - - -
4 Anthraquinones
Borntrager’s test - - - -
5 Steroids
Salkowski’s test + + + +
6 Tannins
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. TestsExtracts
Hexane Acetone Methanol Water
Contd...
Table 8. Total phenol content
% of phenol content µg GAE/µgConcentration
(µg/ml)Hexane extract
Acetone extract
Methanol extract
Water 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
Figure 4. Total phenol content
Table 9. Total flavonoid content
% of phenol content µg GAE/µgConcentration
(µg/ml)Hexane extract
Acetone extract
Methanol extract Water extract
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
Figure 5. Total flavonoid content
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
Rajesh et. al., 2014Tamil Nadu, India
To evaluate the phytochemicals from aqueous, ethanolic and petroleum ether extracts of four fern species
Qualitative and Quantitative Phytochemical Analysis in Four Pteridophytes
Objective
Methodology
Washed & cut into small pieces
Shade dried & then hot air oven drying at 55-60ºC
Dried leaves, stems & roots were pulverized using mixer
Strained through 25 mm mesh diameter sieve to obtain fine dust
Phytochemical screening
Actinopteris radiata
Pityrogramma calomelanos
Drynaria quercifolia
Dryopteris cochleata
Phytochemical Analysis
Qualitative Quantitative
Total tannins Folin-Ciocalteu reagent expressed as
mg tannic acid equivalent per 100g of the sample)
Total phenols Folin-Ciocalteu reagent method
expressed in mg of gallic acid equivalent (GAE) per g of dry weight of plant powdres
Alkaloids (Mayer’s test) Anthocyanin & betacyanin (NaOH test) Cardio glycosides Coumarins Flavonoids (NaOH test) Glycosides Phenols Quinones Saponins (Foam test) Steroids Tannins Terpenoids
Table 10. Qualitative analysis of phytochemicals in four fern species
Table 11. Quantitative analysis of phytochemicals (mg/g) in four fern species
Phytochemicals A.radiata D.quercifolia D.cochieata P.calamelanos
Total tannin 12.189 ± 0.258 6.332 ± 0.187 9.405 ± 0.299 17.181 ± 0.441
Total phenol 10.962 ± 0.327 7.131 ± 0.184 8.912 ± 0.310 13.581 ± 0.481
Results and discussion
Conclusion
The results revealed that ethanolic solvent performed well exhibiting the presence of major phytochemicals compared to aqueous and petroleum ether extracts
Quantitative analysis showed highest content of tannins and phenols in Pityrogramma calomelanos fern extract followed with least content of tannins and phenols in Actinopteris radiata, Dryopteris cochleata and Drynaria quercifolia
Vastrad et. al., 2014Karnataka, India
To screen the presence of phytochemicals in leaf extracts of C. fistula, P. pinnata, T. grandis & J. curcas and assess total phenolic & total flavonoid content
Characterization of Phytoconstituents in Leaf Extracts of Forest Species for Textile Applications
Objective
Methodology
Cleaned, shade dried, chopped & ground in laboratory mortar & pestle
Ground leaf was mixed with the solvent & incubated (200 strokes) in incubator cum shaker
for 24 hours at 25ºC
The extract was centrifuged (5000 rpm) at room temperature & supernatant separated
Residue was re-extracted with the respective solvent
The extract was filtered using Whatman filter paper & yield was measured
Cassia fistula
Pongamia pinnata
Tectona grandis
Jatropha curcas
Phytochemical Analysis
Qualitative Quantitative
Total phenolic content Folin-Ciocalteu assay method
expressed as gallic acid equivalent in milligrams per gram of fresh leaf)
Total flavonoid content Colourimetric method expressed as
milligrams of rutin equivalent (mg RE) per gram of sample)
Tannins & phenolic compounds Flavonoids Alkaloids Saponins
Table 12. Qualitative phytochemical screening of forest species
Figure 6. Total phenolic content (mg per gram of fresh leaf)
Results and discussion
Table 13. Total flavonoid content (TFC) of extracts in forest species
Forest speciesTFC (mg/g of sample)
Ethyl alcohol Methanol Aqueous
Cassia fistula 106.4 126.21 9.49
Pongamia pinnata 121.53 148.33 8.96
Tectona grandis 153.52 179.1 16.71
Jatropha curcas 149.70 138.7 -
Conclusion
The results revealed that phytochemicals viz., alkaloids, flavonoids, tannins and terpenoids were found in appreciable amount in the selected forest species
Methanolic extract of C. fistula, P. pinnata and T. grandis exhibited maximum TPC and TFC content while J. curcas depicted higher amount of TPC and TFC content in ethanolic extract
Therefore it can be concluded that extracts from forest species such as C. fistula, P. pinnata and T. grandis and J. curcas can be used for applying eco friendly and healthy finishes to textile substrates
Geetha and Geetha, 2014Tamil Nadu, India
To carry out qualitative and quantitative phytochemical analysis of lemongrass leaves
Phytochemical Screening, Quantitative Analysis of Primary and Secondary Metabolites of Cymbopogan citratus (DC) Leaves
Objective
Methodology
Leaves of Cymbopogan citratus
Washed & air dried under shade
Preparation of extracts
Powdered leaf Fresh leaf
Extracted with chloroform, methanol & acetone using soxhlet apparatus
Ground using distilled water & filtered; used as an aqueous extract
Concentrated using rotary vacuum evaporator & dried
Phytochemical Analysis
Qualitative
Alkaloids (Wagner’s test) Flavonoids (Shinoda & lead acetate test) Phenols & tannins (Lead acetate, FeCl3 & NaOH test) Steroids & sterols (Salkowski’s test) Carbohydrates (Fehling’s & Benedict’s test) Saponins (Honey comb & foam test) Glycosides (Glycoside test) Protein & amino acids (Biuret & Ninhydrin test) Anthraquinones (Borntragers test)
Quantitative analysis
Total phenolics Expressed as tannic acid equivalents
Total flavonoid content Colourimetric method expressed as
rutin equivalent
Carbohydrates Total chlorophyll content Proteins Total lipid content
Total tannins Tannin (%) = Total phenolics (%) – Non tannin phenolics (%)
Primary metabolites Secondary metabolites
Plant constituentExtracts
TestsCholoroform Methanol Aqueous Acetone
Alkaloids - - - - Wagners testFlavonoids - + - + Shimoda
Lead acetate testPhenolics & tannins - + + - Lead acetate test
Ferric chloride testSteroids & sterols + + - + Salkowski testCarbohydrates - + - - Fehlings test
Benedicts testSaponins - - + + Honeycomb test
Foam testGlycosides - + - - Glycoside testProtein & amino acids
+ + + - Biuret testNinhydrin test
Anthraquinone test + + + - Borntragers test
Table 14. Preliminary phytochemical screening of lemongrass leaves
Results and discussion
Table 15. Quantification of primary metabolites of lemongrass leaves
Sl. No. Primary metabolites Weight (mg/g dw)
1 Carbohydrates 150.63 ± 26.83
2 Chlorophyll 2.03 ± 0.02
3 Protein 105.4 ± 2.78
4 Lipids 0.03 ± 0.001
Figure 7. Quantification of secondary metabolites
Conclusion
The results indicates that lemon grass leaves can be used as a source of useful drugs because of presence of various phytochemical components
Pandey et. al., 2014Uttarakhand, India
To screen the phytochemicals present in Cinnamon zeylanicum aqueous bark extract
Phytochemical Screening of Selected Medicinal Plant Cinnamon zeylanicum Bark Extract
Objective
Methodology
Barks of Cinnamon zeylanicum
Washed, shade dried and powdered
Preparation of extracts
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
Maceration technique
Phytochemical Analysis
Qualitative
Alkaloids (Dragendorff’s test) Steroids (Salkowski test) Tannins & polyphenols (Ferric chloride test) Flavonoids (Shinoda test) Saponins (Froth test) Glycosides (Legal’s test) Cardenoloids (Kellar Killani test)
Table 16. Phytochemical screening of secondary metabolites of plant extracts
Phytoconstituents
ExtractsCold water (15ºC)
Hot water (70ºC)
Warm water (45ºC)
Ethanol Methanol Acetone
Carbohydrates + + + - - -
Steroids + + + + + +
Proteins - - - - - -
Glycosides - - - - - -
Alkaloids + + + + + +
Flavonoids + - - + - +
Saponins + + + + + +
Tannins & phenol - - - + + +
Results and discussion
Conclusion
Characterization and isolation of the active chemical components possessed by traditional plants may lead to the development of a potential drug that may treat various kinds of infections and may lead to full utilization by the local community
Singh and Bag, 2014Manipur, India
To identify and compare the bioactive constituents present in Hedychium species and determine total phenolic content
Phytochemical Analysis and Determination of Total Phenolics Content in Water Extracts of Three Species of Hedychium
Objective
Methodology
Hedychium rubrum
Cleaned, shade dried, mechanically grinded & coarsely powdered
Preparation of extracts
Powdered material
Solvent extraction with hexane, acetone, methanol & water
Extracts were concentrated using Rotary Evaporator
Phytochemical screening
Subjected to
Hedychium spicatum
Hedychium coronarium
Phytochemical Analysis
Qualitative Quantitative
Total phenolic content Folin-Ciocalteu reagent expressed as
gallic acid equivalent (mg of gallic acid equivalent / g of sample)
Alkaloids (Hager’s test) Carbohydrates (Fehling’s & Benedict’s
test) Proteins (Xanthoproteic test) Flavonoids (Alkaline reagent test) Saponins (Foam test) Phenolic compounds (Lead acetate test) Tannins (Lead acetate & FeCl3 test) Steroids & terpenoids (Salkowski’s test) Saponins (Froth test) Cardiac glycosides (Keller Killiani test) Oil Phlobatannin
T = (C x V) M
T = TPC (mg/g plant extract)C = concentration of gallic acid (µg/ml)V = volume of extract (ml)M = weight of plant extract (g)
Table 17. Comparative analysis of phytochemical constituents of three different species of Genus Hedychium
Phytochemical constituents
Chemical tests Water extractH. spicatum H. coronarium H. rubrum
Alkaloids Hager’s test - - -Carbohydrates (reducing sugar)
Benedict’s test
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 - - +
Results and discussion
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
Conclusion
Results revealed that the water extracts of three different species of Hedychium contain a good quantity of phenolic compounds
These plants can be studied further to know their biological effects which could be a beneficial in the treatment and controlling of various diseases
Vastrad et. al., 2015Karnataka, India
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
Identification of Bio-active Components in Leaf Extracts of Aloe vera, Ocimum tenuiflorum (Tulasi) and Tinospora cordifolia (Amrutballi)
Objective
Methodology
Tinospora cordifolia
Cleaned, shade dried, mechanically grinded & coarsely powdered
Preparation of extracts
Powdered material
Solvent extraction with hexane, acetone, methanol & water
Extracts were concentrated using Rotary Evaporator
Phytochemical screening
Subjected to
Aloe vera
Ocimum tenuiflorum
Phytochemical Analysis
Qualitative Quantitative
Total phenolic content Folin-Ciocalteu assay method
expressed as gallic acid equivalent (GAE) in milligrams per gram of fresh leaf
Total flavonoid content Aluminium chloride colourimetric
method expressed as mg rutin equivalent (mg RE) per gram of fresh leaf
Tannins & phenolic compounds Flavonoids Alkaloids Saponins Terpenoids
Figure 9. Yield of extracts
Results and discussion
Table 20. Total phenolic content (TPC) of the plant leaf extracts
Extraction solvent
Total phenolic content (GAE* mg/g)
A. vera O. tenuiflorum T. cordifolia
Aqueous 94.42 ± 4.92 80.82 ± 8.63 465.82 ± 23.04
Ethanol 138.13 ± 6.63 113.07 ± 9.81 264.06 ± 18.41
Methanol 95.20 ± 3.23 114.34 ± 11.86 301.42 ± 29.69GAE = Gallic acid equivalent
Fig 10. Total phenolic content (TPC): Calibration curve
Table 21. Total flavonoid content (TFC) of the plant leaf extracts
RE = Rutin equivalent
Figure 11. Total flavonoid content (TFC): Calibration curve
Extraction solvent
Total flavonoid content (RE* mg/g)
A. vera O. tenuiflorum T. cordifolia
Aqueous 72.28 ± 8.70 61.84 ± 7.25 178.43 ± 6.61
Ethanol 76.50 ± 8.57 95.46 ± 4.12 208.36 ± 2.86
Methanol 88.59 ± 8.38 96.34 ± 5.85 132.59 ± 7.59
Conclusion
The results revealed that alkaloids were found to be present in all the extracts of A. vera, O. tenuiflorum and T. cordifolia.
Flavonoids were present in ethanol, methanol and aqueous extracts of A. vera and T. cordifolia
TPC was high in ethanol extract of Aloe vera, methanol extract of O. tenuiflorum and aqueous extract of T. cordifolia
TFC was high in methanol extract of A. vera, methanol and ethanol extracts of O. tenuiflorum and ethanol extract of T. cordifolia