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CHAPTER 2
36
CHAPTER - 2
2. Studies on selected Indian medicinal plants and phytochemistry
2.1 Introduction
It is evident from a review of literature that plants have secondary metabolites
that may have broad-spectrum bioactivity. Indeed plants have been a major source of
drugs for centuries, with more than 25% of the pharmaceuticals currently used are
derived from plants. This may be presumably due to the growing demand for
alternative medicine since plant products are less toxic. In order to harness the plant
species, which are currently over 4 lakhs, it is necessary to adopt a systematic
selection method.
The review of literature in the preceding chapter reveals that a set of plant
families show wide range of medicinal activity, while some families exhibit a narrow
specific activity. Keeping this in mind the following 19 indigenous plants have been
chosen for the present study.
37
Methods of selecting plants for identifying the lead molecule.
Based on
Ethnobotanical history
Folklore use
Chemotaxonomic knowledge
Literature reports
Selection of the plants
38
List of plants chosen and its photo image
Table 2.1
Family Plant Name Part used Vernacular Name
Papilionaceae Crotalaria retusa Linn. (seed) Kilukiluppai
Papilionaceae Stylosanthes fruticosa (Retz.)
Alston
(whole plant) Saillekampa
Rutaceae Aegle marmelos (l.) Corr. Serr. (bark) Vilvam
Rutaceae Toddalia asiatica Linn. (bark) Milakaranai
Meliaceae Toona ciliata Roemer (heart wood) Santhana vembu
Meliaceae Swietenia macrophylla King (wood) Mahagoni
Malvaceae Sida acuta Burm. (whole plant) Arrival mukku
patchilai
Nyctaginaceae Bougainvilla spectabilis Willd (leaves) Kakithappu
Annonacea Polyalthia longifolia
(Sonn.) Thwaites
(leaves) Asokam
Euphorbiaceae Euphorbia thymifolia Burm. (leaves) Chithrapaladai
Moraceae Ficus glomerata Linn. (leaves, bark) Atti
Zingiberaceae Elittaria cardamomum Maton (leaves) Ellagai
Zingiberaceae Zingiber offcinale Roscoe (rhizome) Inchi
Compositae Sphaeranthus indicus Linn. (whole plant) Kottakaranthai
Cucurbitaceae Melothria maderaspatna (L.)
Cogn.
(leaves) Musumusukkai
Sapotaceae Madhuca longifolia (Koening)
(Macbride)
(leaves) Illupai
Piperaceae Piper nigrum Linn. (leaves) Milakhu
Verbanaceae Clerodendrum phlomidis
(burm. &) O.Kuntze
(leaves) Thaluthalai
Solanaceae Datura metel Linn. (leaves) Vellumathai
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C.retusa S.fruticosa A.marmelos
T.asiatica T.ciliata S.macrophylla
S.acuta B.spectabilis P.longifolia
40
E.thymifolia F.glomerata E.cardamomum
Z.officinale S.indicus M.maderaspatana
M.longifolia P.nigrum C.phlomidis
D.metal
41
2.2 Authentication of plant materials:
Pharmacognostically identified individual plants were collected from
Kancheepuram District and Munar Hills Kerala during September 2000 and the plants
were identified by Prof. P. Jeyaraman, Plant Anatomy Research Centre, Tambaram,
Chennai - 45. Toona ciliata Roxb. (Meliaceae) wood was collected from Western
Ghats South India and authenticated by Dr.Venkatasubramanian, Institute of Forest
Genetics and Tree Breading (IFGTB), Coimbatore. Voucher specimens were
deposited in the Asthagiri Herbal Research Foundation (AHRF 001- 019), Chennai –
600 059.
2.3 Literature review
Before proceeding with the extraction and isolation of the active components
it is necessary to explore the literature reports on these plants. The following Table
2.2 indicates the chemical composition and pharmacological activity of each of the
selected plants.
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Table 2.2
Literature review
S.No Plant Name &
Family Chemical Constituents Isolated / pharmacological activity References
1. C.retusa Linn. Papilionaceae
Seed – Pyrolizidine alkaloid, retronecine, Monocrotaline retusine and spectabiline linoleic acid, palmitic acid, oleic acid, recinoleic acid, stearic acid, cyclo propenoids, sterculic acid, malvic acid from seed oil cardenolide
1-6
Antibiotic 7 2. S.fruticosa (Retz.)
Alston. Papilionaceae
Aminoacid, flavanoid and tannins 8-11
3. A.marmelos (L.) Corr.Serr. Rutaceae
Heart wood – marmesin, β-sitosterol, auraptene, umbelliferone, marmin, lupeol, xanthotoxin, 6,7 – dimethoxy courmarin, scopoletin, tempamide, umbelliferone, skimmianine, skimmin, decursinol, halopine, aegelinol, anthraquinone, marminal and 7-α methylmarmin. Bark - β-sitosterol, aurapten, marmin, umbelliferone, lupeol, carbohydrate, lignan glycoside, 7-geranyloxy coumarin, chloromarmin, aeglin and lupeone. Leaves – phlobatannins, flavan 3-ol, leucoanthocyanins, anthocyanins, flavanoid glycoside, lupeol, β-sitosterol, ageline, rutin, marmelosin triterpenes, coumarins, alkaloids, α and β - amyrin, marmin, marmesin, umbeliferone, xanthotoxin, imperatorin γ-fagarin, skimmianine, marmeline and cinnamamide. Gum – Polysaccharide. Fruits – scoparone, scopoletin, umbelliferone, marmesin, skimmin, β-sitosterol, fatty acids, dictamine, xanthotoxol, xanthotoxin, isopimpinellin, isoimperatorin, marmin, cinnamamide, bergapten, marmesin, osthol, auraptin and C-glucosylated propelargonidin.
12-47
44
S.No Plant Name &
Family Chemical Constituents Isolated / pharmacological activity References
3. A.marmelos (L.) Corr.Serr. Rutaceae
Antidiabetic, Hypoglycemic, antihyperglycemic Antifungal Antibacterial Alkaloidal action on dogs heart Antimalarial activity of alcoholic extract Anti-inflammatory activity of root extract Anti ulcer activity of pyranocoumarin and isocoumarin Inhibit crystallization of cholesterol Fungitoxic effects of essential oil Antitumor and antimutagenic activity of polyphenols
48-64
4. T.asiatica Linn. Rutaceae
Norbradylin, 5,6,8-trimethoxy coumarin, toddacoumalone, toddacoumaquinone, toddaculin, isopimpinellin, 6-formyllimetin, toddasin, (+) toddanol, toddalolactone, toddalenol, toddalosin, 5-methoxy suberenon, toddalonone, 8-formyllimettin, benzophenanthridine alkaloid, des-N-methyl chelerythrine, oxychelerythrine, arnottianamide, oxyavicine, avicine, chelerythrine, β-amyrin, 7,8-dimethoxy-2, 3-methylene dioxy benzophenanthridine, toddaculin, toddanone, isoimperatorin, bocconoline, arnottianamide, γ-fagarine, skimmianine, toddaquinoline, arnottin and alloisoimperatorin
65-69
Mitogenic activity Antiplatelet activity of coumarin, alkaloid, benzoquinone, amine (+) - toddanin-isocoreximine Antimicrobial activity HIV inhibitory of quarternary alkaloids and nitidine Antimalarial nitidine Antiplasmodial Anti-inflammatory activity, spasmolytic activity and activity on myocardial ischaemia
70-79
45
S.No Plant Name &
Family Chemical Constituents Isolated / pharmacological activity References
5. T.ciliata Roemer Meliaceae
Cedrelone, quercetin, β-sitosterol, gallic acid, protocatechuic acid, p-hydroxy benzoic acid, chlorogenic acid, caffeic acid, vanillic acid, syringic acid, ferulic acid, sesquiterpene, cycloartenes stigmasterol, campesterol, apotirucallene, tirucallene, catechin, proanthocyanidin, leucoanthocyanidin, toonacilin, 6-acetoxy toonacilin, toonacilid, geranyl geraniol, δ-cadinone, calamenene, α-calacorene, siderin, deoxy cedrelone, 5α, 6β, 8α - trihydroxy -28 norisotoonafolin, 5α, 6β, 8α, 12α - tetra – hydroxy – 28 norisotoonafolin, 1,2-dihydrocedrelone and bergapten
80 –90
Activity on plant pathogens Antimicrobial activity of stem bark
91-93
6. S.macrophylla King Meliaceae
Sweitenolide, swietenine, stigmasterol glycoside, saponin, limonoids, 8,30 epoxy sweitenine acetate, tiglate, tannin, polysaccharide, tetra nor triterpenoids, augustineolide and 3β-6-dihydroxy dihydrocarapin
94-97
Antimalarial 98,997. S.acuta Burm.
Malvaceae Oleic acid, linoleic acid, palmitic acid, stearic acid, arachidic acid, β-sitosterol, sterculic acid, malvic acid, ephedrine, S- (+), Nb-methyl tryptophan methylester, hypaporine, quinazoline alkaloids vascinone, vasicine, vasicinol, choline, betains, ecdysterone, polysaccharides, campesterol, stigmasterol, saponin, flavanoids, gossypol, hypolaetin, gossypetin, kaempferol, quercetin, luteolin, apigenin, chrysoeriol, heraclenol, daucosterol, syringin, α-amyrin, cryptolepine, quinazoline, carboxylated tryptamine alkaloid.
100-118
46
S.No Plant Name &
Family Chemical Constituents Isolated / pharmacological activity References
7. S.acuta Burm. Malvaceae
Acetylcholine like activity Antifertility Antimicrobial activity of cryptolepine, ephedrine, vasicine Antibacterial activity of alkane, alkanol, sterol Hepatoprotective activity Neutralising capacity of Bothrops atrox venom
119-125
8. B.spectabilis Willd Nyctaginaeceae
Spermine, spermidine, putreseine, pinitol, isorhamnetin, quercetin, protein, esculetin 126-130
Hypoglycemic activity of alcoholic extract are pinitol Insecticidal activity Antimicrobial activity of phenolic, non phenolic fraction and quercetin Antiviral activity Hepatoprotective activity
129-134
9. P.longifolia (Sonn.) Thwaites Annonaceae
Altholactone, proanthocyanidin, morphol, aporphine alkaloid, liriodenine, azafluorene alkaloid, darienine polyfothine, isooncodine, polyalthial dioic acid, kolavanic acid, 16α-hydroxy cleroda – 3,13 (14) Z-dien 15,16, -olide, Clerodane diterpene, lanuginosine, oxostephanine, ent-halimane, 3 β, 5 β, 16α - hydroxy halima-13 (14)-en-15, 16-olide, (-)-8-oxopolyalthianine, α-amyrin, β-amyrin, taraxasterol, querctin and its glycoside, (+)-O-methyl bulbocapnine β-N-oxide, (+)-O-methyl bulbocapnine α-N-oxide, (+)- N-methyl nontigenine β-N-oxide, aporphinoid alkaloid, (-)-oliveroline -N-β-oxide, liriodenine ,anonaine, (+) norlirioferine, tetrahydropotoberberine, (-) stepholidine, onychine, darienine and 6,7 – dimethoxychine.
135-149
Antibacterial activity of β-sitosterol, stigmasterol, campasterol, α, β-amyrin Antifeedant, hypoglycemic, antipyretic, anticancer property & clerodane diterpanoids.
150,151
47
S.No Plant Name &
Family Chemical Constituents Isolated / pharmacological activity References
10. E.thymifolia Burm. Phorbol, diterpene acetate, anthocyanin, sterol, protoacousic pigment, flavanoids, hydrolysable tannins and isomallotinic acid.
152-156
Strengthening skull bones. Antimicrobial activity.
157,158
11. F.glomerata Linn. Moraceae
Sterol, gluonol acetate, lupeol, ceryl behanate, α-amyrin acetate, leucocyanidin 3-0-α-L-rhamnopyranoside, guanol acetate, friedelin and bergenin.
159-164
Hypotensive, anthelmintic. Antidysentry. Respiratory disorder, skin diseases. Hypolipidemic activity. Antidiabetic activity. Analgesic, antipyretic and anti-inflammatory. Hepatoprotective activity. Anti tumor, calcium antagonistic. Anti-microbial, antiviral and diuretic activity.
165-174
12. E.cardamomum Maton Zingiberaceae
Hydroxy cinnamic acid, hydroxy benzoic acid, biphenyl and O-phenyl phenol, salicylates, (E) – 4-8-dimethy-l-3, 7-nonatriene and (E, E) – 4, 8,12-trimethyl 1,3,7,11-trideca tetraone, cardiolipin, sterol, 1,8 – cineole, α-terpenyl acetate, limonene, sabiene, α-terpeneol – palmitic acid, oleic acid, α-tocopherol, linoleic, linolenic, α-pinene, linalol, terpenyl acetate, (+) – (S)-nerolidol and aflotoxin.
175-185
48
S.No Plant Name &
Family Chemical Constituents Isolated / pharmacological activity References
E.Cardamomum maton Zingiberaceae
Cholagogue activity with terpineol and terpenyl acetate. Cholinomimetic activity. Penetration enhancing activity. Antimicrobial activity. Anti-inflammatory. Anti-hypertensive. Analgesic activity.
186-192
13. Z.officinale Roscoe Zingiberaceae
Myrcene, limonone, phellandrene, 18-cineole, gingidiol, methylgingediol sequihujene, α-terpinene, α-terpineol, nerol, zingiberene, nerol, geranial, geraniol, paradols, gingerdiols, ar-curcumene, shogaols, 6-gingersulfonic acid, gingerenones, diarylheptanoids, leaf flavanoids kaempferol, quercetin, proanthocyanidin, myricetin, isorhamnetin gingerdione, syringetin, 1-dehydro gingerdione.
193-202
Antioxidant activity of thymol, carvacrol, 6-gingerol, zingerone Cardiotonic activity Anti ulcer activity of 6-ginger sulfonic acid Treatment of chronic liver injury Anti-inflammatory, antibacterial, hypoglycemic activity Antihyperlipidemic Anti-emetic activity of zinger 6-gingerol Antischistosomal activity of gingerol and shagaol Treatment of migraine Gastrointestinal motility enhancing activity Antifungal activity of gingerone A, B, C Inhibition of serotonin induced hyperthermic and diarrhoea
49
S.No Plant Name &
Family Chemical Constituents Isolated / pharmacological activity References
Z.officinale Roscoe Zingiberaceae
Anti-5HT effect of galanolactone. Antiepileptic activity. Learning and memory Performance. Inhibition of prostaglandin synthetase and arachidonate 5-lipoxygenase. Antitumor activity. Antifilarial effect. Treatment of moderate renal failure. Thromboxane synthetase activity. Hepatoprotective activity. Antiamoebic effect. Prophylaxis of typoid fever. Treatment of hepatitis. Antiplatelet effect. Immunomodulatory. Antiviral activity. Vasodialotor effect. Estrogenic activity. Allergic rhinitis. Antidepressant effect. Anti tubercular. Treatment of rheumatoid arthritis.
203-236
14. S.indicus Linn. Compositae
Methylchavicol, α-ionone, d-cadinone, p-methoxy cinnamaldehyde, ocimene, α-terpene, citral, geraniol, geraniol acetate, β-ionone, sphaerene, indicusone, sphaeranthol, ilicic acid, 7-hydroxy frullanolide, eudesmanolido, cyclopeptide alkaloid, sesquiterpenoids, cryptomeridiol, flavone glycoside, 4-epi-cryptomeridiol and isoflavone glycoside.
237-244
Antimicrobial activity of bicyctic sesquiterpene lactone. 245
50
S.No Plant Name &
Family Chemical Constituents Isolated / pharmacological activity References
15. M.maderaspatana (L.) Cogn. Cucurbitaceae
Columbin, fatty acid, linoleic, oleic, palmitic acid, β-pyrazole, γ-glutamoyl β-pyrazole and citrulline
246-247
Motor activity, uterus contraction, anti-inflammatory activity antispasmodic activity. 248 16. M.longifolia
(Koening) (Macbride) Sapotaceae
Saponin bassianin, protobassic acid, genuine sapogenol, sterol, oleanolic acid, palmitate, myricetin, Mi-saponin A, B, C, aminoacid triterpene glycoside madlongiside, carotene, tocopherol, tocotrienol and triglyceride.
249-254
Spasmolytic, anti-ulcerogenic anti-inflammatory activity and analgesic activity. 255,256 17. P.nigrum Linn.
Piperaceae Piperine, N-isobutyl eicosa-trans-2 trans-4-dienamide, 3,4-dihydroxy phenyl ethanol glucoside, 3,4-dihydroxy-6- (N-ethylamine) benzamide, pipericine, piperittine, piperanine, piperamides, pipericide, guineensine, sarmentine, propenylphenols viz., eugenol, myristicine, safrole, monoterpene, and sesquiterpenes, 1,8 cineole, p-cymene carvone and b-bisabolene.
257-261
Antioxidant activity. Antibacterial activity, carcinogenicity. Thyrogenic activity. Hepatoprotective activity. CNS depressant, antipyretic, analgesic and anti-inflammatory. Anti tubercular activity. Treatment of heart disorder. Ovicidal activity. Inhibition of haemolytic activity of snake and scorpion venoms. Anticonvulsant properties and Inhibition of cytochrome p.450-2B by piperine.
262-274
51
S.No Plant Name &
Family Chemical Constituents Isolated / pharmacological activity References
18. C.phlomidis (burm.&) O.Kuntze Verbanaceae
Sterol, ceryl alcohol, palmitic acid, scuttellarin, pectolinaringenin (4’6-dimethyl scutellarein), clerodin, clerosterol, clerodondrin A, 4, 2’, 4’-trihydroxy-6’-methoxy chalcone 4,4’-diglucoside, 7-hydroxy flavanone 7-0-glucoside, 7-hydroxy-flavoneocleronindicin
275-280 Antifungal activity of flavone glycoside, chalcone glucoside, pectolinarigenin, 7-hydroxy flavone, Antidiarrhoeal and antiviral activity.
281-283
19. D.Metel Linn. Solanaceae
Hyoscine, hyoscyamine, scopalamine, fastusine, fastusinine fastudine, nor-hyoscyamine, daturanolone, fastusic acid, pyrolizidine alkaloid, 3α, 6β-ditigloyloxy tropane, 3α, 6β-ditiglolyoxy-tropane - 7β-ol-tigloidine, apohyoscine, 3α-tigloyloxy tropane, norhyoscine, metaloidine, atropine, nor-hyoscyamine, 3α-acetoxy tropane, cuscohygrine, tropine, ψ-tropine, nicotinamide, sterol, daturadiol, 31-norcycloartanol, cyclo-eucalenol, 31-norlanost-8-enol, 31-norlanosterol, abtusifoliol, 4α, 14α, 24-trimethyl cholesta – 8, 24- dienol, 4α-methyl cholesta-8-enol, lophenol, 24-methyl lophenol, 24-ethyl lophenol, gramisterol, citrostadienol, datumetalin, daturilin , withanolide, datumelin, daturametalin A-H, daturilinol, secowithametalin, anisodine, anisodamine, physalindicanol A, daturasterol, withametalin F,G,H tricyclic diterpene, diaturabietatriene, withasteroid, spingosine, calystegines (A3, A5, B1 – B4, C1 and N1) and withafastuosin D,E,F.
284-306
Anticholinergic activity. Increase of brain lipid peroxidase and catalyse activities. Antimicrobial activity. Antistress activity. Monoamino oxidase inhibition activity. Locomotor activity and anti convulsant activity.
307-311
52
2.4 Preparation of plant extracts
In general the plants were subjected to extraction process using the following
scheme 2.1
Extracted with n-hexane
Extracted with EtOH Marc discarded
Typical extraction procedure
The plant material was air dried under shade and powdered. About 100g of the
powder were used for solvent extraction.
Hexane extracts
First it was extracted with hexane by cold maceration process for 24h. The
extract was filtered using Whatmann filter paper No.1, and the filtrate was evaporated
under reduced pressure. The residue was further dried under high vacuum.
Hexane extract Marc
EtOH extract
Plants air dried and powdered
53
Ethanolic extracts
After extraction with hexane the marc obtained was treated with 95% ethanol
until complete extraction of the herb. The extract was filtered using Whatmann filter
paper No.1, and the filtrate was then evaporated under reduced pressure and dried
under high vacuum.
Extracts for pharmacological activities
Air-dried powdered plant material was extracted by maceration with ethanol
for 72h. Then the extracts were concentrated under vacuum and were investigated on
various pharmacological screening procedures.
Percentage yield of various plant extract was given in the following Table 2.3
Table 2.3
Percentage yield of various plant extract
Plant Name Successive extraction Direct ethanolic
extract Hexane extract Ethanolic extract C.retusa 1.8 7.0 - S.fruticosa 1.1 6.0 8.45 A.marmelos 0.6 6.58 - T.asiatica 0.32 2.0 - T.ciliata 0.4 4.6 6.0 S.macrophylla 1.4 7.2 - S.acuta 0.94 7.45 8.5 B.spectabilis 2.01 8.63 10.1 P.longifolia 2.5 10.5 13.73 E.thymifolia 1.3 9.0 - F.glomerata (bark) 0.75 3.5 4.6 F.glomerata (leaves) 1.6 7.8 9.0 E.cardamomum 1.3 7.0 - Z.offcinale 0.4 4.2 - S.indicus 0.9 8.6 -
Contd….
54
Plant Name Successive extraction Direct ethanolic
extract Hexane extract Ethanolic extract M.maderaspatna 1.9 10.2 - M.longifolia 1.5 9.8 - P.nigrum 1.2 11.2 - C.phlomidis 1.9 9.3 - D.metal 1.4 8.8 -
Fractionation
General protocol for fractionation and weights of different fraction in gm/5gm
of the ethanolic extract were given in the following Scheme 2.2 and Table 2.4
Scheme 2.2
Hexane
Hexane: EA, 9:1
Hexane: EA, 8:2
Hexane: EA, 7:3
Hexane: EA, 1:1
Hexane: EA, 3:7
Hexane: EA, 2:8
Ethyl acetate
Methanol
Ethanolic extract of the plant
Fraction 1
Fraction 3
Fraction 5
Fraction 7
Fraction 8
Fraction 9
Fraction 6
Fraction 4
Fraction 2
55
Table 2.4
Weight of each fraction in gm/5gm of the ethanolic extracts used for
fractionation
Plant name Fr.1 Fr.2 Fr.3 Fr.4 Fr.5 Fr.6 Fr.7 Fr.8 Fr.9 Total
S.acuta 0.1 0.12 0.12 0.2 0.2 0.5 0.5 1.0 1.0 3.74
S.fruticosa 0.5 0.3 0.6 0.2 0.11 0.1 0.2 0.6 1.2 3.81
T.ciliata 0.1 0.1 0.8 0.5 0.1 0.1 0.1 0.15 0.15 2.1
B.spectabilis 0.1 0.4 0.3 0.2 0.4 0.2 0.15 0.2 0.8 2.75
F.glomerata (Bark) 0.45 0.3 0.8 0.25 0.1 0.05 0.1 0.05 1.5 3.6
P.longifolia 0.5 0.2 0.2 0.5 0.3 0.2 0.3 0.15 0.75 3.1
F.glomerata (Leaves) 0.2 0.2 0.5 0.3 0.2 0.48 0.57 0.3 1.22 3.97
2.5 HPTLC PROFILE
HPTLC is a sophisticated advanced and automated version of the thin layer
chromatography. Currently it is the analytical technique widely employed for the
analysis of various plants crude extracts and finished products or individual
compounds. The simplicity, high specificity and wide range of sensitivity make
HPTLC an ideal tool for the analysis of herbal products.
Various steps involved in HPTLC
Sample preparation
Application of the sample/standard
Chromatographic development
Detection of spots
Scanning and documentation
56
Salient features of HPTLC
Sophisticated and automated form of TLC
Simultaneous process of sample and standard under similar condition leads to
better comparison. User friendly
Qualitative analysis to identify the number of constituents presents in the
sample solution and affords quantitative measurement.
HPTLC analysis of potent plant extracts
Finger printing of various extracts and then fractions are furnished in Figure
2.1 to 2.14. The number of peaks and the nature of component present in various
plants extract at 2 different wavelengths are given in Table 2.5. Number of Peaks and
Polarity of the components present in each of the fractions obtained from various
plant extracts are given in Table 2.6
57
FINGER PRINTING OF ETHANOLIC EXTRACTS OF VARIOUS PLANTS
Sample:
10 mg of the ethanolic extract of each plant in 1 ml of CHCl3.
Standard (optional):
5 mg of β-sitosterol in 1 ml of CHCl3.
Chromatographic conditions:
Stationary phase: HPTLC plates 10 x 10 cm Silica gel 60 F254 (Merck) Mobile Phase: Hexane: ethyl acetate (7:3)
Sample application:
With Linomat IV or Automatic TLC Sampler III: 5μl of test solution and standard as 7 mm bands, space 5 mm, 10 mm from lower edge.
Development: 10 x 10 cm Twin Trough Chamber, saturated for 10 minutes (filter paper), 5 ml developing solvent per trough, developing distance 80mm. Dry plates with cold air for 5 minutes.
Detection:
Scanner:
a) UV 254 nm b) UV 366 nm CAMAG TLC Scanner 3
Analysis of Samples
1. Sida acuta (whole plant) (authenticated Indian source)
2. Stylosanthes fruticosa (whole plant) (authenticated Indian source)
3. Toona ciliata (heart wood) (authenticated Indian source)
4. Bougainvilla spectabilis (leaves) (authenticated Indian source)
5. Ficus glomerata (bark) (authenticated Indian source)
6. Polyalthia longifolia (leaves) (authenticated Indian source)
7. Ficus glomerata (leaves) (authenticated Indian source)
Imaging Software: CAMAG VideoStore 2
58
FINGER PRINTING OF VARIOUS PLANT FRACTIONS
Sample:
10 mg of each fraction of the plant extracts in 1 ml of CHCl3.
Chromatographic conditions:
Stationary phase: HPTLC plates 10 x 10 cm Silica gel 60 F254 (Merck)
Mobile Phase: Hexane: ethyl acetate (8:2, 7:3 & 1:1) CHCl3: MeOH (9:1) as mentioned in the figure 2.9 to 2.14
Sample application:
With Linomat IV or Automatic TLC Sampler: 5μl of test solution as 7 mm bands, space 5 mm, 10 mm from lower edge.
Development: 10 x 10 cm Twin Trough Chamber, saturated for 10 minutes (filter paper), 5 ml developing solvent per trough, developing distance 80mm. Dry plate with cold air for 5 minutes.
Detection:
Scanner:
a) UV 254 nm b) UV 270 nm CAMAG TLC Scanner II
Analysis of Samples
1. Sida acuta (whole plant) (authenticated Indian source)
2. Stylosanthes fruticosa (whole plant) (authenticated Indian source)
3. Bougainvilla spectabilis (leaves) (authenticated Indian source)
4. Ficus glomerata (bark) (authenticated Indian source)
5. Polyalthia longifolia (leaves) (authenticated Indian source)
6. Ficus glomerata (leaves) (authenticated Indian source)
Imaging Software: CAMAG software (c) 1998
59
60
61
Fig. 2.1. S.acuta
Fig. 2.2. S.fruticosa
62
63
Fig. 2.5 F.glomerata (bark)
Fig. 2.6 P.longifolia
64
Fig. 2.7 F.glomerata (leaves)
Fig. 2.8 β-sitosterol
65
Fig. 2.9 S.acuta
TLC Solvent System Hexane: EtOAc (8:2)
66
S.acuta
TLC Solvent System CHCl3: MeOH (9:1)
67
Fig. 2.10 S.fruticosa
68
S.fruticosa
69
70
B.spectabilis
71
Fig. 2.12. F.glomerata (bark)
72
73
Fig. 2.13 P.longifolia
74
75
Fig. 2.14. F.glomerata (leaves)
76
77
Table 2.5
Plant Name
Number of spots
At 254 nm At 366 nm
LP MP HP LP MP HP
S.acuta 2 2 - 3 1 1
S.fruticosa 5 - - 3 1 1
T.ciliata 4 - - 2 1 2
B.spectabilis 3 1 - 4 1 1
F.glomerata (bark) 3 1 - 3 - 1
P.longifolia 3 - - 3 - 1
F.glomerata (leaves) 5 - - 3 1 1
β-sitosterol 2 - - 2 - - *LP-Low polar (Rf 0.61-0.99), MP-Medium polar Rf 0.31-0.60), HP-high
polar (Rf 0.01-0.30)
78
2.6 Phytochemistry
2.6.1 Isolation of chemical constituents from the whole plant Stylosanthes
fruticosa
Scheme 2.3
Ethanol extraction 30g of extract VLC on silica gel, 70-325 mesh 15% Ethyl acetate in hexane
Compound (I-1) Compound (I-2) β-sitosterol (yield 2g) linoleic acid (yield 0.35g) M.p.139ºC
Marc EtOH extract (Residue 84.5g)
Fraction 3(6.7g)
Whole plant of S.fruticosa dried and powdered (1Kg)
79
Table 2.6
Number of peaks and polarity of the components present in each of the fractions obtained from various plant extracts
Fraction PlantName
Fraction 2 Fraction 3 Fraction 4 Fraction 5 Fraction 6 Fraction 7 Fraction 8 Fraction 9
LP MP HP LP MP HP LP MP HP LP MP HP LP MP HP LP MP HP LP MP HP LP MP HP S.acuta 1 4 3 1 3 3 - 1 3 - 2 2 - - 5 - 1 2 2 1 4 4 1 5 S.fruticosa 2 2 1 1 4 2 1 1 1 1 - 2 4 1 3 - - 3 - - - - 2 4 B.spectabilis 2 1 4 1 2 2 5 3 3 2 2 3 3 2 3 2 1 4 2 2 5 - 2 7 F.glomerata (bark) 4 5 4 5 3 5 2 3 5 - - - 3 6 4 1 - 2 - - 3 3 2 3
P.longifolia 4 3 4 2 3 5 2 2 4 - - - 1 3 2 3 1 3 2 2 4 1 1 5 F.glomerata (leaves) 2 4 1 3 3 4 1 4 3 2 5 3 3 4 2 2 3 5 1 1 6 - 2 3
LP-Low polar components (Rf 0.61-1.00), MP-Medium polar components (Rf 0.31-.60), HP-High polar components (0.01-0.30)
80
The S.fruticosa whole plant was dried, grounded (1kg) and extracted with
EtOH (1x 5 litres). About 84.5g of a residue were obtained after evaporation of the
ethanolic extract. From this 30g was impregnated on silica gel (60-120 mesh, 50g)
and subjected to vacuum liquid chromatography on a silica gel 70-325, 300g, using
solvents of increasing polarity as per the general protocol given above. The fraction
(Fraction 3) obtained with hexane: EtOAc (80:20) yielded a solid material on
concentration, which showed two spots in TLC at Rf 0.34 & 0.46 [hexane: EtOAc
(7:3)].
When all fractions eluted in this solvent system (20% EtOAc in hexane) were
concentrated under vacuum yielded a residue 6.7g. The residue on further column
chromatography (silica gel 70-325 mesh, 150g) with 15%EtOAc in hexane yielded a
white solid which was purified by recrystallisation with MeOH to yield a pure
compound (I-1) (2g). The compound was identified as β-sitosterol, on comparison
with authentic sample in TLC (hexane: EtOAc, 7:3 system) and spectral data.
Characterization:
HO
bb
db
cc1
bc
ca b
ba
c b
bc
ca b
b
c
b a
ca
a
HO
21
45 6
89
103
7
1112
13
14
17
18
19
20
21 2223
26
27
2829
15
1624
25
(I-1) β-sitosterol
Mol.formula C29 H50O Mol.wt: 414 M.p.139ºC
81
82
83
1H NMR (200 MHz, CDCl3)δ, ppm: a 6 CH3, b 11CH2, c 7CH,50H m,(0.64-2.04):, d
1H,(3.49-3.55), c1 1H s, (5.14).
13C NMR (200 MHz, CDCl3) δ, ppm: 37.2 (C-1), 31.5 (C-2), 71.2 (C-3), 42.2 (C-4),
140.7 (C-5), 121.6 (C-6), 31.8 (C-7), 33.8 (C-8), 50.0 (C-9), 36.1 (C-10), 21.0 (C-11),
39.7 (C-12), 45.7 (C-13), 56.7 (C-14, C-O), 24.2 (C-15), 28.3 (C-16), 56.0 (C-17),
11.9 (C-18), 19.7 (C-19), 36.4 (C-20), 19.3 (C-21), 33.8 (C-22), 26.0 (C-23), 48.2 (C-
24), 29.0 (C-25), 18.9 (C-26), 18.7 (C-27), 23.0 (C-28), 12.2 (C-29).
IR (KBr): 3456, 2919, 2850, 2443, 1789, 1636, 1384, 835, 725 cm-1
Subsequent elution with 15% EtOAc in hexane yielded another white solid,
which on purification by recrystallization with MeOH gave compound (I-2) (0.35g),
which was characterized by 1H NMR and 13C NMR as linoleic acid.
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
COOH1
(I-2) Linoleic acid
Mol.formula : C18H32O2 Mol.wt: 280
The spectral data of linoleic acid were given in Fig.2.16, 2.16a
M.F.C18H32O2 Mol.wt: 280 1H NMR (200 MHz, CDCl3) δ, ppm: 0.87 (1, 3H, CH3), 1.25 (m, 8H, (CH2)4, 1.62 (m,
14H, (CH2)7), 2.34 (t, 2H, (CH2)), 5.36 (q, 4H, (CH=CH2)2) and 9.03 ( q, H,COOH).
13C NMR (200 MHz, CDCl3) δ, ppm: 180.6 (C-1), 34.1 (C-2), 22.6 (C-3), 29.2 (C-4),
25.5 (C-5), 24.6 (C-6), 27.1 (C-7), 29.0 (C-8), 127.8 (C-9), 130.1 (C-10), 29.6 (C-11),
129.9 (C-12), 128.4 (C-13), 31.9 (C-14, C-O), 29.4 (C-15), 29.3 (C-16), 22.5 (C-17),
14.0 (C-18).
84
85
86
IR (KBr): 3456, 2919, 2850, 2443, 1789, 1636, 1460, 835, 725 cm-1
This was the first report on the isolation of these two compounds from S.fruticosa.
2.6.2 Isolation of constituents from the Ficus glomerata (leaves)
Scheme 2.4
Ethanol extraction
VLC on silica gel, 70-325 meshe
20% Ethyl acetate in hexane
Compound (II-1) β-sitosterol (yield 1.2g). M.p: 139oC
Leaves of F.glomerata were dried and grounded (1kg) and then extracted with
EtOH (1X5lit). The residue obtained (90.0g) after evaporations of the ethanolic
extract. From this (30g) was impregnated on silica gel (60-120 mesh, 50g) and was
subjected to vacuum liquid chromatography (silica gel 70-325 mesh, 300g), using
solvents of increasing polarity as per the general protocol given above. The fractions
(Fraction 4) obtained with Hexane: EtOAc (7:3) yielded a solid material on
concentration (Rf 0.36 in Hexane and EtOAc 7:3). Yield: 1.2g.
Marc EtOH extract (Residue 90.0g)
Fraction 4(2.0g)
Ficus glomerata (leaves) dried and powdered (1.0 Kg)
87
The residue on further separation by column chromatography (silica gel 70-325
mesh, 50g) with 20%EtOAc in hexane yielded a white solid which was purified by
recrystallization with cold MeOH to yield a pure compound (II-I)(1.2g). The
compound was identified as β-sitosterol in TLC (hexane and EtOAc 7:3 system). This
compound was already reported from this plant.
Characterization:
HO
bb
db
cc1
bc
ca b
ba
c b
bc
ca b
b
c
b a
ca
a
(II-1) β-sitosterol
Mol.formula: C29H50O Mol.wt: 414 M.p.1390C
The spectral details are given below
1H NMR (200 MHz, CDCl3) δ, ppm: a 6 CH3, b 11CH2, c 7CH, 50H m, (0.67-2.04):
d 1H, (3.44-3.63), c1 1H s, (5.14).
13C NMR (200 MHz, CDCl3) δ, ppm: 37.2 (C-1), 31.5 (C-2), 71.7 (C-3), 42.2 (C-4),
140.7 (C-5), 121.7 (C-6), 31.8 (C-7), 31.8 (C-8), 50.1 (C-9), 36.1 (C-10), 21.0 (C-11),
39.7 (C-12), 42.3 (C-13), 56.7 (C-14, C-O), 23.0 (C-15), 29.1 (C-16), 56.0 (C-17),
11.9 (C-18), 19.3 (C-19), 36.4 (C-20), 18.7 (C-21), 29.6 (C-22), 23.0 (C-23), 45.8 (C-
24), 36.4 (C-25), 18.7 (C-26), 19.0 (C-27), 19.3 (C-28), 11.8 (C-29).
88
2.6.3 Isolation of active molecule from the T.ciliata (heart wood)
Scheme 2.5
Hexane extraction
Purified by CC
Compound (III-1) Cedrelone (yield 3.0g). Mol.formula: C26H30O5 Mol.wt: 422 M.p.213°C
The isolation of natural product, cedrelone was first reported by Parihar and
Dutt (1950) 312, who obtained it, along with an essential oil, by benzene extraction of
the heart wood of Toona ciliata Roxb. Syn. Cedrela toona, these workers assigned the
formula C26H30O5 on the basis of color tests indicates the presence of phenolic
hydroxyl, ketonic carbonyl group, an ethylenic double bond, and an unsaturated
lactone ring.
Later other Indian workers313 have also isolated cedrelone-using heartwood in
pure form and the molecular formula was assigned as above. The spectral properties
were in good agreement with the functional group present.
Dried and powdered heartwood of Cedrela toona was extracted with hexane;
the extract was concentrated under vacuum. Crude cedrelone was about 0.4% of the
Marc Hexane extract (Residue 4.0g)
T.ciliata (heart wood) dried and powdered (1.0 Kg)
89
weight of dried heartwood. The extract was column chromatographed over silica gel
to get pure cedrelone. M.p. 213ºC.
Characterization:
O HOO
O
O23
45 7
89
1 91 1
1 2
1 31 5
1 6
1 8 2 1
2 2 2 3
1
1 0 1 4
6
2 0
1 7
(III-1) Cedrelone
The structure of the compound (III-1) was established on the basis of the
following spectral evidences. IR spectrum showed specific absorptions at 3396 cm-1
(hydroxyl) 1675 cm-1 (α,β unsaturated carbonyl) 1621 cm-1, 1606 cm-1 (-C=C-), 2982
(methyl), 872 cm-1, 1463 cm-1, 3150 cm-1 (furan moiety) 1365 cm-1, 1382 cm-1 (C-O
stretching). The 1H and 13C NMR spectrum (Fig.2.17, 2.17a).
1H NMR (200 MHz, CDCl3) δ, ppm: 6.93 (d, 1H, J=10Hz , C-1), 6.13 (d,1H,
J=10Hz,C-2), 2.94 (m, 1H, C-9), 2.20 (m, 2H, C-11), 2.15-2.66 (m, 2H, C-12), 4.0 (s,
1H, C-15), 2.64-2.80 (m, 2H, C-16), 3.79(m, 1H, C-17), 7.36 (s, 1H, C-21), 6.46 (d,
1H, C-22), 7.33 (s, 1H, C-23), 1.53 (s, 3H, CH3), 1.43 (s, 3H, CH3), 1.29 (s, 3H, CH3),
1.1 (s, 3H, CH3), 0.9 (s, 3H, CH3).
13C NMR (200 MHz, CDCl3) δ, ppm: 152.3 (C-1), 127.2 (C-2), 197.9 (C-3), 48.7 (C-
4), 141.1 (C-5), 133.8 (C-6), 203.6 (C-7), 46.7 (C-8), 41.7 (C-9), 41.9 (C-10), 20.19
(C-11), 35.13 (C-12), 40.2 (C-13), 69.7 (C-14, C-O), 55.0 (C-15), 35.1 (C-16), 43.1
(C-17), 123.1 (C-20), 139.3 (C-21), 110.6 (C-22), 142.9 (C-23), 26.7, 23.8, 23.0, 21.1,
19.4 (5CH3)
90
91
92
2.6.2 Isolation of chemical constituents from the P.longifolia (leaves) Scheme 2.6
Ethanol extraction
VLC on silica gel, 70-325 mesh
20% ethyl acetate in hexane
Compound (IV-1)
β-sitosterol (yield 1.0g). M.p: 139oC
Isolation Procedure
The leaves of P.longifolia were dried, grounded (1.7kg) and then extracted with
EtOH (1x10lit). The residue obtained (137.3g) after evaporation of the ethanolic
extract, from this 30g was taken, admixture with silicagel (60-120 mesh, 30g) and was
subjected to vacuum liquid chromatography (70-325 mesh, 300g), using solvents of
increasing polarity, starting from n-hexane (4x250ml), then combination of n-hexane
and (EtOAc 10%, 20%, 30%, 50%, 70%, 80%) each (4x250ml) and then 100%
EtOAc followed by MeOH 100%. Fractions obtained with n-hexane and EtOAc 30%
yielded a residue (3g). This residue, on further column chromatography (70-325
mesh, 80g) with n-hexane: EtOAc (80:20) yielded a white amorphous solid, which
Marc EtOH extract (Residue 137.3g)
Fraction 4(2.0g)
P.longifolia (leaves) dried and powdered (1.0 Kg)
93
was purified by recrystallisation with MeOH to yield a pure compound (1.0g). The
compound was identical with β-sitosterol and it was confirmed by 1H and 13C NMR
P.longifolia
1H NMR (200 MHz, CDCl3)δ, ppm: a 6 CH3, b 11CH2, c 7CH,50H m,(0.69-1.72):, d
1H,(3.46-3.56), c1 1H s,(5.12).
13C NMR (200 MHz, CDCl3) δ, ppm: 37.2 (C-1), 31.6 (C-2), 71.8 (C-3), 42.2 (C-4),
140.7 (C-5), 121.7 (C-6), 31.9 (C-7), 29.6 (C-8), 50.1 (C-9), 36.5 (C-10), 21.0 (C-11),
39.7 (C-12), 45.8 (C-13), 56.0 (C-14, C-O), 23.0 (C-15), 29.1 (C-16), 56.7 (C-17),
11.9 (C-18), 19.3 (C-19), 36.1 (C-20), 18.7 (C-21), 28.2 (C-22), 24.2 (C-23), 45.8 (C-
24), 33.9 (C-25), 18.7 (C-26), 19.0 (C-27), 19.8 (C-28), 11.8 (C-29).
94
2.7 References
1. Mattocks, A.R. Nature, 191, 1281-2 (1961).
2. Puri, S.C., Sawhney, R.S. and Atal, C.K. Indian J.Pharm., 34, 123-4 (1972).
3. Han, Gui-Qiu; Xu, Shi ming; Liu, Fu-Qin; Wang, and Xu, Pei-Ching I Hsuch Yuan Hsuch Pao, 13,15-18 (1981).
4. Pilbeam, D.J., Lyon-Joyce, A.J., Bell and E.A. J.Nat.Prod., 46, 601-5 (1983).
5. Daulatabad, ChiragMahmoodJehan D., Hosamani, Kallapa, M., Mulla, Gouse Mohaddin, M. J.sci.Food agri., 47, 253-5 (1989).
6. Yadava, R.N., Thakur and Vibha, Fitoterapia, 66,147-8 (1995).
7. Bulhocs, Genisa de castro coitinho., Da meta-e-silva, and Antonio., Pernambuco, 15,45-50(Port).(1977).
8. Casaotto, Meris,Palazzo, Aura, L., Agostini, Getulio, Benshimol and Abraham Levy, Acta Cient.Venez., 35, 248-252 (1984).
9. Ali, M.and Qadry, J.S. J.Indian Chem. Soc., 64, 230-1 (1987).
10. Ogbeide, O.N.and Parvez, M. J.Liq. Chrom., 15, (17): 2989-96 (1992).
11. Mwendia and CharlesWaweru, Diss. Abst. Int ., B, 56, 3518 (1995).
12. Asima Chatterjee and Roy, S.K. J.Indian Chem.Soc., 36, 267-9 (1959).
13. Bhattacharyya, P., Jash, S.S., Dey, A.K.J.Indian Chem.Soc., 66, 424-5 (1989).
14. Chatterjee, A. and Chaudhury, B. J.Indian Chem.Soc., 37, 334-6 (1960).
15. Khaleque, A., Ismail, K., Mohamed, Rahman, A., Kim.Mojibur, Amin, M., Sadrul, Fritz, H. and Besch, E. Bangladesh J.Sci.Ind.Res., 13, 252-5 (1978).
16. Shoeb, Aboo, Kapil, Randhirs, and Popli Satya, P. Phytochemistry, 12, 2071-2 (1973).
17. Basu, D., Sen, R. Phytochemistry, 13, 2329-30 (1974).
18. Banerji, J., Das, A.K., Ghoshal, N. and Das, B. Indian J.Chem., 27B, 594-6 (1988) .
19. Manandhar, Mangala, D., Shoeb, Aboo, Kapil, Randhirs., Popli and Satya, P. Phytochem., 17,1814-15 (1978).
20. Tiwari, S., Bansal, S. Goobios (Jodbpur, India ), 22, 59-61 (1995).
21. Srivastava, S.D., Srivastava, S., Srivastava, S.K. Fitoterapia, 67,83-4 (1996).
95
22. Yang, Xiuwei, Masao, Hattori, Tsuneo, Namba, Chin, J.Pharm.Sci., 5, 132-140 (1996).
23. Tetrahedron Lett., 71(1967).
24. Patra.A., Mukhopathyay, A.K. and Ghosh, A. Indian J.Chem.,Sec.B, 17B, 385- 7 (1979).
25. Basak, Ratan, K., Mukherjee and Amal, K. Carbohydrate.Res., 110, 145-51 (1982).
26. Arun, K., Srivastava, Santosh, K., Srivastava, Savitri, D. J.Indian Chem.Soc., 68, 452-4 (1991).
27. Nema, Durga, Srivastava, Santosh, K. Pract.Nalt.Acad.Sci.India, Sec.A, 61, 465-8 (1991).
28. Ohashi, Kazuyoshi, Watanache, Hisashi, Okumura,Yasuaki, Uji, Tahan, Kitagawa, Isao , Chem.Phar.Bull., 42, 1924-6 (1994).
29. Ohashi, Kazuhoshi, Watanabe, Hishashi, Ohi, Katsuhide, Arimoto, Hirokazu, Okumura, Yasua, K. Chem.Lett., 10, 881-2 (1995).
30. Riyanto, Sugeng, Sukari etal., Maj.Farm.Indones., 11, 17-24 (2000).
31. Bajaj, K.L., Sharma, A.K., Bhatia, I.S. J.Inst.Chem., (Calcutta ) 47, 79-81 (1974).
32. Sharma, B.R., Rattan, R.K. and Sharma, Perveen, Indian J.Chem.Sec.B., 19B, 162 (1980).
33. Siddappa, G.S. Food Sci.(Mysore), 7, 180 (1958).
34. Chatterjee, Majumder, A. Indian J.Chem., 9, 763-6 (1971).
35. Govindachari, T.R., Premila and Manakkel, S. Phytochem., 22, 755-7 (1983).
36. Govindachari, T.R. and Premila, S.M. Phytochemistry, 22, 755 (1983).
37. Roy, Arati, Mukherjee, Amal, K., Rao and Chintala Charuva V.N.Carbohydrate Res., 54, 115-24 (1977).
38. Roy, Pratima, Majumdar, B.C. Sci.Cult., 55, 110-11 (1989).
39. Roy, Susanta, K. and Singh, R.N. Indian food Packer, 32, 3-8 (1978).
40. Banerji, N. and Kumar, R.J. Inst.Chem (India), 52, 59-60(1980).
41. Pal, P.K., Bhattacharya-Dk., Ghosh-S. J. Oil Technologists Association of India, 25, 55-59 (1993).
96
42. Banerji, N. and Maiti, M., J.Indian Chem.Soc., 57, 945-6 (1980).
43. Mandal, Prabir, K., Mukherjee and Amal, K. Carbohydrate Res., 90, 233-41 (1981).
44. Tokitomo, Yukiko, Shimono, Yuko, Kabayashi, Akio, Yamanish and Tei Agri- Biol.Chem., 46, 1873-7 (1982).
45. Ghosh, A.K., Nagar, P.K. and Sircar, P.K. Physiol.Plant, 59, 165-70 (1983).
46. Reisch, J., Hussain, R.A. and Adesin, S.K. Pharmazie, 40, 503-4 (1985).
47. Abeysekera, A.M., Desilva, K.T.D., Samarasinghe, S., Seneviratne, P.A.K., Von DenBerg, A.J.J. Fitoterapia, 67, 367-370 (1996).
48. Alam, M.M, Siddiqui-MB, and Husain-W. Fitoterapia, 61, 240-242 (1990).
49. Rao, V.V., Dweid, S.K., Swarup, D.and Sharma, S.R. Current science, 69, 932-933 (1995).
50. Sharma, S.R., Dwivedi, S.K.and Swarup, D. Indian Journal of Animal Sciences, 66, 1017-1020 (1996).
51. Sharma, S.R., Dwivedi, S.K., Varshney, V.P.and Swarup, Phytotherapy Research, 10, 426-428 (1996).
52. Singh, S.P. Ethnobiology in human welfare: abstracts of the fourth international congresss of ethnobiology, Lucknow, Uttar Pradesh, India, 17-21 November, 248 (1994).
53. Jain, N.K. Indian Drugs Pharm.Ind., 12, 55 (1977).
54. Sasidharan, V.K., Krishnakumar, T.and Manjula, C.B. Philippine Journal of Science, 127, 65-72 (1998).
55. Haravey, S.K. Indian J.med.Res., 56, 327-31 (1968).
56 Misra-P., Pal-N.L., Guru, P.Y, Katiyar-J.C. and Tandon, J.S, International J. of Pharmacognosy, 29, 19-23 (1991).
57. Udupa, S.L., Udupa, A,L.and Kulkarni, D.R. Fitoterapia, 65, 119-123 (1994).
58. Goel, R.K., Maiti, R.N., Manickam, M.and Ray, A.B. Indian Journal of Experimental Biology, 35, 1080-1083 (1997).
59. Venkattaranganna, M.V., Gopu madhavan, S., Sundaram, R.and Mitra, S.K. J. Ethnopharmacology, 63, 187-192 (1998).
60. Saraswathi, N.T.and Gnanam, F.D. J.Cryst.Growth, 179, 611-617 (1997).
97
61. Singh, G., Srivstava, P., Mallavrapu, G.R., Ramesh, S. and Rao G.P. Perfumerie und kosmetik, 74, 686, 714, 716, 719-720 (1993).
62. Annapurani, S.and Priya-R. Indian Journal of Nutrition and Dietetics, 36, 431-435 (1999).
63. Singh, R.P., Banerjee, S.and Rao, A.R. Indian journal of Pharmacy and Pharmacology, 52, 991-1000 (2000).
64. Muruganandan, S., Srinivasan, K., Tandon, S.K., Jawahar Lal, Sureshchandra, Raviprakash, V., Lal, J., Chandra, S., Sushilkumar, Hasan,S.A., Samresh Dwivedi, Kukreja, A.K.M., Ashok Sharma, Singh, A.K., Srikant Sharma and Rakesh Tewari, Journal of Medicinal and Aromatic Plant Sciences, 22-23, 56-58 (2001).
65. Desmukh, M.N., Despande, V.H.and Rao, A.V.Rama, Phytochemistry, 15, 1419-20 (1967).
66. Ishhii, H., Ishikawa, T., Kobayashi, J.I.and Seki, H. Tennen Yuki Kogobutsu Toronkai Koen Yoshishu, 34, 518-525 (1992).
67. Ishii, H., Kobayashi, J., Ishikawa, M., Haginiwa, J.and Ishikawa, T. Journal of the Pharmaceutical Society of Japan, 111, 365-375 (1991).
68. Tsai, IanLih, Fang-SongChwan, Ishikawa, T., ChangChinTeng, Chen-IhSheng, Tsai, I.L., Fang, S.C., Chang, C.T.and Chen, I.S. Phytochemistry, 44,1383-1386, (1997).
69. Tsai, Ian-Lih, Chang, Ron- Girr, Fang, Song-Chwan, Ishikawa, T., Isutoma Chen, and Ih-Sheng, Chin.Pharm.J., (Taipei), 48, 63-75 (1996).
70. Tachibana, Y., Kato, A., Nishiyama, Y., Ikemi, M., Ohoka, K., Kawanishi, K., Juma, F.D., Nganga, J.and Mathenge, S.G. Phytomedicine, 2, 335-339 (1996).
71. Isai, Ian-Lih, Wen, Ming-Feng, Teng, Chi-Ming, Ishikawa, Tsutomu, Chen and Ih-Shen, Phytochemistry, 48, 1377-1382 (1998).
72. Bandara, B.M.R., Hewage, C.M., Jayamanne, D.H.L.W., Karunaratne, V., Adikaram, N.K.B., Bandara, K.A.N.P., Pinto, M.R.M. and Wijesundra, D.S.A. Journal of the Natural science Council of Srilanka, 18, 71-77 (1990).
73. Chakraborty, D.P., Dasgupta and Bose, P.K. Ann.Biochem.and Expt.Med., (Calcutta), 17, 59-62 (1957).
74. Mohammad, A., Gustafson, Kirk, R., Kashman, Yoel, Cardellma, John, H., Mcmahan, James, B., Boyd and Michael, R. Nat.Prod.Lett., 6, 153-156 (1995).
75. Gakunju, D.M.N., Mberu, E.K., Dossaji, S.F., Gray, A.I., Waigh, R.D., Waterman, P.G.and Watkins, W.M. Antimicrobial Agents and Chemotherapy, 39, 2606-2609 (1995).
98
76. Oketch, Rabah, H.A., Mwangi, J.W., Lisgarten, J.and Mberu, E.K.Fitoterapia, 71, 636-640 (2000).
77. Kavimani, S., Vetrichelvan, Ilango, R. and Jeykar, B.Indian Journal of Pharmaceutical Sci., 58, 67-70 (1996).
78. Lakshmi, V., Kapil, R.S.and Patnaik, G.K.Indian Drugs, 26, 706-707 (1989)
79. Ren Xianda, Ite and Xiaoping, Jinan Daxue Xuebao, Ziran Hexue Yu Yixueban, 19, 22-25 (1998).
80. Hodges, R., McGeachin, S.G.and Raphael.R.A.Journal of Chemical Society, 2515-2526 (1963).
81. Kraus, W., Bauman, S., Bokel, M., Cramer, R., Grimminger, W., Handlmeir, R., Keil, E., Keller, A., Klingele, M., Pohnl, H.and Schwinger, M. In “Proceedings of the 1st PrincessChulabhorn Sciences Congress,” 1st Congress on Natural Products Bangkok, Vol.II, p. 554 (1987).
82. Singh, D.P.and Plant, A.K. Fitoterpia, 66, 192 (1995).
83. Da Silva, M., FatimaDas,G.F., Agastinho, Sveli , M.M.,De Paula, Jose, R., Nato, Jao Oiano, Castro-Gamboa, Lan, Filho, EdsonRodrigues, Fernandes, Joao, Vieira, and Paulo, C.Pure.Appl.chem., 71, 1083-1087 (1999).
84. Bhatia, V.K., Madhav, R., Seshadri, and Tiruvenkada, R.Indian J.Chem., 7, 121-2 (1969) .
85. Kraus, Wolfgang, Grimminger, Wolf, Sawitzki, Gisela, Angew Chem., 90, 476-7 (1978).
86. Vlhov, R., Holub, M.and Herout, V.Coll.Czech.Chem.Comm., 32, 822 (1967).
87. Nagasambagi, B.A., Sriraman, M.C., Yankov, L.and SukhDev, Phytochemistry, 14, 673 (1975).
88. Banerji, R.and Mitra, C.R. Planta Medica, 28, 52-55 (1975).
89. Chatterjee, A., Chakrabortty, T.and Chandrasekharan, S. Phyochemistry, 10, 2533-5 (1971).
90. Maia, Beatriaz, H.L.N.S.and dePaula et.al., J.Braz.Chem.Soc., 11, 629-639 (2000).
91. Damayanti, M., Susheela, K.and Sharma, G.J.Cytobios, 86, 155-165 (1996).
92. Govindachari, T.R., Suresh, G., Gopalakrishnan, G., Masilamani, S.and Banumathi, B. Fitoterapia, 71, 317-320 (2000).
93. Rasheduzzaman Chowdhury, Choudhury M.Hasan and Mohammad,A. Rashid, Fitoterapia, 74,155-158 (2003).
99
94. Anjaneyulu, A.S.R., Murthy, Y.L.N., Row and Ramachandra, L. Curr.Sci., 46, 141-2 (1977).
95. Cespedas, Roberto, Chacon, S., Mora, A. and Luis, Ing.Cienc quim., 14, 6-10 (1992).
96. Leonde Pinto, Gladys, Gonzalez deTroconis, Nola, Martinez, Maritza, Clamens, Carmen, Vera, Antonio, Rivas, Carlos, Ocando, and Edger , Ciencia (maracaiba) , 4, 47-52(1996).
97. Kojima, Keisuke, Isaka, Kimio, Ogihara, and Yukir, Chem.Pharm.Bull., 46, 523-525 (1998).
98. Mucloz, V., Sauvain, M., Bourdy, G., Callapa, J., Rojas, I., Vargas, l., Tae, A.and Deharo, J.Ethnopharmacol., 69, 139-55 (2000).
99. Soediro, Iwang, Padmawinata, Kosasih, Wattimena, Johana Rosina, Sekita, and Setsuko, Acta Pharm. Indones., 15, 1-13 (1990).
100. Pande, C.S. and Tewari, J.D. J.Oil.Technol.Associ.India, 16, 25-8 (1960).
101. Khan, M.W., Rashid, M.A., Huq, E., Ahmad and Mesbah, J.Bangladesh Acad . Sci., 13, 55-60 (1989).
102. Ahmad, Moghos, U., Husain, S.K., Ahmad, M., Osman, S.M.and Subbarao, R. J.Am.Oil Chem.Soc., 53, 698-9 (1976).
103. Rao, sundar, K.and Lakshminarayana, G. J.Am.Oil Chem.Soc., 61, 1345-6 (1984).
104. Tapan Dutta, Bull.Reg.Res.Lab. (Jammu, India), 1, 178-182 (1963).
105. Prakash Anand, Varma, R.K.and Ghosal, S. Planta Med., 43, 384-8 (1981).
106. Pandit, S.S., Naik, S.D., Jathar, V.S. and Kulkarni, A.B. IndianJ.Chem., 14B, 907-8 (1976).
107. Nauumenko, V.I., Dudkin, M.S.and Bezusov, A.T. Rastit.Resur., 19, 529-32 (1983).
108. Chowhan, Ugam Kumari, Shukla and Ravindranath, J.Sci.Res. (Bhopal, India), 6, 49-50 (1984).
109. Chugan, and Ugamkumari, Proc.Natl.Acad.Sci, India, Sec.B, 54, 236-9 (1984).
110. Sinha, S.K.Pand Dogra, J.V.V. Int.J.Crude Drugs Res., 23, 77-86, 1985.
111. Goyal, M.M., Rani, and KumKum, Fitoterapia, 60, 2,163-4 (1989).
112. Schmidt, John, H.Wells, and Randy, J.Agri.Food Chem., 38, 505-8 (1990).
100
113. Matlawska, Herba Pol., 36, 65-9 (1990).
114. Cao, Jianhong, Qi and Yiping, Zhongguo Zhongg yao Za zhi, 18, 681-2 (1993).
115. Haq, Q.N.and Haq, M.M. Sci.Res. (Dacca), 7, 175-180 (1970).
116. Rao, R.V., Krishna, Satyanarayana, T., Rao, Kameswara, B.V. Fitoterapia, 55, 249-50 (1984).
117. Bagerhotta, Asha, and Bannerjee, N.R. Cur.Sci., 59, 690-2 (1985).
118. Thampi et.al., Indian J.Pharm., 30, 236 (1968).
119. Prasad, D.N. and Achari, G. Indian J.of Pharm., 28, 241-4 (1966).
120. Kholkute, S.D., Munshi, S.R., Naik, S.D.and Jathar, V.S. Indian J.Exp.Bio., 16, 696-8 (1978).
121. Gunatilaka, Leslie, A.A., Sootheeswaran, S., Balasubramaniam, S., Chandrasekaran, A., Indumathie, Sriyani and Badra, H.T. Planta Med., 39, 66-72 (1980).
122. Goyal, M.M., Rani and KumKum, J.Indian Chem.Soc., 65, 74-6 (1988).
123. Sushikumar, Bagchi, G.D., Darokar, M.P.and Kumar, S.J. Int.J.Pharmacogn., 35, 179-184 (1997).
124. Kurma, S.R.and Mishra, S.H. Indian Drugs, 35, 92-97 (1998).
125. Otero, R., Nunez, V., Jimenez, S.L., Fonnegra, R., Osorio, R.G., Garcia, M.E and Diaz, A. J. Ethnopharmacology, 71, 505-511 (2000).
126. Narayanan, C.R., Joshi, D.D.and Majumdar, A.M. Curr.Sci., 53, 579 (1984).
127. Chemical abstracts. 86, 68506a (1977).
128. Chemical abstracts. 100, 99962h (1984).
129. Bolognesi, Andrea, Poliro, Letizia, Olivera, Fabiola, Valbenesi, Paola; Barbieri, Luigi; Batteli, Gullia, M., Carusi, Vittoria, M., Benvenuto, Eugenio, Del vacchiomarir, Stripe and Fiorenzo, Planta Med., 203, 422-429 (1997).
130. Gilani, A.H., Janbaz, K.H.and Shah, B.H. Pharnacol.Res., 37, 31-35 (1998).
131. Narayanan, C.R., Joshi, D.D., Majumdar, A.M.and Dhekne, V.V. Curr.Sci., 56, 139 (1987).
132. Seshagiri Rao, D. Econ.Botany, 11, 274-76 (1976).
133. Patil, R.B., Gandhi, T.P., Chakravarthy, B.K., Patel, R.J., Pundrikakashudu, K.and Dhyani, H.K., Indian Drugs, 23, 595-597 (1985).
101
134. Chang, Wen Shing; Lee, YiJean; Lu, Fung Jou; Chiang, and Hsuch Ching, Anticancer Res., 13, 2165 (1993).
135. Leoder, John, W., Nearn, and Roland, H. Heterocycles, 7, 113-118 (1977).
136. Agarwal, Sudhir, and Misra, K. Cur.Sci., 48, 141-3 (1979).
137. Dan, M.O.S., Dan, S.S., Mukhopadhayas, P.and Mukherjee, M.K. Sci.Cult., 48, 350-51 (1982).
138. Manzoor-I-Khuda., Hossain, and Golam, M.M. J.Sci.Ind.Res., 17, 134-5 (1982).
139. Wu, Yang Chang, Duh, Chang Yih, wang, Shang Kwei, Chen, Keh Shaw, Yang, Tsang and Heiung, J.Nat.Prod., 53, 1327-31 (1990).
140. Hasan, C.M., Islam, M.O.Rashid, M.A. Pharmazie., 50, 227-8 (1995).
141 Zhao, Gengxian, Jung, Jee, H., Smith., David, Karl, V., Mc Laughlin, and Jerry,L. Planta Med., 57, 380-3 (1991).
142. Chakarabarty, Manas, Nath, and Amar, C. J.Nat.Pro., 55, 256-8 (1992).
143. Zhao, G.X., Jung, J.H., Smith, D.L., Wood, K.V.and McLaughlin, J.L. Planta Med., 57, 380-3 (1991).
144. Ferdous, A.J., Islam, M.O., Hasan, C.M.and Islam, S.N. Fitoterapia, 63, 549-50 (1992).
145. Hara, Neriyuki, Asaki, Hitomi, Fujimoto, Yoshinori, Gupta, Yogesh Kumar, Singh, Ashish Kumar, Sahai Mahendra, Phytochemistry, 38, 189-94 (1995).
146. Chen, C.Y., Chang, F.R., Shih, Y.C., Hseieh, T.J., Chia, Y.C., Tseng, H.Y., Chen, H.C., Chen, S.J., Hsu, M.C. and Wu, Y.C. J.Nat.Prod., 63, 1475-8 (2000).
147. Wu and Yang Chang, Heterocycles, 29, 463-75 (1989).
148. Chakrabarty, Mans, Patra, and Amrendra, Indian J.Chem.Sec.B, 29B, 394-5 (1990).
149. Wu, Yang Chang, Heterocycles, 29, 463-475 (1989).
150. Sobti, A.K., Sharma, O.P.and Bhargava, A.K. Indian Phytopathol., 48, 191-3 (1995)
151. Gengxian Zhao, JeeH.Jung, David L.Smith, Karl V.Wood, and JerryL.McLaughlin, Planta Med., 57, 380-383 (1991).
152. Kinghern, A.D and Evans, F.J. J.Pharm.Pharmacol., 26, 408-12 (1974).
102
153. Baslas, R.K., Agrawal, and Rakesh, Indian J.chem., 19B, 717-8 (1980).
154. Veeranjaneyulu, K.and Das, V.S.D. J.of Exp.Bot., 33, 515-19 (1982).
155. Shieh, Wang Chung, Tsi, Jinn Lai, Kuo Li Chung Hsing Ta Hsuieh, 21, 1-9 (1984).
156. Lee, Seung Ho, Tanaka, Takashi, Nanaka, Genichiro, Nishioka and Itsue, Phytochemistry, 29, 3621-5 (1990).
157. Banerjee, D.K.and Pal, D.C. Ethanobiology in human welfare: abstracts of the fourth international congress of ethnobiology, Lucknow, Uttar Pradesh, India, 17-21 November, 340 (1994).
158. Khan, N.H., Rahman, M.and Nur-e-Kamal, M.S. Indian J.Med.Res., 87, 395-7 (1988).
159. Agrawal, Y.K., Roctz Chem., 51, 1265 (1977).
160. Joshi, Krishna, C., Prakash, Lalit, Shah, and Ramakant, J.Indian Chemical Soc., 54, 1104 (1977).
161. Srivastava, P.N., Mishra, G.S.and Sukla, Y.N. Proct.Nattl.Acad.Sci., India, Sec.A,47, 1-3 (1977).
162. Agrawal, Sudhir, Mishra, and Krishna, Chem.Scr., 12, 37-9 (1977).
163. Merchant, J.R., Bakshi, V.M.and Engineer, A.B. Indian J. Chem., Sec.B, 17, 87-8 (1979).
164. Baslas, R.K.and Agha, R. Himalayan Chem.Pharm.Bull., 2, 13 (1985).
165. Bailey, L. Manual of cultivated plants, Macmillan, New York, p.388 (1958).
166. Trivedi, C., Shine, S., Sharma, R.C. Indian Journal of Medical research, 57, 1070-1074 (1969).
167. Nadkarni, K.M., Nadkarni, A.K. and Chopra, R.N. Indian Materia Medica, Popular Prakashan, Bombay, V.1, p.548-550 (1976).
168. Agarwal, Vinti, and Chuhan, B.M. Plant Foods Hum.Nutr., 38, 189-97 (1988).
169. Zafar, R.and Neerja, Hamdard Medicus, 34, 49-61 (1991).
170. Forestieri, A.M., Monforte, M.T., Ragusa, S., Irovato, A.and Iauk, L. Phytotherapy Research, 10, 100-6 (1996).
171. Mandal, S.C., Maity, T.K., Das, J., Saha, B.P.and Pal, M. J. Ethnopharmacol. 72, 87-92 (2000).
103
172. Mandal, S.C., Maity, T, K., Das, J., Pal, M.and Saha, B.P. Phytother Res., 13, 430-2 (1999).
173. Mousa, O., Vuorela, P., Kiviranta, J., Adbel Wahab, S., Vuorela, H.et al., J. Ethnopharmacology, 41, 71-76 (1994).
174. Bhakuni, D, S., Goel, A.K., Sudha Jain, Mehrotra, B.N., Patnaik, G, K. and Ved Prakash, Indian J.of Experimental Biology, l26, 883-904 (1988).
175. Schul, Z.Juergen, Herrmann, M. and Karl, Univ.Hannover, 171, 193-9 (1980).
176. Gnusowski, B.Zygmunt, B.and Majchrzykowa, Herba Pol., 27, 325-31 (1981).
177. Swain Anne, R., Dutton, Stephan, P., Truswell, and A.Stewart, J.Am.Diel.Assoc., 85, 950-60 (1985).
178. Maurer, Bruno, Hauser, Arnol, Froidevaux, and Jean Claude, Tetrahedron lett., 27, 2111-12 (1986).
179. Kataoka, Eiko, Tokue, Chiyoko, Tanimura, and Wahachiro, Nogaku Shuho, 31, 189-196 (1987).
180. Pieribattesti, Jean, Smadja, S., Jacgueline, Monton, and Jean, M. Dev.Food Sci., 18, 697-706 (1988).
181. Shaban, M.A.E.Kandeel, K.M.Yacout, G.A.and Mehaseb, S. Acta Aliment., 17, 95-101 (1988).
182. Chandrasekar, K., Vijayalakshmi, V.and Deosthale, Y.G. J.Food Sci.Technol., 32, 5, 403-5 (1995).
183. Okugawa, Hitoshi, Moriyasu, Mastaka, Matsushita, Sakae, Saiki, Kayoko, Fujioka, Akihiro Kato, and Atsushi, Shoyakugaku zasshi, 42, 94-7 (1988).
184. Maurer, Bruno, Hauser, and Arnol, Chem.Ind. (London), 18, 587-91 (1988).
185. Elkady, I.A., El-Maraghy, S.S.M., Mostafa, M.and Eman, Folia Microbiol., (Prague) 40, 3, 297-300 (1995).
186. Yamahara, Johji, Kimura, Hitoshi, Kobayashi, Masaaki, Sawada, Tokunosuke, Fujimura, Hajime, Chisaka, and Takashi, Yakugaku Zasshi, 103, 9, 979-85 (1983).
187. Haranth, P.S.R.K., Akther, M.H.and Sharif, S.I. Phytother.Res., 1, 91-2 (1987).
188. Yamahara, Johji, Kashiwa, Hiroyuki, Kishi, Kazumasa, Fujimura, and Hajimo, Chem.Pharm. Bull., 37, 3, 855-6 (1989).
104
189 Kubo, sao, imejima, asaki, Muroi, and Hisae, J.Agri.Food Chem., 39, 1984-6 (1991).
190. Jain, P., Khanna, N.K., Trehan, N., Pendse, V.K.and Godhwani, J.L.Indian J. Experimental Biology, 32, 633-636 (1994).
191 El-Tahir, K.E.H., Shoeb, H. and Al.Shora, H. Saudi pharmaceutical Journal, 5, 96-102 (1997).
192. Al-Zuhair, H.El-Sayeh, B.Ameen, H.A. and Al-Shoora, H.Pharmacological Research, 34, 79-82 (1996).
193. Gopalan, A. and Ratnambal, M.J. Indian perfumes, 33, 63 (1989).
194. Cho, Kang-Jin, Kim, Jin-Weon etal., Agri.Chem.Biotechnology, 44, 12-15 (2001).
195. Govindarajan, V.S., CRC Crit.Rev.Food.Sci.Nutr., 17, 189 (1982).
196. Yoshikawa, M., Yamaguchi, S., Kunimik, K., Matsuda, H., Okaoy, Yamahara, J. and Murakami, N., Chem.Pharm.Bull., 42, 1226 (1994).
197. Endo, K., Kanno, E. and Oshimay, Phytochemistry, 43, 273 (1996).
198. Kikuzahi, H. and Nakatani, N. Phytochemistry, 43, 273-77 (1996).
199. Muhamed bin Zakaria, Ibrahim, Halijah, Malays.J.Sci., 8, 125-8 (1986).
200. Menut, C., Lamaty, G., Bessiere, J.M. and Koudou, J.Journal of essential oil research, 6, 161-164 (1994).
201. Asfaw, N. and Abegaz, B. Sinet, an Ethiopian journal of science, 18, 133-137 (1995).
202. Charles, R., Garg, SN., Sushil-Kumar, and Kumar, S. Fitoterapia, 71, 716-718 (2000).
203. Yu, BoLiang, Yu, B.L. Chemistry and Industry of forest- products, 18, 21-25 (1998).
204. Shoji, N., Iwasa, A., Takemoto, T., Ishida, Y., Ohizumi, Y. J.Pharm.Sci., 71, 1174-5 (1982).
205. Yamahara, J., Mochizuki, M., Huang, Q.R., Matsuda, H., and Fujimura, H. J.Ethnopharmacol., 23, 299-304 (1988).
206. Amagaya-S, Hayakawa-M, Ogihara-Y, Fujiwara-K, Oka-H, Oshio-H, and Kishi, T. J. Ethnopharmacol., 25, 181-187 (1989).
207. Mascolo, N., Jain, R., Jain, S.C.and Capasso, F.J.Ethnopharmacol., 27, 129-140 (1989).
105
208. Watanable, K., Tamaru, N., Takihara, H. and Yoshida, M. J.Ethnopharmacol., 43, 191-196 (1994).
209. Umeda, M., Amagaya, S. and Ogihara, Y. J. Ethnopharmacology, 26, 255-269 (1989).
210. Srivastava, K.C. and Mustafa, T. Med.Hypotheses, 29, 25-8 (1989).
211. Mustafa, T.and Srivastava, K.C. J. Ethnopharmacol., 29, 267-73 (1990).
212. Ikarashi, Y., Yuzurihara, M., Sakakibara, I., Takahashi, A., Ishimaru, A., Ishimaru, H.and Maruyama, Y. Phytomedicine, 8, 8-15 (2001).
213. Yamahara, J., Huang, Q.R., Li, Y.H., Xu, L. and Fujimura, Chemical and Pharmaceutical Bulletin, 38, 430-31 (1990).
214. Endo, K., Kanno, E. and Oshima, Y. Phytochemistry, 29, 797-799 (1990).
215. Huang, Q.R., Matsuda, H., Sakai, k., Yamhara, J. Tamai, Y. J.of the Pharmaceutical Society of Japan, 110, 936-942 (1990).
216. Huang, Q.R., Iwamoto, M., Aoki, S., Tanaka, N., Tajima, K., Yamahara, J., Takaishi, Y., Yoshida, Y., Tomimatsu, T. and Tamai, Y.Chemical and Pharmaceutical Bulletin, 39, 397-399 (1991).
217. Sugaya, E., Ishige, A., Sekiguchi, K., Yuzurihara, T., Iizuka, S., Sugimoto, A.,
Takeda, S., Wakui, Y., Ishihara, K. and Aburada, M. J.Ethnopharmacol., 34, 13-19 (1991).
218. Nishizawa, K., Saito, H. and Nishiyama, N. Phytotherapy Research, 5, 97-102 (1991).
219. Hata, Y., Pancho, L.R., Nojima, H. and Kimura, I. Biological and Pharmaceutical Bulletin, 21, 792-794 (1998).
220. Konoshima, T., Takasaki, M., Kozuka, M. and Tokuda, H. Journal of the Pharmaceutical Society of Japan, 114, 248-256 (1994).
221. Tanushree-Chakraborty, Babu, S.P.S., Sukul, N.C., Chakraborty, T.and Sinha Babu, S.P.Environment and Ecology, 12, 774-776 (1994).
222. Yokozawa, T., Miyauchi, I., Oura, H., Zheng, P.D. and Zhou, J. J.Phytotherapy Research, 8, 456-460 (1994).
223. Lumb, A.B. Thrombosis and Haemostasis, 71, 110-111 (1994).
224. Tsuji, M., Yoshida, Y., Kodama, K., Okazaki, M. and Oguchi, K. Phytotherapy Research, 8, 100-102 (1994).
106
225. Sohini, Y.R., Padmaja-Kaimal, Bhatt, R.M. and Kaimal, P. J Ethnopharmacol., 45, 43-52 (1995).
226. Sohini, Y.R., Padmaja-Kaimal, Bhatt, R.M. and Kaimal, P. J.Ethnopharmacol., 45,141-147 (1995).
227. Belford, R. Australian Journal of Medical Herbalism, 6, 94-98 (1994).
228. Guh-JihHwa, Ko-fengnNien, Jong-TingTing, Ten CheMing, Guh, J.H., Ko, F.N., Jong, T.T. and Teng, C. M. J. Pharmacy and Pharmacology, 47, 329-332 (1995).
229. Sohni, Y.R. and Bhatt, R.M. J. Ethnopharmacol., 54, 119-124 (1996).
230. Parida, M.M., Pandya, G., Bhargawa, R. and Jana, A.M. Indian Journal of virology, 13, 101-105 (1997).
231. Calixito, J.B. and Cabrini, Phytotherapy Research, 11, 32-38 (1997).
232. Vimala, S., Norhanom, A.W. and Yadav, M. British J. of Cancer, 80, 110-116 (1999).
233. Mathur, A.K., Sawhney, M. and Bhushan Kumar, Indian J. Indigenious Medicines, 17, 25-29 (1996).
234. Luo Lan, wang Jian Nong, Kong LingDong, Jiang QingGuang, Tan RenXiang, Luo, L., Wang, J.N., Kong, L.D., Jiang, Q.G. and Tan, R.X. J.Ethnopharmacol., 73, 277-281 (2000).
235. Usha, K., Saroja, S., Sushil Kumar (ed.), Hasan, S.A, SamreshDwivedi (ed.), Kukreja, A.K., Ashok Sharma(ed), Singh, A.K., Srikant Sharma(ed), and Rakesh Tewari, J. of Medicinal Aromatic Plant Sciences, 22-23, 182-184 (2001).
236. Cohen, Stanley, Cannan et al., Arthiritis and Rheumatism, 44, 1984-1992 (2001).
237. Yadava, R.N., Kumar, and Saurabh, Asian J.Chem., 10, 764-766 (1998).
238. Gogte, M.G., Ananthasubramanian, L., Nargund, K.S. and Battacharyya, S.C. Indian J.chem., 25B, 233-8 (1986).
239. Sohani, Jayant.S., Rojatkar, Supada, R., Kulkarni, Mandakini, M., Dhaneshwar, Narayandatta, N., Tavale, Sudam, S., Gururow, Tayur, N., Nagasampagi, and Bhimsen, A. J.Chem.Soc.,Perkins Trans., 2, 157-160 (1988).
240. Atta-ur-Rahman, Shekhani, Mohammed Salah, Perveen, Shahnaz, Habib-ur Rehman, Yasmin, Afshan, ZiaulHaque, Atiya, Shaikh, Dilnawaz, J.Chem.Res. Synop., 3, 68 (1989).
107
241. Shekhani, M.S., Shah, P.M., Khan, K.M.and Atta-ur-Rahman, J.of Natural Products, 54, 882-885 (1991).
242. Chughtai, M.I.D., Khokhar, Irshad, Ahmed, and Ashfaq, Sci.Int. (Lahore), 4, 151-4 (1992).
243. Rojatkar, Supada, R., Nagasampagi, and Bhimsen, A. Phytochemistry, 31, 3270-1 (1992).
244. Yadava, R.N., Kumar, Saurabh, Fitoterapia, 70, 127-129 (1998).
245. Singh, S.K., Saroj, K., Tripathi, V.J., Singh, A.K. and Singh, R.H. International Journal of Crude Drug Research, 26, 235-239 (1988).
246. Chen, Yu.P., Hsu, Hong, Y., Ruo, Tsuen, I., Iguchi , Kazuo, Kakisawa, and Hiroshi, Phytochemistry, 12, 3000 (1973).
247. Rao, K., Sunder, Jones, G.P., Rivett, D.E. and Tucker, D.J. Fett Wiss.Technol., 94, 37-38 (1992).
248. Sinha, B.N., Sasmal, D. and Basu, S.P. Fitoterapia, LXVIII, 75-78 (1997).
249. Proct.Nat.Acad.Sci.India, 41A, 21 (1971).
250. Kitagawa, Isao, Inada, Akira, Yosioka, Itiro, Somanthan, R. and Sulthanbawa, M.U.S. chem.pharm.Bull., 20, 630-32 (1972).
251. Bhatnagar, S.C., Awasthi, Y.C. and Mitra, C.R. Phytochemistry, 11, 465-7 (1972).
252. Macbride, Kitakawa, Isao, Shirakawa, Kiyoharru, Yosikawa, and Masiyaki, Chem.Pharm.Bull., 26, 1100 (1978).
253. Yoshikawa, Kazuko et.al., J.Nat.Prod., 63, 1679 (2000).
254. Tou and Pin, G. Jpn.kokai Tokkyo koho, J.P2002, 194, 381 My Appl., 200,005,886, (2000).
255. Yamahara, Johji, Shintani, YoshiKo, Konoshima, Takao, Sawada, Tokumsuke, Fujimura, and Hajime, Yukugaku Zasshi, 95, 1179-82 (1975).
256. Dinesh Chandra, Indian Journal of Pharmacology, 33, 108-111 (2001).
257. Raina, M.L., Dhar, K.L. and Atal, C.K. Planta Med., 30, 198-200 (1976).
258. Variyar, P.S. and Bandyopadhyay, Chromatograpia, 39, 11-12, 743-746 (1994).
259. Wagner, H. and Bladt, S. Plant Drug analysis, A Thin Layer Chromatography Atlas, Springer, Berlin.2nd ed. 292 (1996).
108
260. Siddiqui, B.S. Begum, S., Uulzar, T., Farhat, and Noor, F. Phytochemistry, 45, 1617 (1997).
261. Indian Herbal Pharmacopoeia, IDMA Mumbai and RRL JammuTawi, V.II, p.93-101 (1999).
262. Al-Jalay, Bushra, Blank, Greg, Mc Connell, and Barry, J.Food Prot., 50, 25-7 (1987).
263. Jain, S.R. and Kar, A. Planta Medica, 20, 118-123 (1971).
264. Islam, S.N., Monira Ahsan, Ferdous, A.J., Faroque, A.B.M. and Ahsa, M. Bangladesh J. of Botany, 19, 99-101 (1990).
265. el Mofty, M.M., Soliman, A.A., Abdel Gawad, A.F., Sakr, S.A. and Shwaireb, M.H. Oncology, 45, 47-52 (1988).
266. Tripathi, P., Tripathi, G.S. and Tripathi, Y.B. Fitoterapia, 60, 539-42 (1989).
267. Rao, P.G.M., Rao, S.G., Vasanthkumar, Ramnarayan, K.Nayak, S.S., Kamath, S.S.K., Srinivasan, K.K. and Kumar, V. Fioterapia, 64, 108 (1993).
268. Lee, E.B., Shin, K.H. and Woo, W.S. Arch.Pharmacol.Res., 7, 127 (1984).
269. Houghton, P.J., Astaniou, A., Grange, J.M. and Yates, M. J. Pharmacy and Pharmacology, 46, 1042 (1994).
270. Sharma, U.D. Sachitra Ayurved., 47, 95-96 (1994).
271. Ghatak, S.S. and Bhusan, T.K. Environment and Ecology, 13, 284-286 (1995).
272. Sallal, A.K.J. and Alkofahi, A. Biomedical Letters, 53, 211-215 (1996).
273. Khajuria.A., Thusu, N., Zutshi, U. and Bedi, K.L. Indian Drugs, 34, 557 (1997).
274. Reen, R.K., Weibel, F.J. and Singh, J. J.Ethnopharmacol., 58, 165 (1997).
275. Bhakuni, D.S., Srivastava, S.N., Sehgal, S.L., Kaul, K.N. J.Sci.Ind.Res., (India), 21B, 48-49 (1962).
276. Subramanian, Sankara, S., Nair, A.G.R. Phytochemistry, 11, 3095-6 (1972).
277. Joshi, Krishna, C., Singh, P. and Mehra Alka, Planta Med., 37, 64-66 (1979).
278. Roy, R., Pandey, V.B. Phytochemistry, 37, 1775-6 (1994).
279. Roy, R., Pandey, V.B. Indian J. Natural Products, 11, 13-14 (1995).
280. Tian, J., Zhao, Q.S., Zhang, H.J., Lin, Z.W., Sun, H.D. J. Natural Products, 60, 766-769 (1997).
109
281. Roy Ruchira, Singh, U.P., Pandey, V.B. Orient Journal of Chemistry, 11, 145-148 (1995).
282. Murugesan, T., Rani, S., Ahamed, N., Rajaram, S., Saluja, R. J. Ethnopharmacol., 68, 315-319 (1999).
283. Khan, M.M.A.A., Singh, N., Dhawan, K.N. National Academy of Science Letters, 19, 145-148 (1996).
284 Shah, C.S. and Khanna, P.N. Indian J.Pharm., 26, 140 (1964).
285. Itch, T., Ishii, T., Tamura, T.Matsumoto, T. Phytochemistry, 17,971-7 (1978).
286. Khaleque, A., Nirmolendu Roy, Wahed Miah, M.A. and Md.SadrulAmin, Sci.Res., (Dacca, Pakistan), 2, 147-51(1955).
287. Khaleque, A., Wahed, Mian, M.A., Ehsanul Huq, and Md.SadrulAmin, Sci.Res., (Dacca, Pakistan), 3, 216 (1966).
288. Khanna, Pushpa, Khanna and Renu, Indian J.Pharm., 38, 120-3 (1976).
289. Noma, Masana, Naguchi, and Masao, Phytochemistry, 15, 1701-2 (1976).
290. Nag and Tej, N. Indian J.Pharm., 38, 80-1 (1976).
291. Bhattacharya, T.K., Vedantham, T.N.C., Subramanian, Sankara, S. Indian J.Pharm., 39, 119-20 (1977).
292. Siddiqui, Salimuzzaman, Sultana, Naheed, Ahmed, Salman, S., Haider and Imtiaz, S. J.Nat.Prod., 49, 511-13 (1986).
293. Siddiqui, Salimuzzaman, Sultana, Naheed, Ahmed, Salman, S., Haider, and Imtiaz, S. Phytochemistry, 26, 2641-3 (1987).
294. Siddiqui, Salimuzzaman; Ahmed, Salman, S., Mahmood Tariq, Pak.J.Sci.India Res., 30, 567-8 (1987,).
295. Shingu, Kazushi, Kajimoto, Tetsuya, Furusawa, Yorika, Nohara and Toshihiro, Chem.Pharm.Bull., 35, 4359-61 (1987).
296. Mahmood, Tariq, Ahmad, S., Salman, Siddiqui, Salmuzzaman, Heterocycles, 27,101-3 (1988).
297. Kundu, Sujata, Sinha, Subbash,C., Bagchi, Anjana, Ray, Anil, B., Phytochemistry, 28, 1769-70 (1989).
298. He, Liyi, Zhang, Guan de, Tong, Yuyi, Sagara, Kazuhika, Oshima, Toshiyuki, Yoshida, Tsuguchika, J.Chromatogr., 481, 428-33 (1989).
299. Shingu, Kazushi, Furusawa, Yoriko, Nohara, Toshihiro, Chem.Pharm.Bull., 37-8, 2132-5 (1989).
110
300. Gupta, Mohini, Baghchi, Anjana, Roy, Anil,B. J.Nat.prod., 54, 559-602 (1991).
301. Gupta, Mohini, Manikam, M., Sinha, Subbash, Sinha Baghchi, Anjana, Ray, Anil,B., Phytochemistry, 31, 2423-5 (1992).
302. Jahromi, M.A., Farboodniay, Manickam, M., Gupta, Mohini, Oshima, Yoshiteru, Hatakeyama, Susumi, Ray, Anil,B. J.Chem.Res, Synop., 16, 234-5 (1993).
303. Manickam, M., Kumar, S., Sinha-Baghchi, A., Sinha, S.C., Ray, A.B., J.Indian Chem.Soc., 71, 393-9 (1994).
304. Ali Mohad, Shuaib, Mohd., Indian J.Pharm.Sci., 58, 243-245 (1996).
305. Sahai, M.,Manickam, M., Gupta, M., Srivastava, A., Ray A.B. Indian,J., Chem.Soc., 76, 95-97 (1999).
306. Manickam, M., Srivastava, A., Ray, A.B. Phytochemistry, 47, 1427-1429, (1998).
307. Decarvalho and Paulo, An Acad.Brazil.Cienc., 42, 245-50 (1970).
308. Rastogi and Mehrotra, Compendium of Indian medicinal plants, PID, New Delhi, Vol.4, p.256 (1993).
309. Khan, F.Z., Alam, M., Saleem, R., Rashid, I. Pak.J.Pharm.Sci., 5, 55-61 (1992).
310. Singh, V.P., Prakash, O.M. and Ray, A.B., Mycology, 29, 96-99 (2001).
311. Manickam, M., Padma, P., Chansouria, J.P.N.and Ray, A.B. Phytother.Res., 11, 384-385 (1997).
312. Dharam Bal Parihar and Sikhibhushan Dutt., J. Indian Chem. Soc., 27, 77-80 (1950).
313. Arigoni, Gopinath, Govindachari, T.R., Parthasarathy, Viswanathan, and Wildmn, Proc. Chem. Soc., 446 (1961).
