SEC_II (Herbal Technology)

Unit_3
(Phytochemistry)

Phytochemistry: Active principle and methods of their testing- identification & utilization of the
medicinal herbs; Catharanthus roseus (cardiotonic), Withania somnifera (drugs acting on nervous
system), Clerodendron phlomoides (anti-rhematic), and Centella asiatica (memory booster)

General Techniques Involved in Phytochemical Analysis

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Identification & utilization of the medicinal herbs; Catharanthus roseus (cardiotonic)

Catharanthus roseus is an evergreen ever blooming herb which has its origin in central
Madagascar island. Cathranthus roseus is one of the best-studied medicinal Plants. It is an erect,
bushy, evergreen and ever blooming herb.

Nomenclature and taxonomy

The name Catharanthus L.G. Don is derived from the latin words Katharos (pure) and
anthos (flower), referring to the neatness and beauty of the flower.

Taxonomic Hierarchy

Kingdom : Plantae
Division : Magnoliophyta
Class : Magnoliopsida
Order : Gentianales
Family : Apocynaeean
Genus : Catharanthus
Species : Catharanthus roseus
Vernacular names
Nayantara (Bengali), periwinkle, Church yard blossom, Dead-man's flower, Ainskati,
Rattanjor, Sadaphul, Sudukad umullai,Billagannerue (Telugu) Bara Massi and Sada Bahar
(Hindi), Gulfering(Kashmiri).

Characteristics

1. It is an erect branched deciduous herb woody at the base.

2. The branching starts from the base.
3. The leaves are short stalked leathery, petiole generally orbate, rarely oblong elliptic
having oblong tips.
4. The leaf blade ranges from 2 to 4 inch.
5. The younger leaves are particularly very soft, light green in colour.

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Alkaloids in Catharanthus:

Alkaloids are large group of naturally occurring, basicallyproduced from plants, bacteria,
fungi, and animals, chemical compounds. They mostly contain basic nitrogen atoms. Alkaloids
have a wide range of pharmacological activities including antimalarial, anticancer, anti-
arrhythmic, analgesic, antibacterial, antihyperglycemic activities etc. Because of their potent
pharmacological effects, alkaloids are the basis for many drugs.

More than 130 alkaloids have been identified from extract of different plant parts of C.
roseus. Because of large number of alkaloids it is very complicated and costly process to extract
and purify vincristine and vinblastine. However after the development of plant tissue cell culture
systems it is more reasonable to isolate these alkaloids as well as increasing their potential yields.

Traditionally, Catharanthus roseus has been used in large scale to treat the diabetes and
high blood pressure. C. roseus has some diuretic action, meaning that it may promote the loss of
urine from the body. This effect may help to relieve high blood pressure, but other diuretics are
both safer and more effective to use. The high risk of serious side effects from Catharanthus
roseus makes taking it inadvisable.

Types of alkaloids
A wide range of alkaloids has been discovered from C. roseus. More than 130 alkaloids
of the indole and the dihydro-indole groups have been isolated and characterized from different
plant organs. A few importantones are described as follows:

Vinblastine:

Vinblastin is very important alkaloidextracted from leaf part of C. roseus. Previously the
cost of exraction and purification of vinblastin was very high but now after the advancement of
various biotechnological approaches, new protocols for rapid production have been developed.
Vinblastin inhibits the cell cycle of cancer cells. It binds with tubulin, and inhibits the formation
of microtubules. Due to the inhibition of formation of microtubules, cell cycle arrest in M phase
because of none separation of chromosomes durin anaphase of mitosis. Vinblastin is mainly used
for treatment of Hodgkin’s disease, lymphocytic lymphoma, histiocytic lymphoma, advanced
testicular cancer, advanced breast cancer, Kaposi’s sarcoma.

Vincristine:

Vincristine formally known as leurocristine, sometimes abbreviated “VCR”, is an

alkaloid extracted from Catharanthus roseus. It is normally used in cancer chemotherapy. The
chemical structure of vinblastine and vincristine are very similar, but their effects are not the
same. Vinblastine is used to treat specific types of cancer, such as Hodgkin’s disease, and
Vincristine is used in the treatment of acute lymphoblastic leukemia (ALL). When vincristine
is added to the treatment regimen for children suffering from acute lymphoblastic leukemia
(ALL), the survival rate reaches eighty percent.

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Other important uses:
1. Anti-diabetic activity:
The “periwinkle tea” prepared from leaf decoction of C. roseus is used for curing
diabetes. Hypoglycemic defects of a number of indole alkaloids such as Lochnerine,
tetrahydroalstonine and Vindolidine have also been revealed.
2. As a cardio-vascular drug:
Ajmalicine (Raubasine) present in root is reported to improve cerebral circulation
when given intra-muscularly to humans suffering from cerebral sclerosis (Jayanthi et al.
2010).
3. Anti-helminthic activity:
The plant has been reported to possess vermifungal properties. Dried leaves,
incorporated into the soil produce nematicidal and ovistatic effects.
4. Anti-bacterial and pesticidal activities:
Antibacterial action against Vibrio cholera, Mycobacterium Salmonella typhi
and Spinach mosaic virus have been revealed by in-vitro studies. Insect growth
inhibition and insect sterilant properties have also been reported in Heliothus armigera,
Spodolptera litura, Achoea janata and Gryllodes sigillatus.

Conclusion:

Catharanthus roseus L. is used for the treatment of various diseases like diabetes,
tumors and as a cardio-vascular drug in all over the world from the last several decades. Every
part of Catharanthus like root, stem, bark and flower are rich sources of several bioactive
compounds.

Identification & utilization of the medicinal herbs; Withania somnifera (drugs acting on
nervous system)

Withania somnifera Dunal belongs to the family solanaceae. It is a xerophytic plant,

found in the drier parts of India, Sri Lanka, Afghanistan, Baluchistan and Sind and is distributed
in the Mediterranean regions, the Canaries and Cape of Good Hope. It is found in high altitude
ascending to 5,500 feet in the Himalayas. This shrub is common in Bombay and Western India,
occasionally met within Bengal. It grows wildly throughout India particularly in hotter parts, on
waste places and on road sides. It is also cultivated for medicinal purposes in fields and open
grounds throughout India.

Withania somnifera (L.) Dunal

Taxonomic Hierarchy

Kingdom : Plantae
Order : Solanales
Family : Solanaceae
Genus : Withania
Species : W. somnifera
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Vernacular names
Ashwagandha, Indian ginseng, poison gooseberry or winter cherry

Characteristics

1. This species is a short, tender perennial shrub growing 35–75 cm (14–30 in) tall.
2. The branching starts from the base.
3. Leaves are dull green, elliptic, usually up to 10–12 cm (3.9–4.7 in) long.
4. The flowers are small, green and bell-shaped.
5. The younger leaves are particularly very soft, light green in colour.
6. The ripe fruit is orange-red.

Phytochemical analysis of W. somnifera revealed the presence of pharmacologically

active steroidal lactones named withanolides. Withanine, a group of alkaloids isolated from the
roots of the plant, forms 38% of the total weight of alkaloids. The principal withanolides
extracted from W. somnifera in India were withanolide D and withaferin A which exhibited
antitumor and cytotoxic properties. In addition to alkaloids, the plant also consisted of steroids,
saponins, phenolics, flavonoids, phytophenols, and glycosides. Also, it is widely used in
traditional medicine formulations as an antipyretic, analgesic, adaptogenic, and anti-
inflammatory agent.

Phytochemistry:
The plants are rich in phytochemicals such as alkaloids, steroids, terpenoids, etc., which
are important parts of food and folk medicine in the history of mankind. Power and Salway
initiated phytochemical studies of W. somnifera in 1911 with the isolation of withaniol,
somnirol, somnitol, withanic acid, phytosterol, ipuranol, and alkaloids from alcoholic extracts of
leaves and roots. Alkaloids isolated in the study mentioned above were named as somniferine,
somnine, somniferinine, withamine, withanmine, pseudowithamine, and withanaminine. It was
investigated that alcoholic extract of the plant contains various phytochemicals such as tropine,
choline, pseudotropine, dl-isopelletierine, cuscohygrine, anahygrine, and anaferine.

Furthermore, a pyrazole alkaloid, withasomnine, was separated from the alcoholic root
extract of W. somnifera. In 1980 the presence of tisopelletierine, 3α-tigloyloxtropine,
cuscohygrine, 3-tropyltigloate, hygrine, dl-isopelletierine, withasomnine, mesoanaferine,
withanine, somniferine, hentriacontane, withananine, visamine, ashwagandhine, and
pseudowithanine in methanolic extract of the plant leaves was reported. Besides, the methanolic
extract of the plant was shown to have withaniol, reducing sugars, acylsteryl glucosides, ducitol,
starch, hantreacotane, iron, and amino acids such as aspartic acid, proline, tyrosine, alanine,
glycine, glutamic acid, cysteine, and tryptophan. Seven new withanosides I-VII were obtained
from methanolic extract of the plant root, and structures were confirmed using Fast Atom
Bombardment-Mass Spectrometry, 13C and 1H nuclear magnetic resonance (NMR), and UV-
Visible spectroscopic techniques.

Withasomidienone, isolated from the methanolic root extract of the plant, showed three
double bonds at 1, 4, and 24 positions, which are a characteristic feature of most of the
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withanolides. In another investigation, withaoxylactone and somnifericin were isolated from the
plant and characterized using various techniques

Folk medicinal uses

Different parts of the plant, such as leaves, roots, flowers, bark, and stem, are
traditionally used to cure heart problems, pain, liver disorders, fever, respiratory infections,
wounds, ulcers, and sex-related diseases. The curing potential of the plants used in ancient
systems of medicines (Unani and Ayurvedic) owed to bioactives such as alkaloids, steroids,
phenolics, flavonoids, etc., The plants of W. somnifera is rich in aforesaid bioactives making it
the first choice of folk healers.

Pharmacological attributes:
Neurotic regeneration activity

The neurodegenerative disorder is actually the selective dysfunctioning or sometimes

death of neural cells in the central nervous system or regeneration of synaptic, neuronal, and
neurotic cells. The plant extract was fed to a group of mice for three weeks. It was declared that
the extracts showed neuroprotective and neuronal growth effects by reversing all
neurodegenerative processes. The methanolic root extract of the plant exhibited prominent
neurite regeneration even at a dose of 1.0 mM on a human neuroblastoma SH-SY5Y cell line.
Different derivatives of withanolides such as (20S,22R)-3,6α-epoxy-4,5β,27-trihydroxy-1-
oxowitha-24-enolide, 27-O-β-D-glucopyranosylpubesenolide 3-O-β-D-glucopyranosyl (1→6)-β-
D-glucopyranoside, 27-O-β-D-glucopyranosyl (1→6)-β-D-glucopyranosylpubesenolide 3-O-β-
D-glucopyranosyl (1→6)-β-Dglucopyranoside, 27-O-β-D-glucopyranosylpubesenolide3-O-β-D-
glucopyranoside, and (20R,22R)-1α,3β,20,27-tetrahydroxywitha-5,24-dienolide 3-O-β-
Dglucopyranoside) isolated from methanolic extract of roots produced positive neurite outgrowth
effects in rats. Sominone, a class of withanoside IV, was the major metabolite responsible for
significant axonal and dendritic regeneration with synaptic reconstruction in Alzheimer’s
disease.

Sitoindosides (VII–X) and withaferin extracted from roots were investigated on brain
cholinergic, gamma-aminobutyric acid-induced allergic and glutamatergic receptors in rats.
Results showed a slight increase in acetylcholinesterase activity in lateral septum with relative
lowering in the vertical diagonal band of lateral septum and globus pallidus. The root extract of
the plant reversed the effects of scopolamine-induced disruption, retention, and attenuation in
amnesia. It was revealed that withanolide A at a dose of 10 mmol/kg of body weight
reconstructed severely damaged neurons in mice.

Cardioprotective activity
Root powder of the plant when orally administered to hypercholesteremic rats reduced
total lipid cholesterol and triglycerides contents in subject animals. However, a significant
increase in HDL cholesterol level, liver bile, and 3-hydroxy-3-methylglutyral-Coenzyme A
reductase activity was also observed. Later on, it was noticed that tuber powder of W.

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somnifera (50 mg/kg of body weight) administered to albino rats showed cardioprotective and
anti-oxidant activities in isoproterenol-induced myocardial infarctions.

The alkaloids isolated from roots of W. somnifera plant exhibited prolonged hypotensive,
respiratory stimulant, and bradycardiac actions in dogs. The cardioprotective effect of hydro-
alcoholic extract of W. somnifera at doses of 25, 50, and 100 mg/kg was investigated in
isoprenaline (isoproterenol) induced myocardial necrosis using rats as model animals. A
significant cardioprotective effect with continuous restoration of the hemodynamic parameter
was observed. Various extracts of the plant had been used to increase the beating ratio of the
heart by lowering the lipid peroxidation level

Antimicrobial and antifungal activities

The plant also exhibited bursal disease virus inhibition in cytopathic effect reduction
assay. Monomeric glycoprotein isolated from root tuber of the plant inhibited the growth of fungi
such as Fusarium oxysporum, Aspergillus flavus, and Fusarium verticilloides, and bacterium
such as Clvibacter michiganensis subsp. Michiganensis. Aqueous and alcoholic root extracts
were examined for bactericidal potential using agar well diffusion assay. Butanolic sub-fraction
of methanolic root/leaf extract of the plant exhibited significant antibacterial potential
against Salmonella typhimurium while oral administration of aqueous leaf/ root extract of W.
somnifera exhibited bacteriostatic effect against S. typhimurium similar to chloramphenicol, the
standard drug. Different leaf, root, and stem extracts of the plant were investigated for
bactericidal activity against six bacterial strains, e.g., Staphylococcus aureus, Bacillius
subtilis, Escherichia coli, Raoultella planticola, Pseudomonas aeruginosa, Enterobacter
aerogens, and fungistatic action against Candida albicans and A. flavus using serial dilution and
disc diffusion methods. Among the extracts, aqueous leaf extract showed the highest activity
against R. planticola, while all others showed moderate antibacterial and fungistatic activities.

Various extracts and pure compounds obtained from the plant acted as potential
antibacterial and antifungal agents. In another investigation, leaves and roots of the plant were
extracted with methanol, diethyl ether, and n-hexane and assessed for antibacterial potential
against S. typhimurium and E. coli using agar plate diffusion assay. The methanol and hexane
based extract showed substantial antimicrobial effects. A considerable rise in antibacterial
activity of Tibrim was observed on co-administering with methanolic and hexane extracts of the
plant.

Conclusion:
The W. somnifera has fascinated the researchers more, owing to the traditional
therapeutic applications, neutraceutical potential, and pharmaceutical attributes. Folk healers
cure various ailments such as cancer, arthritis, diabetes, eyesores, asthma, pyrexia,
inflammations, hemorrhoids, ulcers, hepatitis, and wounds. Results of multiple preclinical trials
such as antidepressant, antibacterial, anti-inflammatory, cardioprotective, anti-oxidant,
antifungal, hepatoprotective, and hypoglycemic in animal models (mice, rats, and rabbits) are
encouraging. The plant further requires the attention of phytochemists for the isolation of
bioactives responsible for the biological activities so that new formulations are developed.

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 Thus, there is a dire need to isolate phytochemicals for the development of commercial
formulations for the obsessive-compulsive disorder. The plant parts have also been appraised for
clinical trials such as male infertility, antianxiety, bone and muscle strengthening potential,
hypolipidemic, and antidiabetic. Further clinical trials of different extracts of the plant are
required for the development of economical and safer drugs.

Identification & utilization of the medicinal herbs; Clerodendron phlomoides (anti-rhematic)

Clerodendrum phlomoides Hort.Ital. ex DC.

Taxonomic Hierarchy
Kingdom : Plantae
Order : Lamiales
Family : Lamiaceae
Genus : Clerodendron
Species : C. phlomoides

Vernacular names
Agnimantha (Sanskrit), Ganiyari, Ganira (bengali), Munja (Malayalam), Arni, Agathu,
Ganiyari, Urni (Hindi)

Characteristics

1. This species is a short, a bushy shrub about 5-15 ft.

2. The branching starts from the base.
3. Leaves deltoid, ovate-cordate.
4. Flowers bisexual, subactinomorphic, fragrant.
5. The younger leaves are particularly very soft, light green in colour.
6. Fruit drupaceous, obovoid, about 1 cm in diameter and black when ripe.

PHARMACOGNOSY:

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Conclusion:

More than 300 chemical constituents have been isolated and identified from the different
genus of Clerodendrum, and pharmacological studies indicated that the crude extracts and some
special monomer compounds of the genus Clerodendrum exert various biological activities, such
as anti-inflammatory and antioxidant, anticancer, antimicrobial, antihypertensive, anti-obesity,
anti-diarrheal, hepatoprotective, memory enhancing, and neuroprotective activities. Terpenes,
including monoterpene and its derivatives, sesquiterpene, diterpenoids, triterpenoids, as the
major characteristic constituents with significant biological activities, have great potential to be
developed into new drugs, especially for anti-inflammatory, antioxidant, anticancer, and
antimicrobial agents.

It can be concluded that phytochemical and pharmacology investigations were mainly

focused on Clerodendrum phlomoides Many other speciesof Clerodendrum are totally unknown
phytochemically and biologically. All in all, the omnibearing study on this genus Clerodendrum
should be performed as soon as possible, which will provide reliable theory evidence for better
exploit and utilize the resources of the species in this genus.

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Identification & utilization of the medicinal herbs; Centella asiatica (memory booster)

Centella asiatica (L.) Urban

Taxonomic Hierarchy
Kingdom : Plantae
Order : Apiales
Family : Apiales
Genus : Centella
Species : C. asiatica

Vernacular names
Thankuni (Bengali), Mandookaparni, Brahmi (Hindi)

Characteristics
1. This species is a short, tender perennial weed.
2. The branching starts from the base.
3. Stem is thin, cylindrical.
4. Leaves are arranged in small cluster of 1 to 5.
5. The flowers are small, green white to dark red.
6. The younger leaves are particularly very soft, light green in colour.
7. The ripe fruit is yellowish brown and slightly lobed.

Phytochemical Content of Centella asiatica:

The plant has long been used as folklore medicine for the treatment of a variety of
diseases. Chemically been identified leading to therapeutic properties. Asiatic acid, asiaticoside,
and madecassoside form the major constituents responsible for pharmacological value apart from
being rich in flavonoids and terpenoids. Centelloid was term given for different constituents of
secondary metabolites produced by plant which mainly comprised of pentacyclic triterpenoid
saponins. P-cymene-(44%) along with other volatile compounds was found to be in a prominent
amount in the essential oil of C. asiatica on analysis with gas chromatography-mass spectrometry
(GC-MS). Centellin, asiatic, and centellicin were isolated from the aerial part of the plant, and
further, their structures had been determined using 2D nuclear magnetic resonance technique.
From plant extract using high-performance liquid chromatography to identify bioactive
compounds, madecassoside, asiaticoside, madecassic acid, and asiatic acid were found in the
significant amount. Aquantitative estimation of triterpenoids showed highest asiaticoside content
(6.42%) in leaf samples collected in Mangoro region. New triterpene and a saponin, 2α,3β,23-
trihydroxyurs-20-en-28-oic acid and 2α,3β,23- trihydroxyurs-20-en-28-oic acid O-α-l-
rhamnopyranosy-(1→4)-O-β-dglucopyranosyl(1→6)-O-β-d glucopyranosyl ester, have been
isolated from the aerial part of C. asiatica, and their structures were determined using spectral
methods.
Cognitive function:

Asiatic acid was found to prevent spatial working memory and reduction of
neurogenesis defects in the hippocampal region caused by 5-FU chemotherapy. Water extract of
C. asiatica was observed to enhance synaptic differentiation and dendritic arborization with
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reference to Aβ which causes cognitive improvement. In a study, gotu-kola extract was
supplemented for weeks in defined concentration results showed to be effective in the treatment
of cognitive function impairment after stroke. Asiatic acid has potential to restore the impairment
of cell proliferation, spatial working memory caused by treatment with valproic acid.

Water extract helped to improve cognitive function by activation of antioxidant response

gene and mitochondrial biogenesis, normalized calcium homeostasis. In another study, asiatic
acid was found to enhance hippocampal neurogenesis which can serve as potent cognitive
enhancer.

Asiaticoside isolated from Hydrocotyle sibthorpioides helps in scavenging free radical

enhancing activity of antioxidant enzymes, improving synaptic plasticit. In cognitive-related
disorders, mitochondrial dysfunction and oxidative stress have major role.

C. asiatica have proved to carry neuroprotective potential. Significant improvement in

cognitive function was observed in the Morris maze test and retained glutathione and lipid
peroxidation and SOD activity in cortex and hippocampus to control levels. In another study C.
asiatica proved to have a protective role against D-glucose-induced biochemical, behavioral, and
mitochondrial dysfunction.

Acute administration of asiatic acid was studied on male Sprague-Dawley rats for
memory and learning, treatment at 30 mg/kg of asiatic acid significantly improved memory. The
healing effect of C. asiatica was observed against colchicine-induced cognitive impairment and
lead acetate-induced changes in oxidative biomarkers in the central nervous system. Cyclic AMP
response element binding protein phosphorylation level was enhanced in primary culture of rat
embryonic cortical cells on treatment with plant extract, thus improving memory function. In
another finding, it was found that high dose of plant extract-enhanced memory and increased
N100 constituent amplitude of event-related potential.

Conclusion:
C. asiatica is potential herb with an array of health-care applications. It is widely
accepted that plant has got neuroprotective activities and helpful in brain improvement. Plants
have proved to bear low toxicity and higher efficacy in clinical treatment with prominent
activities such as anticancer, antibacterial, antifungal, anti-inflammation, neuroprotection,
antioxidant, wound healing, and antidepressant. As C. asiatica is an endangered species using
plant tissue culture mass propagation major can be helpful, and callus and suspension culture
techniques can be harnessed for secondary metabolite extraction, which establish the chemical
compounds responsible for a wide range of therapeutic activity. Due to the presence of wide
bioactive compound, the plant has vast application. The plant can be a safer alternative for the
formulation of new drugs and clinical studies which help to safe application for humans.

Prepared by
Dr. S. A. Kowser Alam Laskar
Department of Botany
M.H.C.M.Sc. College, Algapur
07/08/21 [08:20PM] (Haflong)
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