06_chapter 2.pdf - Shodhganga
Transcript of 06_chapter 2.pdf - Shodhganga
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2. REVIEW OF LITERATURE
Ethno pharmacology is a highly diversified approach to drug discovery
involving the observation, description, and experimental investigation of
indigenous drugs and their biological reactions. It is based on botany, chemistry,
biochemistry, pharmacology and many other disciplines that contribute to the
discovery of natural products with biological activity.
Plants are considered to be medicinal if they possess pharmacological
activities of possible therapeutic use. Indian Materia Medica includes about 2000
drugs of natural origin almost all of which are derived from different traditional
systems. These activities are often recognized as a result of millennia of trial and error
but they have to be carefully investigated if new drugs are to be developed for use in
modern treatment.
2.1. MEDICINAL PLANTS SCENARIO IN INDIA:
The rich biodiversity of Indian subcontinent contributes to the wealth of
medicinal plants, which are very much used in traditional medical treatments
(Chopra et al., 1956). India is one of the 12 mega biodiversity centers with over
18,000 plant species. Over 2,500 species are formally recognized as having true
medicinal value. About 7500 plants have been used in local health traditions in rural
and tribal villages of India. Out of these, the medicinal efficacy of 4000 plants is
either little known or unknown to the mainstream population (Pushpangadan, 1996).
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About 1500 plants with medicinal uses are mentioned in ancient texts such as
Ayurveda, Siddha, Unani and Tibetan medicine (Table: 1). In fact it is the
responsibility of the scientific community to do scientific validation of the medicinal
property of the traditional claims.
TABLE 1: Medicinal plants used in various codified systems of Medicine in India
In recent years, the growing demand for herbal product has led to a quantum
jump in volume of plant materials traded within and across various countries in the
world. Though India has a rich biodiversity, the growing demand is putting a heavy
strain on the existing resources. While the demand for medicinal plants is growing,
some of them are increasingly being threatened in their natural habitat. According to
an all India ethno biological survey carried out by the Ministry of Environment &
S.NO MEDICAL SYSTEM NO.OF PLANTS USED
1 AYURVEDA 1587
2 SIDDHA 1128
3 UNANI 503
4 SOWA-RIGPA 253
5 HOMOEOPATHY 468
6 WESTERN 192
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Forests, Government of India, there are over 8000 species of plants being used by the
people of India for diseases like breast cancer (12%), liver disease (21%), human
immunodeficiency virus (22%), asthma (24%) and rheumatological disorders (26%).
Some of these plants are commonly used by people as folk medicines for hundreds of
years.
Over 17,500 species of higher plants occur in India, of which approximately
9,000 plants are known to be economically useful. Of these, 7,500 species are used for
healthcare by various ethnical communities in India (Arora, 1997). However various
traditional communities in India are using around 800 plants for curing different
ailments (Kamboj, 2000). India is the world’s largest supplier of raw materials. India
is also known to be exporting a number of medicinal plants. Some of the important
medicinal plants with high trade value are Cuscuta epithymum, Glycyrrhiza glabra,
Lavendula stoechas, Operculina turpethum, Pimpinella anisum, Smilax china, Smilax
ornata and Thymus vulgaris. The annual exports of India’s herbal sector add up to Rs.
807 Crores for the year 2004-05 (Ved and Goraya, 2007). This includes exports worth
Rs. 354.80 Crores related to plant raw drugs, Rs. 161 Crores related to plant extracts
and Rs.291 Crores related to medicants of Ayurvedic, Unani, Siddha and
Homoeopathic systems. It shows that the finished herbal products constitute nearly
36% of the total exports of India’s herbal sector and the balance 64% exports are in
the form of raw materials and extracts.
Three of the widely selling herbal medicines in developed countries, namely
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Allium sativum, Aloe barbadensis and Panax species are also available in India. India
is the largest grower of Psyllium (Plantago ovata) and Senna (Cassia senna) plants
and one of the largest growers of Castor (Ricinus communis) plant. The plants
Glycyrrhiza glabra, Piper longum, Adhatoda vasica, Withania somnifera, Cyperus
rotundus, Tinospora cordifolia, Berberis aristata, Tribulus terristris, Holarrhena
antidysenterica and Boerhaavia diffusa have been used in 52 to 141 herbal
formulations and triphala (Terminalia chebula, Terminalia bellerica and Emblica
officinalis) alone have been used in 219 formulations (BCIL, 1996).
The turnover of herbal medicines in India as counter products, ethnical and
classical formulations and home remedies of Ayurveda, Unani and Siddha systems of
medicine is about $1 billion with a meager export of about $80 million. Psyllium
seeds and husk, castor oil and opium extract alone account for 60% of the exports.
80% of the exports to developed countries are of crude drugs. However, only a small
proportion of plant species have been thoroughly investigated for their medicinal
properties (Frame et al., 1998) and undoubtedly there are many more novel
biologically active compounds to be discovered.
2.2. Drugs isolated from traditional Medicinal Plants
Natural products derived from medicinal plants play an important role as useful
tools in pharmacological studies. With the technological advancement of
science the isolation, identification and elucidation of chemical principles from
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natural sources have become much simpler and has contributed significantly to the
development of new drugs from medicinal plants.
There are hundreds of significant drugs and biologically active compounds
developed from the traditional medicinal plants, a few of which are mentioned here;
the antispasmodic agent vasicin isolated from Adhatoda vasica, anticancer agents
such as vincristine, vinblastine and D- Tubocurarine isolated from Catharanthus
roseus (Gurib-Fakim, 2006), antibacterial agents isolated from Diospyros
melanoxylon (Mallavadhani et al, 1998), antimalarial agent isolated from Sida acuta
(Karou et al, 2006), steroid and lancamarone with cardiotonic properties, lantamine
with antipyretic and antispasmodic properties from Lantana camara (Ghisalberti,
2000), antimicrobial agents isolated from Acorus calamus (Chowdhury et al, 1993),
antiviral, antibacterial and anti-inflammatory agents isolated from Urtica dioica
(Harborne and Buxter, 1993), anticancer agents isolated from Andrographis
paniculata, Phyllanthus amarus, Piper longum, Semecarpus anacardium, Withania
somnifera, Moringa oleifera, Aloe vera, Curcuma longa, Allium sativum and
Tinospora cordifolia (Balachandran and Govindarajan, 2005), promising and potent
antimalarial drug artemisinin isolated from Artemesia annua (Dhingra et al, 2000).
Fabricant and Farnsworth (2001) presented 86 important drugs developed from active
compounds and their clinical use, few are listed (Table 2).
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Table: 2 Drugs derived from plants with their ethno medical correlations and sources
Drug Action or Clinical use Plant source
Acetyldigoxin Cardiotonic Digitalis lanata
Aescin Anti-inflammatory Aescuus hippocastanum
Arecoline Anthelmintic Areca catechu
Aesculetin Anti-dysentery Fraxinus rhynchophylla
Ajamalicine Circulatory disorders Rauvolfia serpentiana
Allyl isothiocyanate Rubefacient Brassica nigra
Andrographolide Bacillary dysentery Andrographis paniculata
Anisodarnine Anticholinergic Anisodus tanguticus
Anisodine Anticholinergic Anisodus tanguticus
Asiaticoside Vulnerary Centella asiatica
Atropine Anticholinergic Atropa belladonna
Berberine Bacillary dysentery Berberis vulgaris
Bergenin Antitussive Ardisia japonica
Bromelain Anti-inflammatory Ananas comosus
Caffeine CNS stimulant Camellia sinensis
(+)-Catechin Haemostatic Potentilla fragaroides
Chymopapain Proteolytic Carica papaya
Cocaine Local anaesthetic Erythroxylum coca
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Codeine Analgesic; antitussive Papaver somniferum
Colchicine Antitumor agent Colchicum autumnale
Convallotoxin Cardiotonic Convallaria majalis
Curcumin Choleretic Curcuma longa
Cynarin Choleretic Cynara scolymus
Danthron Laxative Cassia Sps.
Deserpidine Antihypertensive Rauvolfia canesceas
Deslanoside Cardiotonic Digitalis lanata
Digitalin Cardiotonic Digitalis purpurea
Digitoxin Cardiotonic Digitalis purpurea
Digoxin Cardiotonic Digitalis lanata
Emetine Amoebicide; emetic Cephaelis ipecacuanha
Ephedrine Sympathetomimetic Ephedra sinica
Etoposide Antitumour agent Podophyllum peltatom
Gitalin Cardiotonic Digitalis purpurea
Glaucaroubin Amoebicide Simarouba glauca
Glycyrrhizin Sweetener Glycyrrhiza glabra
Gossypol Male contraceptive Gossypium Sps.
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2.3. Traditional uses of different species of Cassia
Cassia species have been of medical interest due to their good therapeutic
value in folk medicine. Cassia genus consists of more than 600 species throughout the
world. Some important species are Cassia fistula, Cassia grandis, Cassia hirsutica,
Cassia sieberiena, Cassia alata, Cassia tora, Cassia occidentalis, C. auriculata, C.
nigricans.
The anti-inflammatory and hepatoprotective activity of Cassia occidentalis, C.
fistula and C. sophera are also reported1 (Jafri et al., 1999; Burkill H.M, 1997). C.
sophora, C. italica, C. pumila are reported to have CNS depressant, anxiolytic and
hypnotic activity (Jain, 1996). The seeds of Cassia tora have good binding and
suspending property and are used as a substitute for coffee. Cassia mimosoides is
reported to have antiobesity activity (Caceres, 1991). Cassia tora, C. auriculata, C.
fistula, C. alata were reported to have anti-diabetic and antioxidant activity (Juvekar,
2006). The seeds of Cassia tora have good binding and suspending property and also
used as a substitute for coffee.
Among the species, Cassia auriculata L. (Leguminaceae) plant known for
curing diabetes, rheumatism, fever, leprosy, skin diseases, conjunctivitis, constipation
etc. has been chosen for detailed investigation of some pharmacological parameters
and toxicity studies. In the present study an attempt has been made to screen the
leaves of Cassia auriculata and to asses various pharmacological activity and toxicity
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of the isolated active molecules.
On the basis of the use of Cassia auriculata by the traditional healers all over
the world, research has been carried out in various pharmacological studies of this
plant such as toxicity, anti-diabetic, anti-inflammatory and antipyretic activities.
2.4. Cassia auriculata L.
Figure 1: Cassia auriculata L.
Cassia auriculata L. belonging to Leguminosae family is native of India and
Sri Lanka. It is commonly known as tangedu in Telugu and tanner’s cassia & avaram
senna in English in India. It is fast growing branched evergreen shrub with reddish
brown branches and vivid yellow flowers (Figure 1). It mainly occurs in the dry
regions of India and Sri Lanka. It is common along the sea coast and the dry zone in
Sri Lanka. The leaves are alternate, stipulate, paripinnately compound, slender,
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pubescent, 2-2.5cms long and 1-1.3cms broad. Its flowers are irregular, bisexual,
bright yellow and large (nearly 5cms across), the pedicels glabrous and 2.5cms long
the fruit is a short legume, 7.5-11cms long, 1.5cms broad, flat, thin, pale brown. 12-20
seeds per fruit are carried each in its separate cavity. This plant is said to contain
cardiac glycoside (sennapicrin) and sap, leaves and bark yield anthraquinones, while
the latter contains tannins.
Cassia auriculata has been widely used in traditional medicine. Its leaves,
flowers, roots and pods are employed in herbal medicine in and around the world. The
leaves have anthelmintic, anti-diabetic, laxative properties (Malhotra and Mishra,
1982). The root is used in decoctions against fevers, diseases of urinary system,
constipation, rheumatism, skin diseases, leprosy, conjunctivitis and diabetes (Joshi,
2000). The dried flowers are used as a substitute for tea in case of diabetes patients
and its leaves are also used for anti-diabetic, antioxidant (Juvekar and Halade, 2006;
Sawhney et al., 1978). Seeds are used in ophthalmia and dysentery (Kirtikar and
Basu, 1980; Vaidya 1998).
2.5. Toxicity
The active principles of medicinal plants may sometimes have deleterious
toxic effect on the physiological functioning of organisms. Hence there is a need for
scientific study of acute and chronic toxicity. Seventy five medicinal plants
of the traditional Ayurvedic pharmacopeia of Sri Lanka have been screened
chemically for alkaloids and pyrrolizidine alkaloids. Of these, Crotolaria juncea L.
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was found to contain pyrrolizidine alkaloids with biological effects consistent with
pyrrolizidine alkaloid toxicity. Mattei et al. (1998) reported the toxic effects and
behavioral level of Paullinia cupana in rats and mice and showed that, percentage
mortality was equivalent in control and treated groups. Similarly Hellion-lbarrola et al.
(1999) studied the acute toxicity and general pharmacological activities of the crude
hydroalcoholic rhizome extract of Kyllinga brevifolia. The LD50 was found to be 575
mg/kg.
Muko and Ohiri (2000) studied the toxicity by oral administration of aqueous
extracts of Emilia sonchifolia in rats arid concluded that the acute toxicity showed
LD50 of 860 mg/kg and 780 mg/kg for methanol and aqueous extracts respectively.
Rezaeipoor et al. (2000) reported the effect of Isatis cappadocica on humoral immune
response in mice. In particular, the dose of 0.25 g/kg suppressed the primary immune
response, while the dose of 0.5 g/kg stimulated the secondary immune response.
Monteiro (2001) evaluated the toxicity levels of Vernonia condensate (Asteraceae) at
a dose of 5000 mg/kg. The results suggested that V. condensate aqueous extracts
contained low acute toxicity and possed neither teratogenic nor mutagenic risks.
Marcelo et al. (2002) investigated the acute toxicity of
Stryphnodendron adstringen, after oral administration to mice, and its effect on certain
biochemical parameters were assessed in plasma of rats after 30 days of
administration. The results showed that the extract administered in a prolonged period
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produced toxic effects in the experimental animals. In another study, the total
alcoholic, total aqueous, and methanolic extracts isolated from the leaves of A.
marmelos were studied for their toxic effects. There were no remarkable changes
noticed in the histopathological studies after 50 mg/kg body weight of the
extracts of A. marmelos when administered intraperitoneally for 14 days
successively (Veerappan et al., 2007).
Toxicity studies were carried out for six plants used in the traditional Arab system
of medicine, namely: Aloe vera, Ammi majus, Areca catechu, Citrullus colocynthis,
Cuminum cyminum and Zizyphus spina-christi. In the acute toxicity test, C. colocynthis
showed a dose dependent toxic effect. On chronic treatment, per cent lethality was
found to be significant in A. catechu and C. colocynthis treated groups where
haematological changes were also observed (Shah, 2003).
The cytotoxicity of aqueous extracts (AE) and hydroalcoholic extracts (HAE) of
Phyllanthus amarus on Caco-2 cells were evaluated using neutral red uptake and
MTT test. A single oral dose of the extracts at 5 g/kg b.wt did not produce mortality
or any significant change in treated animals over a 14 day observation period (Lawson
et al., 2008). Various concentrations of aqueous extract of Phyllanthus niruri
were investigated in diabetic Wistar strain rats by Nwanjo (2007). Alterations were
observed in glucose level and some hepatospecific markers in a dose dependent
manner.
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2.6. Anti-diabetic activity
Diabetes Mellitus (DM) describes a metabolic disorder of multiple etiologies
characterized by chronic hyperglycemia with disturbances of carbohydrate, fat and
protein metabolism resulting from defects in insulin secretion, insulin action, or both
(Andrade-Cetto and Heinrich, 2005). It is caused by inherited and/or acquired
deficiency in the production of insulin by the pancreas, or by ineffectiveness of the
insulin produced and such a deficiency results in increased concentrations of glucose
in the blood, which in turn damage the body system, in particular the blood vessels and
nerves (Mukherjee et al., 2006).
2.6.1. Indigenous Treatment for Diabetes mellitus
The use of plants as therapeutic tools, especially those used to relieve
chronic pathologies, have had a remarkable role in the popular medicine of
different countries (Lee et al., 1999). Plant derivatives with purported hypoglycemic
properties have been used in folk medicine and traditional healing systems around
the world (Yeh et al., 2003) and they represent a vast source of potentially useful
dietary supplements for improving blood glucose control and preventing long-term
complications in type 2 diabetes mellitus (Gallagher et al., 2003). Since time
immemorial, various plants and plant derived compounds have been used in the
treatment of diabetes to control the blood sugar of the patients. In the period of
1907-1988, anti-diabetic activity was reported for about 343 plants at global level
and those are mostly used in the indigenous system of medicine as well as
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scientifically proven plants (Rahman and Zaman, 1989). Before the introduction of
insulin in 1922 the treatment of diabetes mellitus relied heavily on dietary
measures which included the use of traditional plant therapies (Gray and Flatt,
1999).
Before two decades, more than 400 traditional plants were recorded for the
treatment of diabetes mellitus with only a small number of scientific and medical
evaluations to assess their efficacy (Bailey and Day, 1989). In traditional medicine,
diabetes mellitus is treated with diet, physical exercise and medicinal plants, even
though, more than 1200 plants are used around the world in the empirical
control of diabetes mellitus and approximately 30% of the traditionally used
anti-diabetic plants were pharmacologically and chemically investigated (Alarcon-
Aguilar et al., 1998). On the other hand, potential hypoglycaemic agents have also
been detected for more than 100 plants used in anti-diabetic therapy.
Hypoglycemic action from some treatments has been confirmed in animal
models and non-insulin dependent diabetic patients and now a day a number of
hypoglycemic compounds have been identified.
Yeh et al., (2003) reviewed and analyzed, a total of 108 trails examining 36
herbs (single or in combination) and 9 vitamin/ mineral supplements, involving 4,565
patients with diabetes or impaired glucose tolerance. In their study there were 58
controlled clinical trials involving individuals with diabetes or impaired glucose
tolerance and most studies involved patients with type 2 diabetes. Of these 58 trials,
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the direction of the evidence for improved glucose control was positive in 76% and
very few adverse effects were also reported by them. There have been many studies
on hypoglycaemic plants and a great variety of compounds have been isolated (Table
3)(alkaloids, glycosides, terpenes, flavonoids, etc.), but the main fact is the further
development of such leads into clinically useful medicines and especially
phytomedicines or adequate nutritional supplements, which would be of direct
benefits to patients (Andrade-Cetto and Heinrich, 2005).
Table 3: Plants with significant Anti-diabetic compounds and their mode of action
Plant name Class of
Compound
Anti-diabetic
Compound
Mode of action
Abelmoscus moschatus Flavonoid Myricetin
(3,5,7,3’,4’,5’-
hexahydroxyflavone)
Anti-diabetic effect (Liu
et al., 2005)
Alpinia galanga
Polysaccharides
Protein-bound
polysaccharide
Increase serum insulin
levels, reduce blood
glucose levels and
improve glucose tolerance
(Quanhong et al., 2005).
Andrographis paniculata Diterpenoid Andrographolide Hypoglycemic activity
(Yu et al., 2003).
Citrus aurantifolia Polymethoxylated
flavones
Insulin resistance (IR)
model of hamsters
Increases the insulin-
sensitizing effects (Li et
al., 2006)
Curcuma longa
Ferulic acid 4-hydroxy-3-
methoxycinnamic acid
Hypoglycemic effect
(Ohnishi et al., 2004)
Stimulate insulin
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secretion (Nomura et al.,
2003).
Gymnema sylvestre Triterpenoid Gymnemic acid IV Hypoglycemic activity
(Sugihara et al., 2000).
Momordica charantia
Sterol Charantin Insulin-like activity and
stimulate insulin release
(Ng et al., 1986)
Punica granatum Tanins Gallic acid
Enhances cardiac PPAR-γ
mRNA expression and
cardiac glucose
transporter (GLUT)-4.
(Huang et al., 2005)
Tinospora cordifolia
Alkaloid Berberine
Hypoglycemic activity
and Alpha -glucosidase
inhibitor and decrease
glucose transport (Singh
et al., 2003).
Tribulus terrestris
Imidazoline
compounds
Harmane, norharmane
and pinoline
Increase insulin secretion
(Nadkarni, 1976; Cooper
et al., 2003).
In the Chinese traditional medical treatment of diabetes, compound recipes are
often used more than simple recipes (prescription with one medicine) due to
consideration of integrated effects of different medicines. There are hundreds of
prescriptions to aim directly at different symptoms of diabetes, and about 100 of
natural medicines and preparations are used in these prescriptions or folk simple
recipes and diets for diabetes care in China, most of which come from plants (Li et
al., 2004). More than 100 medicinal plants are mentioned in the Indian system of
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medicines including folk medicines for the management of diabetes, which are
effective either separately or in combinations (Kar et al, 2003). In India, the
plants such as Eugenia jambolana, Coccinia indica, Gymnema sylvestre, Momordica
charantia and Tigonella foenum-graecum are widely used for the treatment of diabetes
in traditional as well as modern medicine. At the present time,(Table 4) the plants like
Gymnema, fenugreek, bitter melon, ginseng, nopal, Aloe, bilberry, milk thistle,
garlic and ginkgo are the most active plants and have many active constituents with
anti-diabetic properties (McWhorter, 2001).
Table: 4 Plants with significant anti-diabetic activity in experimental animals
Botanical Name Parts Anti-diabetic mechanism References used
Aloe vera Leaf gel Hypoglycemic effects Roman-Ramos et al.,,
1995.
Annona squamosa Leaf Hypoglycaemic effect Gupta et al., 2005.
Azadirachta indica Leaf Hepatoprotective Chattopadhyay, 2005.
Eugenia jambolana Seed Anti-hyperglycaemic effect Grover et al., 2000.
Gymnema sylvestre Leaf Anti peroxidative,
hypoglycaemic and cortisol
lowering activities
Gholap and Kar, 2003.
Tinospora cordifolia Root Hypoglycaemic and
hypolipidaemic effects
Stanely and Menon,
2003.
Trigonella foenum
graecum
Seeds Anti-hyperglycemic and
hypoglycemic effects
Hannan et al, 2003.
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Panax ginseng Whole
plant
Hypoglycemic and
hypolipidemic effects
Jang et al., 2001.
Rhus barb Rhizome Anti-diabetic Lee et al, 2008.
Experimental diabetes has the advantage that it allows the analysis of the
biochemical, hormonal and morphological events that take place not only during the
induction of a diabetic state but also after it has become established and during its
evolution to a severe insulin deficiency or even death (Silva et al., 2002).
Streptozotocin has been extensively studied and has yielded the vast majority of
information relevant to human diabetes due to its wide range of safety compared to
other chemical agents capable of inducing diabetes. The effective dose (ED50) is 4-5
times lower than the lethal dose (LD50).
Table: 5 Mechanism of action of some Anti-diabetic principles isolated from
medicinal plants
Name of the
Plant
Compound Experimental
Model
Molecular level
functional properties in
the treatment of
diabetes & References
Citrus aurantifolia Polymethoxylated
flavone
Insulin resistance (
IR) model of hamster
Increases the insulin-
sensitizing effects might
occur as a consequence
of adipocytokine
regulation and increases
protein expression of
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PPAR-α and PPar-γ in
the liver(Li et al., 2006).
Ervatamia
microphylla
Conophylline
vinca alkaloid
AR42J-progenitor
cells, Neonatal
Streptozotocin –
treated rats.
Effectively inducing
differentiation of
pancreatic ductal cells
to β–cells (Kojima &
Umezawa, 2006.)
Eucommia
ulmoides
Flavanol
glycoside and
quercitine 3-O-α-
1-arabino
pyranosyl-(1-2)-β-
D-gluco
pyranoside
Bovine serum
albumin (10 mg/dl)
was incubated with
250 mMD-fructose
in the presence or
absence of test
material for 5 days
Inhibits the advanced
advanced glycation in in
vitro condition, and
decrease the secondary
complications (Kim et
al., 2004)
Hyssopus
officinalis
(7S,8S)-
Syringoylglycerol-
9-O-(60-O-
cinnamoyl)-b-d-
gluco pyranoside
Rat small intestine Α-Glucosidase inhibitor
activity(Matsuura et al.,
2004)
Pterocarpus
marsupium
7-O-α-1-rham
nopyranosyl oxy-
4-menthoxy-5-
hydroxy
isoflavone
L6 myotube &
adipocytic cell-line
3T3LI
Activation of PPARγ
through PPARγ agonists
are known to increase
the glucose uptake
through induction of
GLUT-4 mRNA
expression
(Anandharajan et al.,
2005)
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Punica granatum Galic acid (3,4,5-
trihydroxybenzoi
c acid)
Zucker diabetic fatty
rats, a genetic animal
model for type 2
diabetes
Enhanced cardiac
PPAR-γ mRNA
expression and restored
the down regulated
cardiac glucose
transporter(GLUT-4)
mRNA (Desrosis et al.,
2004).
Vitis vinifera Procyanidins
(flavonoids)
Streptozotocin-
induced diabetic rats,
L6E9 myotubes and
3T3-L1 adipocytes
Stimulate the glucose
uptake in adipocytes and
myotubes acts through
glucose transporter,
GLUT-4, in the plama
membrane (Pinent et al.,
2004).
2.6.2. Structure of Streptozotocin
Streptozotocin (STZ; N-nitro derivative of glucosamine) is a naturally
occurring, broad-spectrum antibiotic and cyto-toxic chemical that is particularly toxic
to the pancreatic, insulin producing β-cells The diabetogenicity of streptozotocin has
been correlated with a rapid reduction in pancreatic islet pyridine nucleotide
concentration and subsequent beta cell necrosis (Hayashi et al., 2006).The chemical
name is 2-deoxy-2-(3-methyl-3-nitrosourido)-O-glucopyranose. Molecular formula is
C8H15N3O7 (Fig.2). The structure is composed of a nitrosourea moiety with a methyl
group attached at one end and a glucose molecule at the other end (Weiss, 1982).
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Fig. 2. Structure of Streptozotocin
Diabetes is induced by single intraperitonial injection of freshly prepared
streptozotocin (55 mg kg-1
bw) in 0.1M citrate buffer (PH-4.5) in a volume of 1 ml
Kg-1
rats (Siddique et al., 1987). In case of mice the dose 175-200 mg kg-1
and Dog
15 mg kg-1
for 3 days). Streptozotocin also induces diabetes in Hamster, monkey and
Guinea pig (Chattopadhyay et al., 1997).
2.6.3. Mechanism of action
STZ is diabetogenic because it selectively destroys the insulin-producing beta
cells by inducing necrosis. It is postulated that the selective beta-cell toxicity of STZ
is related to the glucose moiety in its chemical structure, which enables STZ to enter
the cell via the low affinity glucose transporter GLUT2 in the plasma membrane
(Elsner, 2000). It is generally accepted that the cytotoxicity produced by STZ depends
on DNA alkylation and subsequent activation of poly ADP-ribose synthetase causes
CH2 OH
OH OH
O H
OH
NH
C=0
H
H
N-NO
CH3
H
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rapid and lethal depletion of NAD in pancreatic β-islets, thereby causing cell death
(Fig.3).
Several lines of evidences indicate that free radicals, highly reactive carbonium
radicals originating from the decay of STZ molecules might increase the production
of oxygen free radicals including hydroxyl radicals and nitric oxide, may play an
essential role in the mechanism of β-cell damage and diabetogenic effect of STZ
(Halliwell & Gutteridge, 1990).
2.6.4. Treatment for diabetes mellitus
Diabetes is a multifactorial disease leading to several complications, and
therefore demands a multiple therapeutic approach. Currently available drug regimes
for management of diabetes mellitus have certain drawbacks and therefore, there is a
need for safer and more effective anti-diabetic drugs. Many oral hypoglycaemic
agents, such as biguanides and sulfonylurea are available along with insulin for the
Fig. 3: Mechanism of action of Streptozotocin.
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treatment of diabetes mellitus, but these synthetic agents can produce serious side
effects, and in addition, they are not suitable for use during pregnancy. Even though
several therapies are in use for treatment, there are certain limitations due to high cost
and side effects such as development of hypoglycemia, weight gain, gastrointestinal
disturbances, liver toxicity etc (Dey et al., 2002). Based on recent advances and
involvement of oxidative stress in complicating diabetes mellitus, efforts are on to
find suitable anti-diabetic and antioxidant therapy.
2.6.5. Herbal Support for Diabetes
In the late 1980s, Chinese doctors became alarmed by a huge increase in the
incidence of diabetes in China. Observing that this previously rare condition appeared
to be linked with newly adopted Western eating habits, they initially relied on
Western diabetes drugs to treat what was perceived to be a primarily Western
disorder. Eventually medical experts turned away from this approach after noting that,
in addition to serious side effects, modern pharmaceuticals also failed to address the
underlying cause of the problem. A central tenet of Chinese healing is to treat both
acute symptoms and the underlying cause of an illness. Based on this principle,
leading Chinese researchers turned their attention to traditional herbal remedies used
in China for thousands of years (Richards 2007).
2.7. ANTI-INFLAMMATORY ACTIVITY
Inflammation is the body’s way of dealing with infections and tissue damage,
but there is a fine balance between the beneficial effects of inflammation cascades and
30
their potential for long-term tissue destruction (Simmons, 2006). Studies have been
continuing on anti-inflammatory drugs to treat inflammatory diseases in various
countries. Inflammation can be defined as a generalized, on specific, beneficial
response of tissues to injury. The inflammatory process involves a complex interplay
between cells of the blood, the blood vessels and the cells of the involved tissue. The
process can be seen as a coordinated response of a large number of cells to an initial
stimulus. Early changes in inflammatory tissues are involved in the release of various
biologically active materials from polymorph nuclear leukocytes and lysosomal
enzymes.
The vascular effects are primarily mediated by kinins, prostaglandins and vaso-
active amines (e.g. histamine, released by mast cells), which cause increased vascular
permeability leading to plasma exudation .This complex reaction of inflammation is a
consequence of the materials that release into the extra cellular environment. Such
materials include histotoxic agents such as proteases and oxygen metabolites (Henson
and Murphy, 1989).
The inflammatory reaction is characterized by blush, heat, tumor, pain and lost
function (Dassoler et al., 2004). There are many causes for the inflammations, but the
mechanisms are common to all. In spite of the discovery of several newer agents, the
search for better anti-inflammatory drugs continues because they have many known
side effects and none of them is suitable for prolonged use (Ramprasath et al, 2004).
31
Plant drugs with anti-inflammatory, antioxidant activity can bring relief to
conditions like hemorrhoids, varicose veins and other conditions that involve a better
flow of blood. The anti-inflammatory activities are often attributed to the presence of
saponins while the antioxidant activity attributed to the presence of flavonoids and
other molecules having antioxidant activities (Gurib-Fakim, 2006).
Over the past 20 years there has been a significant increase in knowledge about
immunology, both in terms of molecular targets and molecular mechanisms. The
processes of inflammation fall into four major groups: changes in blood flow caused
by changes in smooth muscle cell function causing vasodilatation, alterations in
vascular permeability by cytoskeletal contraction in endothelial cells, migration of
phagocytic leukocytes to the site of inflammation and phagocytosis (Denko, 1992;
Evans and Whicher, 1992).
Inflammatory disorders can be studied in two different models
(1) Acute inflammatory model
(2) Chronic inflammatory model.
Acute models are designed to test drugs that modulate blood flow (erythema),
changes in vascular permeability, leukocyte migration and chemotaxis, phagocytosis
and other phagocytic cells, measurement of local pain, antipyretic activity, local
analgesic action and rat paw edema. Chronic models are designed to find drugs that
may modulate the disease process and these include sponge and pellet implants and
granuloma pouches which deposit granulation tissue (Lewis, 1989). Different
32
approaches used to analyze the anti-inflammatory potential of plants and plant derived
compounds in the past years.
Desmarchelier et al. (2000) reported that the oral administration of methanol
extract of the leaves of Pothomorphe peltata at a dose of 20 mg/kg exhibited
significant anti-inflammatory activity. The leaves of Bouchea fluminensis contain
iridoid and steroid glycosides that are present in the form of crude triterpene mixture
having anti-inflammatory property and the purified fraction of the plant was found to
contain ursolic, oleanolic and micromeric acids (Costa et al., 2003).
The leaves of Gochnatia polymorpha contain sesquiterpenes and lactone
derivatives and the ethanolic fractions at a dose level of 200 mg/ kg showed
significant inhibition on rat paw edema induced by carregeenan (Moreira et al., 2000).
The leaf extract of Hyptis pectinata administered orally at a dose of 600 mg/kg
exhibited a significant antidermatogenic activity (Bispo et al., 2001). The
intraperitoneal administration of the extract at the dose of 300 mg/kg inhibited the rat
paw edema by 33.8% and the results showed that the aqueous extract of H. pectinata
acted on both the cyclooxygenase and lipoxygenase pathways.
Franzotti et al. (2000) studied the aqueous extract of leaves of Sida cordifolia
to assess the anti-inflammatory properties using the Carrageenan and arachidonic acid
induced rat paw edema model. Oral administration at a dose level of 400 mg/kg of the
extracts howed 28.31% reduction in edema,whereas the dose level of 200 mg/kg was
33
ineffective in reducing edema and the dose of 800 mg/kg inhibited 7.55% in the
model of Carrageenan induced edema.
Bani et al. (2000) studied that the hydrosoluble fraction of Euphorbia royleana
latex was active in suppressing the paw edema in both acute and chronic models of
inflammation and this was further supported by the poor action of the extract in cotton
pellet granuloma test in rats which was found to be highly sensitive to steroidal type
of drugs. Anti-inflammatory activity of the leaves of Dalbergia was investigated by
Hajare et al. (2001) and Carrageenan, kaolin and nystatin were used to induce paw
edema. The 90% ethanolic extract of the plant when administered orally at doses of
100, 300 and 1000 mg/kg showed significant inhibition of inflammation.
Leaves of Cassia fistula were tested for anti-inflammatory effects as compared
with phenylbutazone using carrageen, histamine and dextran induced paw edema in
rats and potent anti-inflammatory activity against all phlogistic agents was noted
(Bhakta et al., 2000). The anti-inflammatory activity of heat treated Cassia alata leaf
extract and kaempferol 3-O-gentiobioside isolated from C. alata as an abundant
flavonoid glycoside were studied by evaluating by their activities with sun dried C.
alata leaf extract. The heat treated C. alata leaf extract exhibited stronger inhibitory
effects than the effects of the sun dried leaf extract (Moriyama et al., 2003).
Devi (2004) evaluated the anti-inflammatory activity of the aqueous extract of
Spilanthes acmella in Carrageenan induced rat paw edema. The aqueous extract in
34
doses of 100, 200 and 400 mg/kg showed 52.6, 54.4 and 56.1% inhibition of paw
edema. Karina et al. (2007) investigated the effect of Brazilian polyherbal formulation
for inflammatory conditions. In their investigation, the formulation at a dose of 26
ml/kg inhibited capsaicin induced ear edema.
Ethanolic extracts of the roots of Cissampelos pareira at the doses of 200, 400
mg/kg exhibited significant anti-inflammatory activity (Amresh, 2007). The methanol
extract of the leaves of Jatropha curcas exhibited significant anti-inflammatory
activity in acute Carrageenan induced rat paw edema and cotton pellet induced
granular tissue formation after oral treatment for 7 days in albino rats (Mujumdar and
Misar, 2004).
The methanol extract of Ionidium suffruticosam was evaluated on Carrageenan,
histamine and serotonin-induced rat hind paw edema acute models (Boominathana et
al., 2004). The extract at the doses of 200 and 400 mg/kg has been found to possess
significant anti-inflammatory activity on the tested experimental models.
The anti-inflammatory activity of the leaves of Cleome gynandra was assessed
by paw volume measurement and the extract showed significant effect on inhibiting
the paw edema (Narendrakannan and Subramanian,2007). Anti-inflammatory activity
of the ethanolic extract of the leaves of Morus indica was studied in wistar rats using
the Carrageenan induced pleurisy and cotton pellet induced granuloma model
(Balasubramanian and Ramalingam, 2005). The extract (100 mg/kg) inhibited
35
Carrageenan induced rat paw edema and it also showed an inhibitory effect on
leukocyte migration and a reduction on the pleural exudates as well as reduction on
the granuloma weight in the cotton pellet granuloma method.
Table 6: Plants with potential anti-inflammatory principles
NAME OF THE
PLANT
ACTIVE
COMPOUND
ANTI-INFLAMMATORY
MECHANISM
REFERENCES
Allium sativum Allicin and its
precursor alliin
S-allyl cysteine
Scavenge hydroxyl
radicals inhibit NFB
activation in T cells
Rabinkov et al.
(1998);
Geng et al (1997)
Artocarpus
heterophyllus
Cycloheterophyllin
artonins
Inhibit lipid peroxidation
and directly scavenge
stable free radicals
Ko et al. (1998)
Ginkgo biloba Flavonoids,
terpenoids,
ginkgolide B
Scavenge superoxide
anions, hydroxyl radicals,
and peroxy radicals
Inhibit the activity of key
enzymes involved in the
inflammatory process
Kobuchi et al.
(1992)
Magnolia obovata Magnolol Inhibits lipid peroxidation
Prevents
ischemicreperfusion
injury
Hong et al.(1996)
Panax ginseng Ginseng Inhibits decomposition of
fatty acids, which are
highly cytotoxic, due to
iron-mediated lipid
peroxidation
Zhang et al.
(1996)
36
Suba et al. (2005) studied the anti-inflammatory efficacy of the methanol
extract of the aerial parts of Barleria lupulina in acute and sub-acute inflammatory
models of albino rats. In all the tested dose levels, methanol extract exhibited
significant inhibition of Carrageenan and serotonin induced paw edema volumes. The
analgesic and anti-inflammatory activities of a new alkaloid (5′-Hydroxymethyl-1′-(1,
2, 3, 9-tetrahydro-pyrrole [2, 1-b] quinazolin-1-yl)-heptan-1-one) isolated from Sida
cordifolia was produced significant (p<0.01) analgesic activity in animal models
(Sutradhar et al., 2007).
Emmanuel et al. (2006) studied the anti-inflammatory effect of solasodine,
sobatum and methanol extract of Solanum trilobatum and the results showed
significant anti-inflammatory activity. Ratheesh and Helen (2007) carried out anti-
inflammatory tests in Wistar male rats using aqueous, ethanolic and methanolic
extracts of Ruta graveolens. These extracts were administered in the concentrations of
20 and 50 mg/kg b.wt before Carrageenan injection. Methanolic extracts of R.
graveolens with a concentration of 20 mg/kg b.w. and ethanolic extract with a
concentration of 50 mg/kg b.wt showed maximum (90.9%) inhibition on Carrageenan
induced rat paw edema. Biren et al. (2007) studied the ethanolic extract of the leaves
of Colocasia esculenta in Carrageenan induced rat paw edema and the extract
produced significant (p<0.05) anti-inflammatory activity (100 mg/kg, p.o.) induced by
Carrageenan.
37
Yam et al. (2008) reported that the methanol: water (50:50 vol/vol) extracts of
the leaves of Orthosiphon stamineus significantly reduced the hind paw edema in rats
at 3 and 5 hours after Carrageenan administration at the doses of 500 and 1,000
mg/kg. The aqueous extract of the stems of Gynandropsis pentaphylla showed
significant dose dependent anti inflammatory activity at the doses of 100 mg/kg i.p
(Mule et al., 2008).
Lee et al. (2008) studied the effect of ethanol extract of Asparagus
cochinchinensis on skin inflammation in mice. The extract significantly inhibited the
ear edema of mouse at the doses of 200 mg/kg. Oral administration of the ethanolic
extract (200 and 400 mg/kg.) and its fractions (200 mg/kg each) of the aerial parts of
Cleome rutidosperma showed significant analgesic activity in acetic acid induced
writhing and tail immersion tests, anti-inflammatory effect against Carrageenan
induced inflammation (Bose et al., 2007). Anti-inflammatory activity of ethanol
extract of Dalbergia lanceolaria exhibited significant systemic anti-inflammatory
activity in Carrageenan induced rat paw edema by inhibition of histamine and
prostaglandin phases of acute inflammation (Kale et al., 2007).
Methanol extract of Phyllanthus amarus on different phases of inflammation
were examined using different phlogistic agents induced paw edema, Carrageenan
induced air-pouch inflammation and cotton pellet granuloma in rats. Methanol extract
of Phyllanthus amarus significantly inhibited Carrageenan, bradykinin, serotonin and
prostaglandin E1-induced paw edema (Mahat and Patil, 2007). Anti-inflammatory
38
effect of the extracts and purified lignans obtained from the plant, Phyllanthus amarus
inhibited the Carrageenan induced paw edema (Kassuya et al., 2005).
2.8. ANTIPYRETIC ACTIVITY
In the traditional system of medicine, many medicinal plants have been
reported to possess the potentiality to cure fever. In the Chinese medicine system the
antipyretic plants are of five types (Dong et al., 1998).
a) the herbs that clear intense internal heat
b) the herbs that clear intense heat in the blood systems
c) the herbs that reduce dry internal dampness or heat in the viscera or bowels
d) the herbs that reduce internal noxious heat or toxins
e) the herbs that clear interior heat of the deficient type
The ethanolic extracts of Ailanthus excelsa, Toddalia asiatica and Araucaria
bidwilli showed moderate to significant degree of antipyretic activity in an
experimental rat model of 20% yeast suspension induced hyperthermia (Suresh et al.,
1995). Methanolic extract of the rhizome of Nelumbo nucifera produced a significant
dose dependent lowering of pyretic effect in pyretic rats (Mukherjee et al., 1996).
Rhynchosia cana showed significant antipyretic activity in rats (Vimala et al., 1997).
Alcoholic extract of the roots of Clerodendron serratum showed significant
antipyretic activity and result was found to be almost equal to paracetamol
(Narayanan et al., 1999). Panax ginseng showed hyperthermic effect and attenuated
39
the hypothermic response of reserpine and 5-HTP induced hyperthermia in animals
(Mitra et al., 1996). Significant antipyretic activity was observed in hexane,
chloroform and water soluble extracts of Artemisia absinthium, Viola odorata, Melia
azadirachta and Fumaria parviflora and comparable in potency aspirin. The Pyresis
was induced by subcutaneous yeast injections and the antipyretic activity was more
prominent in hexane soluble portions (Khattak et al., 1985). Some important
medicinal plants with antipyretic activity are listed in Table 7.
Table 7: Some important medicinal plants with Anti-pyretic activity
Botanical Name Parts used References
Adansonia digitata Fruit pulp Ramadan et al. (1994)
Ageratum conyzoides Essential oil Abena et al. (1996)
Astragalus siculus Whole plant Bisignano et al. (1994)
Buddleia cordata Leaves Martinez-Vazquez et al. (1996)
Euphorbia hirta Whole plant Lanhers et al. (1991)
Lawsonia inermis Leaves Ali et al. (1995)
Maytenus boaria Leaves Backhouse et al. (1994)
Michelia champaca Leaves Vimala et al. (1997)
40
Oral administration of Taxus wallichiana produced significant antipyretic
activity in yeast induced pyrexia model at a dose of 200 mg/kg dose (Nisar et al.,
2008). Amiruddin et al. (2008) investigated the leaves of Dicranopteris linearis for its
antinociceptive, anti-inflammatory and antipyretic properties in experimental animals.
The antipyretic activity was measured using brewer’s yeast-induced pyrexia. The
aqueous extract of Dicranopteris linearis was also found to have significant
antipyretic activity.
Salawu et al. (2008) evaluated the methanolic extract of Crossopteryx
febrifuga for analgesic, anti-inflammatory, antipyretic and antiplasmodial activities in
rodents. The extract significantly diminished antipyretic activity in mice and rats in a
dose-related manner. In another investigation it was observed that oral administration
of the ethanolic extract (200 and 400 mg/kg, p.o) and its fractions (200 mg/kg each)
of the aerial parts of Cleome rutidosperma produced significant antipyretic activity
against yeast induced pyrexia (Bose et al., 2007).
Mucuna pruriens Seeds Jauk et al. (1993)
Ocimum sanctum Leaves Godhwani et al. (1987)
Premna herbacea Root Narayanan et al. (2000)
Solanum ligustrinum Leaves Delporte et al. (1998)
Teclea nobilis Whole plant Mascolo et al. (1998)
41
Garcinia hanburyi was assessed for its antipyretic activity by Panthong et al.
(2007) using experimental animal model and they observed that G. hanburyi
possessed significant antipyretic effect when tested in yeast induced hyperthermic
rats. Oral administration of the aqueous extract of the stem of Urtica macrorrhiza
(200–400 mg/kg) was dose dependently suppressed the yeast induced fever in rats
(Yongna et al., 2005).
The antipyretic activity of methanol extract of Cleome viscosa was investigated
by Devi et al. (2003) for its activity on normal body temperature and yeast induced
pyrexia in albino rats. The extract at doses of 200, 300 and 400 mg/kg b.wt showed
significant reduction in normal body temperature and yeast provoked elevated
temperature in a dose dependent manner. Chattopadhyaya et al. (2002) were studied
the antipyretic potential of the methanol extract of Mallotus peltatus on normal body
temperature and yeast induced pyrexia in Wister albino rats. The leaf extract at oral
doses of 100, 200 and 300 mg/ kg showed significant reduction in normal body
temperature and yeast provoked elevated temperature in a dose dependent manner.
Archana et al. (2005) evaluated the antipyretic and analgesic activities of
ethanolic extract of the seed kernel of Caesalpinia bonducella in adult albino rats at a
dose of 100 and 300 mg/kg administered orally. They observed that antipyretic
activity against Brewer's yeast induced pyrexia in rats. The ethanol (100 and
200 mg/kg) and water extracts (200 mg/kg) of Capparis zeylanica leaves showed dose
dependent analgesic and antipyretic activities (Ghule et al., 2007).
42
The chloroform, methanolic and ether extracts of the leaves of Vernonia
cinerea at the doses of 100, 200 and 400 mg/kg intraperitoneally significantly
suppressed the brewer’s yeast induced pyrexia in rats (P<0.05) compared to the
control (Iwalewa et al., 2003). Ahmadiani et al. (2001) examined the Trigonella
foenum-graecum leaves extract for its antipyretic activity. In their study, hyperthermia
was induced by intraperitoneal injection of 20% (w/v) aqueous suspension of brewer's
yeast. Leaf extract of T. foenum-graecum significantly reduced the hyperthermia
induced by brewer's yeast in 1 and 2 hrs after their administration.