As a part of curriculum of
BECHELOR OF SCIENCE
IN
BIOTECHNOLOGY
(2008-09)
by
SUMIT KUMAR
B.Sc IInd Yr. Integrated
Department of Biotechnology
Mahatma Gandhi Institute of applied Sciences, Jaipur
CERTIFICATE
This is to certify that Mr. Sumit Kumar S/O Mr. Shankar lal student of
B.Sc. part II (Biotechnology Integrated) has undergone 8 weeks training
under the supervision of Dr. Ekta Menghani at Mahatama Gandhi
Institute of Applied sciences, Jaipur. The student has completed his work
in Biotechnology laboratory and undertaken the training as per the
curriculum of University of Rajasthan, Jaipur.
Place: Jaipur
Date:
UNDERTAKING
Sumit Kumar
B.Sc. Part-II Biotechnology(Int.)
CONTENTS
4. CONCLUSION
1.Introduction:
Guduchi (Tinospora cordifolia (Willd.) Miers ex Hook. F. & Thoms) is a large
glabrous, deciduous climbing shrub belonging to the family Menispermaceae. It is
distributed throughout tropical Indian subcontinent and China, ascending to an altitude of
300m. In Hindi the plant is commonly known as Giloya, which is a Hindu mythological
term that refers to the heavenly elixir that have saved celestial being from old age and
kept them eternally young. Guduchi is one of the most versatile rejuvenate herbs. It
works on all the tissue elements in the body. The Sanskrit name guduchi means the one,
which protects the body. It is also called as amrta or nectar, as it is extremely useful in
strengthening the immune system of the body and keeping the functions of its various
organs in harmony. It possesses various synonyms like jvaranasi febrifuge, vayastha –
promotes longevity, rasayan- a rejuvenative, tikta – bitter etc. Maharsi Caraka has
categorized it as stanya sodhana – relieve burning sensation of the skin, trptighna – anti
saturative, vayah sthapana – promote longevity and as medhya nervine tonic. Susurta has
mentioned it as sukra sodhana – sperm purifier. Sarngadhara has classified it as
samsamana – pacifier and rasayana – a rejuvenator. It possesses properties like jvaraghna
– antipyretic, kamala nasaka – cures jaundice, amavataghna – mitigates gout, gandamala
nasaka – alleviates glandular swellings. Guduchi is recommended as a drug of the choice
in the treatment of gout.
Different constituents that it includes are bitter principles. A glucoside, alkaloide etc. The
glycoside – giloin and a non-glucoside – gilenin and gilosterol have been found. The
alkaloid tinosporin, tinosporic acid and tinosporol have been identified in the leaves.
Tinosporidine and sitosterol isolated from stem, cordifol, heptacosanol and octacosonal
from leaves a new furanoid diterpene – tinosporide – isolated from stems and its structure
determined.
Guduchi is bitter, pungent and astringent in taste, sweet in the post digestive effect and
hot in potency. It alleviates all the dosas. It possesses light and oily attributes. It has a
special potency as an anti-toxin.
The root, stem, leaves and satva (starch) of guduchi are used for medicinal purpose,
externally; the medicated oil of the plant is effectively used to reduce the pain and
oedema, in gout and skin diseases. In filariasis, the paste of guduci, karuka, sunthi,
devadara and vidanga works well when applied externally.
Internally, guduchi is one of the most effective rasayanas- rejuvenatives. It works well on
all the seven dhatus tissues and keeps the systems in balance. The rasayana accords
longevity, enhances memory, improves health, and bestows youth, betters complexion,
voice, energy and luster of the skin. In the diseases due to vata dosa it is given with ghrta,
in pitta dosa with sugar and kapha dosa with honey. It is immensely helpful in the
digestive ailments like hyperacidity, colitis, worm infestations, and loss of appetite,
abdominal pain, excessive thirst, vomiting and liver disorders like hepatitis.
Guduchi is one of the best bitter tonics useful in fevers especially of pitta origin. It
alleviates body heat, thirst, burning sensation to the skin and vomiting if any, due to pitta.
In tubercular fever, it combines well with ativisa for decoction. The juices of guduchi,
amalaki and haridra act synergistically in urinary problems. In hepatitis, the fresh juice of
guduchi given with rock candy, hastens the recovery. It particularly helps in diseases like
raktapitta, anaemia, cardiac debility, diabetes, sexual debility and splenic disorders, due
to vitiation of pitta.
Guduchi is the drug of choice amongst all the remedies in treating gout vatarakta. The
decoction of guduchi and sunthi is a very effective combination for the treatment of gout
and rheumatic disorders. Its medicinal ghee with kantakari is beneficial in cough Guduchi
juice works well with cow’s milk or lodhra in leucorrhea and with cumin seeds in
burning sensation due to pitta.
The starch (sattva) of guduchi is traditionally used as a household remedy, for chronic
fever, to alleviate it as well as to reduce burning sensation and to increase the energy and
appetite. It is beneficial in tuberculosis and general debility also. Guduchi is
recommended in the skin diseases and its decoction with nimba and vasa effectively
relieves the itching and oozing. It also works well in the cutaneous rashes and
condylomata, in the secondary stage of syphilis. Guduchi should be always used fresh for
good results and the twiner which grows on nimba tree is said to have better results.
So in the present project attempts have been made to made a complete study on Guduchi
Satva where its authentication and validation was worked out in a series of manner. In
standardization of Guduchi satva the extract of the Tinospora cordifolia stem were
extracted from authenticated stem and simultaneously Market sample of Giloy satva were
also performed and the characterization of authentic and market sample were performed
and results were discussed. The validation of guduchi satva and gilov market samples were
simultaneously screened and the results proved that this can be used as a Monograph for
any authentic Validate Pharmacopoeia where the various thin layer chromatograms,
bioefficacies results and microscopic structures will be aid as an identification tools. These
characterizations will be further enhance the quality of the market sample of giloy and their
justification as the potent use as antidiabetic agent even their use as anticancerous agent.
Tinospora cordifolia
Scientific classification
Kingdom Planti
Division: Magnoliophyta
Class: Magnoliopsida
Order: Ranunculales
Family: Menispermaceae
Tinospora cordifolia
Genus: Tinospora
Species: Cordifolia
CONSTITUENTS
The active adaptogenic constituents are diterpene compounds including tinosporone,
tinosporic acid, cordifolisides A to E, syringen, the yellow alkaloid, berberine, Giloin,
crude Giloininand, a glucosidal bitter principle as well as polysaccharides, including
arabinogalactan polysaccharide (TSP).
Its principal constituents are tinosporine, tinosporide, tinosporaside, cordifolide, cordifol,
heptacosanol, clerodane furano diterpene, diterpenoid furanolactone tinosporidine,
columbin and b-sitosterol.
A verity of constituents have been isolated from Tinospora cordifolia plant and their
structures were elucidated. They belong to different classes such as alkaloids, diterpenoid
lactones, glycosides, steroids, sesquiterpenoid, phenolics, aliphatic compounds and
polysaccharides.
Leaves of this plant are rich in protein (11.2%) and are fairly rich in calcium and
phosphorus. Studies on the physical characteristic and chemical composition of starch
obtained from Guduchi Satwa (extract) were carried out and the polysaccharide was
found to consist chiefly of 1g4 linked glucan with occasionally branched points. Its
similarities and differences from amylase were elucidated. An arabinogalactan had been
isolated from the dried stems of T. cordifolia .
PARTS USED
Roots, Stem, Leaves and satva (starch)
• Guduchi acts as a diuretic and found to be effective against Renal obstruction like
calculi and other urinary disorders.
• Guduchi acts as a memory booster, develops inteligence, promotes mental clarity.
It is described as one of the Medhya Rasayana (mental rejuvenative) in the
Charak Samhita (The oldest and most potent book of Ayurvedic Medicine).
• Guduchi is regarded as a liver protector.
• Guduchi is considered helpful in eye disorders as a tissue builder and promotes
mental clarity.
• The stem of guduchi is used in general debility, dyspepsia and urinary diseases.
• Guduchi is anti-pyretic and act as a tonic after fever, also has action against
alternative fever like Malaria.
According to the 1918 United States Dispensatory edited by Joseph Remington, Horatio
Wood et al.:
Tinospora. Br. Add. 1900.—"The dried stem of Tinospora cordifolia Miers (Fam.
Menispermaceae), collected in the hot season." Br. Add., 1900. Tinospora has long been
used in India as a medicine and in the preparation of a starch known as gilae-ka-sat or as
palo. It is said to be a tonic, antiperiodic, and a diuretic. Flückiger obtained from it traces
of an alkaloid and a bitter glucoside. The Br. Add., 1900, recognized an infusion
(Infusum Tinosporae Br. Add., 1900, two ounces to the pint), dose one-half to one
fluidounce (15-30 mils); a tincture (Tinctura Tinosporae Br. Add., 1900, four ounces to
the pint), dose, one-half to one fluidrachm (1.8-3.75 mils); and a concentrated solution
[Liquor Tinosporae Concentratus Br. Add., 1900), dose, one-half to one fluidrachm (1.8-
3.75 mils). Tinospora crispa Miers (more), which is abundant in the Philippines, is used
freely by the natives under the name of makabuhay (that is, "You may live"), as a
panacea, especially valuable in general debility, in chronic rheumatism, and in malarial
fevers. It may be prepared in the same way and given in the same doses as Tinospora
cordifolia.
Tinospora cordifolia is used in Ayurvedic herbal medicine as a hepatoprotectant,
protecting the liver from damage that may occur following exposure to toxins. Recent
research has demonstrated that a combination of T. cordifolia extract and turmeric extract
is effective in preventing the hepatotoxicity which is otherwise produced as a side effect
of conventional pharmaceutical treatments for tuberculosis using drugs such as isoniazid
and rifampicin.
The stem of Tinospora cordifolia is one of the constituents of several ayurvedic
preparations used in general debility, dyspepsia, fever and urinary disease. The stem is
bitter, stomachic, diuretic, stimulates bile secretion, causes constipation, allays thirst,
burning sensation, vomiting, enriches the blood and cures jaundice. The extract of its
stem is useful in skin diseases. The root and stem of T. cordifolia are prescribed in
combination with other drugs as an anti-dote to snake bite and scorpion sting. Dry barks
of T. cordifolia has anti-spasmodic, antipyretic, anti-allergic, anti-inflammatory and anti-
leprotic properties.
The aqueous extract of the stem antagonizes the effect of agonists such as 5-
hydroxytriptamine, histamine, bradykinine and prostaglandins E1 and E2 on the rabbit
smooth muscle, relaxes the intestinal, uterine smooth muscle and inhibits the constrictor
response of histamine and acetylcholine on smooth muscle. In travenous exposure to
aqueous extract of T. cordifolia in doses of 5.0, 10.0 and 15.0 mg/kg body weight
produces a temporary but marked fall in blood pressure and bradycardia in anaes-thetized
dogs.
It is reported that the daily administration of either alcoholic or aqueous extract of
T. cordifolia decreases the blood glucose level and increases glucose tolerance in rodents.
Aqueous extract also caused a reduction in blood sugar in alloxan induced hyperglycemia
in rats and rabbits in the dose of 400 mg/kg. However, histological examination of
pancreas has not revealed any evidence of regeneration of b-cells of islets of Langerhans
and the possible mode of action of the plant is through glucose metabolism. The aqueous
extract has also exhibited some inhibitory effect on adrenaline-induced hyperglycemia.
Ethyl acetate extract of its roots has afforded a pyrrolidine derivative with hypoglycemic
activity in rabbits. Another study has also revealed significant hypoglycemic effect of
extract of leaves in normal and alloxan diabetic rabbits. However, the extract had no
significant effect on total lipid levels in normal or treated rabbits.
T. cordifolia is reported to benefit the immune system in a variety of ways. The alcoholic
and aqueous extracts of T. cordifolia have been tested successfully for immuno-
modulatory activity. Pre-treatment with T. cordifolia was to impart protection against
mortality induced by intra-abdominal sepsis following coecal ligation in rats. It has also
significantly reduced the mortality from E. coli induced peritonitis in mice. In a clinical
study, it has afforded protection in cholestatic patients against E. coli infection. These
activities are not due to its anti-bacterial activity as shown by the negative in-vitro anti-
bacterial activity of the plant extract. It is reported that the treatment in rats had resulted
in significant leucocytosis and predominant neutrophilia. It has been also observed that it
stimulates the macrophages as evidenced by an increase in the number and %
phagocytosis of S.aureaus by peritoneal macrophages in rats. Other workers have also
supported these observations. The phagocytic and Intra-cellular killing capacity of
polymorphs in rats, tested at 3.5 h after E. coli infection were significant.
The anti-stress and tonic property of the plant was clinically tested and it was found that
it brought about good response in children with moderate degree of behaviour Disorders
and mental deficit. It has also significantly improved the I.Q. levels.
The hepatoprotective action of T. cordifolia was reported in one of the experiment in
which goats treated with T. cordifolia have shown significant clinical and hemato-
biochemical improvement in CCl4 induced hepatopathy. Extract of T. cordifolia has also
exhibited in vitro inactivating property against Hepatitis B and E surface antigen.
The aqueous extract of T. cordifolia exerted a significant anti-inflammatory effect on
cotton pellet granuloma and formalin induced arthritis models. Its effect was comparable
with Indomethacin and its mode of action appeared to resemble that of a non-steroidal
anti-inflammatory agent. The dried stem of T. cordifolia produced significant anti-
inflammatory effect in both acute and subacute models of inflammation. T. cordifolia was
found to be more effective than acetylsalicylic acid in acute inflammation. But in
subacute inflammation, the drug was inferior to phenylbutazone. In a clinical evaluation,
a compound preparation 'Rumalaya' containing T. cordifolia was reported to significantly
reduce the pain in patients suffering from rheumatoid arthritis.
The aqueous extract of roots of T. cordifolia has shown the anti-oxidant action in alloxan
diabetes rats. The administration of the extract of T. cordifolia roots (2.5, 50 mg/kg body
weight) for 6 weeks resulted in a significant reduction of serum and tissue cholesterol,
phospholipids and free fatty acids in alloxan diabetic rats.
Jagetia et al., have found that guduchi killed the HeLa cells very effectively in vitro and
thus it indicates that guduchi needs attention as an anti-neoplastic agent. In this study
exposure of HeLa cells to 0, 5, 10, 25, 50 and 100 mg/ml of guduchi extract (methanol,
aqueous and methylene chloride) resulted in a dose dependent but significant increase in
cell killing when compared to non drug treated controls.
Ether extract of the stem distillate of aerial part of cordifolia has inhibited the in vitro
growth Mycobacterium tuberculosis at 1:50,000 dilution. Ethanolic extract has exhibited
significant antipyretic activity in experimental rats. 'Septilin' syrup, compound
preparation containing T. cordifolia (7.82% in 5 ml of syrup) was found to elicit good
clinical response in children suffering from upper respiratory tract infection and chronic
otitis media. The Ayurveda literature reports that it can cause constipation, if taken
regularly in high doses; it has no side effect and toxicity. Yet the safety and the potential
indications in human beings have to established using modern methods.
PHARMACOLOGICAL ACTION
Cholagogue, detoxicant, immune modulator, anti-inflammatory, diuretic, anthelminthic,
nervine tonic. The aqueous extract of guduchi stem has shown the presence of
arabinogalactan that showed immunological activity. The stem is used in dyspepsia,
fevers and urinary diseases. The bitter principle present shows antiperiodic,
antispasmodic, anti-inflammatory and antipyretic properties. Guduchi has been reported
to treat throat cancer in humans. Dichloromethane extract is the most promising one as
far as cytotoxic effect is concerned. These preliminary studies were done to establish the
cytotoxic effect of guduchi at higher doses. Extracts of Tinospora cordifolia (TCE) have
been shown to possess anti-tumor properties and was shown to upregulate antitumor
activity of tumor-associated macrophages (TAM). Evidence has shown that an alcoholic
extract of Tinospora cordifolia (ALTC) enhances the differentiation of TAM to dendritic
cells (DC) in response to granulocyte/macrophage-colony-stimulating factor, interleukin-
4, and tumour necrosis factor.
INDICATION
Liver: Liver damage, viral hepatitis or toxicity from alcohol, chemicals and medicinal
drugs. Useful in repairing fibrosis and regenerating liver tissue. Applied in all conditions
of aggravated ranjakapitta and pitta in the blood. Joints: Gout (vatarakta), arthritis
(amavata) and other inflammatory joint conditions. It acts by clearing pitta toxins and
uric acid via the urinary system that have accumulated in raktavahasrotas. It also removes
ama toxins from the system without destabilising any of the other dosha.
Immunity: All auto-immune diseases causing inflammation. Applicable in degenerative
diseases such as cancer, AIDS and arthritis as it boosts the immune system. Use to offset
the ulcerative and toxic effects of chemo-radiotherapy.
Skin: Supperative and inflammatory skin conditions such as eczema, psoriasis, Systemic
Lupus Erythmatosus. Useful when there is high tejas and pitta that has burnt immune
protecting ojas away resulting in inflammatory skin conditions. Skin problems from
excessive alcohol, recreational drug and pharmaceutical drug use may indicate the use of
Guduchi. Specific for burning sensations on the skin (daha).
GIT: Guduchi heals a bowel affected with constipation, intestinal bleeding, haemorrhoids
or dysentery. Useful at redressing intestinal floral imbalance with candida-like symptoms
(krimi, grahani) such as bloating, flatulence and malabsorption. Its bitter yet heating
qualities are used to stimulate raktadhatvagni and strengthen digestion in pitta types.
Metabolic: It regulates blood sugar levels via its direct effect on rakta and medas-dhatu
thus benefiting diabetes and hypoglycaemia. Guduchi is very calming to vata and the
nervous system via its unctuous nature soothing nervous irritation.
Reproductive: Its ability to clear heat is applied when sexual dysfunction is caused by a
hyper-heat condition. It is often used in formulas for male sexual dysfunction caused by
pitta imbalance as its sweet post-digestive effect nourishes shukradhatu.
Rejuvenating Properties
The term "Rasayana" accords to longevity, enhances memory, better complexion, voice
energy and luster of skin thus bestows youth.
Anupana (Carrier of Drug) Internally it can be given with Ghee, Sugar and honey in
Vata, Pitta and Kapha Dosha respectively.( Nayampalli S, Ainapure SS, Nadkarni PM,
1982).
REFERENCE
1. Anonymous. Wealth of India: Raw materials, Vol X. New Delhi: CSIR; 1976.
2 . Nadkarni KM, Nadkarni AK, editors. Indian Materia Medica, Vol 1. 3rd ed. Mumbai:
M/S Popular Prakasan Pvt. Ltd;1976.
3. Kirtikar KR, Basu BD, editors. Indian Medicinal Plants, Vol 1. 2nd ed. New
Connaught Place, Dehra Dun: M/S Bishen Singh, Mahendra Pal Singh; 1975.
4. Chopra RN, Nayar SL, Chopra IC, editors. Glossary of Indian Medicinal plants. New
Delhi: CSIR; 1956.
5. Chopra RN, Chopra LC, Handa KD, Kapur LD, editors. Indigenous Drugs of India.
2nd ed. Kolkata: M/S Dhar VN & Sons; 1982.
6 Zhao TF, Wang X, Rimando AM, Che C. Folkloric medicinal plants: Tinospora
sagittata var. cravaniana and Mahonia bealei. Planta Med 1991;57:505.
7. Nayampalli S, Ainapure SS, Nadkarni PM. Study of antiallergic acid Bronchodilator
effects of Tinospora cordifolia. Indian J Pharm 1982;14:64-6.
2.1. Collection:
2.2. Identification:
All the samples were authenticated and were given identification number. The
identification were as follows:
• Tinospora cordifolia(Powdered Stem)
• Giloy satva (White stem extract)
These samples were authenticated and submitted in Department of Biotechnology,
MGiaS, Jaipur (Rajasthan).
I. Microscopic methods
tissues, cells and cell contents by microscope. Methods of preparing specimens of crude
materials for microscopical studies vary depending on the part used like leaf, stem, root,
bark, flower, fruit and also on the nature of the material i.e. entire, cut or powdered.
Preparation of sections: Drugs which are hard to cut, are boiled for 20 to 30 min in
water. Cross section or transverse section with a razor blade was taken. Thin materials
such as leaves, slender stems or flat seeds are placed in a potato slit and sections were
taken with ordinary blade. The section was placed on a slide, clear with chloral hydrates,
Powder microscopy
Likewise, powder of the selected species and their adulterants were subjected to
leaves) in chloral hydrate on a slide was warmed over a low flame or on a hot-plate for a
short time, covered it with a cover glass and observed under microscope (4, 10, 20 or 45
X). Disintegration of hard and woody tissues: The material was cut into pieces and few
pieces transferred to a test tube containing 4 ml of nitric acid and heated to boiling. Later,
powdered potassium chlorate was gently added, warmed and allowed to react leading to
disintegration of the tissue. When completely bleached, pressure was applied with glass
rod for complete disintegration of the tissues. The material was allowed to settle down,
decant the liquid and the settled material washed repeatedly with water until the acidity is
removed. The material was transferred on to a slide, a drop of glycerol added, covered
Microscopic test
1 Starch – For examining the presence of starch, the specimen was taken in I2
3 Fixed oil – For examining the presence of fixed oil, specimen was stained
with sudan red resulting in the droplet of fixed oil to become pink coloured.
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Fig E: Antibacterial activity of Tinospora cordifolia against E. coli
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Fig L: Antibacterial activity of Giloy satva against P. aeruoginosa
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4.Conclusions:
Standardization of ayurvedic drugs and plant materials is the need of the day. Several
pharmacopoeias containing monographs on plant materials describe only the physico-
chemical parameters. Hence, modern methods describing the identification and
quantification of active compounds in the plant material may be useful for proper
standardization of herbs and their formulation. Tinospora cordifolia (Menispermaceae) is
one such plant which is widely used in indigenous system of medicine. The stems have
been used for centuries, in ayurvedic preparations for the treatment of jaundice, diabetics,
skin diseases and anemia. The plant has been shown to possess anti-inflammatory and
anti-allergic properties. More recently, the immunomodulatory properties , antioxidant
and antineoplastic activities have been reported. Among the complex mixture of
biologically active compounds in the plant, microscopic identification, TLC, and bio
efficacies can be used as an analytical characterization to determine the quality of plant
material of different sources. It is very surprising that the simple tests which are not
useful in identification sometimes become very beneficial for the characterization and
validation of important Ayurvedic drugs. Hence, the microscopic characterization is
simply a authentic and easy tool for validation of Guduchi satva. The use of safe drug is
the motto of our healthy being so if we use the correct parameters for the identification of
the drug it will be very useful for Ayurvedic drugs to not only enhance their quality and
efficacy but also their therapeutic potentials as drugs.