Abhi

IN-VITRO ANTI-HELMENTHIC ACTIVITY OF ETHANOLIC LEAF EXTRACT OF
URTICA DIOICA
Dissertation submitted to the
Jawaharlal Nehru Technological University Kakinada (JNTUK)

In partial fulfillment of the requirement for the award of the Degree of
BACHELOR OF PHARMACY
Submitted By
V. LAVANYA (208N1R0003) SK. SHARMILA (208N1R0004)
D. ABHISHEK (208N1R0026) G. YUVARAJU (208N1R0030)
M. VIMALA JYOTHI (208N1R0053) N. DIVYA MOUNIKA (208N1R0059)
P. RAMU (208N1R0062) P. GAYATRI (208N1R0070)
UNDER THE GUIDANCE OF

KANIKELLA. SIVAJI, M.Pharm.
Professor – Department of Pharmacology
A.K.R.G. COLLEGE OF PHARMACY
Approved by PCI, New Delhi

Affiliated to JNTU-K & Recognized by APSCHE, AP
NALLAJERLA-534112, East Godavari Dist., Andhra Pradesh, India.
2024
DECLARATION BY THE CANDIDATE(S)
We hereby declare that the dissertation entitled “IN-VITRO ANTI-HELMENTHIC ACTIVITY OF

ETHANOLIC LEAF EXTRACT OF URTICA DIOICA” is a bonafied and genuine research work
carried out by us under the guidance of KANIKELLA. SIVAJI, M.Pharm. Professor, Department of
Pharmaceutical Chemistry, A.K.R.G. College of Pharmacy.

Date: /04/2024
Place: Nallajerla
2024
CERTIFICATE BY THE GUIDE
This is certify that the dissertation entitled “IN-VITRO ANTI-HELMENTHIC ACTIVITY OF
ETHANOLIC LEAF EXTRACT OF URTICA DIOICA” is a bonifide research work done by V. Lavanya
(208N1R0003), SK. Sharmila (208N1R0004), D. Abhishek (208N1R0026), G. Yuvaraju (208N1R0030), M. Vimala
jyothi (208N1R0053), N. Divya mounika (208N1R0059), P. Ramu (208N1R0062), P. Gayatri (208N1R0070) in the
partial fulfillment of the requirement for the degree of “BACHELOR OF PHARMACY”
Date: Signature of the Research guide

Place: Nallajerla KANIKELLA. SIVAJI M.Pharm.
Professor
Department of Pharmacology
2024
CERTIFICATE BY THE PRINCIPAL
This is certify that the dissertation entitled “IN-VITRO ANTI-HELMENTHIC ACTIVITY OF
ETHANOLIC LEAF EXTRACT OF URTICA DIOICA” is a bonafide research work done by V. Lavanya
(208N1R0003), SK. Sharmila (208N1R0004), D. Abhishek (208N1R0026), G. Yuvaraju (208N1R0030), M. Vimala
jyothi (208N1R0053), N. Divya mounika (208N1R0059), P. Ramu (208N1R0062), P. Gayatri (208N1R0070) under
the guidance of KANIKELLA. SIVAJI, M.Pharm Professor, Department of Pharmaceutical Chemistry,
Raghavendra Institute of Pharmaceutical Education and Research (RIPER) in partial fulfilment of the
Requirement for the degree of “BACHELOR OF PHARMACY”.
Date: Signature of the Principal
Place: Nallajerla Dr. Pemmadi Raghuveer Varma M.Pharm., Ph.D.

Principal
A.K.R.G. College of Pharmacy
ACKNOWLEDGEMENTS
First of all, I would like to thank God for showering his blessings on me throughout my life and giving me
strength for completing my research work.
I am indebted to Shri Chava Gokul, Chairman of A.K.R.G Educational Institutions for encouraging us through
providing the necessary facilities and infrastructure.
I would like to offer my sincere gratitude to my research supervisor, mentor and guru Dr.
Raghuveer Varma Pemmadi, Principal & Professor, A.K.R.G College of Pharmacy whose guidance has shown
me new dimensions in medicinal chemistry and drug design. His excellent vision in this field and the optimistic
nature towards life has helped me in many situations.
I owe my sincere regards to our teaching staff Dr.KLN Mallikarjuna Rao, Dr.N.Srinivas, Mr.N.Saikrishna,
MRS.T.Anjal, Mrs.B.DevakiDevi, Mrs.N.Priyanka, who bestowed us with knowledge and wisdom on various
concepts of Pharmacy.
I owe my special thanks to our non-teaching staff f Mrs.S. Aruna Lakshmi, , Mr.J.Hanumanth, and Librarian
Mr.Amar for their continuous assistance and timely support throughout our work.
I thank my classmates for their unending support, love and care who made this 4 years enjoyable and
memorable and cherishable.
My personal acknowledgements are due to my mom & dad and other family members for their unconditional
love, invaluable affection and care they showered on me.

Dedicated to
My beloved parents &
Guide
TABLE OF CONTENTS
S.NO TITTLE NAME PAGE

NO
1 INTRODUCTION 1-15
16-19
2 REVIEW OF LITERATURE
20
3 AIM & OBJECTIVE
21-29
4 EXPERIMENTAL DESIGN
30-32
5 RESULTS
33-34
6 DISCUSSIONS
35
7 CONCLUSIONS
36-38
8 BIBILOGRAPHY
LIST OF TABLES
S.N TITTLE NAME PAGE NO

O
1.1 LIST OF ANTHELMINTIC DRUGS
1.2 TEST OF ALKALOIDS 22
1.3 TEST FOR CARBOHYDRATES 23
1.4 TEST FOR CARDIAC GLYCOSIDES 24
1.5 TEST FOR ANTHRAQUINONE 24
1.6 TEST FOR GUMS AND MUCILAGE 24
1.7 TEST FOR PROTEINS AND AMINO 25
ACIDS
1.8 TEST FOR TANNINS AND PHENOLIC 26
COMPOUNDS
1.9 TEST FOR FLAVONOIDS 26
2.0 TEST FOR STEROIDS AND STEROLS 27
2.1 TEST FOR TRITERPENOIDS 27
2.2 TEST FOR SAPONINS 28
2.3 TREATMENT SCHEDULE 29
2.4 PHYTOCHEMICAL SCREENING 30
2.5 EVALUATION OF ANTHELMINTIC 31
ACTIVITY
LIST OF FIGURES
S.NO TITTLE NAME PAGE NO

1.1 Life cycle of Helminths
1.2
Types of internal parasitic
worms
1.3 Classification of Helminths 32
1.4 Mechanism of action of 34
anthelmintic Drugs
Introduction
Helminthiasis is a macro parasitic disease of human and animals in which a part of body is infested with the
parasitic worms such as pinworm, roundworm or tapeworm. Intestinal.
parasitic helminths such as roundworms (Ascaris lumbricoides), hookworms

(Ancylostoma doudenale and Necator americanus) and whipworm (Trichuris
trichiura) are common in the developing world. The prevalence of intestinal worm
infestation in India varies from 5% to 76%, which is similar to that in other
developing countries. These parasitic infestations are acquired by ingestion,
inhalation or penetration of the
skin by the infective forms. Typically, the worms reside in the gastrointestinal tract but
may also burrow into the liver and other organs. Infected people excrete helminth
eggs in their faeces, which then contaminate the soil in areas with inadequate
sanitation. Other people can then be infected by ingesting eggs or larvae in
contaminated food, or through penetration of the skin by infective larvae in the soil
(hookworms). Infestation can cause morbidity and sometimes death, by
compromising nutritional status, affecting cognitive processes, including tissue
reaction, such as granuloma, and provoking intestinal obstruction or rectal prolapse.
A.K.R.G COLLEGE OF PHARMACY 1

Introduction
Helminth infections are among the most widespread infections in humans,

distressing huge population of the world. About 3.5 billion people in the world are
affected and 450 million are ill as a result of parasitic infection. The diseases
caused by helminth
infection is often called tropical diseases. They are in fact diseases of under
development since the common feature of the societies in which helminth infection
are highly prevalent is low socioeconomic status. In tropical regions, where
prevalence is greatest, simultaneous infection with more than one type of
helminth is
3
common . The majority of the infections due to helminthes are generally restricted to
tropical regions and cause enormous hazards to health and contribute to the
prevalence of undernourishment, anemia, eosinophilia and pneumonia. Some
examples of the effects of the helminth infection in man:
(a) Mechanical obstruction- large ascaris burden can cause mechanical or
spastic
(b) obstruction of the intestine and produce serious or fatal illness.
(c) Hypersensitivity- the antigens shed by ascaris larvae as they migrate

through the lungs can induce antibody formation and cause severe
immediate hypersensitivity or asthma.
(d) Anaemias – hookworm infection leads to morbidity by blood loss, iron

deficiency anaemia and hypoproteinaemia which can be serious in
individuals already malnourished.
(e) Malnutrition – helminth infection can have adverse effect on the nutritional
status especially of individuals who are already suffering from starvation.

Introduction
The mechanism by which helminth infection can adversely affect nutritional

status are complex, but may include: loss of appetite; decreased absorption
of essential nutrients such as iron and vitamins; host-parasite competition
for food materials; and diarrhoea.
(f) Upper gastrointestinal haemorrhage due to portal hypertension caused by

S. mansoni is a major cause of mortality among adult males in Egypt.
(g) Carcinoma of the bladder associated with chronic Schistosoma h a e m a t o b i u m

infection is a common malignancy in Egyptian men.
Parasitic diseases cause ruthless morbidity affecting principally population in

endemic areas. Prevalence is commonly combined with worm burden (also referred
to as the
―intensity of infection‖), which is commonly measured by the number of eggs per
gram (EPGs) of feces for intestinal helminths and schistosomes. Based on EPGs and
their association with morbidity, individuals are classified into categories of light,
moderate, and heavy infection by the WHO. Furthermore, in the case of soil-
transmitted helminths, the WHO recommends use of both prevalence and intensity
of infection to classify communities into transmission categories — category I
(high), category II (medium), and category III (low). These transmission categories
are assigned according to both the number of heavily infected people in the
community (greater or less than 10%) and the
prevalence of infection (greater or less than 50%). The gastro intestinal helminthes
become resistant to currently available anthelmintic drugs.

Introduction
ROUNDWORMS
Roundworms are a member of the nemathelminth’s phylum or group of animals. The

hookworm, pinworm and trichinella are part of this group. They are more advanced than
flatworms but less advanced than earthworms. They have thin round bodies, with none of
the pieces or segments that earthworms have in their bodies.
Roundworms live in salt water, fresh water and the soil. Many of them are harmful to
man as they are parasites.
Scientific classification
Kingdom: Animalia
clade: Nematoid
Phylum: Nematoda
Anatomy and physiology of roundworms.
Muscular Skeletal - A roundworm has no skeleton.
Digestion - A roundworm has a definite digestive system that runs the length of their
bodies. It has a mouth, pharynx, intestine and anus. Many are parasites and live off other
animals and plants.
Nervous - A roundworm has two nerve cords that transmit impulses in the roundworm.
Circulation - A roundworm has no heart or formal blood vessels.
Respiration - A roundworm has no formal respiratory system.
Reproduction - A roundworm reproduces sexually. The female has an ovary, holds eggs
in an oviduct and then passes them to the uterus, where they are fertilized. When it is time
to reproduce, the sperm cells pass through the spicule. Over 200,000 eggs can be
deposited at once in the soil once they are fertilized.

Introduction
Excretion - A roundworm has an anus at its rear end and a series of excretory tubes that
end in an excretory pore.
Symmetry - A roundworm has bilateral symmetry.
Appearance - A roundworm is thin, round, smooth and can be up to four feet in length.

Introduction
Life cycle
Fig.1.1 Life cycle of Helminths
Step 1: Adult worms live in the lumen of the small intestine. A female may produce up to
200,000 eggs per day, which are passed in the feces.
Step 2 and 3: Fertile eggs embryonate and become infective after 18 days to several
weeks, depending on the environmental conditions (optimum: moist, warm, and shaded
soil).
Step 4: The eggs of the worm are found in soil contaminated by human feces or in
uncooked food contaminated by soil containing eggs of the worm. Humans are infected

Introduction
when they ingest the contaminated soil found in their food, on their
fingers, or in their drinks.
Step 5: The eggs hatch into larvae within the person’s intestine.
Step 6: The larvae penetrate the intestine wall and reach the lungs
through the blood stream.
Step 7: The larvae mature further in the lungs (10 to 14 days), penetrate the
alveolar walls, ascend the bronchial tree to the throat, and are swallowed.
Upon reaching the small intestine, they develop into adult worms. The
female adult worm, which can grow to more than 30 cm in length, lays eggs
that are then passed into the feces. If soil is polluted with human or animal
feces containing eggs, the cycle begins again. An adult Ascaris may live up
to one and a half years. Humans are the only reservoir, but the eggs
may
remain viable in soil for years.
EARTHWORM
It is the common name for the largest members of Oligochaeta (which is
either a class or subclass depending on the author) in the phylum Annelida.
Folk names for the earthworm include "dew-worm", "Rainworm", "night
crawler" and "angleworm".
ANATOMY OF EARTHWORMS
The basic body plan of an earthworm is a tube, the digestive system, within
a tube, the muscular slimy, moist outer body. The body is annular, formed
of segments that are most
specialized in the anterior. Earthworms have a simple closed circulatory

Introduction
system. They have two main blood vessels that extend through the length of their
body: a ventral blood vessel which leads the blood to the posterior end, and a dorsal
blood vessel which leads to the
anterior end. The dorsal vessel is contractile and pumps blood forward,
where it is pumped into the ventral vessel by a series of "hearts" (aortic
arches) which vary in number in the different taxa. The blood is distributed
from the ventral vessel into capillaries on the body wall and other organs
and into a vascular sinus in the gut wall, where gases and nutrients are
exchanged. This arrangement may be complicated in the various groups by
sub esophageal, paraoesophageal, parietal and neural vessels, but the basic
arrangement holds in all earthworms. Most earthworms are decomposers
feeding on
10
undecayed leaf and other plant matter, others are more geophageous .
HOOKWORMS
The hookworm is a parasitic nematode that lives in the small intestine of its
host, which may be a mammal such as a dog, cat, or human. Two species of
hookworms commonly infect humans, Ancylostoma duodenale and Necator
americanus
Hookworms are much smaller than the larger roundworm Ascaris

lumbricoides, and the complications of tissue migration and mechanical
obstruction so frequently observed with roundworm infestation are less
frequent in hookworm infestation. The most significant risk of hookworm
infection is anemia, secondary to loss of iron (and protein) in the gut. The
worms suck blood voraciously and damage the mucosa. However, the blood
loss in
the stools are not visibly apparent.
WHIPWORMS
The human whipworm (Trichuris trichiura or Trichocephalus trichiuris) is a

roundworm, which causes trichuriasis when it infects a human large
intestine. The name whipworm refers to the shape of the worm; they look
like whips with wider "handles” at the posterior end.

Introduction
Trichuris trichiura has a narrow anterior esophageal end and shorter and
thicker posterior anus. These pinkish-white worms are threaded through the
mucosa. They attach to the host through their slender anterior end and feed
on tissue secretions instead of blood.
Roundworms Hookworms
Tapeworms Whipworms
Fig.1.2 Types of internal parasitic worms
LIFE CYCLE OF HELMINTH PARASITES

Helminthes have complicated life cycles in which man is either a primary
or secondary host. The primary (or definitive) host harbours the adult
sexually mature worms. The intermediate host harbors other stages in the
life cycle. In many cases, man becomes
infected by ingesting the parasite in the form of the cysterici (encysted

larvae) in inadequately cooked flesh (beef, pork, or fish) or by ingesting
ova deposited by the primary host. Man is infected by the free-living larvae
after penetrating the skin the microfilariae worms are transmitted by biting
flies, etc.

Introduction
TREATMENT
The aim in the anthelmintic chemotherapy, as in bacterial and protozoal
chemotherapy, is to introduce into the infected person or domestic animals
a drug which are toxic to the helminth parasite but not to the host. The
drugs should selectively interfere with physiological or biological processes
essential for the functional integrity of the worms. Selective toxicity can
also be achieved in case of helminthes residing in the lumen, by using
orally active, non-absorbable drugs which affect parasite function by direct
contact into gut. One difference between helminthes and other microbial
infection having a bearing on chemotherapy is that the most helminthes
parasites do not multiply in the host as do protozoa or bacteria.
Consequently, severity of helminthes infection depends on number of eggs
or larvae entering the host. Therefore, inhibition of growth, good strategy in
the chemotherapy of the bacterial infection, is not a useful approach in
helminthes chemotherapy. Rather the aim here is to weaken the worms and
4
expel it or to kill it .
Anthelmintic are those agents that expel parasitic worms (helminthes)

from the body, by either stunning or killing them. Anthelmintics are drugs
that act either locally to expel worms from gastrointestinal tract or
systematically to eradicate adult helminthes or development from that
invade organs and tissues. Because of increasing anthelmintic resistance
and impact of conventional anthelmintic on the environment, it is important
to look for anthelmintic strategies against gastrointestinal nematodes. It has
been well
accepted that due to the limited availability and affordability of modern

medicines most of the world’s population depends to a greater extent on
traditional medical remedies. Herbs have always been principal forms of
medicine in India and presently they are becoming popular throughout
developing countries. It has been well evidenced that the traditional
medicine including plants and plant derived preparation hold a great
promise as a source of easily available effective anthelmintic agents to the
people. Number of synthetic drugs used to control and prevent the

Introduction
infestation related to worms like mebendazole, fenbendazole, albendazole,

piperazine and pyrantel, almost mebendazole used as broad-spectrum
anthelmintic drug.

Introduction
HERBAL MEDICINAL PLANTS USED AS ANTHELMENTIC
Many herbs are used as natural anthelmintics like
Roots of plants like:
-Thespesia lampus (Ranibhendi)

-Chloroxylon swietenia (yellow wood)
-Carissa spinarum (karaunda)
-Baliospermum montanum (Danti)
plants like:
-Cocos nucifera (Kalijiri)

-Trigonella foenum-graecum (fenugreek, methi)
-Coriandrum sativum (coriander)
-Azadirachta indica (neem)
-Caesalpinia crista
Leaves of plants like:
-Cassia tora (sickle senna)
-Camellia sinensis (Tea)
-Gmelina arborea (Gambhari)
Bark of plants like:
-Ficus racemosus
-Alstonia scholaris (Saptaparni)
Fruits of plants like:
-Embelia ribes (vidang).

Introduction
PHARMACOTHERAPY 21
Anthelmintic drugs may be classified according to their spectrum of

Anthelmintic activity as follows:
1. Broad Spectrum anthelmintics:
a) Drugs effective in roundworms, threadworms, hookworms,

tapeworms and flukes.
e.g., Mebendazole.
b) Drugs effective in roundworms, threadworms and
hookworms
e.g., Thiabendazole, Pyrantel pamoate and Albendazole.
2. Narrow Spectrum anthelmintics:
a) Drugs effective in roundworms and threadworms

e.g., Piperazine, Tetramisole or Levamisole, Ivermectin,
Santonin and Hexylresorcinol.
b) Drugs effective in hookworm infection e.g.,
Bephenium, Tetrachloroethylene, Thymol and
Chenopodium.
c) Drugs effective in filariasis e.g., Diethylcarbamazine.
d) Drugs effective in tapeworm e.g., Niclosamide, dichlorophen,
Maleferns, Mepacrine and Chloroquine.
e) Drugs effective in flukes e.g., Praziquantel, Bithionol,
Metrifonale and Oxamniquine.

Introduction
fig .1.3 Classification of Helminths

Introduction
Anti-inflammatory: Stinging nettle contains compounds like flavonoids and phenolic acids that
have been found to possess anti-inflammatory properties. This makes it useful in traditional
medicine for conditions such as arthritis and allergic rhinitis.
Diuretic: The plant has been traditionally used as a diuretic, promoting urine production and
helping to flush out toxins from the body. This diuretic activity is beneficial for conditions such as
urinary tract infections and edema.
Antioxidant: Stinging nettle contains antioxidants like flavonoids, which help neutralize free
radicals in the body, reducing oxidative stress and inflammation. This antioxidant activity
contributes to its overall health-promoting effects.
Antimicrobial: Some studies have shown that extracts of stinging nettle possess antimicrobial
properties, inhibiting the growth of certain bacteria and fungi. This activity may contribute to its
traditional use in treating infections.
Hypoglycemic: Research suggests that stinging nettle may have hypoglycemic effects, helping to
lower blood sugar levels. This potential activity could be beneficial for managing diabetes and
related complications.
Hypotensive: There is evidence to suggest that stinging nettle may have hypotensive effects,
meaning it can help lower blood pressure. This activity may be attributed to its vasodilatory
properties.
Overall, Urtica dioica exhibits a range of pharmacological activities, making it a subject of interest
for further research and potential therapeutic applications. However, it's essential to consult with
a healthcare professional before using stinging nettle for medicinal purposes, especially if you
have any underlying health conditions or are taking medications.

Literature review
Taxonomy of Urtica dioica

Kingdom: Plantae
Phylum: Angiosperms
Class: Eudicots
Order: Rosales
Family: Urticaceae
Genus: Urtica
Species: Urtica dioica
Synonyms:
Urtica major
Urtica urens
Urtica galeopsifolia
Urtica procera
Urtica hirsuta
Urtica gracilis
Common names of Urtica dioica include:
Stinging nettle
Common nettle
Nettle
Stinger
Nettles
Nettle leaf
European nettle
The classification of Urtica dioica is as follows:
Kingdom: Plantae
Clade: Tracheophytes (vascular plants)
Clade: Angiosperms (flowering plants)
Literature review
Clade: Eudicots
Clade: Rosids
Order: Rosales
Family: Urticaceae
Genus: Urtica
Species: Urtica dioica
URTICA DIOICA

Literature review
DESCRIPTION
Urtica dioica, commonly known as stinging nettle, is a perennial herbaceous
plant characterized by its serrated, heart-shaped leaves and its ability to deliver a
painful sting upon contact with its tiny, hair-like structures containing irritants. The
plant typically grows to a height of 1 to 2 meters and thrives in disturbed areas,
woodlands, meadows, and along streams.
The leaves of stinging nettle are arranged oppositely along the stem and are
covered with small, stinging hairs. Despite its defensive mechanism, stinging nettle
is valued for its medicinal properties and as a food source. The plant produces small,
greenish flowers arranged in clusters called inflorescences.
Stinging nettle has a long history of use in traditional medicine and cuisine. Its
leaves are edible when cooked or dried and are rich in vitamins, minerals, and
protein. Medicinally, it has been used to treat a variety of ailments such as arthritis,
allergies, urinary tract infections, and skin conditions due to its anti-inflammatory
and diuretic properties.
Overall, Urtica dioica is a versatile plant with both beneficial and defensive
attributes, making it an important species in various ecological and cultural
contexts.
Urtica dioica has a rich ethnobotanical history and various traditional uses:
Medicinal Uses: Stinging nettle has been used medicinally for centuries. It is
believed to have diuretic, anti-inflammatory, and antioxidant properties. It has been
used to treat conditions such as arthritis, allergies, urinary tract infections, and skin
irritations.
Food Source: Despite its stinging hairs, stinging nettle is edible when cooked or
dried. Its young leaves can be harvested and used in soups, teas, or as a spinach
substitute. It is rich in vitamins, minerals, and protein.
Fiber Source: Historically, the fibres from stinging nettle stems have been used to
make textiles. The plant's fibres are strong and have been used to make cloth, rope,
and paper.
Cultural and Ritual Uses: In some cultures, stinging nettle has been used in rituals or
as a protective charm. Its stinging properties were believed to ward off evil spirits or
provide protection against negative energies.

Literature review
Wildlife Habitat: Stinging nettle provides habitat and food for various insects,
including butterfly larvae, which feed on its leaves.
Overall, Urtica dioica has played a significant role in traditional medicine,
Chemical constituents of plant:
 Flavonoids: Quercetin, kaempferol, rutin, and others. Flavonoids contribute to the plant's
antioxidant properties.
 Phenolic compounds: Chlorogenic acid, caffeic acid, ferulic acid, and others. These compounds
have antioxidant and anti-inflammatory properties.
 Amino acids: Histidine, lysine, phenylalanine, serine, and others. Amino acids are essential building
blocks for proteins and play various roles in metabolic processes.
 Minerals: Calcium, magnesium, iron, potassium, and others. These minerals are essential for
various physiological functions in the human body.
 Vitamins: Vitamin A, vitamin C, and vitamin K. These vitamins contribute to overall health and
immune function.
 Lignans: Secoisolariciresinol and others. Lignans have been studied for their potential health
benefits, including hormone-balancing effects.
 Triterpenes: β-sitosterol, stigmasterol, and others. Triterpenes have been investigated for their
anti-inflammatory and anti-cancer properties.
 Acids: Linoleic acid, linolenic acid, palmitic acid, stearic acid, and others. These fatty acids play roles
in lipid metabolism and cellular function.
These constituents contribute to the medicinal properties of Urtica dioica, including its anti-
inflammatory, antioxidant, and diuretic effects, among others.
Nutrition, and cultural practices across different regions.

Drug profile
Drug profile
 Fenbendazole is a member of the class of benzimidazoles that is 1H-benzimidazole which is substituted
at positions 2 and 5 by (methoxycarbonyl)amino and phenylsulfamide groups, respectively. A broad-
spectrum anthelmintic, it is used, particularly in veterinary medicine, for the treatment of
Nematoda infections
 IUPAC name Methyl N-(6-phenylsulfanyl-1H-benzoimidazol-2-yl) carbamate
 MEDICINAL USES
1. Anthelmintic: Fenbendazole is primarily used as an anthelmintic medication to treat parasitic

worm infections in animals. It is effective against various types of worms including roundworms,
hookworms, whipworms, and certain types of tapeworms.
2. Veterinary Medicine: Fenbendazole is commonly used in veterinary medicine to treat parasitic

infections in dogs, cats, horses, livestock, and other animals.
3. Anticancer Properties: Preliminary studies and anecdotal reports suggest fenbendazole may have
potential anticancer properties, but further research is needed to fully understand its safety and
efficacy for this purpose in humans.
4. Fenbendazole is a broad-spectrum anthelmintic (anti-parasitic) drug commonly used in veterinary

medicine to treat various parasitic infections in animals. It belongs to the benzimidazole class of
compounds. Here is a drug profile of fenbendazole:
5. Mechanism of Action: Fenbendazole acts by disrupting the microtubule structure in the cells of
parasites, thereby inhibiting their ability to divide and reproduce. This leads to the death of the
parasites.
6. Administration: Fenbendazole is typically administered orally as a tablet, paste, or liquid

suspension. The dosage and duration of treatment vary depending on the type of infection and the
species of the animal being treated.
7. Side Effects: Although fenbendazole is generally considered safe when used as directed, some
animals may experience side effects such as:
8. Vomiting
9. Diarrhea
10. Loss of appetite
11. Lethargy
12. In rare cases, allergic reactions or neurotoxicity may occur.

Drug profile
MOA OF FENBENDAZOLE
Mechanism of Action:
Fenbendazole acts by disrupting the microtubule structure in the cells of parasites, thereby inhibiting their
ability to divide and reproduce. This leads to the death of the parasites

AIM AND OBJECTIVES
Aim and Objectives
AIM:
The aim of the present study is to evaluate invitro anthelmintic activity of ethanolic extract of Urtica dioica.
OBJECTIVES:
 Following are the objectives of the study
 To Identify plant Urtica dioica and authentication of plant
 To collect all equipment’s and laboratory chemicals required for study Collect leaves of Urtica
dioica and prepare ethanolic leaf extract
 Drying of ethanolic leaf extract by rotary evaporator
 To carry out phytochemical screening for ethanolic plant extract
 To collect Indian earthworms
 To carry out in-vitro study on earthworms.

EXPERIMENTAL DESIGN
Experimental Designs
PLANT COLLECTION:
 Plants are collected from the nearby institution surroundings.
 Plants claimed with anthelmintic activity as selected for the proposed study and suitable plant part
leaf ethanolic extract is prepared.
ANIMALS
The anthelmintic activity was performed on adult Indian earth worm “Pheretima posthuma” it has
anatomical and physiological resemblance with the intestinal round worm sims of human beings’ Indian
earthworms are collected from vermicompost area of Nallajerla, east Godavari district. All the Earth worms
are handled with gloves and placed in sterilized Petri dishes.
DRUGS AND CHEMICALS:
Fenbendazole tablets was procured in surrounding pharmacies of Nallajerla All other laboratory chemicals
required ethanol 90%for phytochemical screening was procured National Scientific products (NSP).
INSTRUMENTS:
Weighing balance, gloves, whattsman filter paper, and mortar and pestle, petridishes, droppers, heating
mantle, filtration equipment.
METHODS:
Preparation of plant extract
The crude ethanolic extract was prepared by the simple decoction, method using 20 g of triturated dried
leaves for 300ml of ethanol After 30- mints of decoction the whole content was blended in a domestic
blender filtered by whattsman filter papers extract obtained was evaporated in rotary evaporator.



PRELIMINARY PHYTOCHEMICAL SCREENING:

Ethanolic extract obtained by above procedure is subjected to phytochemical screening of qualitative
analysis of phytoconstituents
TEST FOR ALKALOIDS:

SN TEST OBSERVATION INFERENCE
O
1 Wagner’s test Reddish brown Presence of alkaloid

precipitate
1.27 gm of iodine +2 gm of potassium
iodide made up to 200 ml
2 Mayer’s Test White to buff precipitate Presence of alkaloid
1.36 gm of mercuric chloride +5 gm of

potassium iodide in 60 ml of distilled
water and made up to 200 ml
3 Dragendroff’s Test
14 gm of sodium iodide +5.2 gm of Orange – brown colour Presence of alkaloid

bismuth carbonate in 50 ml glacial acetic precipitate
acid. 40 ml filtrate+160 ml of acetate
+1ml of water. 10 ml of stock +20 ml of
acetic acid made up to 100 ml with water
4 Hager’s Test Crystalline precipitate Presence of alkaloids

with many precipitates
Saturated aqueous picric acid
Table No. 1.2 Test for Alkaloid

TEST FOR CARBOHYDRATES:

SNO TEST t’s OBSERVATION INFERENCE
1 Benedict’s Test Brick red precipitate at Presence of

bottom of the test tube carbohydrates
5 ml of benedict’s reagent (benedict’s A+
benedict’s B) +3ml of test solution on a
water bath
2 Fehling’s Test Yellow or brick red Prescence of

precipitate at the bottom carbohydrates
2 ml of Fehling A+2 ml of Fehling B + 2ml
of test tube
of extract were boiled
3 Molisch’s Test A violet ring at the Presence of

junction of two liquids carbohydrates
Aqueous or alcoholic solution of the
extract +10% alcoholic solution of α -
naphthol+ concentrated sulphuric acid
Table No.1.3 Test for carbohydrates
TEST FOR CARDIAC GLYCOSIDES:

SNO TEST OBSEVATION INFERENCE
1 Keller -Killiani Test A reddish- brown colour is Presence of cardiac

formed at the junction of glycosides
Extract of the drug in glacial acetic
the two layers and upper
acid+ few drops of ferric chloride +
layer turns bluish green
concentrated sulphuric acid
2 Legal Test Change from pink to red Presence of cardiac

colour glycosides
Extract + pyridine sodium
nitroprusside solution and + sodium
hydroxide solution.

TEST FOR ANTHRAQUINONE GLYCOSIDES:

TEST OBSERVATION INFERENCE
SNO
Bontrager ‘s Test
1 Boil 0.1 gm of the drug with 5 ml of Presence of

10 % sulphuric acid filtrate and anthraquinone
Lower ammonia layer will
shake with benzene layer is glycosides
show red pink colour
separated and 10% aqueous
ammonia was added and shaken
gently
Modified Borntrager’s Test The lower ammonia layer Presence of

shows pink colour anthraquinone
2 Boil 0.1 gm of the drug +dilute HCL
glycosides
+5ml of 5% solution of ferric
chloride. shake the filtrate with
benzene; add dilute ammonia
solution to benzene layer
Table No.1.5 Test for Anthraquinone Glycosides

TEST FOR GUMS AND MUCILAGES
SNO TEST OBSEVATION INFERENCE
1 Ruthenium Red Test Solution turns to red colour Presence of mucilage
Drug + 0.08 Gm of Ruthenium Red

+10 ml of 10%solution of lead
acetate
2 Molisch’s Test l A violet ring at the junction of Presence of

two liquids carbohydrates, gums
Aqueous or alcoholic solution of the
and mucilage
extract +10%alcoholic solution α −¿
napthol+ concentrated sulphuric
acid along the side of the test tube
Table No.1.6 Test for Gums and mucilage’s

TEST FOR PROTEINS AND AMINO ACIDS:

SNO TEST OBSERVATION INFERENCE
1 Biuret Test The appearance of violet or Confirms the

purple colour presence of proteins
2 ml of the extract +2 ml of 10% NaOH
solution +2-3 drops of 1%CuSo4 solution
were mixed
2 Ninhydrin Test The appearance of blue Confirms the

colour presence of proteins
0.5 ml of ninhydrin solution +2 ml of the
extract boiled for 2 minutes
3 Xanthoproteic Test The appearance of orange Confirms the

colour solution presence of proteins
2 ml of extract +1 ml of conc. HNO3
boiled and cooled add 40% NaOH
solution added drop by drop to it
4 Mallon’s Test The appearance of red Confirms the

precipitate and red coloured presence of proteins
2ml of extract +2 ml of million’s reagent
solution
were boiled and cooled then add few
drops of NaNo2
Table No.1.7 test for proteins and amino acids

TEST FOR TANNINS AND PHENOLIC COMPOUNDS:

1 Test with lead acetate White precipitate is Presence of tannins.

formed.
1 ml of the extract +lead acetate.
2 Test with Ferric chloride Precipitate is formed. Presence of phenols.
5 ml of the extract +5% W/V solution of

ferric chloride in 90% alcohol.
3 Test with gelatine solution A white buff precipitate is Presence of tannins.

formed.
5 ml of the extract +1% aqueous
solution of gelatine +10% sodium
chloride.
Table No:18 Test for Tannins and Phenolic compounds
TESTS FOR FLAVONOIDS
1 Test with NaOH A yellow colour Confirms the presence of

flavonoids.
The extract was first dissolved with appears.
water.it was filtered and the filtrate was
treated with sodium hydroxide.
2 Shinoda Test Appearance of pink The presence of

colour
Extract +a pinch of magnesium powder flavonoids bioflavonoids
was added followed by conc. HCL.
Table No:1.9Test for Flavonoids

TEST FOR STEROIDS AND STEROLS:
1 Salkowski’s Test the upper chloroform The presence of

layer showing a play of steroids and sterols
5 ml of extract in chloroform +conc.
colours first form bluish
Sulphuric acid along with side of the
red to gradually violet and
test tubes
lower acid layer showing
yellow colour with green
fluorescence
2 Liebermann Burchard reagent Test An emerald green colour The presence of

develops. steroids and sterols
5 ml of the extract in chloroform +2
ml of acetic anhydride + 2-3 drops of
conc. Sulphuric acid and allow to
stand for few minutes
Table No.2.0Test for steroids and sterols
TEST FOR TRITERPENOIDS:

1 Test with TIN AND THIONYL CHLORIDE Pink colour was The presence of
developing. triterpenoids
Extract was dissolved in chloroform +
apiece of metallic tin +1 drop of thionyl
chloride was added to it.
TABLE NO.2.1 TEST FOR TRITERPENOID

TEST FOR SAPONINS:
S. TEST OBSERVATION INFERENCE

No
1 1 cm layer of foam after The presence of

FOAM TEST standing for 30 minutes saponins
1ml of alcoholic and aqueous

extract was diluted separately with distilled
water make the volume up to 10 ml, and
shaken in a graduated cylinder for 15
minutes and kept aside.
TABLE NO.2.2TEST FOR SAPONINS:

EXPERIMENTAL DESIGN
Experimental was carried out on 42 adult Indian earthworms randomly divided into nine groups with
distilled water, extract fenbendazole (Brand name Pana cur).
TREATMENT SCHEDULE FOR ANTHELMINTIC ACTIVITY:

GROUPS TREATMENT CONCENTRATION PURPOSE
normal saline (Nacl) To serve as a control

I. -
Standard 10%
II.
(Fenbendazole) 20% TO SERVE AS A
30% Standard
40%
50%
TEST 10% To accesses

III. anthelmintic activity
(Plant extract) 20%
30%
TREATMENT SCHEDULE FOR ANTHELMINTIC ACTIVITY:

GROUPS TREATMENT CONCENTRATION PURPOSE
normal saline (Nacl) To serve as a control

I. -
Standard 10%
II.
(Fenbendazole) 20% TO SERVE AS A
30% Standard
40%
50%
TEST 10% To accesses

III.

(Plant extract) 20% anthelmintic activity
30%
 Group I receive normal saline (Nacl) water in the study acts as a

control.
 Group Il receiving standard drug fenbendazole is sub divided into
groups receiving 10%, 20%, 30%, 40%, 50% of standard drug.
 Group III receiving test dose of plant extract are further separated into
group receiving 10% 20% 30% of ethanolic plant extract.
All the animals treated with control, standard and plant extract was observed for
time taken for paralysis when no movement of any sort could be observed except
when the worms were shaken vigorously. Time for death of worms were recorded
after ascertaining that worms neither moved when shaken vigorously nor when
dipped in warm water All the results were shows in Table 1 and expressed as a
mean SEM of six worms in each group.

Results
PERCENTAGE YIELD OF EXTRACT:
S.n Extracts Colour Consistency Yield

o
01 Ethanolic extract Dark green More sticky 7.5%
The percentage yield was obtained using this formula:
W2-W1/W0×100(Anokwuru, c.p, c.p et al)
 Where W2 is the weight of the extract and the container.

 W1 the weight of the alone.
 W0 the weight of the initial dried sample.
PRELIMINARY PHYTOCHEMICAL SCREENING:
S. No Class of the compound Plant part(leaf) Test performed
01 Alkaloids + Dragendroff’s test,

Mayer’s test
02 Carbohydrates + Fehling test
03 Glycosides + Keller Killiani test
04 Proteins and amino acids + Xanthoproteic test
05 Flavonoids + Ammonia test
06 Saponins + With water Na2co3
07 Phenolic compounds and tannins + Ferric chloride text
Table No.2.6 Phytochemical screening

Results

Results

Results
EVALUATION OF ANIHELMINTIC ACTIVITY OF AQUEOUS LEAF EXTRACT OF

URTICA DIOICA:
Group Treatment Concentration of treatment Time taken in minutes Sem
Paralysis Death
Group1 Normal saline (Nacl) ------- --------- --------

water
Group2 Fenbendazole 20% ---------
40% ----------
60% 191± 5.89 219 ± 3.78
Group3 Ethanolic leaf extract of 20% 30 ± 19 ± 1.93***

Urtica dioica 1.50***
40% 16± 19± 0.99***

1.12***
60% 10± 14± 0.39****

0.72***
***p< 0.0001 significant when compared to the control group using Dunnett’s method of comparison.
Evaluation of anihelmintic activity of aqueous leaf extract of URTICA DIOICA
400
300
200
Time taken in minutes Sem
100
0
) ) ) )
0% 0% 0% 0%
(6 (2 (4 (6
ol
e ca ca ca
az oi oi oi
di di di
end ca ca ca
en
b
U rti U rti U rti
F of of of
a ct a ct a ct
r r r
xt xt xt
a fe a fe a fe
le le le
li c li c li c
no no no
ha ha ha
Et Et Et
Treatment
Paralysis Death

Results
Evaluati on of anihelminti c acti vity of aqueous

leaf extract of URTICA DIOICA
219 Paralysis Death
191
30
19
19
16
14
10
F en b en d azo l e (6 0 % ) Et h an o l i c l eaf ex t r ac t Et h an o l i c l eaf ex t r ac t Et h an o l i c l eaf ex t r ac t
o f U r ti c a d i o i c a ( 2 0 % ) o f U r ti c a d i o i c a ( 4 0 % ) o f U r ti c a d i o i c a ( 6 0 % )
Paralysis
10 Fenbendazole (60%)
16 Ethanolic leaf extract of
Urtica dioica (20%)
Urtica dioica (40%)
Ethanolic leaf extract of
Urtica dioica (60%)
191
Death
Fenbendazole (60%)
19 Urtica dioica (20%)
Urtica dioica (40%)
Ethanolic leaf extract of
Urtica dioica (60%)
219

Discussion
Helminth infections are among the widespread infections in distressing a huge population of the world:
Intestinal worm infestations are widely prevalent in tropical and subtropical countries and occur where
there is poverty and poor sensation. Soil-transmitted helminth (STH) infections form the most important
group of intestinal worms affecting two billion people worldwide and the main species, which infect are
Ascarislumbricoides, (roundworms). Trichuristrichiura, (whipworm and Necatoramericanus
Ancylostomaduodenale (hookworms) According to World Health Organisation (WHO), globally there are
1221 e1472 million cases of Ascariasis, 75001050 million cases of Trichiniasis and 740el 300 million cases of
hookworm infestation.
It was found that various problems like drug resistance occurred to several families of chemical
anthelmintic drugs hence the present study is carried to evaluate the ant homothetic potential of some
traditional plant of Urtica dioica the leaves of Urtica dioica is selected to evaluate anthelmintic activity.
Aqueous leaf extract of Urtica dioica was prepared and anthelmintic activity was evaluated by in-vitro
made on earthworms.
The study results showed that phytochemical screening of Urtica dioica leaf ethanolic extract contain
the following class of phytoconstituents alkaloids, tannins, phenolic compounds proteins amino acids,
flavonoids glycosides.
The antihelminth etic activity of Urtica dioica leaf ethanolic extract (CGLE) was evaluated by In-vitro
method on Indian earthworms divided into three groups receiving distilled water, standard and test doses
of drug in 20%.40% and 60%. The study results of test and standard are compared by observing time taken
for paralysis of worms when no movement of any sort could be observed except when the worms were
shaken vigorously. Time for death of worms were recorded after ascertaining that worm neither moved
when shaken vigorously nor when dipped in warm water. All the results were expressed as a mean SEM of
six worms in each group.
The current study results showed that earthworms (Group II) treated with standard drug fenbendazole at
doses of 20% 40%, does not exhibit any paralysis and mortality of worms It is also notified from the present
study results that earthworm administered with standard drug at 60% of fenbendazole shrewd paralysis
and mortality of worm after 4 hours of drug administration. A dove dependent of standard drug was from
the study result
It was observed from the study results that Group II treated with test drug with 20% and 40% exhibit
paralysis and mortality with 30s1:45 354191 and 16x1.06, 19:0.94 minutes. It is also observed from the
study results that Group II treated with test drug at doses of 60% also exhibit paralysis and mortality with in
10:0.69, 1440.26 minutes. From the results it was observed that Group Il treated with test doses of drug
exhibit a significant paralysis and mortality of earthworms when compared to standard group
The present investigation exhibited that significant anthelmintic activity was showed by test drug when
compared to standard drug. The better anthelmintic activity of leaf extract of Urtica dioica compared to
standard drug may due to the presence of phytoconstituents in leaf extract which are revealed in
phytochemical screening. The plant is reported to contain phytoconstituents like tannins phenolic
compounds and alkaloids which might be the reasons for possible and better antihelminth etic activity of
leaf extract compared to standard drug fenbendazole (Piyush Jain, et. al.)

Discussion
Fig NO.1.4 mechanism of action anthelmintic Drugs

Conclusion
From the present study results it was concluded that ethanolic extract Urtica
dioica showed good in-vitro anthelminthic activity. The study results when
compared to standard drug fenbendazole ethanolic extract of Urtica dioica possess
better significant antiemetic activity compared to standard drug fenbendazole .
The current research also provides a scope for further expansion for
conducting the preclinical and clinical phases of evaluation

1. Ananthakrishnana S, Nalini P, Pani SP, ―Intestinal geohelminthiasis in the developing
World‖, Natural Medicinal Journal of India, March-April 1997, 10(2), 67-71.
2. WHO. World Health Report, Conquering suffering enriching humanity 2000.
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5. Peter J. Hotez, Paul J. Brindley, Jeffrey M. Bethony, Charles H. King, Edward J. Pearce and
Julie Jacobson, ― Helminth infection: the great neglected tropical diseas.
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7. http://en.wikipedia.org/wiki/Nematode.
8. http://www.mswdn.org/Animals/Roundworms.html
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269.
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IN-VITRO ANTI-HELMENTHIC ACTIVITY OF ETHANOLIC LEAF EXTRACT OF

Jawaharlal Nehru Technological University Kakinada (JNTUK)

D. ABHISHEK (208N1R0026) G. YUVARAJU (208N1R0030)

M. VIMALA JYOTHI (208N1R0053) N. DIVYA MOUNIKA (208N1R0059)

P. RAMU (208N1R0062) P. GAYATRI (208N1R0070)

UNDER THE GUIDANCE OF

A.K.R.G. COLLEGE OF PHARMACY

Approved by PCI, New Delhi

Approved by PCI, New Delhi

Affiliated to JNTU-K & Recognized by APSCHE, AP

NALLAJERLA-534112, East Godavari Dist., Andhra Pradesh, India.

DECLARATION BY THE CANDIDATE(S)

We hereby declare that the dissertation entitled “IN-VITRO ANTI-HELMENTHIC ACTIVITY OF

V. LAVANYA (208N1R0003) SK. SHARMILA (208N1R0004)

Affiliated to JNTU-K & Recognized by APSCHE, AP

NALLAJERLA-534112, East Godavari Dist., Andhra Pradesh, India.

CERTIFICATE BY THE GUIDE

This is certify that the dissertation entitled “IN-VITRO ANTI-HELMENTHIC ACTIVITY OF

(208N1R0003), SK. Sharmila (208N1R0004), D. Abhishek (208N1R0026), G. Yuvaraju (208N1R0030), M. Vimala

partial fulfillment of the requirement for the degree of “BACHELOR OF PHARMACY”

Date: Signature of the Research guide

Approved by PCI, New Delhi

Affiliated to JNTU-K & Recognized by APSCHE, AP

NALLAJERLA-534112, East Godavari Dist., Andhra Pradesh, India.

CERTIFICATE BY THE PRINCIPAL

This is certify that the dissertation entitled “IN-VITRO ANTI-HELMENTHIC ACTIVITY OF

(208N1R0003), SK. Sharmila (208N1R0004), D. Abhishek (208N1R0026), G. Yuvaraju (208N1R0030), M. Vimala

the guidance of KANIKELLA. SIVAJI, M.Pharm Professor, Department of Pharmaceutical Chemistry,

Requirement for the degree of “BACHELOR OF PHARMACY”.

Date: Signature of the Principal

Place: Nallajerla Dr. Pemmadi Raghuveer Varma M.Pharm., Ph.D.

V. LAVANYA (208N1R0003) SK. SHARMILA (208N1R0004)

My beloved parents &

S.NO TITTLE NAME PAGE

S.N TITTLE NAME PAGE NO

1.3 TEST FOR CARBOHYDRATES 23

1.4 TEST FOR CARDIAC GLYCOSIDES 24

1.5 TEST FOR ANTHRAQUINONE 24

1.6 TEST FOR GUMS AND MUCILAGE 24

1.7 TEST FOR PROTEINS AND AMINO 25

2.0 TEST FOR STEROIDS AND STEROLS 27

2.1 TEST FOR TRITERPENOIDS 27

2.2 TEST FOR SAPONINS 28

2.3 TREATMENT SCHEDULE 29

2.4 PHYTOCHEMICAL SCREENING 30

2.5 EVALUATION OF ANTHELMINTIC 31

S.NO TITTLE NAME PAGE NO

1.4 Mechanism of action of 34

parasitic helminths such as roundworms (Ascaris lumbricoides), hookworms

A.K.R.G COLLEGE OF PHARMACY 1

Helminth infections are among the most widespread infections in humans,

(c) Hypersensitivity- the antigens shed by ascaris larvae as they migrate

(d) Anaemias – hookworm infection leads to morbidity by blood loss, iron

A.K.R.G COLLEGE OF PHARMACY 2

The mechanism by which helminth infection can adversely affect nutritional

(f) Upper gastrointestinal haemorrhage due to portal hypertension caused by

(g) Carcinoma of the bladder associated with chronic Schistosoma h a e m a t o b i u m

Parasitic diseases cause ruthless morbidity affecting principally population in

A.K.R.G COLLEGE OF PHARMACY 3

Roundworms are a member of the nemathelminth’s phylum or group of animals. The

Anatomy and physiology of roundworms.

Muscular Skeletal - A roundworm has no skeleton.

Circulation - A roundworm has no heart or formal blood vessels.