Laboratory Manual
Pharmacology I
Salman Bin Abdulaziz University
COLLEGE OF PHARMACY
2014-15 / 1435-36
Lab No.
TABLE OF CONTENTS
PAGE
Laboratory Orientation
2-4
1
Study of Different Laboratory Animals and Their
Application
5-6
2
Study of Different Stages of Anesthesia
7-8
3
Study of Some Basic Instruments Used for Isolated
Tissue Experiments
9-10
4
Routes of Drug Administration to Laboratory Animals
11
5
To Evaluate the Analgesic Potency of Drug by 12-14
Formalin Test
6
To Evaluate the Analgesic Potency of Drug by Tail 15-16
Flick Method
7
To Evaluate the Analgesic Potency of Drug by Hot 17-18
Plate Method
8
Screening of Different Diuretics Using Laboratory 19-20
Animals
9
Screening of Anti-inflammatory Drug Using Hind Paw 21-22
Method
1
LABORATORY ORIENTATION
Introduction:
The laboratory portion of this course is designed to study the different laboratory animals, their
applications and screening of different pharmacological agents thoroughly than it is presented in
lecture. Core learning will come from practical and tissue studies. This method of ‘hands on’ learning
should also enhance and strengthen the knowledge you gain in lectures.
At times you will be working individually, in pairs or in groups of three or four. Each lab period is loosely
structured to begin with a short introduction to the exercise that highlights the activities of the day,
what materials are available for use and any changes in procedures. After that you will work
independently to learn the material.
There is never enough time in lab to go over each and every item that you are assigned. The lab is a
designated a time when you have access to materials that you will not have available during home study
time. Some of the information assigned in lab you can learn at home, particularly animals, instruments,
method, procedure, application, mechanism of action of drugs etc.
General Lab Rules:
1. Read the lab exercise before you come to lab. There is not time to review every aspect of each
exercise and still give you time to work on your own. I will assume that you know what the exercise
covers in general and I will only review changes or specific materials that you will use.
2. Before each lab, use the terminology list to mark the items in your manual’s text and illustrations that
you are responsible for learning.
2. Read and memorize the laboratory safety rules of the lab below. The preservatives are irritants and
some of you may be allergic to them. Gloves must be used during dissections and will be provided. Your
dissecting tools will be provided for you as well.
2
Pharmacology Laboratory: Safety, Procedures, Emergencies
These are minimum safety requirements. Instructors may institute additional policies at their discretion.
1. No open food or drink is permitted at any time, whether a lab is in progress or not. No eating,
drinking, candy, cough drops, chewing gum or tobacco is permitted. All beverage and food containers
must be put away in a backpack/bag/purse or left outside of the lab. There are shelves outside in the
hallway to store food and beverages during lab. Never taste anything at all while in the lab rooms,
unless it is a part of the lab activity (such as PTC paper). Also, do not apply cosmetics in lab (this includes
lip balm).
2. Visitors under 16 years of age and children are not allowed in the lab rooms at any time. Visitors over
16 may be allowed at the instructor’s discretion, as long as the lab activity does not involve hazardous
materials.
3. Know the locations of the eye wash and shower stations, fire alarm, fire extinguisher, first aid kit, and
emergency exits. Do not block access to these with trash cans, recycle bins, etc.
4. Safety instructions are given at the beginning of each lab period. Always arrive on time so that you
know what you are supposed to do and are informed of any specific safety concerns or safety
equipment associated with the day’s lab activity.
5. Wear any required personal protective equipment (lab coat, apron, goggles, etc).
6. Stash book bags safely so that they won’t trip people.
7. Report all illnesses, injuries, breakages, or spills to your laboratory instructor immediately.
8. Clean broken glass (glass that is not contaminated with any chemical reagents, blood, or bacteria) can
be swept up using the dust pan and placed in the broken glass container. If the glass is contaminated in
any way, keep the area clear to prevent tripping or laceration hazards, and consult your instructor for
proper disposal guidance. A broken glass flow-chart is available in the lab to help you decide what to do.
9. Notify your instructor if any of the equipment is faulty.
10. Clean up your entire work area before leaving. Put away all equipment and supplies in their original
places and dispose of reagents and infectious materials in the designated receptacles. Disinfect your
work surface if the lab activity involved any infectious materials. Otherwise, wipe the entire work
surface down with a clean, wet paper towel (no soap).
11. Use the appropriate waste containers provided for any infectious or hazardous materials used in lab.
12. Safety information reagents used in the lab activities can be found in the Material Safety Data Sheets
(MSDS), which are available in a binder in the lab. Know the location of the MSDS binder. We (faculty
and students) should be fully aware of the properties of the reagents we are using. Please use the
3
MSDSs. If you cannot find the MSDS for the reagent you are using in lab, inform your instructor. They
are also relatively easy to find online. A Google keyword example is “Sodium Chloride MSDS.”
13. Use caution with the lab chairs. Because they are on casters, they can roll away when you are
standing at your workstation. Make sure your chair is where you expect it to be before sitting down. Do
not use your chair as a means of moving from one part of the lab to the other.
14. Wash your hands before leaving the lab room.
15. If class is held at an alternate location (e.g. a field trip), you will be expected to conduct yourself
appropriately and follow lab safety rules where applicable.
GENERAL OBJECTIVES OF THE COURSE
At the end of the practical training in general, and experimental pharmacology the learner shall be
able to:
1. List the various dosage forms and enumerate their advantages and disadvantages.
2. Advise patients about the proper use of medication devices, storage of medicines etc.
3. Retrieve drug information from appropriate sources.
4. Appreciate the role of good laboratory practice in promotion of rational diagnostics, therapy, and
experimentation.
5. Realize the cardinal role of ethics in experimentation.
6. Order monitoring of drug levels where indicated and take appropriate remedial measures.
7. Prescribe rationally and in an individualized pattern.
8. Plan and carry out experiments to demonstrate the effect of drugs in experimental animals and
isolated tissues.
9. Critically appraise drug advertisements.
10. Apply fundamental statistical tests to experimental data and interpret results.
4
LAB 1: STUDY OF DIFFERENT LABORATORY ANIMALS AND THEIR
APPLICATION
AIM: To study the advantages, disadvantages and experimental uses of different most
commonly used laboratory animals.
BACKGROUND INFORMATION:
PHARMACOLOGY is the branch of science which deals with study of drugs on living systems.
EXPERIMENTAL PHARMACOLOGY: deals with study of effect of various Pharmacological agents
on different animal species.
OBJECTIVES OF PHARMACOLOGY:
▫
To find out the therapeutic agent suitable for human use
▫
To study the toxicity of the drugs
▫
To study the mechanism and site of action of drugs
LABORATORY ANIMALS: Animals those can be breaded and handled in laboratory.
Examples: Rat, Mice, Guinea pig, Rabbits, Frogs, Cat, Dog, Monkey, Pigeon etc.
1. RATS (Rattus norvegicus )
•
Albino rats of Wistar strain are commonly used
•
Other strains –
▫
Wistar kyoto rat
▫
Sprague Dawley rat
▫
Biobreeding (BBDP) rat
▫
Long-Evans rat
▫
Zucker rat
▫
Hairless rats (Rowett nude, Fuzzy, Shorn)
▫
RCS rats
5
2. MICE( Mus musculus)
•
Swiss albino mice are commonly used species
•
Other strains are – Balb/C and C-57
ADVANTAGES AND CHARACTERISTICS:
▫
Smallest
▫
Less drug required
▫
Easy to handle
▫
Cheap
3. GUINEA PIGS ( Cavia porcellus)
ADVANTAGES AND CHARACTERISTICS:
▫
Docile animals
▫
Highly susceptible to TB and anaphylaxis
▫
Highly sensitive to histamine, penicillin
▫
Required exogenous vitamin C in diet
4. RABBITS (Lupas cuniculus)
ADVANTAGES AND CHARACTERISTICS:
▫
Docile animal with large ears
▫
New Zealand white strains are widely used
▫
It has huge caceum and long appendix
▫
Enzyme atropine esterase is present in rabbit liver and plasma so it can tolerate
large doses of belladona (atropine)
5. FROGS (Rana tigrina)
ADVANTAGES AND CHARACTERISTICS:
▫
Used before 200 years
▫
Easily available during rainy season
▫
Amphibian animal and safe to handle
▫
Cannot breed in laboratory
6
LAB 2: STUDY OF DIFFERENT STAGES OF ANESTHESIA
AIM: To study the different stages of anesthesia and calculate onset and duration of action of
different anesthetic agent.
ANIMALS: Wistar albino rats
APPARATUS: Desiccator, Syringes, needle
DRUGS AND SOLUTIONS: Diethyl ether, chloral hydrate
BACKGROUND INFORMATION:
General anesthesia was absent until the mid-1800’s. William Morton administered ether to a
patient having a neck tumor, removed at the Massachusetts General Hospital, Boston, in
October 1846. The discovery of the diethyl ether as general anesthesia was the result of a
search for means of eliminating a patient’s pain perception and responses to painful stimuli.
Anesthesia is defined as partial or complete loss of sensation with or without loss of
consciousness as a result of disease, injury, or administration of an anesthetic agent, usually by
injection or inhalation.
STAGES OF ANESTHESIA:
 STAGE 1 (ANALGESIA/ONSET/INDUCTION):
 STAGE 2 (EXCITEMENT/DELIRIUM):
 STAGE 3 (Surgical Anesthesia):
 STAGE 4 (Impending Death/ Stage of Danger):
PROCEDURE:
Two Wistar albino rats will be weigh and keep in different cages. One rat will be injected by
intraperitoneal route with chloral hydrate in a dose of 400 mg/kg and another rat will be
inhaled by diethyl ether. Onset and duration of action of both anesthetic agents will be
recorded. Above mentioned different stages of anesthesia will be observed.
7
RESULTS:
S. No.
Name of Anesthetic agent
Onset of action
CONCLUSION:
8
Duration of Action
LAB 3: STUDY OF SOME BASIC INSTRUMENTS USED FOR ISOLATED
TISSUE EXPERIMENTS
AIM: To study the basic instruments used for in-vitro experiments.
BACKGROUND INFORMATION:
 Isolated tissue preparations are commonly used to study the effects of drugs on specific
type of receptors. These preparations are also used:
 For bioassay of drugs,
 Characterization of specific receptor or its subtypes,
 To determine concentration response curve of an agonist,
 To study antagonism of drug and in new drug discovery.
 The in vitro isolated preparation represents an isolated organ or a piece of living tissue
from a freshly killed animal.
 Optimum ionic environment
 Adequate supply of nutrition and oxygen
 And a stable temperature
 These basic requirements should be provided if one has to maintain the isolated
tissue in living state.
BASIC REQUIREMENTS:
 Physiological Salt Solutions
 Instrumentation
 Procedures and drugs – to render the animal unconscious
 Tissue: isolated or whole
PHYSIOLOGICAL SALT SOLUTIONS:
 Optimal pH 7.0 – 7.2
 Commonly used PSS are:
 Frog Ringer (for heart, rectus abdominis and other preparations of frog)
 Tyrode (for guinea pig ileum, rat ileum, rabbit ileum etc.)
 De Jaon(for rat uterus)
 Kreb’s solution (for rat fundus strip, tracheal preparations, vas deferens) etc.
ISOLATED ORGAN BATH:
 The apparatus providing the basic requirements of life to the tissue and facilitating the
record of response.
 Outer water bath of perspex or glass
9
 Inner organ bath (single or multiple)-glass (15-100 ml): Organ tube
 Tissue holder cum Aeration (oxygen) tube
 Glass coil connected to organ bath, Marriott's bottle:
Reservoir for PSS
 Electric heater
 Preheating coil
Sherrington Recording Drum and Drum Cylinder:
 It is an instrument on which physiological responses are recorded.
 Heavy base
 Base hoofs (legs) with adjustable leveling screws
 Gear rod
 Vertical shaft
 Drum Cylinder
 Brass or iron
It has 8 variable speeds. The speed is measured in mm per second. Eight variable speeds are 0.
12, 0.25, 1.25, 2.5, 12.5, 25, 320 and 640 mm per second.
Levers:
 Isometric contractions: change in tension (force). E.g. spring
lever
 Isotonic contractions: change in length at uniform tension.
E.g. isotonic frontal writing lever (light aluminum or stainless
steel rod)
Useful Hints for Students:
 Step by step
 Setup the assembly first
 Assembly should be on right hand side and drum should be on left hand side
 The drum should move from right to left or away from the writing point
 The speed of drum should be minimum
 The writing lever should be horizontal and the writing point should touch the drum.
 All tracing should begin after the overlap of the paper.
 Always start with the minimum concentration of drug.
 Write the name & amount of drug. Name of experiment and student with student ID
and signature of teacher before varnishing.
10
LAB 4: ROUTES OF DRUG ADMINISTRATION TO LABORATORY
ANIMALS
AIM: To demonstrate different routes of drug administration using Wistar albino rats.
ANIMALS: Wistar albino rats
APPARATUS: Disposable needle and syringes
DRUGS AND SOLUTIONS: Normal saline
BACKGROUND INFORMATION:
The possible routes of drug entry into the body may be divided into following classes:
I. ENTERAL ROUTE
1. Sublingual: under the tongue
2. Oral administration (P.O.)
3. Rectal or vaginal
II. PARENTERAL ROUTE: administration of medications by needle
1. Intravenous (I.V.): into vein
Fastest
2. Subcutaneous (S.C.): in the subcutaneous tissue
Slowest
3. Intramuscular (I.M.)
Medium
4. Intraperitoneal (I.P.) : into the peritoneum (body cavity)
5. Intraarterial: direct inject into artery
6. Intradermal: under the epidermis or into dermis
7. Intraosseous: into the bone
III. PULMONARY ROUTES: Inhalation into lungs
IV. TOPICAL
I. Nasal
II.
Skin
III.
Eye
11
LAB 5: STUDY OF ANALGESICS BY CHEMICAL METHOD
AIM: To evaluate the analgesic activity of drugs by using formalin test.
Chemical Method: Formalin induced writhing test
Animals: Wistar albino Rats
Apparatus: Syringe (with 100 divisions) 26g needle, stop watch
Drug & Solutions:
Aspirin
100 mg/kg, p.o.
Formalin
10 %, 0.1 ml
Normal Saline
0.9%
BACKGROUND INFORMATION:
 Analgesics: painkillers, a drug that selectively relive pain by acting in the CNS and
peripheral pain mechanism without significantly altering consciousness.
 Types of analgesics
1. Opioid or narcotic analgesics: relive visceral pain.
Uses: deep pain e.g. Cancer, myocardial infarction, anginal pain
Examples: 1- natural (as morphine, codeine) 2- semi synthetic (heroin) 3- synthetic
(pethidine) 4- endogenous opiates (endorphins & encephalin)
2. Non opioid or non-narcotic analgesics: relieve somatic pain.
Uses: dull pain e.g. Headache, toothache, backache
Examples: aspirin, paracetamol, diclofenac, piroxicam
PROCEDURE:
Rats weighing 180-300 gm will be administer with 0.1 ml of 10%
Formalin into the dorsal portion of the front paw. The test drug is
administered simultaneously either s.c. or orally. Each individual Rat is
placed into a clear plastic cage for observation. Reading is taken for a
period of 60 minutes. Pain response is indicated by elevation or favoring
of paw. Analgesic response is indicated if both paws are resting on the
floor with no obvious favoring of injected paw.
12
PAIN RATING SCALE:
0
1
2
3
Both forepaws are
placed on the floor
and weight is evenly
distributed.
The injected paw is
rested lightly on the
floor or on another
part of the animal
body and little or no
weight is placed on
it.
The injected paw is
elevated and not in
contact with any
surface.
The
uninjected paw is
placed firmly on
floor.
The injected paw is
licked, bitten or
shaken while the
uninjected paw is
not.
Attempt to sleep by
curling up with both
paws off the floor is
also given this rating.
Attempts to sleep off
by curling up with
only the injected
paw off the floor,
even when it is
tucked under the
body is also given
this rating.
Normal
grooming
(during which both
paws are elevated),
licking and rearing
(Forepaws are not in
contact with floor,
although forepaws
may be in contact
with the wall).
During locomotion
there
is
no
discernible favoring
of the injected paw
This behavior is quite
distinct from normal
grooming although
transition between
the two is common.
During locomotion
there is not obvious
limp.
EVALUATION:
 Shows the prolongation of latency time by comparing value before and after
administration of test compound.
 Comparison done by statistical analysis using t-test.
13
RESULTS:
RESOURCES AND HINTS FOR TEACHERS:
1. Check work books after students calculate the volume of drug to be injected. Check
volumes in syringes prior to injection. Restrain the animal for the student to inject. Timings
of injections should be noted. Only one animal to be observed at a time.
2. Formalin should be freshly prepared for each class.
3. The experiment can be completed in 90 minutes. This can be followed by a small group
discussion on evaluation of analgesics in humans.
14
LAB 6: TO EVALUATE THE ANALGESIC POTENCY OF DRUG BY TAIL
FLICK METHOD
AIM: To evaluate the analgesic activity of drugs by using Tail flick analgesiometer.
Method: Thermal
Animals: Wistar albino Rats
Apparatus: Oral feeding needle, Tail flick analgesiometer, Mouse restraint
Drug & Solutions:
Aspirin
100 mg/kg, p.o.
Normal Saline
0.9%
BACKGROUND INFORMATION: The tail flick assay is a pain
receptive assay in which a mouse is placed within a
restraining tube with its tail protruding.
PRINCIPLE: Record the response of an animal to a painful stimulus before and after
administration of analgesics.
PURPOSE: The method is based on the reaction of the rat to heat stimulus applied to a small
area of the tail. The time until this response occurs is prolonged after administration of
analgesics.
PROCEDURE:
Albino rats of either sex weighing between 100-150 g are used. The
rat is held and its tail is placed on a level surface, a radiant heat is
applied to the tail at a point not more than 3 cm from its tip. After
an interval the animal withdraws its tail with a sudden and
characteristic flick. Time of flicking is recorded, before drug
administration. The reaction time are thereafter recorded after 15,
30, 45 and 60 minutes after administration of the test drug
15
OBSERVATIONS TABLE:
Reaction time in second
Rat No.
Control
Minutes after drug administration
15
30
45
60
1.
2.
3.
4.
5.
6.
EVALUATION:
 Shows the prolongation of latency time by comparing value before and after
administration of test compound.
 Comparison done by statistical analysis using t-test.
RESULTS:
RESOURCES AND HINTS FOR TEACHERS:
1. Check work books after students calculate the volume of drug to be injected. Check
volumes in syringes prior to injection. Restrain the animal for the student to inject. Timings
of injections should be noted. Only one animal to be observed at a time.
2. The experiment can be completed in 60 minutes. This can be followed by a small group
discussion on evaluation of analgesics in humans.
16
LAB 7: TO EVALUATE THE ANALGESIC POTENCY OF DRUG BY HOT
PLATE METHOD
AIM: To evaluate the analgesic activity of drugs by using Hot Plate analgesiometer.
METHOD: Thermal
ANIMALS: Wistar albino Rats
APPARATUS: Oral feeding needle, thermostatically controlled Hot Plate
DRUG & SOLUTIONS:
Aspirin
100 mg/kg, orally
Normal Saline
2 ml/kg, orally
BACKGROUND INFORMATION: Eddy’s Hot Plate: Animal is placed on hot plate and time for
jumping from plate is noted before and after administration of drug.
Hind paw licking (4-6 sec) or jump response (2-3 sec)
PRINCIPLE: Record the response of an animal to a painful
stimulus before and after administration of analgesics.
PURPOSE: The paws of rats are very sensitive to heat at
temperature. So the responses are jumping, withdrawal of
paws and licking of the paws. The time until this response occurs is prolonged after
administration of centrally acting analgesics.
PROCEDURE: Wistar albino rats of either sex with weight of about 200-250 g. The hot plate consists
of electrically heated surface with controlled temperature 55±1°C. Animals are place on hot
plate and time of flicking or jumping is recorded by stop watch, before drug administration. The
latency time is recorded after 15,30,45,60 minutes following oral or subcutaneous
administration of the test compound
17
OBSERVATIONS TABLE:
Reaction time in second
Rat No.
Control
Minutes after drug administration
15
30
45
60
1.
2.
3.
4.
5.
6.
EVALUATION:
 Shows the prolongation of latency time by comparing value before and after
administration of test compound.
 Comparison done by statistical analysis using t-test.
 The value which is exceeding the value before administration for 50% or 100% can be
regarded as positive response.
RESULTS:
RESOURCES AND HINTS FOR TEACHERS:
1. Check work books after students calculate the volume of drug to be injected. Check
volumes in syringes prior to injection. Restrain the animal for the student to inject. Timings
of injections should be noted. Only one animal to be observed at a time.
2. The experiment can be completed in 60 minutes. This can be followed by a small group
discussion on evaluation of analgesics in humans.
18
LAB 8: SCREENING OF DIURETICS
AIM: To study the Effect of different Diuretics on the Production of Urine in Laboratory Animals
ANIMALS: Wistar albino Rats
APPARATUS: Oral feeding needle, metabolic cages, measuring cylinder, funnel
DRUG & SOLUTIONS:
Furosemide
(5 mg/kg; oral)
Urea
(900 mg/kg; oral)
Normal Saline
(2 ml/kg; oral)
BACKGROUND INFORMATION:
 Some of pathological conditions associated with retention of sodium and water in the body
e.g. Congestive Heart failure, pulmonary edema, renal edema and hypertension.
 Diuretics are the drugs that cause a net loss of Na+ and water in urine.
 The first important site with regard to renal function is the glomerulus itself. Drugs which
increase glomerular filtration rate (Xanthines, cardiac glycosides, saline) produces diuresis.
THE FORMATION OF URINE:
 In summary, three processes occurring in
successive portions of the nephron
accomplish the function of urine formation:
 Filtration of water and dissolved
substances out of the blood in the
glomeruli and into Bowman's capsule;
 Reabsorption of water and dissolved
substances out of the kidney tubules
back into the blood (note that this
process prevents substances needed
by the body from being lost in the
urine);
19
 Secretion of hydrogen ions (H+), potassium ions (K+), ammonia (NH3), and certain
drugs out of the blood and into the kidney tubules, where they are eventually
eliminated in the urine.
PRINCIPLE: Diuretics are the compounds which increase the flow of urine. Normal urine output
in rats is very small (1-2 ml/rat/day). Hence, to get the measurable quantity the animals are
first hydrated. The urine output is increased after administration of diuretics like urea and
furosemide. Increase in volume of urine is measured with the help of measuring cylinder and
compared with the normal urine output.
PROCEDURE: Albino rats of either sex weighing between 150-200 g are fasted (deprived of food &
water) overnight and saline is administered orally. These rats are divided into three groups as
follows:
1. Only normal saline
2. Saline + Urea
3. Saline + Furosemide
After administration of drugs, rats are placed in the three different metabolic cages. Urine is
collected in measuring cylinder. Time when the first drop of urine is collected in a cylinder for
each group is noted and the volume is recorded at intervals of 15 min for 3-4 Hours. The
difference in the volume collected at different time interval and total volume can be compared
with various diuretics.
OBSERVATIONS TABLE:
Amount of urine
collected (ml)
Groups
After 15 Min
After 30 Min
After 1 Hr.
Total Volume
I
II
III
EVALUATION: Calculate percentage increase in volume of urine taking control value as 100%.
RESULTS:
20
LAB 9: SCREENING OF ANTI-INFLAMMATORY DRUGS USING HIND
PAW METHOD
AIM: To Evaluate the Anti-inflammatory Activity of the Drug (acetyl salicylic acid) Using Hind
Paw Method
MODEL: Acute inflammatory
METHOD: Rat hind paw method
ANIMALS: Wistar albino Rats
APPARATUS: Oral feeding needle, Plethysmometer,
DRUG & SOLUTIONS:
Aspirin
(100 mg/kg; oral)
BACKGROUND INFORMATION:
Inflammation is a protective response to injury. It occurs in three phases:
(a) The first phase being oedema and swelling with accompanying pain. These effects are
produced as a result of the dilation and increased permeability of the blood vessels
(veins) due to the release of certain mediators such as histamine, serotonin and kinins
etc.
(b) In the second phase, leukocytes migrate to this area and mopping up operations starts.
(c) Second
phase
is
followed
by
repair,
which
is
ushered
in
by
the
proliferation of fibroblasts and synthesis of connective tissue.
The ability of a compound to reduce the local oedema induced in rat paw by various irritants is
the most widely used test to screen new non-steroidal anti-inflammatory drugs. Many
compounds like formalin, carrageenan, Kaolin, yeast and dextran have been used as irritants to
produce oedema.
PROCEDURE: Twelve healthy male albino rats weighing 100-200 gms will be selected and made into
two groups of six animals each. All the animals will be kept on fasting for 18 hours. The hind paw
of the rats will be marked at the level of tibio tarsal junction of hind leg, so that while
measuring the volume, the dipping will be done to the same level. One group serve as control,
21
0.3 ml of Normal saline will be given orally. Another group
will receive the test drug, Acetyl salicylic acid 300 mg/Kg.
After 30 minutes of the administration of drug, 0.1 ml of
1% Carrageenan will be administered to the rats into the
plantar surface of the right hind limb to induce paw
oedema. The volume will be measured immediately and
after
3
hours
using
plethysmometer.
The change in the paw volume was compared with the
control animals. The percentage of oedema compared to
the control by the test drug was calculated using the
formula
CALCULATION:
Vc - Vt
% oedema inhibition = ---------------
X 100
Vc
Where:
Vt- means increase in paw volume in rats treated with test drug;
Vc-means increase in paw volume in control group of rats.
OBSERVATION TABLE:
Group
Initial
Reading 3 hr after
Difference in
Percentage
Reading
carrageenan
volume
oedema
Control
Test
RESULT: The % oedema inhibition was found to be ______________
22