The document describes screening methods used to evaluate potential anti-inflammatory drugs. It discusses both acute and chronic inflammation screening approaches. For acute inflammation, methods described include carrageenan-induced paw edema in rats, croton oil-induced ear edema, and oxazolone-induced ear edema, which measure increased swelling at the site of inflammation. Chronic screening models involve cotton wool or glass rod implants, which induce granuloma formation over longer periods. The document provides details on procedures and evaluations for several common preclinical screening methods used to assess anti-inflammatory activity.

1. KARNATAKA COLLEGE OF PHARMACY
DEPARTMENT OF PHARMACOLOGY
PHARMACOLOGICAL AND TOXICOLOGICAL SCREENING METHOD
PRESENTATION ON
SCREENING METHOD OF ANTIINFLAMMATORY DRUGS
Prepared by sudarshan singh
M pharmacy ( dept; pharmacology)

2. Inflammation is defined as local response of living
mammalian tissues to injury due to any agent. It is a
defense reaction in order to eliminate or limit the spread
of injurious agent, followed by removal of the necroses
cells and tissues.

3. Agents causing inflammation
1. Infective agents: bacteria, virus, and other toxins,
fungi
2. Immunological agents: cell mediated and antigen-
antibody reaction
3. Physical agents: heat, cold, radiation, mechanical
trauma
4. Chemical agents: organic and inorganic poisons
5. Inert materials: foreign bodies

4. Phases of inflammation
 Acute: vasodilatation & increased capillary
permeability
 Delayed: infiltration of leukocytes and
phagocyte cells
 Chronic proliferative:
tissue degeneration and fibrosis

5. Steps of the inflammatory
response
(1) Recognition of the injurious agent
(2) Recruitment of leukocytes
(3) Removal of the agent
(4 ) Regulation of the response
(5) Resolution

6. Different types of inflammation
 1 Acute and
 2 Chronic inflammations
Cardinal sign of inflammation
 1 redness (rubor)
 2 pain (dolor)
 3 hot ( color)
 4 swollen (tumour)
 Loss of function (function losis)

7. Comparison between acute and
chronic inflammation:
Types Acute Chronic
Causative agent Bacterial pathogens,
injured tissues
Persistent due to non-
degradable pathogens,
viral infection,
Major cells involved neutrophils ,basophils,
and eosinophils
monocytes,
macrophages,
lymphocytes, plasma
cells), fibroblasts
Primary mediators Vasoactive amines,
eicosanoids
IFN-γ and other
cytokines, growth
factors,
Onset Immediate Delayed
Duration Few days Up to many months, or
years
Outcomes Resolution, abscess formation,
chronic inflammation
Tissue destruction,

9. Classification of anti inflamentary
drugs
1 Salicylates
Aspirin (acetylsalicylic acid), Salicylic acid and
other salicylates , Salsalate
2 Propionic acid derivatives
 Ibuprofen ,Dexibuprofen ,Naproxen
3Acetic acid derivatives
 Indomethacin ,Tolmetin ,Diclofenac
4Sulfonanilides
 Nimesulide

10. 5 Acetic acid derivatives
 Indomethacin ,Tolmetin ,Diclofenac
6 Enolic acid (Oxicam) derivatives
 Piroxicam ,Meloxicam ,Tenoxicam,Phenylbutazone
7 Selective COX-2 inhibitors
 Celecoxib, Rofecoxib,Etoricoxib
8 Anthranilic acid derivatives
 Mefenamic acid
 Meclofenamic acid
 Flufenamic acid
 Tolfenamic acid

15. The percentage of HRBC membrane stabilization or protection was
calculated using the formula:
% 100- OD of test solution –OD of product control x 100
 OD of test control

20. Result for In-vitro study (Table 1) :
 The extract at concentration range of 6.3 -100 mg/ml protect the
human erythrocyte membrane against lysis induced by hypotonic
solution.
 At concentration of 100µg/ml, the extract produced 47.36 %
inhibition of RBC haemolysis as compared with 48.14% produced by
diclofenac sodium.

21. Result for In-vivo study (Table 2) :
 The extract at concentration 200 mg/kg and 400 mg/kg protect the human
erythrocyte membrane against carrageen induced.
 At concentration 200 mg/kg and 400 mg/kg, the extract produce 36.02 % and
42.63 % inhibition of inflammation as compared with 58.13 % produced by
diclofenac sodium.

22. Types of screening methods
 Acute(or)transient phase: In this phase vasodilation and
increased capillary permeability are observed.
 Sub acute phase : Infiltration of leucocytes and
phagocytes in blood.
 Chronic inflammatory phase: granuloma formation is
observed in this phase.

23. ACUTE PHASE
 Acute phase: The methods that include acute phase
are as follows:
 Carrageenan induced paw oedema in rats
 Croton-oil induced ear edema
 Oxazolone induced ear edema
 U V erythema in guinea pigs
 Pleurisy in rats
 Granuloma air pouch technique
 Vascular permeability

24. CHRONIC PHASE
The methods that include this chronic phase are:
 Cotton wool granuloma
 Glass rod granuloma
 Sponge implantation technique

25. ACUTE METHOD
 1 Carrageenan induced paw oedema in rats
 2Croton-oil induced ear edema
 3Oxazolone induced ear edema in mice

26. Carrageenan induced paw oedema in rats
 a body weight between 150 and 170g are used . the
animals are starved for overnight .
 Thirty minutes later, the rats are challenged by a
subcutaneous injection of 0.05ml of 1% solution of
carrageenan
 The paw is marked with ink at the level of lateral
malleolus and immersed in the mercury up to this
mark.

27.  EVALUATION
 The paw volume was measured using plenthysmometer at 3
and 6 hr
 The increase in paw volume at 3 and 6 hr is calculated
 The percentage increase in paw volume was measured by
comparing the difference of the average values between
treated group animals and control group
 A dose-response curve is run for the active drugs and ED50
values can be determined.

28. 2Croton-oil induced ear edema
 Both rats and mice are used
 In this methodology 75microg of croton oil is applied
to the inner surface of the right ear of each mouse.
 The animals are previously, anaesthetised with diethyl
ether
 Varying dose levels of test drug are applied to inner
surface of the right ear of each mouse inducing
inflammation.

29.  The animals are sacrificed by cervical dislocation, and a
plug of 8mm diameter is removed from each of the ear.
 The difference in weight between the two plugs is taken as
the measure of edematous response

30. Evaluation:
 This methodology is mainly used to detect antiinflammatory
activity of steroids.
 The % antiinflammatory activity is calculated by,
% anti inflammatory activity = ( wt of treated ear-wt of
untreated ear/wt of control ear)*100

31. CHRONIC METHOD
1 Cotton wool induced granuloma
2 Glass rod granuloma technique

32. Cotton wool induced granuloma
 Male rats weighing about 180-200g are used.
 Drugs are administered orally once on a dosage regimen for
7 days and the control group received the vehicle.
 Two sterilized pellets of cotton wool were implanted
subcutaneously, one on each side of abdomen
 The rats were sacrificed on the 8th day, the implanted pellet
was dissected out and the wet weight was recorded
 Both of these were dried at 600 c for 18 hrs and the dry
weight was recorded.

33. Evaluation:
Evaluation:
 The weight of transudate and the granuloma as well as
the percent granuloma inhibition of the test drugs
were calculated.
 The body weight gain was also recorded.

34. Glass rod granuloma technique
Methodology:
 In this technique, glass rod with a diameter of 6mm and 40
mm length are selected.
 Male sprague-dawley rats with an initial weight of 130g are
selected.
 From an incision in the caudal region a subcutaneous tunnel
is formed in the cranial direction with a closed blunted
forceps.
 The glass rod is introduced in to this tunnel which lies on the
back of the animal.The incision wound is closed by sutures.

35.  The animals are kept in separate cages, the rods remain
insitu for 20 or 40 days.
 At the end, the animals are sacrificed under CO2
anaesthesia.
 The glass rod is prepared with surrounding connective tissue
which forms a tube around the glass rod.
 Wet weight of the granuloma tissue is recorded, finally it is
dried and the dry weight is also recorded

36. Evaluation:
 Granuloma weight reduced by the test compound is
compared with that of the standard.

37. Be miserable.
Or motivate yourself.
Whatever has to be
done, it's always your
choice.