anti inflammatory test
DESCRIPTION
credit also given to our lecturer,demonstraters, and my lovely friends -Nur Aqilah -Anis Ayuni -NurhayatiTRANSCRIPT
MINI PROJECT: ANTI-INFLAMMATORY TEST
PHARMACOLOGY & TOXICOLOGY UNIT.
Group Members↝ Anis Ayuni bt. Ismail (170607)↝ Nur Aqilah bt. Fauzi (170646)↝ Nurfarah Aqilah bt. Ahmad Nizam (170657)↝ Nurhayati bt. Mohd Yusoff (170921)
IntroductionANTI-INFLAMMATORY•Inflammation is the body's attempt at self-protection to remove harmful stimuli, including damaged cells, irritants, or pathogens - and begin the healing process. •The inflammatory response is usually quantified by increase in paw size (oedema) and is modulated by inhibitors of specific molecules within the inflammatory cascade.•In this experiment, we want to determine the anti-inflammation activity of Ficus Deltoidea in relieving the inflammation, so carrageenan-induced paw oedema experiment was conducted.•There are three groups:
1. Treatment group: Ficus Deltoidea
2. Positive group: Indomethacin
3. Negative group: Saline water
• Our group is a control or negative group.
• We gives the rat saline water by force feed.
• It is a procedure that differentiate the three group in this experiment.
• Every groups injected 0.1ml of 1% carrageenan and saline water respectively whether in left or right hind paw onto subcutaneous of plantar surface to induce oedema.
• The injection of Carrageenan and saline water will caused the inflammation the rat’s paws.
• The increase in percentage of paw volume was calculated based on the volume difference between the normal and abnormal paws (with or without carrageenan injection).
Introduction
Methods
1% carrageenan, 100 mg/kg plant extract, 10 mg/kg indomethacin in saline were prepared.
A line was marked above the ankle joint of the hind
limbs.
The solutions were administered to each
respective rats.
Both hind paws were then dipped
into small container of plethysmometer
to measure base volume.
After 30 minutes, subcutaneous injection
of 0.1 ml of 1% carrageenan was
performed onto plantar surface of right hind paw
of all rats.
Another subcutaneous injection of 0.1 ml
saline was performed onto plantar surface of
left hind paws.
Volume of both hind paws in all group was measured
on the next 30 minutes with 30 minutes interval
for 5 hours.
Hind paw swelling was calculated using the given
formula.
A. Carrageenan Induced Oedema Using Plethysmometer.
B. Carrageenan Induced Hyperalgesia Using Plantar Test.
After measuring base volume, the rats were
placed inside the container on Plantar Test.
Thermocouple was placed under the heels of
both hind paws, and withdrawal latencies
were recorded as base line.
Withdrawal latencies of both hind paws after
subcutaneous injection was performed were also
recorded.
The readings were recorded every 30
minutes interval for 5 hours.
Graphs for both experiment were plotted.
* Anti-hyperalgesia was detected when the right paw prolong the withdrawal time compared to the baseline values.
Plethysmometer.
During Plantar Test.
Results
RESULTSA)
Time (minutes) Right Paw Volume (Carrageenan)
Left paw Volume(Saline)
-30 1.29 1.23
0 1.85 1.63
30 1.9 1.53
60 2.25 1.76
90 2.50 2.28
120 2.34 1.90
150 3.06 2.02
180 3.14 2.32
210 2.42 1.54
240 2.02 1.69
Formula
% swelling = Vr – Vr0 - Vl – Vl0 X 100
Vr0 Vl0
Vr = Right Paw Volume
Vr0 = Right Paw initial Volume
Vl = Left Paw Volume
Vl0 = Left paw initial Volume
0 50 100 150 200 250 3000
10
20
30
40
50
60
70
80
Graph of percentage of Hind Paw swelling Against Time%
Sw
ellin
g
ResultsB)
Time (minutes) Right Paw ( Carrageenan) Left Paw ( Saline)
-30 14.9 11.9
0 17.5 15.1
30 15.6 7.4
60 26.7 10.9
90 13.9 11.9
120 14.9 14.6
150 14.1 8.9
180 9.9 7.3
210 9.9 6.4
240 8.9 7.8
Graph of Withdrawal Latencies vs Time (min)
-50 0 50 100 150 200 250 3000
5
10
15
20
25
30
Right Paw (Carrageenan)
Left Paw (Saline)Wit
hdra
wal
Lat
anci
es
Time (min)
Data Analysis
% S
wel
ling
Time (Minutes)
Data Analyses
• The paw swelling ratio is the percent increase of paw volume. The time-dependent curve
has shows that the injected carrageenan had cause the paw to swell.
• The graph show that the swelling have two phase in the given time. The different
phases(two different peaks) indicates cascade of biochemical events propagates to
matures the inflammatory response.
Graph of Withdrawal Latencies vs Time (min)
Time(min)
With
draw
al L
atan
cies
Data Analyses
• The graph shows the withdrawal latencies of rat’s hind paw.
• The inflamed paws by carrageenan have greater initial resting paw
temperatures than the paws receiving injections of saline. Also,
stimulation of the inflamed paws results in longer paw withdrawal
latencies than stimulation of saline-treated paws, and this longer latency
• There is some error in our result in which actually the theory that we found is the
inflamed paw should result in shorter paw withdrawal latencies.
Discussion
• The inflammatory effect was evaluated by carrageenan induced hind paw oedema. Carrageenan-induced paw oedema has been commonly used as an experimental animal model for acute inflammation and is believed to be biphasic.
• The oedema was induced by injection of 1% solution of carrageenan into rat’s plantar region. Different group of animal were administered with different solutions which are plant extract (100mg/kg) for treatment group, 1ml saline for negative control negative control group and 10mg/kg indomethacin for positive control group.
• Rat’ paw volume were measured by Plethysmometer. By using plantar test that measures response to infrared heat stimulus after the subcutaneous injection were performed, anti hyperalgesia can be detected.
• The graph show the value increased and decreased and have two phase.
• The early phase of the carrageenan model is mainly mediated by histamine, serotonin and increased synthesis of prostaglandins in the damaged tissue surroundings. While the late phase involved prostaglandin release mediated by the tissue macrophages.
• Apparatus used to measure the oedema:
• Plethysmometer
• Can measure the small volumes changes that occur.
• The water displacement produced by the immersion of the animal paw in the measuring tube is reflected into the second tube, inducing a change in the conductance between the two platinum electrodes.
• Plantar test
• Enables to discern the oedema to thermal stimulation caused by inflammatory in the unrestrained rat.
Conclusion
• The volume of paw that have two phases indicates the cascade of the
different biochemical compound in body to respond to the inflammation.
• The inflamed paw has higher threshold temperature.
• Our part of experiment show the unstable result maybe influenced by
surrounding.
• Why must inject carrageenan and saline water?
• It is because carrageenan will cause the reaction of inflammation(oedema) by stimulating the release of specific antibody in the blood, while the saline water will not stimulate the reaction of inflammation but the inflammation still occur because of the tissue damage when we injecting the solution. Actually, saline water act as a blank because we can measure the inflammation(oedema) caused by cell injury and we can find the inflammation(oedema) caused by carrageenan compound
Thank You.