· INTRODUCTION
Bioassay of histamine using guinea pig ileum by matching method.
The bioassay of histamine using the guinea pig ileum is a pharmacological method to evaluate histamine's biological activity. The guinea pig ileum, a smooth muscle, responds to histamine by contraction, which is used as an indicator of its potency. In this bioassay, tissue preparations from the ileum are exposed to various concentrations of histamine, and the degree of contraction is measured. The method helps in determining the concentration-response relationship and the potency of histamine or histamine antagonists. It is commonly used for evaluating histamine's role in allergic reactions or testing the efficacy of antihistamine drugs.
· EQUIPMENT REQUIRED
Animal :- Guinea pig
Drug:- Histamine solution (10 μg/ml)
Instrument:- Student Organ Bath, kymograph.
Physiological salt solution:- Tyrode’s solution
· PRINCIPLE
The principle of the bioassay of histamine using the guinea pig ileum is based on the contraction of smooth muscle tissue in response to histamine. When histamine is introduced to the isolated guinea pig ileum, it binds to histamine receptors (H1 receptors) on the smooth muscle, causing a series of biochemical events that lead to muscle contraction. The degree of contraction is proportional to the concentration of histamine. By comparing the response of the ileum to known concentrations of histamine, the potency of the substance being tested can be determined. This method is used to assess the physiological effects of histamine and evaluate the efficacy of antihistamines.
PROCEDURE:
1) Isolate guinea pig ileum tissue and mount in organ bath
2) Tie ends to aeration tube and isotonic lever.
3) Stabilize tissue and allow it for 30min to stabilize and at interval of every 10min change PSS solution
4) Take baseline of 30sec and keep contact time of 60sec after recording effect of histamine wash tissue for 60sec Two times.
5) Maintain total time cycle of 180sec.
6) Add histamine dose 0.4mL and take baseline of 60sec
7) Maintain contact time of 60sec and washing period of 60sec twice.
8) Total time cycle of 180sec
9) After that add test drug (T1) 0.1mL and take baseline timing for 30sec
10) Maintain contact time of 60sec and washing period of 60sec twice.
11) Total time cycle 180sec.
12) After that add test drug (T2) 0.2mL and take baseline timing for 30sec
13) Maintain contact time of 60sec and washing period of 60sec twice.
14) Total time cycle 180sec.
15) Then add histamine (S1) 0.1 mL.
16) Maintain contact time of 60sec and washing period of 60sec twice.
17) Total time cycle 180sec.
18) Then add histamine (S2) 0.15 mL.
19) Maintain contact time of 60sec and washing period of 60sec twice.
20) Total time cycle 180sec.
CALCULATION & INTERPRETATION:
A1*X = A2 * Y
A1 = Conc. of Test
X= Dose of Test Selected
A2 = Conc. Of standard
Y= Dose of standard that matches with the response height of dose of test.
A1 = ( A2 * Y)/ X
In this case the
T2* 0.2 = S2 * 0.15
T2 = (0.15 * S2 ) /0.2
T2 = (0.15 * 10 ) /0.2
T2 = 7.5 μg
So, the conc. Of the test histamine is 7.5 μg
CONCLUSION:
In conclusion, the bioassay of histamine using the guinea pig ileum is a reliable and effective method to study the pharmacological effects of histamine on smooth muscle contraction. It provides valuable data on histamine’s potency and allows for the evaluation of antihistamines or other compounds that may modulate histamine activity. The contraction response of the ileum serves as an indicator of histamine's biological effects, and through careful analysis of the concentration-response curve, researchers can quantify histamine’s activity and assess the therapeutic potential of histamine-related drugs.
IDEAL OBSERVATION
Sr. No.
Conc. Of histamine
(μg/ml)
Amoun1t added in organ2 bath
μg/ml (Organ bath contains 20ml sol.)
Response
(in mm)
% Response
In m4L
In μg
1.
10
0.1
1
0.05
7
16.66
2.
3.
0.2
2
16
38.09
4.
0.4
4
22
52.38
5.
0.8
8
34
80.95
6.
1.6
42
100.00
7.
3.2
32
Matching Bioassay
Dose (ml)
Drug Used
Response (mm)
Histamine (10 μg – sub max response
Test Drug (T1)
Test Drug (T2)
12
Histamine (10 μg) S1
0.15
Histamine (10 μg) S2
· RESULT:
The result of a histamine bioassay using the guinea pig ileum typically includes a concentration-dependent increase in the contraction of the ileum as histamine is administered. The response is measured by recording the amplitude or frequency of muscle contractions at various histamine concentrations. A clear dose-response curve is often generated, showing the relationship between histamine concentration and the degree of contraction. The potency of histamine can be determined by the minimum concentration required to elicit a noticeable response. Additionally, if antihistamines are tested, the inhibition of contraction at varying doses of the antagonist can be evaluated to assess its efficacy in blocking histamine’s effects.
DISCUSSION:
The guinea pig ileum bioassay effectively measures histamine's ability to induce smooth muscle contraction, demonstrating a dose-dependent response. This assay is valuable for assessing histamine potency and evaluating the effectiveness of antihistamines. Variations in results can arise due to factors like tissue preparation and experimental conditions. Despite newer techniques in pharmacology, this bioassay remains a reliable method for studying histamine's physiological effects and the efficacy of histamine-related drugs.
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