This document summarizes different types and methods of bioassay. There are two main types of bioassay: qualitative and quantitative. Qualitative bioassay determines the type of biological action, while quantitative determines the amount of active constituent. Common methods include direct methods like matching and bracket methods using frog rectus muscle, and indirect methods like graphical interpolation and 3-4 point bioassays which construct dose-response curves to calculate potency ratios. The principle of bioassay is to compare a test substance to a standard preparation and determine the dose needed to produce the same biological effect.

1. NETES Institute of Pharmaceutical Sciences, Mirza
Presented by-
Jubair Sikdar
Roll no-04
M. Pharm 2nd semester
Department of Pharmaceutical Chemistry
BIOASSAY

2. CONTENTS
 Principle
 Types of bioassay
 Methods of bioassay

3. Bioassay
Definition:
Bioassay is defined as the estimation of the potency of an active principle in a unit
quantity of preparation or detection and measurement of the concentration of the
substance in a preparation using biological methods.
Principle:
The basic principle of bioassay is to compare the test substance with the International
Standard preparation of the same and to find out how much test substance is required
to produce the same biological effect, as produced by the standard.

4. Types of Bioassay
There are two types of Bioassays. They are
(i) Qualitative Bioassay.
(ii) Quantitative Bioassay.
Qualitative bioassay:
The objective is to determine what type of action a drug produces, by comparing its
action with those of other substance of known activity. It can be used to detect small
amounts of biologically active substance present at disease sites or released in
pathology.
Quantitative bioassay:
The objective is determined as the amount if active constituent present in the drug
preparation, for this purpose another preparation containing a known amount of active
constituents is used as standard. Quantitative Bioassay is typically analyzed using the
methods of biostatistics.

5. Methods of bioassays
(A) Direct methods:
1. Matching method
2. Bracket method
(B) Indirect methods:
1. Graphical or interpolation bioassay.
2. 2. Three point bioassay
3. Four point bioassay
(A) Direct methods:
1. Matching method: This test is performed by using frog's rectus abdominis muscle
which placed in organ bath.
• Main aim of this method is to find out potency (or concentration) of the test sample by
comparing the doses of standard acetylcholine and test acetylcholine.

6. • In this method, Frog rectus abdominous muscle is placed in organ bath, with that DRC
(Dose Response Curve) of drug (standard drug after that test drug) is carried out at
various doses.
2. Bracket method:
• Bracketing Bioassay method is performed by selecting two standard doses, which will
give a close bracket between lower dose and higher dose of standard when test dose is
given.
• The method also carried out on frog rectus abdominous muscle.
• By taking of DRC of drug (for both standard and test) select two response of standard
such that response to unknown dose is between these responses.

7. • The potency of drug is calculated by using following equation.
Potency of drug =
𝑆1+𝑆2
2
mg/ml or µg/ml
where S1 = Standard dose of 1
S2 = Standard dose of 2
(B) Indirect methods:
1. Graphical or Interpolation bioassay:
• This method is less time consuming and get reliable as compared to matching type of
bioassay.
• Main advantage of this method is that the sensitivity of the tissue is determined by
prior plotting of concentration response curve with known against as is the case with
standard drug.

8. • Three point bioassay:
• This method is performed on isolated frog rectus abdominous muscle.• In this method
two standard doses were selected which will give close bracket on either side of
response produced by unknown. The response of 2 doses of standard ( S1, S2) the test
is obtained by changing the order in successive cycle.
• The potency of the substance is determined by using the following formula.
• Potency of test solution= S
𝑛1
𝑡
𝑎𝑛𝑡𝑖𝑙𝑜𝑔[
(𝑇1−𝑆1)
𝑆2−𝑆1
]log
𝑛2
𝑛1
Where n1= Lower standard dose
n2-Higher standard dose
S1 and S2 = Standard responses
t = Test does (ml)
T1= Test response

9. Four point bioassay:
In four point bioassay comprises are based on analysis of dose response curve and the
matching dose of standard and unknown are calculated by following formula.
Potency ratio:
𝑥1
𝑥2
𝑎𝑛𝑡𝑖𝑙𝑜𝑔[
𝑇1−𝑆1 +(𝑇2−𝑆2)
𝑇2−𝑇1 +(𝑆2−𝑆1)
log
𝑥1
𝑥2
Where, x1= Lower dose of standard drug
x2= Higher dose of standard drug
y1= Lower volume of test
S1, S2, T1, T2= Mean responses of standard and test

10. Reference
1. Srinivasa A. and Lakshmi N.: Pharmacological Screening Methods and Toxicology,
First edition, 2022:101-115

11. OPEN TO DISCUSSION

12. THANK YOU