Determination of LD 50

and ED50
Grace Marie C. Alunan, RPh, CPS, MSc
Definition of Terms

Lethal Dose
- A dose at which a given percentage
of the subjects die. Usually expressed
as median lethal dose
Median Lethal Dose (LD 50)

- The dose required to kill half

member of the tested population after
a specific test duration
Definition of Terms

Effective Dose
- A dose at which a pharmacologic
agent will exhibit therapeutic effect

Median Effective Dose (ED50)

- The amount or dose required to
produce a response in 50% of the
population
Brine Shrimp

 A general assay to determine the LD50 of

a test solution
 Usually employed in the lethality of
pesticides for environmental toxicological
testing of invertebrates
 Provides estimation of toxicant that may
potentially contaminate aquatic ecological
system
OECD Fixed Dose
 Aims to identify the appropriate hazard
class for new chemicals
 It does not provide a point of estimate of
the LD 50
 Animals are tested sequentially at one of the 4
doses (5, 50, 300 or 2000mg/kg BW)
 Once clear sign of toxicity appear, additional
animals are dosed at the same level for a total
of 5 animals
 Subsequent groups of animals may receive a
dose higher or lower, depending on the outcome
of the previous group
Toxicological Testing
Brine Shrimp
OECD Fixed Dose
OECD Acute Toxic Class
OECD Up and Down
Acute oral testing/ Classical
LD50
OECD Acute Toxic Class

Provides a range for lethality rather

than a point of estimate
 Group of 3 animals receive one of the 4 or 5
doses 5,50,300 or 2000mg/kg
 Depending on the survival or mortality of the
first group of animals, 3 or more animals may
receive the same or higher or lower dose
 The number of animals that survive or dies
determines the classification decision
OECD Up and Down

 Employs a sequential dosing, using a

single animal at each step
 The dosage depends on whether the
previously dosed animal lives or dies
 Provides a point of estimate of lethality
and confidence interval and can be used
to evaluate lethality up to 5000mg/kg
Classical Median Lethal Dose

 Logarithmic expansion from a starting

dose
 Starting dose is usually 10mg/kg BW
 Each dose level necessitates the use of a
group of animals; if many dose level are
to be tested, then the classical LD 50
presents the following disadvantages
 In expensive
 Time consuming
Linear Regression Method
Dose Log # Mice % Probit
(mg/kg Dose Responded (Y1) Value
BW) (Y2)
215 2.3324 0/9 0 -
454.36 2.6574 2/9 22.22 4.2345
960.16 2.9824 3/9 33.33 4.5684
2,029.55 3.3074 4/9 44.44 4.8592
4,289.43 3.6324 5/9 55.55 5.1383
9,065.67 3.9574 9/9 100 8.7190

Log Dose interval 0.325

Linear Regression Method

Y = xb + a

 Equation 1  Equation 2
4.8592 = 3.3074b + a 5.1383= 3.6324b + a

4.8592 = 3.3074b + a
-1(5.1383 = 3.6324b + a)
-0.2791 = -0.325b
-0.325 -0.325

b = 0.8588
 Linear Regression Method
Y = xb + a
 Equation 1
4.8592 = 3.3074(0.8588) + a
4.8592 – 3.3074(0.8588) = a
4.8592 – (2.8404) = a
2.0188 = a
 Equation 2
5.1383= 3.6324(0.8588) + a
5.1383 – 3.6324(0.8588) = a
5.1383 – 3.1195 = a
2.0188 = a
 Linear Regression Method

Y = xb + a
 Equation
5 = x(0.8588) + 2.0188
5 – 2.0188 = x (0.8588)
2.9812 = x(0.8588)
0.8588 0.8588
X = 3.4714 = 2960.74mg/kg
Graphical Method