Sterilization

on a practical basis means the absence of any detectable

viable organism.
: only the desired organism is detectably present.
Sterilization methods:
Medium & equipment:
Heat-sensitive materials in medium:
sizes<0.2m).

.
(microfiltrator pore

Heat-sensitive equipment: UV radiation (energy intensive) or

chemical agents (toxic to the desired culture. e.g.70% ethanolwater, and ethylene oxide (gas).
Gas (air): sterilized through adiabatic compression process for
supplying oxygen (~2200C) followed by
(e.g. glass wool
filter).

Sterilization
Thermal deactivation
The probability of an unsuccessful sterilization:
1- P0(t)
The probability of extinction (successful sterilization):
P0(t)=[1-p(t)]N0
where p(t) is the probability that an individual will still be viable
at time t. N0 is the number of individuals (cells or spores)
initially present.
Assume first order death rate:
p(t)=e-kdt
1- P0(t)=1-[1-p(t)]N0 = 1-[1-e-kdt]N0
where the specific death rate kd = e-E0d/RT(1/time), constant at specified
T. Eod is the activation energy for the death of the organism.

Sterilization
(D): time for the number
of viable cells to decrease tenfold.
E[N(t)] = N0 p(t)
E[N(t)] = N0 e-kdt
0.1= e-kdD
D=2.303/kd
E[N(t)] is the expected value of the number of
individuals present at time t.

Sterilization
1- P0(t)= 1-[1-e-kdt]N0
kd = e-E0d/RT
From the above equation:
- Known N0, T, t, determine Kd, the probability of
an unsuccessful sterilization is determined.
- Given N0, T, acceptable probability of failure e.g.
10-3, required time can be determined
- Higher Kd tends to achieve low probability of
sterilization failure. Normally at 121oC.
Kd of vegetative cells > 1010 min-1, spores 0.5-5
min-1. The major concern is spores.

Sterilization Chart
e.g. N0=108,
Kd=1min-1 (1210C), t=26min
Kdt=26
Probability of failure:
1-P=0.001
e.g. N0=108,
Kd=1min-1 (1210C), 1-P=0.001
Kdt=26
Required sterilization time:
t=kdt/kd=26 min

(M. Shular, Textbook, p.319)

Batch Sterilization
holding
he
ati

ng

ling
co o

T (K)

t time
Temperature verse time in a batch sterilization process

Kd2
T (K)

Kd1

Kd3

t time
Simplified calculations for deactivation of spores and medium components

Sterilization
Degradation of components in the medium in sterilization
process.
vitamin and growth factor
The inactivation of viability is much more sensitive to
temperature changes than the degradation of
important growth factors in the medium
It is important to assure complete killing of foreign
organisms without the destruction of important
components in the medium.

Sterilization
Degradation of components in medium
Assume the degradation rate of such components is
first order.
dC/dt=-kdC
The degradation rate constant kd can also be related
to temperature by Arrhenius equation.
Integrating the above equation,
ln C/C0=-kdt or C=C0e-kdt
where C0 is the initial concentration of the component.
To determine the components remaining active:
the temperature T determine kd with known t,
determine C.

Sterilization
Degradation of components in the medium.
C1 t2 C2 Kd,2
T (K)

kd = e-E0d/RT

C0 t1 C1
Kd,1

C2 t3 C3 Kd,3

t time

C1 C0 e

k d ,1t1

C2 C1e

k d , 2 t1

C3 C 2 e

k d , 3 t3