▪ As per PDA TR1 : Sterilization – A process used to render a product

free of viable organisms with specified probability.

▪ As per ISO: “Validated Process used to render a product free of all

forms of viable microorganisms”

▪ In General, the term Sterilization for pharmaceutical preparations,

means the complete destruction of all living organisms and their

spores or their complete removal from the preparation.

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▪ Below mentioned sterilization processes are described in the USP/EMA guidelines:

1. Steam Sterilization

2. Dry heat sterilization

3. Ionization radiation sterilization

4. Gas sterilization

5. Sterile filtration

6. Aseptic processing

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▪ D-value:

✓ The D-value is defined as the decimal (or decadal) decay (or reduction) time: i.e. it is

the time required, at a specified temperature T, to reduce the microbial population

being considered by one logarithmic value, i.e. from 100% to 10% of the initial value.

✓ The D-value is a measure of the heat resistance of a particular microorganism. The D-

value represents the time in minutes that is required to effect a 1-log reduction in the
population of a particular microorganism. Most naturally occurring microorganisms
have a D-value < 1.0.

▪ Z-value: The z-value is the temperature change required to effect a 1 log reduction in

the D-value.

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▪ F0 – Value OR Equivalent Exposure Time At 121°C :

▪ F0, as defined by USP (10) at a particular temperature other than 121 °C, is the time

(in minutes) required to provide the lethality equivalent to that which is provided at
121 ° C for a stated time.

✓ A mathematical method of defining the equivalent time at 121.1 °C for a given

temperature (in minutes).

✓ F0 provides a relationship between sterilization temperature and biological “kill”.

✓ F0 provides a method of determining the Sterility Assurance Level (SAL).

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▪ Sterility Assurance Level (SAL):

✓ A sterility-assurance level (SAL) is defined as the probability of an item being

nonsterile after it has been exposed to a validated terminal sterilization process.

✓ Implantable medical devices are terminally sterilized to achieve a SAL of 10−6,

which implies a probability of finding not more than one viable microorganism in
one million sterilized items of the final product.

▪ Overkill Method:

✓ According to USP, “overkill sterilization can be defined as a method in which the

destruction of a high concentration of a resistant microorganism supports the

destruction of reasonably anticipated bioburden present in routine processing.”
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▪ Sterilization of product and equipment by saturated steam (moist heat) is one of the
most widely used treatment in the parenteral drug industry.

▪ As per EMA Guideline on sterilization of the medicinal product, active substance,

excipient and primary container

✓ All steam sterilization processes require a minimum lethality of F0 ≥ 8 minutes and a

minimum process hold temperature of 110 °C.

▪ Advantages:

✓ 1st choice for sterilization for products

✓ Very efficient procedure

✓ Quick and economical

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▪ Mechanism of Steam sterilization:

✓ Denaturation and coagulation of some of the organism's essential protein.

✓ The presence of the hot moisture within the microbial cell permits destruction at

relatively low temperature.

▪ As per EMA Guideline on sterilization of the medicinal product, active substance,

excipient and primary container, Steam sterilization performed with finished

product temperature below 115 °C during the holding phase is an exceptional case
and should be scientifically justified and supported by additional data.

▪ If temperatures below 110 °C are included (during heat-up and cool-down) in the

determination of F0, this should be justified.

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▪ Dry heat sterilization is widely used to sterilize glassware and equipment parts in
manufacturing areas for parenteral products.

▪ Is usually carried out in sterilizing ovens specifically designed for this purpose.

▪ Dry heat is less effective in killing microorganisms than is moist heat

▪ Higher temperature and longer period of exposure are required.

▪ Dry heat kill microorganisms primarily through oxidation.

▪ The most common time-temperature relationship for dry heat

sterilization are-

1.160 °C for 2hrs

2.220 °C for validated time is used to sterilized glassware & PPM.

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▪ Radiation sterilization utilizes the lethal effect of various forms of radiation as a

means of microbial destruction.

▪ Ionizing radiation (gamma, x-ray, or beam) sterilization is used extensively for the

sterilization of medical devices and for a variety of other materials and products.

▪ The advantages of sterilization by irradiation include:

✓ Simplicity

✓ Absence of mechanical complexity

✓ Reproducibility and

✓ Overall efficiency.

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▪ Gamma sterilization needs the use of a specifically designed facility where items to be

sterilized are exposed to a Co60 radiation source in a manner that ensures uniform dosing.

▪ Highly penetrating photons (gamma rays) are emitted from Co60 as it decays to Ni60.

▪ The half-life for this isotope is 5.27 years, which means that over the course of each year the

source loses about 12% of its radioactivity.

▪ This steady reduction in radioactivity requires that radiation process operators adjust their

process controls (typically exposure time) to maintain the established dose required.

▪ It is also used to sterile the heat sensitive APIs when other suitable methods are not
applicable.

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▪ Generally,gas sterilization is only acceptable if no other method of sterilization is possible.

▪ The use of sterilizing gases for the preparation of materials and equipment is commonly

used for items that are susceptible to damage by heat or radiation processes.

▪ To ensure adequate sterility, sufficient penetration by gas and moisture is essential.

▪ The majority of gas sterilization processes employ ethylene oxide (EO) gas for sterilization

because of its ability to penetrate through polymers, cellulosics, and other materials allows it
to be used for the terminal sterilization of medical devices in their final packaging.

▪ Other gases like Ozone, chlorine dioxide, nitrogen dioxide may used for sterilization but EO

is first choice by parenteral manufacturers.

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▪ EO is a powerful alkylating agent that destroys microorganisms by chemical reaction, primarily
with cell DNA.

▪ The destructive mechanism largely follows first-order kinetics and depends on concentration,
humidity, and temperature.

▪ The usual EO process follows a sequence of prehumidification, air removal, rehumidification in

the chamber, gas exposure, gas removal from the chamber, and post exposure aeration.

▪ The pre exposure steps ensure that adequate moisture is present on and within the items
being sterilized. The post exposure steps provide time for the diffusion of EO and its byproducts
out of the materials and packaging.
Sterilization method BI Organism
Bacillus stearothermophilus
Moist Heat (121°C for 15 min)
(Geobacillus stearothermophilus)
Dry Heat (160 °C for 2h) Bacillus subtilis
Irradiation (25KGy) Bacillus pumilus
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Ethylene Oxide Bacillus subtilis
▪ Sterilization by filtration, depends upon the physical removal of microorganisms by

adsorption on the filter medium or by sieving mechanisms, is used for sterilization

of heat-sensitive solutions.

Representative image for Parenteral filtration set up 14

▪ Aseptic processing is not considered to be a sterilization process but concerns the

usage of technologies to process sterile components avoiding addition of

microbiological contaminants, e.g. use of an isolator.

▪ For aseptic processing, information on the bulk holding time before filling and on

the filling time should be stated and appropriately supported by data. The times
should be minimized.

▪ The grounds for holding and filling times longer than 24 hours should be justified

and supported by a risk assessment.

▪ It should be verified that the results of the media simulations support the proposed

holding and processing times.

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▪ Finished products intended to be sterile should be terminally sterilized in their

final container whenever possible, as clearly stated in the Ph. Eur., general chapter
5.1.1.

▪ Active substances, excipients and containers when required to be sterile should be

packed before they are sterilized whenever possible.

▪ When terminal sterilization by heat is not possible, the application of an alternative

method of terminal sterilization, sterilizing filtration and/or aseptic processing may

be considered.

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