Analysis of the resistance characteristics
The resistance characteristic of biological indicators is not solely dependent on the test organism’s species but also varies between production batches. For this reason, ISO 11138 standard series claims minimum requirements for the resistance of biological indicators. The resistance of biological indicators is described by the D value. This value specifies the time required to kill 90 % of the initial spore population under defined sterilization conditions.
As the D value has a stronger influence on the resistance characteristic of the indicator than the population, it is advisable for users to verify the D value. The resistance characteristics of biological indicators are also influenced by carrier materials or suspension media. This applies for instance when spore suspensions are used for inoculation of product surfaces or liquids.
In those cases users complying with USP have to determine not only the population but also the D value of spores in the inoculated products.
For determining D values SAL GmbH is equipped with resistometers according to ISO 18472 (BIER vessel). These differ from other sterilizers in their ability to produce highly precise sterilization conditions within a few seconds. Thus, resistometers allow a reproducible analysis of resistance characteristics of biological indicators. We also offer the analysis of resistance characteristics on inoculated products (covering carrier effects) and in liquid products (covering suspension media effects). Biological indicators can be tested in the following sterilization processes:
- ethylene oxide
- steam
- dry heat
- formaldehyde
- hydrogen peroxide (feasibility dependent on individual process conditions)
You can choose between different methods for analyzing the resistance characteristics of biological indicators.
One method for determining the D value is the survivor curve. The analysis is carried out according to the ISO 11138 standard series. It is a quantitative test method. Indicators are subjected to graded exposures to sterilization conditions and the number of surviving spores is determined. From this data the D value can be calculated.
This test method requires relatively few samples (30 indicators are sufficient) and provides results within 2 days. Additional information about the homogeneity of the spore population, a quality feature, can be deduced from the survivor curve.
Terms like “Holcomb-Spearman-Karber Method“, “limited Holcomb-Spearman-Karber Method“ or „Stumbo-Murphy-Cochran Method“ describe different types of the fraction negative method for D value determinations. The test is described by the ISO 11138 standard series. It is a qualitative test method. Indicators are submitted to graded exposures of sterilization conditions and are evaluated for viable spores (growth / no growth). From this data the D value can be calculated.
This test method requires a relatively large number of samples (usually 144 indicators) and can be evaluated not earlier than after one week. On the other hand, test results are statistically sound due to the large overall samples which are analyzed. Most manufacturers of biological indicators use the fraction negative method for analyzing their products. The choice of method to determine the D value can influence the test result under certain circumstances. For this reason, USP recommends the use of the manufacturer’s test method for verifying D values.
Should you have any questions regarding the different D value determination methods we will gladly assist you in the choice of a suitable test method.
The survival-kill response verifies known population and resistance specifications. Indicators are sterilized for two different exposure times and evaluated for viable spores (growth / no growth). Exposure times are selected to yield microbiological growth after the shorter exposure (survival time) whereas after the longer exposure indicators should be inactivated completely (kill time).
The test is described by the ISO 11138 standard series, USP and European Pharmacopeia. Procedure and sample requirements vary between different compendia ranging from as few as 15 samples up to 110 samples.
Should you have any more questions we will gladly assist you in the choice of a suitable test method.
You would like to use a biological indicator in a steam or dry heat sterilization process at other temperatures than 121 °C or 160 °C? Then you can calculate the D value of your indicator at other temperatures by means of the z value. The z value specifies the required increase in temperature to reduce the D value to 10 % of its initial value.
This z value is an important characteristic of biological indicators because it defines how the indicator will perform in a real sterilization process with prolonged heating-up and cool-down times and other sterilization temperatures than 121 °C for steam or 160 °C for dry heat.
Users complying with USP who inoculate liquid products with spore suspension for validation or routine monitoring of terminal sterilization processes have to determine the D value and z value of spores in the inoculated product.
We offer z value determinations according to ISO 11138 standard series.
Specifications of biological indicators vary with the carrier materials or suspension media. For this reason, manufacturers of biological indicators have to check each production batch by a thorough analysis.
If you are using spore suspensions for inoculating product surfaces or liquids, the resistance characteristics of the spore suspension might not apply to the inoculated product due to the influence of carrier material or suspension medium.
To avoid using self-inoculated products which do not fulfil minimum resistance requirements you should verify resistance characteristics of your self-inoculated products.
Users complying with USP who inoculate solid products have to determine at least the population and the resistance. When using spore suspensions for inoculating liquid products, also the z value of spores in the inoculated product has to be determined.
We offer the determination of resistance and z value of biological indicators on inoculated products (covering carrier effects) and in liquid products (covering suspension media effects).
