The phrase sterility simply means the absence of living organisms including bacteria, fungi, viruses, protozoa and other vegetative cells of microbes in a product. Sterility is usually achieved through the process of sterilization. Sterilization is defined as the process by which objects, materials or environments can be rendered sterile (i.e. free from living organisms). Sterilization can be achieved by several methods including autoclaving, filtration, heating, use of ethylene oxide gas and by ionizing radiation. Microbiological and/or biochemical tests carried out on biological products to test their sterility are necessary in order to ensure that the products are not just efficacious but also safe for human and/or animal consumption. The term sterility is usually an absolute term which implies the complete absence of viable microorganisms in a product or in an environment. But in practice, the sterility of a product is defined by the absence of viable and actively multiplying microorganisms when tested in specified culture (nutrient) media that supports the growth of the suspected microbes. Usually, culture (nutrient) media that support the growth of suspected contaminants or pathogens including bacteria and fungi are used for the isolation of microbes from the tested samples that are representatives of the batch of product to be tested.
Sterility tests are microbiological and/or biochemical tests carried out on biological products, food and other pharmaceutical products in order to critically assess their freedom from contaminating microorganisms including mycoplasma, bacteria, fungi and viruses. Such tests are usually intended to detect the possible presence of viable microorganisms in a given biological or pharmaceutical products. Several products ranging from medicines, food and vaccines are subjected to series of sterility tests that ascertain their freedom from contamination or pathogenic microbes. Sterility tests are critical in the production of biological products since they are intended for parenteral (systemic) administration, and in which case they either confer health benefits or therapeutic effects on the consumers. Because sterility is an absolute term, it is vital to ensure that the working environments where sterility tests are done are free from contaminating microbes; and it is also important to avoid any accidental contamination of the product being tested. The principle supporting the sterility tests of biological products is simply based on the fact that microorganisms are ubiquitous and thus could find their way into products during production. And if this is the case, when such contaminated product is subjected to sterility tests using appropriate culture media, it is expected that the contaminating organisms will grow and produce colonies (in the case of solid culture media plates) or turbidity and/or gas production (for broth cultures) that gives impetus to the presence of contaminating microorganisms.
Sterility tests are adequately designed in such a manner that they reveal the presence of contaminating microorganisms present in the test samples used for the experiment. It is noteworthy that sterility test is not performed on all the products but on some representative samples of the whole lot or batch since it is not practicably possible to test all the samples or products in a batch of product. Nevertheless, a sufficient number of product samples from each batch of the product are subjected to sterility testing in order to give “an acceptable degree of confidence” in the ultimate results obtained from the sterility test. This helps to certify the batch of the product as free from contaminating microorganisms. A sampling technique based on probability profile is used to select samples from a batch of product for sterility testing since not all the samples or products in a batch can be tested at the same time.
Sampling is defined as the statistical process of selecting a part or portion of a whole product batch to represent the entire batch. Samples are picked or selected in a random manner to serve as representative samples of the whole lot. The use of membrane filtration technique and direct inoculation using culture (nutrient) media are often the two most important principal sterility testing technique used to determine the sterility of a product. Membrane filtration technique is indispensable in sterility testing because of the assumption that microorganisms especially bacteria will not pass through a membrane filter of 45 mm (0.45 µm) porosity. However, there also exist some rapid techniques employed for the determination of the sterility of a product. Some of these rapid techniques include the ATP-bioluminescence, colorimetric growth detection technique, autofluorescence and the use of cytometry. Sterility tests only detect gross contamination of those microorganisms which will produce visible turbidity in broth culture media (as evidence of microbial growth) or produce gas or acid as a result of microbial activity. Factors that affect the sterility tests include number of samples used, and testing conditions such as media used and incubation conditions for growth. The sterility test will only detect or show the presence of those microorganisms that will grow under the provided test conditions even when testing products suspected of heavy microbial contamination. These test conditions include the type of culture media used, incubation temperature, diluents used and recovery methods.
Generally, sterility testing is a regulatory requirement for the release of biological and pharmaceutical products that cannot be terminally sterilized (i.e. products that are heat-labile and thus susceptible to destruction by heat). Sterility test still remain a pass for the release of biological products for public consumption. Since sterility testing cannot on its own certify the absolute assurance of freedom of a product from microbial contamination, it is vital that every manufacturing processes (especially those meant for the production of biological products) ensures a continued and strict compliance to Good Manufacturing Practices (GMPs) at every production stage. This implies that the possible microbial contamination of the process and/or product should be prevented before it happens. The quality systems of production and/or manufacturing units of production industries should ensure aseptic process, proper sterilization techniques, sterility assurance, quality control and assurance. The microbiological and physical parameters of the production process should be continuously monitored during production to minimize contamination of the finished biological products.
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