Opinion Review Article | Open Access | Published 16th July 2024
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Microbial control of the process: Do you know where your raw materials come from?
Tim Sandle, Ph.D., CBiol, FIScT | EJPPS | 292 (2024) | Click to download pdf
Microbial contamination control is technically challenging and there exists the potential for adverse patient impact should pharmaceutical products become contaminated. One source of contamination is with the raw materials used in the manufacturing process¹.
Raw materials are those substances which can be brought into a manufacturing unit either for further processing or to aid in such processing. In the microbiological control of pharmaceutical raw materials there is one primary aim: To exclude any microorganism which may subsequently result in deterioration of the product or may harm the patient.
Contaminants in raw materials are important not only because they may contaminate the product directly, but also because they may contaminate the manufacturing plant, possibly giving rise to problems in the future.
Mechanisms of control
Mechanisms of control involve the sampling of raw materials and the testing of part or the entire sample for the presence of microorganisms, with special reference to pathogenic species, and hence the need to consider screening for specific indicator organisms. In terms of the relative risk of different raw materials², a logical approach must be used. Some raw materials are more likely to be contaminated than others, and some never contain any detectable contamination.
The criteria which may be used to identify materials to be examined are:
Is the product of natural origin?
Is it synthetic?
Is it produced in a manner likely to reduce or increase the level of contamination?
Are microorganisms likely to multiply in it?
Materials of natural origin
Ideally, pharmaceutical preparations should be formulated with raw materials that are unlikely to be sources of contamination. To do so would mean avoiding raw materials originating from plants, animals, or mineral material (e.g. gums, sugars, gelatine talc etc.). Unfortunately, this is not always practicable. There is value in performing bioburden testing for process control, however data is limited by factors such as inter-laboratory variation, problems associated with sampling, quantification of viable but non-culturable bacteria and the dynamics of microbial growth.
Microorganisms from plant material
As with any natural material, the microbial count from a plant origin will reflect the natural flora. Generally, this will comprise a mixture of yeasts, moulds, Gram-positive bacteria and bacterial spore-formers³. Hence, the microflora of plant materials such as gum acacia and tragacanth, agar, powdered rhubarb and starches will be indigenous to the respective plants and may include bacteria such as Erwinia spp., Pseudomonas spp., Lactobacillus spp., Bacillus spp. and streptococci, and moulds such as Cladosporium spp., Alternaria spp., and Fusarium spp.
The numbers of microorganisms present on the plant material may also reflect storage and harvest conditions and should be taken into account when performing any bioburden estimation. In addition, some microbial species have inherent antimicrobial properties which must be considered when testing. If fungal growth on the material is suspected, then the presence and significance of fungal toxins must be evaluated. It is not unreasonable to expect bacterial numbers of anywhere between 100 and one million per gram of plant material. Whilst the material from roots, tubers, bulbs etc. is low in bioburden in the healthy plant, these regions are likely to be contaminated with soil organisms, predominantly Gram-positive spore forming bacteria. There, numbers may vary from 100/g to 10,000/g.
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Microorganisms from animal sources
Historically, animal derived products include vaccines, hormones and growth factors, sutures and organ explants and implants. However, with concerns over prions, the number of animal products used as raw materials has vastly reduced. Those that are still obtained from animals are required to be sourced from approved suppliers who have given sound (documented and audited) consideration to hygiene and microbiological control in the design of their production (e.g. farming, cultivation, extraction etc.) and distribution practices.
In order to trace the source of any possible contaminant, each organ / extract effectively becomes its own batch number. Products from animal sources such as gelatine, desiccated thyroid, pancreas and cochineal may be contaminated with animal-borne pathogens. For this reason, statutory bodies require freedom of such materials from E. coli and Salmonella spp. at a stated level before they can be used in the preparation of pharmaceutical products. Such raw materials must also comply with a total viable count limit in some cases.
Microorganisms from mineral-derived materials
Mined sources of minerals such as talcum and gypsum may also contain microorganisms. Often, they reflect the microbiota of the surrounding soil, or the method of extraction, but can harbour potentially harmful microorganisms. Clostridium tetani rarely occurs in pharmaceutical products, but when present in e.g. contaminated talcum powder, has been known to cause serious wound infections and several cases of neonatal death. Generally speaking, mineral-derived materials present only minor sources of microbiological contamination. However, it is essential that they be sourced from suppliers who have given sound consideration to hygiene and microbiological control in the design of their manufacturing and distribution practices.
Synthetic raw materials
Synthetic raw materials are usually free from all but incidental microbial contamination. Although microorganisms may contaminate all types of raw material, the potential for growth in synthetic materials is very low. In general, these incidental organisms are considered to be of little practical significance. Contamination by pathogens is extremely unlikely. Exceptions to this rule can occur, for example, where one of the preparation stages involves washing with, or crystallizing from, water of poor microbiological quality. In relation to these types of materials, microorganisms cannot thrive in nonaqueous environments and many of the organic solvents commonly used in the manufacture of products are inherently bactericidal, although not sporicidal due to the protection dipicolinic acid affords bacterial endospores.
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Preparation and storage
Raw materials account for a high proportion of the microorganisms introduced during the manufacture of pharmaceuticals, and the selection of materials of good microbiological quality aids in the control of contamination in both products and the environment. An important consideration is that the method of preparation of the raw materials may in itself lead to increased levels of contamination. Hence, a knowledge of the method of preparation of raw materials is essential in order to determine the full extent of microbiological testing for any material. For example, some refining processes modify the microflora of raw materials. Drying may concentrate the level of spore-forming bacteria, whilst some solubilization processes may introduce waterborne bacteria.
Storage
The careful storage of raw materials, particularly hygroscopic substances, is important in order to prevent growth of the organisms and spoilage of the material. If stable, natural products with a high microbial count may be sterilized. Staff handling raw materials must be given adequate training to prevent cross-contamination. However, for the majority of raw materials pre-product sterilization is not an option. An important parameter for storage is the moisture content of the environment. Any increase in water activity (Aw) above the minimum required for growth will encourage population growth especially of moulds and yeasts, as will insect or rodent infestation during storage⁴. Water activity is defined as the ratio of the vapour pressure of water in a material (p) to the vapour pressure of pure water (po) at the same temperature. Relative humidity of air is defined as the ratio of the vapour pressure of air to its saturation vapour pressure. The water activity scale extends from 0 (bone dry) to 1.0 (pure water). Many bacteria will not grow below a value of 0.8, and at 0.6 growth is considered impossible even for the most extremophilic fungi (xerophilies).
Precautions should be taken to ensure that dry materials are held below these levels. Hence, it is important to hold raw materials at a constant temperature in order to prevent evaporation and condensation occurring. It is worth noting that some materials such as oils and sugar solutions may contain local pockets of free water where microbial proliferation may take place. Care should also be taken with packaging as some types (e.g. unlined paper sacks) may absorb moisture and may itself be subject to microbial deterioration and so contaminate the contents.
In this article, a risk consideration for different types of raw materials has been presented. This is based on the origin of the material and in consideration of the relative microbiological control of pharmaceutical raw materials. To exclude any organisms which may subsequently result in deterioration of the product or may harm the patient account needs to be taken of the bioburden and specific pathogen risk, and understanding where a raw material has come from and what its likely quality is going to be, presents an important step in this risk assessment process.
Tim Sandle’s microbiology website is Pharmaceutical Microbiology Resources.
References
01. Sandle, T. (2014) The Risk of Bacillus cereus to Pharmaceutical Manufacturing, American Pharmaceutical Review, 17 (6)
02. Sutton, S, and Jimenez, L. “A Review of Reported Recalls Involving Microbiological Control 2004-2011 with Emphasis on FDA Considerations of ‘Objectionable Organisms.’” American Pharmaceutical Review 15 (2012): 42–57
03. Cundell, T. “Mold Monitoring and Control in Pharmaceutical Manufacturing Areas.” American Pharmaceutical Review 19 (2016) (Accessed July 18, 2017)
04. He, Y.; Li, Y.; Salazar, J. K.; Yang, J.; Tortorello, M. L.; Zhang, W. (2013). "Increased Water Activity Reduces the Thermal Resistance of Salmonella enterica in Peanut Butter". Applied and Environmental Microbiology. 79 (15): 4763–4767
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Author Information
Corresponding Author: Tim Sandle, Head of Microbiology
Bio Products Laboratory ,
UK Operations, England
Email: timsandle@btinternet.com
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