• Medicines are vulnerable to contamination and spoilage by microorganisms. The two words have different meanings; contamination is the entry of microorganisms into the product, whilst spoilage describes the product damage that results from microbial growth.

• Microorganisms should be excluded from medicines not only because they represent an infection hazard, but also because they may degrade the active pharmaceutical ingredient, reduce the product’s physical stability or make it unacceptable to the patient.

• Anhydrous medicines are not normally susceptible to spoilage because microbial reproduction will not occur in the absence of water. The amount of free water available for microbial growth in a product is determined by the water activity (A_w). Reducing the water activity of a product is therefore a means of protecting it against spoilage.

• Microbial contamination of medicines arises from three principal sources: the raw materials (particularly water), the manufacturing environment and from personnel.

• Raw materials of animal, vegetable or mineral origin normally have a higher level of contamination than those made by chemical synthesis where heat, extremes of pH or organic solvents tend to kill microorganisms.

• The ‘Orange Guide’ describes manufacturing practices that are designed to minimize both the contamination introduced into a product and the opportunities for those contaminants to reproduce whilst the product is being made.

• Preservative chemicals are required for most water-containing non-sterile products and for multidose sterile products.

• The range of available preservatives is quite limited, and they all suffer from one or more of the following faults: only possessing good activity against bacteria or fungi, but not both; exhibiting reduced antimicrobial activity in certain pH ranges – several only working well in acid conditions; causing skin sensitivity reactions; and interacting with other common excipients – several preservatives lose activity in the presence of surfactants.

The need to protect medicines against microbial spoilage

The need to protect foods against microbial spoilage is well appreciated since microbial growth results in obvious signs of deterioration. However, there is a much lower level of awareness among the general public of the need to similarly protect cosmetics, toiletries and medicines. Although most medicines present a less favourable environment for microbial growth than foods, a wide variety of potentially hazardous organisms are nevertheless capable of growing to high concentrations in unprotected products. The subject of preservation is therefore an important aspect of medicine formulation, simply because patients taking medicines are, by definition, unhealthy and so quite possibly more vulnerable to infection.

It is important to distinguish between the terms contamination and spoilage because they are sometimes used synonymously, which is incorrect.

Contamination, in this context, means the introduction of microorganisms into a product, i.e. it describes microbial ingress. Contaminating organisms can arise from many sources (considered later in this chapter) during the course of both product manufacture and subsequent use of the product. The procedures of good manufacturing practice (GMP, also considered later) are used to limit the first of these (see Rules and Guidance for Pharmaceutical Manufacturers and Distributors, 2007), but contamination arising from the patient is largely out of the control of the manufacturer except in the context of container design and labelling. There has been a trend in recent years to adopt containers that minimize contact between the patient’s body and the product, e.g. collapsible tubes are used for creams and ointments rather than open-mouthed tubs or jars into which fingers can be inserted. Similarly, single-dose eye drops may be preferred to bottles where the dropper can come into contact with an infected eye and then be replaced in the eye drop solution. Despite this, contamination by the patient is still a problem to be considered in container design and product preservation.

Spoilage follows contamination and describes the process and consequences of microbial growth in the product. Considering the potential for product spoilage and taking appropriate steps to minimize the risk of it occurring are very much the responsibility of the formulation scientist and the manufacturer.

There are three principal reasons why microorganisms should be excluded totally from medicines or their presence subjected to stringent limits set by pharmacopoeias or regulatory agencies, such as the United States Food and Drugs Administration (FDA), the European Medicines Agency (EMA) or the UK’s Medicines and Healthcare products Regulatory Agency (MHRA):

It is quite obvious that medicines should not contain pathogenic organisms that represent a source of infection. However, specifying the species and numbers of organisms representing an infection hazard is not straightforward. Certain pathogens are recognized as ‘objectionable organisms’ and must be totally excluded from particular raw materials or product types (see Chapter 14), but the risk of infection is influenced not just by the number and type of organism but by other factors too. For example, an organism may be present at a concentration that would be regarded as relatively harmless to a healthy individual but which may pose a problem for patients with impaired immunity.

In the 1960s and 1970s there were several reports in the pharmaceutical literature of infection occurring as a result of medicines containing pathogenic species, e.g. salmonellae, clostridia and Pseudomonas aeruginosa, but such reports became far less frequent towards the end of the last century with the adoption of more rigorous quality standards and regulatory control of manufacture. However, contaminated medicines are by no means a thing of the past. The FDA publishes details of product recalls on its website, and in March 2011 alone, there were three unrelated product recalls due to concerns about potential or confirmed microbial contamination. It should be emphasized that there is the possibility of an infection arising from the use of a product contaminated with a concentration of organisms that is too low to be detectable by sight or smell. This situation is potentially much more hazardous than that of a patient confronted with a medicine in which microbial growth is clearly evident.

Quite apart from representing an infection hazard, microorganisms may damage the medicine by degrading either the active ingredient or one or more excipients, thus compromising the quality and fitness for use of the product.

Degradation is usually due to either hydrolysis or oxidation, but decarboxylation, racemization and other reactions may also occur. Active ingredients known to be susceptible to microbial attack include steroids, alkaloids, analgesics and antibiotics. Much of the literature on this topic has been reviewed by Spooner (1996) and Bloomfield (2007). The numbers and variety of excipients that have been reported to be degraded are at least as great as those of active ingredients. Thus, most categories of excipients contain materials that have been shown to be susceptible to microbial enzymes, acids or other metabolic products.

Common examples of product instability or deterioration include emulsion phase separation due to surfactant degradation, loss of viscosity due to microbial effects on gums, mucilages and cellulose derivatives employed as thickening agents, and alcohol and acid accumulation following fermentation of sugars. Despite the fact that the very purpose of their use is to restrict microbial growth, even some preservatives are vulnerable to inactivation by microorganisms that, in exceptional cases, use them as a carbon and energy source. Nor should it be assumed that products whose very purpose is to kill microorganisms will necessarily be self-sterilizing: two of the three FDA product recalls mentioned above involved antiseptic wipes containing alcohol and iodine which were designed to decontaminate skin prior to injection or surgery, but the wipes were, themselves, sources of microbial contamination.

If microbial growth within the product is sufficiently extensive, it is possible for the presence of the organisms to be detectable by:

Clearly, any product manifesting such changes would be unlikely to be used by the patient. This may result in short-term problems for the patient of obtaining alternative supplies, and possible longer term problems for the manufacturer in terms of customer complaints, product recalls, adverse publicity and possible legal action.

Products and materials vulnerable to spoilage

Spoilage, in the sense of detectable physical or chemical change within a pharmaceutical product, nearly always follows growth and reproduction of the contaminating organisms. The pharmacopoeial and regulatory limits for the maximum permissible numbers of microorganisms in manufactured products or raw materials are typically not more than 100–1000 colony-forming units (cfu) per mL or gram. Whilst these concentrations may allow some pathogens to initiate infections, they are not normally sufficient to cause detectable changes in chemical composition, physical appearance or stability. Bacteria and fungi are just the same as all other living organisms in requiring water for growth (although not necessarily for mere survival). This means that only products containing sufficient water to permit such growth are vulnerable to spoilage. Consequently, spoilage is not normally a problem in anhydrous products like ointments and dry tablets or capsules, although hygroscopic materials like gelatin and glycerol may absorb enough water from the atmosphere to enable moulds (but not normally bacteria) to grow. Similarly, cellulosic materials, particularly paper and other packaging, may show mould growth if stored in humid atmospheres, e.g. in tropical climates.

The fact that a product contains water and is obviously a liquid does not necessarily mean that the water is available to participate in chemical reactions and enable microorganisms to grow. Some of the water present in a solution is bound to the solute due to hydrogen bonding or other mechanisms. Thus a parameter that indicates the proportion of ‘free’ or available water is a useful guide to the ease with which microorganisms might grow in the product. Such a parameter is water activity (A_w) which is the ratio of water vapour pressure of a solution to the water vapour pressure of pure water at the same temperature. A_w is expressed on a scale from zero to 1 with a value of 1.00 representing pure water. As the concentration of solute in a solution is increased, A_w falls proportionately and the range of organisms able to grow in the solution progressively diminishes. Thus, it is possible to construct a table indicating the minimum A_w values that permit the growth of different types of microorganism (Table 50.1).

To a certain extent, the values of A_w in Table 50.1 are reflective of the natural habitats of the organisms concerned; pseudomonads and other waterborne organisms therefore tend to require high A_w values for optimum growth, whilst skin organisms like staphylococci and micrococci that exist in relatively high salt concentrations (from sweat glands) will tolerate significantly lower values. Pharmaceutical materials that have high solute concentrations, e.g. syrups, may to a large extent be self-preserving just like salted foods. Syrup BP, for example, is 67% by weight sucrose, has an A_w value of 0.86 and so is not susceptible to bacterial growth, but may contain chemical preservatives to protect it from mould spoilage.

Variations in water activity may arise within a single container of a manufactured medicine by, for example, water evaporating from the bulk liquid during storage in high temperatures and that water vapour condensing on cool glass round the neck of a bottle as the storage temperature drops, then running back to dilute the surface layer of product. It is for this reason that syrups should not be stored in fluctuating temperatures.

The possibility also exists for contaminants to grow and generate water from respiration and so produce localized increases in A_w