• Types and size of contaminant inoculum.
• Nutritional factors.
• Water.
• Storage temperature.
• pH.
• Oxidation-reduction balance (Redox potential).
• Package design.
Types and Size of Contaminant Inoculum
Very low levels of contaminants that are unable to replicate
in a product might not cause appreciable spoilage but, if a higher contaminant
bio-burden occurs, the built-in protection could be insufficient and spoilage
occurs. This higher contamination arises if:
1. Raw materials were unusually contaminated;
2. A problem of the plant-cleaning protocol occurs;
3. Biofilm detached itself from within supplying pipework;
4. There was demolition or maintenance work in the vicinity
of the manufacturing site
5. Misuse of the product occurs during administration.
Nutritional Factors
Many spoilage microorganisms have simple nutritional
requirements and metabolic adaptability which enables them to utilize many
formulation components as substrates for biosynthesis, growth, and also trace
materials contained in them. The use of animal products and crude vegetable
materials in a formulation provides an additionally nutritious environment.
Demineralized water prepared by ion-exchange methods, contains sufficient
nutrients to allow significant growth of many water-borne Gram-negative
bacteria like Pseudomonas spp.
Water
It is the most important cause of the survival and growth of
micro-organisms. Some solute-rich medicines such as syrups appear to be 'wet',
microbial growth in them may be difficult since the microbes have to compete
for water molecules with the large numbers of sugar and other molecules of the
formulation which also interact with water via hydrogen bonding. An estimate of
the proportion of the non-complexed water in a formulation that is available to
equilibrate with any microbial contaminants and facilitate growth can be
obtained by measuring its water activity (Aw).
Aw = Vapour pressure of formulation / Vapour
pressure of water under similar conditions
The greater the solute concentration, the lower the water
activity. Most microorganisms grow best in dilute solutions (high Aw)
and, as solute concentration rises (lowering Aw), growth rates
decline until a minimal, growth-inhibitory Aw is reached. The Aw of
aqueous formulations are lowered to increase resistance to microbial attack by
the addition of high concentrations of sugars or polyethylene glycols.
Condensed water films accumulate on the surface of 'dry’ products such as
tablets or bulk oils following storage in damp atmospheres with fluctuating
temperatures, resulting in high localized Aw to initiate fungal
growth.
Storage Temperature
The actual storage temperature determines the spoilage by
particular types of microorganisms. Spoilage of pharmaceuticals occurs
potentially over the range of about 20°C to 60°C. Storage in a deep freeze at
−20°C or lower is used for long-term storage of foodstuffs and some
pharmaceutical raw materials and dispensed total parenteral nutrition feeds are
stored in hospitals for short periods at −20°C to even further minimize the
risk of growth of any contaminants that are introduced during their aseptic
compounding. Reconstituted suspensions and multi-dose eye drop packs are
sometimes dispensed with the instruction to 'store in a cool place' such as a
domestic fridge (8°-12°C), or “store in the refrigerator “ partly to reduce the
risk of in-use contamination growing before the expiry date. Water for
Injections is recommended to be held at 80°C or above after distillation and before
packing and sterilization to prevent possible regrowth of Gram-negative
bacteria, and the release of endotoxins.
pH
Extremes of pH prevent microbial attack. They grow at
neutral pH, therefore acidic or alkaline formulations are less susceptible to
spoilage. Around neutrality bacterial spoilage is more likely, with reports of
pseudomonas and related Gram-negative bacteria growing in antacid mixtures,
flavored mouthwashes, and in distilled or demineralized water. Above pH 8, spoilage
is rare for soap-based emulsions. Products with low pH levels such as fruit
juice-flavoured syrups are attacked by mould or yeast. Yeasts are metabolized by
organic acids and raise the pH to levels where secondary bacterial growth
occurs. In the food industry, low pH adjustment is made to preserve foodstuffs
only.
Redox Potential
The ability of microbes to grow in an environment is
influenced by their oxidation-reduction balance since they require compatible
terminal electron acceptors to permit the function of their respiratory
pathways. The redox potential in viscous emulsion is high due to the high
solubility of oxygen in most fats and oils.
Packaging Design
It has a major influence on the microbial stability of some
formulations in controlling the access of contaminants during both storage and
use. The most important dosage form such as parenteral drugs is protected
because of the high risks of infection by this route. Self-sealing rubber
closures are used to prevent microbial entry into multi-dose injection
containers following withdrawals with a hypodermic needle. Wide-mouthed cream
jars are replaced with narrow nozzles and flexible screw-capped tubes to remove
the likelihood of operator-introduced contamination during use. For medicines that
rely on their low Aw to prevent spoilage, packaging such as strip foils must be
of water vapor-proof materials with fully efficient seals. Sacking, cardboard,
card liners, corks, and paper are unsuitable for packaging pharmaceuticals
because they are heavily contaminated with bacterial or fungal spores. These
are replaced by non-biodegradable plastic materials. In the past, packaging in
hospitals was frequently reused for economic reasons.
Other factors affecting microbial spoilage of pharmaceutical
products include:
• Relative Humidity
• Oxygen Availability
• Osmotic Pressure
• Surface Tension