Introduction
It is often difficult to prevent the contamination of a pharmaceutical product during manufacture or contamination of individual components used for the manufacture of the final product. Also, a pharmaceutical product can be contaminated during used by the patient, during dispensing by the pharmacist and even by the medical staff.
The physicochemical and chemical parameters of a product influence microbial spoilage to a large extent. Therefore, their manipulation during formulation development can be favourably used to discourage conditions of spoilage and induce in an environment unfavourable for the survival or growth of microbes. A contaminant may sufficiently modify a formulation’s properties as to allow subsequent attack by other contaminant earlier discouraged by stringent initial conditions such as the following
1. Size and type of contaminant inoculum of the microorganism
There will be no risk of contamination or spoilage of pharmaceutical product (medicine) if the medicines are presented in sterile, impervious, single-dosage units. However, pyrogens can survive heat sterilization, and sterile filtration will not usually eliminate the pyrogens, soluble toxins or extracellular spoilage enzymes such as lipases. Upon entry of an aggressive contaminant into a product, there is the usual adaptive lag period before rapid spoilage ensures, which is usually extensive for very low microbial numbers but decreases disproportionately with increasing contaminate loads.
In non-sterile product or multidose pharmaceuticals, the nature and extent of microbial spoilage is determined by the type and quantity of contaminants remaining after manufacturing processes or entering during use of the product, and the degree to which they find the particular microenvironment conducive to growth or survival. Therefore, a detailed consideration of the contaminant lod which would enter the product is necessary to assist in the design of a potentially stable formulation.
2. Protection of Microorganisms within pharmaceutical products
It is known that components of formulations and products tend to increase the resistance and longevity of contaminant microoganisms in addition to reducing directly the efficiency of antimicrobial preservatives. The substances incriminated in this this phenomenon are certain proteins, suspending agents, low concentrations of surfactants such as polysorbates and the cetomacrogols. Adsorption of microorganism onto suspended particles such as kaolin, magnesium trisilicate or aluminum hydroxide gel may increase their longevity. Formulation pH will often influence the sensitivity of many organisms towards preservatives by affecting the following parameters
- The rate of growth of the inoculum
- The potency of the antimicrobial agent itself
- The ability of the drug to combine with site on the bacterial cell surface.
3. Nutritional Factors
It is known that several pathogens require specific growth factors that are not often present in pharmaceutical formulations. As a result of this they may not be able to grow and multiply in the pharmaceutical formulations because of the limited growth requirement and may survive in them for a short while with eventual extinction.
The nutritional requirements of saprophytic spoilage microorganism are simple and uncomplicated. As a result of this, they can easily utilise a wide variety of the single organic and inorganic trace ingredient present in most medicine as substrates for their biosynthetic and cell division processes. It has been demonstrated that good quality distilled water often contains sufficient nutrient to facilitate the growth of many Gram-negative bacteria including Ps.aeruginosa. Demineralized water produced by ion-exchange methods can support the growth of microbial organisms. When microorganism fail to grow in a medicine, it may not be due to their limited presence of nutrients, but largely due to other non-supportive, physicochemical or toxic properties of the medicinal preparations.
The pH of a liquid formulation is very important in the preservation of such medicinal liquid formulation. Although extreme of pH prevent microbial attack, it is known that feeble growth have been observed even a dilute hydrochloric acid. The following effect of pH have been observed.
Mildly alkaline conditions i.e. pH 8-9 as found in soap emulsions and mixtures such as those of magnesium hydroxide or aluminum hydroxide gel can discourage microbial growth. Acidic conditions favour fungal and yeast proliferation. Inadequately prevserved fruit flavoured phamarcetuical products at pH 3.5 can support good growth. Yeasts can metabolise organic acids raising the pH to allow secondary bacterial attach. Protein degradation in acidic formations can raise the pH by decarboxylase activity or lower the pH in slightly alkaline formulations by deaminase activity.
4. Storage temperature
The storage temperature of a pharmaceutical product activity can greatly influence the shelf-life of the product. Most spoilage of pharmaceutical product occur between-20o to 60oC, although the spoilage occurs much less at the extremes of temperature. However storage within specific, narrower temperature ranges will encourage the growth of particular groups of spoilage organisms. Below – 20oc microbial growth and spoilage of pharmaceuticals are minimal.
Note: For domestic use, in temperate countries, a room temperature of 20ºC (70ºF) is considered hot, in Nigeria, room temperature is about 25ºC. A home refrigerator may be an “warm” as 12ºC.
Distilled water for the preparation of water for injections is recommended to be maintained at 80ºC prior to sterilization to prevent pyrogen production. Hence storage of an extemporaneously prepared solution in a warm place, rather than in a cool place could increase its microbial deterioration. Therefore the effect of transportation and storage of products at local temperatures in the tropics or subtropics should always be considered in respect to the of effect of the environment temperature.
6. Moisture Content of the Product: Water activity (Aw)
Water activity (Aw) is a measure of the proportion of water molecules (remaining uncomplexed during formulation (via hydrogen bonding, with the discoloured components of the aqueous formulation) that is available in the pharmaceutical products to contaminant microbes for metabolism.
The greater the solute concentration in the product the lower the Aw. Most microorganisms grow best in dilute solutions of (high Aw) when the water activity is high and as the solute concentration increases (lowering of Aw), the growth rate declines until a minimum growths inhibitory Aw is reached.
Limiting Aw values are of the order of:
Gram-negative rods, 0.95
Staphylococci, 0.9
Micrococci 0.9
Lactobacilli 0.9
Yeasts 0.88
Syrup-fermenting osmo-tolerant yeasts can occasionally grow at low Aw levels as low as 0.73 while some filamentous fungi can grow at even lower levels, with Aspergillus glaucus as low as 0.61.
Pharmaceutical formulations are best preserved by reducing the Aw of formulations below these levels (above) by the addition of high concentrations of sugar or polyethylene glycols. BP syrup that is officially 66% w/v sucrose has Aw of 0.86. The BP syrup can fail to inhibit the more osmoloterant yeasts occasionally encountered.
Note: The BP intention to remove sucrose from its formulations will definitely cause problems as alternative solutes for lowering Aw such as sodium chloride (for food stuffs) may produce intolerant taste or toxicity.
Generally, Aw can be lowered by drying although the dry often hygroscopic, products (tablets, biscuits, capsules, powders) will require suitable packaging to prevent resorption of water molecules and consequent microbial growth.
It has been noted that condensed moisture films can form on the surface of supposedly otherwise “dry” products such as tablets or bulk oils following exposure to a damp atmosphere and fluctuating storage temperatures, resulting in sufficiently high localized Aw to initiate fungal growth. In addition, metabolically product water in the cytoplasm may raise Aw and initiate and allow more growth. It has also been found that dilute aqueous films similarly formed on the surface of visous syrups, or exuded from hydrogels by syneresis, can and do reach sufficiently high Aw to permit yeast and fungal spoilage.
7. Product life (shelf-life)
Modest levels of non-pathogenic microorganisms may be tolerated in an extemporaneous product with a short-life of several days. In the same vein, similar initial levels might be quite unacceptable in a factory-produced formulation which could be many weeks old and the product could have suffered marked microbial attack before it is administered to the patient.
Prolonged storage of a parenterial solution containing (as a consequence of an inadequate heat sterilization procedure) an extremely low level of viable but damaged bacteria, might enable recovery, marked multiplication, and pyrogen release to transform a relatively innocuous preparation into a potentially lethal one.
8. Redox Potential
Spoilage prevention of pharmaceuticals involving physical removed of oxygen is not always practicable. The inhibitory effect of pressurized C02 for soft drinks is due to the specific microbicidal action of carbonic acid rather than low oxygen content. The reductive and viscous nature of some food distuffs that may encourage anaerobic spoilage are uncommon with pharmaceuticals.
Therefore the oxidation reduction balance of a formulation is determined partly by its oxygen content and partly by its ingredients.
9. Package design
It could have been better in terms of eliminating the probability of microbial contamination to use unit-dosage packs than the use of multidose containers. However, the cost of packaging in unit dosages prohibitively outweigh the cost of dispensing in multidosage container. Faulty packages can be a source of microbial contamination that will eventually degrade the medicament. Multidose injection containers must be properly sealed by self-sealing rubber substance following probable puncture by a hypodermic needle. Moisture vapour and oxygen can penetrate inferior quality packaging plastics. Cap liners of cork or cardboard can easily absorb moisture and support microbial growth if not impregnate with a preservative. Wide mouthed ointment and cream jars can readily allow the entry of contaminants and microbial nutrients via contact with bare fingers or occasional direct coughs etc into the containers during usage.