The author does not know of any regulatory-defined microbiological standards for the manufacture of nonsterile liquids, ointments and semisolids. Standards for sterile manufacture are, on the other hand, well defined.

At first it might seem that the direction for defining microbiological standards for nonsterile manufacturing environments would be to "scale down" those already defined for sterile manufacture. However, this is not as easy as it would appear. Environmental microbiology standards for sterile manufacture are mainly quantitative (cfu per m3, cfu per plate per hour, cfu per cm2, etc.) and are limited to extremely low numbers, often zero. The extremely highly specified environmental controls applied in sterile manufacture make it possible to set and achieve these limits. When environmental controls are less stringent (as in nonsterile manufacture), the numbers of microorganisms not only increase, they also show more variability.

The consequences of the variability in numbers of microorganisms recovered from nonsterile manufacturing areas, and the relative weakness of the environmental control mechanisms, diminishes the effectiveness of quantitative standards. Limits are either set so high to accommodate the variability that they become meaningless, or set so strictly that they do not accommodate the intrinsic variability. They therefore result in frequent out-of-specification (OOS) conditions that cannot be sensibly rectified. In this latter case, the microbiological environmental monitoring programme most often falls into disrepute.

On the other hand, there are good arguments for setting limits for absence of the microorganisms, which indicate diminishing standards of hygiene, operator malpractice, system breakdown, etc. Gram-negative microorganisms are the greatest risk to liquids, ointments and semisolids. Although Gram-negative organisms are ubiquitous in nature in water and in drains, they are not all identically significant as potential contaminants of these preparations. Known Gram-negative pathogens should not be tolerated in manufacturing areas. However, there are other Gram-negative types, e.g., Enterobacter agglomerans, which need not always be of great concern.

The nonsterile environmental monitoring programme should balance the

(a) Types of Gram-negative organism (which may or may not be tolerable against the locations monitored)

(b) Type of product manufactured

(c) Severity of action required when they are isolated (see the scheme shown in Table 4.4)

Table 4.4 Suggested Approach to Monitoring Liquids, Ointments and Semisolids Manufacturing Areas for Gram-Negative Microorganisms

Location Actions Required in the Event of

Detection of P aeruginosa, Detection of other Gram

E. coli, Salmonella or other negative species confirmed pathogen

Protected hygienically controlled locations such as manufacturing equipment, filling machines, etc.

Other less critical areas where sources of microbiological contamination might be anticipated, e.g., areas where water is used

• Suspension of manufacture

• Action on product (rejection, recall)

• Corrective/preventive actions

• Suspension of manufacture if persistent

• Corrective/preventive actions

• Suspension of manufacture if persistent

Restrictions are not recommended because some isolation of these species is likely to be expected

Known Gram-negative pathogens are customarily addressed via attempted isolation of the indicator organisms, Pseudomonas aeruginosa, E. coli and Salmonella. Selective media are best used to ensure that these indicator organisms are not obscured by other, more resilient, microorganisms, which grow more rapidly and extensively on general media. The consequences of finding Pseudomonas aeruginosa, E. coli or Salmonella or other pathogens on manufacture or filling equipment are likely to be severe — batch rejection is a possibility, but this must be considered on a product-by-product, incident-by-incident basis. The consequences of finding them in a wash-bay, for instance, should not be as severe. Isolation of indicator organisms from these less critical areas should be seen as an early warning of a contamination source, which could lead eventually to manufacturing equipment or product contamination. Corrective and preventive actions against sources of contamination should be mandatory, and manufacture may have to be suspended if the problems persist.

Table 4.5 Recommended Frequencies for Nonsterile Environmental Monitoring

Type of Monitoring

Aqueous Inhalations

Creams and Lotions

Oral Liquids

Ointments and Gels

Microbial monitoring of surfaces by swabbing, or by contact plates

On each day of manufacture or filling

On each day of manufacture or filling


Every two weeks

Viable microorganisms in air recovered by active sampling

On each day of manufacture or filling


Every two weeks



3D O

All Gram-negative microorganisms should be restricted from manufacturing equipment, filling machines, etc. General media are best used for this purpose, alongside the selective media used for the indicator organisms. The consequences of finding nonpathogenic Gram-negative species on manufacturing and filling equipment merit corrective and preventive action against the sources of contamination, but not necessarily action against product. As they may be indicative of the presence of pathogenic species at levels below the sensitivity of the detection methods used, manufacture may have to be suspended. It is to be expected that some Gram-negative microorganisms will be isolated from less critical areas, particularly those in which water is present. Unless isolates are of the indicator types or other confirmed pathogens, their importance should not be exaggerated.

Gram-positive types may be similarly dealt with, concentrating on isolation of Staphylococcus aureus on suitable selective media. Some incidence of ubiquitous environmental Gram-positive types (such as Bacillus spp. and Micrococcus spp.) is inevitable, and unless excessive, should not give too much cause for concern. Grampositive types should be considered of greatest significance in locations from which personnel are excluded and locations (e.g., within filling cabinets) where personnel are expected to disinfect after rare and unusual intrusions. Isolation of Staphylococcus aureus in such locations, for example, means that something is not happening in the manner intended.

Table 4.5 indicates suitable frequencies and methods for microbiological environmental monitoring of facilities for manufacture of liquids, ointments and semisolids.

Actions and sanctions should be expected and applied when there are infringments to microbiological limits applying to the manufacturing environment for liquids, ointments and semisolids. The most likely actions arising out from out-of-specification or atypical nonsterile environmental results are those relating to the control of the process, or relating to operators and facilities.

The most frequently required actions are disinfection of an area or piece of equipment, or counselling to retrain personnel. These are typical corrective actions. Corrective actions are defined in terms of fixing the immediate problem. Additionally, attention must be given to preventive actions, for instance, replacing a defective item of equipment. Preventive actions are defined in terms of making sure the problem cannot arise again.

The most serious actions that can arise from an out-of specification result from microbiological environmental monitoring of nonsterile manufacturing areas are for product withdrawal (recall) or rejection. Neither action is likely to be required as a result of environmental data with no evidence of actual product contamination. Nonetheless it would be difficult to justify continuing manufacture of, for example, a paediatric syrup in equipment from which E. coli or Salmonella is repeatedly isolated, or to manufacture an inhalation in equipment from which Pseudomonas cepacia is isolated. Suspension of manufacture is a viable and probable option pending thorough investigation, diagnosis of the root cause of the problem and implementation of adequate corrective and preventive action.


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3. Casey, W.M., Muth, H., Kirby, J., Allen, P. Use of Nonselective Preenrichment Media for the Recovery of Enteric Bacteria from Pharmaceutical Products. Pharmaceutical Technology, 22, 1998.

4. Enigl, D.C., Sorrels, K.M. Water Activity and Self-Preserving Formulas. In Preservative-Free and Self-Preserving Cosmetics and Drugs: Principles and Practice (eds. J.J. Kabara, D.S. Orth). Marcel Dekker, New York, 1997.

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