770 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS inoculation and range of fungal organisms is generally the same as used in the testing of creams and lotions. Moulds isolated from spoiled products should also be used. After inoculation, incubate test and control samples at 22-25 ø in a moist atmosphere as previously described. Inspect regularly for evidence of fungal growth for a test period of not less than 3 months. 3.523 Saliva test A test which simulates the customary practice of the user in moistening such products as eyeliners and mascara with saliva, thereby contributing large numbers of micro-organisms to the preparation, may be carried out in the laboratory. For this purpose, pool together 4-6 samples of fresh saliva. The mixed sample will carry a rich microbial flora that is normally indigenous to saliva, including large numbers of Streptococci, Staphylococci, and Neisseria. Spread an inoculum of about 0.2 ml of the pooled saliva over the surface of duplicate preparations. Incubate the test samples, and a control, in a moist atmosphere at 22 ø. Periodically moisten dry samples with 0.1-0.2 ml of sterile distilled water. Observe bactericidal action by culturing the infected samples on Blood Agar at 37 ø. Sub-culture at regular intervals according to the schedule previously described for bacteria. If a complete kill has been achieved, the challenge test may suitably be repeated on the same sample with fresh saliva. 4. CHOICE OF PRESERVATIVES 4.1 Requirements Modern cosmetic preparations, unless adequately preserved, often present a favourable environment for microbial growth. Apart from the health hazard, this may cause separation of emulsions, discoloration, changes in viscosity, the formation of gas and of off-odours. The extent of microbial growth depends largely on the type of formulation, water content, phase composition of an emulsion, pH, nutritive value, amount and type of perfume, and the type of container in which the product is packed. After manufacture, most products are likely to be exposed to further con- tamination from filling machines, containers and caps. Fungal growth on the surface of cosmetic creams has often been traced to infected cap-liners. Products packed in wide-mouth jars are easily contaminated by the user's fingers. Flexible bottles which draw air back into them are liable to suffer airborne contamination during use. Products packed in collapsible tubes and bottles with small apertures are less prone to contamination in use but

HYGIENIC MANUFACTURE AND PRESERVATION 771 they may exhibit spoilage owing to the difficulties experienced in cleaning these packaged components effectively before the product is filled. Creams and lotions generally need to be protected against attack by moulds, yeasts, aerobic and anaerobic bacteria by means of a suitable pre- servative. Organisms of particular concern from the public health aspect and which are most likely to be present as contaminants are Pseudomonas aeruginosa, Klebsiella species, and coagulase-positive Staphylococcus aureus. The dark pigmented fungi, such as Cladosporium and coloured moulds, can be a particular nuisance since they so obviously disfigure the product. Although they do not present a serious public health hazard, they could possibly serve as allergens. Anionic detergents in many shampoos are particularly vulnerable to bacterial decomposition. Shampoo prepared from such materials must be protected against gram-negative bacteria generally, and Pseudomonads in particular. Preparations intended for application to or near the eye such as solid or liquid eyeliners, eyelash materials, and pigmented materials for make-up around the eye require particular attention. These preparations should be sterile at the time of manufacture and should also contain a preservative known to be bactericidal to Pseudomonas aeruginosa. Similar precautions are necessary for products to be used on newborn infants or for application to broken skin. Powders, such as raw talcs and mineral earth pigments, may contain large numbers of fungal spores, aerobic and anaerobic bacteria including Clostridial spore-bearing species when used for cosmetic purposes they may need to be subjected to an efficient form of presterilization. Reliance, in this connection, should not be placed on the incorporation of a preserva- tive. 4.2 Recommendations A preservative should always be regarded as an additional aid in pro- longing the shelf-life of a product. It must never be used to mask deficiencies in hygiene. In the avoidance of spoilage, good housekeeping is essential to cosmetic manufacture just as it is to food production. Notwithstanding good manufacturing conditions, unforeseen and exceptional contamination may occur during routine manufacture which will overwhelm an in- adequate preservative system and render it ineffective. The preservative capacity of a product should therefore exceed the minimum requirements