Cosmetic products should be safe under normal or reasonably foreseeable conditions of use. The regulation on cosmetic products (EC 1223/2009) requires for the cosmetic product safety report the results of preservation challenge test to prove the microbiological safety. The regulation itself didn’t specify the test procedure for the challenge test. Beside the test from the EU and US pharmacopoeia in-house test protocols are established since many years. The applicability of the schülke KoKo test for the evaluation of the microbiological safety of finished cosmetic products is assessed in this paper.
Demands for challenge testing are specified only in the SCCS notes of guidance. In paragraph 4-4 “Guidelines on Microbiological of the Finished Cosmetic Product“ in the 8th revision (December 2012)  of the SCCS publication “Notes of Guidance for the Testing of Cosmetic Ingredients and their Safety Evaluation” is an obligation for carrying out a preservation test with all cosmetic products which could be contaminated under normal storage and usage conditions or if an infection risk for the consumer exists.
The 7th Amendment to the Cosmetics Directive has introduced a new requirement on the labelling of cosmetic products. Products with a minimum durability of more than 30 months must indicate the period of time after opening for which the product can be used without any harm to the consumer. This information shall be indicated by the symbol and the period itself in months and/or years. For products with a minimum durability of more than 30 months, it is not allowed to indicate the minimum durability instead of the “Period after Opening”.
A variety of relevant methods may be used to support the period indicated on a product, including those used during product development, since there is no officially sanctioned methodology that could be used. Examples of sources of information for assessing a product’s PaO may include:
- microbiological challenge tests
- stability data - analytical data (e.g. preservative analysis)
- type of packaging - experience with similar formulations and products
- consumer habits and practices.
In the first step some tissues are removed from the original wet wipe packaging. From these tissues a stack with 20 tissues à 1g (total weight 19.9 – 20.1g) is produced and then placed in a seal bag (LD-PE, 90µm, 120 x 170 mm). This test packaging is then inoculated with 0.2 ml (0.1 ml on top side and 0.1 ml on bottom side) of a solution containing the following test organisms. The titre of this inoculation solution is 106 –107 germs/ml.
The inoculum contains pathogenic microorganisms as germs which are well known for product spoilage. All species have to be cultivated separately and mixed directly before the addition, to ensure a constant composition and germ count of the inoculum. After inoculation the whole packaging is kneaded properly and, thereafter, incubated for three weeks at 30°C. The evaluation is done by placing pieces of tissue (approx. 3 x 3 cm, cut out of the tissue pack in one block) on CS and SA agar. In every case double determinations are done. The growth on the tissues is evaluated after three days of incubation at 30°C (for bacteria) and seven days at 25°C (for moulds). Fungi are classified into yeasts and the different mould types.
A sample can be called well preserved, if it passes a period of three weeks under the above described laboratory conditions without showing microbial growth on the test batches. That means even after inoculation no microbial growth can be observed. From many years of experience in the use of this test method these results can state the microbiological stability of 30 months which is recommended for cosmetic products.
In this test, the schülke KoKo test, a mixture of bacteria, yeast and moulds are inoculated 6 times (once a week) into the test material (see graph), with the goal of keeping the test material germ free for this period. The inoculum contains pathogenic microorganisms as gems which are well known for product spoilage. All species have to be cultivated separately and mixed directly before the addition, to ensure a constant composition and germ count of the inoculum. Its germ count is approximately 106-7 cfu/ml, which means a germ count of approximately 105 cfu/ml in the sample.
A sample can be called well preserved, if it passes a period of six weeks under the above described laboratory conditions without showing microbial growth on the test batches. That means even after the sixth inoculation no microbial growth can be observed. From many years of experience in the use of this test method these results can state the microbiological stability of 30 months which is recommended for cosmetic products.