Real-time qPCR to evaluate bacterial contamination of cosmetic cream and the efficiency of protective ingredients.

AIMS: The absence of objectionable micro-organisms in cosmetics and the efficiency of preservatives are still mainly assessed by time-consuming cultivation-based methods. We explored the applicability of real-time quantitative polymerase chain reaction (qPCR) and reported on the behaviour of different bacteria in artificially contaminated creams. METHODS AND RESULTS: Real-time qPCR on DNA from Burkholderia cepacia, Pluribacter gergoviae, Pseudomonas aeruginosa and Sphingomonas paucimobilis identified specific primer pairs that amplify accurately and efficiently two strains/isolates of each species. Using DNeasy mericon Food Kit, we detected bacterial growth in an inoculated cosmetic cream and persistency of DNA from heat-inactivated bacteria. We were also able to monitor the growth inhibitory effect of caprylyl glycol and EDTA, also showing how different bacterial species interact depending on the presence/absence of these ingredients. Finally, creams supplemented with the protective cosmetic ingredients revealed the various behaviour of five strains/isolates from P. aeruginosa. CONCLUSIONS: Successfully extracting bacterial DNA from artificially contaminated cosmetic creams, we could perform real-time qPCR to identify and follow the growth of various strains of 4 bacteria species under different conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: Real-time qPCR appears as a promising method to detect bacterial contamination in cosmetic creams and/or to monitor growth inhibition by ingredients.

In vitro study to evaluate the antimicrobial activity of various multifunctional cosmetic ingredients and chlorphenesin on bacterial species at risk in the cosmetic industry.

AIMS: We evaluated the activity of the preservative chlorphenesin and of four antimicrobial cosmetic multifunctional ingredients against various strains of gram-negative and gram-positive human opportunistic pathogens. METHODS AND RESULTS: Growth kinetics, modelling growth parameters and statistical analyses enabled comparing bacterial behaviour in the presence and in the absence of the compound. Whatever compound tested (i.e. chlorphenesin, phenylpropanol, hexanediol, ethylhexylglycerin, hydroxyacetophenone) and strain origin (i.e. clinical versus industrial), the growth of 42 strains belonging to Acinetobacter spp., Burkholderia cepacia complex and Stenotrophomonas maltophilia, was totally inhibited. On the opposite all of the P. aeruginosa strains (n = 13) as well as 4 and 6 out of 10 strains of Pluralibacter gergoviae grew in the presence of chlorphenesin and ethylhexylglycerin, respectively. Some P. gergoviae and Staphylococcus hominis strains withstand hydroxyacetophenone. Within a species, the different strains show variable latency phase, growth rate (r) and carrying capacity (K). They can be similar, lower or higher than those measured in control conditions. CONCLUSIONS: Data showed differences in the antimicrobial activity of compounds. Upon exposure, strains differed in their behaviour between and within species. Whatever species and strains, compound sensitivity could not be related to antibiotic resistance. SIGNIFICANCE AND IMPACT OF THE STUDY: Most multifunctional ingredients showed significant antimicrobial properties against the wide panel of species and strains evaluated. This will help adjusting preservation strategies in the cosmetic industry.

Rapid assessment and prediction of the efficiency of two preservatives against S. aureus in cosmetic products using High Content Screening-Confocal Laser Scanning Microscopy.

Most cosmetic products are susceptible to microbiological spoilage due to contaminations that could happen during fabrication or by consumer's repetitive manipulation. The composition of cosmetic products must guarantee efficient bacterial inactivation all along with the product shelf life, which is usually assessed by challenge-tests. A challenge-test consists in inoculating specific bacteria, i.e. Staphylococcus aureus, in the formula and then investigating the bacterial log reduction over time. The main limitation of this method is relative to the time-consuming protocol, where 30 days are needed to obtain results. In this study, we have proposed a rapid alternative method coupling High Content Screening-Confocal Laser Scanning Microscopy (HCS-CLSM), image analysis and modeling. It consists in acquiring real-time S. aureus inactivation kinetics on short-time periods (typically 4h) and in predicting the efficiency of preservatives on longer scale periods (up to 7 days). The action of two preservatives, chlorphenesin and benzyl alcohol, was evaluated against S. aureus at several concentrations in a cosmetic matrix. From these datasets, we compared two secondary models to determine the logarithm reduction time (Dc) for each preservative concentration. Afterwards, we used two primary inactivation models to predict log reductions for up to 7 days and we compared them to observed log reductions. The IQ model better fits datasets and the Q value gives information about the matrix level of interference.

Development of a standardized method to evaluate the protective efficiency of cosmetic packaging against microbial contamination.

Doubts surrounding the potential adverse effects of antimicrobial preservatives have modified the demand of consumers, who increasingly insist on the production of low-level and even preservative-free cosmetics. Protection of the product against microbial contamination is therefore focused on the packaging. This has prompted the emergence of a highly diverse array of so-called "protective", "overprotective", and "barrier" packaging. However, these designations are not normalized and the choice of the right packaging adapted to each cosmetic product is still essentially empirical, hazardous, and time consuming. The Cosmetic Valleys cluster has launched a commission to define a complete and experimentally-validated method to classify the level of protection of cosmetic packaging against microbial contamination. As reported herein, this required the development a specific bacteriostatic medium that can be used for 7 days and an in vitro procedure that reproduces in-use contamination and consumer practices. Based on tests performed on over 800 packages of different origin and performance characteristics, we propose a classification, divided into six grades, to differentiate the protective efficiency of cosmetic packaging. This work can be considered as a first step towards a regulatory text.