Discovering Bacterial Secrets in Cosmetics: Fast and Accurate Detection with Molecular Methods
The presence of unwanted bacteria in cosmetic products is a serious health threat to consumers worldwide. In addition, bacterial growth negatively impacts the quality of cosmetic products.
Bacterial contamination of cosmetic products can occur due to various components in cosmetics that support bacterial growth, such as sugar, vitamins, proteins, oils, and water. The risk of contamination increases if cosmetic manufacturing practices are not carried out properly during the production process.
Contaminated cosmetics are at risk of being damaged and harming consumers. Therefore, it is very important to ensure that cosmetic products are free from bacteria.
Over the past 30 years, the implementation of good manufacturing practices (GMP) has become the basis for improving quality control analysis in the cosmetic industry. As part of GMP, the microbial limit test by the United States Pharmacopoeia (USP) provides a method for determining the total number of bacteria, yeast, or mold in products to be manufactured.
USP sets 4 bacterial indicators, namely Salmonella spp., Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli. Based on its regulations, cosmetic products must be free from contamination by these bacteria.
Detection of bacterial contaminants has so far been carried out using cultivation-based methods. However, now there are methods available with a molecular approach that can detect bacteria in contaminated samples more quickly. One method that is often used is Polymerase Chain Reaction (PCR) technology.
Polymerase Chain Reaction (PCR) as a Method for Detecting Bacterial Contamination
The PCR method has been developed and validated as a relatively rapid method for detecting bacteria in cosmetic samples. This method will show the presence/absence of certain genetic sequences.
In the PCR reaction, the detection of the presence of bacterial contaminants is carried out by DNA polymerase which is able to amplify specific target genetic sequences.
Identification of bacterial DNA by PCR utilizes a special gene called the 16S rRNA gene. The use of this gene has become a standard molecular method, both in the laboratory and in the clinical environment. The 16S rRNA gene is very specific for each type of bacteria, making it an ideal target for identification.
Although the 16S rRNA gene is often used for bacterial identification by PCR, there are times when this gene is the same in two closely related species. In such cases, other genes such as rpoB, tuf, gyrA, gyrB, and heat shock proteins are used as targets for identification.
The PCR device automatically conditions different thermal cycles based on the process. The sample will be heated to 95ºC to start the reaction. At the same temperature, an additional 30 seconds are carried out in order to separate the DNA strands.
The temperature is lowered to 65ºC for 1 minute to allow the primers to attach to the target DNA strand. The temperature is raised again to 72ºC and left for 90 seconds for the DNA amplification process, the steps are repeated 35 times.
After the PCR results have been obtained, further analysis to separate and examine DNA fragments can be carried out using the agarose gel electrophoresis technique. Then, the DNA is sequenced and BLAST analysis is carried out on the bacterial DNA sequence to identify them.
There are several other methods that have the same working principle, namely real-time PCR and Random Amplification of Polymorphic DNA (RAPD-PCR).
PCR analysis can provide a rapid evaluation of the quality of cosmetic products so that corrective actions can also be taken earlier
Advantages of Using Molecular-Based Methods
Molecular-based methods, especially PCR, allow for rapid, sensitive, and accurate detection of microbial strains and types. This method works by detecting important molecules, such as DNA (deoxyribonucleic acid).
PCR is known as a very sensitive technique for detecting bacteria because it can rapidly amplify target nucleic acids from very small initial amounts.
PCR is able to analyze the presence of bacteria quickly, which only takes 24 hours or 27 hours.
Burkholderia cepacia is one of the most frequently isolated bacterial contaminants in cosmetic samples worldwide. Standard biochemical test methods for the isolation and identification of this bacteria take 5–6 days to complete.
Burkholderia cepacia has genetic variations that make it difficult to detect with conventional biochemical tests. However, the PCR method can quickly detect this bacteria within 27 hours using PCR Ready-To-Go beads. This method is simpler because it only requires adding samples and specific DNA primers for Burkholderia cepacia.
The PCR method is not only faster than conventional methods that use culture, but it can also identify bacteria that are difficult to grow in the laboratory.
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