In pharmaceutical, medical devices, cosmetics or food production, it is not only a requirement to clean but also to disinfect facilities. The type of disinfectants used and the particular procedures and techniques that are suitable depend upon the classification of the relevant cleanrooms and the products manufactured there. Taking the classifications set down in the EU Guidelines for Good Manufacturing Practice as the basis, the following picture emerges:
Annex 1 Grade D and C
In these areas commercially available disinfectants are used. There are no further requirements concerning their filling and packaging beside the classic requirements for biocidals. The company's sanitation protocol must contain the disinfectants to be used — inspectors usually expect a positive list of detergents and disinfectants contained in the sanitation protocol or the relevant standard operating procedures (SOPs). The intervals between disinfection, the application concentrations/exposure times and the equipment to be used, such as mops or wipes, must also be defined.
The efficacies of the disinfectants should also be tested. This means that the disinfectants must at least be included in the national lists of disinfectants, such as the VAH (www.vah-online.de) in Germany or AFNOR (www.afnor.org) in France for example — or there have to be comparable assessments carried out according to the European test methods.
- EN 1276 Test for bacteria with an expected 5log reduction
- EN 1650 Test for fungi with an expected 4log reduction
- EN 13704 Test for spores with an expected 3log reduction
- EN 13697 Quantitative non-porous surface test for the evaluation of bactericidal and/or fungicidal activity of chemical disinfectants
Usually the cleanroom operator will re-test the efficacy prior to release for use in the production environment, and may also test efficacy against microorganisms found during monitoring of that environment.
Annex 1 Grade A and B
The requirements are substantially stricter for these grade areas, i.e. in the area of aseptic production or the immediate vicinity. The EU Guidelines on Good Manufacturing Practice (Annex 1) as well as the US FDA’s Aseptic Guide require the use of disinfectants that are “sterile prior to use”. This means in practice that the products used are sterile-filtered at filling, are double- or triple-bagged and are irradiated after packaging to ensure the sterility of the primary and secondary packaging.
Furthermore a comprehensive validation of the efficacy is required for these disinfectants. This means that the standard tests using the reference germs for the national listings according to VAH (DGHM) or the European EN standards are not sufficient. Moreover, the efficacy has to be tested using microorganisms from ‘in-house’ monitoring on the particular surface materials to be used. These additional tests, including efficacy against spores, which usually the cleanroom operator’s lab can not carry out and which can cost between €10,000– €50,000, depending on their extent.
Calculation of laboratory costs for a disinfectant used in a CLASS A/B cleanroom (data from a contract lab)
| For one disinfectant at only one concentration, the following is required: |
| 24 agar plates |
| inhibition materials |
| 24 surface plates at 5 cm x 5 cm |
| glass balls, pipettes etc |
| Time taken |
| About 2 hrs for the test, along with the appropriate time for nutrition tests, buffer and inhibitor mixing etc. |
| Calculated cost approximately €150 |
| Example costs for the validation of four surface disinfectants |
| x 5 surfaces (e.g. Pharmaterrazzo, glass, polycarbonate glass, stainless steel, melamine) |
| x 2 concentrations / dwell times |
| x 8 microorganisms (5 reference microorganisms and 3 isolates |
| x €150 |
| Calculated cost approximately €48,000 |
It is also required that the disinfectant active substances are changed (in rotation) for the aseptic areas. Furthermore, the availability of at least one disinfectant with efficacy against bacterial spores is required for this area. Traditional biocides, such as alcohols, quaternary ammonium compounds, phenols and amphoteric surfactants are effective against bacteria in their vegetative phase but ineffective against bacterial spores. Products containing hydrogen peroxide, peracetic acid (peracids), hypochlorite, chlorine dioxide or formaldehyde as active substance are effective against spores, but caution is needed as some of these products can cause corrosion on metal surfaces and unpleasant odours!
It must also be observed that chemical or physical reactions might occur among single active substances, which can lead to discolourations or in the worst case scenario, to inactivation. Amino compounds and aldehydes may lead to a colour reaction and quaternary ammonium bases react with anionic surfactants (e.g. from the preliminary cleaning).
If the disinfectant containers are used more than once (such as canisters or spray bottles containing an alcoholic solution), evidence of the sterility after opening is required.
In all cases, the product must be accompanied by the required batch documents and certificates of sterility, irradiation and where available, conformity. For these cleanroom classes, all measures and products used for cleaning and disinfecting must also be defined and documented in writing. This includes the products and application devices used, the frequency of disinfection measures, the frequency of rotation, the responsible person(s), the description of the procedure and the concentrations and exposure times.
Summary of the most important requirements
| Sterile filtration (0.2 µm) |
| Filling under cleanroom conditions |
| Double- or triple-bagged |
| Gamma irradiated |
| Certificate for each batch (if necessary online retrieval) |
| Certificate of sterility after opening for ready-to-use solutions (shelf-life) |
It is necessary to carry out a validation test with regard to residues for surfaces near products or for surfaces with product contact (according to PIC/S PI 006-1) to prevent contamination of the product with chemical residues from the disinfectant.
Exposure times
We are asked repeatedly whether a disinfected area has to be kept wet during the exposure time prescribed. This statement seems to be circulating on the market that this is a regulatory requirement and the suggestion is that this is described in the EN method. But this is not stipulated in any European guideline known to us. Principally, the EN does not stipulate that the surface has to remain wet during exposure time, but according to EN 13697 the exposure time for bacteria is 5 min and 15 min for yeasts and moulds. Since 100 µl of disinfectant is applied on the surfaces, these almost always remain wet during these short exposure times. Hence, the standards do not specify the wetting time but the amount of disinfectant to be applied for the test. After applying the product, it is left to react during the exposure time (irrelevant of whether the surface dries during this time or remains wet).
If the exposure times required are longer, such as in national listings, e.g. in the list of disinfectants of the VAH or in the product descriptions of many disinfectants, no rewetting is carried out either. If there are concerns in practice that the use in premises with an increased air change rate will lead to significantly quicker drying times than those prescribed in the test procedures (or in the validation in the laboratory) and could actually adversely affect the efficacy of the disinfection procedure, the monitoring data should be used to show whether efficacy is lacking in the case of existing procedures that have already be in use for a long time.
In this case or in the implementation of a new disinfection procedure, one could actually determine on-site the time needed for drying and review the reduction during this time in the laboratory testing/validation. This would mean that in laboratory testing, the disinfectant would be inactivated after the time determined for drying in practice, and its efficacy tested. Should the reduction prove to be insufficient, an increase in concentration could be a possible means to reduce the exposure time.
But in principal it is true that an active disinfection ingredient continues to have an effect on a microorganism’s cells once applied, even after the surroundings have become dry. And it has to be taken into consideration that, usually, the microbial concentrations applied for testing in the laboratory (depending on the test, at least 106 cells) are much higher than the microbial concentrations that usually are present in a cleanroom. This means that, even in the case of a smaller reduction in laboratory, the limits set are usually not exceeded in practice and the monitoring data should comply with the requirements.
For all important aspects of controlling disinfection visit Contamination Control on 15-17 November 2017 in Barcelona, Spain. www.gmp-compliance.org