ASTM F1608−21. Standard Test Method for Microbial Ranking of Porous Packaging Materials (Exposure Chamber Method).

Test with the Certificate of Good Laboratory Practices (GLPs).

ASTM F1608 describes a test to determine the penetration of airborne bacteria through porous materials used in the packaging of sterile medical devices. Porous packaging material is understood as a material used in medical packaging, which is intended to provide an environmental and biological barrier, while allowing sufficient airflow for gaseous sterilization procedures, such as, for example, carbon dioxide. ethylene, steam or gas plasma.

This assay is designed to evaluate materials under conditions that allow detection of the passage of bacterial spores through the material under test. To do this, the material is exposed to an aerosol of Bacillus atrophaeus spores. The exposure is made in a chamber that consists of two main parts, one upper and one lower. The spores are nebulized at the upper part of the chamber with a nebulizer that generates a bacterial aerosol that is distributed throughout the chamber with a fan. The bottom of the test chamber has several outlet ports for intended to place the test samples and controls. In each of the outlet ports, there is a filtration system, each with its own filtration membrane. At the outlet sample ports, the test sample is placed on the filtration membrane. By establishing an airflow through the samples at 2.8 L/min for 15 minutes, if spores pass through a porous sample, they are collected on the filter membranes. A minimum of two samples for each material should be tested, although it is recommended that a larger number of samples of each material be used. One of the outlet sample ports is used as inoculum control and negative growth control. To do this, two superimposed filtering membranes are placed in this outlet port, one on top of the other. The upper filtering membrane corresponds to the inoculum control (N0) since it is used to determine the inoculum of spores to which each sample is exposed (as the spores are retained in this first membrane), while the lower filtering membrane corresponds to the control growth negative, since spores should not reach it because they are retained in the upper membrane. Also, a positive penetration control port can be included, in which a material known to allow the passage of a certain level of spores is placed. After exposure, each of the filtering membranes of each one of the samples is recovered, the spores contained in them are extracted, and the number of spores on each membrane is determined by quantitative culture. With the count obtained, the CFUs that have passed through each sample of test material (N1), those collected in the inoculum control membrane (N0), in the negative control, and those that have passed through the positive control, if included, are determined. The ability of a packaging material to resist the passage of microorganisms is expressed as the Log Reduction Value (LRV). The LRV is calculated by comparing the logarithm of the number of spores that pass through the porous material with the logarithm of the inoculum control (LVR= log N0 - log N1). Results can also be expressed as the percentage of spores retained by a test material.

The data obtained from this test is useful in evaluating the relative potential of a porous material to allow loss of sterility of the contents of the container. If a packaging material does not allow any passage of spores, all that can be stated is that it has resistance to penetration at the test conditions.

The standard establishes a set of parameters to perform the test: a microbial inoculum of 1 x 106 CFU per sample port, a flow rate through the test sample of 2.8 L/min, and an exposure time of 15 min. However, it establishes that, after performing the test under these conditions, these test conditions can be modified, if other conditions are considered more suitable to determine if the porous test material is appropriate for the intended use of the product.

To perform the test with the minimum number of test samples (two samples), enough quantity of the test material is required to prepare a minimum of 12 samples of an approximate size of 50 mm in diameter, or a higher number of samples proportional to the number of exposure tests requested. Sample disks are generally sterilized prior to testing using a sterilization method appropriate for the material tested, which should be specified by the customer. The appropriate method for sterilization must be indicated on the request form or the discs of test material must be sent already prepared and sterilized. Materials can also be tested before or after they are subjected to certain environmental conditions, such as heat or cold, relative humidity, different sterilization processes, or accelerated aging. If it is required to use a control of porous material permeable to microorganisms, it must be provided by the client.