Antifungal Product Testing – ASTM Test Methods

 Antifungal Test Methods ASTM G 21, D 5590, and D 3273

Antifungal products are those designed to provide resistance to the growth or effects of fungi, including mold, mildew and other forms of fungi.  The range of products that should be treated with an antifungal is extensive and can include the components of raw materials, formulation additives, or the ultimate finished product.

A common example for the use of an antifungal treatment would be drywall, a long-term building component that receives extensive exposure to environmental challenges, especially moisture or high humidity that promotes growth of fungi.  Most building surfaces are exposed to the many types of fungal spores prevalent in every day air.  The degree of exposure is both seasonal and geography-dependent but typically, it can be assumed that there are fungal spores present that will germinate and grow once the appropriate conditions are met.  For drywall the materials are generally comprised of a paper outer layer (or several layers) and a gypsum core.  Both the outer layer and core can have additives put into them to supplement their physical properties, and one of these additives could be an antifungal agent.  Depending on the use environment or performance desired by the product, different amounts and types of antifungal additives could be used.

The Rule of 3 provides guidance on whether or not fungi will begin to grow on a given material.  For product developers, how to best optimize their products will depend on the product development process and implementation of an appropriate testing strategy. There are many test methods used to make this assessment, but the most common methods used in North America are those provided by the American Society for Testing and Materials (ASTM).  ASTM provides a standardized series of test methods that are rigorously updated and harmonized for global applicability.  For our laboratory, the most frequently requested test methods include the ASTM G 21, ASTM D 3273, and ASTM D 5590.  These antifungal test methods are targeted at specific product types (such as coatings for ASTM G21 or ASTM D5590, or drywall for ASTM D 3273), but are also used as standards in the development of a wide range of product types.

Laboratories experienced in microbiological testing know that there is inherent variability in the testing of microorganisms (Upcoming  – Microbiological Testing and Inherent Variability).  Qualitative testing, such as the ASTM test methods discussed above, do a good job of providing Yes/No results–but not as good in providing degrees of Yes or No.  We like to say that ‘It’s the nature of the beast’ but the reality is that finished products are complex and deriving an analytically accurate results is most often not possible or cost prohibitive.

For each of the ASTM Antifungal test methods, I will briefly discuss their intended purpose, principle attributes, useful applications and some test limitations.  All test methods have limitations because it is simply not possible to precisely replicate the in situ (in use) environment for a given products’ lifespan, in a laboratory setting.   That being said, manufacturers and microbial laboratories can improve the test value by including critical ‘durability’ factors and incorporate into their product testing those variables that most represent the intended use environment.

ASTM G 21Standard Practice for Determining Resistance of Synthetic Polymeric Materials to Fungi

Intended Purpose of the ASTM G 21 Antifungal Test Method: Qualitative determination of inherent fungal resistance for a test coating, with supplementation of essential minerals but no carbon energy source (see – Rule of 3). –

Principle Attributes: This test will demonstrate if a tested material provides the needed carbon source that will support the growth of fungi and whether an additive or product attribute will protect the material from the fungal growth.

Useful Applications and Limitations: The ASTM G 21 test provides a straight forward measurement of the intrinsic fungal resistant property of the material.  As the fungi are applied to the material, they will either grow or not and this result is directly compared to a control sample that will grow fungi.  This is not a good method for assessment of durability unless a durability parameter is applied to the sample prior to the test.  However, ASTM G 21 is a very useful method to demonstrate antifungal properties for many types of materials, including paper, textiles, plastics, coatings, foams, wood, and others.  Test samples for the  ASTM G 21 must be  relatively  thin and flat to have contact with the test media, for coatings this is not often a challenge, but for molded materials that could be assessed using this method, sample shape can be an issue.  The most common fungi or mold used in the test are Aspergillus niger, and Penicillium citrinum. 

ASTM D 5590Determining the Resistance of Paint Films and Related Coatings to Fungal Defacement by Accelerated Four-Week Agar Plate Assay  –

Intended Purpose of the ASTM D 5590 Antifungal Test Method:  The ASTM D 5590 is similar to the ASTM G 21, with one critical exception; the test material used to test the samples contains a complete nutrient spectrum.  Therefore, the basis of the test is that fungi will always grow unless the test material provides an antifungal component preventing fungal growth.  This is a very significant difference, as fungi applied to material tested in the ASTM G 21, which does not satisfy the Rule of 3 typically by carbon limitation, will not grow; but, in the ASTM D 5590, the Rule of 3 is always satisfied and without an appropriate antimicrobial the test sample will fail and fungi will grow.

Principle Attributes:This test will demonstrate if a tested material provides active antifungal protection of a treated product, and represents a higher level of challenge to the test sample..

Useful Applications and Limitations: The usefulness of the ASTM D 5590 is targeted at a higher stringency antifungal performance.  Long-term use or outdoor exposure of a product always results in ‘soiling’ of any product or material, leading to the need for an antifungal additive.  This test method is a quick way of evaluating antifungal performance.  Much like the ASTM G 21, there is no inherent durability assessment incorporated into the test method.  The results will provide a solid indication of a products resistance or antifungal performance, but cannot provide an indication of durability.  If durability is a factor in the product performance, a specific products’ use environment should guide the test lab to include appropriate durability parameters.  Also as with the ASTM G 21, test samples must be essentially flat to fit into the test plate. Again, the most common organisms tested are Aspergillus niger, and Penicillium citrinum.

ASTM D 3273 Standard Test Method for Resistance to Mold on the Surface of Interior Coatings in an Environmental Chamber

Intended Purpose of the ASTM D 3273 Antifungal Test Method:  Is unique in that it is designed to simulate a harsh environmental exposure of the test material.  Principally, high humidity and temperature are the conditions most likely to promote fungal growth.  In addition, the test samples are continuously exposed to the fungal spores by virtue of a soil bed resting beneath the samples.  It is primarily intended for testing of building products, but there are many similar test methods employing similar strategies for other product types.

Principle Attributes of the ASTM D 3273:Test sample are suspended in an environmental chamber setup to grow fungi in a bed of soil.  Having both high humidity >95% and high temperature (32°C / 90°F), the samples experience a degree of exposure lacking in other antifungal test methods by virtue of water condensation and dripping from the samples over the 28 day incubation period. In addition to testing inherent antifungal resistance, the ASTM D 3273 will also determine if the products antimicrobial additive is not appropriate or appropriately formulated for high humidity and temperature.  For example, this test will reveal if the additive it too highly mobile and thus can “leach” out of the treated sample.  By contrast to either the ASTM G 21 or D 5590, additive mobility is not significantly challenged.

Useful Applications and Limitations of ASTM D 3273: Environmental chambers provide a unique perspective on the ability of a test material to resist fungal growth by providing the optimum environment and essentially continual inoculation of the sample to the fungal spores.  An advantage of this test is that non-standard samples can be tested eliminating the need for a flat uniform sample.  The main limitation is that the growth environment, while permissive to fungal growth on the sample, is not enriched for fungal growth.  Commonly, this results in partial coverage or growth of fungi, which can affect reproducibility of results within replicate test samples and from test-to-test.  The records for the level of fungal growth benefit from an experienced laboratory that can conclusively differentiate fungal growth and have a standardized method for assuring test sample reproducibility.

Footnote:  Each of these ASTM methods are scored by the laboratory to provide a degree of differentiation of the test samples, but don’t be mislead.  The scores provided are not representative of an actual number or metric which correlates to the degree of antifungal performance.  They are simply related to the degree of antifungal performance within a test set and will likely be subject to low reproducibility.  On the other hand, the Pass/Fail determination (Yes or No for antifungal performance) should be highly reproducible and useable for quality control, manufacturing, and product development purposes.  The ASTM D3273 specified both Aspergillus niger, and Penicillium citrinum, and Aureobasidium pullulans.

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