Designation: G21 − 15

Standard Practice for

Determining Resistance of Synthetic Polymeric Materials to
Fungi1
This standard is issued under the fixed designation G21; the number immediately following the designation indicates the year of original
adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. A superscript
epsilon (´) indicates an editorial change since the last revision or reapproval.
This standard has been approved for use by agencies of the U.S. Department of Defense.

1. Scope D785 Test Method for Rockwell Hardness of Plastics and

1.1 This practice covers determination of the effect of fungi Electrical Insulating Materials
on the properties of synthetic polymeric materials in the form D882 Test Method for Tensile Properties of Thin Plastic
of molded and fabricated articles, tubes, rods, sheets, and film Sheeting
materials. Changes in optical, mechanical, and electrical prop- D1003 Test Method for Haze and Luminous Transmittance
erties may be determined by the applicable ASTM methods. of Transparent Plastics
D1708 Test Method for Tensile Properties of Plastics by Use
1.2 The values stated in SI units are to be regarded as the of Microtensile Specimens
standard. The inch-pound units given in parentheses are for E96/E96M Test Methods for Water Vapor Transmission of
information only. Materials
1.3 This standard does not purport to address all of the E308 Practice for Computing the Colors of Objects by Using
safety concerns, if any, associated with its use. It is the the CIE System
responsibility of the user of this standard to establish appro- 2.2 TAPPI Standard:
priate safety and health practices and determine the applica- Test Method T 451-CM-484 Flexural Properties of Paper3
bility of regulatory limitations prior to use. 2.3 Federal Standards:
FED STD 191 Method 5204 Stiffness of Cloth, Directional;
2. Referenced Documents Self Weighted Cantilever Method4
2.1 ASTM Standards:2 FED STD 191 Method 5206 Stiffness of Cloth Drape and
D149 Test Method for Dielectric Breakdown Voltage and Flex; Cantilever Bending Method4
Dielectric Strength of Solid Electrical Insulating Materials
3. Summary of Practice
at Commercial Power Frequencies
D150 Test Methods for AC Loss Characteristics and Permit- 3.1 The procedure described in this practice consists of
tivity (Dielectric Constant) of Solid Electrical Insulation selection of suitable specimens for determination of pertinent
D257 Test Methods for DC Resistance or Conductance of properties, inoculation of the specimens with suitable
Insulating Materials organisms, exposure of inoculated specimens under conditions
D495 Test Method for High-Voltage, Low-Current, Dry Arc favorable to growth, examination and rating for visual growth,
Resistance of Solid Electrical Insulation and removal of the specimens and observations for testing,
D618 Practice for Conditioning Plastics for Testing either before cleaning or after cleaning and reconditioning.
D638 Test Method for Tensile Properties of Plastics NOTE 1—Since the procedure involves handling and working with
D747 Test Method for Apparent Bending Modulus of Plas- fungi, it is recommended that personnel trained in microbiology perform
tics by Means of a Cantilever Beam the portion of the procedure involving handling of organisms and
inoculated specimens.

4. Significance and Use

1
This practice is under the jurisdiction of ASTM Committee G03 on Weathering
and Durability and is the direct responsibility of Subcommittee G03.04 on
4.1 The synthetic polymer portion of these materials is
Biological Deterioration. usually fungus-resistant in that it does not serve as a carbon
Current edition approved June 1, 2015. Published July 2015. Originally approved
in 1961. Last previous edition approved in 2013 as G21 – 13. DOI: 10.1520/G0021-
3
15. Available from Technical Association of the Pulp and Paper Industry (TAPPI),
2
For referenced ASTM standards, visit the ASTM website, www.astm.org, or 15 Technology Parkway South, Norcross, GA 30092, http://www.tappi.org.
4
contact ASTM Customer Service at service@astm.org. For Annual Book of ASTM Available from Standardization Documents Order Desk, DODSSP, Bldg. 4,
Standards volume information, refer to the standard’s Document Summary page on Section D, 700 Robbins Ave., Philadelphia, PA 19111-5098, http://
the ASTM website. dodssp.daps.dla.mil.

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 G21 − 15
source for the growth of fungi. It is generally the other cate glass, or baking dishes up to 400 by 500 mm (16 by 20 in.)
components, such as plasticizers, cellulosics, lubricants, in size, covered with squares of window glass.
stabilizers, and colorants, that are responsible for fungus attack 5.2 Incubator—Incubating equipment for all test methods
on plastic materials. To assess materials other than plastics, use shall maintain a temperature of 28 to 30°C (82.4 to 86°F) and
of this test method should be agreed upon by all parties a relative humidity not less than 85 %. Automatic recording of
involved. It is important to establish the resistance to microbial wet and dry-bulb temperature is recommended.
attack under conditions favorable for such attack, namely, a
temperature of 2 to 38°C (35 to 100°F) and a relative humidity 6. Reagents and Materials
of 60 to 100 %.
6.1 Purity of Reagents—Reagent grade chemicals shall be
4.2 The effects to be expected are as follows: used in all tests. Unless otherwise indicated, it is intended that
4.2.1 Surface attack, discoloration, loss of transmission all reagents shall conform to the specifications of the Commit-
(optical), and tee on Analytical Reagents of the American Chemical Society,
4.2.2 Removal of susceptible plasticizers, modifiers, and where such specification are available.6 Other grades may be
lubricants, resulting in increased modulus (stiffness), changes used, provided it is first ascertained that the reagent is of
in weight, dimensions, and other physical properties, and sufficiently high purity to permit its use without lessening the
deterioration of electrical properties such as insulation accuracy of the determination.
resistance, dielectric constant, power factor, and dielectric
strength. 6.2 Purity of Water—Unless otherwise indicated, references
to water shall be understood to mean distilled water or water of
4.3 Often the changes in electrical properties are due prin- equal or higher purity.
cipally to surface growth and its associated moisture and to pH
changes caused by excreted metabolic products. Other effects 6.3 Nutrient-Salts Agar—Prepare this medium by dissolving
include preferential growth caused by nonuniform dispersion in 1 L of water the designated amounts of the following
of plasticizers, lubricants, and other processing additives. reagents:
Attack on these materials often leaves ionized conducting Potassium dihydrogen orthophosphate (KH2PO4) 0.7 g
Magnesium sulfate (MgSO4·7H2O) 0.7 g
paths. Pronounced physical changes are observed on products Ammonium nitrate (NH4NO3) 1.0 g
in film form or as coatings, where the ratio of surface to Sodium chloride (NaCl) 0.005 g
volume is high, and where nutrient materials such as plasticiz- Ferrous sulfate (FeSO4·7H2O) 0.002 g
Zinc sulfate (ZnSO4·7H2O) 0.002 g
ers and lubricants continue to diffuse to the surface as they are Manganous sulfate (MnSO4·H2O) 0.001 g
utilized by the organisms. Agar 15.0 g
Dipotassium monohydrogen orthophosphate (K2HPO4) 0.7 g
4.4 Since attack by organisms involves a large element of
chance due to local accelerations and inhibitions, the order of 6.3.1 Sterilize the test medium by autoclaving at 121°C
reproducibility may be rather low. To ensure that estimates of (250°F) for 20 min. Adjust the pH of the medium so that after
behavior are not too optimistic, the greatest observed degree of sterilization the pH is between 6.0 and 6.5.
deterioration should be reported. 6.3.2 Prepare sufficient medium for the required tests.
6.3.3 Nutrient– Salts Broth—Prepare using the formula in
4.5 Conditioning of the specimens, such as exposure to 6.3, omitting the agar. Broth may be filter sterilized to avoid the
leaching, weathering, heat treatment, etc., may have significant precipitation of the salts that occurs with autoclaving.
effects on the resistance to fungi. Determination of these effects
is not covered in this practice. 6.4 Mixed Fungus Spore Suspension:
NOTE 2—Since a number of other organisms may be of specific interest
5. Apparatus for certain final assemblies or components, such other pure cultures of
5.1 Glassware—Glass or plastic vessels are suitable for organisms may be used if agreed upon by the purchaser and the
manufacturer of the plastic. Reference (1)7 illustrates such a choice.
holding specimens when laid flat. Depending on the size of the
specimens, the following are suggested:
5.1.1 For specimens up to 75 mm (3 in.) in diameter, 100 by 6
Reagent Chemicals, American Chemical Society Specifications, American
100 mm (41⁄4 by 41⁄4 in.) plastic boxes5 or 150-mm (6-in.) Chemical Society, Washington, DC. For suggestions on the testing of reagents not
covered Petri dishes, and listed by the American Chemical Society, see Analar Standards for Laboratory
5.1.2 For 75 mm (3 in.) and larger specimens, such as Chemicals, BDH Ltd., Poole, Dorset, U.K., and the United States Pharmacopeia
tensile and stiffness strips, large Petri dishes, trays of borosili- and National Formulary, U.S. Pharmaceutical Convention, Inc. (USPC), Rockville,
MD.
7
The boldface numbers given in parentheses refer to a list of references at the
5
Available from Tri-State, Inc., Henderson, KY. end of the practice.

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 G21 − 15
6.4.1 Use the following test fungi in preparing the cultures: 7. Viability Control
Fungi ATCC No.A 7.1 With each daily group of tests place each of three pieces
Aspergillus brasiliensisB 9642
of sterilized filter paper, 25 mm (1 in.) square, on hardened
Penicillium funiculosumC 11797 nutrient-salts agar in separate Petri dishes. Inoculate these,
Chaetomium globosum 6205 along with the test items, with the spore suspension by
Trichoderma virensD 9645
Aureobasidium pullulans 15233 spraying the suspension from a sterilized atomizer8 so that the
entire surface is moistened with the spore suspension. Incubate
A
Available from American Type Culture Collection, 12301 Parklawn Drive, these at 28 to 30°C (82 to 86°F) at a relative humidity not less
Rockville, MD 20852. than 85 % and examine them after 14 days’ incubation. There
B
Historically known as A. niger.
C
Historically known as P. pinophilum.
shall be copious growth on all three of the filter paper control
D
Historically known as Gliocladium virens. specimens. Absence of such growth requires repetition of the
test.
6.4.1.1 Maintain cultures of these fungi separately on an
appropriate medium such as potato dextrose agar. The stock 8. Test Specimens
cultures may be kept for not more than four months at
8.1 The simplest specimen may be a 50 by 50-mm (2 by
approximately 3 to 10°C (37 to 50°F). Use subcultures
2-in.) piece, a 50-mm (2-in.) diameter piece, or a piece (rod or
incubated at 28 to 30°C (82 to 86°F) for 7 to 20 days in
tubing) at least 76 mm (3 in.) long cut from the material to be
preparing the spore suspension.
tested. Completely fabricated parts or sections cut from fabri-
6.4.1.2 Prepare a spore suspension of each of the five fungi cated parts may be used as test specimens. On such specimens,
by pouring into one subculture of each fungus a sterile 10-mL observation of effect is limited to appearance, density of
portion of water or of a sterile solution containing 0.05 g/L of growth, optical reflection or transmission, or manual evaluation
a nontoxic wetting agent such as sodium dioctyl sulfosucci- of change in physical properties such as stiffness.
nate. Use a sterile platinum, plastic, or nichrome inoculating
wire to gently scrape the surface growth from the culture of the 8.2 Film-forming materials such as coatings may be tested
test organism. in the form of films at least 50 by 25 mm (2 by 1 in.) in size.
Such films may be prepared by casting on glass and stripping
6.4.2 Pour the spore charge into a sterile flask or tube
after cure, or by impregnating (completely covering) filter
containing 45 mL of sterile water with wetting agent and 10 to
paper or ignited glass fabric.
15 solid glass beads. Cap and shake the flask vigorously to
liberate the spores from the fruiting bodies and to break the 8.3 For visual evaluation, three specimens shall be inocu-
spore clumps. lated. If the specimen is different on two sides, three specimens
6.4.3 Alternatively, the spore charge can be poured into a of each, face up and face down, shall be tested.
sterile glass tissue grinder and gently ground to break up the NOTE 3—In devising a test program intended to reveal quantitative
spore clumps and liberate the spores from the fruiting bodies. changes occurring during and after fungal attack, an adequate number of
specimens should be evaluated to establish a valid value for the original
6.4.4 Filter the shaken or ground suspension through a thin property. If five replicate specimens are required to establish a tensile
layer of sterile glass wool in a glass funnel into a sterile flask strength of a film material, the same number of specimens shall be
in order to remove mycelial fragments. removed and tested for each exposure period. It is to be expected that
6.4.5 Centrifuge the filtered spore suspension aseptically, values of physical properties at various stages of fungal attack will be
variable; the values indicating the greatest degradation are the most
and discard the supernatant liquid. Resuspend the residue in an significant (see 4.4). Reference (2) may be used as a guide.
aliquot of sterile water and centrifuge.
6.4.6 If large mycelia fragments or clumps of agar were 9. Procedure
dislodged during the harvesting, wash the spores in this manner 9.1 Inoculation—Pour sufficient nutrient-salts agar into suit-
three times to remove possible nutrient carryover from the able sterile dishes (see 5.1) to provide a solidified agar layer
original cultures. Dilute the final washed residue with sterile from 3 to 6 mm (1⁄8 to 1⁄4 in.) in depth. After the agar is
nutrient-salts solution (see 6.3.3) in such a manner that the solidified, place the specimens on the surface of the agar.
resultant spore suspension shall contain 1 000 000 6 200 000 Inoculate the surface, including the surface of the test
spores/mL as determined with a counting chamber. specimens, with the composite spore suspension by spraying
6.4.7 Repeat this operation for each organism used in the the suspension from a sterilized atomizer8 so that the entire
test and blend equal volumes of the resultant spore suspension surface is moistened with the spore suspension.
to obtain the final mixed spore suspension.
9.2 Incubation Conditions:
6.4.8 The mixed spore suspension may be prepared fresh 9.2.1 Incubation—Cover the inoculated test specimens and
each day or may be held in the refrigerator at 3 to 10°C (37 to incubate at 28 to 30°C (82 to 86°F) and not less than 85 %
50°F) for not more than four days. The individual spore relative humidity.
suspensions may be held in the refrigerator at 3 to 10°C (37 to
50°F) for not more than fourteen days. NOTE 4—Covered dishes containing nutrient agar are considered to

8
DeVilbiss No. 163 atomizer or equivalent has been found satisfactory for this
purpose.
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have the desired humidity. Covers on large dishes may be sealed with much visual growth, hence some measure of change in physical property
masking tape. selected from those cited in the appendix is recommended.
9.2.2 Incubation Duration—The standard length of the test 9.4 Effect on Physical, Optical, or Electrical Properties—
is 28 days of incubation. The test may be terminated in less Wash the specimens free of growth, immerse in an aqueous
than 28 days for samples exhibiting a growth rating of two or solution of mercuric chloride (1 + 1000) for 5 min, rinse in tap
more. The final report must detail the actual duration of water, air dry overnight at room temperature, and recondition
incubation. at the standard laboratory conditions defined in Practice D618,
9.3 Observation for Visible Effects—If the test is for visible 23 6 1°C (73 6 2°F) and 50 6 5 % relative humidity, and test
effects only, remove the specimens from the incubator and rate according to the respective methods used on control specimens
them as follows: (see the appendix).
Observed Growth on Specimens NOTE 6—For certain electrical tests, such as insulation and arc
(Sporulating or Rating resistance, specimens may be tested in the unwashed, humidified condi-
Non-Sporulating, or Both) tion. Test values will be affected by surface growth and its associated
moisture.
None 0
Traces of growth (less than 10 %) 1 10. Report
Light growth (10 to 30 %) 2
Medium growth (30 to 60 %) 3 10.1 Report the following information:
Heavy growth (60 % to complete coverage) 4 10.1.1 Organisms used,
9.3.1 Specimens are rated after the fourth week. At Week 4, 10.1.2 Time of incubation,
a rating of trace or no growth (one or less) is confirmed with 10.1.3 Visual rating of fungus growth according to 9.3,
the stereoscope using oblique lighting and the magnification is including magnification for rating of 1 or less, and
recorded. Growth includes sporulating and non-sporulating 10.1.4 Table of progressive change in physical, optical, or
hyphae. Traces of growth may be defined as scattered, sparse electrical property against time of incubation. Give the rating
fungus growth such as might develop from a mass of spores in for each replicate.
the original inoculum, or extraneous contamination such as 11. Precision and Bias
fingermarks, insect feces, etc. Continuous cobwebby growth
11.1 A precision and bias statement cannot be made for this
extending over the entire specimen, even though not obscuring
practice at this time.
the specimen, should be rated as two. When non-test organisms
are present, include all growth of test and non-test organisms in 12. Keywords
the final rating. 12.1 fungal biosusceptibility; fungal decay; microbiological
NOTE 5—Considerable physical change in plastics may occur without assay; microbiological susceptibility

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 G21 − 15

APPENDIX

(Nonmandatory Information)

X1. TEST METHODS FOR EVALUATION OF EFFECT OF FUNGI ON SYNTHETIC POLYMERIC MATERIALS

X1.1 For evaluation of the effect of fungi on mechanical, TABLE X1.1 Recommended Test Methods
optical, and electrical properties, the following ASTM and Property Test Methods
other test methods are recommended. Tensile strength
D638, D882, D1708A
Stiffness D747A
TAPPI Test Method T 451-M-45A
Fed. Std. No. 191, Method 5204A
(Clark Stiffness Test)
Fed. Std. No. 191, Method 5206A
(Cantilever Bend Method) Hard-
ness
D785A
Optical transmission E308A
Haze D1003A
Water vapor transmission E96/E96MA
Dielectric strength D149A
Dielectric constant-power factor D150A
Insulation resistance D257A
Arc resistance D495A
A
These designations refer to the test methods given in Section 2.

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REFERENCES

(1) Bagdon, V. J., Military Specification Mil-P-43018(CE), “Plastic (5) Berk, S., Ebert, H., and Teitell, L., “Utilization of Plasticizers and
Sheets: Polyethylene Terephthalate, Drafting, Coated,” June 13, 1961. Related Organic Compounds by Fungi,” Industrial and Engineering
(2) ASTM Manual on Presentation of Data and Control Chart Analysis, Chemistry, Vol 49, No. 7, July 1957, pp. 1115–1124.
ASTM STP 15D, ASTM. (6) Brown, A. E., “Problem of Fungal Growth on Synthetic Resins,
(3) Baskin, A. D., and Kaplan, A. M., “Mildew Resistance of Vinyl- Plastics, and Plasticizers,” Modern Plastics, Vol 23, 1946, p. 189.
Coated Fabrics,” Applied Microbiology, Vol 4, No. 6, November (7) Ross, S. H., “Biocides for a Strippable Vinyl Plastic Barrier Material,”
1956. Report PB-151-119, U.S. Department of Commerce, Office of Tech-
(4) Berk, S., “Effect of Fungus Growth on Plasticized Polyvinyl Chloride nical Services.
Films,” ASTM Bulletin, No. 168, September 1950, p. 53 (TP 181).

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