1、Designation: D5589 09 (Reapproved 2013)Standard Test Method forDetermining the Resistance of Paint Films and RelatedCoatings to Algal Defacement1This standard is issued under the fixed designation D5589; the number immediately following the designation indicates the year oforiginal adoption or, in t
2、he case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon () indicates an editorial change since the last revision or reapproval.1. Scope1.1 This test method covers an accelerated method fordetermining the relative resistance
3、of a paint or coating film toalgal growth.NOTE 1It is hoped that a ranking of relative performance would besimilar to that ranked from outdoor exposures. However, this test methodshould not be used as a replacement for exterior exposure since manyother factors, only a few of which are listed will af
4、fect those results.1.2 The values stated in SI units are to be regarded as thestandard. The values given in parentheses are for informationonly.1.3 This standard does not purport to address all of thesafety concerns, if any, associated with its use. It is theresponsibility of the user of this standa
5、rd to establish appro-priate safety and health practices and determine the applica-bility of regulatory limitations prior to use.2. Referenced Documents2.1 ASTM Standards:2D822 Practice for Filtered Open-Flame Carbon-Arc Expo-sures of Paint and Related CoatingsD4141 Practice for Conducting Black Box
6、 and Solar Con-centrating Exposures of CoatingsD4587 Practice for Fluorescent UV-Condensation Expo-sures of Paint and Related CoatingsD5031 Practice for Enclosed Carbon-Arc Exposure Tests ofPaint and Related CoatingsD6695 Practice for Xenon-Arc Exposures of Paint andRelated Coatings3. Summary of Tes
7、t Method3.1 This test method outlines a procedure to (1) prepare asuitable specimen for testing, (2) inoculate the specimen witha mixture of the proper algal species, (3) expose the inoculatedsamples under the appropriate conditions for growth, and (4)provide a schedule and guidelines for visual gro
8、wth ratings.This test method is not designed to include all the necessaryprocedures to maintain the proper microbiological techniquesrequired to provide the most accurate results.4. Significance and Use4.1 Defacement of paint and coating films by algal growth isa common phenomenon under certain cond
9、itions. It is gener-ally known that differences in the environment, lighting,temperature, substrate, and other factors in addition to thecoating composition affect the susceptibility of a given paintedsurface. This test method attempts to provide a means tocomparatively evaluate different coating fo
10、rmulations for theirrelative performance under a given set of conditions. It doesnot imply that a coating that resists growth under theseconditions will necessarily resist growth in the actual applica-tion (see Note 1).4.2 Familiarity with microbiological techniques is required.This test method shou
11、ld not be used by persons without at leastbasic microbiological training.5. Apparatus and Materials5.1 Balance, capable of weighing to 0.10 g.5.2 Incubator, or other device capable of maintaining aconstant temperature between 25 6 2C, relative humidity of85 %, and having a constant fluorescent light
12、 source.5.3 Refrigerator.5.4 Petri Dishes, 100 by 15 mm (3.9 by 0.6 in.).5.5 Autoclave.5.6 Paint Brush, coarse bristle, 12 to 19 mm (12 to34 in.).5.7 Test Substrate, filter paper, either regular paper or glassfiber, 4.2 cm (1.65 in.) in diameter, or drawdown paper(unlaquered chart paper) 21.6 by 28.
13、0 cm (8.5 by 11 in.), cutinto ten 21.6 by 2.8-cm (8.5 by 1.1-in.) strips may be used.5.8 Tissue Grinder.5.9 Atomizer or Chromatography Sprayer.1This test method is under the jurisdiction of ASTM Committee D01 on Paintand Related Coatings, Materials, and Applications and is the direct responsibility
14、ofSubcommittee D01.28 on Biodeterioration.Current edition approved Oct. 1, 2013. Published October 2013. Originallyapproved in 1994. Last previous edition approved in 2009 as D5589 09. DOI:10.1520/D5589-09R13.2For referenced ASTM standards, visit the ASTM website, www.astm.org, orcontact ASTM Custom
15、er Service at serviceastm.org. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States15.10 Sterile Glass Rods, Forceps, 250-
16、mL Glass ErlenmeyerFlask, and other routine microbiological equipment.5.11 Allens Medium3or Bolds Basal Medium4ingredients(see 6.3).5.12 Distilled Water.6. Reagents and Materials6.1 Purity of ReagentsReagent grade chemicals should beused in all tests. Unless otherwise indicated, it is intended thata
17、ll reagents should conform to the specifications of theCommittee on Analytical Reagents of the American ChemicalSociety, where such specifications are available.5Other gradesmay be used, provided they are first ascertained to be ofsufficiently high purity to permit use without decreasing theaccuracy
18、 of the determination.6.2 Purity of WaterUnless otherwise indicated, referencesto water are understood to mean distilled water or water ofequal or higher purity.6.3 Allens MediumPrepare liquid medium by dissolvingin 1000 mL of water the following reagents in the designatedamounts:Reagent Amount, g/1
19、000 mLNaNO31.5K2HPO40.039MgSO47H2O 0.075CaCl22H2O 0.027Na2CO30.020Na2SiO39H2O 0.058Citric acid 0.006EDTAA0.006Allens trace element solution 1.0 mLBDistilled water to 1000 mFerric citrate (see Note 2) 0.006 (see Note 2)AEthylenediaminetetraacetic acid, disodium saltBAllens Trace-Element Solution:Diss
20、olve in 500 mL of distilled water:Reagent Amount, gH3BO32.86MnCl24H2O 1.81ZnSO47H2O 0.222Na2MoO42H2O 0.391CuSO45H2O 0.079Co(NO3)26H2O 0.0494NOTE 2The ferric citrate must be autoclaved separately. The ferriccitrate should be added after the medium has cooled from beingautoclaved.6.3.1 Adjust the pH o
21、f the medium to 7.8 using 1.0 MNaOH/1.0 M HCl and autoclave at 121C (without ferriccitrate added) to 45 to 50C before aseptically adding the ferriccitrate (see Note 2).6.3.2 Allens AgarPrepare by dissolving 15 g of agar in1000 mL Allens Medium before autoclaving. Cool to 45 to50C before aseptically
22、adding the ferric citrate. After mixing,pour the media into petri dishes.6.4 Bolds Basal MediumPrepare ten individual stocksolutions in distilled water as indicated:Stock Solutions g/400 mL1. NaNo310.02. MgSO47H2O3.03. NaCl 1.04. K2HPO43.05. KH2PO47.06. CaCl22H2O1Trace Element Solutions: g/L7. ZnSO4
23、7H2O 8.82MnCl24H2O 1.44MoO30.71CuSO45H2O 1.57Co(NO3)26H2O 0.49Distilled Water to 1 LAutoclave to dissolve.8. H3BO311.429. EDTAKOH solution:EDTA 50.0KOH 31.010. FeSO47H2O 4.98H2SO4(concentrate) 1.0 mL6.4.1 Combine 10 mL each of Stock Solutions 1 through 6with 1 mL each of Stock Solutions 7 through 10
24、 in 936 mLdistilled water. Autoclave at 121C.6.5 A variety of algal cultures, including wild strains iso-lated from paint films, may be used in this protocol. Choosestrains from the following list, use field isolates or use otherstrains found to grow satisfactorily under the protocol condi-tions. It
25、 is recommended to choose at least one culture fromeach type. The choice of strains should be agreed upon betweenthe parties involved in the testing.Algae Collection/StrainAUnicellular GreenChlorella sp. ATCC 7516Chlorella vulgaris ATCC 11468Filamentous GreenUlothrix gigas ATCC 30443Trentepohlia aur
26、ea UTEX 429Trentepohlia odorata CCAP 483/4Colony-forming GreenScenedesmus quadricauda ATCC 11460Filamentous BluegreenOscillatoria sp. ATCC 29135Calothrix sp. ATCC 27914AAvailable from the following culture collections and found suitable for this test:American Type Culture Collection (ATCC), 12301 Pa
27、rklawn Drive, Rockville, MD20852; University of Texas (UTEX), Department of Botany, The University of Texasat Austin, Austin, TX 78713-7640; Culture Collection of Algae and Protozoa(CCAP), Institute of Freshwater Ecology,The Windermere Laboratory, Far Sawrey,Ambleside, Cumbria LA22 OLP, U.K. Grow pu
28、rchased cultures in media and underincubation conditions recommended by culture collection.6.6 Cultures should be maintained separately in liquidmedia recommended by the culture supplier. Allens Medium(6.3) is commonly used for bluegreen and other algae. The3Bold, H. C., Wynne, M. J., “Introduction
29、to the Algae,” Prentiss-Hall,Englewood Cliffs, NJ, 1978, pp. 5745.4Kirsop B. E., and Snell J. J. S., “Maintenance of Microorganisms,” AcademicPress, Harcourt Brace Jovanovich, Orlando, FL, 1984, p. 158.5Reagent Chemicals, American Chemical Society Specifications, AmericanChemical Society, Washington
30、, DC. For suggestions on the testing of reagents notlisted by the American Chemical Society, see Analar Standards for LaboratoryChemicals, BDH Ltd., Poole, Dorset, U.K., and the United States Pharmacopeiaand National Formulary, U.S. Pharmacopeial Convention, Inc. (USPC), Rockville,MD.D5589 09 (2013)
31、2recipe for Bolds Basal Medium, which supports the growth ofa wide range of algae is given in 6.4. If preferred, individualcultures may be maintained on solid media prepared bydissolving 1 to 1.5 % agar in liquid medium before autoclav-ing.6.6.1 Cultures should be actively growing prior to use. Usea
32、 tissue grinder to homogenize filamentous algae beforepreparing inoculum. Adjust each culture to approximately onemillion cells per millilitre in sterile water or to a light greencolor. Combine equal volumes of individual cultures for amixed inoculum.6.6.2 If preferred, harvest algae from an agar pe
33、tri dishculture by pouring 10 mL of distilled water on the agar surface.Gently scrape the algae with a sterile glass rod or pipet. Pipetthe suspension into a sterile 250-mL glass Erlenmeyer flask.Repeat for all the cultures by pipetting into the same flask (tryto obtain approximately equal amounts o
34、f each species, andabout the same total amount between runs of this test methodto make correlation of data between test runs easier). Bring themixed volume of suspension up to 100 mL with sterile water.Retain for later use as inoculum in 8.1.NOTE 3The previous procedure gives a mixed inoculum.Altern
35、atively, each sample could be inoculated separately with individualcultures as agreed upon between the parties involved.7. Preparation of Test Specimens7.1 A set of coatings to be tested should contain a controlpaint without algicide (blank). If available, a formulationknown to perform satisfactoril
36、y in this test method should alsobe included.Aset of paper filter disks or the draw-down paperswithout coating may be suitable growth controls (see 5.7).7.2 Handle the disks or drawdown sections with steriletongs or tweezers.NOTE 4Sterilization or aseptic handling of the test material, or both,avoid
37、s bacterial or other contamination that may interfere with the testresults.7.3 Coatings to be tested will be applied to the chosen testsubstrate (5.7) by brush coating the strips of drawdownpaperboard or filter disks with each sample in duplicate. Takecare to apply a thin, even coating with the same
38、 thickness forall coating samples.NOTE 5One or both sides of the substrate (drawdown strips or filterpaper) may be coated as agreed upon between the parties involved.NOTE 6With the drawdown strips, this can be conveniently accom-plished by punching a hole in the top of the strip and suspending the s
39、tripfrom a drying rack with string or a twist tie. The label for each strip canbe written in the top 12.7 mm (12 in.) of the strip (near the hole) and thecoating applied below that 12.7-mm strip. Another 12.7-mm area can beleft uncoated at the bottom of the strip to permit holding the strip whilebru
40、shing. This would still leave sufficient coated area for six 28 by 28-mm(1.1 by 1.1-in.) test squares from each strip. With the filter disks, a holecan be punched near the edge of the disk.7.4 After application, suspend the sample disks or stripsfrom drying racks and allow them to air dry for 24 to
41、72 h atroom temperature.7.5 When accelerated weathering, heat aging, or otherpreconditioning of samples is to be done, a minimum of 2additional test pieces must be prepared from each coating foreach type of preconditioning used. The results from thepreconditioned samples may be compared with those f
42、rom theunconditioned samples.NOTE 7There are a variety of methods that can be used to simulateaccelerated effects of weathering (sunlight or rain, or both), on thesamples. For example, a leach test that is frequently used to simulate theeffects of rainwater (an important factor for algae growth) is
43、outlined inNote 8. Conditioning of specimens by artificial weathering can be doneaccording to one of the following practices: D822, D4141, D4587, D5031,or D6695. These practices use very different test conditions and may beexpected to produce different test results. Therefor, they cannot be usedinte
44、rchangeably unless equivalency of test results is demonstrated.NOTE 8A leaching test may be conducted as follows. One of thereplicate sets is leached with distilled water for 24 h, then allowed to airdry. The coated substrate can be leached by suspension for 24 h in 4-L(1-gal) containers of distille
45、d water with a flow rate such that there are sixchanges in 24 h (or other flow rate as agreed upon between the partiesinvolved). Note differences in the integrity of the coatings after thisleaching. The test panels are then air dried for 24 h under the sameconditions as the unleached samples, as in
46、7.4.7.6 If the drawdown strips are being used, cut them intoroughly 28-mm (1.1-in.) squares. Place these specimensquares, or the coated filter disks, on the center of pre-pouredAllens (or appropriatesee 6.6) agar plates. The plates shouldbe prepared at least 24 h in advance, but no longer than onewe
47、ek. If the plates were stored in the refrigerator, allow themto equilibrate to room temperature prior to placement of thesamples.8. Procedure8.1 Inoculation of the Test Specimens:8.1.1 Place test specimens in the center of solidified Allens(or appropriate) Agar plates. If drawdown strips are used, f
48、irstcut into 28-mm (1.1-in.) squares.8.1.2 Transfer the mixed algal inoculum from the flask(from 6.6.2) into a sterile atomizer or chromatography sprayer.8.1.3 Apply a thin coat of algae suspension to eachspecimen, making sure the surface is covered, but not over-saturating the samples.Also, be cert
49、ain the amount of inoculumapplied is the same between the various samples under test(this should be done by the same applicator at the same time forall samples).8.1.4 Transfer the inoculated plates to an incubator with aconstant fluorescent light source, humidity 85 %, and atemperature setting to maintain 25 6 2C.NOTE 9If the capability is available, a cycle of 14-h light and 10-hdarkness can improve the growth of the algae.8.1.5 Incubate the samples under the specified conditionsjust stated and examine weekly for growth. Growth
