1、Designation: D 5589 97 (Reapproved 2002)Standard Test Method forDetermining the Resistance of Paint Films and RelatedCoatings to Algal Defacement1This standard is issued under the fixed designation D 5589; the number immediately following the designation indicates the year oforiginal adoption or, in
2、 the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon (e) indicates an editorial change since the last revision or reapproval.1. Scope1.1 This test method covers an accelerated method fordetermining the relative resistan
3、ce 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
4、 affect 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 sta
5、ndard 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:D 822 Practice for Filtered Open-Flame Carbon-Arc Expo-sures of Paint and Related Coatings2D 4141 Practice for Conducting Blac
6、k Box and Solar Con-centrating Exposures of Coatings2D 4587 Practice for Fluorescent UV-Condensation Expo-sures of Paint and Related Coatings2D 5031 Practice for Enclosed Carbon-Arc Exposure Testsof Paint and Related Coatings2G 53 Practice for Operating Light- and Water-ExposureApparatus (Fluorescen
7、t UV-Condensation Type) for Expo-sure of Nonmetallic Materials33. Summary of Test 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 appropriat
8、e conditions for growth, and (4)provide a schedule and guidelines for visual growth 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
9、paint and coating films by algal growth isa common phenomenon under certain conditions. 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
10、method attempts to provide a means tocomparatively evaluate different coating formulations 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
11、.2 Familiarity with microbiological techniques is required.This test method should 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 be
12、tween 25 6 2C, relative humidity of$85 %, and having a constant fluorescent light 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
13、 cm (1.65 in.) in diameter, or drawdown paper(unlaquered chart paper) 21.6 by 28.0 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.5.10 Sterile Glass Rods, Forceps, 250-mL Glass ErlenmeyerFlask, and other ro
14、utine microbiological equipment.1This test method is under the jurisdiction of ASTM Committee D01 on Paintand Related Coatings, Materials, and Applications and is the direct responsibility ofSubcommittee D01.28 Biodeterioration.Current edition approved July 10, 1997. Published September 1997. Origin
15、allypublished as D 5589 94. Last previous edition D 5589 94.2Annual Book of ASTM Standards, Vol 06.01.3Annual Book of ASTM Standards, Vol 14.02.1Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959, United States.5.11 Allens Medium4or Bolds Basal Medium5
16、ingredients(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 thatall reagents should conform to the specifications of theCommittee on Analytical Reagents of the American Chemica
17、lSociety, where such specifications are available.6Other gradesmay be used, provided they are first ascertained to be ofsufficiently high purity to permit use without decreasing theaccuracy of the determination.6.2 Purity of WaterUnless otherwise indicated, referencesto water are understood to mean
18、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/1000 mLNaNO31.5K2HPO40.039MgSO47H2O 0.075CaCl22H2O 0.027Na2CO30.020Na2SiO39H2O 0.058Citric acid 0.006EDTAA0.006A
19、llens trace element solution 1.0 mLBDistilled water to 1000 mLFerric citrate (see Note 2) 0.006 (see Note 2)AEthlenediaminetetraacetate.BAllens Trace-Element Solution:Dissolve in 500 mL of distilled water:Reagent Amount, gH3BO32.86MnCl24H2O 1.81ZnSO47H2O 0.222Na2MoO42H2O 0.391CuSO45H2O 0.079Co(NO3)2
20、6H2O 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 of 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
21、 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 adding the ferric citrate. After mixing,pour the media into petri dishes.6.4 Bolds Basal MediumPrepare ten individual stocksoluti
22、ons in distilled water as indicated:Stock Solutions g/400 mL1. NaNo310.02. MgSO47H2O 3.03. NaCl 1.04. K2HPO43.05. KH2PO47.06. CaCl22H2O 1.0Trace Element Solutions: g/L7. ZnSO47H2O 8.82MnCl24H2O 1.44MoO30.71CuSO45H2O 1.57Co(NO3)26H2O 0.49Distilled Water to 1 LAutoclave to dissolve.8. H3BO311.429. EDT
23、AKOH 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 in 936 mLdistilled water. Autoclave at 121C.6.5 A variety of algal cultures, including wild strains iso-lated from paint fil
24、ms, 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 is recommended to choose at least one culture fromeach type. The choice of strains should be agreed upon betweenthe parties
25、involved in the testing.Algae Collection/StrainAUnicellular GreenChlorella sp. ATCC 7516Chlorella vulgaris ATCC 11468Filamentous GreenUlothrix gigas ATCC 30443Trentepohlia aurea UTEX 429Trentepohlia odorata CCAP 483/4Colony-forming GreenScenedesmus quadricauda ATCC 11460Filamentous BluegreenOscillat
26、oria sp. ATCC 29135Calothrix sp. ATCC 27914AAvailable from the following culture collections and found suitable for this test:American Type Culture Collection (ATCC), 12301 Parklawn Drive, Rockville, MD20852; University of Texas (UTEX), Department of Botany, The University of Texasat Austin, Austin,
27、 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 purchased cultures in media and underincubation conditions recommended by culture collection.6.6 Cultures should be maintained
28、 separately in liquidmedia recommended by the culture supplier. Allens Medium(6.3) is commonly used for bluegreen and other algae. Therecipe for Bolds Basal Medium, which supports the growth ofa wide range of algae is given in 6.4. If preferred, individual4Bold, H. C., Wynne, M. J., “Introduction to
29、 the Algae,” Prentiss-Hall,Englewood Cliffs, NJ, 1978, pp. 5745.5Kirsop B. E., and Snell J. J. S., “Maintenance of Microorganisms,” AcademicPress, Harcourt Brace Jovanovich, Orlando, FL, 1984, p. 158.6Reagent 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.D 5589 97 (2002)2
31、cultures 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 tissue grinder to homogenize filamentous algae beforepreparing inoculum. Adjust each culture to approximately onemillion c
32、ells 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 petri dishculture by pouring 10 mL of distilled water on the agar surface.Gently scrape the algae with a sterile glass rod or
33、 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 of each species, andabout the same total amount between runs of this test methodto make correlation of data between test run
34、s 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. Alterna-tively, each sample could be inoculated separately with individual culturesas agreed upon between the parties involved.7
35、. 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 satisfactorily in this test method should alsobe included. A set of paper filter disks or the draw-down paperswithout coating may be s
36、uitable 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,avoids bacterial or other contamination that may interfere with the testresults.7.3 Coatings to be tested will be applied to
37、 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 thickness forall coating samples.NOTE 5One or both sides of the substrate (drawdown strips or filterpaper) may be coat
38、ed 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 stripfrom a drying rack with string or a twist tie. The label for each strip canbe written in the top 12.7 mm (12 in.) o
39、f 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 whilebrushing. This would still leave sufficient coated area for six 28 by 28-mm(1.1 by 1.1-in.) test squares from each strip.
40、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 72 h atroom temperature.7.5 If accelerated weathering, heat aging, or other precon-ditioning of samples is also to be r
41、un, prepare a separate set ofduplicate sample disks or strips. The results from these samplesmay be compared with those from the unweathered or uncon-ditioned samples.NOTE 7There are a variety of methods that could be used to simulateaccelerated effects of weathering (sunlight or rain, or both), on
42、thesamples. For example, a leach test that is frequently used to simulate theeffects of rainwater (an important factor for algae growth) is outlined inNote 8. Conditioning of specimens by artificial weathering may be doneaccording to one of the following practices: D 822, D 4141, D 4587, orD 5031.NO
43、TE 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 distilled water with a flow rate such that there are sixchanges in 24 h (
44、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 7.4.7.6 If the drawdown strips are being used, cut them intorough
45、ly 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 oneweek. If the plates were stored in the refrigerator, allow themto e
46、quilibrate 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, firstcut into 28-mm (1.1-in.) squares.8.1.2 Transfer the mixed alg
47、al 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 each speci-men, making sure the surface is covered, but not oversaturatingthe samples. Also, be certain the amount of inoculum appliedis the same between the variou
48、s samples under test (this shouldbe done by the same applicator at the same time for allsamples).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
49、 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 will appearas the typical green algae-like discoloration of the coating.Other species may show different colors.9. Evaluation of Results9.1 Rate the growth on the specimen weekly for three weeksaccording to the following:Observed Growth on Specimens RatingNone 0Traces of growth (10 %) 1Light growth (1030 %) 2Moderate growth (3060 %) 3Heavy growth (60 % to compl
