1、Designation: D4783 01 (Reapproved 2013)Standard Test Methods forResistance of Adhesive Preparations in Container to Attackby Bacteria, Yeast, and Fungi1This standard is issued under the fixed designation D4783; the number immediately following the designation indicates the year oforiginal adoption o
2、r, in the 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.This standard has been approved for use by agencies of the Department of Defense.1. Scope1
3、.1 These test methods cover the determination of theresistance of liquid adhesive preparations to microbial attack inthe container by challenging adhesive specimens with culturesof bacteria, yeast, or fungi, and checking for their ability toreturn to sterility. These test methods return qualitative
4、results.1.2 The values stated in SI units are to be regarded as thestandard. The values in parentheses are for information only.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 standard to establish
5、appro-priate safety and health practices and determine the applica-bility of regulatory limitations prior to use. These test methodsare designed to be used by persons trained in correct micro-biological technique. Specific precautionary statements aregiven in Section 8.2. Referenced Documents2.1 AST
6、M Standards:2D907 Terminology of AdhesivesD4299 Test Method for Effect of Bacterial Contamination onPerformance of Adhesive Preparations and AdhesivesFilms (Withdrawn 1990)3D4300 Test Methods for Ability of Adhesive Films toSupport or Resist the Growth of FungiE640 Test Method for Preservatives in W
7、ater-ContainingCosmeticsNOTE 1Test Method E640 is under the jurisdiction of ASTMCommittee E35 on Pesticides. The procedure in this method outlines aserial dilution method of determining plate count using a pour platetechnique.2.2 TAPPI Method:T 487 Fungus Resistance of Paper and Paperboard42.3 CSMA:
8、Cosmetics Preservation, Method 3853. Terminology3.1 DefinitionsMany terms in these test methods aredefined in Terminology D907.3.2 Definitions of Terms Specific to This Standard:3.2.1 adhesive preparation, nthe adhesive as packaged fordistribution, storage, and use.3.3 Abbreviations:3.3.1 PBSphospha
9、te buffered saline.3.3.2 PDApotato dextrose agar.3.3.3 YMPGyeast malt peptone glucose (agar).4. Summary of Test Method4.1 The adhesive specimen is challenged by inoculationwith a culture of bacteria, yeast, or fungi, which may be asingle species or a mixed culture of several species, followingthe gu
10、idelines given in Note 6. The inoculated adhesivespecimen is stored at 21 to 27C (70 to 80F) for 7 days, duringwhich time cultures (streak plates) are made at preset intervals.See Note 2. At any point in the series of challenges, if theinoculated specimen shows microbial growth on the streakplates m
11、ade during the week following the challenge (indicat-ing that it has not returned to sterility), the test is discontinued,and the sample is reported as not resistant to attack in thecontainer by the species or combination of species used as theinoculum. If the cultures show no growth, the test is re
12、peatedwith up to four challenges. If the specimen tests out as sterilefollowing the fourth challenge, it is reported to be resistant toattack in the container by the species or combination of speciesof bacteria, fungi, or yeast used as the inoculum. At the1These test methods are under the jurisdicti
13、on of ASTM Committee D14 onAdhesives and are the direct responsibility of Subcommittee D14.30 on WoodAdhesives.Current edition approved Oct. 1, 2013. Published October 2013. Originallyapproved in 1988. Last previous edition approved in 2008 as D4783 01 (2008).DOI: 10.1520/D4783-01R13.2For referenced
14、 ASTM standards, visit the ASTM website, www.astm.org, orcontact ASTM Customer Service at serviceastm.org. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.3The last approved version of this historical standard is referenced onwww.a
15、stm.org.4Available from Technical Association of the Pulp and Paper Industry (TAPPI),15 Technology Parkway South, Norcross, GA 30092, http:/www.tappi.org.5This method is the same as Test Method E640.Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. U
16、nited States1discretion of the biological laboratory, the test may be discon-tinued after the second or third challenge. See Section 16 forfurther interpretation.4.2 The time necessary to kill is determined by noting theearliest streak plate to read sterile. If the 4-h plate is positiveand the 24-h
17、plate is negative, the kill time could be narroweddown further by repeating the challenge and making streakplates at intervals of 4, 8, 12, and 24 h following the challenge.4.3 The testing laboratory has the option of changing thetiming of the challenges, the sterility checks, and the incuba-tion pe
18、riod.NOTE 2Two proposed schedules for the challenging and sterilitychecks are shown in Table 1 and Table 2, Schedule A for bacteria andyeast, and Schedule B for fungi. The exact format to be followed will vary,according to the convenience of the schedule to the testing laboratory andspecial circumst
19、ances relating to the problem being addressed.NOTE 3A serial-dilution plate-count method of checking for sterilitymay be used when numerical information is needed on the population ofviable organisms or the reduction in population with increasing levels ofbiocide. Letheen broth is recommended for th
20、e diluent and Letheen agarfor the pour plate. See Note 1.5. Significance and Use5.1 These test methods are used to demonstrate whether anadhesive preparation is sufficiently protected with biocide toresist attack by bacteria, yeast, and fungi during its storage life.They are patterned after methods
21、used by biological laborato-ries serving the adhesive industry.5.2 These test methods may also be used to determine theefficacy of different biocide systems against specific microor-ganisms.5.3 These test methods are especially useful when testedagainst wild-type microorganisms which have been isola
22、tedfrom contaminated adhesives as an aid in determining theamount and type of biocide necessary to kill or inhibit thegrowth of the contaminants. If an isolated microorganism notgenerally used as a challenge organism, is chosen as theTABLE 1 Schedule AProposed Bacteria and Yeast Testing, Covering 4-
23、h, 24-h, 48-h, 72-h, and 7-Day TestsDay of Week Day no. First Challenge Second Challenge Third Challenge Fourth ChallengeMonday (1) inoculate fresh bacterialor yeast culture. . .Tuesday 0 prepare suspension . . .Tuesday 0 inoculate specimens . . .Tuesday (0 + 4 h) streak 4-h plate . . .Wednesday 1 s
24、treak 24-h plate . . .Thursday 2 streak 48-h plate . . .Friday 3 streak 72-h plate . . .Sat./Sun. 45 . . . .Monday 6 . inoculate fresh bacterialor yeast culture. .Tuesday 7 . prepare suspension . .Tuesday 7 streak 7-day plate inoculate specimens . .Tuesday (7 + 4 h) read 4-h plate streak 4-h plate .
25、 .Wednesday 8 read 24-h plate streak 24-h plate . .Thursday 9 read 48-h plate streak 48-h plate . .Friday 10 read 72-h plate streak 72-h plate . .Sat./Sun. 1112 . . . .Monday 13 . . inoculate fresh bacterialor yeast culture.Tuesday 14 . . prepare suspension .Tuesday 14 read 7-day plate streak 7-day
26、plate inoculate specimens .Tuesday (14 + 4 h) . read 4-h plate streak 4-h plate .Wednesday 15 . read 24-h plate streak 24-h plate .Thursday 16 . read 48-h plate streak 48-h plate .Friday 17 . read 72-h plate streak 72-h plate .Sat./Sun. 1819 . . . .Monday 20 . . . inoculate freshbacterial or yeastcu
27、ltureTuesday 21 . . . prepare suspensionTuesday 21 . read 7-day plate streak 7-day plate inoculate specimensTuesday (21 + 4 h) . . read 4-h plate streak 4-h plateWednesday 22 . . read 24-h plate streak 24-h plateThursday 23 . . read 48-h plate streak 48-h plateFriday 24 . . read 72-h plate streak 72
28、-h plateSat./Sun. 2526 . . . .Monday 27 . . . .Tuesday 28 . . read 7-day plate streak 7-day plateTuesday (28 + 4 h) . . . read 4-h plateWednesday 29 . . . read 24-h plateThursday 30 . . . read 48-h plateFriday 31 . . . read 72-h plateSat./Sun. 3233 . . . .Monday 34 . . . .Tuesday 35 . . . read 7-day
29、 plateD4783 01 (2013)2inoculum, it is important to identify the organism and deter-mine on which medium and under what conditions it will grow,in order to demonstrate the efficacy of the biocide.5.4 The results obtained when using the procedures given inthese methods apply only to the species which
30、are used for thetesting. The test species listed in Section 9 are frequently usedby laboratories to test for antimicrobial properties, but they arenot the only ones which could be used. Selection of the speciesto use for these test methods requires informed judgment by thetesting laboratory or by th
31、e party requesting the tests. It is alsoimportant that species which commonly attack adhesives beused. See 9.4.5.5 The presence of an active biocide carried over from theadhesive specimen to the agar could have an inhibiting effecton the growth of microorganisms, resulting in no growthduring the spa
32、n of a normal incubation period, when in fact,viable microorganisms are present, but their growth has beenslowed down or held in stasis. The use of Letheen agar andbroth is recommended to neutralize the effect of this carry-over.NOTE 4Letheen agar may be used for the streak plates, or if anotheragar
33、 is chosen for testing, a Letheen agar plate could be streaked as acontrol to test against the neutralizing effect. Even more effective wouldbe diluting the challenged adhesive specimen with Letheen broth andrunning Letheen agar pour plates. See Note 1 and Note 3. Extending theincubation period of n
34、egative plates would be another safeguard. Toneutralize thiazoline-based preservatives, 10 to 50 ppm of sodiumthioglycolate can be added to the medium.5.6 These test methods are dependent upon the physiologi-cal action of living microorganisms under a reported set ofconditions. Conclusions about the
35、 resistance of the test adhe-sive to microbiological attack can be drawn by comparing theresults to simultaneously run controls of known resistance. SeeX5.2 for statements regarding test repeatability.6. Apparatus6.1 In addition to the standard equipment found in any fullyequipped microbiological la
36、boratory, the following items aresometimes needed:6.1.1 Autoclave, capable of producing 103 kPa of steampressure at 121C (250F) and maintaining it for a minimum of15 min.TABLE 2 Schedule BProposed Fungi Testing, Covering 4-h, 24-h, 48-h, 72-h, and 7-Day TestsDay of Week Day no. First Challenge Secon
37、d Challenge Third Challenge Fourth ChallengeFriday (10) inoculate fresh fungalculture. . .Friday (3) . inoculate freshfungal culture. .Monday 0 prepare sporesuspension. . .Monday 0 inoculate specimens . . .Monday (0 + 4 h) streak 4-h plate . . .Tuesday 1 streak 24-h plate . . .Wednesday 2 streak 48-
38、h plate . . .Thursday 3 streak 72-h plate . . .Friday 4 . . inoculate fresh fungalculture.Sat./Sun. 5, 6 . . . .Monday 7 . prepare sporesuspension. .Monday 7 streak 7-day plate inoculate specimens . .Monday (7 + 4 h) read 4-h plate streak 4-h plate . .Tuesday 8 read 24-h plate streak 24-h plate . .W
39、ednesday 9 read 48-h plate streak 48-h plate . .Thursday 10 read 72-h plate streak 72-h plate . inoculate freshfungal cultureFri./Sat./Sun. 11, 12, 13 . . . .Monday 14 . . prepare sporesuspension.Monday 14 read 7-day plate streak 7-day plate inoculate specimens .Monday (14 + 4 h) . read 4-h plate st
40、reak 4-h plate .Tuesday 15 . read 24-h plate streak 24-h plate .Wednesday 16 . read 48-h plate streak 48-h plate .Thursday 17 . read 72-h plate streak 72-h plate .Fri./Sat./Sun. 18, 19, 20 . . . .Monday 21 . . . prepare sporesuspensionMonday 21 . read 7-day plate streak 7-day plate inoculate specime
41、nsMonday (21 + 4 h) . . read 4-h plate streak 4-h plateTuesday 22 . . read 24-h plate streak 24-h plateWednesday 23 . . read 48-h plate streak 48-h plateThursday 24 . . read 72-h plate streak 72-h plateFri./Sat./Sun. 25, 26, 27 . . . .Monday 28 . . read 7-day plate streak 7-day plateMonday (28 + 4 h
42、) . . . read 4-h plateTuesday 29 . . . read 24-h plateWednesday 30 . . . read 48-h plateThursday 31 . . . read 72-h plateFri./Sat./Sun. 32, 33, 34 . . . .Monday 35 . . . read 7-day plateD4783 01 (2013)36.1.2 Cell Counting Chamber, Petroff-Hausser, cell depth0.02 mm (or equivalent).66.1.3 Bottles, Sc
43、rewcap, approximately 375 mL, BostonRounds of flint glass. Mold-A-7232-D, Finish 28-400, andBlack Artmold Caps BM-8041, Size 28-400, with rubber ringliners fastened to caps with steamproof adhesive.76.1.4 Constant Temperature Chamber, capable of beingmaintained at 35 6 0.5C (95 6 1F) for bacteria, o
44、r 30 60.5C (86 6 1F) for fungi, or two chambers if neededsimultaneously.6.1.5 Glass Rods, 305 mm in length having a diameter of6.3 mm.6.1.6 Hemacytometer, Levy Counting Chamber, cell depth0.1 mm, Newbauer rulings.66.1.7 Hood, laminar flow type.86.1.8 Jar, Screw Cap, round, approximately 1 L (1-qt ma
45、sontype) for samples.6.1.9 Pasteur Pipets.66.1.10 Pipet, 1 mL, disposable, sterile.6.1.11 Pipettes, automatic Oxford or Eppindorf, sterile, withsterile tips.66.1.12 Refrigerator, capable of maintaining temperature of 46 1C (39 6 2F).6.1.13 Spectrophotometer, capable of measuring cell countat a wavel
46、ength of 425 nm.67. Materials7.1 Beef Extract.7.2 Deionized or Distilled Water, sterile.7.3 Disinfectant Solution, Amphyll, Alcide, or equivalent.67.4 Glucose.7.5 Letheen Agar, (Difco or equivalent).67.6 Letheen Broth, (Difco or equivalent).67.7 Mycophil Agar, pH 4.7 (BBL or equivalent).67.8 Nutrien
47、t Agar, (BBL or equivalent).67.9 Potato Dextrose Agar (PDA, Dehydrated), (Difco orequivalent).67.10 Phosphate buffered saline (PBS), sterile.7.11 Physiological Saline Solution, 0.85 % NaCl, sterile.7.12 Sorbitan mono-oleate polyoxyethylene.97.13 Tryptone.7.14 Tryptone Glucose Extract Agar, dehydrate
48、d (Difco orequivalent).7.15 Yeast Malt Peptone Glucose Agar (YMPGA), dehy-drated (Difco or equivalent).8. Precautions8.1 These test methods employ live cultures of bacteria,fungi, and yeast, some of which are capable of causing disease,and others allergic reaction in some humans. In addition toother
49、 precautions, assign laboratory personnel trained in cor-rect microbiological techniques to run these tests. Use propermicrobiological procedures in order to prevent contaminationof the cultures or of the work area. Clean and sterilize in anapproved manner all spills and all equipment coming intocontact with the cultures and the inoculated adhesive speci-mens. Also sterilize in an approved manner all cultures andcontaminated disposable equipment before discarding. See 1.3.9. Test Organisms109.1 Cultures of one or more of the following bacterialspecies
