1、Designation: E 1428 99 (Reapproved 2004)Standard Test Method forEvaluating the Performance of Antimicrobials in or onPolymeric Solids Against Staining by Streptoverticilliumreticulum (A Pink Stain Organism)1This standard is issued under the fixed designation E 1428; the number immediately following
2、the designation indicates the year oforiginal adoption or, in 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 is u
3、sed for determining the perfor-mance of antimicrobial agents used in or on synthetic poly-meric solids against staining by the actinomycete Streptover-ticillium reticulum. This organism has been chosen as anindicator organism, although other organisms have been knownto cause undesirable staining in
4、polymeric solids.1.2 This test method is not suitable for evaluating dark-pigmented test samples.1.3 A knowledge of microbiological techniques is recom-mended for these procedures.1.4 This standard does not purport to address all of thesafety concerns, if any, associated with its use. It is therespo
5、nsibility of the user of this standard to establish appro-priate safety and health practices and determine the applica-bility of regulatory limitations prior to use.2. Summary of Test Method2.1 Test specimens are placed on an agar surface inoculatedwith Streptoverticillium reticulum and incubated. A
6、fter incuba-tion, test specimens are rated visually by percentage of samplearea stained.3. Significance and Use3.1 This test method provides a technique for evaluatingantimicrobials in or on polymeric solids against staining byStreptoverticillium reticulum, and should assist in the predic-tion of pe
7、rformance of treated articles under actual fieldconditions.3.2 Conditioning of the specimens, such as exposure toleaching, weathering, and heat treatment, may have significanteffects on performance of antimicrobials against staining.Determination of these effects is not included in this testmethod.4
8、. Interferences4.1 An interference may be caused by contamination ofplates and agar by unwanted organisms that settle from theenvironment.4.2 Dark pigments mask observation of the pink stain.5. Apparatus5.1 Petri dishes, 100 mm diameter.25.2 Cotton swabs, sterile.5.3 IncubatorIncubating equipment fo
9、r this test methodshall maintain a temperature of 29 6 1C.5.4 Autoclave.5.5 Sterilizer, ethylene oxide (optional).6. Reagents and Materials6.1 Yeast Malt Extract Agar (ISP Medium 2)3Prepare thismedium according to manufacturers directions.6.2 InoculumStreptoverticillium reticulum ATCC25607.4Maintain
10、 stock cultures on yeast malt extract agar. Thestock may be kept for not more than 12 months at approxi-mately 3 to 10C. Subcultures, incubated at 29 6 1C for 7 to14 days, shall be used for inoculation.7. Test Specimens7.1 From each test unit,5cut duplicate 0.75 in. diameterdiscs. If the test unit i
11、s of different construction on each side,two specimens of each, two face up and two face down, shallbe tested.7.2 A test unit containing no biocide should be included asa positive stain control.1This test method is under the jurisdiction of ASTM Committee E35 onPesticides and Alternative Control Age
12、nts and is the direct responsibility ofSubcommittee E35.15 on Antimicrobial Agents.Current edition approved Oct, 1, 2004. Published October 2004. Originallyapproved in 1991. Last previous edition approved in 1999 as E 1428 99.2Presterilized and disposable plastic petri dishes are available from most
13、laboratory supply houses.3The sole source of supply of Bacto-Yeast Malt ExtractAgar, Stock No. 0770-01known to the committee at this time is Difco Labs, P.O. Box 1058A, Detroit, MI28232. If you are aware of alternative suppliers, please provide this information toASTM International Headquarters. You
14、r comments will receive careful consider-ation at a meeting of the responsible technical committee1, which you may attend.4American Type Culture Collection, 10801 University Blvd., Manassas, VA20110.5Atest unit is a solid in the form of plastic sheets, films, coated fabrics or similarpolymeric mater
15、ials.1Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959, United States.7.3 A test unit known to inhibit staining by Streptoverticil-lium reticulum should be included as a negative stainingcontrol.7.4 Heavily-soiled specimens should be rinsed vigorousl
16、y inwater.7.5 If sterilization of test specimens is considered necessary,ethylene oxide is the sterilant of choice. Ethylene oxide residuemay affect test outcome. Proper aeration after sterilization isnecessary.8. Procedure8.1 InoculationPour sufficient yeast malt extract agar intosuitable sterile d
17、ishes (see 5.1) to provide a solidified agar layerfrom 5 to 8 mm in depth. After the agar has solidified, moistenthe agar surface by streaking with a cotton swab dipped insterile water.8.2 Add 3 mL of sterile saline or phosphate buffered salineto the stock culture plate and use the premoistened swab
18、 toharvest the Streptoverticillium cells.8.3 Streak the cell-laden swab on the agar surface bystreaking in a manner which will ensure total coverage by theorganism. This swabbing technique typically provides suffi-cient growth to yield measurable pink stain on positive staincontrols.8.4 Use sterile
19、forceps to place the sterile test specimens,two to a plate, on the agar surface. There shall be good contactestablished between the test specimen and the inoculated agarsurface. Should the specimens tend to curl, weights (such as58in. sterile stainless steel nuts) may be placed on top ofspecimens to
20、 maintain good contact.8.5 IncubationCover the inoculated test plates containingthe specimens and incubate at 29 6 1C for 14 days.NOTE 1Covered petri dishes containing yeast malt extract agar areconsidered to have the desired humidity.9. Interpretation of Results9.1 Observe the degree of staining on
21、 the samples bycomparing with the inoculated, untreated control.9.2 If staining is not seen on the surface of the specimen, usesterile forceps to lift and turn the specimen over and inspect theunderside for stain. The degree of staining is determined by theamount of sample surface stained rather tha
22、n the intensity ofthe color. Any possible sample discoloration that is not pink incolor should be disregarded when determining the degree ofstain. However, pigmented samples that show a color changeassociated with pink (that is, a blue pigmented sample chang-ing to purple) should be rated according
23、to degree of stain. Ratethe degree of stain in accordance with Table 1.9.3 Leachability of AntimicrobialsA“ zone of inhibition”may be observed where no organisms grow on the agaradjacent to the specimen. This indicates that the antimicrobialmay be leaching from the specimen.10. Report10.1 Report the
24、 visual rating of stain in accordance with 9.1.11. Precision and Bias11.1 A precision and bias statement cannot be made for thistest method at this time.12. Keywords12.1 antimicrobial efficacy; pink stain; plastics; polymericsolids;Streptoverticillium reticulumASTM International takes no position re
25、specting the validity of any patent rights asserted in connection with any item mentionedin this standard. Users of this standard are expressly advised that determination of the validity of any such patent rights, and the riskof infringement of such rights, are entirely their own responsibility.This
26、 standard is subject to revision at any time by the responsible technical committee and must be reviewed every five years andif not revised, either reapproved or withdrawn. Your comments are invited either for revision of this standard or for additional standardsand should be addressed to ASTM Inter
27、national Headquarters. Your comments will receive careful consideration at a meeting of theresponsible technical committee, which you may attend. If you feel that your comments have not received a fair hearing you shouldmake your views known to the ASTM Committee on Standards, at the address shown b
28、elow.This standard is copyrighted by ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959,United States. Individual reprints (single or multiple copies) of this standard may be obtained by contacting ASTM at the aboveaddress or at 610-832-9585 (phone), 610-832-955
29、5 (fax), or serviceastm.org (e-mail); or through the ASTM website(www.astm.org).TABLE 1 Degree of Stain RatingObserved Stain on Specimens RatingNo Stain 0Trace of Stain (less than 10 % coverage) 1Slight Stain (10 to 30 % coverage) 2Moderate Stain (30 to 50 % coverage) 3Heavy Stain (50 % to complete coverage) 4E 1428 99 (2004)2
