1、Designation: E 723 07Standard Test Method forEfficacy of Antimicrobials as Preservatives for Aqueous-Based Products Used in the Paper Industry (BacterialSpoilage)1This standard is issued under the fixed designation E 723; the number immediately following the designation indicates the year oforiginal
2、 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 laboratory test method is used to determine theefficacy of an
3、 antimicrobial for preventing bacterial spoilage ofin-process aqueous-based products used in the paper industry.For information on fungal spoilage, see Test Method E 875.This test method should be performed by persons who havehad basic microbiological training.1.2 This standard does not purport to a
4、ddress all of thesafety concerns, if any, associated with its use. It is theresponsibility 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. Referenced Documents2.1 ASTM Standards:2D 1193 Spec
5、ification for Reagent WaterE 875 Test Method for Efficacy of Fungal ControlAgents asPreservatives for Aqueous-Based Products Used in thePaper IndustryE 1054 Test Methods for Evaluation of Inactivators ofAntimicrobial AgentsE 1326 Guide for Evaluating Nonconventional Microbio-logical Tests Used for E
6、numerating Bacteria3. Terminology3.1 Definitions of Terms Specific to This Standard:3.1.1 antimicrobial, nchemical or physical agent that killsor inactivates microorganisms or suppresses their growth orreproduction.3.1.2 bactericide, na physical or chemical agent that killsbacteria, but not necessar
7、ily bacterial spores.3.1.3 preservatives, na chemical agent(s) added to aproduct to reduce or prevent microbial growth.4. Summary of Test Method4.1 Aqueous material to be preserved is inoculated with anappropriate bacterial inoculum followed by addition of abactericide that will reduce populations o
8、f bacteria and preventthe growth of survivors for a specified period of time. Bacterialnumbers in the sample are determined at various time periodsand compared to a contol without any bactericide. The properlevel of antimicrobial is one that reduces and keeps theorganisms to an acceptable level in t
9、he test material.5. Significance and Use5.1 This test method should be used to determine if anantimicrobial preserves pigment suspensions, dye solutions,pulp slurries, starch solutions, polymers, sizing agents, latexemulsions, and other aqueous-based materials used in the paperindustry from bacteria
10、l spoilage.6. Apparatus6.1 BalanceTwo balances: one should be sensitive to 0.1g at a load of 200 g and have a platform to accommodatebottles being used in the test. The second balance (analytical)should be sensitive to 0.1 mg and should be employed to weighthe candidate preservative to be used in th
11、e preparation of thestock solutions.6.2 BottlesBorosilicate glass milk dilution bottles or othersuitable containers fitted either with screw caps or Escherrubber stoppers. These bottles are used for water blanks andaqueous-based samples.1This test method is under the jurisdiction of ASTM Committee E
12、35 onPesticides and Alternative Control Agents and is the direct responsibility ofSubcommittee E35.15 on Antimicrobial Agents.Current edition approved April 1, 2007. Published April 2007. Originallyapproved in 1980. Last previous edition approved in 2002 as E 723 97 (2002).2For referenced ASTM stand
13、ards, 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.1Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, P
14、A 19428-2959, United States.6.3 Colony CounterAny one of several types may be usedas the Quebec, Buck, and Wolfhuegel. A hand tally for therecording of the bacterial count is recommended if manualcounting is done. Alternatively, an automated video colonycounter may also be used.6.4 Culture Tube Clos
15、uresAppropriate nontoxic closuresshould be selected.6.5 Culture TubesRecommended size is 15 by 125 mm or18 by 150 mm without lip, and preferably of borosilicate glass.6.6 BlenderAny blender that will assure proper agitationand blending.6.7 Flaming EquipmentDepending upon circumstances,either an alco
16、hol lamp or bunsen burner may be used to flameinoculating needles and other equipment.6.8 Incubators, capable of maintaining temperatures of 28to 70 61C to provide proper incubation temperatures. Tem-perature should be consistent with the temperature of theproduct to be preserved.6.9 Petri Dishes, 1
17、00 by 15-mm, plastic or borosilicateglass, sterile.6.10 pH MeasurementAny reliable pH meter is suitable tostandardize the pH of the culture medium. Nonbleeding testsstrips are recommended for samples.6.11 Pipets, 1.1 or 2.2-mL milk dilution type, 1.0-mLgraduated in 0.01 mL, 10-mL graduated in 0.1 mL
18、 andappropriately calibrated pipettors may be used. Serologicalpipets and pipettors should not be used for highly viscousmaterials.6.12 Sterilizers, pressurized steam sterilizer or hot air ovencapable of 180 6 2C for 2 6 0.2 h.7. Microbicides and Materials7.1 Freshly prepared test solutions of the a
19、ntimicrobial shallbe used in all tests.7.2 Purity of WaterAll references to water as diluent orreagent shall mean distilled water or water of equal purity,unless otherwise noted (see Specification D 1193, Type III).7.3 Test MaterialsFreshly prepared pigment slurries, ad-hesives, dye rosin, polymer,
20、sizing solutions, and other mate-rials to be preserved should be used as the substrate.7.4 Culture Medium:7.4.1 Standard dehydrated tryptone glucose extract agar orother medium that is known to recover organisms from thematerial to be tested is recommended.7.4.2 For some substrates it may be necessa
21、ry to add a smallamount of nutrient material to ensure growth of the organismsin the material to be studied for preservation. For bacterialpreservation studies, add 5.0 mL/L of 0.2 % nutrient broth tothe test material to assure sufficient populations.8. Test Organisms8.1 The test organisms will vary
22、 with the material to bepreserved and the purpose of the test. For specific materialsthat are contaminated, that material will serve as the inoculum.For general screening of activity or preventative evaluations,the inoculum may consist of a single or mixed culture oforganisms that are known to cause
23、 problems in the material tobe preserved. The viability of the microorganisms in thematerial to be tested should be verified prior to initiating thetest.8.2 To provide a uniformly inoculated substrate, the inocu-lum should be added to the entire quantity of the test substrateat one time, mixed thoro
24、ughly, and then dispensed into theseparate test bottles.8.3 The material under test should be inoculated withsufficient microorganisms either from pure cultures or con-tained in the spoiled material used as the inoculum to give abacterial count of 13106CFU/mL or higher.9. Procedure9.1 Dispense 50-g
25、aliquots (or any other suitable quantity)of the inoculated test material aseptically into sterile bottles (ifnecessary add nutrient). Treat the samples immediately withappropriate concentrations of the antimicrobial. Set up controlsin duplicate. Note appropriate physical characteristics such aspH, c
26、olor, odor, viscosity etc., of all test samples at this time.9.1.1 Make the following additions aseptically to eachbottle in the order named and shake vigorously after eachaddition, using 20 complete cycles in a vertical motion.9.1.2 Add the desired volume of the stock solution of theantimicrobial t
27、o be tested to give the desired concentration inparts per million or percent. Stock solution of the antimicrobialshould be of such strength so that the volume of antimicrobialsolution added is no more than 1 % of the total volume ofsample in each bottle. Do not add an antimicrobial to thecontrol. In
28、clude in each test a minimum of five concentrationsof the antimicrobial under test. Suggested antimicrobial con-centrations are 50, 100, 200, 300, 400, and 600 ppm, orwhatever range of concentration may be suitable for thematerial to be tested. Record the pH of all samples at thebeginning of the exp
29、eriment, using nonbleeding test strips.9.1.3 Incubate all samples at 28 to 37C (or other tempera-ture at which the test material will be stored, such as 65C forstarch solutions) with the bottle capped tightly to avoidevaporation. The organisms being tested need to be viable andstable at the test tem
30、perature.9.1.4 Determine concentration of viable organisms in thecontrols at the time of biocide addition and of all samples atperiodic intervals after biocide addition (see Practices E 1054).This can be done with standard plate counts or other acceptedalternative means of determining concentrations
31、 of organisms(see Guide E 1326). All plates or recovery medium should beincubated at the same temperature as the test.9.1.5 Time intervals for determining the level of organismsremaining in the sample depend on the length of time the testmaterial needs to be preserved in actual use and the acceptabl
32、elevel of contamination when the material is to be used. Thus,samples can be taken at 3 h, 8 h, 24 h, 48 h, 72 h, or weekly,depending on how rapidly and to what extent the inoculumneeds to be killed.9.1.6 If the material can be re-inoculated while it is pre-served or to determine the number of re-in
33、oculations a givenpreservative level will be able to handle, the samples should bere-inoculated on a weekly or biweekly interval with $13106CFU/mL (see 8.3).E7230729.1.7 Typical test protocols range from biweekly inocula-tions with sampling 24 h postinoculation to weekly inocula-tions with sampling
34、7 days postinoculation. For comparativestudies, these intervals and the inoculum must be constant. Asin any lab test, it is difficult to duplicate the conditions thatmight exist in an actual production facility.9.1.8 At each time interval (see 9.1.5) or after reinoculation(see 9.1.6), mix the sample
35、 thoroughly and immediatelydetermine the level of microorganisms in the sample (see9.1.4). The controls must maintain a high count throughout thestudy or show visual signs of deterioration. Either criterion canbe used to indicate spoilage and the validity of the test.10. Calculation10.1 At each samp
36、ling time and at the end of each test,calculate the number of bacteria killed at each microbicideconcentration tested as follows:% kill 5control plate count 2 test plate count!control plate count3 10010.1.1 The percent kill at any given time is indicative of theeffectiveness of the antimicrobial und
37、er test. The proper levelof antimicrobial to use for the material being tested is the onethat decreases the level of organisms to the acceptable level oforganisms for the test material in an acceptable amount of time.Visual deterioration and other signs of degradation, such aschanges in pH, color, o
38、dor, loss of viscosity, and so forth,should also be used to judge the degree of preservationobtained.NOTE 1Typically, the level of organisms should be reduced to lessthan 13103CFU/mL in a 24 h period of time. This is equivalent to a99.9 % kill when the inoculum provides 13106CFU/mL.11. Precision and
39、 Bias11.1 A precision and bias statement cannot be made.12. Keywords12.1 bacterial; bactericide; paper based products; preserva-tive; pulp; spoilageASTM International takes no position respecting the validity of any patent rights asserted in connection with any item mentionedin this standard. Users
40、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 standard is subject to revision at any time by the responsible technical committee and must be reviewed every fi
41、ve 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 International Headquarters. Your comments will receive careful consideration at a meeting of theresponsible technical
42、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 below.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), 610-832-9555 (fax), or serviceastm.org (e-mail); or through the ASTM website(www.astm.org).E723073
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