ASTM E1054-21

Standard Test Practices for Evaluation of Inactivators of Antimicrobial Agents

SIGNIFICANCE AND USE
5.1 The effectiveness of antimicrobial agents incorporated into disinfectants, sanitizers, and antiseptics is measured by their ability to kill microorganisms within a specified contact time. Hence, accurate determination of antimicrobial effectiveness requires complete and immediate inactivation (neutralization) of the antimicrobial agent. Inefficient or incomplete neutralization will permit killing or inactivation of microorganisms to continue beyond the experimental exposure time, resulting in an overestimation of antimicrobial activity.  
5.2 The neutralization methods commonly used in antimicrobial effectiveness evaluations are chemical inactivation, dilution, and filtration. All critical parameters of an antimicrobial effectiveness evaluation—for example, media, equipment, microorganism(s), and temperature of solutions—must be duplicated in the performance of selected neutralization procedure.  
5.3 The neutralization evaluation must include at least three replications (five replications in Section 9) so that a statistical analysis of the microbial recovery data can be performed. The number of replicates used in the evaluation depends on the statistical significance required for the expected results, the variability encountered in the data, and the relative effectiveness of the neutralization procedure.  
5.4 A limitation of these evaluation procedures is that they use microorganisms that have not been exposed to an antimicrobial agent. Under experimental conditions, cells exposed to neutralization procedures are likely to be damaged to different degrees by the antimicrobial agent. Sublethal injury may be a factor in recovery, and the effect of the neutralization procedure on recovery of injured organisms should be examined. This method is not intended to assess recovery of injured organisms.
Note 3: Ideally, all microorganisms used in the antimicrobial effectiveness evaluation should be tested in the neutralization assay. However, representative organism...
SCOPE
1.1 These test procedures are used to determine the effectiveness of methodologies procedures and materials intended for inactivating (neutralizing, quenching) the microbicidal properties of antimicrobial agents; to ensure that no components of the neutralizing procedures and materials, themselves, exert an inhibitory effect on microorganisms targeted for recovery; and to demonstrate that the antimicrobial chemistry tested is microbicidal.  
1.2 Knowledge of microbiological and statistical techniques is required for these procedures.
Note 1: These methods are not suitable when testing the virucidal activity of microbicides (see Test Method E1482).  
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.  
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use.  
1.5 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.

General Information

• Status: Historical
• Publication Date: 31-Mar-2021
• ICS: 11.080.20 - Disinfectants and antiseptics
• ICS: 71.100.35 - Chemicals for industrial and domestic disinfection purposes
• Technical Committee: E35 - Pesticides, Antimicrobials, and Alternative Control Agents
• Drafting Committee: E35.15 - Antimicrobial Agents

Standards Content (Sample)

NOTICE: This standard has either been superseded and replaced by a new version or withdrawn.
Contact ASTM International (www.astm.org) for the latest information
Designation: E1054 − 21
Standard Test Practices for
1
Evaluation of Inactivators of Antimicrobial Agents
This standard is issued under the fixed designation E1054; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision.Anumber in parentheses indicates the year of last reapproval.A
superscript epsilon (´) indicates an editorial change since the last revision or reapproval.
1. Scope toxicity Reduction and Neutralization
E2756Terminology Relating toAntimicrobial andAntiviral
1.1 These test procedures are used to determine the effec-
Agents
tiveness of methodologies procedures and materials intended
for inactivating (neutralizing, quenching) the microbicidal
3. Terminology
properties of antimicrobial agents; to ensure that no compo-
3.1 Definition of Terms: For definition of terms not listed
nentsoftheneutralizingproceduresandmaterials,themselves,
below, refer to E2756, Standard Terminology Relating to
exert an inhibitory effect on microorganisms targeted for
Antimicrobial and Antiviral Agents.
recovery; and to demonstrate that the antimicrobial chemistry
tested is microbicidal. 3.2 Definitions of Terms Specific to This Standard:
3.2.1 antimicrobial effectiveness evaluation, adj/n—a deter-
1.2 Knowledgeofmicrobiologicalandstatisticaltechniques
mination of microbicidal properties of an antimicrobial agent
is required for these procedures.
by methods, such as Test Methods E645 and E1115.
NOTE 1—These methods are not suitable when testing the virucidal
activity of microbicides (see Test Method E1482).
3.2.2 CFU/mL (abbrev.)—colony-forming units of a micro-
organism per millilitre of fluid.
1.3 The values stated in SI units are to be regarded as
standard. No other units of measurement are included in this
3.2.3 neutralizer effectiveness, adj/n—ability of a neutral-
standard.
ization procedure to inactivate or quench the microbicidal
properties of an antimicrobial agent.
1.4 This standard does not purport to address all of the
safety concerns, if any, associated with its use. It is the
3.2.4 neutralizer toxicity, adj/n—any inhibitory effects a
responsibility of the user of this standard to establish appro-
neutralization procedure may have on the survival of a micro-
priate safety, health, and environmental practices and deter-
bial population.
mine the applicability of regulatory limitations prior to use.
3.2.5 test material control, adj/n—an evaluation of the
1.5 This international standard was developed in accor-
activity of a test material in reducing a known population of
dance with internationally recognized principles on standard-
microorganisms.
ization established in the Decision on Principles for the
3.2.6 test organism viability, adj/n—the population of a
Development of International Standards, Guides and Recom-
challenge microorganism used in a neutralization assay.
mendations issued by the World Trade Organization Technical
Barriers to Trade (TBT) Committee. 3.2.7 viability, n—the ability of a challenge microorganism
to form colonies or grow on a nutrient medium.
2. Referenced Documents 3.2.7.1 Discussion—In the context of these test methods,
2 “viability” is understood to be synonymous with cultivability.
2.1 ASTM Standards:
E1115Test Method for Evaluation of Surgical Hand Scrub
4. Summary of Test Methods
Formulations
NOTE 2—The neutralization test procedure selected must be consistent
E1482Practice for Use of Gel Filtration Columns for Cyto-
with the methods of testing used in the antimicrobial effectiveness
evaluation.
4.1 Neutralization Assay with Recovery on Semi-solid
1
These test practices are under the jurisdiction of ASTM Committee E35 on
Medium—Neutralization assay for antimicrobial effectiveness
Pesticides, Antimicrobials, and Alternative Control Agents and are the direct
tests that recover and quantify microbial populations on
responsibility of Subcommittee E35.15 on Antimicrobial Agents.
semi-solid (agar) media. This method is appropriate for anti-
Current edition approved April 1, 2021. Published May 2021. Originally
microbial agents that are chemically inactivated or diluted to
approved in 1985. Last previous edition approved in 2013 as E1054–13. DOI:
10.1520/E1054-21.
sub-inhibitory levels and performed entirely in vitro or includ-
2
For referenced ASTM standards, visit the ASTM website, www.astm.org, or
ing an in vivo component to verify neutralization of an
contact ASTM Customer Service at [e-mail]. For Annual Book of ASTM
antimicrobial formulation sampled from the skin of a human
Standards volume information, refer to the standard’s Document Summary page on
the ASTM website. volunteer.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
1

---------------------- Page: 1
...

This document is not an ASTM standard and is intended only to provide the user of an ASTM standard an indication of what changes have been made to the previous version. Because
it may not be technically possible to adequately depict all changes accurately, ASTM recommends that users consult prior editions as appropriate. In all cases only the current version
of the standard as published by ASTM is to be considered the official document.
Designation: E1054 − 08 (Reapproved 2013) E1054 − 21
Standard Test MethodsPractices for
1
Evaluation of Inactivators of Antimicrobial Agents
This standard is issued under the fixed designation E1054; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. A
superscript epsilon (´) indicates an editorial change since the last revision or reapproval.
1. Scope
1.1 These test methodsprocedures are used to determine the effectiveness of methodologies procedures and agents materials
intended for inactivating (neutralizing, quenching) the microbicidal properties of antimicrobial agents, and agents; to ensure that
no components of the neutralizing procedures and agents,materials, themselves, exert an inhibitory effect on microorganisms
targeted for recovery.recovery; and to demonstrate that the antimicrobial chemistry tested is microbicidal.
1.2 Knowledge of microbiological and statistical techniques is required for these procedures.
NOTE 1—Knowledge of microbiological and statistical techniques is required for these procedures. These methods are not suitable when testing the
virucidal activity of microbicides (see Test Method E1482).
1.3 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.
1.4 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility
of the user of this standard to establish appropriate safety safety, health, and healthenvironmental practices and determine the
applicability of regulatory limitations prior to use.
1.5 This international standard was developed in accordance with internationally recognized principles on standardization
established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued
by the World Trade Organization Technical Barriers to Trade (TBT) Committee.
2. Referenced Documents
2
2.1 ASTM Standards:
E645 Practice for Evaluation of Microbicides Used in Cooling Water Systems
E1115 Test Method for Evaluation of Surgical Hand Scrub Formulations
E1482 Practice for Use of Gel Filtration Columns for Cytotoxicity Reduction and Neutralization
E2756 Terminology Relating to Antimicrobial and Antiviral Agents
3. Terminology
3.1 Definition of Terms: For definition of terms not listed below, refer to E2756, Standard Terminology Relating to Antimicrobial
and Antiviral Agents.
3.2 Definitions of Terms Specific to This Standard:
1
These test methodspractices are under the jurisdiction of ASTM Committee E35 on Pesticides, Antimicrobials, and Alternative Control Agents and are the direct
responsibility of Subcommittee E35.15 on Antimicrobial Agents.
Current edition approved April 1, 2013April 1, 2021. Published April 2013May 2021. Originally approved in 1985. Last previous edition approved in 20082013 as
E1054 – 08.E1054 – 13. DOI: 10.1520/E1054-08R13.10.1520/E1054-21.
2
For referenced ASTM standards, visit the ASTM website, www.astm.org, or contact ASTM Customer Service at [e-mail]. For Annual Book of ASTM Standards
volume information, refer to the standard’s Document Summary page on the ASTM website.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
1

---------------------- Page: 1 ----------------------
E1054 − 21
3.1.1 antimicrobial, adj—describes an agent that kills or inactivates microorganisms or suppresses their growth or reproduction.
3.2.1 antimicrobial effectiveness evaluation, adj/n—a determination of microbicidal properties of an antimicrobial agent by
methods, such as Test Methods E645 and E1115.
3.2.2 CFU/mL (abbrev.)—colony-forming units of a microorganism per millilitre of fluid.
3.1.4 neutralization, n—a physical or chemical procedure that inactivates or quenches the microbicidal properties of an
antimicrobial agent.
3.2.3 neutralizer effectiveness, adj/n—ability of a neutralization procedure to inactivate or quench the microbicidal properties of
an antimicrobial agent.
3.2.4 neutralizer toxicity, adj/n—any inhibitory effects a neutralization procedure may have on the survival of a microbial
population.
3.2.5 test material control, adj/n—an evaluation of the activity of a test material in reducing a known population of
microorganisms.
3.2.6 test organism viability, adj/n—the population of a challenge microorganism used in a neutrali
...