ASTM E2361-04
(Guide)Standard Guide for Testing Leave-On Products Using In-Situ Methods
Standard Guide for Testing Leave-On Products Using In-Situ Methods
SCOPE
1.1 This guide covers test methods and sampling procedure options for leave-on products for consumer and hospital personnel. Leave-on products, such as alcohol hand rubs and lotions containing antimicrobial ingredients, are increasingly marketed and used by consumers and health care personnel. These products are distinguished from conventional washing and scrubbing preparations in that they do not rely on the rinsing, physical removal, and antimicrobial action in determining their effectiveness. Although agitation and friction may serve to release organisms from the skin and folds and crevices, organisms are then killed in situ and are not rinsed from the skin surface before sampling. Appropriate test methods for the hands have been published, while other sampling methods will be needed for testing body areas other than the hands.
1.2 Methods of recovery after application of the contaminating organisms to a part of the body other than by the agitation/rubbing of the hands against a glass petri plate also need examination. Consideration should be given to contact plating, controlled swabbing with a template, and cup scrubbing (detergent/agitation used) since the target organisms for recovery are likely to be on the superficial layers of skin.
1.3 The values stated in SI units are to be regarded as the standard.
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 and health practices and determine the applicability of regulatory requirements prior to use.
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Designation:E2361–04
Standard Guide for
Testing Leave-On Products Using In-Situ Methods
This standard is issued under the fixed designation E 2361; 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 (e) indicates an editorial change since the last revision or reapproval.
1. Scope nel Handwash Formulations by Utilizing Fingernail Re-
gions
1.1 This guide covers test methods and sampling procedure
2.2 European Standard:
options for leave-on products for consumer and hospital
EN1500 Chemical Disinfectants and Antiseptics-Hygienic
personnel. Leave-on products, such as alcohol hand rubs and
Handrub-Test Method Requirements (phase 2/step 2)
lotions containing antimicrobial ingredients, are increasingly
approved by CEN (Comité Européen de Normalisation)
marketed and used by consumers and health care personnel.
These products are distinguished from conventional washing
3. Summary of Guide
and scrubbing preparations in that they do not rely on the
3.1 In this guide, choices of recovery techniques after the
rinsing, physical removal, and antimicrobial action in deter-
use of antimicrobial products will be considered. By the nature
mining their effectiveness.Although agitation and friction may
of the distribution of the skin flora, these sampling techniques
servetoreleaseorganismsfromtheskinandfoldsandcrevices,
estimate the flora remaining after antimicrobial use; some of it
organisms are then killed in situ and are not rinsed from the
is superficial and some hidden. An appropriate sampling
skin surface before sampling.Appropriate test methods for the
method can be selected depending on product use and the
hands have been published, while other sampling methods will
importance of superficial (transient) and hidden or deep
be needed for testing body areas other than the hands.
(mostly resident) flora.
1.2 Methods of recovery after application of the contami-
3.2 This guide was written because ASTM Subcommittee
nating organisms to a part of the body other than by the
E35.15 worked on its own test method for leave-on products
agitation/rubbing of the hands against a glass petri plate also
used without water, but found that the EN1500 protocol
need examination. Consideration should be given to contact
encompassed the test method that had been developed.
plating, controlled swabbing with a template, and cup scrub-
3.3 This CEN type of test methodology is widely used in
bing (detergent/agitation used) since the target organisms for
European and Scandinavian countries but has not been widely
recovery are likely to be on the superficial layers of skin.
used in the United States, although the use of alcohol/alcohol
1.3 The values stated in SI units are to be regarded as the
gel hand rubs has expanded greatly here in the last few years.
standard.
The underlying question is whether a test method designed for
1.4 This standard does not purport to address all of the
a leave-on product like alcohol or the conventional hand
safety concerns, if any, associated with its use. It is the
washingfollowedbysamplinginagloveorplasticbagismore
responsibility of the user of this standard to establish appro-
appropriate. There have been criticisms of test methods, such
priate safety and health practices and determine the applica-
as EN1500, which was based on Rotter’s methods (1), but
bility of regulatory requirements prior to use.
published data confirm that the test is highly reliable in
2. Referenced Documents showing consistent reduction levels with low variation from
2 subject to subject. Leave-on products that are not rinsed or
2.1 ASTM Standards:
washed off in use are primarily represented by alcohol-based
E 1174 Test Method for Evaluation of the Effectiveness of
hand rubs. However, other leave-on formulations have been
Health Care Personnel or Consumer Handwash Formula-
introduced and, undoubtedly, their number will increase in the
tions
future. Often test methods designed for washing/rinsing pro-
E 1327 Test Method for Evaluation of Health Care Person-
cedures have been used for these products. When different
more specific methods are required for testing, questions of
methodology become clearer, and the selection of a new or
This guide is under the jurisdiction ofASTM Committee E35 on Pesticides and
different sampling method is necessary.
Alternative ControlAgents and is the direct responsibility of Subcommittee E35.15
on Antimicrobial Agents.
Current edition approved April 1, 2004. Published May 2004.
2 3
For referenced ASTM standards, visit the ASTM website, www.astm.org, or Available from British Standards Institute (BSI), 389 Chiswick High Rd.,
contact ASTM Customer Service at service@astm.org. For Annual Book of ASTM London W4 4AL, U.K.
Standards volume information, refer to the standard’s Document Summary page on The boldface numbers in parentheses refer to the list of references at the end of
the ASTM website. this standard.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959, United States.
E2361–04
3.4 When a typical hand-washing product is used, the hands product application. Since changes have been made in Test
are wet; scrubbing and manipulation are pursued, often vigor- Method E 1327 to sample only after the first and last applica-
ously; and rinsing follows. Agitation here is to remove organ- tions, the applicability of this test method for products rubbed
isms and particulate and oily soil physically. Any residue of into the skin and used repeatedly without water may not be
applicable for these leave-on products.
active ingredient remaining on the skin is a small fraction of
the amount applied and assumed to be attached to the stratum 3.9 EN1500 is an adaptation of a test developed by Rotter
known as the Vienna Model (6).
corneum. The residual may also be absorbed over time.
Ultimately, the reduction in microbial count is a combination 3.10 There are many publications describing and evaluating
fingertip-sampling methods. One of the major criticisms of the
of kill from the antimicrobial and the physical removal by
methods is the procedure used for sampling. The tips of the
agitation and rinsing.
fingers and thumb are sampled by rubbing against the bottom
3.5 In contrast, leave-on products, such as alcohol products
of a glass petri dish to release contaminating bacteria from
intended to be applied and not rinsed off, present a different
these areas before and after treatment. The sampled areas are
situation.There are two distinct techniques when sampling: (1)
only portions of the areas treated. However, published results
sampling by washing target organisms off with detergent,
have shown consistent, statistically valid data. With the
assuming that most of removal is transient flora, and (2)
EN1500 test procedure, sampling is performed after a single
sampling in situ, for example, the cup scrub, swab, contact
use of the product (divided into two portions for application).
plate, or velvet block/pad that sample bacteria by impression
and contact or by using fluid to remove samples so that the
4. Significance and Use
volume of the sample is restricted to a very small size. These
4.1 The United States has concentrated attention and testing
different sampling methods disturb the deep or hidden flora to
efforts on surgical scrubbing far more than on hand care in
differing degrees. There has been an overwhelming concentra-
patient-to-patient routines. Great Britain, the originators of
tion of the cup-scrub sampling method as various test methods
infection control nursing, have always had their focus on
have been developed. The combination of detergent and
infection transmission. In the United States, published articles
agitation attempts to remove as much remaining flora as
have documented the short exposure time for health care
possible.Thebesteffort,however,onlyremovesabout15 %of
personnel who do wash their hands between patients. The
the full thickness flora (2). When other contact sampling or
average is less than 10 s.The ideal product for the reduction of
tape stripping are used, the distribution of bacterial colonies on
transient flora is one that rapidly kills or removes or both the
the skin are mirrored as they occur; whereas, if detergent/
microbial load acquired during health care activities. The
scrubbing techniques are used, the microcolonies are dispersed
emphasis on rapidity is essential simply because health care
yielding higher counts. Washing/scrubbing methods stir up the
personnel will not take the necessary time when using conven-
cells and bacteria from the deeper skin layers and release more
tional hand-washing products. The use of products not in-
of the hidden flora (described by Reybrouck (3)). This is also
tended for use with water has increased dramatically and their
true of the cup scrub method that uses detergent/surfactants to
use is common in European countries largely because of
detach bacteria from the skin. Contact methods sample the
convenience and effectiveness. A second characteristic is the
flora that can easily be transferred and that is conceded to be
level of antimicrobial action. The use of a rapid and potent
the most important in disease transmission. Williams (4) has
active product to reduce work-acquired microbial flora is ideal.
stated that, “although the distinction between residents and
4.2 Since the change from strictly in-vitro testing of topical
transients must certainly be a real one, it is not to allocate the
antimicrobials for use on skin to simulated use testing in hand
various bacterial species to one or other class with regularity.”
washing, prepping, site access testing, and sampling, emphasis
3.6 There has been a long-time focus on the cup-scrub
has always been on washing hands, agitating, rubbing, and
technique only, and it would be beneficial to look at sampling
brushing with liquid on the skin site to estimate bacteria
specific areas, such as Test Method E 1327, which samples
removed after testing.
around the fingernail region using a toothbrush, or the use of
4.3 The use of hard agitation has diminished with surgical
direct contact plating when washing is not involved (5),asin
scrubs without brushes or with only mild agitation and friction.
skin prepared for surgery. This guide is intended to assess the
4.4 There is a history of microbial dispersal(7) and increase
effectiveness of application of products rubbed into the skin or
insurfacebacteriafromdeeperlayersresultingfromshowering
on the hands when these sites are not washed between uses.
(8-10), washing, scrubbing, and agitation. In the normal
3.7 Superficially, the testing method is the same as with
situation on the skin, there is a superficial, surface flora and a
products that are used to scrub and wash the hands or skin in
deeper or hidden flora (3). The proportion of one to the other
that the hands are contaminated with a recoverable transient
has been addressed by Selwyn (2) and his judgment is that
organismandthetestproductapplied.Thesimilarityendshere.
from 20 to 50 % of the flora is “deep.” The intent in skin
3.8 If the hands are sampled after application of organisms sampling has almost always been to scrub, agitate, and use
and the test product in sequence, they are dried or gloved wet surfactant to remove as many organisms as we can. In doing
and are sampled after extensive rinsing. The stripping solution this, we have completely ignored the two types of flora.
is then added for sampling to increase the release of viable 4.5 Further, when the skin is treated with a cleansing agent
organisms to be recovered. In contrast, in testing for hand rubs or an antimicrobial that is subsequently rinsed away, the
or leave-on products, glove sampling would seem appropriate “deep” or “hidden” flora is pushed to the surface as the sebum
only if sampling were performed after each contamination and replenishes the sebum from the sebaceous glands removed in
E2361–04
washing. Many early investigators have looked at simpler comparing a sampling method to the culture results of a full
sampling methods that we now recognize were sampling thickness biopsy of the skin designated as 100 %. Another
primarily the superficial transient flora. technique developed and published by Leyden et al (13) may
enhance the use of some contact methods. This test method
5. Published Testing Methods for Leave-On Products
involves improved counting methodology using computer
5.1 Hygienic Hand Rub—Vienna Model:
imaging of colonies on contact plates from fingerprint/
5.1.1 When viable organisms are captured in the sampling
handprint techniques.
fluid after exposure to a test product, sampling like that used in
5.2.4 At any rate, we are sampling only a fraction of the
theglovejuicetestusesamuchlargeramountoffluidfollowed
total skin flora and sampling the hands presents even a bigger
by microbiological analysis on a small sample. While in the
dilemma. There are nail folds, cuticles, and fingernail spaces
test method in EN1500 and Rotter’s procedure (6), the volume
that collect bacteria. Price (14) himself showed that bacteria
in the plate after rubbing the fingertips on the plate’s bottom is
are released in each of a sequential series of twelve basins, so
either cultured in toto or sampled and diluted.
that a technique like the glove fluid sampling method is still
5.1.2 With this in-situ procedure, only the fingertips are
only releasing a fractional portion of the total flora. The
sampled in contrast to the whole hand in the glove juice
fingertips are the part of the hand most frequently in contact
procedure. The agitation to the fingertips in the in-situ testing
with people and hard surfaces. The important question be-
is more intense than 1 min of massage of the whole hand.
comes, “Is the fraction removed consistent?” for both pre- and
5.1.3 This test method has been legally mandated as the
post-sampling. The procedure in EN1500 does give consistent
officialCENmethodfortheirmembercountries.Itisdescribed
recovery, when statistical analyses are performed.
in this international standard as simulating practical conditions
5.2.5 In selecting a test method for assessing products used
for whether a product designed as a hygienic hand rub reduces
with repeated applications, the reason and frequency for the
the release of transient flora in use.The criteria specified in the
applications must be considered. The following standards are
standard require that the mean reduction shall not be less than
published test methods that have been used to test leave-on
achieved by a reference hand rub with pr
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