ASTM E1327-90(2000)

NOTICE: This standard has either been superseded and replaced by a new version or withdrawn.
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Designation:E1327–90(Reapproved 2000)
Standard Test Method for
Evaluation of Health Care Personnel Handwash
Formulations by Utilizing Fingernail Regions
This standard is issued under the fixed designation E 1327; 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 subtilis may be utilized to study (1) degree of physical removal
by handwashing techniques, and (2) the recovery and precision
1.1 This test method covers the determination of ability of
aspects of the test method.
antimicrobial handwashing agents to give reductions of tran-
sient microbial flora (bacterial flora) when used in a hand
4. Significance and Use
washing procedure.
4.1 The procedure should be used to test the degerming
1.2 A knowledge of microbiological techniques is required
effectiveness of antimicrobial hand washing products used by
for these procedures.
health care personnel that are intended for frequent use, and
1.3 This standard does not purport to address all of the
that are intended to reduce the level of contamination acquired
safety concerns, if any, associated with its use. It is the
through contact with contaminated objects or people.
responsibility of the user of this standard to establish appro-
4.2 Performance of these procedures requires the knowl-
priate safety and health practices and determine the applica-
edge of regulations pertaining to the protection of human
bility of regulatory limitations prior to use. For more specific
subjects.
hazard statements, see Note 1.
5. Apparatus
2. Referenced Documents
2 5.1 Colony Counter—Anyofseveraltypesmaybeused,for
2.1 ASTM Standards:
example, Quebec Colony Counter.
E 1054 Practices for Evaluating Inactivators of Antimicro-
5.2 Incubators—One incubator capable of maintaining a
bial Agents Used in Disinfectant, Sanitizer, Antiseptic, or
temperature of 25 6 2°C (this temperature required to ensure
Preserved Products
pigment production of Serratia); a second incubator capable of
3. Summary of Test Method maintaining 37 6 2°C is used for E. coli and B. subtilis
incubation is acceptable.
3.1 This test method, involving an improved method of
5.3 Water Bath—Capable of maintaining temperature of 80
recovering bacteria from hands, is used to study the effects of
6 2°C for heat shocking of B. subtilis spores is needed.
antimicrobial health care personnel handwash products. The
5.4 Sterilizer—Any suitable steam sterilizer capable of
groupofvolunteerpanelistsneednotrefrainfromusingtopical
producing the conditions of sterilization is acceptable.
antimicrobials(suchasdeodorantsoaps)beforeparticipatingin
5.5 Timer—Any stop-watch that can be read in minutes and
the study. All subjects wash their hands with a nonantimicro-
seconds is required.
bial hand soap prior to testing to remove any residual hand
5.6 Handwashing Sink—A sink of sufficient size to permit
lotionsandtolowerthenumbersofresidentskinflora.Activity
panelists to wash without touching hands to sink surface or
of products is measured by comparing the numbers of marker
other panelists is needed.
bacteria recovered from artifically contaminated fingernail
5.6.1 Water Faucet(s), to be located above the sink at a
regions after use of the handwashing formulations to the
height that permits the hands to be held higher than the elbow
numbers recovered from the artificially contaminated but
during the washing procedure.
unwashed fingernail regions. Broth cultures of Serratia marce-
5.6.2 Tap Water Temperature Regulator and Temperature
scens (a red pigmented bacterial species) and Escherichia coli
Monitor, to monitor and regulate water temperature of 40 6
(which produces fluorescent colonies on a special agar me-
2°C.
dium) are used as test bacteria.Aspore suspension of Bacillus
5.7 Quad Petri plates, 100 by 15 mm, plastic, sterile,
disposable.
This test method is under the jurisdiction of ASTM Committee E35 on
Pesticides and is the direct responsibility of Subcommittee E35.15 onAntimicrobial
and Antiviral Agents. Federal Register, Vol 46, No. 17, Jan. 27, 1981.
Current edition approved Jan. 26, 1990. Published March 1990. Presterilized disposable quad plastic petri plates, the two sizes of glass petri
Withdrawn. plates and other equipment are available from most local laboratory supply houses.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959, United States.
E1327–90 (2000)
5.8 Small Petri Plates, 60 by 15 mm, glass. 7.3 Bacillus subtilis,ATCC No. 19659. Grow in tryptic soy
5.9 Large Petri Plates, 150 by 15 mm, glass. broth at 37°C.
5.10 Tooth Brushes: 7.4 Preparation of Spore Suspension—Inoculate each sur-
5.10.1 Young Size. faceoftwotrypticsoyagarplates(30mLagarin150-mmpetri
5.10.2 Battery Operated. plates) with 1 mL of B. subtilis tryptic soy broth culture.
5.11 Ultraviolet Lamp, having separate short wave and long Spread over the entire surface of the agar. Incubate for 5 to 10
wave bulbs. days at 37°C. Suspend the growth in 20 mL of 0.1 % tryptone
8 11
5.12 Germicidal Lamp Monitor Strips. water by rubbing the agar surface with a sterile rubber
policeman.Add ethanol to the suspension to a final concentra-
6. Reagents and Materials
tion of 80 % (wt/wt) and store in a refrigerator.
7.5 Other bacteria containing adequate markers to enable
6.1 Bacteriological Pipettes, 10.0 mL, sterile.
distinction from normal flora and of known safety may also be
6.2 Pipettors and Pipette Tips, Eppendorf, MLA or similar
used for testing purposes.
types.
6.3 Disposable Analyzer Cusp, 2 mL, plastic, not sterile.
NOTE 1—Warning: Theapplicationofmicroorganismstotheskinmay
6.4 Sampling Solution—Dissolve 0.4 g KH PO , 10.1 g
involve a health risk. Prior to applying S. marcescens or other bacteria to
2 4
Na PO and 1.0 g isooctylphenoxypolyethoxyethanol in1L the skin, the antibiotic sensitivity profile of the strain should be deter-
2 4
mined. If the Serratia strain is not sensitive to gentamycin, it should not
distilled water. Adjust pH to 7.8 with 0.1 N HCl or 0.1 N
be used. If an infection occurs, the antibiotic sensitivity profile should be
NaOH. Dispense in 100 mL-volumes and sterile for 20 min at
made available to an attending clinician. Following the panelist’s con-
121°C.
tamination and testing for the day, the panelist’s hands should be
6.5 Dilution Fluid—The sampling fluid may be used for
decontaminated with a 70 % ethanol solution. Care should be taken to
dilutions or use Butterfields sterile phosphate buffered water
decontaminate around the fingernail regions.
adjusted to pH 7.2 with suitable inactivator for the antimicro-
7.6 Preparation of Marker Culture Suspension—Inoculatea
bial. Adjust pH with 0.1 N HCl or 0.1 N NaOH (see Practices
10-mLtryptic soy broth tube with each of the test bacteria and
E 1054).
incubateeachtubeatthetemperatureindicated.Whenstudying
6.6 Agar, Tryptic soy agar or equivalent. Include the appro-
mixed inocula, mix equal volumes of the cultures into a sterile
priate inactivator if needed.
test tube; an equivalent volume of B. subtilis spore suspension
6.7 Agar with MUG—Tryptic soy agar with 60 to 80 µg/mL
(that is prepared by centrifuging the alcoholic suspension and
4-methylumbelliferyl-b-D-glucuronide (MUG) is required.
resuspending cells in water) may be added for bacterial
6.8 Test Formulations—Directions for use of test formula-
physical removal determinations. Keep mixed suspension on
tion should be included if available. If these are not available,
ice during the days testing.
liquid antimicrobial soap formulations are tested by same
routine as the nonantimicrobial control (10.5); alcoholic lotion
8. Panelists
type formulations are rubbed to dryness and then sampled for
8.1 Recruit a sufficient number of healthy adult human
survivors (10.7).
volunteers who have no clinical evidence of dermatosis, open
6.9 Nonantimicrobial Control Soap, a liquid castile soap or
wounds, hangnail or other skin disorders. The number of
other liquid soap containing no antimicrobials.
people needed for a trial is dependent on the number of
6.10 Broth—Tryptic soy broth or equivalent is required.
treatments within a study.
8.2 Volunteers are asked to maintain their normal use of
7. Test Organisms
soaps, shampoos, etc.They are asked to refrain from the use of
7.1 Serratia marcescens American Type Culture Collec-
acids, bases, solvents on the hands during the test period.
tion,ATCCNo.14756istobeusedasamarkerorganism.This
Gloves should be provided for use where exposure to these
is a strain having stable pigmentation. Grow in tryptic soy
agents is unavoidable.
broth at 28°C.
7.2 Escherichia coli, ATCC No. 11229 is used as another
9. Experimental Design
gram negative marker organism. Grow in tryptic soy broth at
9.1 Each fingernail of a volunteer may be assayed sepa-
37°C.
rately; therefore, 10 test determinations (replicates) may be
obtained from one volunteer. For the comparison of several
products during a single study, a design such as a Latin Square
Oral B-30 tooth brushes, av
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