SIST EN ISO 20150:2019

SLOVENSKI STANDARD
SIST EN ISO 20150:2019
01-maj-2019
2EXWHYLQVHVWDYQLGHOLREXWYH.YDQWLWDWLYQDSUHVNXVQDPHWRGD]DRFHQMHYDQMH
SURWLJOLYLþQHJDGHORYDQMD,62
Footwear and footwear components - Quantitative challenge test method to assess
antifungal activity (ISO 20150:2019)
Schuhe und Schuhbestandteile - Qualitative Prüfverfahren zur Bestimmung der Aktivität
gegen Pilzbefall (ISO 20150:2019)
Chaussures et composants de chaussure - Méthode de test d'épreuve quantitatif pour
évaluer l'activité antifongique (ISO 20150:2019)
Ta slovenski standard je istoveten z: EN ISO 20150:2019
ICS:
61.060 Obuvala Footwear
SIST EN ISO 20150:2019 en,fr,de
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

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SIST EN ISO 20150:2019

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SIST EN ISO 20150:2019


EN ISO 20150
EUROPEAN STANDARD

NORME EUROPÉENNE

February 2019
EUROPÄISCHE NORM
ICS 61.060
English Version

Footwear and footwear components - Quantitative
challenge test method to assess antifungal activity (ISO
20150:2019)
Chaussures et composants de chaussure - Méthode de Schuhe und Schuhbestandteile - Quantitatives
test d'épreuve quantitatif pour évaluer l'activité Challengetestverfahren zur Bestimmung der
antifongique (ISO 20150:2019) antimykotischen Wirksamkeit (ISO 20150:2019)
This European Standard was approved by CEN on 28 December 2018.

CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this
European Standard the status of a national standard without any alteration. Up-to-date lists and bibliographical references
concerning such national standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN
member.

This European Standard exists in three official versions (English, French, German). A version in any other language made by
translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC Management
Centre has the same status as the official versions.

CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia,
Finland, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania,
Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland,
Turkey and United Kingdom.





EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION

EUROPÄISCHES KOMITEE FÜR NORMUNG

CEN-CENELEC Management Centre: Rue de la Science 23, B-1040 Brussels
© 2019 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 20150:2019 E
worldwide for CEN national Members.

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SIST EN ISO 20150:2019
EN ISO 20150:2019 (E)
Contents Page
European foreword . 3

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SIST EN ISO 20150:2019
EN ISO 20150:2019 (E)
European foreword
This document (EN ISO 20150:2019) has been prepared by Technical Committee ISO/TC 216
"Footwear" in collaboration with Technical Committee CEN/TC 309 “Footwear” the secretariat of which
is held by UNE.
This European Standard shall be given the status of a national standard, either by publication of an
identical text or by endorsement, at the latest by August 2019, and conflicting national standards shall
be withdrawn at the latest by August 2019.
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. CEN shall not be held responsible for identifying any or all such patent rights.
According to the CEN-CENELEC Internal Regulations, the national standards organizations of the
following countries are bound to implement this European Standard: Austria, Belgium, Bulgaria,
Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia,
France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta,
Netherlands, Norway, Poland, Portugal, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland,
Turkey and the United Kingdom.
Endorsement notice
The text of ISO 20150:2019 has been approved by CEN as EN ISO 20150:2019 without any modification.

3

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SIST EN ISO 20150:2019

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SIST EN ISO 20150:2019
INTERNATIONAL ISO
STANDARD 20150
First edition
2019-01
Footwear and footwear components —
Quantitative challenge test method to
assess antifungal activity
Chaussures et composants de chaussure — Méthode de test d'épreuve
quantitatif pour évaluer l'activité antifongique
Reference number
ISO 20150:2019(E)
©
ISO 2019

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SIST EN ISO 20150:2019
ISO 20150:2019(E)

COPYRIGHT PROTECTED DOCUMENT
© ISO 2019
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting
on the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address
below or ISO’s member body in the country of the requester.
ISO copyright office
CP 401 • Ch. de Blandonnet 8
CH-1214 Vernier, Geneva
Phone: +41 22 749 01 11
Fax: +41 22 749 09 47
Email: copyright@iso.org
Website: www.iso.org
Published in Switzerland
ii © ISO 2019 – All rights reserved

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SIST EN ISO 20150:2019
ISO 20150:2019(E)

Contents Page
Foreword .v
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Principle . 2
5 Safety . 2
6 Apparatus . 2
7 Reagents and culture medium . 3
7.1 General . 3
7.2 Water . 3
7.3 Malt medium . 3
7.3.1 Composition . 3
7.3.2 Preparation . 3
7.4 Malt extract agar (MEA) medium . 4
7.4.1 Composition . 4
7.4.2 Preparation . 4
7.5 Physiological saline (sodium chloride solution) . 4
7.5.1 Composition . 4
7.5.2 Preparation . 4
7.6 Wetting agent (nonionic surfactant) . 4
7.7 Buffer solution . 4
7.7.1 Buffer stock . 4
7.7.2 Preparation of buffer stock . 5
7.7.3 Preparation of buffer solution . 5
8 Test microorganisms . 5
9 Preparation of test inoculums . 5
9.1 Indications for use of strains . 5
9.2 Preparation of inoculums of Candida albicans . 6
9.3 Preparation of test spore suspension of filamentous micro-fungi . 6
10 Preparation of test specimens . 6
10.1 General . 6
10.2 Test specimen . 7
10.3 Pre-treatment of test specimen . 7
11 Test procedure . 7
11.1 Summary of test methods . 7
11.2 Dynamic challenge tests . 7
11.2.1 Inoculation . 7
11.2.2 Neutralization and elution after inoculation (time 0 h) . 8
11.2.3 Incubation . 8
11.2.4 Neutralization and elution after incubation (time 24 h) . 8
11.2.5 Determination of the number of viable micro-fungi . 8
11.3 Static challenge test . 8
11.3.1 Inoculation . 8
11.3.2 Neutralization and elution after inoculation (time 0 h) . 9
11.3.3 Incubation . 9
11.3.4 Neutralization and elution after incubation (time 24 h) . 9
11.3.5 Determination of the number of viable micro-fungi . 9
12 Expression of results . 9
12.1 Calculation of the number of viable micro-fungi . 9
12.2 Judgement of test effectiveness . 9
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12.3 Calculation of antifungal activity ratio .10
13 Test report .10
Bibliography .12
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SIST EN ISO 20150:2019
ISO 20150:2019(E)

Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out
through ISO technical committees. Each member body interested in a subject for which a technical
committee has been established has the right to be represented on that committee. International
organizations, governmental and non-governmental, in liaison with ISO, also take part in the work.
ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of
electrotechnical standardization.
The procedures used to develop this document and those intended for its further maintenance are
described in the ISO/IEC Directives, Part 1. In particular, the different approval criteria needed for the
different types of ISO documents should be noted. This document was drafted in accordance with the
editorial rules of the ISO/IEC Directives, Part 2 (see www .iso .org/directives).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
any patent rights identified during the development of the document will be in the Introduction and/or
on the ISO list of patent declarations received (see www .iso .org/patents).
Any trade name used in this document is information given for the convenience of users and does not
constitute an endorsement.
For an explanation of the voluntary nature of standards, the meaning of ISO specific terms and
expressions related to conformity assessment, as well as information about ISO's adherence to the
World Trade Organization (WTO) principles in the Technical Barriers to Trade (TBT) see www .iso
.org/iso/foreword .html.
This document was prepared by Technical Committee ISO/TC 216, Footwear.
Any feedback or questions on this document should be directed to the user’s national standards body. A
complete listing of these bodies can be found at www .iso .org/members .html.
© ISO 2019 – All rights reserved v

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SIST EN ISO 20150:2019

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SIST EN ISO 20150:2019
INTERNATIONAL STANDARD ISO 20150:2019(E)
Footwear and footwear components — Quantitative
challenge test method to assess antifungal activity
CAUTION — Test methods specified herein require the use of micro-fungi. These tests are only
to be carried out in facilities with containment techniques for handling microorganisms and by
persons with training and experience in the use of microbiological techniques.
1 Scope
This document specifies quantitative challenge test methods for evaluating the antifungal activity of
footwear and footwear components.
This document is applicable only to footwear and components that claim to have antifungal (antimycotic)
properties or antimicrobial properties.
Two methods can be applied. The choice of method depends on the material properties and test
microorganisms. Dynamic challenge test method can be applied to all types of materials. For single
absorbent materials, static challenge test method is recommended. Brief descriptions of each method
are given in 11.2 and 11.3.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
ISO 7218, Microbiology of food and animal feeding stuffs — General requirements and guidance for
microbiological examinations
ISO 19952, Footwear — Vocabulary
3 Terms and definitions
For the purpose of this document, the terms and definitions given in ISO 19952 and the following apply.
ISO and IEC maintain terminological databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https: //www .iso .org/obp
— IEC Electropedia: available at http: //www .electropedia .org/
3.1
antifungal activity
antimycotic activity
efficacy of a material or finish used to prevent or mitigate the growth of micro-fungi, to reduce the
number of micro-fungi or to kill micro-fungi
3.2
control specimen
material identical to the test material but without antifungal treatment
Note 1 to entry: If no control specimen is available, sterilized conical flask can be used as control specimen.
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3.3
neutralizer
chemical agents used to inactivate, neutralize, or quench the antifungal properties of antifungal agents
[SOURCE: ISO 20743:2013, 3.7, modified — “antibacterial” has been replaced with “antifungal”.]
4 Principle
The test specimens and control specimens are inoculated with a spore suspension of a selected test
strain of micro-fungi specified or claimed. Two test methods are available to assess antifungal activity.
Antifungal performance is quantitatively determined by counting the number of viable micro-fungi and
calculating the antifungal activity ratio.
5 Safety
The handling of microorganisms which are potentially hazardous requires a high degree of technical
competence and can be subject to current national legislation and regulations. Only personnel trained
in microbiological techniques should carry out such tests.
NOTE: Refer to country-specific codes of practice for personal hygiene, disinfection and sterilization.
It is recommended that the person who perform the test should consult IEC 60068-2-10:2005,
Appendix A, and ISO 7218.
6 Apparatus
6.1 General
Disposable apparatus is an acceptable alternative to re-usable glassware and plastic if it has suitable
specifications.
Usual microbiological laboratory equipment in accordance with ISO 7218 and in particular the
following.
6.2 Biological safety cabinet.
6.3 Incubator, capable of maintaining a temperature of (28 ± 2) °C.
6.4 Autoclave, capable of maintaining a temperature of (121 ± 2) °C and a pressure of (103 ± 5) kPa,
for wet sterilization, used in accordance with ISO 7218.
6.5 Humidity chamber, capable of maintaining a temperature of (28 ± 2) °C and a relative humidity of
(85 ± 5) %.
6.6 Ultraviolet lamp.
6.7 Wide mouth jars, with cap, 100 ml, capable of being used with an autoclave (6.4).
6.8 Vortex mixer.
6.9 Centrifugal machine, 2 000 × g.
6.10 Dimensional shaker, two dimensional or three dimensional, capable of adjusting to 50 r/min.
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6.11 Shaking incubator, capable of maintaining a temperature of (28 ± 2) °C and a rotational frequency
of (120 ± 10) r/min.
6.12 Glass beads, 2 mm to 3 mm in diameter, 10 beads to 15 beads per conical flask, for preparation of
fungal spore solutions.
6.13 Glass wool or medical gauze (double layers), for preparation of fungal spore solutions.
6.14 Oven, for dry sterilization.
6.15 pH-meter, capable of measuring to ±0,2 units.
6.16 Balance, capable of weighing to ±0,01 g.
6.17 Spectrophotometer, capable of measuring at 500 nm to 660 nm wavelength, or McFarland’s
nephelometer.
6.18 Petri dishes, that have been sterilized, made of glass or plastic, in diameter sizes of 90 mm to
100 mm or 55 mm to 60 mm.
6.19 Pipette, having the most suitable volume for each use.
7 Reagents and culture medium
7.1 General
The preparation and test shall be freshly prepared in order to ensure the culture quality.
NOTE This could be done according to ISO 11133, or according to national standards or regulations.
Reagents used in tests shall be of analytical grade and/or suited for microbiological purposes.
7.2 Water
Water used in tests shall be analytical-grade water for microbiological media preparation, which is
freshly distilled and/or ion-exchanged and/or ultra-filtered and/or filtered with reverse osmosis.
It shall be free from all toxic or microorganism inhibitory substances.
7.3 Malt medium
7.3.1 Composition
Malt extract 30,0 g
Soya peptone 3,0 g
Water 1 000 ml
7.3.2 Preparation
Dissolve the designated amounts of components in distilled water, stir and adjust pH to (5,5 ± 0,2) at
room temperature, sterilize at (121 ± 2) °C for 15 min in an autoclave (6.4) with saturated water vapour.
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ISO 20150:2019(E)

7.4 Malt extract agar (MEA) medium
7.4.1 Composition
Malt extract 30,0 g
Soya peptone 3,0 g
Agar 15,0 g
Water 1 000 ml
7.4.2 Preparation
After mixing, stir and adjust pH to (5,5 ± 0,2) at room temperature. Heat with stirring on a hotplate
or in a boiling-water bath until the components are completely dissolved, sterilize at (121 ± 2) °C for
15 min in an autoclave (6.4) with saturated water vapour. Cool and shake solution well, then pour into
the Petri dishes.
NOTE The potato dextrose agar (PDA) can also provide a complete medium for the growth of micro-fungi.
The standard PDA medium can be obtained as a commercial product thereby avoiding the preparation steps
of cooking and the variation in the composition of potato species. The commercial PDA medium with standard
composition can be used to avoid the influence of composition of potato and operation when boiling it, and Malt
Extract Agar (MEA) medium can be obtained from commercial source.
7.5 Physiological saline (sodium chloride solution)
7.5.1 Composition
Sodium chloride, NaCl 8,5 g
Water 1 000 ml
7.5.2 Preparation
After well mixing, adjust pH to (6,9 ± 0,2) at room temperature and sterilize at (121 ± 2) °C for 15 min.
7.6 Wetting agent (nonionic surfactant)
To be used for harvesting the spores, does not react with other reagents and does not cause a
reduction or increase in micro-fungi number, such as polysorbate 80 (TWEEN 80), N-methyltauride,
1)
TM
Triton X-100 or polyglycol ether and so on.
NOTE Wetting agent (nonionic surfactant) can be used when the specimen has coating.
7.7 Buffer solution
7.7.1 Buffer stock
Potassium dihydrogen phosphate, KH PO 34,0 g
2 4
Water 1 000 ml
1) TritonTM X-100 is the trade name of a product supplied by SIGMA-ALDRICH. This information is given for
the convenience of users of this document and does not constitute an endorsement by ISO of the product named.
Equivalent products may be used if they can be shown to lead to the same results.
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7.7.2 Preparation of buffer stock
Prepare phosphate buffer solution by placing 34,0 g of potassium dehydrogenate phosphate in a
1 000 ml volumetric flask. Add 500 ml of water and mix to dissolve. Adjust the pH to 7,2 ± 0,2 at room
temperature with sodium hydroxide. Add distilled water to make up to 1 000 ml.
7.7.3 Preparation of buffer solution
Transfer 1 ml of buffer stock solution and 0,08 g of wetting agent (nonionic surfactant) (7.6), corresponds
to 0,01 % and dilute to 800 ml with distilled water. After mixing well, sterilize at (121 ± 2) °C for 15 min.
NOTE If wetting agent (nonionic surfactant) is not required, it might be omitted.
8 Test microorganisms
The strain used shall be stated in the test report.
The species that shall be used in antifungal activity tests are listed in Table 1.
Table 1 — Test strains
a ®
Micro-fungi Name WDCM No. CGMCC No. ATCC No.
TM b
Yeast Candida albicans 00054 AS 1.2031 ATCC® 10231
TM b
Aspergillus niger 00144 AS 3.4463 ATCC® 6275
Filamentous micro-fungi/Mould
TM b
Aspergillus brasiliensis 00053 AS 3.5487 ATCC® 16404
TM b
Filamentous micro-fungi/Dermato- Trichophyton menta- 00191 — ATCC® 9533
phyte grophytes
Key
WDCM:  World Data Centre for Microogranisms
CGMCC:  China General Microbiological Culture Collection Centre
ATCC®:  American Type Culture Collection
NOTE 1  Refer to WDCM and website: http: //refs .wdcm .org.
NOTE 2  Other micro-fungi (appropriate species or other appropriate strains) can be used after appropriate validation.
a
Test strains shall be obtained from agencies of the World Federation of Culture Collection (WFCC). The micro-fungal
species and their supply sources shall be stated in the test reports.
b TM TM TM TM
ATCC® 10231 , ATCC® 6275 , ATCC® 16404 and ATCC® 9533 are examples of suitable products available
commercially. This information is given for the convenience of users of this document and does not constitute an
endorsement by ISO of these products.
Testing of yeast (Candida albicans) is mandatory.
If activity against mould is claimed, testing of one mould strain shall be performed.
If activity against dermatophyte is claimed, testing of Trichophyton mentagrophytes shall be performed.
Strains can be preserved in accordance with the supplier’s instructions or EN 12353.
9 Preparation of test inoculums
9.1 Indications for use of strains
Apply strains within 4 generations.
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ISO 20150:2019(E)

9.2 Preparation of inoculums of Candida albicans
9.2.1 Using a sterile inoculating loop, transfer one Candida albicans colony into 20 ml of malt medium
(7.3) and incubate in the shaking incubator at (28
...