EN ISO 4973:2023

Cosmetics - Microbiology - Quality control of culture media and diluents used in cosmetics standards (ISO 4973:2023)

This document specifies the minimum requirements for quality control of microbiological culture media and diluents in order to demonstrate their ability to detect microorganisms and to ensure reliability of the microbiological test methods described in the ISO cosmetics microbiology standards.
This document describes mainly growth promotion and microbial control tests and is applicable to both commercially ready-to-use culture media and culture media prepared from dehydrated culture media or basic constituents in the user’s laboratory.
Other methods can be substituted provided that their equivalence has been demonstrated.

Kosmetische Mittel - Mikrobiologie - Qualitätskontrolle von Nährmedien und Verdünnungsmitteln, die in den Normen für kosmetischen Mittel verwendet werden (ISO 4973:2023)

Dieses Dokument legt die Mindestanforderungen an die Qualitätskontrolle mikrobiologischer Nährmedien und Verdünnungsmitteln fest, um deren Fähigkeit zum Nachweis von Mikroorganismen nachzuweisen und die Zuverlässigkeit der in den ISO Normen zur Mikrobiologie kosmetischer Mittel beschriebenen mikrobiologischen Untersuchungsverfahren sicherzustellen.
Dieses Dokument beschreibt hauptsächlich Untersuchungen auf Wachstumsförderung und zur mikrobiellen Kontrolle und ist sowohl für handelsübliche Fertignährmedien als auch für Nährmedien, die im Labor des Anwenders aus Trockennährmedien oder Grundbestandteilen hergestellt werden, anwendbar.
Andere Verfahren können ersatzweise verwendet werden, sofern ihre Gleichwertigkeit nachgewiesen wurde.

Cosmétiques - Microbiologie - Contrôle qualité des milieux de culture et des diluants utilisés dans les normes relatives aux cosmétiques (ISO 4973:2023)

Le présent document spécifie les exigences minimales relatives au contrôle qualité des milieux de culture microbiologique et des diluants afin de démontrer leur capacité à assurer la détection de micro-organismes et de garantir la fiabilité des méthodes d'essais microbiologiques décrites dans les normes ISO relatives à la microbiologie cosmétique.
Le présent document décrit principalement des essais de fertilité et de contrôle microbien; il est applicable à la fois aux milieux de culture prêts à l'emploi disponibles dans le commerce et aux milieux de culture préparés à partir de milieux de culture déshydratés ou de constituants de base dans le laboratoire de l'utilisateur.
D'autres méthodes peuvent être utilisées sous réserve que leur équivalence ait été démontrée.

Kozmetika - Mikrobiologija - Kontrola kakovosti gojišč in razredčil, ki se uporabljajo v standardih za kozmetiko (ISO 4973:2023)

Ta dokument določa minimalne zahteve za kontrolo kakovosti mikrobioloških gojišč in razredčil, da se dokaže njihova sposobnost zaznavanja mikroorganizmov in zagotovi zanesljivost mikrobioloških preskusnih metod, opisanih v mikrobioloških standardih ISO za kozmetiko. Dokument opisuje predvsem preskuse spodbujanja rasti in mikrobiološkega nadzora ter se uporablja tako za gojišča, ki so komercialno pripravljena za uporabo, kot gojišča, pripravljena iz dehidriranih gojišč ali osnovnih sestavin v uporabnikovem laboratoriju.
Druge metode je mogoče nadomestiti, če je bila dokazana njihova enakovrednost.

General Information

• Status: Published
• Publication Date: 03-Oct-2023
• ICS: 07.100.40 - Cosmetics microbiology
• Technical Committee: CEN/TC 392 - Cosmetics
• Drafting Committee: CEN/TC 392/WG 2 - Microbiological methods
• Current Stage: 6060 - Definitive text made available (DAV) - Publishing
• Start Date: 04-Oct-2023
• Due Date: 16-Jul-2024
• Completion Date: 04-Oct-2023
• Ref Project: SIST EN ISO 4973:2023 - Cosmetics - Microbiology - Quality control of culture media and diluents used in cosmetics standards (ISO 4973:2023)

Standards Content (Sample)

SLOVENSKI STANDARD
01-december-2023
Kozmetika - Mikrobiologija - Kontrola kakovosti gojišč in razredčil, ki se
uporabljajo v standardih za kozmetiko (ISO 4973:2023)
Cosmetics - Microbiology - Quality control of culture media and diluents used in
cosmetics standards (ISO 4973:2023)
Kosmetische Mittel - Mikrobiologie - Qualitätskontrolle von Nährmedien und
Verdünnungsmitteln, die in den Normen für kosmetischen Mittel verwendet werden (ISO
4973:2023)
Cosmétiques - Microbiologie - Contrôle qualité des milieux de culture et des diluants
utilisés dans les normes relatives aux cosmétiques (ISO 4973:2023)
Ta slovenski standard je istoveten z: EN ISO 4973:2023
ICS:
07.100.40 Kozmetika - mikrobiologija Cosmetics microbiology
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

EN ISO 4973
EUROPEAN STANDARD
NORME EUROPÉENNE
October 2023
EUROPÄISCHE NORM
ICS 07.100.40
English Version
Cosmetics - Microbiology - Quality control of culture media
and diluents used in cosmetics standards (ISO 4973:2023)
Cosmétiques - Microbiologie - Contrôle qualité des Kosmetische Mittel - Mikrobiologie - Qualitätskontrolle
milieux de culture et des diluants utilisés dans les von Nährmedien und Verdünnungsmitteln, die in den
normes relatives aux cosmétiques (ISO 4973:2023) Normen für kosmetischen Mittel verwendet werden
(ISO 4973:2023)
This European Standard was approved by CEN on 26 September 2023.

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, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway,
Poland, Portugal, Republic of North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Türkiye 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
© 2023 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 4973:2023 E
worldwide for CEN national Members.

Contents Page
European foreword . 3

European foreword
This document (EN ISO 4973:2023) has been prepared by Technical Committee ISO/TC 217
"Cosmetics" in collaboration with Technical Committee CEN/TC 392 “Cosmetics” the secretariat of
which is held by AFNOR.
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 April 2024, and conflicting national standards shall be
withdrawn at the latest by April 2024.
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.
Any feedback and questions on this document should be directed to the users’ national standards
body/national committee. A complete listing of these bodies can be found on the CEN website.
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, France, Germany, Greece, Hungary, Iceland,
Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Republic of
North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Türkiye and the
United Kingdom.
Endorsement notice
The text of ISO 4973:2023 has been approved by CEN as EN ISO 4973:2023 without any modification.

INTERNATIONAL ISO
STANDARD 4973
First edition
2023-09
Cosmetics — Microbiology — Quality
control of culture media and diluents
used in cosmetics standards
Cosmétiques — Microbiologie — Contrôle qualité des milieux
de culture et des diluants utilisés dans les normes relatives aux
cosmétiques
Reference number
ISO 4973:2023(E)
ISO 4973:2023(E)
© ISO 2023
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: [phone]
Email: [e-mail]
Website: www.iso.org
Published in Switzerland
ii
ISO 4973:2023(E)
Contents Page
Foreword .v
Introduction . vi
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Principle . 3
4.1 General information. 3
4.2 pH . 3
4.3 Absence of microbiological growth . 3
4.4 Growth promotion. 3
4.5 Selective and indicative properties. 3
5 Diluents, neutralizers and culture media . 4
5.1 General . 4
5.2 Diluents and neutralizers . 4
5.3 Culture media . 4
6 Apparatus and glassware .4
7 Strains of microorganisms . 4
8 Procedure .6
8.1 General recommendation . 6
8.2 Preparation of strains . 6
8.2.1 General . 6
8.2.2 Preparation of bacterial and Candida albicans suspensions . 9
8.2.3 Preparation of Aspergillus brasiliensis spore stock suspension . 9
8.2.4 Control of the concentration of the calibrated suspension . 10
8.3 Absence of microbial growth . 10
8.3.1 Solid culture media . 10
8.3.2 Liquid culture media and diluents . 10
8.4 Growth promotion. 10
8.4.1 Solid culture media . 10
8.4.2 Liquid culture media . 11
8.5 Selective properties  . 11
8.5.1 Solid culture media — For indicative properties . 11
8.5.2 Solid culture media — For inhibition properties . 11
9 Expression of results .11
9.1 Absence of microbial growth . 11
9.1.1 Solid culture media . 11
9.1.2 Liquid culture media . 11
9.2 Growth promotion. 11
9.2.1 Solid culture media . 11
9.2.2 Liquid culture media . 11
9.3 Selective and indicative properties.12
9.3.1 Solid culture media — For indicative properties .12
9.3.2 Solid culture media — For inhibition properties .12
10 Interpretation and acceptance criteria .12
10.1 General .12
10.2 Absence of microbial growth .12
10.3 Growth promotion.12
10.3.1 Solid culture media .12
10.3.2 Growth promotion — Liquid media culture .13
10.4 Selective properties of solid culture media . 13
iii
ISO 4973:2023(E)
10.4.1 For indicative properties . 13
10.4.2 For inhibition properties . . 13
Annex A (normative) Specifications for growth promoting, inhibitory and indicative
properties of culture media .14
Annex B (informative) Quality control report for culture media.19
Bibliography .21
iv
ISO 4973:2023(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 document 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).
ISO draws attention to the possibility that the implementation of this document may involve the use
of (a) patent(s). ISO takes no position concerning the evidence, validity or applicability of any claimed
patent rights in respect thereof. As of the date of publication of this document, ISO had not received
notice of (a) patent(s) which may be required to implement this document. However, implementers are
cautioned that this may not represent the latest information, which may be obtained from the patent
database available at www.iso.org/patents. ISO shall not be held responsible for identifying any or all
such patent rights.
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 217, Cosmetics, in collaboration with
the European Committee for Standardization (CEN) Technical Committee CEN/TC 392, Cosmetics, in
accordance with the Agreement on technical cooperation between ISO and CEN (Vienna Agreement).
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.
v
ISO 4973:2023(E)
Introduction
The quality of culture media used in the current standards for cosmetic microbiology is an essential
part of microbiological analysis reliability and needs to be verified.
Checking different parameters of culture media such as growth promotion, absence of microbial growth
for non-inoculated culture media, physical characteristics, can help to assess their quality.
This document is intended to provide methods to assess the quality of the media used in cosmetics
microbiology standards and define the minimum acceptance criteria required to ensure their
performance.
This applies to:
a) commercially ready-to-use culture media;
b) culture media prepared from dehydrated culture media plus additional ingredients, or only
ingredients.
vi
INTERNATIONAL STANDARD ISO 4973:2023(E)
Cosmetics — Microbiology — Quality control of culture
media and diluents used in cosmetics standards
1 Scope
This document specifies the minimum requirements for quality control of microbiological culture
media and diluents in order to demonstrate their ability to detect microorganisms and to ensure
reliability of the microbiological test methods described in the ISO cosmetics microbiology standards.
This document describes mainly growth promotion and microbial control tests and is applicable to
both commercially ready-to-use culture media and culture media prepared from dehydrated culture
media or basic constituents in the user’s laboratory.
Other methods can be substituted provided that their equivalence has been demonstrated.
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 21148, Cosmetics — Microbiology — General instructions for microbiological examination
3 Terms and definitions
For the purposes of this document, the following terms and definitions 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 https:// www .electropedia .org/
3.1
culture medium
mixture of ingredients, in liquid or solid form, prepared according to a formula and intended to support
the growth of microorganisms under specific conditions
Note 1 to entry: There are different types of culture media suitable for growing different types of microorganisms
depending on different included nutrients and chemicals present in the formulation.
3.1.1
batch of culture medium
lot of culture medium
homogenous and fully traceable unit of culture medium referring to a defined amount of bulk, which
has been produced within one defined production period, having been assigned the same batch number
3.1.2
ready-to-use culture medium
sterile liquid culture medium (3.1.3) or solid culture medium (3.1.4) that is supplied in plates, tubes, or
other containers in ready-to-use form
ISO 4973:2023(E)
3.1.3
liquid culture medium
culture medium (3.1) consisting in aqueous solution of one or more constituents, such as peptone water
or nutrient broth
Note 1 to entry: Liquid culture media in tubes, flasks or bottles are commonly called “broths”.
Note 2 to entry: Enrichment culture media are generally liquid media which, due to their composition, provide
favourable conditions for microorganisms’ multiplication.
3.1.4
solid culture medium
culture medium (3.1) containing solidifying substances (e.g. agar, gelatin) in different concentrations
3.1.5
selective culture medium
liquid culture medium (3.1.3) or solid culture medium (3.1.4) which allows specifically the growth of
a selected microorganism while inhibiting partially or totally the growth of different, non-target
microorganisms which can be in the product to be tested
Note 1 to entry: It may have indicative properties with growth of characteristic aspect of colonies if this is a solid
culture medium.
3.2
diluent
liquid phase designed to separate microorganisms from a solid culture medium (3.1.4) and/or to reduce
their concentration by dilution without multiplication or inhibition during the time of contact
Note 1 to entry: Diluent can contain neutralizing agent to inactivate the antimicrobial properties of the product.
3.3
strain
test microorganism used for quality control of the culture medium (3.1)
3.3.1
reference strain
test microorganism provided by a reference culture collection centre
3.3.2
reference stock culture
stored reference strain
set of separate identical cultures obtained by a single subculture from the reference strain (3.3.1)
Note 1 to entry: The reference stock culture or stored reference strain can be stored in a seed lot system (e.g.
single-use vials or beads) to maintain reference strains in the laboratory.
3.3.3
stock culture
subculture from a reference stock culture (3.3.2)
3.3.4
working culture
subculture from a reference stock culture (3.3.2) or stock culture (3.3.3) and is often kept as slants or
plates, used for preparation of calibrated microbial suspension
3.3.5
subculture
passage, i.e. transfer of organisms from a viable culture to fresh medium with growth of the
microorganisms
Note 1 to entry: Any form of subculturing is considered to be a transfer/passage.
ISO 4973:2023(E)
4 Principle
4.1 General information
The quality control of culture media refers to different parameters such as:
— pH;
— absence of microbial growth;
— growth promotion;
— selective and indicative properties (when relevant).
These are the key parameters to ensure and control the quality of the culture media. However,
particular attention should also be paid to:
— the manufacturer’s instructions;
— preparing conditions (volume, weighing, water quality);
— sterilization conditions (cycle time, temperature, pressure, packaging);
— storage conditions (temperature, duration).
Failure to comply with these instructions and conditions can affect appearance and functional
characteristics provided in the manufacturer’s guidance such as colour, gel consistency, clarity and
homogeneity.
NOTE For sterilization conditions and/or other conditions, see ISO 21148 and ISO 11133.
The user’s laboratory should ensure its own preparation process is accurate.
4.2 pH
pH is an essential physical parameter of all culture media.
The target pH value should be reached after autoclaving when culture media are prepared from
dehydrated media or basic constituents in the user’s laboratory.
4.3 Absence of microbiological growth
The purpose of this test is to check that the medium does not contain any microbiological contamination
which can interfere with microbial tests results.
4.4 Growth promotion
The purpose of this test is to ensure the ability of microorganisms to grow on the culture media.
NOTE Growth promotion is also called ‘productivity’ of culture medium.
4.5 Selective and indicative properties
The purpose of this test is to verify the ability of the culture medium to allow the growth of target
microorganisms and/or to confirm its colony morphology within the range of incubation time and
temperature.
An additional purpose of this test is to ensure that there is no growth of the target inhibited
microorganism(s).
ISO 4973:2023(E)
5 Diluents, neutralizers and culture media
5.1 General
The diluents, neutralizers and culture media suitable for enumeration and detection of microorganisms
are described in ISO 11930, ISO 16212, ISO 18415, ISO 18416, ISO 21149, ISO 21150, ISO 21322,
ISO 22717 and ISO 22718. Other diluents, neutralizers and culture media may be used if they have been
demonstrated to be suitable for use.
Use the general instructions given in ISO 21148. When water is mentioned in this international
standard, use water as specified in ISO 21148.
5.2 Diluents and neutralizers
Diluents may be used for preparation and dilutions of calibrated microbial suspensions and to disperse
the samples. In this case, it is required that a diluent contains neutralizers if the sample to be tested
has antimicrobial properties or contains preservatives. Diluents and neutralizers are described in
ISO 11930, ISO 16212, ISO 18415, ISO 18416, ISO 21149, ISO 21150, ISO 21322, ISO 22717 and ISO 22718.
5.3 Culture media
Culture media for enumeration and/or detection of microorganisms are described in ISO 11930,
ISO 16212, ISO 18415, ISO 18416, ISO 21149, ISO 21150, ISO 21322, ISO 22717 and ISO 22718.
Culture media may be prepared from dehydrated culture media using specific standards instructions
or the instructions provided by the manufacturer of the culture media.
Already prepared commercially manufactured ready-to-use culture media in liquid or solid form may
also be used if they meet the criteria described in this document.
6 Apparatus and glassware
The laboratory equipment, apparatus and glassware are described in ISO 21148.
7 Strains of microorganisms
The culture should be reconstituted according to the procedures provided by the supplier of the
refere
...