SIST EN ISO 15088:2009

SLOVENSKI STANDARD
SIST EN ISO 15088:2009
01-maj-2009
.DNRYRVWYRGH'RORþHYDQMHDNXWQHWRNVLþQRVWLRGSDGQLKYRGDQDMDMþHFDFHEULF
'DQLRUHULR,62
Water quality - Determination of the acute toxicity of waste water to zebrafish eggs
(Danio rerio) (ISO 15088:2007)
Wasserbeschaffenheit - Bestimmung der akuten Toxizität von Abwasser auf Zebrafisch-
Eier (Danio rerio) (ISO 15088:2007)
Qualité de l'eau - Détermination de la toxicité aiguë des eaux résiduaires vis-à-vis des
oeufs de poisson-zèbre (Danio rerio) (ISO 15088:2007)
Ta slovenski standard je istoveten z: EN ISO 15088:2008
ICS:
13.060.30 Odpadna voda Sewage water
13.060.70 Preiskava bioloških lastnosti Examination of biological
vode properties of water
SIST EN ISO 15088:2009 en,fr,de
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

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SIST EN ISO 15088:2009

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SIST EN ISO 15088:2009
EUROPEAN STANDARD
EN ISO 15088
NORME EUROPÉENNE
EUROPÄISCHE NORM
November 2008
ICS 13.060.70

English Version
Water quality - Determination of the acute toxicity of waste water
to zebrafish eggs (Danio rerio) (ISO 15088:2007)
Qualité de l'eau - Détermination de la toxicité aiguë des Wasserbeschaffenheit - Bestimmung der akuten Toxizität
eaux résiduaires vis-à-vis des oeufs de poisson-zèbre von Abwasser auf Zebrafisch-Eier (kursiv(Danio rerio)) (ISO
(Danio rerio) (ISO 15088:2007) 15088:2007)
This European Standard was approved by CEN on 29 October 2008.
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 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 Management Centre has the same status as the
official versions.
CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Cyprus, Czech Republic, Denmark, Estonia, Finland,
France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal,
Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland and United Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
Management Centre: rue de Stassart, 36  B-1050 Brussels
© 2008 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 15088:2008: E
worldwide for CEN national Members.

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SIST EN ISO 15088:2009
EN ISO 15088:2008 (E)
Contents Page
Foreword.3

2

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SIST EN ISO 15088:2009
EN ISO 15088:2008 (E)
Foreword
The text of ISO 15088:2007 has been prepared by Technical Committee ISO/TC 147 “Water quality” of the
International Organization for Standardization (ISO) and has been taken over as EN ISO 15088:2008 by
Technical Committee CEN/TC 230 “Water analysis” the secretariat of which is held by DIN.
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 May 2009, and conflicting national standards shall be withdrawn at the
latest by May 2009.
Attention is drawn to the possibility that some of the elements of this document may be the subject of patent
rights. CEN [and/or CENELEC] 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, Cyprus, Czech
Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia,
Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain,
Sweden, Switzerland and the United Kingdom.
Endorsement notice
The text of ISO 15088:2007 has been approved by CEN as a EN ISO 15088:2008 without any modification.

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SIST EN ISO 15088:2009

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SIST EN ISO 15088:2009

INTERNATIONAL ISO
STANDARD 15088
First edition
2007-10-01

Water quality — Determination of the
acute toxicity of waste water to zebrafish
eggs (Danio rerio)
Qualité de l'eau — Détermination de la toxicité aiguë des eaux
résiduaires vis-à-vis des œufs de poisson-zèbre (Danio rerio)




Reference number
ISO 15088:2007(E)
©
ISO 2007

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SIST EN ISO 15088:2009
ISO 15088:2007(E)
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©  ISO 2007
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means,
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ii © ISO 2007 – All rights reserved

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SIST EN ISO 15088:2009
ISO 15088:2007(E)
Contents Page
Foreword .iv
Introduction.v
1 Scope.1
2 Normative references.1
3 Terms and definitions .1
4 Interferences .2
5 Principle.3
6 Reagents.3
7 Apparatus.3
8 Procedure.4
9 Determination of toxicological endpoints.7
10 Validity criteria.8
11 Determination of LID .9
egg
12 Expression of results.9
13 Test report.9
14 Reproducibility .9
Annex A (informative) Scheme for layout and distribution of eggs in microplates.11
Bibliography.12

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SIST EN ISO 15088:2009
ISO 15088:2007(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.
International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2.
The main task of technical committees is to prepare International Standards. Draft International Standards
adopted by the technical committees are circulated to the member bodies for voting. Publication as an
International Standard requires approval by at least 75 % of the member bodies casting a vote.
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.
ISO 15088 was prepared by Technical Committee ISO/TC 147, Water quality, Subcommittee SC 5, Biological
methods.
iv © ISO 2007 – All rights reserved

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SIST EN ISO 15088:2009
ISO 15088:2007(E)
Introduction
Fish play a major role in the aquatic food web. They are high-order consumers (often final consumers) and
have important functions of regulation in aquatic ecosystems. They are a confirmed part within test concepts
regarding aquatic organisms from different trophic levels.
The eggs of the zebrafish (Danio rerio Hamilton-Buchanan) are used as test material. The zebrafish belongs
to the family of Cyprinidae (carp-related fish) within the class of Osteichthyes (teleost fish). Zebrafish are easy
to keep and produce transparent, nonadhesive eggs (diameter about 1 mm) throughout the whole year. Their
embryonic development is well described. The zebrafish is one of the most important model fish in research
on the developmental biology of vertebrates and is recommended as a test fish, i.e. in the OECD Guidelines
203, 204 and 210.
The development of fertilized fish eggs can be affected by water constituents and effluents. Death of embryos
and certain defined disturbances of embryonic development, which finally lead to death, are considered
effects.

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SIST EN ISO 15088:2009

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SIST EN ISO 15088:2009
INTERNATIONAL STANDARD ISO 15088:2007(E)

Water quality — Determination of the acute toxicity of waste
water to zebrafish eggs (Danio rerio)
WARNING — Persons using this International Standard shall be familiar with normal laboratory
practice. This International Standard does not purport to address all of the safety problems, if any,
associated with its use. It is the responsibility of the user to establish appropriate safety and health
practices and to ensure compliance with any national regulatory conditions.
IMPORTANT — It is absolutely essential that tests conducted according to this International Standard
be carried out by suitably qualified staff. When applying this International Standard, it is necessary in
each case to determine if and to what extent additional conditions should be established.
1 Scope
This International Standard specifies a method for the determination of degrees of dilution or of concentrations
as a measure of the acute toxic effect of waste water to fish eggs within 48 h. This International Standard is
also applicable to treated municipal waste water and industrial effluents.
NOTE This International Standard has been elaborated as a substitute for the acute fish toxicity test. Applied to
waste water, it gives the same or similar results as achieved from the acute fish toxicity test (e.g. ISO 7346-1 or
ISO 7346-2). If used for single substances, different sensitivities from both test systems are possible.
2 Normative references
The following referenced documents are indispensable for the application 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 5667-16, Water quality — Sampling — Part 16: Guidance on biotesting of samples
ISO 5814, Water quality — Determination of dissolved oxygen — Electrochemical probe method
ISO 7346-1, Water quality — Determination of the acute lethal toxicity of substances to a freshwater fish
(Brachydanio rerio Hamilton-Buchanan (Teleostei, Cyprinidae)] — Part 1: Static method
ISO 7346-2, Water quality — Determination of the acute lethal toxicity of substances to a freshwater fish
(Brachydanio rerio Hamilton-Buchanan (Teleostei, Cyprinidae)] — Part 2: Semi-static method
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1
control
dilution water with fertilized fish eggs
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SIST EN ISO 15088:2009
ISO 15088:2007(E)
3.2
dilution level
D
reciprocal value of the volume fraction of waste water in dilution water in which the test is conducted
EXAMPLE 250 ml of waste water in a total volume of 1 000 ml (volume fraction of 25 %) represents dilution level
D = 4.
NOTE See ISO 5667-16.
3.3
EC
50
concentration at which there is an effect on 50 % of the organisms in line with the test criterion
NOTE In this International Standard, LC is the lethal concentration at which 50 % of the organisms are dead.
50
3.4
fish egg
generally used term for all development stages of an egg cell (inside the chorion) within this International
Standard
NOTE If necessary, the following qualifiers for this term are used: “not fertilized”, immediately after spawning; “freshly
fertilized”, between 4-cell stage and 128-cell stage; “embryo”, if the developing embryo is visible inside the egg before
hatching.
3.5
lowest ineffective dilution
dilution factor
LID
lowest ineffective dilution tested, expressed as dilution level D (3.2), at which no inhibition, or only effects not
exceeding the test-specific variability, are observed
3.6
LID
egg
lowest ineffective dilution within a test batch in which at least 90 % of the fish eggs do not show any effect
according to this International Standard
NOTE See Clauses 11 and 12 and Figure 2.
3.7
test batch
defined dilutions with fertilized fish eggs
4 Interferences
The differentiation of eggs can be difficult in the case of heavily coloured and/or turbid waste water. In such
cases, the following modified preparation order is used before starting the exposure:
⎯ spawning;
⎯ differentiation into fertilized and unfertilized eggs (8.2.2);
⎯ transfer into dilution water (7.10, crystallization bowls);
⎯ transfer into microplates without previous exposure.
While transferring into microplates, the error in dilution caused by adherent water should be kept as small as
possible.
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SIST EN ISO 15088:2009
ISO 15088:2007(E)
5 Principle
In a dilution series, waste water graduated to integral volume ratio is mixed with dilution water giving defined
dilution levels, D,. After exposure of fertilized fish eggs to the
...