SIST ISO 10706:2010
Water quality - Determination of long term toxicity of substances to Daphnia magna Straus (Cladocera, Crustacea)
Water quality - Determination of long term toxicity of substances to Daphnia magna Straus (Cladocera, Crustacea)
This International Standard describes a method for the determination of the long term sublethal toxicity to Daphnia
magna Straus (Cladocera, Crustacea) of:
a) chemical substances, which are soluble under the conditions of the test, or can be maintained as stable
suspensions or dispersions under the conditions of the test,
b) industrial or sewage effluents, treated or untreated, after decanting, filtration, or centrifugation,
c) surface or ground waters.
NOTE This International Standard is adapted from OECD Guideline 211[1].
Qualité de l'eau - Détermination de la toxicité à long terme de substances vis-à-vis de Daphnia magna Straus (Cladocera, Crustacea)
La présente Norme internationale décrit une méthode pour la détermination de la toxicité sublétale à long terme
vis-à-vis de Daphnia magna Straus (Cladocera, Crustacea) des substances suivantes:
a) substances chimiques solubles dans les conditions de l'essai ou pouvant être maintenues en suspension ou
en dispersion stable dans les conditions de l'essai;
b) effluents industriels ou urbains, épurés ou non, s'il y a lieu après décantation, filtration ou centrifugation;
c) eaux de surface ou eaux souterraines.
NOTE La présente Norme internationale est adaptée de la Ligne directrice 211 de l'OCDE [1].
Kakovost vode - Določevanje dolgotrajne strupenosti snovi z Daphnia magna Straus (Cladocera, Crustacea)
Ta mednarodni standard opisuje metodo določevanja dolgotrajne subletalne strupnosti snovi z Daphnia magnaStraus (Cladocera, Crustacea) za:
a) kemične substance, ki so topne pod pogoji preskusa oziroma se lahko ohranjajo kot stabilne suspenzije ali disperzije pod pogoji preskusa,
b) industrijske ali komunalne odplake, obdelane ali neobdelane, po pretočitvi, filtraciji ali centrifugiranju,
c) površinske in podtalne vode.
General Information
Standards Content (Sample)
SLOVENSKI STANDARD
01-september-2010
.DNRYRVWYRGH'RORþHYDQMHGROJRWUDMQHVWUXSHQRVWLVQRYL]'DSKQLDPDJQD
6WUDXV &ODGRFHUD&UXVWDFHD
Water quality - Determination of long term toxicity of substances to Daphnia magna
Straus (Cladocera, Crustacea)
Qualité de l'eau - Détermination de la toxicité à long terme de substances vis-à-vis de
Daphnia magna Straus (Cladocera, Crustacea)
Ta slovenski standard je istoveten z: ISO 10706:2000
ICS:
13.060.70 Preiskava bioloških lastnosti Examination of biological
vode properties of water
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
INTERNATIONAL ISO
STANDARD 10706
First edition
2000-04-01
Water quality — Determination of long term
toxicity of substances to Daphnia magna
Straus (Cladocera, Crustacea)
Qualité de l'eau — Détermination de la toxicité à long terme de substances
vis-à-vis de Daphnia magna Straus (Cladocera, Crustacea)
Reference number
©
ISO 2000
PDF disclaimer
This PDF file may contain embedded typefaces. In accordance with Adobe's licensing policy, this file may be printed or viewed but shall not
be edited unless the typefaces which are embedded are licensed to and installed on the computer performing the editing. In downloading this
file, parties accept therein the responsibility of not infringing Adobe's licensing policy. The ISO Central Secretariat accepts no liability in this
area.
Adobe is a trademark of Adobe Systems Incorporated.
Details of the software products used to create this PDF file can be found in the General Info relative to the file; the PDF-creation parameters
were optimized for printing. Every care has been taken to ensure that the file is suitable for use by ISO member bodies. In the unlikely event
that a problem relating to it is found, please inform the Central Secretariat at the address given below.
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means, electronic
or mechanical, including photocopying and microfilm, without permission in writing from either ISO at the address below or ISO's member body
in the country of the requester.
ISO copyright office
Case postale 56 � CH-1211 Geneva 20
Tel. + 41 22 749 01 11
Fax + 41 22 734 10 79
E-mail copyright@iso.ch
Web www.iso.ch
Printed in Switzerland
ii © ISO 2000 – All rights reserved
Contents Page
Foreword.iv
Introduction.v
1 Scope .1
2 Normative references .1
3 Principle.1
4 Test environment.1
5 Reagents and materials .2
6 Apparatus .3
7 Treatment and preparation of samples .3
7.1 Special precautions for sampling and transportation of water or effluent samples.3
7.2 Preparation of solutions of substances to be tested.3
8 Procedure .4
8.1 Controls .4
8.2 Selection of exposure concentrations .4
8.3 Renewal of exposure solutions.5
8.4 Introduction of animals into the test system .5
8.5 Feeding of organisms .5
8.6 Observations and measurements.5
8.7 Validity of the test.6
9 Data analysis and expression of results .6
10 Test report .7
Annex A (informative) OECD M4 and M7 media — Preparation and acclimation of Elendt M4 and M7
media.9
Annex B (informative) ASTM reconstituted hard fresh water.12
Annex C (informative) Data collection sheet .13
Annex D (informative) Calculation of time-weighted mean concentration.15
Bibliography.17
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 3.
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 International Standard may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights.
International Standard ISO 10706 was prepared by Technical Committee ISO/TC 147, Water quality,
Subcommittee SC 5, Biological methods.
Annexes A, B, C and D of this International Standard are for information only.
iv © ISO 2000 – All rights reserved
Introduction
This International Standard defines a procedure for the determination of the long term sublethal toxicity of
chemicals, waters and waste waters to the water-flea Daphnia magna Straus. The methodology is adapted from a
guideline produced by the Organisation for Economic Co-operation and Development (OECD Guideline 211, see
reference [1] in the Bibliography).
INTERNATIONAL STANDARD ISO 10706:2000(E)
Water quality — Determination of long term toxicity of substances
to Daphnia magna Straus (Cladocera, Crustacea)
WARNING — Activated sludge and sewage contain potentially pathogenic organisms. Therefore,
appropriate precautions should be taken when handling them. Toxic test compounds and those whose
properties are unknown should be handled with care.
1 Scope
This International Standard describes a method for the determination of the long term sublethal toxicity to Daphnia
magna Straus (Cladocera, Crustacea)of:
a) chemical substances, which are soluble under the conditions of the test, or can be maintained as stable
suspensions or dispersions under the conditions of the test,
b) industrial or sewage effluents, treated or untreated, after decanting, filtration, or centrifugation,
c) surface or ground waters.
[1]
NOTE This International Standard is adapted from OECD Guideline 211 .
2 Normative references
The following normative documents contain provisions which, through reference in this text, constitute provisions of
this International Standard. For dated references, subsequent amendments to, or revisions of, any of these
publications do not apply. However, parties to agreements based on this International Standard are encouraged to
investigate the possibility of applying the most recent editions of the normative documents indicated below. For
undated references, the latest edition of the normative document referred to applies. Members of ISO and IEC
maintain registers of currently valid International Standards.
ISO 5667-2, Water quality — Sampling — Part 2: Guidance on sampling techniques.
ISO 5667-16, Water quality — Sampling — Part 16: Guidance on biotesting of samples.
3Principle
Daphnia magna females, less than 24 h old, are exposed for a period of 21 days to a test substance, industrial or
sewage effluent or surface/ground water added to dilution water in a range of concentrations in a semi-static or
flow-through system. The survival of parents is recorded together with the number of live offspring produced per
live parent at the end of the test.
The survival is reported and the reproductive output of exposed parents living at the end of the test (expressed in a
number of terms) is compared to those of the control parents.
4 Test environment
The test atmosphere shall be free from vapours or dusts which may be toxic to Daphnia magna Straus.
The exposure solutions shall not be aerated.
The dissolved oxygen concentration in the exposure solutions shall be above 3 mg/l and the pH shall be within the
range of pH 6 to pH 9 and not change by more than 1,5 pH units for the duration of the test. Hardness shall be
above 140 mg/l (as CaCO ), which has been demonstrated to be necessary to promote reproductive performance
that shall meet the validity criteria (see 8.7).
The photoperiod of the test shall be 16 h of light and 8 h of darkness. The intensity shall be in the range 600 lux to
800 lux but not exceeding 1 200 lux.
The temperature of the test exposures shall be maintained within the range of 18 °C to 22 °C and the temperature
for the test shall not vary by more than 2 °C (for instance 18 °C to 20 °C, 19 °C to 21 °C or 20 °C to 22 °C)
throughout the test.
5 Reagents and materials
5.1 Test organism: Daphnia magna Straus (Cladocera, Crustacea), hereafter referred to as D. magna, obtained
by acyclical parthenogenesis for at least three generations under specified culture conditions.
The age and the source (including clone, if possible) of the D. magna culture shall be indicated in the test report,
since the sensitivity of D. magna to toxicants can be affected by the source of the culture.
The animals used for the test shall be less than 24 h old and be from the second to fifth brood. The D. magna shall
be from a healthy stock showing no signs of stress such as mortality � 20 %, presence of males, ephippia, or
discoloured animals and there shall be no delay in the production of the first brood.
The stock animals shall be maintained in culture conditions (light, temperature, medium, feeding and animals per
unit volume) similar to those in the test. If the culture conditions differ from test conditions, it is recommended that
one generation be acclimated under the test conditions for about three weeks so as to avoid stressing the parent
animals.
5.2 Dilution water, synthetic as described in 5.3 or uncontaminated natural waters with comparable pH and
hardness characteristics to these dilution waters for culturing and testing.
5.3 Synthetic dilution water, prepared using one of the following methods:
a) OECD M4 and M7 media (see informative annex A);
b) ASTM reconstituted hard freshwater (see informative annex B).
The dilution water shall be aerated until the dissolved oxygen concentration has reached 95 % saturation and the
pH has stabilized. If necessary, adjust the pH to 8,0� 0,5 by adding sodium hydroxide (NaOH) solution or
hydrochloric acid (HCl) solution. The dilution water prepared in this way shall not be further aerated before use.
It is recommended that the TOC levels be� 5 mg/l before the addition of algae. The use of dilution water containing
EDTA (for instance M4 and M7) is not recommended when testing compounds containing metals because
chelation may reduce the toxicity of the compound.
If the test has to be performed for purposes necessitating the use of a dilution water with characteristics differing
from those described above, mention is necessary in the test report of the main characteristics (for instance pH,
hardness, TOC, COD) of the synthetic dilution water used.
2 © ISO 2000 – All rights reserved
...
INTERNATIONAL ISO
STANDARD 10706
First edition
2000-04-01
Water quality — Determination of long term
toxicity of substances to Daphnia magna
Straus (Cladocera, Crustacea)
Qualité de l'eau — Détermination de la toxicité à long terme de substances
vis-à-vis de Daphnia magna Straus (Cladocera, Crustacea)
Reference number
©
ISO 2000
PDF disclaimer
This PDF file may contain embedded typefaces. In accordance with Adobe's licensing policy, this file may be printed or viewed but shall not
be edited unless the typefaces which are embedded are licensed to and installed on the computer performing the editing. In downloading this
file, parties accept therein the responsibility of not infringing Adobe's licensing policy. The ISO Central Secretariat accepts no liability in this
area.
Adobe is a trademark of Adobe Systems Incorporated.
Details of the software products used to create this PDF file can be found in the General Info relative to the file; the PDF-creation parameters
were optimized for printing. Every care has been taken to ensure that the file is suitable for use by ISO member bodies. In the unlikely event
that a problem relating to it is found, please inform the Central Secretariat at the address given below.
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means, electronic
or mechanical, including photocopying and microfilm, without permission in writing from either ISO at the address below or ISO's member body
in the country of the requester.
ISO copyright office
Case postale 56 � CH-1211 Geneva 20
Tel. + 41 22 749 01 11
Fax + 41 22 734 10 79
E-mail copyright@iso.ch
Web www.iso.ch
Printed in Switzerland
ii © ISO 2000 – All rights reserved
Contents Page
Foreword.iv
Introduction.v
1 Scope .1
2 Normative references .1
3 Principle.1
4 Test environment.1
5 Reagents and materials .2
6 Apparatus .3
7 Treatment and preparation of samples .3
7.1 Special precautions for sampling and transportation of water or effluent samples.3
7.2 Preparation of solutions of substances to be tested.3
8 Procedure .4
8.1 Controls .4
8.2 Selection of exposure concentrations .4
8.3 Renewal of exposure solutions.5
8.4 Introduction of animals into the test system .5
8.5 Feeding of organisms .5
8.6 Observations and measurements.5
8.7 Validity of the test.6
9 Data analysis and expression of results .6
10 Test report .7
Annex A (informative) OECD M4 and M7 media — Preparation and acclimation of Elendt M4 and M7
media.9
Annex B (informative) ASTM reconstituted hard fresh water.12
Annex C (informative) Data collection sheet .13
Annex D (informative) Calculation of time-weighted mean concentration.15
Bibliography.17
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 3.
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 International Standard may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights.
International Standard ISO 10706 was prepared by Technical Committee ISO/TC 147, Water quality,
Subcommittee SC 5, Biological methods.
Annexes A, B, C and D of this International Standard are for information only.
iv © ISO 2000 – All rights reserved
Introduction
This International Standard defines a procedure for the determination of the long term sublethal toxicity of
chemicals, waters and waste waters to the water-flea Daphnia magna Straus. The methodology is adapted from a
guideline produced by the Organisation for Economic Co-operation and Development (OECD Guideline 211, see
reference [1] in the Bibliography).
INTERNATIONAL STANDARD ISO 10706:2000(E)
Water quality — Determination of long term toxicity of substances
to Daphnia magna Straus (Cladocera, Crustacea)
WARNING — Activated sludge and sewage contain potentially pathogenic organisms. Therefore,
appropriate precautions should be taken when handling them. Toxic test compounds and those whose
properties are unknown should be handled with care.
1 Scope
This International Standard describes a method for the determination of the long term sublethal toxicity to Daphnia
magna Straus (Cladocera, Crustacea)of:
a) chemical substances, which are soluble under the conditions of the test, or can be maintained as stable
suspensions or dispersions under the conditions of the test,
b) industrial or sewage effluents, treated or untreated, after decanting, filtration, or centrifugation,
c) surface or ground waters.
[1]
NOTE This International Standard is adapted from OECD Guideline 211 .
2 Normative references
The following normative documents contain provisions which, through reference in this text, constitute provisions of
this International Standard. For dated references, subsequent amendments to, or revisions of, any of these
publications do not apply. However, parties to agreements based on this International Standard are encouraged to
investigate the possibility of applying the most recent editions of the normative documents indicated below. For
undated references, the latest edition of the normative document referred to applies. Members of ISO and IEC
maintain registers of currently valid International Standards.
ISO 5667-2, Water quality — Sampling — Part 2: Guidance on sampling techniques.
ISO 5667-16, Water quality — Sampling — Part 16: Guidance on biotesting of samples.
3Principle
Daphnia magna females, less than 24 h old, are exposed for a period of 21 days to a test substance, industrial or
sewage effluent or surface/ground water added to dilution water in a range of concentrations in a semi-static or
flow-through system. The survival of parents is recorded together with the number of live offspring produced per
live parent at the end of the test.
The survival is reported and the reproductive output of exposed parents living at the end of the test (expressed in a
number of terms) is compared to those of the control parents.
4 Test environment
The test atmosphere shall be free from vapours or dusts which may be toxic to Daphnia magna Straus.
The exposure solutions shall not be aerated.
The dissolved oxygen concentration in the exposure solutions shall be above 3 mg/l and the pH shall be within the
range of pH 6 to pH 9 and not change by more than 1,5 pH units for the duration of the test. Hardness shall be
above 140 mg/l (as CaCO ), which has been demonstrated to be necessary to promote reproductive performance
that shall meet the validity criteria (see 8.7).
The photoperiod of the test shall be 16 h of light and 8 h of darkness. The intensity shall be in the range 600 lux to
800 lux but not exceeding 1 200 lux.
The temperature of the test exposures shall be maintained within the range of 18 °C to 22 °C and the temperature
for the test shall not vary by more than 2 °C (for instance 18 °C to 20 °C, 19 °C to 21 °C or 20 °C to 22 °C)
throughout the test.
5 Reagents and materials
5.1 Test organism: Daphnia magna Straus (Cladocera, Crustacea), hereafter referred to as D. magna, obtained
by acyclical parthenogenesis for at least three generations under specified culture conditions.
The age and the source (including clone, if possible) of the D. magna culture shall be indicated in the test report,
since the sensitivity of D. magna to toxicants can be affected by the source of the culture.
The animals used for the test shall be less than 24 h old and be from the second to fifth brood. The D. magna shall
be from a healthy stock showing no signs of stress such as mortality � 20 %, presence of males, ephippia, or
discoloured animals and there shall be no delay in the production of the first brood.
The stock animals shall be maintained in culture conditions (light, temperature, medium, feeding and animals per
unit volume) similar to those in the test. If the culture conditions differ from test conditions, it is recommended that
one generation be acclimated under the test conditions for about three weeks so as to avoid stressing the parent
animals.
5.2 Dilution water, synthetic as described in 5.3 or uncontaminated natural waters with comparable pH and
hardness characteristics to these dilution waters for culturing and testing.
5.3 Synthetic dilution water, prepared using one of the following methods:
a) OECD M4 and M7 media (see informative annex A);
b) ASTM reconstituted hard freshwater (see informative annex B).
The dilution water shall be aerated until the dissolved oxygen concentration has reached 95 % saturation and the
pH has stabilized. If necessary, adjust the pH to 8,0� 0,5 by adding sodium hydroxide (NaOH) solution or
hydrochloric acid (HCl) solution. The dilution water prepared in this way shall not be further aerated before use.
It is recommended that the TOC levels be� 5 mg/l before the addition of algae. The use of dilution water containing
EDTA (for instance M4 and M7) is not recommended when testing compounds containing metals because
chelation may reduce the toxicity of the compound.
If the test has to be performed for purposes necessitating the use of a dilution water with characteristics differing
from those described above, mention is necessary in the test report of the main characteristics (for instance pH,
hardness, TOC, COD) of the synthetic dilution water used.
2 © ISO 2000 – All rights reserved
6 Apparatus
Ordinary laboratory apparatus and in particular the following.
6.1 Environmental controls, for controlling temperature, photoperiod and light intensity.
6.2 Measuring apparatus and/or instruments, for measuring dissolved oxygen, pH, hardness, total organic
carbon, chemical oxygen demand, light intensity and temperature.
6.3 Test containers, of chemically inert material and of sufficient capacity for the tests (for example 50 ml or
100 ml glass test tubes or beakers) and which are clean and uncontaminated.
7 Treatment and preparation of samples
7.1 Special precautions for sampling and transportation of water or effluent samples
Sampling of water or effluent shall be carried out in accordance with the general procedure specified in ISO 5667-2.
Bottles shall be completely filled to exclude air.
The preservation and storage of water or effluent samples shall be carried out in accordance with ISO 5667-16; the
following is only a summary. Carry out the toxicity test as
...
NORME ISO
INTERNATIONALE 10706
Première édition
2000-04-01
Qualité de l'eau — Détermination de la
toxicité à long terme de substances
vis-à-vis de Daphnia magna Straus
(Cladocera, Crustacea)
Water quality — Determination of long term toxicity of substances to
Daphnia magna Straus (Cladocera, Crustacea)
Numéro de référence
©
ISO 2000
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Fax. + 41 22 749 09 47
E-mail copyright@iso.ch
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Imprimé en Suisse
ii © ISO 2000 – Tous droits réservés
Sommaire Page
Avant-propos.iv
Introduction.v
1 Domaine d'application.1
2Références normatives .1
3 Principe.1
4 Environnement de l'essai.2
5Réactifs et produits .2
6 Appareillage .3
7 Traitement et préparation des échantillons .3
7.1 Précautions particulières concernant le prélèvement et le transport des échantillons d'eaux ou
d'effluents.3
7.2 Préparation des solutions des substances à expérimenter.3
8 Mode opératoire.4
8.1 Témoins .4
8.2 Sélection des concentrations d'essai.4
8.3 Renouvellement des concentrations d'essai.5
8.4 Introduction des animaux dans le système d'essai.5
8.5 Alimentation des organismes.5
8.6 Observations et mesures.6
8.7 Validité des résultats.6
9 Analyse des données et expression des résultats .7
10 Rapport d'essai .8
Annexe A (informative) Milieux M4 et M7 de l'OCDE — Préparation et acclimatation des milieux M4
Elendt et M7.10
Annexe B (informative) Eau dure reconstituée ASTM.13
Annexe C (informative) Fiche de collecte de données.14
Annexe D (informative) Calcul de la concentration moyenne pondérée dans le temps.16
Bibliographie .18
Avant-propos
L'ISO (Organisation internationale de normalisation) est une fédération mondiale d'organismes nationaux de
normalisation (comités membres de l'ISO). L'élaboration des Normes internationales est en général confiéeaux
comités techniques de l'ISO. Chaque comité membre intéressé par une étude aledroit de fairepartie ducomité
technique créé à cet effet. Les organisations internationales, gouvernementales et non gouvernementales, en
liaison avec l'ISO participent également aux travaux. L'ISO collabore étroitement avec la Commission
électrotechnique internationale (CEI) en ce qui concerne la normalisation électrotechnique.
Les Normes internationales sont rédigées conformément aux règles données dans les Directives ISO/CEI, Partie 3.
Les projets de Normes internationales adoptés par les comités techniques sont soumis aux comités membres pour
vote. Leur publication comme Normes internationales requiert l'approbation de 75 % au moins des comités
membres votants.
L’attention est appelée sur le fait que certains des éléments delaprésente Norme internationale peuvent faire
l’objet de droits de propriété intellectuelle ou de droits analogues. L’ISO ne saurait être tenue pour responsable de
ne pas avoir identifié de tels droits de propriété et averti de leur existence.
La Norme internationale ISO 10706 a étéélaboréepar le comité technique ISO/TC 147, Qualité de l'eau,
sous-comité SC 5, Méthodes biologiques.
Les annexes A, B, C et D de la présente Norme internationale sont données uniquement à titre d’information.
iv © ISO 2000 – Tous droits réservés
Introduction
La présente Norme internationale définit une méthode pour la détermination de la toxicité sublétale à long terme
des substances chimiques, des eaux et des effluents vis-à-vis de la puce d’eau Daphnia magna Straus. La
méthodologie est adaptéed’une ligne directrice élaborée par l’Organisation de coopération et de développement
économiques (Ligne directrice 211 de l’OCDE, voir référence [1] de la Bibliographie).
NORME INTERNATIONALE ISO 10706:2000(F)
Qualité de l'eau — Détermination de la toxicitéà long terme de
substances vis-à-vis de Daphnia magna Straus (Cladocera,
Crustacea)
AVERTISSEMENT — Les boues activées et les eaux usées peuvent contenir des organismes
potentiellement pathogènes. Il est par conséquent recommandé de prendre les précautions d'usage lors de
leur manipulation. Il convient de manipuler avec soin les composés d'essai toxiques et ceux dont les
propriétés sont inconnues.
1 Domaine d'application
La présente Norme internationale décrit une méthodepour ladétermination de la toxicité sublétale à long terme
vis-à-vis de Daphnia magna Straus (Cladocera, Crustacea) des substances suivantes:
a) substances chimiques solubles dans les conditions de l'essai ou pouvant être maintenues en suspension ou
en dispersion stable dans les conditions de l'essai;
b) effluents industriels ou urbains, épurés ou non, s'il y a lieu aprèsdécantation, filtration ou centrifugation;
c) eaux de surface ou eaux souterraines.
NOTE La présente Norme internationale est adaptéedelaLigne directrice 211del'OCDE[1].
2Références normatives
Les documents normatifs suivants contiennent des dispositions qui, par suite de la référence qui y est faite,
constituent des dispositions valables pour la présente Norme internationale. Pour les références datées, les
amendements ultérieurs ou les révisions de ces publications ne s’appliquent pas. Toutefois, les parties prenantes
aux accords fondés sur la présente Norme internationale sont invitées à rechercher la possibilité d'appliquer les
éditions les plus récentes des documents normatifs indiqués ci-après. Pour les références non datées, la dernière
édition du document normatif en référence s’applique. Les membres de l'ISO et de la CEI possèdent le registre des
Normes internationales en vigueur.
ISO 5667-2, Qualité de l’eau —Échantillonnage — Partie 2: Guide général sur les techniques d’échantillonnage.
ISO 5667-16, Qualité de l’eau —Échantillonnage — Partie 16: Lignes directrices pour les essais biologiques des
échantillons.
3Principe
Exposition pendant 21 jours de Daphnia magna femelles âgées de moins de 24 heures à une substance d'essai, à
des effluents industriels ou urbains ou à des eaux de surface/souterraines ajoutés à de l'eau de dilution, dans une
gamme de concentrations d'un système semi-statique ou à renouvellement continu. Le nombre de parents
survivants ainsi que le nombre de descendants vivants issus de parents vivants à lafindel'essaisontconsignés.
La survie est notée et la capacité reproductrice des parents mis en expérimentation, vivants en fin d’essai
(exprimée en nombre d'individus) est comparée à celle des parents témoins.
4 Environnement de l'essai
L'enceinte d'essai doit être exempte de vapeurs ou de poussièrestoxiquesvis-à-vis de Daphnia magna Straus.
Les solutions d'essai ne doivent pas être aérées.
La concentration en oxygène dissous dans les solutions d'essai doit être supérieure à 3 mg/l et le pH doit se situer
dans la gamme comprise entre 6 et 9 et ne doit pas varier de plus de 1,5 unités au cours de l'essai. La dureté doit
avoir une valeur supérieure à 140 mg/l (expriméeenCaCO)ayant démontré qu'elle favorise la capacité
reproductrice pour êtreconformeaux critères de validité (voir 8.7).
Le cycle d'essai doit être de 16 h de lumière et de 8 h d’obscurité.L’intensité doit de préférence être comprise
entre 600 lux et 800 lux mais ne doit pas excéder 1 200 lux.
La température des solutions d'essai doit être maintenue dans une gamme comprise entre 18 °Cet 22 °Cet ne
pas varier de plus de 2 °C (par exemple 18 °C à 20 °C, 19 °C à 21 °Cou20 °C à 22 °C) pendant toute la duréede
l'essai.
5Réactifs et produits
5.1 Organismes pour essai: Daphnia magna Straus (Cladocera, Crustacea), notée D. magna dans la suite du
texte, de troisième génération au moins, obtenue par parthénogenèse acyclique dans des conditions d'élevage
définies.
La sensibilité de D. magna aux toxiques étant fonction de l'origine de l'élevage, l'âge et la provenance (y compris
les clones, si possible) de l'élevage de D. magna doivent être mentionnés dans le rapport d'essai.
Les animaux utilisés pour l'essai doivent être âgés de moins de 24 h et provenir de la seconde à la cinquième
portée. Les D. magna doivent provenir d’un lot sain ne présentant aucun signe de stress tel que mortalité >20%,
présencedemâles, ephippia ou animaux décolorés, et doivent produire sans retard la première génération.
Le lot d’animaux doit être maintenu dans des conditions d'élevage (lumière, température, milieu, alimentation et
nombre par unité de volume) similaires à celles de l'essai. Si les conditions d'élevage diffèrent de celles de l'essai,
il est recommandé d'acclimater une génération aux conditions d'essai pendant environ trois semaines afin d'éviter
toute source de stress aux animaux parents.
5.2 Eau de dilution: eau synthétique comme décrit en 5.3 ou des eaux naturelles non contaminées ayant un pH
et des caractéristiques de dureté similaires aux eaux de dilution utilisées pour l'élevage et les essais.
5.3 Eau de dilution synthétique,préparée au moyen de l'une des méthodes suivantes:
a) milieux M4 et M7 de l’OCDE (voir annexe A);
b) eau de source dure reconstituée ASTM (voir annexe B).
L'eau de dilution doit être aérée jusqu'à ce que la concentration en oxygène dissous ait atteint la valeur de
saturation de 95 % et jusqu'à stabilisation du pH. Si nécessaire, ajuster le pH à 8,0� 0,5 par ajout d'une solution
d'hydroxyde de sodium (NaOH) ou d'acide chlorhydrique (HCI). L'eau de dilution ainsi préparéenedoit plus être
aérée avant emploi.
Il est recommandé que les niveaux de COT (carbone organique total) soient < 5 mg/l avant l'ajout d'algues. Il n'est
pas recommandé d'utiliser de l'eau de dilution contenant de l'EDTA (par exemple M7 et M4) pour analyser des
composés contenant des métaux dans la mesure où la chélation peut réduire la toxicité du composé.
Si l'essai doit être conduit en vue de besoins qui impliquent l'utilisation d'une eau de dilution dont les
caractéristiques diffèrent de celles décrites ci-dessus, il convient de mentionner dans le rapport d'essai les
principales caracté
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