EN ISO 5667-16:2017

Water quality - Sampling - Part 16: Guidance on biotesting of samples (ISO 5667-16:2017)

ISO 5667-16:2017 gives practical guidance on sampling, pre-treatment, performance and evaluation of environmental samples in the context of performing biological tests. Information is given on how to cope with the problems of biotesting arising from the sample and the suitability of the test design.
It is intended to convey practical experience concerning precautions to be taken by describing methods successfully proven to solve or to circumvent some of the experimental problems of biotesting of, for example, waters.
Primarily dealt with are substance-related problems concerning sampling and pre-treatment of environmental samples (e.g. waste water samples) for the performance of biotests.
This guidance is on ecotoxicological testing with organisms (single-species biotests; in vivo and in vitro). Some features addressed in this document also apply to biotests using single-cell systems (in vitro bioassays) and biodegradation studies as far as sampling and sample preparations are concerned. Testing of substances in the water solubility range is also addressed.
Reference has been made as far as possible to existing International Standards and guidelines. Information taken from published papers or oral communication has been utilized as well.
ISO 5667-16:2017 is applicable to biological tests for determining the effect of environmental samples like treated communal and industrial waste water, groundwater, fresh water, aqueous extracts (e.g. leachates, eluates), pore water of sediments and whole sediments. This document is also applicable to chemical substances.
ISO 5667-16:2017 is not applicable to bacteriological examination of water. Appropriate methods for bacteriological examination are described in other documents (see ISO 19458[17]).

Wasserbeschaffenheit - Probenahme - Teil 16: Anleitung zur Probenahme und Durchführung biologischer Testverfahren (ISO 5667-16:2017)

Dieses Dokument gibt praktische Hinweise zur Probenahme, Vorbehandlung, Durchführung und Bewertung von Umweltproben in Verbindung mit der Durchführung von biologischen Testverfahren. Es werden Informationen gegeben, wie bei biologischen Testverfahren mit Problemen umzugehen ist, die das Probenmaterial und das Testdesign mit sich bringen.
Es ist das Ziel, praktische Erfahrung hinsichtlich zu treffender Vorkehrungen zu vermitteln, indem erfolgreich bewährte Verfahren beschrieben werden, um einige der experimentellen Probleme bei der Untersuchung von z. B. Wässern mit biologischen Testverfahren zu lösen oder zu umgehen.
Vor allem werden substanzbezogene Probleme in Bezug auf die Probenahme und Vorbehandlung von Umweltproben (z. B. Abwasserproben) bei der Durchführung von Biotests berücksichtigt.
Diese Anleitung behandelt ökotoxikologische Testverfahren mit Organismen (Tests mit jeweils einer Art; in vivo und in vitro). Einige Ausführungen in diesem Dokument beziehen sich auch auf Biotests, die einzellige Mikroorganismen (in-vitro Biotests) verwenden, und auf Untersuchungen zum biologischen Abbau, sofern die Probenahme und die Probenvorbereitung betroffen sind. Ebenso wird auf die Untersuchung von Substanzen im Bereich ihrer Wasserlöslichkeit eingegangen.
Soweit möglich, wird auf bestehende Internationale Normen und Anleitungen verwiesen. Die Informationen aus Veröffentlichungen und mündliche Mitteilungen sind ebenfalls berücksichtigt worden.
Dieses Dokument gilt für biologische Testverfahren zur Bestimmung der Wirkung von Umweltproben, wie behandeltes kommunales und Industrieabwasser, Grundwasser, Süßwasser, wässrige Extrakte (z. B. Sickerwasser, Eluate), Porenwasser von Sedimenten und Gesamtsedimente. Dieses Dokument ist auch auf chemische Substanzen übertragbar.
Dieses Dokument ist nicht anwendbar auf die bakteriologische Untersuchung von Wasser. Entsprechende Verfahren sind in anderen Dokumenten beschrieben (siehe ISO 19458) [17].

Qualité de l'eau - Échantillonnage - Partie 16: Lignes directrices pour les essais biologiques des échantillons (ISO 5667-16:2017)

ISO 5667-16:2017 fournit des lignes directrices pour l'échantillonnage, le prétraitement, la mise en ?uvre et l'évaluation des échantillons environnementaux dans le cadre de la réalisation d'essais biologiques. Des informations sont données sur la manière de traiter les problèmes, pour les essais biologiques, liés à l'échantillon et à l'adaptation du dispositif expérimental.
Elles sont destinées à diffuser une expérience pratique concernant les précautions à prendre en décrivant des méthodes qui se sont révélées satisfaisantes pour résoudre ou éliminer certains problèmes expérimentaux au cours des essais biologiques des eaux, par exemple.
ISO 5667-16:2017 traite en premier lieu des problèmes liés aux substances en ce qui concerne l'échantillonnage et le prétraitement des échantillons environnementaux (par exemple échantillons d'eaux résiduaires) pour la réalisation d'essais biologiques.
Ces lignes directrices portent sur les essais écotoxicologiques réalisés avec des organismes (essais biologiques monospécifiques; in vivo et in vitro). Certaines caractéristiques traitées dans le présent document s'appliquent également à l'échantillonnage et la préparation d'échantillons pour les essais biologiques utilisant des systèmes unicellulaires (essais biologiques in vitro) et les études de biodégradation. Les essais sur substances dans la gamme de solubilité dans l'eau sont également traités.
Il a été fait référence aux Normes internationales et aux directives existantes dans toute la mesure du possible. Des informations provenant de publications ou de communications orales ont également été utilisées.
ISO 5667-16:2017 s'applique aux essais biologiques destinés à déterminer l'effet des échantillons environnementaux tels que les eaux résiduaires communales et industrielles traitées, les eaux souterraines, les eaux douces, les extraits aqueux (par exemple lixiviats et éluats), l'eau interstitielle des sédiments et les sédiments entiers. Il s'applique également aux substances chimiques.
ISO 5667-16:2017 n'est pas applicable à l'examen bactériologique des eaux. Des méthodes appropriées pour l'examen bactériologique sont décrites dans d'autres documents (voir l'ISO 19458[17]).

Kakovost vode - Vzorčenje - 16. del: Navodilo za biološko preskušanje vzorcev (ISO 5667-16:2017)

Ta dokument podaja praktične smernice glede vzorčenja, predhodne obdelave, zmogljivosti in ocenjevanja okoljskih vzorcev za biološke preskuse. Vsebuje informacije o tem, kako se soočiti s težavami biološkega preskušanja, ki so posledica vzorca, in o primernosti zasnove preskusa.
Namenjen je podajanju praktičnih izkušenj v zvezi s previdnostnimi ukrepi, ki jih je treba izvesti, in sicer z opisovanjem metod, ki so se izkazale za uspešne pri reševanju ali preprečevanju nekaterih poskusnih težav, na primer pri biološkem preskušanju vod.
Zajema predvsem težave zaradi snovi, ki so povezane z vzorčenjem in predhodno obdelavo okoljskih vzorcev (npr. vzorci odpadne vode) za biološko preskušanje.
Smernice podajajo informacije o ekotoksikološkem preskušanju z organizmi (biološko preskušanje posamezne vrste; in vivo in in vitro). Nekatere značilnosti, obravnavane v tem dokumentu, veljajo tudi za biološke preskuse, ki uporabljajo enocelične sisteme (biološki preskusi in vitro) in študije biorazgradljivosti v povezavi z vzorčenjem in pripravo vzorcev.
Obravnavano je tudi preskušanje topnosti snovi v vodi.
Dodani so bili sklici na obstoječe mednarodne standarde in smernice, in sicer v največjem možnem obsegu.
Uporabljene so bile tudi informacije iz objavljenih študij ali ustne komunikacije. Ta dokument se uporablja za biološko preskušanje za določanje učinka okoljskih vzorcev, kot so obdelana javna in industrijska odpadna voda, podtalnica, sveža voda, vodni ekstrakti (npr. izcedne vode, izlužki), voda v porah sedimentov in celotnih sedimentov. Ta dokument se uporablja tudi za kemične snovi.
Ta dokument se ne uporablja za bakteriološko preskušanje vode. Metode, ki so primerne za bakteriološko preskušanje, so opisane v drugih dokumentih (glej ISO 19458[17]).

General Information

• Status: Published
• Publication Date: 30-Aug-2017
• Withdrawal Date: 29-Nov-2017
• ICS: 13.060.10 - Water of natural resources
• ICS: 13.060.45 - Examination of water in general
• ICS: 13.060.70 - Examination of biological properties of water
• Technical Committee: CEN/TC 230 - Water analysis
• Drafting Committee: CEN/TC 230 - Water analysis
• Current Stage: 6060 - Definitive text made available (DAV) - Publishing
• Start Date: 10-May-2017
• Completion Date: 10-May-2017
• Ref Project: SIST EN ISO 5667-16:2017 - Water quality - Sampling - Part 16: Guidance on biotesting of samples (ISO 5667-16:2017)

Relations

• Replaces: EN ISO 5667-16:1998 - Water quality - Sampling - Part 16: Guidance on biotesting of samples (ISO 5667-16:1998)
• Effective Date: 17-May-2017

Standards Content (Sample)

SLOVENSKI STANDARD
SIST EN ISO 5667-16:2017
01-julij-2017
1DGRPHãþD
SIST EN ISO 5667-16:2000
.DNRYRVWYRGH9]RUþHQMHGHO1DYRGLOR]DELRORãNRSUHVNXãDQMHY]RUFHY
,62
Water quality - Sampling - Part 16: Guidance on biotesting of samples (ISO 5667-
16:2017)
Wasserbeschaffenheit - Probenahme - Teil 16: Anleitung zur Probenahme und
Durchführung biologischer Testverfahren (ISO 5667-16:2017)

Qualité de l'eau - Échantillonnage - Partie 16: Lignes directrices pour les essais

biologiques des échantillons (ISO 5667-16:2017)
Ta slovenski standard je istoveten z: EN ISO 5667-16:2017
ICS:
13.060.70 Preiskava bioloških lastnosti Examination of biological
vode properties of water
SIST EN ISO 5667-16:2017 en,fr,de

2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

---------------------- Page: 1 ----------------------
SIST EN ISO 5667-16:2017
---------------------- Page: 2 ----------------------
SIST EN ISO 5667-16:2017
EN ISO 5667-16
EUROPEAN STANDARD
NORME EUROPÉENNE
May 2017
EUROPÄISCHE NORM
ICS 13.060.45 Supersedes EN ISO 5667-16:1998
English Version
Water quality - Sampling - Part 16: Guidance on biotesting
of samples (ISO 5667-16:2017)

Qualité de l'eau - Échantillonnage - Partie 16: Lignes Wasserbeschaffenheit - Probenahme - Teil 16:

directrices pour les essais biologiques des échantillons Anleitung zur Probenahme und Durchführung

(ISO 5667-16:2017) biologischer Testverfahren (ISO 5667-16:2017)
This European Standard was approved by CEN on 9 February 2017.

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: Avenue Marnix 17, B-1000 Brussels

© 2017 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 5667-16:2017 E

worldwide for CEN national Members.
---------------------- Page: 3 ----------------------
SIST EN ISO 5667-16:2017
EN ISO 5667-16:2017 (E)
Contents Page

European foreword ....................................................................................................................................................... 3

---------------------- Page: 4 ----------------------
SIST EN ISO 5667-16:2017
EN ISO 5667-16:2017 (E)
European foreword

This document (EN ISO 5667-16:2017) has been prepared by Technical Committee ISO/TC 147 “Water

quality” in collaboration with 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 November 2017, and conflicting national standards

shall be withdrawn at the latest by November 2017.

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.
This document supersedes EN ISO 5667-16:1998.

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 5667-16:2017 has been approved by CEN as EN ISO 5667-16:2017 without any

modification.
---------------------- Page: 5 ----------------------
SIST EN ISO 5667-16:2017
---------------------- Page: 6 ----------------------
SIST EN ISO 5667-16:2017
INTERNATIONAL ISO
STANDARD 5667-16
Second edition
2017-04
Water quality — Sampling —
Part 16:
Guidance on biotesting of samples
Qualité de l’eau — Échantillonnage —
Partie 16: Lignes directrices pour les essais biologiques des
échantillons
Reference number
ISO 5667-16:2017(E)
ISO 2017
---------------------- Page: 7 ----------------------
SIST EN ISO 5667-16:2017
ISO 5667-16:2017(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2017, Published in Switzerland

All rights reserved. Unless otherwise specified, 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
Ch. de Blandonnet 8 • CP 401
CH-1214 Vernier, Geneva, Switzerland
Tel. [phone]
Fax [phone]
[e-mail]
www.iso.org
ii © ISO 2017 – All rights reserved
---------------------- Page: 8 ----------------------
SIST EN ISO 5667-16:2017
ISO 5667-16:2017(E)
Contents Page

Foreword ..........................................................................................................................................................................................................................................v

Introduction ................................................................................................................................................................................................................................vi

1 Scope ................................................................................................................................................................................................................................. 1

2 Normative references ...................................................................................................................................................................................... 1

3 Terms and definitions ..................................................................................................................................................................................... 1

4 General guidance regarding test design ....................................................................................................................................... 5

4.1 General ........................................................................................................................................................................................................... 5

4.2 Replicates ..................................................................................................................................................................................................... 5

4.2.1 General...................................................................................................................................................................................... 5

4.2.2 Lowest ineffective dilution (LID) ....................................................................................................................... 5

4.2.3 Hypothesis testing — two-sample comparisons ................................................................................. 6

4.2.4 Concentration and dilution response relationship ............................................................................ 6

5 Evaluation .................................................................................................................................................................................................................... 7

5.1 General ........................................................................................................................................................................................................... 7

5.2 Statistical analysis ................................................................................................................................................................................ 7

6 Sampling and transportation .................................................................................................................................................................. 7

6.1 General ........................................................................................................................................................................................................... 7

6.2 Sampling equipment .......................................................................................................................................................................... 8

6.2.1 General...................................................................................................................................................................................... 8

6.2.2 Sample container ............................................................................................................................................................. 8

6.3 Filling status of sample containers ........................................................................................................................................ 9

6.4 Sample identification and records ......................................................................................................................................... 9

6.5 Sub-sampling ............................................................................................................................................................................................ 9

6.6 Transportation .....................................................................................................................................................................................10

6.7 Contamination during sampling ...........................................................................................................................................10

6.8 Sampling quality control techniques ................................................................................................................................10

7 Pre-treatment .......................................................................................................................................................................................................11

7.1 General ........................................................................................................................................................................................................11

7.2 Preservation and storage ............................................................................................................................................................11

7.3 Thawing .....................................................................................................................................................................................................12

7.4 Homogenization ..................................................................................................................................................................................12

7.5 Separation of soluble and particulate matter ............................................................................................................12

7.6 Preconcentration ................................................................................................................................................................................13

7.6.1 General...................................................................................................................................................................................13

7.6.2 Extraction methods ....................................................................................................................................................14

7.7 pH adjustment ......................................................................................................................................................................................14

8 Apparatus and equipment .......................................................................................................................................................................14

8.1 Selection of apparatus ...................................................................................................................................................................14

8.2 Cleaning of apparatus and equipment .............................................................................................................................15

9 Impairment of test performance ......................................................................................................................................................15

9.1 Problems and preventive measures for samples containing removable ingredients .............15

9.1.1 General...................................................................................................................................................................................15

9.1.2 Volatilization ...................................................................... ...............................................................................................16

9.1.3 Foaming ................................................................................................................................................................................16

9.1.4 Adsorption .................. .................................................... ....................................................................................................16

9.1.5 Precipitation/flocculation ....................................................................................................................................16

9.1.6 Degradation .......................................................................................................................................................................16

9.2 Problems and preventive measures concerning coloured and/or turbid samples ..................17

10 Preparation of stock solutions and test batches ..............................................................................................................17

10.1 Water-soluble substances ..........................................................................................................................................................17

© ISO 2017 – All rights reserved iii
---------------------- Page: 9 ----------------------
SIST EN ISO 5667-16:2017
ISO 5667-16:2017(E)

10.2 Poorly soluble substances ..........................................................................................................................................................17

10.2.1 General...................................................................................................................................................................................17

10.2.2 Testing in the water solubility range ...........................................................................................................17

10.2.3 Dispersions and emulsions ..................................................................................................................................18

10.2.4 Special problems with mixtures of substances or technical products ...........................18

10.2.5 Limit test ..............................................................................................................................................................................19

11 Quality assurance for biotesting .......................................................................................................................................................19

11.1 General ........................................................................................................................................................................................................19

11.2 Quality assurance in the context of the investigation of environmental samples .....................19

12 Reporting ...................................................................................................................................................................................................................20

Bibliography .............................................................................................................................................................................................................................23

iv © ISO 2017 – All rights reserved
---------------------- Page: 10 ----------------------
SIST EN ISO 5667-16:2017
ISO 5667-16:2017(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 on 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 the following

URL: w w w . i s o .org/ iso/ foreword .html.

This document was prepared by ISO/TC 147, Water quality, Subcommittee SC 6, Sampling (general

methods).

This second edition cancels and replaces the first edition (ISO 5667-16:1998), which has been technically

revised.
A list of all parts in the ISO 5667 series can be found on the ISO website.
© ISO 2017 – All rights reserved v
---------------------- Page: 11 ----------------------
SIST EN ISO 5667-16:2017
ISO 5667-16:2017(E)
Introduction

Biological tests are suitable for determining the effect of environmental samples or chemical substances

on the respective test organism under the specific standardized test conditions. Environmental samples

are e.g. treated communal and industrial waste water, fresh water, aqueous extracts of solid material

(e.g. leachates, eluates), pore water of sediments. The effect can be stimulative or inhibiting, and can be

determined by the reaction of the test organism (e.g. death, growth, morphological and physiological

changes or generally, changes in molecular mechanisms of action). Inhibiting effects can be triggered

by toxic water constituents or by other noxious influences.

The toxicity measurable in the biological test is the result of the interaction between a single toxic

substance, a mixture of substances or the constituents of an environmental sample and the test

organism. The protective potential of the biological system, i.e. the test organism, for instance by

metabolic detoxification and excretion, is an integral part of the biological test.

Apart from the direct toxic effect of one or more sample constituents, biological effects can be exerted

by the combined action of all constituents of a sample. Such a combined effect includes the impact of, for

example, substances which are not toxic per se but affect the chemical or physical properties of the test

batches by interfering with the test specific additives (e.g. nutrients, salts) and, consequently, the living

conditions for the test organisms. This applies for instance to oxygen-depleting substances, coloured

substances or turbid matter which reduce light exposure.

Biological tests also include those tests which examine the effect of organisms on substances (e.g.

microbial degradation studies).

The results of the biological test refer primarily to the organism used in the test and the defined

conditions stipulated for the test procedure. A harmful effect stated by means of standardized biological

tests can justify concern that aquatic organisms and biocoenosis might be endangered. The results,

however, do not permit direct or extrapolative conclusions as to the occurrence of similar effects in

the aquatic environment. This applies in particular to suborganismic tests, as important properties

and physiological functions of intact organisms (e.g. protective integuments, repair mechanisms) are

removed or deactivated.

In principle there is no test organism which can be used to test all the effects on the biocoenosis or

the ecosystem possible under the various combinations of abiotic and biotic conditions. Only a few

(“model”) species representing relevant ecological functions can be tested in practice.

Besides these fundamental and practical limitations in the selection of test organisms some issues

should be taken into account during sampling and sample treatment in order to avoid a change in the

sample properties. This applies to the method of sampling, including the sampling equipment and

sample container as well as the transport to the laboratory. The method of sample pre-treatment and

storage, as well as the preparation of, for example, stock solutions, may have an influence on the test

result as well.

Furthermore, the sample to be tested can pose experimental problems on biotesting. Environmental

samples (e.g. waste water, eluates) are complex mixtures and may contain, for example, sparingly

soluble, volatile, unstable, coloured substances or suspended, sometimes colloidal, particles. The

complexity and heterogeneity of materials give rise to a variety of experimental problems when

performing biotests.

Special problems are related to the instability of the test material due to reactions and processes such as

— physical (e.g. phase separation, sedimentation, volatilization),
— chemical (e.g. hydrolysis, photodegradation, precipitation), and/or

— biological (e.g. biodegradation, biotransformation, biological uptake in organisms).

Other problems, especially if spectrometric measurements are applied, relate to turbidity and colour of

the test batch.
vi © ISO 2017 – All rights reserved
---------------------- Page: 12 ----------------------
SIST EN ISO 5667-16:2017
ISO 5667-16:2017(E)

The statistical analysis of the data from biological testing of environmental samples should be

conducted according to the current state of the art if not stipulated by the specific biotest standard.

Finally, it is recommended to implement and maintain a quality management system regardless if a

laboratory is involved in testing of substances or environmental samples.

This document is one of a group of International Standards dealing with the sampling of waters and

sediments and is intended to be read in conjunction with the other parts of the ISO 5667 series, in

particular with ISO 5667-1, ISO 5667-3 and ISO 5667-15.
© ISO 2017 – All rights reserved vii
---------------------- Page: 13 ----------------------
SIST EN ISO 5667-16:2017
---------------------- Page: 14 ----------------------
SIST EN ISO 5667-16:2017
INTERNATIONAL STANDARD ISO 5667-16:2017(E)
Water quality — Sampling —
Part 16:
Guidance on biotesting of samples
1 Scope

This document gives practical guidance on sampling, pre-treatment, performance and evaluation of

environmental samples in the context of performing biological tests. Information is given on how to

cope with the problems of biotesting arising from the sample and the suitability of the test design.

It is intended to convey practical experience concerning precautions to be taken by describing methods

successfully proven to solve or to circumvent some of the experimental problems of biotesting of, for

example, waters.

Primarily dealt with are substance-related problems concerning sampling and pre-treatment of

environmental samples (e.g. waste water samples) for the performance of biotests.

This guidance is on ecotoxicological testing with organisms (single-species biotests; in vivo and in

vitro). Some features addressed in this document also apply to biotests using single-cell systems (in

vitro bioassays) and biodegradation studies as far as sampling and sample preparations are concerned.

Testing of substances in the water solubility range is also addressed.

Reference has been made as far as possible to existing International Standards and guidelines.

Information taken from published papers or oral communication has been utilized as well.

This document is applicable to biological tests for determining the effect of environmental samples

like treated communal and industrial waste water, groundwater, fresh water, aqueous extracts (e.g.

leachates, eluates), pore water of sediments and whole sediments. This document is also applicable to

chemical substances.

This document is not applicable to bacteriological examination of water. Appropriate methods for

[17]
bacteriological examination are described in other documents (see ISO 19458 ).
2 Normative references
There are no normative references in this document.
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:

— IEC Electropedia: available at http:// www .electropedia .org/
— ISO Online browsing platform: available at http:// www .iso .org/ obp
3.1
blank
mixture of water and nutrients without test organism
© ISO 2017 – All rights reserved 1
---------------------- Page: 15 ----------------------
SIST EN ISO 5667-16:2017
ISO 5667-16:2017(E)
3.2
cell density
number of cells per unit volume of medium
Note 1 to entry: Cell density is expressed in cells per millilitre.
[6]
[SOURCE: ISO 10253:2016, 3.1]
3.3
control

control medium (3.4), or control sediment (3.5), including organisms used in the test, without test sample

3.4
control medium
combination of dilution water and/or nutrient medium used in the test
[18]
[SOURCE: ISO 20079:2005, 3.6]
3.5
control sediment
defined artificial or natural sediment used in the test
3.6
dilution level

reciprocal value of the volume fraction of test sample in dilution water (3.7) 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.
[13]

[SOURCE: ISO 15088:2007, 3.2, modified — “waste water” replaced by “test sample”]

3.7
dilution water
water added to the test sample to prepare a series of defined dilutions

Note 1 to entry: The composition of the water is specified in the respective standard.

[18]
[SOURCE: ISO 20079:2005, 3.7, modified — “Note 1 to entry” has been added]
3.8
effective concentration

concentration of the test material in water or sediment that causes x % change in response during a

specified time interval

[SOURCE: ISO/TS 20281:2006, 3.8.1, modified — “quantal” has been removed from the term and

abbreviated term; “soil” and “(e.g. immobility)” have been removed from the definition; the EXAMPLE

[20]
and Notes 1 and 2 to entry are not included]
3.9
field blank

container prepared in the laboratory using reagent water or other blank matrix and sent with the

sampling personnel for exposure to the sampling environment to verify possible contamination during

sampling
[9]
[SOURCE: ISO 11074:2015, 4.5.3]
2 © ISO 2017 – All rights reserved
---------------------- Page: 16 ----------------------
SIST EN ISO 5667-16:2017
ISO 5667-16:2017(E)
3.10
growth rate
proportional rate of increase in biomass per unit of time: (1/day)

[SOURCE: ISO 10253:2016, 3.2, modified — “specific grow rate” replaced by “growth rate

...