ISO 5430:2023
(Main)Plastics — Ecotoxicity testing scheme for soluble decomposition intermediates from biodegradable plastic materials and products used in the marine environment — Test methods and requirements
Plastics — Ecotoxicity testing scheme for soluble decomposition intermediates from biodegradable plastic materials and products used in the marine environment — Test methods and requirements
This document specifies test methods and evaluation criteria by addressing potential ecotoxicological adverse effects on marine organisms. Adverse effects on marine species can be caused by soluble degradation products of plastic materials such as intermediates or remaining residues resulting from the biodegradation of plastic materials that are used in products for marine applications (e.g. nets for fish farming, dolly ropes, floats, buoys and other non-fishing applications) and which are used in different marine zones, e.g. eulittoral, sublittoral or pelagic zones. The ecotoxicity testing scheme covers marine organisms from four trophic levels, primary producer, primary and secondary consumers and decomposer: — toxicity to marine algae, — toxicity to marine invertebrates, — toxicity to marine fish, — toxicity to marine microorganisms. This document is not suitable for the assessment of adverse effects caused by solid materials of any size.
Plastiques — Méthodes d'essai d'écotoxicité pour les intermédiaires de décomposition solubles à partir de matériaux et produits plastiques biodégradables utilisés dans le milieu marin — Méthodes d'essai et exigences
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INTERNATIONAL ISO
STANDARD 5430
First edition
2023-05
Plastics — Ecotoxicity testing
scheme for soluble decomposition
intermediates from biodegradable
plastic materials and products used
in the marine environment — Test
methods and requirements
Plastiques — Méthodes d'essai d'écotoxicité pour les intermédiaires
de décomposition solubles à partir de matériaux et produits
plastiques biodégradables utilisés dans le milieu marin — Méthodes
d'essai et exigences
Reference number
ISO 5430:2023(E)
© ISO 2023
---------------------- Page: 1 ----------------------
ISO 5430:2023(E)
COPYRIGHT PROTECTED DOCUMENT
© 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: +41 22 749 01 11
Email: copyright@iso.org
Website: www.iso.org
Published in Switzerland
ii
© ISO 2023 – All rights reserved
---------------------- Page: 2 ----------------------
ISO 5430:2023(E)
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 2
4 General . 2
5 Test methods . 3
5.1 Preparation of marine matrices for ecotoxicity testing . 3
5.1.1 General . 3
5.1.2 Enriched cultures . 4
5.2 Determination of ecotoxicological effects on marine algae (mandatory) . 4
5.3 Determination of ecotoxicological effects on marine invertebrates (marine
copepods) (mandatory) . 5
5.4 Determination of ecotoxicological effects on marine fish (optional) . 5
5.5 Determination of ecotoxicological effects on marine microorganisms (mandatory) . 5
6 Test report . 5
Annex A (normative) Determination of ecotoxicological effects on the marine algae
Skeletonema sp. and Phaeodactylum tricornutum . 6
Annex B (normative) Determination of ecotoxicological effects on marine copepods
(Copepoda, Crustacea) . 8
Annex C (informative) Determination of ecotoxicological effects on marine fish .10
Annex D (normative) Determination of ecotoxicological effects on the marine
microorganism Aliivibrio fischeri .12
Bibliography .14
iii
© ISO 2023 – All rights reserved
---------------------- Page: 3 ----------------------
ISO 5430:2023(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 61, Plastics, Subcommittee SC 14,
Environmental aspects.
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.
iv
© ISO 2023 – All rights reserved
---------------------- Page: 4 ----------------------
ISO 5430:2023(E)
Introduction
There is a growing interest in using biodegradable materials in products used in the marine environment
(e.g. farming and fishing gears, floats, buoys and other non-fishing materials or products). These
products are subject to wear and tear and, therefore, tend to be sources of macro- and microplastics.
Biodegradability is a factor that, in principle, mitigates the environmental impacts of fragmentation,
thanks to persistence times in the environment that are shorter than that of non-biodegradable
materials. Therefore, test methods to measure the level of biodegradation and disintegration of plastic
materials in different marine habitats have been established by ISO/TC 61/SC 14 in recent years to
better characterize the degradation of plastics in these very particular environments:
— The test standards like ISO 18830 (or ISO 19679), ISO 22404, ISO 23977-1 (or ISO 23977-2) are suited
to investigate the biodegradation of plastic materials exposed to marine environmental samples
(sediments and seawater).
— The ISO standard specification ISO 22403 specifies test methods and requirements to assess
the intrinsic biodegradability of materials exposed to marine inocula under mesophilic aerobic
laboratory conditions.
— The ISO standard ISO 22766 describes methods for the determination of the degree of disintegration
of biodegradable plastic materials exposed to sublittoral and eulittoral habitats under real field
conditions.
— The ISO standard ISO 23832 describes methods for the determination of the degradation rate
and disintegration degree of plastic materials exposed to marine environmental matrices under
laboratory conditions.
Besides data on the biodegradability of plastics materials, tests on ecotoxicological effects of potential
soluble decomposition intermediates of the biodegradation process to marine organisms are necessary
to enable developer and manufacturer of materials to evaluate and to exclude negative effects on
marine organisms. In addition, in combination with data on biodegradability, data on ecotoxicological
effects can be used for e.g. risk assessment purposes.
This document specifies test methods and requirements for assessing potential adverse effects on
different marine organisms caused by soluble decomposition intermediates (degradation products)
resulting from the decomposition of plastic materials that are intentionally used in marine areas.
Comprehensive ecotoxicity testing schemes and evaluation criteria are already part of ISO standard
specifications like ISO 17088 and ISO 23517. The scheme and criteria given in ISO 23517 are equivalent
to the requirements specified in the CEN-standard EN 17033. The CEN-document EN 17427 on carrier
bags suitable for treatment in well-managed home composting installations includes an ecotoxicity
testing scheme that follows the same basic principles as laid down in above mentioned ISO- and EN-
standard specifications: adverse effects are assessed based on results from three tests covering
organisms representing different trophic levels.
This document aims to provide a suitable ecotoxicity testing scheme for marine organisms.
v
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---------------------- Page: 5 ----------------------
INTERNATIONAL STANDARD ISO 5430:2023(E)
Plastics — Ecotoxicity testing scheme for soluble
decomposition intermediates from biodegradable plastic
materials and products used in the marine environment —
Test methods and requirements
1 Scope
This document specifies test methods and evaluation criteria by addressing potential ecotoxicological
adverse effects on marine organisms. Adverse effects on marine species can be caused by soluble
degradation products of plastic materials such as intermediates or remaining residues resulting from
the biodegradation of plastic materials that are used in products for marine applications (e.g. nets
for fish farming, dolly ropes, floats, buoys and other non-fishing applications) and which are used in
different marine zones, e.g. eulittoral, sublittoral or pelagic zones.
The ecotoxicity testing scheme covers marine organisms from four trophic levels, primary producer,
primary and secondary consumers and decomposer:
— toxicity to marine algae,
— toxicity to marine invertebrates,
— toxicity to marine fish,
— toxicity to marine microorganisms.
This document is not suitable for the assessment of adverse effects caused by solid materials of any size.
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 472, Plastics — Vocabulary
ISO 5667-16, Water quality — Sampling — Part 16: Guidance on biotesting of samples
ISO 10210, Plastics — Methods for the preparation of samples for biodegradation testing of plastic
materials
ISO 10253:2016, Water quality — Marine algal growth inhibition test with Skeletonema sp. and
Phaeodactylum tricornutum
ISO 11348 (all parts), Water quality — Determination of the inhibitory effect of water samples on the light
emission of Vibrio fischeri (Luminescent bacteria test)
ISO 14669, Water quality — Determination of acute lethal toxicity to marine copepods (Copepoda,
Crustacea)
ISO 17088, Plastics — Organic recycling — Specifications for compostable plastics
ISO 18830, Plastics — Determination of aerobic biodegradation of non-floating plastic materials in a
seawater/sandy sediment interface — Method by measuring the oxygen demand in closed respirometer
1
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ISO 5430:2023(E)
ISO 19679, Plastics — Determination of aerobic biodegradation of non-floating plastic materials in a
seawater/sediment interface — Method by analysis of evolved carbon dioxide
ISO 22404, Plastics — Determination of the aerobic biodegradation of non-floating materials exposed to
marine sediment — Method by analysis of evolved carbon dioxide
ISO 23977-1, Plastics — Determination of the aerobic biodegradation of plastic materials exposed to
seawater — Part 1: Method by analysis of evolved carbon dioxide
ISO 23977-2, Plastics — Determination of the aerobic biodegradation of plastic materials exposed to
seawater — Part 2: Method by measuring the oxygen demand in closed respirometer
OECD 2019), Test No. 203 Fish, Acute Toxicity Test, OECD Guidelines for the Testing of Chemicals,
Section 2
3 Terms and definitions
For the purposes of this document, the terms and definitions given in ISO 472, ISO 17088 and the
following apply.
ISO and IEC maintain terminological databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https:// www .iso .org/ obp
— IEC Electropedia: available at https:// www .electropedia .org/
3.1
biodegradable plastic
plastic material which, under controlled laboratory conditions, is intrinsically biodegradable
Note 1 to entry: The relevant controlled laboratory conditions are presented in ISO 22403.
4 General
The marine ecotoxicity testing scheme uses test methods with several marine organisms belonging to
different taxonomic groups and trophic levels (see Table 1). The selected test methods, in combination
with defined test acceptability criteria, allow the characterization of toxic properties of soluble
degradation products of biodegradable plastic materials (i.e. intermediates or remaining residues) to
marine organisms.
Table 1 — Ecotoxicity testing scheme for evaluation of adverse effects on marine organisms
that represent different trophic levels in marine aquatic food webs
Marine taxa – species Ecological process Test method
Algae
Marine algal growth inhibition test accord-
— Skeletonema sp
Primary producer ing to ISO 10253 with modifications speci-
fied in Annex A
— Phaeodactylum tricornutum
Invertebrates
— Acartia tonsa
Acute lethal toxicity test to marine copepods
Primary consumer according to ISO 14669 with modifications
— Tisbe battagliai
specified in Annex B
— Nitocra spinipes
2
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ISO 5430:2023(E)
TTabablele 1 1 ((ccoonnttiinnueuedd))
Marine taxa – species Ecological process Test method
Fish (optional)
— Cyprinodon variegatus
Fish acute toxicity test according to
Secondary consumer OECD 203 with modifications specified in
— Dicentrarchus labrax
Annex C
— Pagrus major
Micoorganisms Determination of inhibitory effect on light
emission of luminescent bacteria according
Decomposer
— Aliivibrio fischeri (old name:
to ISO 11348 (all parts) with modifications
Vibrio fischeri)
specified in Annex D
Ecotoxicological effects on marine organisms shall be determined by comparing marine matrices from
preceding marine biodegradation tests produced with or without the addition of a test material.
5 Test methods
5.1 Preparation of marine matrices for ecotoxicity testing
5.1.1 General
Plastic materials preferably in the form of film or powder with a maximum thickness or diameter of
500 µm shall be exposed to marine inocula under mesophilic laboratory test conditions following ISO
standard test method ISO 18830 (or ISO 19679), ISO 22404 or ISO 23977-1 (or ISO 23977-2), for up to
two years. Powder shall be prepared in accordance with ISO 10210.
The minimum initial test item concentration in the marine biodegradation tests shall be the
concentration recommended in the standard test method used to produce the matrices in order to
ensure comparable initial test item concentrations and proper biodegradation. For that reason, the
minimum test item concentrations shall be at least 75 mg/100 g of sediment for ISO 22404 and at least
100 mg/l of seawater plus sediment for ISO 18830 or ISO 19679 and 100 mg/l seawater for ISO 23977-1
or ISO 23977-2.
The same applies to the reference material. Use microcrystalline cellulose or ashless cellulose filters as
1)
a reference material . If possible, the form, size and the amount of carbon added in form of the polymer
should be comparable to that of the test material.
In addition to marine matrices exposed to plastic materials (test sample) or to reference material
(reference sample), marine matrices not exposed to plastic materials (control sample) are incubated in
parallel.
The reactors for preparing the matrices for the test samples shall be run until constant biodegradation
per time unit (e.g. 5 % biodegradation in one month) is reached and more than 50 % of total
biodegradation is observed so that degradation products might have released into the seawater.
There are two options to test the ecotoxicity of soluble decomposition intermediates from biodegradable
plastic materials:
a) Option 1 – limit test: the marine test samples for ecotoxicity testing should be used for the indicated
ecotoxicity tests when the absolute mineralization level reaches 50 %.
1) Microcrystalline Cellulose "Avicel" produced by Merck or Laboratory filter paper Whatman n° 42 has been
found satisfactory for this purpose and are examples of suitable products available commercially. This information
is given for the convenience of users of this document and does not constitute an endorsement by ISO of these
products.
3
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ISO 5430:2023(E)
b) Option 2 – time dependent test: the marine test samples for ecotoxicity testing should be used for
the indicated ecotoxicity tests when absolute mineralization reaches 20 %, 40 % and a plateau
(equal or greater than 60 %).
NOTE 1 Option 2 can be used in cases where an adaption of bacterial activity in biodegradation is expected,
when the constituents are unevenly distributed in the test material or when the rate of biodegradation of
different constituents is expected to be different.
For both options, the biodegradation should be monitored following existing ISO standard test method
ISO 18830 (or ISO 19679), ISO 22404 or ISO 23977-1 (or ISO 23977-2).
WARNING — The biodegradation methods used in this document are exclusively intended to be
used to generate a testing matrix for ecotoxicity testing of biodegradable plastic materials or
products that already fulfil the requirements (see 3.1) to be biodegradable. They shall not be
used to assess the biodeg
...
DRAFT INTERNATIONAL STANDARD
ISO/DIS 5430
ISO/TC 61/SC 14 Secretariat: DIN
Voting begins on: Voting terminates on:
2022-05-20 2022-08-12
Plastics — Marine ecotoxicity testing scheme for soluble
decomposition intermediates from biodegradable plastic
materials in products intentionally used in the marine
environment – Test methods and requirements
ICS: 13.020.40; 83.080.01
THIS DOCUMENT IS A DRAFT CIRCULATED
FOR COMMENT AND APPROVAL. IT IS
THEREFORE SUBJECT TO CHANGE AND MAY
This document is circulated as received from the committee secretariat.
NOT BE REFERRED TO AS AN INTERNATIONAL
STANDARD UNTIL PUBLISHED AS SUCH.
IN ADDITION TO THEIR EVALUATION AS
BEING ACCEPTABLE FOR INDUSTRIAL,
TECHNOLOGICAL, COMMERCIAL AND
USER PURPOSES, DRAFT INTERNATIONAL
STANDARDS MAY ON OCCASION HAVE TO
BE CONSIDERED IN THE LIGHT OF THEIR
POTENTIAL TO BECOME STANDARDS TO
WHICH REFERENCE MAY BE MADE IN
Reference number
NATIONAL REGULATIONS.
ISO/DIS 5430:2022(E)
RECIPIENTS OF THIS DRAFT ARE INVITED
TO SUBMIT, WITH THEIR COMMENTS,
NOTIFICATION OF ANY RELEVANT PATENT
RIGHTS OF WHICH THEY ARE AWARE AND TO
PROVIDE SUPPORTING DOCUMENTATION. © ISO 2022
---------------------- Page: 1 ----------------------
ISO/DIS 5430:2022(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2022
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting on
the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address below
or ISO’s member body in the country of the requester.
ISO copyright office
CP 401 • Ch. de Blandonnet 8
CH-1214 Vernier, Geneva
Phone: +41 22 749 01 11
Email: copyright@iso.org
Website: www.iso.org
Published in Switzerland
ii
© ISO 2022 – All rights reserved
---------------------- Page: 2 ----------------------
ISO/DIS 5430:2022(E)
Contents Page
Foreword .iv
Introduction .v
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 2
4 General . 2
5 Test methods . 3
5.1 Preparation of marine matrices for ecotoxicity testing . 3
5.1.1 Enriched cultures . 4
5.2 Determination of ecotoxicological effects on marine algae (mandatory) . 4
5.3 Determination of ecotoxicological effects to the marine invertebrates (marine
copepods) (mandatory) . 4
5.4 Determination of ecotoxicological effects to the marine fish (optional) . 5
5.5 Determination of ecotoxicological effects on marine microorganisms (mandatory) . 5
6 Test report . 5
Annex A (normative) Determination of ecotoxicological effects on the marine algae
Skeletonema sp. and Phaeodactylum tricornutum . 6
Annex B (normative) Determination of ecotoxicological effects on marine copepods
(Copepoda, Crustacea) . 8
Annex C (informative) Determination of ecotoxicological effects on marine fish .10
Annex D (normative) Determination of ecotoxicological effects on the marine
microorganism Vibrio fischeri .12
Bibliography .14
iii
© ISO 2022 – All rights reserved
---------------------- Page: 3 ----------------------
ISO/DIS 5430:2022(E)
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out
through ISO technical committees. Each member body interested in a subject for which a technical
committee has been established has the right to be represented on that committee. International
organizations, governmental and non-governmental, in liaison with ISO, also take part in the work.
ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of
electrotechnical standardization.
The procedures used to develop this document and those intended for its further maintenance are
described in the ISO/IEC Directives, Part 1. In particular, the different approval criteria needed for the
different types of ISO documents should be noted. This document was drafted in accordance with the
editorial rules of the ISO/IEC Directives, Part 2 (see www.iso.org/directives).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
any patent rights identified during the development of the document will be in the Introduction and/or
on the ISO list of patent declarations received (see www.iso.org/patents).
Any trade name used in this document is information given for the convenience of users and does not
constitute an endorsement.
For an explanation of the voluntary nature of standards, the meaning of ISO specific terms and
expressions related to conformity assessment, as well as information about ISO's adherence to
the World Trade Organization (WTO) principles in the Technical Barriers to Trade (TBT), see
www.iso.org/iso/foreword.html.
This document was prepared by Technical Committee ISO/TC 61, Plastics, Subcommittee SC 14,
Environmental aspects.
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.
iv
© ISO 2022 – All rights reserved
---------------------- Page: 4 ----------------------
ISO/DIS 5430:2022(E)
Introduction
There is a growing interest in using biodegradable materials in products used in the marine
environment (e.g. farming and fishing gears, floats, buoys). These products are subject to wear and
tear and, therefore, tend to be sources of macro- and microplastics. Biodegradability is a factor that
in principle mitigates the environmental impacts of fragmentation, thanks to persistence times in
the environment that are shorter than that of non-biodegradable materials. Therefore, test methods
to measure the level of biodegradation and disintegration of plastic materials in different marine
habitats have been established by ISO/TC 61/SC 14 in recent years in order to better characterize the
degradation of plastics in these very particular environments:
— The test standards like ISO 18830 (or ISO 19679), ISO 22404, ISO 23977-1 (or ISO 23977-2) are suited
to investigate the biodegradation of plastic materials exposed to marine environmental samples
(sediments and seawater).
— The ISO standard specification ISO 22403 specifies test methods and requirements to assess
the intrinsic biodegradability of materials exposed to marine inocula under mesophilic aerobic
laboratory conditions.
— The ISO standard ISO 22766 describes methods for the determination of the degree of disintegration
of biodegradable plastic materials exposed to sublittoral and eulittoral habitats under real field
conditions.
Beside data on the biodegradability of plastics materials, tests on ecotoxicological effects of potential
soluble decomposition intermediates of the biodegradation process to marine organisms are necessary
to enable developer and manufacturer of materials to evaluate and to exclude negative effects on
marine organisms. In addition, in combination with data on biodegradability, data on ecotoxicological
effects may be used for e.g., risk assessment purposes.
This document specifies test methods and requirements for assessing potential adverse effects on
different marine organisms caused by soluble decomposition intermediates (degradation products)
resulting from the decomposition of plastic materials that are intentionally disposed to marine areas.
Comprehensive ecotoxicity testing schemes and evaluation criteria are already part of ISO standard
specifications like ISO 17088 and ISO 23517. The scheme and criteria given in ISO 23517 are equivalent
to the requirements specified in the CEN-standard EN 17033. The CEN-document prEN 17427:2020
on carrier bags suitable for treatment in well-managed home composting installations includes an
ecotoxicity testing scheme that follows the same basic principles as laid down in above mentioned ISO-
and EN-standard specifications: adverse effects are assessed based on results from three tests covering
organisms representing different trophic levels.
This document is aiming to provide a suitable ecotoxicity testing scheme for marine organisms.
v
© ISO 2022 – All rights reserved
---------------------- Page: 5 ----------------------
DRAFT INTERNATIONAL STANDARD ISO/DIS 5430:2022(E)
Plastics — Marine ecotoxicity testing scheme for soluble
decomposition intermediates from biodegradable plastic
materials in products intentionally used in the marine
environment – Test methods and requirements
1 Scope
This document specifies test methods and evaluation criteria by addressing potential ecotoxicological
adverse effects on marine organisms. Adverse effects on marine species may be caused by soluble
degradation products of plastic materials such as intermediates or remaining residues resulting from
the biodegradation of plastic materials that are used in products for marine applications (e.g. nets for
fish farming, dolly ropes, floats, buoys, etc.) and which are intentionally disposed to coastal areas, e.g.
eulittoral, sublittoral or pelagic zones.
The ecotoxicity testing scheme covers marine organisms from four trophic levels, primary producer,
primary and secondary consumer and decomposer:
— toxicity to marine algae,
— toxicity to marine invertebrates,
— toxicity to marine fish,
— toxicity to marine microorganisms.
This document is not suitable for the assessment of adverse effects caused by solid, non-biodegradable
plastic materials of any size.
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 5667-16, Water quality — Sampling — Part 16: Guidance on biotesting of samples
ISO 10210, Plastics — Methods for the preparation of samples for biodegradation testing of plastic
materials
ISO 10253, Water quality — Marine algal growth inhibition test with Skeletonema sp. and Phaeodactylum
tricornutum
ISO 11348-1, Water quality — Determination of the inhibitory effect of water samples on the light emission
of Vibrio fischeri (Luminescent bacteria test) — Part 1: Method using freshly prepared bacteria
ISO 11348-2, Water quality — Determination of the inhibitory effect of water samples on the light emission
of Vibrio fischeri (Luminescent bacteria test) — Part 2: Method using liquid-dried bacteria
ISO 11348-3, Water quality — Determination of the inhibitory effect of water samples on the light emission
of Vibrio fischeri (Luminescent bacteria test) — Part 3: Method using freeze-dried bacteria
ISO 14669, Water quality — Determination of acute lethal toxicity to marine copepods (Copepoda,
Crustacea)
1
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---------------------- Page: 6 ----------------------
ISO/DIS 5430:2022(E)
ISO 18830, Plastics — Determination of aerobic biodegradation of non-floating plastic materials in a
seawater/sandy sediment interface — Method by measuring the oxygen demand in closed respirometer
ISO 19679, Plastics — Determination of aerobic biodegradation of non-floating plastic materials in a
seawater/sediment interface — Method by analysis of evolved carbon dioxide
ISO 22404, Plastics — Determination of the aerobic biodegradation of non-floating materials exposed to
marine sediment — Method by analysis of evolved carbon dioxide
ISO 23977-1, Plastics — Determination of the aerobic biodegradation of plastic materials exposed to
seawater — Part 1: Method by analysis of evolved carbon dioxide
ISO 23977-2, Plastics — Determination of the aerobic biodegradation of plastic materials exposed to
seawater — Part 2: Method by measuring the oxygen demand in closed respirometer
OECD (2019), Test No. 203 Fish, Acute Toxicity Test, OECD Guidelines for the Testing of Chemicals,
Section 2
3 Terms and definitions
For the purposes of this document, the terms and definitions given in ISO 472:2013 and ISO 17088:2021
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
biodegradable plastics
plastics capable of being broken down or mineralized by microorganisms
4 General
The marine ecotoxicity testing scheme uses test methods with several marine organisms belonging to
different taxonomic groups and trophic levels (see Table 1). The selected test methods, in combination
with defined test acceptability criteria, allow the characterization of toxic properties of soluble
degradation products of biodegradable plastic materials (i.e. intermediates or remaining residues) to
marine organisms.
2
© ISO 2022 – All rights reserved
---------------------- Page: 7 ----------------------
ISO/DIS 5430:2022(E)
Table 1 — Ecotoxicity testing scheme for evaluation of adverse effects on marine organisms
that represent different tropic levels in marine aquatic food webs.
Marine taxa – species Ecological process Test method
Algae
Marine algal growth inhibition test accord-
— Skeletonema costatum
Primary producer ing to ISO 10253 with modifications speci-
fied in Annex A
— Phaeodactylum tricornutum
Invertebrate (marine copepod)
— Acartia tonsa
Acute lethal toxicity test to marine copepods
Primary consumer according to ISO 14669 with modifications
— Tisbe battagliai
specified in Annex B
— Nitocra spinipes
Fish (optional)
— Cyprinodon variegatus
Fish acute toxicity test according to
Secondary consumer OECD 203 with modifications specified in
— Dicentrarchus labrax
Annex C
— Pagrus major
Micoorganisms Determination of inhibitory effect on light
emission of luminescent bacteria according
Decomposer
— Aliivibrio fischeri (old name:
to ISO 11348 (all parts) with modifications
Vibrio fischeri)
specified in Annex D
Ecotoxicological effects on marine organisms shall be determined by comparing marine matrices from
preceding marine biodegradation tests produced with or without the addition of a test material.
5 Test methods
5.1 Preparation of marine matrices for ecotoxicity testing
Plastic materials preferably in the form of film or powder with dimension <500 µm shall be exposed
to marine inocula under mesophilic laboratory test conditions following ISO standard test method
ISO 18830 (or ISO 19679), ISO 22404 or ISO 23977-1 (or ISO 23977-2), respectively for up to two years.
Powder can be prepared according to ISO 10210.
The minimum initial test item concentration in the marine biodegradation tests shall be the
concentration recommended in the standard test method used for the production of the matrices in
order to ensure proper biodegradation (e.g. at least 25 mg/100 g of sediment for ISO 22404, at least
100 mg/l of seawater plus sediment for ISO 19679).
Same applies to a reference material. Use microcrystalline cellulose or ashless cellulose filters as
1)
a reference material . If possible, the TOC, form, and size should be comparable to that of the test
material.
In addition to marine matrices exposed to plastic materials (test sample) or to reference material
(reference sample), marine matrices not exposed to plastic materials (control sample) are incubated in
parallel.
The reactors for preparing the matrices for the test samples shall be run until significant biodegradation
is clearly identifiable and degradation products might have released into the seawater. There are two
options to test for ecotoxicity of soluble decomposition intermediates from biodegradable plastic
materials:
1) Microcrystalline Cellulose "Avicel" produced by Merck or Laboratory filter paper Whatman n° 42 has been
found satisfactory for this purpose and are examples of suitable products available commercially. This information
is given for the convenience of users of this document and does not constitute an endorsement by ISO of these
products.
3
© ISO 2022 – All rights reserved
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ISO/DIS 5430:2022(E)
Option 1 – limit test (to be used for pure polymers): the marine test samples for ecotoxicity testing may
be used for the indicated ecotoxicity tests when the mineralization level reaches 50 %.
Option 2 – time depending test (to be used for blends): the marine test samples for ecotoxicity testing
may be used for the indicated ecotoxici
...
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