EN ISO 14851:2019

Determination of the ultimate aerobic biodegradability of plastic materials in an aqueous medium - Method by measuring the oxygen demand in a closed respirometer (ISO 14851:2019)

This document specifies a method, by measuring the oxygen demand in a closed respirometer, for the determination of the degree of aerobic biodegradability of plastic materials, including those containing formulation additives. The test material is exposed in an aqueous medium under laboratory conditions to an inoculum from activated sludge.
If an unadapted activated sludge is used as the inoculum, the test simulates the biodegradation processes which occur in a natural aqueous environment; if a mixed or pre-exposed inoculum is used, the method is used to investigate the potential biodegradability of a test material.
The conditions used in this document do not necessarily correspond to the optimum conditions allowing maximum biodegradation to occur, but this document is designed to determine the potential biodegradability of plastic materials or give an indication of their biodegradability in natural environments.
The method enables the assessment of the biodegradability to be improved by calculating a carbon balance (optional, see Annex E).
The method applies to the following materials.
—          Natural and/or synthetic polymers, copolymers or mixtures thereof.
—          Plastic materials which contain additives such as plasticizers, colorants or other compounds.
—          Water-soluble polymers.
—          Materials which, under the test conditions, do not inhibit the microorganisms present in the inoculum. Inhibitory effects can be determined using an inhibition control or by another appropriate method (see, for example, ISO 8192[2]). If the test material is inhibitory to the inoculum, a lower test concentration, another inoculum or a pre-exposed inoculum can be used.

Bestimmung der vollständigen aeroben Bioabbaubarkeit von Kunststoff-Materialien in einem wässrigen Medium - Verfahren mittels Messung des Sauerstoffbedarfs in einem geschlossenen Respirometer (ISO 14851:2019)

Dieses Dokument legt ein Verfahren fest, mit dem durch Messung des Sauerstoffbedarfs in einem geschlossenen Respirometer der Grad der aeroben Bioabbaubarkeit von Kunststoffen, einschließlich solcher Kunststoffe, die Formulierungs Additive enthalten, bestimmt werden kann. Die Prüfsubstanz wird in einem wässrigen Medium unter Laborbedingungen einem Inokulum aus Belebtschlamm ausgesetzt.
Wird ein nicht adaptierter Belebtschlamm als Inokulum verwendet, werden durch die Prüfung die Bioabbauprozesse simuliert, die in einer natürlichen wässrigen Umgebung auftreten; wird ein gemischtes oder voradaptiertes Inokulum verwendet, dient das Verfahren dazu, die potentielle Bioabbaubarkeit einer Prüfsubstanz zu untersuchen.
Die in diesem Dokument angewendeten Bedingungen entsprechen nicht notwendigerweise den optimalen Bedingungen, die das Auftreten eines maximalen Bioabbaus zulassen, sondern dieses Dokument ist für die Bestimmung der potentiellen Bioabbaubarkeit von Kunststoffen ausgelegt oder dafür vorgesehen, Erkenntnisse hinsichtlich deren Bioabbaubarkeit in natürlichen Umgebungen zu liefern.
Das Verfahren ermöglicht durch die Berechnung einer Kohlenstoffbilanz (wahlweise, siehe Anhang E) die Verbesserung der Beurteilung der Bioabbaubarkeit.
Das Verfahren ist auf folgende Substanzen anwendbar:
   natürliche und/oder synthetische Polymere, Copolymere oder Gemische aus diesen;
   Kunststoffe, die Additive, wie z. B. Weichmacher, Farbstoffe oder andere Verbindungen, enthalten;
   wasserlösliche Polymere;
   Substanzen, die unter den Prüfbedingungen die im Inokulum vorhandenen Mikroorganismen nicht inhibieren. Inhibierungswirkungen können durch eine Inhibierungskontrolle oder ein anderes geeignetes Verfahren bestimmt werden (siehe zum Beispiel ISO 8192 [2]). Wirkt die Prüfsubstanz inhibierend auf das Inokulum, kann eine geringere Prüfkonzentration, ein anderes oder ein voradaptiertes Inokulum verwendet werden.

Évaluation de la biodégradabilité aérobie ultime des matériaux plastiques en milieu aqueux - Méthode par détermination de la demande en oxygène dans un respiromètre fermé (ISO 14851:2019)

Le présent document spécifié une méthode d'évaluation du taux de biodégradation aérobie des matériaux plastiques, y compris ceux contenant des additifs, par la détermination de la demande d'oxygène dans un respiromètre fermé. Le matériau d'essai en milieu aqueux est exposé dans des conditions de laboratoire à un inoculum provenant de boues activées.
La méthode simule les processus de biodégradation d'un environnement aquatique naturel si l'on utilise, par exemple, des boues activées non adaptées; si on utilise un inoculum mélangé ou pré-exposé, la méthode permet d'étudier la biodégradabilité potentielle du matériau d'essai.
Les conditions utilisées dans le présent document ne correspondent pas nécessairement aux conditions optimales  permettant d'obtenir le taux maximal de biodégradation; cependant, le présent document est conçu pour déterminer la biodégradabilité potentielle ou pour donner une indication de la biodégradabilité des matériaux plastiques dans le milieu naturel.
La méthode permet d'affiner l'évaluation de la biodégradabilité par le calcul d'un bilan de carbone (facultatif, voir l'Annexe E).
La présente méthode s'applique aux matériaux suivants:
—          polymères naturels et/ou synthétiques, copolymères ou mélanges de ceux-ci;
—          matériaux plastiques contenant des additifs tels que plastifiants, colorants ou tout autre composé;
—          polymères hydrosolubles;
—          matériaux n'ayant pas d'effet inhibiteur dans les conditions d'essai sur les micro-organismes présents dans l'inoculum. Les effets inhibiteurs peuvent être déterminés en utilisant un dispositif de contrôle de l'inhibition ou par toute autre méthode appropriée (voir, par exemple, l'ISO 8192[2]). Si le matériau d'essai a un effet inhibiteur vis-à-vis de l'inoculum, il est possible d'utiliser une plus faible concentration, un autre inoculum ou un inoculum pré-exposé.

Določanje končne aerobne biorazgradljivosti polimernih materialov v vodnem mediju - Metoda z merjenjem porabe kisika v zaprtem respirometru (ISO 14851:2019)

Ta dokument določa metodo za določanje stopnje aerobne biorazgradljivosti polimernih materialov, vključno s takimi, ki vsebujejo formulacijske dodatke, z merjenjem porabe kisika v zaprtem respirometru. Preskusni material je v vodnem mediju pod laboratorijskimi pogoji izpostavljen inokulumu iz aktivnega blata.
Če se kot inokulum uporabi neprilagojeno aktivno blato, preskus simulira postopke biorazgradnje, ki potekajo v naravnem vodnem okolju; če se uporabi mešan ali predhodno izpostavljen inokulum, se metoda uporabi za preiskavo potencialne biorazgradljivosti preskusnega materiala.
Pogoji, uporabljeni v tem dokumentu, niso nujno enaki optimalnim pogojem, pri katerih pride do največje stopnje biorazgradnje, vendar je ta dokument zasnovan za merjenje potencialne biorazgradljivosti polimernih materialov in podaja oceno njihove biorazgradljivosti v naravnih okoljih.
Metoda omogoča izboljšanje ocene biorazgradljivosti z izračunom ravnovesja ogljika (izbirno, glej dodatek E).
Metoda se uporablja za naslednje materiale:
– naravne in/ali sintetične polimere, kopolimere ali njihove mešanice;
– polimerne materiale, ki vsebujejo dodatke, kot so mehčala, barvila ali druge spojine;
– polimere, topne v vodi;
– materiale, ki v preskusnih pogojih ne zavirajo delovanja mikroorganizmov, prisotnih v inokulumu. Zaviralne učinke je mogoče določiti na podlagi spremljanja zaviranja ali z drugo ustrezno metodo (glej npr. standard ISO 8192[2]). Če preskusni material zavira inokulum, se lahko uporabi manjša preskusna koncentracija, drug inokulum ali predhodno izpostavljeni inokulum.

General Information

• Status: Published
• Publication Date: 09-Apr-2019
• Withdrawal Date: 30-Oct-2019
• ICS: 83.080.01 - Plastics in general
• Technical Committee: CEN/TC 249 - Plastics
• Drafting Committee: CEN/TC 249 - Plastics
• Current Stage: 6060 - Definitive text made available (DAV) - Publishing
• Start Date: 10-Apr-2019
• Completion Date: 10-Apr-2019
• Ref Project: SIST EN ISO 14851:2019 - Determination of the ultimate aerobic biodegradability of plastic materials in an aqueous medium - Method by measuring the oxygen demand in a closed respirometer (ISO 14851:2019)

Relations

• Replaces: EN ISO 14851:2004 - Determination of the ultimate aerobic biodegradability of plastic materials in an aqueous medium - Method by measuring the oxygen demand in a closed respirometer (ISO 14851:1999)
• Effective Date: 17-Apr-2019

Overview

EN ISO 14851:2019 (CEN) - also published as ISO 14851:2019 - specifies a laboratory respirometric method for determining the ultimate aerobic biodegradability of plastic materials in an aqueous medium. The method measures oxygen demand in a closed respirometer while the test material is exposed to an inoculum from activated sludge. It is designed to determine the potential biodegradability of polymers (including formulated plastics and water‑soluble polymers) and to provide an indication of biodegradation behaviour in natural aquatic environments.

Key topics and technical requirements

• Test principle: Measurement of oxygen uptake in a closed respirometer to quantify aerobic biodegradation.
• Inoculum: Activated sludge from wastewater treatment; either unadapted (simulates natural aqueous environments) or pre‑exposed/mixed (used to probe potential biodegradability).
• Applicable materials:
  • Natural and synthetic polymers, copolymers or blends
  • Plastics containing additives (plasticizers, colorants, etc.)
  • Water‑soluble polymers
  • Materials that do not inhibit microbial activity under test conditions
• Controls and validity:
  • Blank controls and reference materials required
  • Use of inhibition controls or methods (e.g., ISO 8192) if inhibitory effects are suspected
  • Test conditions reflect potential, not necessarily maximum, biodegradation
• Optional analyses and annexes:
  • Carbon balance calculation (Annex E) to improve assessment
  • Theoretical oxygen demand (ThOD, Annex A)
  • Nitrification correction (Annex B)
  • Closed manometric respirometer and two‑phase closed‑bottle variants (Annexes C and D)
  • Methods for quantifying residual insoluble polymer and molecular mass (Annex F)
  • CO2 absorption procedures (Annex G)
• Health/safety: Activated sludge may contain pathogenic organisms - appropriate precautions are required.

Applications and practical value

• Product development & R&D: Polymer formulators and materials scientists use EN ISO 14851:2019 to assess biodegradation potential during design and formulation stages.
• Environmental testing laboratories: Standardized respirometric tests provide reproducible data for research, comparative studies and life‑cycle assessments.
• Regulatory & compliance: Supports claims about aquatic biodegradability and informs risk assessment, waste management and labeling decisions.
• Quality control & certification: Helps manufacturers demonstrate biodegradability performance of plastics and additives under controlled aqueous conditions.

Who should use this standard

• Polymer manufacturers, converters and compounders
• Environmental and analytical testing laboratories
• Regulatory bodies, eco‑labeling and certification organizations
• Waste‑management and water‑quality researchers

Related standards

• ISO 8192 (for guidance on inhibition controls) - referenced in EN ISO 14851:2019.
• Consult other ISO/CEN biodegradability standards for soil or compost testing (note: EN ISO 14851 excludes soil/compost inocula).

Keywords: EN ISO 14851:2019, ISO 14851, aerobic biodegradability, closed respirometer, oxygen demand, plastic biodegradability test, activated sludge, carbon balance.

Standards Content (Sample)

SLOVENSKI STANDARD
01-junij-2019
Nadomešča:
SIST EN ISO 14851:2004
Določanje končne aerobne biorazgradljivosti polimernih materialov v vodnem
mediju - Metoda z merjenjem porabe kisika v zaprtem respirometru (ISO
14851:2019)
Determination of the ultimate aerobic biodegradability of plastic materials in an aqueous
medium - Method by measuring the oxygen demand in a closed respirometer (ISO
14851:2019)
Bestimmung der vollständigen aeroben Bioabbaubarkeit von Kunststoff-Materialien in
einem wässrigen Medium - Verfahren mittels Messung des Sauerstoffbedarfs in einem
geschlossenen Respirometer (ISO 14851:2019)
Évaluation de la biodégradabilité aérobie ultime des matériaux plastiques en milieu
aqueux - Méthode par détermination de la demande en oxygène dans un respiromètre
fermé (ISO 14851:2019)
Ta slovenski standard je istoveten z: EN ISO 14851:2019
ICS:
83.080.01 Polimerni materiali na Plastics in general
splošno
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

EN ISO 14851
EUROPEAN STANDARD
NORME EUROPÉENNE
April 2019
EUROPÄISCHE NORM
ICS 83.080.01 Supersedes EN ISO 14851:2004
English Version
Determination of the ultimate aerobic biodegradability of
plastic materials in an aqueous medium - Method by
measuring the oxygen demand in a closed respirometer
(ISO 14851:2019)
Évaluation de la biodégradabilité aérobie ultime des Bestimmung der vollständigen aeroben
matériaux plastiques en milieu aqueux - Méthode par Bioabbaubarkeit von Kunststoff-Materialien in einem
détermination de la demande en oxygène dans un wässrigen Medium - Verfahren mittels Messung des
respiromètre fermé (ISO 14851:2019) Sauerstoffbedarfs in einem geschlossenen
Respirometer (ISO 14851:2019)
This European Standard was approved by CEN on 12 March 2019.

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: Rue de la Science 23, B-1040 Brussels
© 2019 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 14851:2019 E
worldwide for CEN national Members.

Contents Page
European foreword . 3

European foreword
This document (EN ISO 14851:2019) has been prepared by Technical Committee ISO/TC 61 "Plastics"
in collaboration with Technical Committee CEN/TC 249 “Plastics” the secretariat of which is held by
NBN.
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 October 2019, and conflicting national standards shall
be withdrawn at the latest by October 2019.
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. CEN shall not be held responsible for identifying any or all such patent rights.
This document supersedes EN ISO 14851:2004.
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 14851:2019 has been approved by CEN as EN ISO 14851:2019 without any modification.

INTERNATIONAL ISO
STANDARD 14851
Second edition
2019-03
Determination of the ultimate aerobic
biodegradability of plastic materials
in an aqueous medium — Method by
measuring the oxygen demand in a
closed respirometer
Évaluation de la biodégradabilité aérobie ultime des matériaux
plastiques en milieu aqueux — Méthode par détermination de la
demande en oxygène dans un respiromètre fermé
Reference number
ISO 14851:2019(E)
©
ISO 2019
ISO 14851:2019(E)
© ISO 2019
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting
on the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address
below or ISO’s member body in the country of the requester.
ISO copyright office
CP 401 • Ch. de Blandonnet 8
CH-1214 Vernier, Geneva
Phone: [phone]
Fax: [phone]
Email: [e-mail]
Website: www.iso.org
Published in Switzerland
ii © ISO 2019 – All rights reserved

ISO 14851:2019(E)
Contents Page
Foreword .iv
Introduction .vi
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Principle . 3
5 Test environment . 3
6 Reagents . 3
6.1 Distilled or deionized water . 4
6.2 Test medium . 4
6.2.1 Standard test medium . 4
6.2.2 Optimized test medium . . 4
6.3 Pyrophosphate solution . 6
6.4 Carbon dioxide absorber . 6
7 Apparatus . 6
8 Procedure. 6
8.1 Test material . 6
8.2 Reference material . 7
8.3 Preparation of the inoculum . 7
8.4 Test . 8
9 Calculation and expression of results . 9
9.1 Calculation . 9
9.2 Expression and interpretation of results .10
10 Validity of results .10
11 Test report .11
Annex A (informative) Theoretical oxygen demand (ThOD).12
Annex B (informative) Correction of BOD values for interference by nitrification .13
Annex C (informative) Principle of a closed manometric respirometer .15
Annex D (informative) Two-phase closed-bottle version of the respirometric test.17
Annex E (informative) Example of the determination of a carbon balance .20
Annex F (informative) Example of a determination of the amount of water-insoluble
polymer remaining at the end of a biodegradation test and the molecular mass of
the polymer .22
Annex G (informative) Example of the determination of the CO absorbed in the absorbent .23
Bibliography .25
ISO 14851:2019(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).
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: www .iso .org/iso/foreword .html.
The committee responsible for this document is ISO/TC 61, Plastics, Subcommittee SC 14, Environmental
aspects.
This second edition cancels and replaces the first edition (ISO 14851:1999), which has been technically
revised. It also incorporates the Technical Corrigendum ISO 14851:1999/Cor.1:2005. The main changes
compared to the previous edition are as follows:
— the footnotes have been renumbered;
— in Annex C, errors in the key to Figure C.1 have been corrected and minor improvements made to
the figure itself;
— in scope and Clause 8, soil and compost have been excluded for the inoculums used in this document;
iv © ISO 2019 – All rights reserved

ISO 14851:2019(E)
— in 8.4, numbers of test flask for the test material and blank control have been changed from two
to three;
— references in this document have been updated for latest active version;
— the Bibliography has been updated.
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.
ISO 14851:2019(E)
Introduction
With the increasing use of plastics, their recovery and disposal have become a major issue. As a first
priority, recovery is promoted. Complete recovery of plastics, however, is difficult. For example, plastic
litter, which comes mainly from consumers, is difficult to recover completely. Additional examples of
plastics which are difficult to recover are fishing tackle, plastic microbeads in personal care products
and water-soluble polymers. These plastic materials tend to leak from closed waste-management
cycles into the environment. Biodegradable plastics are now emerging as one of the options available
to solve such environmental problems. Plastic materials, such as products or packaging, which are sent
to composting facilities are expected to be potentially biodegradable. Therefore, it is very important
to determine the potential biodegradability of such materials and to obtain an indication of their
biodegradability in natural environments.
vi © ISO 2019 – All rights reserved

INTERNATIONAL STANDARD ISO 14851:2019(E)
Determination of the ultimate aerobic biodegradability
of plastic materials in an aqueous medium — Method by
measuring the oxygen demand in a closed respirometer
WARNING — Sewage, activated sludge, soil and compost may 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 document specifies a method, by measuring the oxygen demand in a closed respirometer, for the
determination of the degree of aerobic biodegradability of plastic materials, including those containing
formulation additives. The test material is exposed in an aqueous medium under laboratory conditions
to an inoculum from activated sludge.
If an unadapted activated sludge is used as the inoculum, the test simulates the biodegradation
processes which occur in a natural aqueous environment; if a mixed or pre-exposed inoculum is used,
the method is used to investigate the potential biodegradability of a test material.
The conditions used in this document do not necessarily correspond to the optimum conditions
allowing maximum biodegradation to occur, but this document is designed to determine the potential
biodegradability of plastic materials or give an indication of their biodegradability in natural
environments.
The method enables the assessment of the biodegradability to be improved by calculating a carbon
balance (optional, see Annex E).
The method applies to the following materials.
— Natural and/or synthetic polymers, copolymers or mixtures thereof.
— Plastic materials which contain additives such as plasticizers, colorants or other compounds.
— Water-soluble polymers.
— Materials which, under the test conditions, do not inhibit the microorganisms present in the
inoculum. Inhibitory effects can be determined using an inhibition control or by another appropriate
[2]
method (see, for example, ISO 8192 ). If the test material is inhibitory to the inoculum, a lower test
concentration, another inoculum or a pre-exposed inoculum can be used.
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 8245, Water quality — Guidelines for the determination of total organic carbon (TOC) and dissolved
organic carbon (DOC)
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
ISO 14851:2019(E)
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 http: //www .electropedia .org/
3.1
ultimate aerobic biodegradation
breakdown of an organic compound by microorganisms in the presence of oxygen into carbon dioxide,
water and mineral salts of any other elements present (mineralization) plus new biomass
3.2
activated sludge
biomass produced in the aerobic treatment of waste water by the growth of bacteria and other
microorganisms in the presence of dissolved oxygen
3.3
concentration of suspended solids in an activated sludge
amount of solids obtained by filtration or centrifugation of a known volume of activated sludge (3.2)
and drying at about 105 °C to constant mass
3.4
biochemical oxygen demand
BOD
mass concentration of the dissolved oxygen consumed under specified conditions by the aerobic
biological oxidation of a chemical compound or organic matter in water
Note 1 to entry: It is expressed as milligrams of oxygen uptake per milligram or gram of test compound.
3.5
theoretical oxygen demand
ThOD
theoretical maximum amount of oxygen required to oxidize a chemical compound completely, calculated
from the molecular formula
Note 1 to entry: It is expressed as milligrams of oxygen uptake per milligram or gram of test compound.
3.6
total organic carbon
TOC
amount of carbon bound in an organic compound
3.7
dissolved organic carbon
DOC
part of the organic carbon in water which cannot be removed by specified phase separation, for example
–2
by centrifugation at 40 000 m·s for 15 min or by membrane filtration using membranes with pores of
0,2 µm to 0,45 µm diameter
3.8
lag phase
time, measured in days, from the start of a test until adaptation and/or selection of the degrading
microorganisms is achieved and the degree of biodegradation of a chemical compound or organic
matter has increased to about 10 % of the maximum level of biodegradation (3.9)
3.9
maximum level of biodegradation
degree of biodegradation, measured in per cent, of a chemical compound or organic matter in a test,
above which no further biodegradation takes place during the test
2 © ISO 2019 – All rights reserved

ISO 14851:2019(E)
3.10
biodegradation phase
time, measured in days, from the end of the lag phase (3.8) of a test until about 90 % of the maximum
level of biodegradation (3.9) has been reached
3.11
plateau phase
time, measured in days, from the end of the biodegradation phase (3.10) until the end of a test
3.12
pre-exposure
pre-incubation of an inoculum in the presence of the chemical compound or organic matter under test,
with the aim of enhancing the ability of the inoculum to biodegrade the test material by adaptation
and/or selection of the microorganisms
3.13
pre-conditioning
pre-incubation of an inoculum under the conditions of the subsequent test in the absence of the chemical
compound or organic matter under test, with the aim of improving the test by acclimatization of the
microorganisms to the test conditions
4 Principle
The biodegradability of a plastic material is determined using aerobic microorganisms in an aqueous
system. The test mixture contains an inorganic medium, the organic test material (the sole source of
carbon and energy) with a concentration between 100 mg/l and 2 000 mg/l of organic carbon, and
activated sludge as the inoculum. The mixture is stirred in closed flasks in a respirometer for a period
not exceeding 2 months. The carbon dioxide evolved is absorbed in a suitable absorber in the headspace
of the flasks. The consumption of oxygen (BOD) is determined, for example by measuring the amount
of oxygen required to maintain a constant volume of gas in the respirometer flasks, or by measuring
the change in volume or pressure (or a combination of the two) either automatically or manually. An
example of a respirometer is given in Annex C. Alternatively, the two-phase closed-bottle version
[3]
described in ISO 10708 may be used (see Annex D).
The level of biodegradation is determined by comparing the BOD with the theoretical amount (ThOD)
and expressed in per cent. The influence of possible nitrification processes on the BOD has to be
considered. The test result is the maximum level of biodegradation determined from the plateau
phase of the biodegradation curve. Optionally, a carbon balance may be calculated to give additional
information on the biodegradation (see Annex E). Moreover, also the absorbed carbon dioxide in the
adsorber at the end of the test may be determined to give additional information on the biodegradation
(see Annex G).
[6]
Unlike ISO 9408 , which is used for a variety of organic compounds, this document is specially designed
for the determination of the biodegradability of plastic materials. The special requirements necessary
affect the choice of the inoculum and the test medium, and there is the possibility of improving the
evaluation of the biodegradability by calculating a carbon balance.
5 Test environment
Incubation shall take place in the dark or in diffuse light in an enclosure which is free from vapours
inhibitory to microorganisms and which is maintained at a constant temperature, preferably between
20 °C and 25 °C, to an accuracy of ±1 °C, or at any other appropriate temperature depending on the
inoculum used and the environment to be assessed.
6 Reagents
Use only reagents of recognized analytical grade.
ISO 14851:2019(E)
6.1 Distilled or deionized water
Distilled or deionized water, free of toxic substances (copper in particular) and containing less than
2 mg/l of DOC.
6.2 Test medium
Depending on the purpose of the test, different test media may be used. For example, if simulating
a natural environment use the standard test medium (6.2.1). If a test material is used at higher
concentrations, use the optimized test medium (6.2.2) with higher buffering capacity and nutrient
concentrations.
6.2.1 Standard test medium
6.2.1.1 Solution A
Dissolve the following in water (6.1) and make up to 1 000 ml.
anhydrous potassium dihydrogen phosphate (KH PO ) 8,5 g
2 4
anhydrous dipotassium hydrogen phosphate (K HPO ) 21,75 g
2 4
disodium hydrogen phosphate dihydrate (Na HPO ·2H O) 33,4 g
2 4 2
ammonium chloride (NH Cl) 0,5 g
The correct composition of the solution can be checked by measuring the pH, which should be 7,4.
6.2.1.2 Solution B
Dissolve 22,5 g of magnesium sulfate heptahydrate (MgSO ·7H O) in water (6.1) and make up to
4 2
1 000 ml.
6.2.1.3 Solution C
Dissolve 36,4 g of calcium chloride dihydrate (CaCl ·2H O) in water (6.1) and make up to 1 000 ml.
2 2
6.2.1.4 Solution D
Dissolve 0,25 g of iron(III) chloride hexahydrate (FeCl ·6H O) in water (6.1) and make up to 1 000 ml.
3 2
Prepare this solution freshly before use to avoid precipitation, or add a drop of concentrated
hydrochloric acid (HCl) or a drop of 0,4 g/l aqueous solution of ethylenediaminetetraacetic acid (EDTA).
6.2.1.5 Preparation
To prepare 1 l of test medium, add the following to about 500 ml of water (6.1),
— 10 ml of solution A;
— 1 ml of each of solutions B to D.
Make up to 1 000 ml with water (6.1). Prepare the test medium freshly before use. The solutions A up to
C may be stored up to 6 months in the dark at room temperature.
6.2.2 Optimized test medium
This optimized medium is highly buffered and contains more inorganic nutrients. This is necessary to
keep the pH constant in the system during the test, even at high concentrations of the test material. The
4 © ISO 2019 – All rights reserved

ISO 14851:2019(E)
medium contains about 2 400 mg/l of phosphorus and 50 mg/l of nitrogen and is therefore suitable
for concentrations in the test material of up to 2 000 mg/l of organic carbon. If higher test-material
concentrations are used, increase the nitrogen content to keep the C:N ratio at about 40:1.
6.2.2.1 Solution A
Dissolve the following in water (6.1) and make up to 1 000 ml.
anhydrous potassium dihydrogen phosphate (KH PO ) 37,5 g
2 4
disodium hydrogen phosphate dihydrate (Na HPO ·2H O) 87,3 g
2 4 2
ammonium chloride (NH Cl) 2,0 g
6.2.2.2 Solution B
Dissolve 22,5 g of magnesium sulfate heptahydrate (MgSO ·7H O) in water (6.1) and make up to
4 2
1 000 ml.
6.2.2.3 Solution C
Dissolve 36,4 g of calcium chloride dihydrate (CaCl ·2H O) in water (6.1) and make up to 1 000 ml.
2 2
6.2.2.4 Solution D
Dissolve 0,25 g of iron(III) chloride hexahydrate (FeCl ·6H O) in water (6.1) and make up to 1 000 ml
3 2
(see second paragraph of 6.2.1.4).
6.2.2.5 Solution E (trace-element solution, optional)
Dissolve in 10 ml of aqueous HCl solution (25 %, 7,7 mol/l), in the following sequence:
70 mg of ZnCl , 100 mg of MnCl ·4H O, 6 mg of H BO , 190 mg of CoCl ·6H O, 3 mg of CuCl ·2H O,
2 2 2 3 3 2 2 2 2
240 mg of NiCl ·6H O, 36 mg of Na MoO ·2H O, 33 mg of Na WO ·2H O and 26 mg of Na SeO ·5H O.
2 2 2 4 2 2 4 2 2 3 2
Make up to 1 000 ml with water (6.1).
6.2.2.6 Solution F (vitamin solution, optional)
Dissolve in 100 ml of water (6.1) 0,6 mg of biotine, 2,0 mg of niacinamide, 2,0 mg of p-aminobenzoate,
1,0 mg of panthotenic acid, 10,0 mg of pyridoxal hydrochloride, 5,0 mg of cyanocobalamine, 2,0 mg of
folic acid, 5,0 mg of riboflavin, 5,0 mg of DL-thioctic acid and 1,0 mg of thiamine dichloride or use a
solution of 15 mg of yeast extract in 100 ml of water (6.1). Filter the solution for sterilization using
membrane filters (see 7.4).
Solutions E and F are optional and are not required if a sufficient concentration of the inoculum is used,
for example, activated sludge. It is recommended that 1 ml portions be prepared and kept refrigerated
until use.
6.2.2.7 Preparation
To prepare 1 l of test medium, add, to about 800 ml of water (6.1):
— 100 ml of solution A;
— 1 ml of each of solutions B to D and, optionally, E and F.
Make up to 1 000 ml with water (6.1) and measure the pH.
ISO 14851:2019(E)
The correct composition of the test medium can be checked by measuring the pH, which should be
7,0 ± 0,2.
6.3 Pyrophosphate solution
Dissolve 2,66 g of anhydrous sodium pyrophosphate (Na P O ) in water (6.1) and make up to 1 000 ml.
4 2 7
6.4 Carbon dioxide absorber
Preferably soda lime pellets or another suitable absorbant.
7 Apparatus
Ensure that all glassware is thoroughly cleaned and, in particular, free from organic or toxic matter.
Required is usual laboratory equipment, plus the following.
7.1 Closed respirometer, including test vessels (glass flasks) fitted with stirrers and all other
necessary equipment, and located in a constant-temperature room or in a thermostated apparatus (e.g.
water-bath). For an example, see Annex C.
NOTE Any respirometer able to determine with sufficient accuracy the biochemical oxygen demand is
suitable, preferably an apparatus which measures and replaces automatically and continuously the oxygen
consumed so that no oxygen deficiency and no inhibition of the microbial activity occurs during the degradation
process. Instead of an ordinary respirometer, the two-phase closed-bottle version can be used (see Annex D).
7.2 Analytical equipment for measuring total organic carbon (TOC) and dissolved organic
carbon (DOC) according to ISO 8245.
7.3 Analytical equipment for measuring nitrate and nitrite concentrations.
A qualitative test is recommended first to decide if any nitrification has occurred. If there is evidence
of nitrate/nitrite in the medium, a quantitative determination using a suitable method (for example ion
chromatography) is required.
7.4 Centrifuge, or filtration device with membrane filters (0,45 µm pore size) which neither adsorb
nor release organic carbon significantly.
7.5 Analytical balance (usual laboratory equipment).
7.6 pH meter (usual laboratory equipment).
8 Procedure
8.1 Test material
The test material shall be of known mass and contain sufficient carbon to yield a BOD that can be
adequately measured by the respirometer used. Calculate from the chemical formula or determine
by elemental analysis the ThOD (see Annex A) and the TOC (using, for example, ISO 8245). Use a test-
material concentration of at least 100 mg/l, corresponding to a ThOD of about 170 mg/l or a TOC of about
60 mg/l. Use lower concentrations only if the sensitivity of the respirometer is adequate. The maximum
amount of test material is limited by the oxygen supply to the respirometer and the test medium used.
When using the optimized test medium (6.2.2), the test-material concentration shall be such that the
TOC does not exceed about 2 000 mg/l, i.e. a C:N ratio of about 40:1. If higher concentrations are to be
tested, increase the amount of nitrogen in the test medium.
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ISO 14851:2019(E)
If biodegradation processes in natural environments are to be simulated, the use of the standard test
medium and a test-material concentration of 100 mg/l are recommended.
The test material should preferably be used in powder form, but it may also be introduced as films,
pieces, fragments or shaped articles. The form and shape of the test material may influence its
biodegradability. Similar shapes should preferably be used if different kinds of plastic material are
to be compared. If the test material is used in the form of a powder, particles of known, narrow size
distribution should be used. A particle-size distribution with the maximum at 250 µm diameter is
recommended. Also, the size of the test equipment used may depend on the form of the test material.
It should be ascertained that no substantial mechanical aberrations occur due to the test conditions,
for example due to the type of stirring mechanism used. Processing of the test material (e.g. the use of
powder in the case of composites) should not influence significantly the degradation behaviour of the
material. Optionally, record the hydrogen, oxygen, nitrogen, phosphorus and sulfur contents and the
molecular mass of a polymeric test material, using for example liquid exclusion chromatography (see,
[1]
for example, ASTM D 3536–91 or any other applicable standard method). Preferably, plastic materials
without additives such as plasticizers should be tested. When the material does contain such additives,
information on their biodegradability will be needed to assess the biodegradability of the polymeric
material itself.
[7]
For details on how to handle poorly water-soluble compounds, see ISO 10634 .
8.2 Reference material
Use aniline and/or a well-defined biodegradable polymer (for example microcrystalline cellulose
powder, ashless cellulose filters or poly-β-hydroxybutyrate) as a reference material. If possible, the
TOC, form and size should be comparable to that of the test material.
As a negative control, a non-biodegradable polymer (e.g. polyethylene) in the same form as the test
material can optionally be used.
8.3 Preparation of the inoculum
Activated sludge from a sewage-treatment plant treating predominantly domestic sewage is a suitable
source of the inoculum. It is obtained from an active aerobic environment and is available over a wide
geographical area in which a broad range of plastic materials needs to be tested. When biodegradation
in a specific waste-treatment system is to be determined, collect the inoculum from that environment.
Take a sample of activated sludge collected from a well-operated sewage-treatment plant or a laboratory
plant handling predominantly domestic sewage. Mix well, keep the sample under aerobic conditions
and use preferably on the day of collection (at least within 72 h).
[4]
Before use, determine the concentration of suspended solids (use, for example, ISO 11923 ). If
necessary, concentrate the sludge by settling so that the volume of sludge added to the test assay is
minimal. Add a suitable volume to obtain suspended solids in the range 30 mg/l to 1 000 mg/l in the
final mixture.
If the endogenous respiration of the inoculum is too high, stabilize the inoculum by aeration before use.
Harmonize the test temperature with the inoculum used.
It may be useful to determine the colony-forming units (cfu) of the inoculum used. The test mixture
3 6
should preferably contain about 10 cfu/ml to 10 cfu/ml.
When biodegradation processes in a natural environment are to be simulated or when a carbon balance
determination (see Annex E) is to be carried out, an inoculum concentration of 30 mg/l suspended
solids is recommended. As solid matter can interfere with the carbon balance determination, the
following procedure for preparing the inoculum is recommended. Take 500 ml of the activated sludge
and homogenize for 2 min at medium speed in a blender or in a suitable high-speed mixer. Allow to
settle until the supernatant liquid contains no significant amounts of suspended matter, but in any case
for at least 30 min. Decant a sufficient volume of the supernatant liquid and add it to the test flasks to
ISO 14851:2019(E)
obtain a concentration of a volume fraction of 1 % to a volume fraction of 5 % in the test medium. Avoid
carrying over sludge particles.
An inoculum may be pre-conditioned, but normally no pre-exposed inoculum should be used, especially
in the case of standard tests simulating biodegradation behaviour in natural environments. Depending
on the purpose of the test, a pre-exposed inoculum may also be used, provided this is clearly stated
in the test report (e.g. per cent biodegradation = x %, using pre-exposed inocula) and the method of
pre-exposure detailed in the test report. Pre-exposed inocula can be obtained from suitable laboratory
[8]
biodegradation tests (see ISO/TR 15462 ) conducted under a variety of conditions or from samples
collected from locations where relevant environmental conditions exist (e.g. contaminated areas or
industrial treatment plants).
8.4 Test
Provide a number of flasks, so that the test includes at least the following:
a) three test flasks for the test material (symbol F );
T
b) three test flasks for the blank control (symbol F );
B
c) two test flasks for checking the inoculum activity using a reference material (symbol F ).
C
And, if required:
d) One flask for checking for possible abiotic degradation or non-biological change in the test material
such as by hydrolysis (symbol F ). The test solution in F shall be sterilized, for example by
S S
autoclaving or by the addition of a suitable inorganic toxic compound to prevent microbial activity.
Use, for example, 5 ml/l of a solution containing 10 g/l of mercury(II) chloride (HgCl ). Add the
same amount of the toxic substance during the test if required.
e) One flask as a negative control (symbol F ) using a non-biodegradable polymeric substance (e.g.
N
polyethylene) in the same form as the test material.
f) One flask for checking the possible inhibiting effect of the test material on microbial activity
(symbol F ). Take care that the ratio of carbon in the test and reference material to nitrogen in the
I
medium is at least about C:N = 40:1. Add nitrogen if required.
Add appropriate amounts of the test medium (6.2) and the inoculum (see 8.3) to the test flasks as
indicated in Table 1.
Measure the pH in the flasks and adjust to 7 if necessary. Add carbon dioxide absorber (6.4) to the
absorber compartments of the respirometer (see Annex C). Add the test material (see 8.1), the reference
material and the material for the negative control (see 8.2) to the respective flasks as indicated in Table 1.
If a carbon balance is to be run (see Annex E), remove a known sufficient volume of the inoculated test
medium from each flask or from additional separate flasks for DOC and biomass determination at the
beginning and the end of the incubation period. Consider the removed volume when adjusting the final
volume or when calculating the test results.
Place the flasks in a constant-temperature environment (see Clause 5) and allow all vessels to reach the
desired temperature. Make any necessary connections, seal the flasks, place them in the respirometer
and start the stirrer.
Take the necessary readings on the manometers (if manual) and verify that the recorder of oxygen
consumption is functioning properly (automatic respirometer). As an alternative, the two-phase closed-
bottle version described in Annex D may be used.
8 © ISO 2019 – All rights reserved

ISO 14851:2019(E)
Table 1 — Final distribution of test and reference materials
Reference mate-
Flask Test material Inoculum
rial
F Test + – +
T
F Test + – +
T
F Test + – +
T
F Blank – – +
B
F Blank – – +
B
F Blank – – +
B
F Inoculum check – + +
C
F Inoculum check – + +
C
F Abiotic degradation check (optional) + – –
S
F Inhibition control (optional) + + +
I
F Negative control (optional) – + +
N
When a constant level of BOD is attained (plateau phase reached) and no further biodegradation is
expected, the test is considered to be completed. The test period should not typically exceed 2 months.
However, if significant biodegradation is still observed and the plateau phase has not been reached
after this length of time, then the test may be extended, but not longer than 6 months. In the case of long
test durations, special attention shall be paid to the technical system (e.g. tightness of the test vessels
and connections).
At the end of the test, measure the pH and determine the concentrations of nitrate and nitrite
immediately in flasks F (see paragraph below), or take suitably preserved samples. Use the values to
T
correct the calculated degree of biodegradation for nitrification (see Annex B).
Allylthiourea can only inhibit nitrification during short incubation periods, as it is biodegradable.
Therefore, addition of allylthiourea to prevent nitrification is not recommended. Experience shows,
however, that with low inoculum concentrations (about a volume fraction of 1 %) nitrification will not
occur, even during long incubation periods, when no inhibitor is used.
Optionally determine additionally the absorbed carbon dioxide in the adsorber at the end of the test
(see Annex G).
9 Calculation and expression o
...