Plastics - Determination of the ultimate anaerobic biodegradation of plastic materials in an aqueous system - Method by measurement of biogas production (ISO 14853:2016)

ISO 14853:2016 specifies a method for the determination of the ultimate anaerobic biodegradability of plastics by anaerobic microorganisms. The conditions described in ISO 14853 do not necessarily correspond to the optimum conditions for the maximum degree of biodegradation to occur. The test calls for exposure of the test material to sludge for a period of up to 90 d, which is longer than the normal sludge retention time (25 to 30 d) in anaerobic digesters, although digesters at industrial sites can have much longer retention times.
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 e.g. ISO 13641). If the test material is inhibitory to the inoculum, a lower test concentration, another inoculum or a pre-exposed inoculum can be used.

Kunststoffe - Bestimmung der vollständigen anaeroben Bioabbaubarkeit von Kunststoff-Materialien in einem wässrigen Medium - Verfahren mittels Analyse der Biogasentwicklung (ISO 14853:2016)

Plastiques - Évaluation de la biodégradabilité anaérobie ultime des matériaux plastiques en milieu aqueux - Méthode par détermination de la production de biogaz (ISO 14853:2016)

L'ISO 14853:2016 spécifie une méthode pour la détermination de la biodégradabilité anaérobie ultime des plastiques par des micro-organismes anaérobies. Les conditions décrites dans l'ISO 14853:2016  ne correspondent pas nécessairement aux conditions optimales permettant d'obtenir le taux maximal de biodégradation. L'essai exige que le matériau d'essai soit exposé aux boues pendant une période allant jusqu'à 90 j, ce qui est plus long que le temps de rétention normal de la boue (25 j à 30 j) dans les digesteurs anaérobies, bien que les digesteurs sur les sites industriels puissent avoir des temps de rétention beaucoup plus longs.
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 autres composés;
-      polymères hydrosolubles;
-      matériaux qui, dans les conditions d'essai, n'ont pas d'effet inhibiteur sur les micro-organismes présents dans l'inoculum. Les effets inhibiteurs peuvent être déterminés en utilisant une substance de contrôle de l'effet inhibiteur ou par toute autre méthode appropriée (voir, par exemple, l'ISO 13641). 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.

Polimerni materiali - Ugotavljanje dokončne anaerobne biodegradacije plastičnih materialov v vodnem sistemu - Metoda z merjenjem proizvodnje bioplina (ISO 14853:2016)

Standard ISO 14853:2016 določa metodo za ugotavljanje končne anaerobne biorazgradljivosti polimernih materialov, ki jo povzročajo anaerobni mikroorganizmi. Pogoji, ki so opisani v standardu ISO 14853, niso nujno enaki idealnim pogojem, pri katerih pride do največje stopnje biorazgradljivosti. Preskus zahteva izpostavljenost preskusnega materiala blatu do 90 d, kar je dlje od običajnega časa hranjenja blata (25 do 30 d) v anaerobnih gniliščih, čeprav imajo lahko gnilišča na industrijskih površinah veliko daljši čas hranjenja.
Ta 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 nadzora zaviranja ali z drugo ustrezno metodo (glej npr. standard ISO 13641). Če preskusni material zavira inokulum, se lahko uporabi manjša preskusna koncentracija, drug inokulum ali predhodno izpostavljeni inokulum.

General Information

Status
Published
Public Enquiry End Date
31-Aug-2017
Publication Date
08-Jan-2018
Current Stage
6060 - National Implementation/Publication (Adopted Project)
Start Date
19-Dec-2017
Due Date
23-Feb-2018
Completion Date
09-Jan-2018

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Standards Content (Sample)

SLOVENSKI STANDARD
SIST EN ISO 14853:2018
01-februar-2018
3ROLPHUQLPDWHULDOL8JRWDYOMDQMHGRNRQþQHDQDHUREQHELRGHJUDGDFLMHSODVWLþQLK
PDWHULDORYYYRGQHPVLVWHPX0HWRGD]PHUMHQMHPSURL]YRGQMHELRSOLQD ,62

Plastics - Determination of the ultimate anaerobic biodegradation of plastic materials in
an aqueous system - Method by measurement of biogas production (ISO 14853:2016)
Kunststoffe - Bestimmung der vollständigen anaeroben Bioabbaubarkeit von Kunststoff-
Materialien in einem wässrigen Medium - Verfahren mittels Analyse der
Biogasentwicklung (ISO 14853:2016)
Plastiques - Évaluation de la biodégradabilité anaérobie ultime des matériaux plastiques
en milieu aqueux - Méthode par détermination de la production de biogaz (ISO
14853:2016)
Ta slovenski standard je istoveten z: EN ISO 14853:2017
ICS:
83.080.01 Polimerni materiali na Plastics in general
splošno
SIST EN ISO 14853:2018 en,fr,de
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

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SIST EN ISO 14853:2018

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SIST EN ISO 14853:2018


EN ISO 14853
EUROPEAN STANDARD

NORME EUROPÉENNE

December 2017
EUROPÄISCHE NORM
ICS 83.080.01
English Version

Plastics - Determination of the ultimate anaerobic
biodegradation of plastic materials in an aqueous system -
Method by measurement of biogas production (ISO
14853:2016)
Plastiques - Évaluation de la biodégradabilité Kunststoffe - Bestimmung der vollständigen anaeroben
anaérobie ultime des matériaux plastiques en milieu Bioabbaubarkeit von Kunststoff-Materialien in einem
aqueux - Méthode par détermination de la production wässrigen Medium - Verfahren mittels Analyse der
de biogaz (ISO 14853:2016) Biogasentwicklung (ISO 14853:2016)
This European Standard was approved by CEN on 17 October 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 14853:2017 E
worldwide for CEN national Members.

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SIST EN ISO 14853:2018
EN ISO 14853:2017 (E)
Contents Page
European foreword . 3

2

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SIST EN ISO 14853:2018
EN ISO 14853:2017 (E)
European foreword
The text of ISO 14853:2016 has been prepared by Technical Committee ISO/TC 61 “Plastics” of the
International Organization for Standardization (ISO) and has been taken over as EN ISO 14853:2017 by
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 June 2018, and conflicting national standards shall be
withdrawn at the latest by June 2018.
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.
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 14853:2016 has been approved by CEN as EN ISO 14853:2017 without any modification.

3

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SIST EN ISO 14853:2018

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SIST EN ISO 14853:2018
INTERNATIONAL ISO
STANDARD 14853
Second edition
2016-07-15
Plastics — Determination of the
ultimate anaerobic biodegradation of
plastic materials in an aqueous system
— Method by measurement of biogas
production
Plastiques — Évaluation de la biodégradabilité anaérobie ultime des
matériaux plastiques en milieu aqueux — Méthode par détermination
de la production de biogaz
Reference number
ISO 14853:2016(E)
©
ISO 2016

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SIST EN ISO 14853:2018
ISO 14853:2016(E)

COPYRIGHT PROTECTED DOCUMENT
© ISO 2016, 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. +41 22 749 01 11
Fax +41 22 749 09 47
copyright@iso.org
www.iso.org
ii © ISO 2016 – All rights reserved

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SIST EN ISO 14853:2018
ISO 14853:2016(E)

Contents Page
Foreword .v
Introduction .vi
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Principle . 3
5 Reagents and materials . 3
6 Apparatus . 5
6.1 Laboratory equipment . 5
6.2 Apparatus for use when biogas is measured by a manometric method . 6
6.3 Apparatus for use when biogas is measured by a volumetric method . 6
7 Procedure. 6
7.1 General . 6
7.2 Digested sludge . 6
7.3 Preparation of the inoculum . 7
7.4 Preparation of test suspensions and controls . 7
7.5 Incubation and gas measurement . 8
7.6 Test duration . 9
7.7 Measurement of inorganic carbon . 9
7.8 Specific analyses . 9
8 Calculation and expression of results . 9
8.1 Amount of carbon in headspace . 9
8.2 Calculation of amount of carbon in headspace when manometric measurement
method is used .10
8.3 Calculation of amount of carbon in headspace when volumetric measurement
method is used .11
8.4 Amount of inorganic carbon in the liquid .11
8.5 Total amount of carbon converted to gas .11
8.6 Amount of carbon in test material .12
8.7 Calculation of percentage biodegradation .12
9 Validity of results .12
9.1 Maintenance of anaerobic conditions .12
9.2 Inhibition of degradation .12
9.3 Validity of the test .12
10 Test report .13
Annex A (informative) Example of apparatus for determining the amount of biogas
produced by measuring the increase in gas pressure .14
Annex B (informative) Example of apparatus for determining volumetrically the amount of
biogas produced .15
Annex C (informative) Example of a biodegradation curve .17
Annex D (informative) Examples of data sheets for anaerobic biodegradability tests .18
Annex E (informative) Table of water vapour pressures at various temperatures .21
Annex F (informative) Calculation of theoretical carbon dioxide (ThCO ) and theoretical
2
methane (ThCH ) production .22
4
Annex G (informative) Example of determination of recovery rate .23
Annex H (informative) Example of a workflow scheme .26
© ISO 2016 – All rights reserved iii

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ISO 14853:2016(E)

Bibliography .28
iv © ISO 2016 – All rights reserved

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SIST EN ISO 14853:2018
ISO 14853:2016(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 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 5, Physical-
chemical properties.
This second edition cancels and replaces the first edition (ISO 14853:2005), which has been technically
revised. It also incorporates the Technical Corrigendum ISO 14853:2005/Cor.1:2009.
© ISO 2016 – All rights reserved v

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SIST EN ISO 14853:2018
ISO 14853:2016(E)

Introduction
With the increasing use of plastics, their recovery and disposal have become a major issue. As a first
priority, recovery should be promoted. For example, plastic litter, which originates mainly from
consumers, is difficult to recover completely. Additional examples of materials difficult to recover
are found in the disposal of fishing tackle, agricultural mulch films and water-soluble polymers.
These plastic materials tend to leak from closed waste management infrastructures into natural
environments. Biodegradable plastics are now emerging as one of the available options to solve such
environmental issues. Plastic materials, such as products or packaging, which are sent to anaerobic
treatment facilities should be potentially biodegradable. Therefore, it is very important to determine
the potential biodegradability of such materials and to obtain a quantitative measure of their
biodegradability in anaerobic environments.
vi © ISO 2016 – All rights reserved

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SIST EN ISO 14853:2018
INTERNATIONAL STANDARD ISO 14853:2016(E)
Plastics — Determination of the ultimate anaerobic
biodegradation of plastic materials in an aqueous system
— Method by measurement of biogas production
WARNING — Sewage and activated sludge may contain potentially pathogenic organisms.
Therefore, appropriate precautions should be taken when handling them. Digesting sewage
sludge produces flammable gases which present fire and explosion risks. Care should be taken
when transporting and storing quantities of digesting sludge. Toxic test chemicals and those
whose properties are not known should be handled with care and in accordance with safety
instructions. The pressure meter and microsyringes should be handled carefully to avoid needle
stick injuries. Contaminated syringe needles should be disposed of in a safe manner.
1 Scope
This International Standard specifies a method for the determination of the ultimate anaerobic
biodegradability of plastics by anaerobic microorganisms. The conditions described in this
International Standard do not necessarily correspond to the optimum conditions for the maximum
degree of biodegradation to occur. The test calls for exposure of the test material to sludge for a period
of up to 90 d, which is longer than the normal sludge retention time (25 to 30 d) in anaerobic digesters,
although digesters at industrial sites can have much longer retention times.
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 e.g. ISO 13641). 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
There are no normative references in this document.
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1
ultimate anaerobic biodegradation
breakdown of an organic compound by microorganisms in the absence of oxygen to carbon dioxide,
methane, water and mineral salts of any other elements present (mineralization) plus new biomass
3.2
primary anaerobic biodegradation
structural change (transformation) of a chemical compound by microorganisms, resulting in the loss of
a specific property
© ISO 2016 – All rights reserved 1

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SIST EN ISO 14853:2018
ISO 14853:2016(E)

3.3
digested sludge
mixture of settled sewage and activated sludge which have been incubated in an anaerobic digester at
about 35 °C to reduce the biomass and odour and to improve the dewaterability of the sludge
Note 1 to entry: Digested sludge contains an association of anaerobic fermentative and methanogenic bacteria
producing carbon dioxide and methane.
3.4
concentration of suspended solids in digested sludge
amount of solids obtained by filtration or centrifugation of a known volume of activated sludge and
drying at about 105 °C to constant mass
3.5
dissolved organic carbon
DOC
organic carbon in the water phase 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
3.6
inorganic carbon
IC
inorganic carbon which is dissolved or dispersed in the aqueous phase of a liquid and is recoverable
from the supernatant liquid after the sludge has been allowed to settle
3.7
total dry solids
amount of solids obtained by taking a known volume of test material or inoculum and drying at about
105 °C to constant mass
3.8
theoretical amount of evolved biogas
Thbiogas
maximum theoretical amount of biogas (CH + CO ) evolved after complete biodegradation of an
4 2
organic material under anaerobic conditions, calculated from the molecular formula and expressed as
millilitres of biogas evolved per milligram of test material under standard conditions
3.9
theoretical amount of evolved carbon dioxide
ThCO
2
maximum theoretical amount of carbon dioxide evolved after complete oxidation of an organic material,
calculated from the molecular formula and expressed as milligrams of carbon dioxide per milligram of
test material
3.10
theoretical amount of evolved methane
ThCH
4
maximum theoretical amount of methane evolved after complete reduction of an organic material,
calculated from the molecular formula and expressed as milligrams of methane evolved per milligram
of test material
3.11
lag phase
lag period
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
2 © ISO 2016 – All rights reserved

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ISO 14853:2016(E)

3.12
plateau phase
time, measured in days, from the end of the biodegradation phase until the end of the test
3.13
biodegradation phase
time, measured in days, from the end of the lag phase of a test until about 90 % of the maximum level of
biodegradation has been reached
3.14
maximum level of biodegradation
degree of biodegradation, measured in percent, of a chemical compound or organic matter in a test,
above which no further biodegradation takes place during the test
4 Principle
The biodegradability of a plastic material is determined using anaerobic conditions in an aqueous
system. Test material with a concentration of 20 mg/l to 200 mg/l organic carbon (OC) is incubated
at (35 ± 2) °C in sealed vessels together with digested sludge for a period normally not exceeding 90 d.
Before use, the digested sludge is washed so that it contains very low amounts of inorganic carbon
(IC) and diluted to 1 g/l to 3 g/l total solids concentration. The increase in headspace pressure or the
volumetric increase (depending on the method used for measuring biogas evolution) in the test vessels
resulting from the production of carbon dioxide (CO ) and methane (CH ) is measured. A considerable
2 4
amount of CO will be dissolved in water or transformed to bicarbonate or carbonate under the
2
conditions of the test. This inorganic carbon (IC) is measured at the end of the test. The amount of
microbiologically produced biogas carbon is calculated from the net biogas production and the net IC
formation in excess of blank values. The percentage biodegradation is calculated from the total amount
of carbon transformed to biogas and IC and the measured or calculated amount of carbon added as
test material. The course of biodegradation can be followed by making intermediate measurements
of biogas production. As additional information, the primary biodegradability can be determined by
specific analyses at the beginning and end of the test.
This test method is designed to determine the biodegradability of plastic materials under anaerobic
conditions. Optionally, the assessment of the recovery rate may also be of interest (see Annex G).
5 Reagents and materials
5.1 Distilled or deionized water, free of toxic substances, containing less than 2 mg/l of DOC.
5.2 Test medium, prepared using only reagents of recognized analytical grade.
Prepare the test medium to contain the following constituents in the stated amounts:
Anhydrous potassium dihydrogen phosphate KH PO 0,27 g
2 4
Disodium hydrogen phosphate dodecahydrate Na HPO ⋅12H O 1,12 g
2 4 2
Ammonium chloride NH Cl 0,53 g
4
Calcium chloride dihydrate CaCl ⋅2H O 0,075 g
2 2
Magnesium chloride hexahydrate MgCl ⋅6H O 0,10 g
2 2
Iron (II) chloride tetrahydrate FeCl ⋅4H O 0,02 g
2 2
Resazurin (oxygen indicator) 0,001 g
a
Disodium sulfide nonahydrate Na S⋅9H O 0,1 g
2 2
Stock solution of trace elements (optional) 10 ml
Stock solutions of vitamins (optional) Vitamin solution No. 1 0,5 ml
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SIST EN ISO 14853:2018
ISO 14853:2016(E)

Anhydrous potassium dihydrogen phosphate KH PO 0,27 g
2 4
Vitamin solution No. 2 0,5 ml
Add water (5.1) (oxygen-free) to 1 l
a
  Use freshly prepared sodium sulfide, or wash and dry it before use, to ensure sufficient reductive capacity.
In order to ensure strictly anaerobic conditions, it is recommended that a small amount of sodium dithionite be
added to the medium after it has been prepared until it becomes colourless. Do not use more than 10 mg/l because
higher concentrations may produce inhibitory effects.
Adjust the pH of the medium with dilute mineral acid or alkali, if necessary, to 7 ± 0,2.
To ensure oxygen-free conditions, purge the water with nitrogen for about 20 min immediately before use.
5.3 Trace-element solution (optional).
It is recommended that the test medium be supplemented with the following trace elements to improve
the anaerobic degradation process, especially if low inoculum concentrations are used:
Manganese chloride tetrahydrate MnCl ⋅4H O 0,05 g
2 2
Boric acid H BO 0,005 g
3 3
Zinc chloride ZnCl 0,005 g
2
Copper (II) chloride CuCl 0,003 g
2
Disodium molybdate dihydrate Na MoO ⋅2H O 0,001 g
2 4 2
Cobalt chloride hexahydrate CoCl ⋅6H O 0,1 g
2 2
Nickel chloride hexahydrate NiCl ⋅6H O 0,01 g
2 2
Disodium selenite Na SeO 0,005 g
2 3
Disodium tungstate dihydrate Na WO ⋅2H O 0,002 g
2 4 2
Add water (5.1) (oxygen free) to 1 l
Use 10 ml of trace-element solution per litre of test medium.
5.4 Vitamin solutions (optional).
5.4.1 Vitamin solution No. 1
4-Aminobenzoic acid 40 mg
d-Biotin 10 mg
Dissolve in hot water (5.1) 500 ml
Allow to cool and add:
d-Pantothenic acid, calcium salt 50 mg
Pyridoxamine dihydrochloride 150 mg
Thiamine dichloride 100 mg
Filter the solution through a membrane filter (pore size 0,45 µm) that neither adsorbs nor releases
organic carbon in significant amounts, and store in the dark at 4 °C.
Use 0,5 ml of vitamin solution per litre of test medium.
5.4.2 Vitamin solution No. 2
Cyanocobalamin (vitamin B12) 10 mg
Dissolve in water (5.1) 100 ml
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ISO 14853:2016(E)

Filter the solution through a membrane filter (pore size 0,45 µm) that neither adsorbs nor releases
organic carbon in significant amounts, and store in the dark at 4 °C.
Use 0,5 ml of vitamin solution per litre of test medium.
5.5 Barrier solution.
NaCl 200 g
Dissolve in water (5.1) 1 000 ml
Acidify with citric acid 5 g
Add a pH indicator such as bromophenol blue or methyl orange in order to be able to verify that the
solution remains acid during the test.
5.6 Test material.
The test material is usually added directly as solid to give a concentration of 20 mg/l to 200 mg/l
organic carbon. The test material (plastic) should be used in powdered form, if possible.
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
biodegrad
...

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