Soil quality - Determination of potential nitrification and inhibition of nitrification - Rapid test by ammonium oxidation (ISO 15685:2012)

This International Standard specifies a rapid method for the determination of the potential rate of ammonium
oxidation and inhibition of nitrification in soils. This method is suitable for all soils containing a population of
nitrifying microorganisms. It can be used as a rapid screening test for monitoring soil quality and quality of
wastes, and is suitable for testing the effects of cultivation methods, chemical substances [except volatiles,
i.e. H > 1 (Henry’s constant)], extracts of biosolids and pollution in soils.

Bodenbeschaffenheit - Bestimmung der potentiellen Nitrifizierung und Hemmung der Nitrifizierung - Schnellverfahren mittels Ammoniumoxidation (ISO 15685:2012)

Diese Internationale Norm legt ein Schnellverfahren zur Bestimmung der potentiellen Ammoniumoxidation und der Hemmung der Nitrifizierung in Böden fest. Dieses Verfahren ist geeignet für die Anwendung bei allen Böden, die eine Population an nitrifizierenden Mikroorganismen enthalten. Es kann als schnelles Übersichtsverfahren zur Überwachung der Bodenbeschaffenheit und der Beschaffenheit von Abfällen verwendet werden und ist für die Prüfung der Einflüsse von Kultivierungsverfahren, zusätzlichen Chemikalien [außer flüchtige Substanzen, d. h. H > 1 (Henry-Konstante)], Eluaten von Biofeststoffen und Verunreinigungen von Böden geeignet.

Qualité du sol - Détermination de la nitrification potentielle et inhibition de la nitrification - Essai rapide par oxydation de l'ammonium (ISO 15685:2012)

L'ISO 15685:2012 spécifie une méthode rapide de détermination de la vitesse potentielle d'oxydation de l'ammonium et d'inhibition de la nitrification des sols. Cette méthode convient à tous les sols contenant une population de micro-organismes nitrifiants. Elle peut être utilisée comme essai de criblage rapide pour surveiller la qualité des sols et la qualité des déchets et elle convient pour l'évaluation des effets des pratiques agricoles, des substances chimiques [sauf les composés volatils c'est-à-dire H > 1 (constante de Henry)], des extraits de biosolides et de la pollution des sols.

Kakovost tal - Določevanje potencialne nitrifikacije in zaviranja nitrifikacije - Hitri preskus z oksidacijo amonija (ISO 15685:2012)

General Information

Status
Published
Public Enquiry End Date
02-Feb-2020
Publication Date
23-Sep-2020
Technical Committee
Current Stage
6060 - National Implementation/Publication (Adopted Project)
Start Date
25-May-2020
Due Date
30-Jul-2020
Completion Date
24-Sep-2020

Buy Standard

Standard
EN ISO 15685:2020
English language
18 pages
sale 10% off
Preview
sale 10% off
Preview
e-Library read for
1 day
Draft
prEN ISO 15685:2020
English language
15 pages
sale 10% off
Preview
sale 10% off
Preview
e-Library read for
1 day

Standards Content (Sample)

SLOVENSKI STANDARD
SIST EN ISO 15685:2020
01-november-2020
Kakovost tal - Določevanje potencialne nitrifikacije in zaviranja nitrifikacije - Hitri
preskus z oksidacijo amonija (ISO 15685:2012)
Soil quality - Determination of potential nitrification and inhibition of nitrification - Rapid
test by ammonium oxidation (ISO 15685:2012)
Bodenbeschaffenheit - Bestimmung der potentiellen Nitrifizierung und Hemmung der
Nitrifizierung - Schnellverfahren mittels Ammoniumoxidation (ISO 15685:2012)
Qualité du sol - Détermination de la nitrification potentielle et inhibition de la nitrification -
Essai rapide par oxydation de l'ammonium (ISO 15685:2012)
Ta slovenski standard je istoveten z: EN ISO 15685:2020
ICS:
13.080.30 Biološke lastnosti tal Biological properties of soils
SIST EN ISO 15685:2020 en,fr,de
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

---------------------- Page: 1 ----------------------
SIST EN ISO 15685:2020

---------------------- Page: 2 ----------------------
SIST EN ISO 15685:2020


EN ISO 15685
EUROPEAN STANDARD

NORME EUROPÉENNE

May 2020
EUROPÄISCHE NORM
ICS 13.080.30
English Version

Soil quality - Determination of potential nitrification and
inhibition of nitrification - Rapid test by ammonium
oxidation (ISO 15685:2012)
Qualité du sol - Détermination de la nitrification Bodenbeschaffenheit - Bestimmung der potentiellen
potentielle et inhibition de la nitrification - Essai rapide Nitrifizierung und Hemmung der Nitrifizierung -
par oxydation de l'ammonium (ISO 15685:2012) Schnellverfahren mittels Ammoniumoxidation (ISO
15685:2012)
This European Standard was approved by CEN on 13 April 2020.

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, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway,
Poland, Portugal, Republic of North Macedonia, 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
© 2020 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN ISO 15685:2020 E
worldwide for CEN national Members.

---------------------- Page: 3 ----------------------
SIST EN ISO 15685:2020
EN ISO 15685:2020 (E)
Contents Page
European foreword . 3

2

---------------------- Page: 4 ----------------------
SIST EN ISO 15685:2020
EN ISO 15685:2020 (E)
European foreword
The text of ISO 15685:2012 has been prepared by Technical Committee ISO/TC 190 "Soil Quality” of the
International Organization for Standardization (ISO) and has been taken over as EN ISO 15685:2020 by
Technical Committee CEN/TC 444 “Environmental characterization of solid matrices” the secretariat of
which is held by NEN.
This European Standard shall be given the status of a national standard, either by publication of an
identical text or by endorsement, at the latest by November 2020, and conflicting national standards
shall be withdrawn at the latest by November 2020.
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, France, Germany, Greece, Hungary, Iceland,
Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Republic of
North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the
United Kingdom.
Endorsement notice
The text of ISO 15685:2012 has been approved by CEN as EN ISO 15685:2020 without any modification.

3

---------------------- Page: 5 ----------------------
SIST EN ISO 15685:2020

---------------------- Page: 6 ----------------------
SIST EN ISO 15685:2020
INTERNATIONAL ISO
STANDARD 15685
Second edition
2012-07-15
Soil quality — Determination of potential
nitrification and inhibition of nitrification —
Rapid test by ammonium oxidation
Qualité du sol — Détermination de la nitrification potentielle et inhibition
de la nitrification — Essai rapide par oxydation de l’ammonium
Reference number
ISO 15685:2012(E)
©
ISO 2012

---------------------- Page: 7 ----------------------
SIST EN ISO 15685:2020
ISO 15685:2012(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2012
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means,
electronic or mechanical, including photocopying and microfilm, without permission in writing from either ISO at the address below or ISO’s
member body in the country of the requester.
ISO copyright office
Case postale 56 • CH-1211 Geneva 20
Tel. + 41 22 749 01 11
Fax + 41 22 749 09 47
E-mail copyright@iso.org
Web www.iso.org
Published in Switzerland
ii © ISO 2012 – All rights reserved

---------------------- Page: 8 ----------------------
SIST EN ISO 15685:2020
ISO 15685:2012(E)
Contents Page
Foreword .iv
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Principle . 2
5 Reagents . 2
6 Apparatus . 3
7 Sampling, storage and characterization of samples . 3
8 Procedure . 4
8.1 Testing soils . 4
8.2 Testing the effect of chemicals . 4
8.3 Testing polluted soils . 4
8.4 Testing water extracts of biosolids . 5
9 Calculations . 5
10 Test report . 5
Annex A (informative) Results of a laboratory ring test . 7
Annex B (informative) Micro-scale measurement of potential nitrification rates. 9
Bibliography . 11
© ISO 2012 – All rights reserved iii

---------------------- Page: 9 ----------------------
SIST EN ISO 15685:2020
ISO 15685:2012(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.
International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2.
The main task of technical committees is to prepare International Standards. Draft International Standards
adopted by the technical committees are circulated to the member bodies for voting. Publication as an
International Standard requires approval by at least 75 % of the member bodies casting a vote.
Attention is drawn to the possibility that some of the elements of this document may be the subject of patent
rights. ISO shall not be held responsible for identifying any or all such patent rights.
ISO 15685 was prepared by Technical Committee ISO/TC 190, Soil quality, Subcommittee SC 4, Biological methods.
This second edition cancels and replaces the first edition (ISO 15685:2004), which has been technically revised.
iv © ISO 2012 – All rights reserved

---------------------- Page: 10 ----------------------
SIST EN ISO 15685:2020
INTERNATIONAL STANDARD ISO 15685:2012(E)
Soil quality — Determination of potential nitrification and
inhibition of nitrification — Rapid test by ammonium oxidation
1 Scope
This International Standard specifies a rapid method for the determination of the potential rate of ammonium
oxidation and inhibition of nitrification in soils. This method is suitable for all soils containing a population of
nitrifying microorganisms. It can be used as a rapid screening test for monitoring soil quality and quality of
wastes, and is suitable for testing the effects of cultivation methods, chemical substances [except volatiles,
i.e. H > 1 (Henry’s constant)], extracts of biosolids and pollution in soils.
2 Normative references
The following referenced documents are indispensable for the application 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 10381-6, Soil quality — Sampling — Part 6: Guidance on the collection, handling and storage of soil under
aerobic conditions for the assessment of microbiological processes, biomass and diversity in the laboratory
ISO 10390, Soil quality — Determination of pH
ISO 10694, Soil quality — Determination of organic and total carbon after dry combustion (elementary analysis)
ISO 11260, Soil quality — Determination of effective cation exchange capacity and base saturation level using
barium chloride solution
ISO 11261, Soil quality — Determination of total nitrogen — Modified Kjeldahl method
ISO 11277, Soil quality — Determination of particle size distribution in mineral soil material — Method by
sieving and sedimentation
ISO 11465, Soil quality — Determination of dry matter and water content on a mass basis — Gravimetric method
ISO 14238, Soil quality — Biological methods — Determination of nitrogen mineralization and nitrification in
soils and the influence of chemicals on these processes
ISO 14256-2, Soil quality — Determination of nitrate, nitrite and ammonium in field-moist soils by extraction
with potassium chloride solution — Part 2: Automated method with segmented flow analysis
EN 14735, Characterization of waste — Preparation of waste samples for ecotoxicity tests
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1
inhibitory dose
ID
amount of a chemical added to soil that effectively inhibits biological activity by a stated percentage after a
given time, in comparison with an untreated control
NOTE It is expressed as a percentage. For example, ID25 and ID50 indicate a 25 % and 50 % inhibition of biological
activity, respectively.
© ISO 2012 – All rights reserved 1

---------------------- Page: 11 ----------------------
SIST EN ISO 15685:2020
ISO 15685:2012(E)
3.2
biosolids
organic products used in agriculture, which include sewage sludge, compost, manure and some industrial wastes
4 Principle
Ammonium oxidation, the first step in autotrophic nitrification in soil, is used to assess the potential activity of
microbial nitrifying populations. Autotrophic ammonium-oxidizing bacteria are exposed to ammonium sulfate
in a soil slurry buffered at pH 7,2. Oxidation of the nitrite performed by nitrite-oxidizing bacteria in the slurry
is inhibited by the addition of sodium chlorate. The subsequent accumulation of nitrite is measured over a 6 h
incubation period, and is taken as an estimate of the potential activity of ammonium-oxidizing bacteria. As the
generation time of ammonia-oxidizing bacteria is long (> 10 h), the method provides a measure of the potential
activity of the nitrifying population at the time of sampling. It does not measure growth of the nitrifying population.
Chemical test substances and extracts of biosolids can be tested by adding them to the slurry at various
concentrations. The test can be used for comparison of different soils (e.g. polluted and reference soils).
Effects of polluted soils, waste samples and other solid materials on nitrification can be evaluated by measuring
the method on mixtures of these samples with a reference soil.
Substances known to be active at pH < 7,2 should be added in due time before starting the test to allow
substances to exhibit their effects on nitrifying bacteria.
5 Reagents
5.1 Distilled water.
5.2 Potassium dihydrogenphosphate, c(KH PO = 0,2 mol/l).
2 4
5.3 Dipotassium hydrogenphosphate, c(K HPO = 0,2 mol/l).
2 4
5.4 Sodium chlorate, c(NaClO = 0,5 mol/l).
3
5.5 Diammonium sulfate, (NH ) SO .
4 2 4
5.6 Sodium hydrogencarbonate, c(NaHCO = 5 mmol/l).
3
5.7 Potassium chloride, c(KCl = 4 mol/l).
5.8 Stock solution A.
Prepare stock solution A as follows.
Potassium dihydrogenphosphate, KH PO (5.2) 28 ml
2 4
Dipotassium hydrogenphosphate, K HPO (5.3) 72 ml
2 4
Distilled water (5.1) 100 ml
5.9 Test medium.
Prepare the test medium as follows.
Stock solution A (5.8) 10 ml
Sodium chlorate, NaClO (5.4) 10 ml to 30 ml
3
Diammonium sulfate, (NH ) SO (5.5) 0,198 g
4 2 4
2 © ISO 2012 – All rights reserved

---------------------- Page: 12 ----------------------
SIST EN ISO 15685:2020
ISO 15685:2012(E)
Dilute to 1 000 ml with distilled water (5.1).
The test medium contains 1 mmol/l of potassium phosphate buffer, 5 mmol/l to 15 mmol/l of sodium chlorate
and 1,5 mmol/l of diammonium sulfate, and has a pH of approximately 7,2.
The sodium chlorate concentration selected within the range given should be sufficient for effective inhibition
of biological nitrate formation, while not having negative effects on ammonium oxidation. If necessary, test the
influence of the sodium chlorate concentration.
Test chemicals to be added to the test medium shall be dissolved in the phosphate buffer described and added
before diluting to 1 l. For test chemicals with low water solubility, prepare solutions by mechanical dispersion
or by using vehicles such as organic solvents, emulsifiers or dispergents of low toxicity to ammonium-oxidizing
bacteria. If any vehicle is used to add a test chemical to the slurry or a soil, additional control shall be performed
to assess possible effects of this vehicle on ammonium oxidation.
Add extracts of biosolids, when tested, separately to the test medium before dilution to 1 l.
NOTE When a carbon source is needed, e.g. when testing biosolids with high nitrate levels, add 10 ml of
NaHCO (5.6) to the test medium before diluting to 1 l.
3
6 Apparatus
6.1 Ordinary laboratory equipment.
6.2 Orbital shaking incubator, thermostatically controlled.
7 Sampling, storage and characterization of samples
Guidance on sampling, sample processing and storage is given in ISO 10381-6. For the testing of chemicals
or biosolids, or for mixing tests with polluted soils, experimental soil with an ammonium-oxidizing activity of
between 200 ng N/g and 800
...

SLOVENSKI STANDARD
oSIST prEN ISO 15685:2020
01-januar-2020
Kakovost tal - Določevanje potencialne nitrifikacije in inhibicije nitrifikacije - Hitri
preskus z oksidacijo amonija (ISO 15685:2012)
Soil quality - Determination of potential nitrification and inhibition of nitrification - Rapid
test by ammonium oxidation (ISO 15685:2012)
Qualité du sol - Détermination de la nitrification potentielle et inhibition de la nitrification -
Essai rapide par oxydation de l'ammonium (ISO 15685:2012)
Ta slovenski standard je istoveten z: prEN ISO 15685
ICS:
13.080.30 Biološke lastnosti tal Biological properties of soils
oSIST prEN ISO 15685:2020 en,fr,de
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

---------------------- Page: 1 ----------------------
oSIST prEN ISO 15685:2020

---------------------- Page: 2 ----------------------
oSIST prEN ISO 15685:2020
INTERNATIONAL ISO
STANDARD 15685
Second edition
2012-07-15
Soil quality — Determination of potential
nitrification and inhibition of nitrification —
Rapid test by ammonium oxidation
Qualité du sol — Détermination de la nitrification potentielle et inhibition
de la nitrification — Essai rapide par oxydation de l’ammonium
Reference number
ISO 15685:2012(E)
©
ISO 2012

---------------------- Page: 3 ----------------------
oSIST prEN ISO 15685:2020
ISO 15685:2012(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2012
All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means,
electronic or mechanical, including photocopying and microfilm, without permission in writing from either ISO at the address below or ISO’s
member body in the country of the requester.
ISO copyright office
Case postale 56 • CH-1211 Geneva 20
Tel. + 41 22 749 01 11
Fax + 41 22 749 09 47
E-mail copyright@iso.org
Web www.iso.org
Published in Switzerland
ii © ISO 2012 – All rights reserved

---------------------- Page: 4 ----------------------
oSIST prEN ISO 15685:2020
ISO 15685:2012(E)
Contents Page
Foreword .iv
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Principle . 2
5 Reagents . 2
6 Apparatus . 3
7 Sampling, storage and characterization of samples . 3
8 Procedure . 4
8.1 Testing soils . 4
8.2 Testing the effect of chemicals . 4
8.3 Testing polluted soils . 4
8.4 Testing water extracts of biosolids . 5
9 Calculations . 5
10 Test report . 5
Annex A (informative) Results of a laboratory ring test . 7
Annex B (informative) Micro-scale measurement of potential nitrification rates. 9
Bibliography . 11
© ISO 2012 – All rights reserved iii

---------------------- Page: 5 ----------------------
oSIST prEN ISO 15685:2020
ISO 15685:2012(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.
International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2.
The main task of technical committees is to prepare International Standards. Draft International Standards
adopted by the technical committees are circulated to the member bodies for voting. Publication as an
International Standard requires approval by at least 75 % of the member bodies casting a vote.
Attention is drawn to the possibility that some of the elements of this document may be the subject of patent
rights. ISO shall not be held responsible for identifying any or all such patent rights.
ISO 15685 was prepared by Technical Committee ISO/TC 190, Soil quality, Subcommittee SC 4, Biological methods.
This second edition cancels and replaces the first edition (ISO 15685:2004), which has been technically revised.
iv © ISO 2012 – All rights reserved

---------------------- Page: 6 ----------------------
oSIST prEN ISO 15685:2020
INTERNATIONAL STANDARD ISO 15685:2012(E)
Soil quality — Determination of potential nitrification and
inhibition of nitrification — Rapid test by ammonium oxidation
1 Scope
This International Standard specifies a rapid method for the determination of the potential rate of ammonium
oxidation and inhibition of nitrification in soils. This method is suitable for all soils containing a population of
nitrifying microorganisms. It can be used as a rapid screening test for monitoring soil quality and quality of
wastes, and is suitable for testing the effects of cultivation methods, chemical substances [except volatiles,
i.e. H > 1 (Henry’s constant)], extracts of biosolids and pollution in soils.
2 Normative references
The following referenced documents are indispensable for the application 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 10381-6, Soil quality — Sampling — Part 6: Guidance on the collection, handling and storage of soil under
aerobic conditions for the assessment of microbiological processes, biomass and diversity in the laboratory
ISO 10390, Soil quality — Determination of pH
ISO 10694, Soil quality — Determination of organic and total carbon after dry combustion (elementary analysis)
ISO 11260, Soil quality — Determination of effective cation exchange capacity and base saturation level using
barium chloride solution
ISO 11261, Soil quality — Determination of total nitrogen — Modified Kjeldahl method
ISO 11277, Soil quality — Determination of particle size distribution in mineral soil material — Method by
sieving and sedimentation
ISO 11465, Soil quality — Determination of dry matter and water content on a mass basis — Gravimetric method
ISO 14238, Soil quality — Biological methods — Determination of nitrogen mineralization and nitrification in
soils and the influence of chemicals on these processes
ISO 14256-2, Soil quality — Determination of nitrate, nitrite and ammonium in field-moist soils by extraction
with potassium chloride solution — Part 2: Automated method with segmented flow analysis
EN 14735, Characterization of waste — Preparation of waste samples for ecotoxicity tests
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1
inhibitory dose
ID
amount of a chemical added to soil that effectively inhibits biological activity by a stated percentage after a
given time, in comparison with an untreated control
NOTE It is expressed as a percentage. For example, ID25 and ID50 indicate a 25 % and 50 % inhibition of biological
activity, respectively.
© ISO 2012 – All rights reserved 1

---------------------- Page: 7 ----------------------
oSIST prEN ISO 15685:2020
ISO 15685:2012(E)
3.2
biosolids
organic products used in agriculture, which include sewage sludge, compost, manure and some industrial wastes
4 Principle
Ammonium oxidation, the first step in autotrophic nitrification in soil, is used to assess the potential activity of
microbial nitrifying populations. Autotrophic ammonium-oxidizing bacteria are exposed to ammonium sulfate
in a soil slurry buffered at pH 7,2. Oxidation of the nitrite performed by nitrite-oxidizing bacteria in the slurry
is inhibited by the addition of sodium chlorate. The subsequent accumulation of nitrite is measured over a 6 h
incubation period, and is taken as an estimate of the potential activity of ammonium-oxidizing bacteria. As the
generation time of ammonia-oxidizing bacteria is long (> 10 h), the method provides a measure of the potential
activity of the nitrifying population at the time of sampling. It does not measure growth of the nitrifying population.
Chemical test substances and extracts of biosolids can be tested by adding them to the slurry at various
concentrations. The test can be used for comparison of different soils (e.g. polluted and reference soils).
Effects of polluted soils, waste samples and other solid materials on nitrification can be evaluated by measuring
the method on mixtures of these samples with a reference soil.
Substances known to be active at pH < 7,2 should be added in due time before starting the test to allow
substances to exhibit their effects on nitrifying bacteria.
5 Reagents
5.1 Distilled water.
5.2 Potassium dihydrogenphosphate, c(KH PO = 0,2 mol/l).
2 4
5.3 Dipotassium hydrogenphosphate, c(K HPO = 0,2 mol/l).
2 4
5.4 Sodium chlorate, c(NaClO = 0,5 mol/l).
3
5.5 Diammonium sulfate, (NH ) SO .
4 2 4
5.6 Sodium hydrogencarbonate, c(NaHCO = 5 mmol/l).
3
5.7 Potassium chloride, c(KCl = 4 mol/l).
5.8 Stock solution A.
Prepare stock solution A as follows.
Potassium dihydrogenphosphate, KH PO (5.2) 28 ml
2 4
Dipotassium hydrogenphosphate, K HPO (5.3) 72 ml
2 4
Distilled water (5.1) 100 ml
5.9 Test medium.
Prepare the test medium as follows.
Stock solution A (5.8) 10 ml
Sodium chlorate, NaClO (5.4) 10 ml to 30 ml
3
Diammonium sulfate, (NH ) SO (5.5) 0,198 g
4 2 4
2 © ISO 2012 – All rights reserved

---------------------- Page: 8 ----------------------
oSIST prEN ISO 15685:2020
ISO 15685:2012(E)
Dilute to 1 000 ml with distilled water (5.1).
The test medium contains 1 mmol/l of potassium phosphate buffer, 5 mmol/l to 15 mmol/l of sodium chlorate
and 1,5 mmol/l of diammonium sulfate, and has a pH of approximately 7,2.
The sodium chlorate concentration selected within the range given should be sufficient for effective inhibition
of biological nitrate formation, while not having negative effects on ammonium oxidation. If necessary, test the
influence of the sodium chlorate concentration.
Test chemicals to be added to the test medium shall be dissolved in the phosphate buffer described and added
before diluting to 1 l. For test chemicals with low water solubility, prepare solutions by mechanical dispersion
or by using vehicles such as organic solvents, emulsifiers or dispergents of low toxicity to ammonium-oxidizing
bacteria. If any vehicle is used to add a test chemical to the slurry or a soil, additional control shall be performed
to assess possible effects of this vehicle on ammonium oxidation.
Add extracts of biosolids, when tested, separately to the test medium before dilution to 1 l.
NOTE When a carbon source is needed, e.g. when testing biosolids with high nitrate levels, add 10 ml of
NaHCO (5.6) to the test medium before diluting to 1 l.
3
6 Apparatus
6.1 Ordinary laboratory equipment.
6.2 Orbital shaking incubator, thermostatically controlled.
7 Sampling, storage and characterization of samples
Guidance on sampling, sample processing and storage is given in ISO 10381-6. For the testing of chemicals
or biosolids, or for mixing tests with polluted soils, experimental soil with an ammonium-oxidizing activity of
between 200 ng N/g and 800 ng N/g of dry mass of soil/h, determined from a preliminary experiment, should
be chosen. When testing chemicals, the soil should have a low adsorption capacity (e.g. sandy soils low in
C < 1 %); on the other hand, the soil should have a pH between 5,5 and 7,5.
org
Soil samples can usually be stored in a refrigerator at approximately 4 °C for periods of up to three months prior
to use (ISO 10381-6). Some soils can only be kept in a refrigerator for up to two weeks. Storage of soil samples
in a freezer (‑20 °C) is not generally recommended.
It has been shown for a number of soils from temperate climates that storage at ‑20 °C for up to 12 months does
not inhibit ammonium oxidation activity (Reference [4]). Where such information is available for a particular soil,
storage under such conditions is permissible.
Measurement of the following variables is recommended for characterization of the soil:
— particle size distribution (ISO 11277);
— water content (ISO 11465);
— water-holding capacity (ISO 14238);
— pH (ISO 10390);
— effective cation exchange capacity (ISO 11260);
— organic matter content (ISO 10694);
— total nitrogen content (ISO 11261).
© ISO 2012 – All rights reserved 3

---------------------- Page: 9 ----------------------
oSIST prEN ISO 15685:2020
ISO 15685:2012(E)
8 Procedure
8.1 Testing soils
8.1.1 General
The test design should include at least three replicates of approximately 25 g of moist soil. The water content
of soil for the calculation of dry mass is determined gravimetrically according to ISO 11465.
8.1.2 Initial incubation
Mix soil samples or waste materials with test medium (5.9) to form slurries. The volume of test medium should
be adjusted to give a precise total liquid volume, e.g. 100 ml. Calculate the volume of medium to be added by
subtracting the volume of water in the initial soil or waste sample from the desired liquid volume, e.g. 100 ml.
Incubate the slurries in 250 ml flasks, placed upright on the orbital shaking incubator (6.2), thermostatically
controlled at (25 ± 2) °C. Rotation should be sufficient to keep solids suspended (1
...

Questions, Comments and Discussion

Ask us and Technical Secretary will try to provide an answer. You can facilitate discussion about the standard in here.