Fertilizers - Determination of chelating agents in fertilizers by chromatography - Part 3: Determination of [S,S]-EDDS by ion pair chromatography

The document specifies a method for the chromatographic determination of the chelating agent [S,S]-EDDS in fertilizers. The method allows the identification and the determination of the total concentration of the water soluble fraction of this chelating agent. It does not allow to distinguish between the free form and the metal bound form of the chelating agent. This method is applicable to EC-fertilizers containing chelates of one or more of the following micro-nutrients: cobalt, copper, iron, manganese and zinc covered by Regulation (EC) No 2003/2003. It is applicable to a mass fraction of the metal chelated of at least 0,35 %.

Düngemittel - Bestimmung von Chelatbildnern in Düngemitteln mit Chromatographie - Teil 3: Bestimmung von [S,S]-EDDS mit Ionen-Paarchromatographie

Diese Europäische Norm legt ein Verfahren zur chromotographischen Bestimmung des Chelatbildners [S,S]-EDDS in Düngemitteln fest. Das Verfahren erlaubt die Identifizierung und die Bestimmung der Gesamtkonzentration der wasserlöslichen Fraktionen dieses Chelatbildners. Es dient jedoch nicht dazu, zwischen der freien Form und der metallgebundenen Form des Chelatbildners zu unterscheiden.
Dieses Verfahren ist anwendbar für EG-Düngemittel, die einen oder mehrere der folgenden Spurennährstoffe enthalten: Kobalt, Kupfer, Eisen, Mangan und Zink, die der Verordnung (EG) Nr. 2003/2003 [4] unterliegen. Es gilt für einen Massenanteil an chelatisiertem Metall von mindestens 0,35 %.
ANMERKUNG 1   Die Substanz EDDS (Ethylendiamin-N,N'-Bernsteinsäure) existiert in mehreren unterschiedlichen stereo-isomeren Formen. [S,S] (mit CAS Nummer 20846-91-7), [R,R] und [R,S] optische Isomere sind möglich. [S,S] und [R,R] sind Spiegelbilder mit gleichen chemischen Eigenschaften. Jedoch ist nur das [S,S] Isomer biologisch abbaubar. Wenn beide vorliegen, bilden sie ein racemisches Gemisch. Das [R,S] Isomer (das Meso-Isomer) ist nur langsam biologisch abbaubar.
Nur das [S,S]-EDDS Isomer ist durch die Verordnung (EG) Nr 2003/2003 zulässig. Da das Cu Chelat des [R,R]-EDDS Isomers die gleiche Stabilität aufweist wie das [S,S]-EDDS, sollten beide koeluieren. Das Fehlen von [R,S] EDDS weist darauf hin, dass nur ein reines Isomer ([S,S] oder [R,R]) vorliegt. Eine zusätzliche, auf Polarimetrie basierende, Prüfung kann angewendet werden, um die isomere Eigenschaft der Probe oder des Standards festzustellen.

Engrais - Détermination des agents chélatants dans les engrais par chromatographie - Partie 3: Détermination du [S,S]-EDDS par chromatographie d'appariement d'ions

La présente Norme européenne spécifie une méthode de détermination de l’agent chélatant [S,S]-EDDS dans les engrais par chromatographie. La méthode permet l’identification et la détermination de la concentration totale de la fraction soluble dans l’eau de cet agent chélatant. Elle ne permet pas de faire la distinction entre la forme libre et la forme liée du métal de l’agent chélatant.
Cette méthode est applicable aux engrais CE contenant des chélates d’un ou de plusieurs des oligo-éléments suivants : cobalt, cuivre, fer, manganèse et zinc, visés par le Règlement (CE) no 2003/2003 [4]. Elle est applicable à une fraction massique du métal chélaté d’au moins 0,35 %.
NOTE 1   La substance EDDS (acide éthylènediamine-N,N´-disuccinique) existe sous différentes formes stéréo-isomériques. Des isomères optiques [S,S] (portant le numéro CAS 20846-91-7), [R,R] et [R,S] sont possibles. Les isomères [S,S] et [R,R] sont des images miroirs avec des caractéristiques chimiques équivalentes. Cependant, seul l’isomère [S,S] est biodégradable. Lorsque les deux sont présents, ils forment le mélange racémique. L’isomère [R,S] (isomère méso) n’est biodégradable que sur le long terme.
Seul l’isomère [S,S]-EDDS est autorisé par le Règlement (CE) no 2003/2003. Comme le chélate de Cu de l’isomère [R,R]-EDDS présente la même stabilité que le [S,S]-EDDS, il convient que les deux co-éluent. L’absence de [R,S]-EDDS indique que seul un isomère pur ([S,S] ou [R,R]) existe. Un essai supplémentaire basé sur la polarimétrie peut être utilisé pour vérifier la caractéristique isomérique de l’échantillon ou de la solution de calibration.

Gnojila - Določevanje sredstev za kelatiziranje v gnojilih s kromatografijo - 3. del: Določevanje [S,S]-EDDS s kromatografijo ionskih parov

Ta dokument določa metodo za kromatografsko določanje sredstva za kelatiziranje [S,S]-EDDS v gnojilih. Metoda omogoča opredelitev in določevanje skupne koncentracije v vodi topnega dela tega sredstva za kelatiziranje. Ne dopušča razlikovanja med prosto obliko sredstva za kelatiziranje in obliko sredstva za kelatiziranje, vezano na kovino. Ta metoda se uporablja za gnojila EC, ki vsebujejo kelate enega ali več od naslednjih mikrohranil: kobalt, baker, železo, mangan in cink, ki jih zajema Uredba (ES) št. 2003/2003. Uporablja se za masni delež kovinskih kelatov najmanj 0,35 %.

General Information

Status
Published
Public Enquiry End Date
06-Nov-2016
Publication Date
17-May-2018
Technical Committee
Current Stage
6060 - National Implementation/Publication (Adopted Project)
Start Date
04-May-2018
Due Date
09-Jul-2018
Completion Date
18-May-2018

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SLOVENSKI STANDARD
SIST EN 13368-3:2018
01-junij-2018

*QRMLOD'RORþHYDQMHVUHGVWHY]DNHODWL]LUDQMHYJQRMLOLKVNURPDWRJUDILMRGHO

'RORþHYDQMH>66@(''6VNURPDWRJUDILMRLRQVNLKSDURY

Fertilizers - Determination of chelating agents in fertilizers by chromatography - Part 3:

Determination of [S,S]-EDDS by ion pair chromatography

Düngemittel - Bestimmung von Chelatbildnern in Düngemitteln mit Chromatographie -

Teil 3: Bestimmung von [S,S]-EDDS mit Ionen-Paarchromatographie

Engrais - Détermination des agents chélatants dans les engrais par chromatographie -

Partie 3: Détermination du [S,S]-EDDS par chromatographie d'appariement d'ions
Ta slovenski standard je istoveten z: EN 13368-3:2017
ICS:
65.080 Gnojila Fertilizers
SIST EN 13368-3:2018 en,fr,de

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

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SIST EN 13368-3:2018
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SIST EN 13368-3:2018
EN 13368-3
EUROPEAN STANDARD
NORME EUROPÉENNE
December 2017
EUROPÄISCHE NORM
ICS 65.080
English Version
Fertilizers - Determination of chelating agents in fertilizers
by chromatography - Part 3: Determination of [S,S]-EDDS
by ion pair chromatography

Engrais - Détermination des agents chélatants dans les Düngemittel - Bestimmung von Chelatbildnern in

engrais par chromatographie - Partie 3: Détermination Düngemitteln mit Chromatographie - Teil 3:

du [S,S]-EDDS par chromatographie d'appariement Bestimmung von [S,S]-EDDS mit Ionen-

d'ions Paarchromatographie
This European Standard was approved by CEN on 25 September 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 13368-3:2017 E

worldwide for CEN national Members.
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SIST EN 13368-3:2018
EN 13368-3:2017 (E)
Contents Page

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

1 Scope .................................................................................................................................................................... 4

2 Normative references .................................................................................................................................... 4

3 Terms and definitions ................................................................................................................................... 4

4 Principle ............................................................................................................................................................. 4

5 Interferences .................................................................................................................................................... 5

6 Reagents ............................................................................................................................................................. 5

7 Apparatus ........................................................................................................................................................... 6

8 Sampling and sample preparation ............................................................................................................ 7

9 Procedure........................................................................................................................................................... 7

9.1 Preparation of the sample solution .......................................................................................................... 7

9.2 Preparation of the calibration solutions ................................................................................................ 7

9.3 Chromatographic analysis ........................................................................................................................... 8

10 Expression of the results ........................................................................................................................... 11

10.1 Mass fraction of the chelating agent [S,S]-EDDS ................................................................................ 11

10.2 Fe chelated by [S,S]-EDDS (only for Fe-[S,S]-EDDS straight fertilizers) .................................... 12

11 Precision .......................................................................................................................................................... 12

11.1 Inter-laboratory test ................................................................................................................................... 12

11.2 Repeatability .................................................................................................................................................. 12

11.3 Reproducibility ............................................................................................................................................. 12

12 Test report ...................................................................................................................................................... 13

Annex A (informative) Statistical results of the inter-laboratory tests performed in 2016 .......... 14

A.1 Test samples .................................................................................................................................................. 14

A.2 Inter-laboratory test procedure ............................................................................................................. 14

A.3 Results and statistical interpretation ................................................................................................... 14

Annex B (informative) General procedure for the determination of the titrimetric purity of

the chelating agent using a photometric automatic titrator ........................................................ 16

Annex C (informative) Polarimetric evaluation of isomeric purity of the EDDS commercial

fertilizers ......................................................................................................................................................... 17

Annex D (informative) Complete names of chelating agents .................................................................... 19

Bibliography ................................................................................................................................................................. 20

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SIST EN 13368-3:2018
EN 13368-3:2017 (E)
European foreword

This document (EN 13368-3:2017) has been prepared by Technical Committee CEN/TC 260 “Fertilizers

and liming materials”, the secretariat of which is held by DIN.

This European Standard shall be given the status of a national standard, either by publication of an

identical text or by endorsement, at the latest by 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.

This document has been prepared under a mandate given to CEN by the European Commission and the

European Free Trade Association.

The European Standard EN 13368 Fertilizers — Determination of chelating agents in fertilizers by

chromatography consists of the following parts:
— Part 1: Determination of EDTA, HEEDTA and DTPA by ion chromatography

— Part 2: Determination of Fe chelated by [o,o] EDDHA, [o,o] EDDHMA and HBED, or the amount of

chelating agents, by ion pair chromatography
— Part 3: Determination of [S,S]-EDDS by ion pair chromatography

According to the CEN-CENELEC Internal Regulations, the national standards organisations 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.
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SIST EN 13368-3:2018
EN 13368-3:2017 (E)
1 Scope

This European Standard specifies a method for the chromatographic determination of the chelating

agent [S,S]-EDDS in fertilizers. The method allows the identification and the determination of the total

concentration of the water-soluble fraction of this chelating agent. It does not allow distinguishing

between the free form and the metal bound form of the chelating agent.

This method is applicable to EC fertilizers containing chelates of one or more of the following micro-

nutrients: cobalt, copper, iron, manganese and zinc, covered by Regulation (EC) No 2003/2003 [4]. It is

applicable to a mass fraction of the metal chelated of at least 0,35 %.

NOTE The substance EDDS (ethylenediamine-N,N'-disuccinic acid) exists as several different stereo isomeric

forms. [S,S] (with CAS Number 20846–91–7), [R,R] and [R,S] optical isomers are possible. [S,S] and [R,R] are mirror

images with equal chemical characteristics. However only the [S,S] isomer is biodegradable. When both are

present, they form the racemic mixture. The [R,S] isomer (the meso isomer) is only slowly biodegradable.

Only the [S,S]-EDDS isomer is allowed by the Regulation (EC) No 2003/2003. Since the Cu chelate of the

[R,R]-EDDS isomer presents the same stability as the [S,S]-EDDS, both should coelute. The absence of

[R,S]-EDDS indicates that only a pure isomer ([S,S] or [R,R]) exists. An additional test based on

polarimetry can be used to ascertain the isomeric characteristic of the sample, or the standard.

2 Normative references

The following documents, in whole or in part, are normatively referenced in this document and are

indispensable for its application. For dated references, only the edition cited applies. For undated

references, the latest edition of the referenced document (including any amendments) applies.

EN 1482-2, Fertilizers and liming materials - Sampling and sample preparation - Part 2: Sample

preparation

EN 12944-1:1999, Fertilizers and liming materials and soil improvers - Vocabulary - Part 1: General terms

EN 12944-2:1999, Fertilizers and liming materials and soil improvers - Vocabulary - Part 2: Terms

relating to fertilizers

EN ISO 3696, Water for analytical laboratory use - Specification and test methods (ISO 3696)

3 Terms and definitions

For the purposes of this document, the terms and definitions given in EN 12944-1:1999 and

EN 12944-2:1999 apply.
4 Principle

The micronutrients associated with the [S,S]-EDDS present in an aqueous extract of the sample are

replaced by Cu(II). The Cu(II) chelates are separated and determined by isocratic ion-pair high-

performance liquid chromatography. When a copper chelate (anion) is added to a polar fluid (eluent),

containing a large cation, an ion pair is formed. This ion pair is retained by an apolar solid phase

(stationary phase). The strength of the retention depends on the molecular size and its acidity. Then,

each copper chelate presents a characteristic retention time depending on the chelating agent, and it is

separated from the other substances present in the sample. The separation is carried out on a reverse

phase silica column and an aqueous solution of TBA (tetrabutylammonium) and copper acetate and

methanol as eluent. The detection is based on photometry at 254 nm.

A derivatization method should be used to form the Cu chelates, so the chelating agent is determined by

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SIST EN 13368-3:2018
EN 13368-3:2017 (E)
the isocratic ion-pair high-performance liquid chromatography here specified.
5 Interferences

No interferences have been detected. Metal chelates with [o,o] EDDHA, [o,o] EDDHMA, HBED, EDDHSA,

EDTA, DTPA, HEEDTA, IDHA, [o,p] EDDHA, lignosulfonates and heptagluconate as well as the chelating

agents do not interfere since after the Cu derivatization they are separated from Cu-[S,S]-EDDS. Since

retention times depend on the column type, any interference should be checked if a mixture with other

chelating or complexing agents is suspected.

For the complete names of the abbreviations of chelating agents used in this document, see Annex D.

6 Reagents
6.1 General.

All reagents shall be of recognized analytical grade. All water used for the preparation of eluents,

standards, and sample solutions shall conform to EN ISO 3696, grade 1 and shall be degassed and free

of organic contaminants. If products with a declared purity of less than 99 % are used for the

preparation of standard solutions, a correction should be made in order to obtain exactly the required

concentration in the solution.

If there is any doubt on the purity of the standard substances, it is necessary to determine it.

For the determination of its concentration, a titrimetric method can be used. See Annex B for a

photometric method using an automatic titrator. Manual titration can also be adequate.

The isomeric purity can be checked using the method indicated in Annex C.
6.2 Sodium hydroxide solution, c(NaOH) = 0,15 mol/l.

Dissolve 3,0 g of NaOH in pellet form in a 500 ml volumetric flask with water (6.1). Dilute to the mark

and homogenize.

The incorporation of CO from the atmosphere should be carefully avoided. Otherwise, the dissolution

of the chelating agent (see 6.6) can be incomplete.
6.3 Phosphoric acid solution, c(H PO ) = 1,0 mol/l.
3 4

Dilute 68 ml of phosphoric acid (mass fraction 85 % H PO ) to 1 000 ml with water.

3 4
6.4 Phosphoric acid solution, c(H PO ) = 0,1 mol/l.
3 4
Dilute 50 ml of phosphoric acid 1,0 mol/l (6.3) to 500 ml with water.
6.5 Cu acetate solution (for derivatization), c(Cu) approximately 0,02 mol/l.

Dissolve 4,00 g of copper acetate monohydrate (Cu(CH -COO) ·H O) in 1 l of water.

3 2 2
6.6 [S,S]-EDDS solution, c(EDDS) = 8,95 mmol/l.

If the EDDS acid form is used (CAS# 20846-91-7), dissolve 26,16/P g (where P is the purity of the solid

standard in percentage of EDDS acid form), in 50 ml of water and 20 ml of NaOH (6.2) in a 100 ml

beaker and make up to 100 ml in a volumetric flask with water. The standard obtained in this way may

be stored in darkness in a fridge (at about 5 °C) for 6 months. If a significant yellowing appears (due to

photodecomposition) a fresh standard should be prepared.
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SIST EN 13368-3:2018
EN 13368-3:2017 (E)

If EDDS trisodium salt solution is used instead of the acid form then dissolve 32,07/P g (where P is the

purity of the standard in percentage of EDDS trisodium salt) in 50 ml of water in a 100 ml beaker and

make up to 100 ml in a volumetric flask with water.

NOTE This concentration is equivalent to ρ(Fe) = 500 mg/l. Since most commercial samples are Fe chelates,

equivalent Fe concentration is given to simplify final calculations.
6.7 Eluent for the determination.

Dissolve 1,00 g of copper acetate monohydrate (Cu(CH -COO) ·H O) (5,00 mmol) in 900 ml of water.

3 2 2

Add 2 ml of TBAOH (mass fraction 40 % Tetrabutylammonium hydroxide solution in water). Adjust to

pH 2,8 with phosphoric acid solution (6.3 and 6.4). Add 40 ml of methanol (HPLC grade) and make up to

volume in a 1 l volumetric flask with water. Filter through a 0,2 µm membrane filter [7.4 b)] and degas.

NOTE TBACl or TBABr can be used, providing that pH is adjusted to 2,8 with H PO .

3 4
7 Apparatus
Usual laboratory equipment, glassware and the following:
7.1 Magnetic stirrer.
7.2 Chromatograph, equipped with:
a) an isocratic pump delivering the eluent at a flow rate of 1,0 ml/min;
b) an injection valve with a 20 µl injection loop;

c) a C-18 column , dp = 5 µm, > 99 % end capped; recommended internal diameter: 3,9 mm and

column length: 150 mm;
d) a C-18 guard column (recommended);
e) a UV/VIS-detector with a 254 nm-filter;
f) an integrator.
7.3 Balance, with an accuracy of ± 0,1 mg.
7.4 Membrane filters, including:

a) micro membrane filters resistant to aqueous solutions, with porosity of 0,45 µm.

b) micro membrane filters resistant to organic solutions (e.g. polyamide 66 micro membrane filters),

with porosity of 0,2 µm.
1 TM

) SYMMETRY C18, from WATERS, LiChroCART® Purospher® RP-18, from MERCK, or equivalent are

examples of suitable products available commercially. This information is given for the convenience of users of

this European Standard and does not constitute an endorsement by CEN of these products.

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SIST EN 13368-3:2018
EN 13368-3:2017 (E)
8 Sampling and sample preparation

Sampling is not part of the method specified in this document. A recommended sampling method is

given in EN 1482-1 [1].
Sample preparation shall be carried out in accordance with EN 1482-2.
For the size reduc
...

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