CEN/TC 260/WG 5 - Determination of chelating agents

This document specifies a method for the determination of the chelated iron content and the chelated fraction of iron, in UVCB chelats, EDDHA, EDDHMA, HBED, EDDHSA, in inorganic micronutrient fertilizers by the treatment with a cation exchange resin.
The limit of determination of the chelated iron content highly depends on the specific electrical conductivity of the sample, on the amount of nutrient present, and varies between 0,005 % in simple matrices with high amounts of micronutrient and 0,5 % in more complex cases (see 9.1).

This document specifies a method for the determination of the soluble metal that remains in solution at different pHs after the application of a solution of the fertilizer substance containing micronutrient chelates in a tap water solution used as a reference.
The method applies to fertilizing products containing chelated micronutrients.

This document specifies two methods required for the identification of lignosulfonate by UV-Vis spectrophotometry (method A) and gravimetry (method B) in organo-mineral fertilizers.
NOTE   Lignosulfonate, as a complexing agent, is a natural polymer produced as a by-product of the sulfite method for manufacturing paper from wood pulp in the paper industry. As a natural polymer, it presents a poorly defined and variable chemical structure. It is an intricate mixture of small- to moderate-sized polymeric compounds with sulfonate groups attached to the molecule, and diverse complexing capacity.

This document specifies the references to the methods for the determination of stability of chelating and complexing agents for CMC 1 as specified in the Regulation (EU) 2019/1009. The document specifies references to the methods and requirements for inorganic micronutrient fertilizers in accordance with PFC 1(C)(II) as specified in the Regulation (EU) 2019/1009 [1].
Inorganic micronutrient materials for this purpose are micronutrient chelates or complexes and mixtures of them, in powder or granular form, aqueous or suspension preparations.

This document specifies the method for the determination of free, chelated or complexed micronutrients and chelating and/or complexing agents present in compound inorganic micronutrient fertilizers.
This method applies to compound inorganic micronutrient fertilizers when micronutrients are chelated and/or complexed.
The method is based on the determination of the following specific parameters :
-   the water-soluble micronutrient concentration;
-   the fraction of chelated micronutrients in relation;
-   identification of chelating agents EDTA, DTPA, HEEDTA, IDHA, [S,S]–EDDS, [o,o] EDDHA, [o,o] EDDHMA, [o,p] EDDHA, HBED and EDDHSA;
-   the fraction of complexed micronutrients;
-   identification of complexing agents (lignosulfonates, heptagluconic acid (HGA)).
The method is based on
-   ICP (inductive coupled plasma) or FAAS (flame atomic absorption spectrometry) measurement of the concentration of water-soluble micronutrients according to EN 16963 or EN 16965 after extraction according to EN 16962;
-   LC (liquid chromatography) measurement of the chelating agents according to EN 15950, EN 13368-1, EN 13368-2, EN 13368-3, EN 15451, EN 15452;
and/or complexing agents according to EN 16109 and EN 16847;
-   determination of the concentration of chelated micronutrients by CEN/TS 17786-1 and/or CEN/TS 17786-2;
-   determination of the complexed micronutrients by EN 15962.
To avoid duplication of the analytical methods, CEN/TS 17786-2 describes the determination of micronutrients and the identification and determination of chelating agents.

This document specifies a method for the determination of the soluble metal that remains in solution after the application of a solution of the fertilizer substance containing micronutrient complexes in water and adjusting the pH to 6 and pH 7 for at least one day.
The method applies to fertilizing products containing micronutrient complexes.

This document specifies a method for the determination of the chelated micronutrient content and the chelated fraction of a micronutrient, in organo-mineral fertilizers, having an organic matrix based on vegetal residues (cocoa shells, grape residue, soybean residue, etc), algae extract, and animal meal (feather, bones, blood, etc) and containing UVCB, EDDHA, EDDHMA, HBED, EDDHSA micronutrients by the treatment with a cation exchange resin.
The limit of determination of the chelated micronutrient content highly depends on the specific electrical conductivity of the sample, on the amount of nutrient present, and varies between 0,005 % in simple matrices with high amounts of micronutrient, and 0,5 % in more complex cases (see 9.1).

This document specifies a method for the determination by ion chromatography of the total amount of each of the individual chelating agents EDTA, HEEDTA, and DTPA in organo-mineral fertilizers, having an organic matrix based on vegetal residues (cocoa shells, grape residue, soybean residue, ...), algae extract, and animal meal (feather, bones, blood, ...) and containing one or more of these substances.
The method allows the identification and the determination of the total water-soluble fraction of each of these chelating agents. It does not allow to distinguish between the free form and the metal bound form of the chelating agents.
This method applies to organo-mineral fertilizers containing chelates of one or more of the following micronutrients: cobalt, copper, iron, manganese, zinc and with a mass fraction of at least 0,1 %.

This document specifies a method for the determination of the chelated fraction of micronutrients for fertilizers containing one or many micronutrients chelated by EDTA, DTPA, HEEDTA, IDHA or [S,S]-EDDS in fertilizers.
This method is used for inorganic micronutrient fertilizers when micronutrients are chelated only by EDTA, DTPA, HEEDTA, IDHA or [S,S]-EDDS or for mixtures in which EDTA, DTPA, HEEDTA, IDHA or [S,S]-EDDS is one of the chelating agents.
The method is applicable to all inorganic micronutrient fertilizers containing EDTA, DTPA, HEEDTA, IDHA or [S,S]-EDDS as chelating agent for contents > 0,1 % (w/w).
The method is based on ICP or AAS measurement of the concentration of micronutrients according to EN 16963 or EN 16965 after water extraction according to EN 16962 and LC measurement of the chelating agents according to EN 15950, EN 13368-1 and EN 13368-3.

This document specifies references to the methods for the determination of specific micronutrients, chelating and complexing agents. The document specifies references to the methods and requirements for inorganic micronutrient fertilizers in accordance with PFC 1 (C) (II) as specified in the Regulation (EU) 2019/1009 [1].
Inorganic micronutrient materials for this purpose are micronutrient salts or oxide and hydroxides, or micronutrient chelates or complexes and mixtures of them, in powder or granular form, aqueous or suspension preparation.

This document specifies references to the methods for the determination of chelating and complexing agents in organo-mineral fertilizers. The document specifies references to the methods and requirements for organo-mineral fertilizers in accordance with PFC 1 (B) as specified in the Regulation (EU) 2019/1009 [1].
Organo-mineral materials for this purpose are organic fertilizers containing micronutrient chelates or complexes and/or mixtures of them, in powder or granular form, aqueous or suspension preparations.

This document specifies a method for the determination by ion pair chromatography of the iron chelated by each individual ortho(hydroxy)-ortho(hydroxy) isomer of the chelating agents [o,o] EDDHA, [o,o] EDDHMA and by HBED in organo-mineral fertilizers, having an organic matrix based on vegetal residues (cocoa shells, grape residue, soybean residue, ...), algae extract, and animal meal (feather, bones, blood, ...) and containing one or more of these substances, except for [o,o] EDDHMA and HBED mixes.
The method allows the identification and the determination of the total concentration of water soluble iron chelates of these chelating agents. Also, after derivatization with Fe, the soluble amount of the chelating agents can be determined when other micronutrients beside Fe are present in organo-mineral fertilizers containing [o,o] EDDHA, [o,o] EDDHMA or HBED.
This method is applicable to a mass fraction of the metal chelated of at least 0,625 %.
NOTE 1   The substances EDDHA and EDDHMA exist as several different isomeric forms. Positional isomers for the hydroxyl or methyl groups (in ortho, meta, and para positions) as well as stereo isomers (meso and dl-racemic forms) are known. Both meso and dl-racemic forms of the [ortho,ortho] EDDHA and [ortho,ortho]. Since para, meta and ortho methyl positional isomers of the EDDHMA present quite similar stability, they could be grouped: in the method here described the para, meta and ortho methyl positional isomers of the [o,o] EDDHMA are considered together. HBED (N,N'-bis(2-hydroxybenzyl)-ethylenediamine-N,N'-diacetic acid) does not present isomeric forms.
NOTE 2   At present, analytically pure standards only exist for [ortho,ortho] EDDHA, [ortho,ortho] EDDHMA and HBED. All other substances being unavailable as a standard, the influence of their eventual presence in the samples (with respect to the sensitivity and the selectivity of this method) has not been studied.
NOTE 3   The meso and the dl-racemic forms of [o,o] EDDHA and [o,o] EDDHMA can be determined separately by this method.

This document specifies a chromatographic method which allows the identification of heptagluconic acid (HGA) in organo-mineral fertilizers containing heptagluconic acid metal complexes.
NOTE   For the complete names of the chelating agents mentioned in this document, see Annex D.

This document specifies a general method for the determination of the micronutrients complexed by complexing agents in organo-mineral fertilizers. The method allows the determination of the total concentration of each complexed micronutrient in complexes after subtraction of the chelated micro-nutrients content, but it does not identify the individual complexing agents.
This procedure concerns EU organo-mineral fertilizing products which contain complexed micro-nutrients covered by Regulation (EU) 2019/1009 [6]. The method is applicable to a mass fraction of the metal complexed of at least 0,07 %, 0,006 % and 0,035 % of Fe, Mn and Zn respectively (see [7]). A lower limit of quantification has not been established for Cu and Co.

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 1   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 than the [S,S]-EDDS, both should coelute. The absence of [R,S]-EDDS indicates that only a pure isomer ([S,S] or [R,R]) exist. An additional test based on polarimetry can be used to ascertain the isomeric characteristic of the sample, or the standard.

This European Standard specifies a method for the chromatographic determination of the iron chelated by each individual ortho(hydroxy)-ortho(hydroxy) isomer of the chelating agents [o,o] EDDHA, [o,o] EDDHMA and by HBED in fertilizers containing one or more of these substances, except for [o,o] EDDHMA and HBED mixes. The method allows the identification and the determination of the total concentration of water soluble iron chelates of these chelating agents. Also, after derivatization with Fe, the soluble amount of the chelating agents can be determined when other micro-nutrients, beside Fe are present in fertilizers containing [o,o] EDDHA, [o,o] EDDHMA or HBED.
This method is applicable to EC fertilizers covered by Regulation (EC) No 2003/2003 [4]. It is applicable to a mass fraction of the metal chelated of at least 0,625 %.
NOTE 1   The substances EDDHA (ethylenediamine-N,N'-di[(hydroxyphenyl)acetic acid] and EDDHMA (ethylenediamine-N,N'-di[(hydroxymethylphenyl)acetic acid] exist as several different isomeric forms. Positional isomers for the hydroxyl or methyl groups (in ortho, meta, and para positions) as well as stereo isomers (meso and dl-racemic forms) are known. Both meso and dl-racemic forms of the [ortho,ortho] EDDHA and [ortho,ortho] EDDHMA are positional isomers for the hydroxyl groups allowed by the Regulation (EC) No 2003/2003. Since para, meta and ortho methyl positional isomers of the EDDHMA present quite similar stability, they could be grouped: in the method here described the para, meta and ortho methyl positional isomers of the [o,o] EDDHMA are considered together. HBED (N,N'-bis(2-hydroxybenzyl)-ethylenediamine-N,N'-diacetic acid) does not present isomeric forms.
NOTE 2   At present, analytically pure standards only exist for [ortho,ortho] EDDHA, [ortho,ortho] EDDHMA and HBED. All other substances being unavailable as a standard, the influence of their eventual presence in the samples (with respect to the sensitivity and the selectivity of this method) has not been studied.
NOTE 3   The meso and the dl-racemic forms of [o,o] EDDHA and [o,o] EDDHMA can be determined separately by this method.

This European Standard specifies a chromatographic method which allows the identification of heptagluconic acid (HGA) in fertilizers containing heptagluconic acid metal complexes.
This method is applicable to EC fertilizers containing complexed micro-nutrients, which are covered by Regulation (EC) No 2003/2003 [1].

This European Standard specifies a method for the chromatographic determination of the total amount of each of the individual chelating agents EDTA, HEEDTA, and DTPA in fertilizers containing one or more of these substances. The method allows the identification and the determination of the total water soluble fraction of each of these chelating agents. It does not allow to distinguish between the free form and the metal bound form of the chelating agents.
NOTE   EDTA, HEEDTA and DTPA are abbreviations used in this European Standard for the sake of simplicity. For complete names see Annex A.
This method applies to fertilizers containing chelates of one or more of the following micro-nutrients: cobalt, copper, iron, manganese, zinc and with a mass fraction of at least 0,1 %.

This document specifies two complementary methods (method A and method B) that allow lignosulfonates to be indentified as soluble complexing agents in fertilizers.
NOTE   Lignosulfonate, as a complexing agent, is a natural polymer produced as a by-product of the sulfite method for manufacturing paper from wood pulp in the paper industry. As a natural polymer, it presents a poorly defined and variable chemical structure. It is an intricate mixture of small- to moderate-sized polymeric compounds with sulfonate groups attached to the molecule, and diverse complexing capacity.
The methods are applicable to EC fertilizers covered by Regulation (EC) No 2003/2003 [1].

This European Standard specifies a general method for the determination of the micronutrients complexed by complexing agents in fertilizers. The method allows the determination of the total concentration of each complexed micronutrient in complexes after subtraction of the chelated micro-nutrients content, but it does not identify the individual complexing agents.
This procedure concerns EC-fertilizers which contain complexed micro-nutrients covered by Regulation (EC) No 2003/2003 [1]. The method is applicable to a mass fraction of the metal complexed of at least 0,07 %, 0,006 % and 0,035 % of Fe, Mn and Zn respectively (see [2]). A lower limit of quantification has not been established for Cu and Co.

This European Standard specifies a method for the determination of N-(1,2-dicarboxyethyl)-D,L-aspartic acid (Iminodisuccinic acid (IDHA)) in fertilizers.
The method is applicable to all fertilizers containing IDHA as chelating agent for contents > 0,5 % (g/100 g).

This Technical Report covers the results from two ring tests (A and B, see Introduction) to compare the repeatability and reproducibility of the standardized method for the determination of EDDHA and EDDHMA by ion chromatography (EN 13368 2:2001) with a non-standardized alternative method using ion-pair chromatography ("Lucena-method", given in Annex A). This Technical Report also provides the results of an univariate multifactorial ANOVA test and F-test for ring test A and F-test for ring test B to verify the significance of the reproducibility and the repeatability data.

This standard defines a method for determination of the chelated trace element content and the chelated fraction of the trace elements (micronutrients) cobalt, copper, iron, manganese, and zinc in fertilizers. This method applies to fertilizers, covered by Directive 89/530/EEC, containing one or more of the trace elements cobalt, copper, iron, manganese, and zinc, chelated by one or more chelating agents of the group of the polyamino polycarboxylic acids.

This document specifies a method for the chromatographic determination of the iron chelated by each individual ortho(hydroxy)-ortho(hydroxy) isomer of the chelating agents [o,o] EDDHA, [o,o] EDDHMA and by HBED in fertilizers containing one or more of these substances, except for [o,o] EDDHMA and HBED mixes. The method allows the identification and the determination of the total concentration of water soluble iron chelates of these chelating agents. It does not determine the free form of the chelating agents.
This method is applicable to EC fertilizers covered by Regulation (EC) No 2003/2003 [4]. It is applicable to a mass fraction of the metal chelated of at least 0,625 %.
NOTE 1   The substances EDDHA (ethylenediamine-N,N’-di[(hydroxyphenyl)acetic acid] and EDDHMA (ethylenediamine-N,N’-di[(hydroxymethylphenyl)acetic acid] exist as several different isomeric forms. Positional isomers for the hydroxyl or methyl groups (in ortho, meta, and para positions) as well as stereo isomers (meso and dl-racemic forms) are known. Both meso and dl-racemic forms of the ortho,ortho-EDDHA and ortho,ortho-EDDHMA are positional isomers for the hydroxyl groups allowed by the Regulation (EC) No 2003/2003. Since para, meta and ortho methyl positional isomers of the EDDHMA present quite similar stability, they could be grouped: in the method here described the para, meta and ortho methyl positional isomers of the [o,o] EDDHMA are considered together. HBED (N,N´-bis(2-hydroxybenzyl)-ethylenediamine-N,N´-diacetic acid) does not present isomeric forms.
NOTE 2   At present, analytically pure standards only exist for ortho,ortho-EDDHA, ortho,ortho-EDDHMA and HBED. All other substances being unavailable as a standard, the influence of their eventual presence in the samples (with respect to the sensitivity and the selectivity of this method) has not been studied.
NOTE 3   The meso and the dl-racemic forms of [o,o] EDDHA and [o,o] EDDHMA can be determined separately by this method.

This method describes the procedure for the ion chromatographic determination of the total amount of each individual chelating agent EDTA, HEDTA, and DTPA in fertilizers, containing one or more of these substances. The method allows the identification and the determination of the total water soluble fraction of each of these chelating agents. It does not allow to distinguish between the free form and the metal bound form of the chelating agent.

This document specifies a method for the chromatographic determination of the iron chelated by each individual ortho(hydroxy)-ortho(hydroxy) isomer of the chelating agents o,o-EDDHA and o,o-EDDHMA in fertilizers containing one or both of these substances. The method allows the identification and the determination of the total concentration of water soluble iron chelates of these chelating agents. It does not determine the free form of the chelating agents.
NOTE 1   The substances EDDHA (ethylene diamine di(hydroxy phenyl) acetic acid) and EDDHMA (ethylene diamine di(hydroxy methyl phenyl)acetic acid) exist as several different isomeric forms. Positional isomers for the hydroxyl or methyl groups (in ortho, meta, and para positions) as well as stereo isomers (meso and dl-racemic forms) are known. Both meso and dl-racemic forms of the ortho,ortho-EDDHA and ortho,ortho-EDDHMA are positional isomers for the hydroxyl groups allowed by the Regulation (EC) No 2003/2003. Since para, meta and ortho methyl positional isomers of the EDDHMA present quite similar stability, they could be grouped: in the method here described the para, meta and ortho methyl positional isomers of the o,o-EDDHMA are considered together.
NOTE 2   At present, analytically pure standards only exist for ortho,ortho-EDDHA and ortho,ortho-EDDHMA. All other substances being unavailable as a standard, the influence of their eventual presence in the samples (with respect to the sensitivity and the selectivity of this method) has not been studied.
NOTE 3   The meso and the dl-racemic forms of o,o-EDDHA and o,o-EDDHMA can be determined separately by this method.
This procedure concerns EC fertilizers covered by Regulation (EC) No 2003/2003 [5]. It is applicable to a mass fraction of the metal chelated of at least 0,625 %.

This Technical Specification specifies a method for the chromatographic determination of the total amount of iron chelated by EDDHSA in commercial products.

This Technical Specification specifies a method for the chromatographic determination of the amount of iron chelated by each of the individual isomers of the chelating agent ortho para EDDHA (o,p-EDDHA) in fertilizers. The method allows the identification of this chelating agent and the determination of the water soluble fraction of iron chelated by this chelating agent. The method is not applicable for the determination of the amount of free chelating agent.
NOTE 1   This method has been shown to be also suitable for the determination of the amount of iron chelated by each of the individual isomers of the chelating agent ortho ortho EDDHA (o,o-EDDHA) in fertilizers.
NOTE 2   o,o-EDDHA and o,p-EDDHA are abbreviations used in this standard for the sake of simplicity. For complete names see Annex A.
NOTE 3   The substances o,o-EDDHA and o,p-EDDHA both exist as different stereoisomers. For o,o-EDDHA a meso form and a d/l pair (the racemic isomers) exist, for o,p-EDDHA two different d/l pairs exist. All four stereoisomers are observed separately in this method.
NOTE 4   Presently, an analytically pure standard only exists for o,o-EDDHA. The method for o,p-EDDHA has been developed with an o,p-EDDHA standard containing an uncertain concentration of o,p-EDDHA.

This method describes the procedure for the ion chromatographic determination of the total amount of each individual ortho-ortho isomer of the chelating agents EDDHA and EDDHMA in fertilizers, containing one or both of these substances. The method allows the identification and the determination of the total water soluble fraction of each of these chelating agents. It does not allow to distinguish between the free form and the metal bound form of the chelating agents.