ISO 25705-2016 Fertilizers - Determination of urea condensates using high-performance liquid chromatography (HPLC) - Isobutylidenediurea and crotonylidenediurea.pdf

1、 ISO 2016 Fertilizers Determination of urea condensates using high- performance liquid chromatography (HPLC) Isobutylidenediurea and crotonylidenediurea (method A) and methylen-urea oligomers (method B) Engrais Dosage des condensats dure par chromatographie liquide haute performance (CLHP) Isobutyli

2、dne diure et crotonylidne diure (mthode A) et oligomres de mthylne-ure (mthode B) INTERNATIONAL STANDARD ISO 25705 First edition 2016-04-01 Reference number ISO 25705:2016(E) ISO 25705:2016(E)ii ISO 2016 All rights reserved COPYRIGHT PROTECTED DOCUMENT ISO 2016, Published in Switzerland All rights r

3、eserved. 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

4、 ISO at the address below or ISOs 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 copyrightiso.org www.iso.org ISO 25705:2016(E)Foreword iv Introduction v 1 Scope . 1 2 Normat

5、ive references 1 3 T erms and definitions . 1 4 Sampling and sample preparation 1 5 Method A: Determination of CDU and IBDU . 1 5.1 Principle 1 5.2 Reagents 1 5.3 Apparatus 2 5.4 Procedure 2 5.4.1 System parameters of HPLC . 2 5.4.2 Calibration . 2 5.4.3 Preparation of the test portion 3 5.4.4 Measu

6、rement 3 5.4.5 Important annotations . 3 5.5 Calculation 3 6 Method B: Determination of methylene-urea oligomers (MU) 4 6.1 Principle 4 6.2 Reagents 4 6.3 Apparatus 4 6.4 Procedure 5 6.4.1 System parameters of HPLC . 5 6.4.2 Calibration . 5 6.4.3 Preparation of the test solution 6 6.4.4 Measurement

7、7 6.4.5 Important annotations . 7 6.5 Calculations 7 7 Precision method A and method B 8 7.1 Inter-laboratory test. 8 7.2 Repeatability . 8 7.3 Reproducibility . 8 8 Test report . 9 Annex A (informative) Results of the inter-laboratory tests 10 Annex B (informative) Chromatogram and calibration curv

8、es method A 12 Annex C (informative) Chromatogram and calibration curves method B.14 Bibliography .17 ISO 2016 All rights reserved iii Contents Page ISO 25705:2016(E) Foreword ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies (ISO member

9、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

10、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

11、 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 d

12、rawn 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 o

13、n 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 asses

14、sment, as well as information about ISOs adherence to the WTO principles in the Technical Barriers to Trade (TBT) see the following URL: Foreword - Supplementary information ISO 25705 was prepared by CEN/TC 260 (as EN 15705:2010) and was adopted by Technical Committee ISO/TC 134, Fertilizers and soi

15、l conditioners. The following modifications were made. The references to EN 1482-1 and EN 1482-2 were changed to ISO 14820-1 and ISO 14820-2. In 5.2.1 and 6.2.1, the general text (not related to the water) was moved directly under 5.2 and 6.2, respectively. In Table 1, the word “approximate” was add

16、ed to the column headers for IBDU and CDU. In 5.4.3 and 6.4.3, “the sample grounded” was changed to “the ground sample”. In 6.4.3, “pieces of glass” and “pieces” were replaced by “boiling stones”. In 6.4.2.1, 6.4.2.2, 6.4.2.3, and 6.4.2.4, “by placing the flask in the” was added before “ultrasonic b

17、ath”. In 6.4.2.1, 6.4.2.2, 6.4.2.3, 6.4.2.4, 6.4.2.5 and 6.4.3, “homogenize” was changed to “mix thoroughly”. In 6.4.2.5, “before transferring into” was changed to “before transferring in” three times. In the keys for Figures B.2, B.3, B.4, C.2, C.3, C.4 and C.5, the units for the areas were added.i

18、v ISO 2016 All rights reserved ISO 25705:2016(E) Introduction Fertilizers containing the condensates of urea and specified aldehydes (with crotonaldehyde called crotonyliden diurea or CDU, with isobutyraldehyde called isobutylidene diurea or IBDU, with formaldehyde called urea formaldehyde or methyl

19、ene urea or MU) are covered by Regulation (EC) 2003/2003, Annex I 4as nitrogenous fertilizers. The methods described in this International Standard enable the quantitative determination of these condensates and the determination of the solubility of the MU-oligomers according to the Regulation. ISO

20、2016 All rights reserved v Fertilizers Determination of urea condensates using high-performance liquid chromatography (HPLC) Isobutylidenediurea and crotonylidenediurea (method A) and methylen-urea oligomers (method B) 1 Scope This International Standard specifies methods for the determination of is

21、obutylidene diurea (IBDU), Crotonylidene diurea (CDU) (method A) and methylene-urea oligomers (MU) (method B) in fertilizers using high-performance liquid chromatography (HPLC). The method is applicable to all fertilizers which do not contain interfering organic compounds. 2 Normative references The

22、 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. ISO 3696,

23、Water for analytical laboratory use Specification and test methods ISO 14820-2, Fertilizers and liming materials Sampling and sample preparation Part 2: Sample preparation EN 12944-1, Fertilizers and liming materials and soil improvers Vocabulary Part 1: General terms EN 12944-2, Fertilizers and lim

24、ing materials and soil improvers Vocabulary Part 2: Terms relating to fertilizers 3 T erms a nd definiti ons For the purposes of this document, the terms and definitions given in EN 12944-1 and EN 12944-2 apply. 4 Sampling and sample preparation Sampling is not part of the method specified in this I

25、nternational Standard. A recommended sampling method is given in ISO 14820-1. Sample preparation shall be carried out in accordance with ISO 14820-2. 5 Method A: Determination of CDU and IBDU 5.1 Principle The sample is extracted with water and, after appropriate dilution, analysed using a suitable

26、HPLC system. 5.2 Reagents Use only reagents of recognized analytical grade, unless otherwise specified. 5.2.1 Water, distilled or demineralized water (grade 3 according to ISO 3696). INTERNATIONAL ST ANDARD ISO 25705:2016(E) ISO 2016 All rights reserved 1 ISO 25705:2016(E) 5.2.2 Acetonitrile, HPLC-g

27、rade. 5.2.3 Isobutylidene diurea and Crotonylidene diurea, in their pure form. 5.3 Apparatus 5.3.1 Laboratory equipment and glassware, for preparation of solutions and dilutions. 5.3.2 Analytical balance, capable of weighing to an accuracy of 0,1 mg. 5.3.3 HPLC-system, with UV-detector. 5.3.4 Ultras

28、onic bath. 5.3.5 Magnetic stirrer. 5.3.6 Disposable filt er, 0,45 m. 5.4 Procedure 5.4.1 System parameters of HPLC Analytical/separating column: silica column with C18 reverse phase 1) Detection wavelength: 200 nm Eluent: acetonitrile/water: 10/90 (volume fraction) Flow rate: 1 ml/min Temperature: a

29、mbient temperature Injection volume: 20 l 5.4.2 Calibration 5.4.2.1 Stock solution IBDU (IBDU) = 100 mg/l Weigh 100/R mg of IBDU (5.2.3), where R is the purity of IBDU, into a 1 000 ml flask and add about 900 ml of water (5.2.1). Dissolve in an ultrasonic bath (5.3.4) for about 10 min, followed by s

30、tirring on a magnetic stirrer (5.3.5) for about 1 h. Make up to volume. Filtration is not necessary. 5.4.2.2 Stock solution CDU (CDU) = 100 mg/l Weigh 100/R mg of CDU (5.2.3), where R is the purity of CDU, into a 1 000 ml flask and add about 900 ml of water (5.2.1). Dissolve in an ultrasonic bath (5

31、.3.4) for about 10 min, followed by stirring on a magnetic stirrer (5.3.5) for about 1 h. Make up to volume. Filtration is not necessary. 5.4.2.3 Calibration solution For calibration, prepare three solutions according to Table 1 using one-mark (bulb) pipettes and dilute to the mark with water (5.2.1

32、). 1) LiChrosorb RP-18 7 m 250/4 mm is an example of a suitable product available commercially. This information is given for the convenience of users of this International Standard and does not constitute an endorsement by ISO of this product.2 ISO 2016 All rights reserved ISO 25705:2016(E) For the

33、 determination of the retention time, dilute 10 ml of the stock solution (5.4.2.1) or respectively (5.4.2.2) into two 100 ml flasks and make up to volume with water (5.2.1). The evaluation of calibration is carried out manually or by means of a suitable PC-aided (computerized) calculation method. Ta

34、ble 1 Preparation of calibration solutions Parameter Amount of stock solution IBDU/CDU ml (to be added to the 100 ml flask) Content of IBDU mg/l (approximate) Content of CDU mg/l (approximate) Standard 1 10 10,0 10,0 Standard 2 25 25,0 25,0 Standard 3 50 50,0 50,0 5.4.3 Preparation of the test porti

35、on Weigh 1 g of the ground sample to 0,2 mm to the nearest 0,1 mg and quantitatively transfer into a 1 000 ml volumetric flask with water (5.2.1). Fill the flask to an amount of approximately 900 ml and treat it for 10 min in the ultrasonic bath (5.3.4). Then make up to the mark and stir for 1 h at

36、room temperature on a magnetic stirrer (5.3.5). Dilute 10 ml of the solution in a 100 ml volumetric flask and filter into the HPLC injection vial through a disposable filter (5.3.6). 5.4.4 Measurement Measurement is performed manually or by means of an automatic sample loading system (autosampler).

37、5.4.5 Important annotations IBDU is able to form urea in aqueous solution. Therefore, the measurement of the calibration and sample solutions shall be completed within one working day. The concentrations of CDU and IBDU in the sample solutions shall be kept within the calibration limits (5.4.2) to e

38、nsure sufficient reproducibility. 5.5 Calculation The calculation can be performed manually or by means of a PC using the calibration parameters in respect to the amount used. In the case of PC-aided (computerized) calculation and application of Table 1 regarding the amounts of stock solution, the c

39、ontent of IBDU/CDU in milligrams per litre will be calculated by the system. The calculated values are equal to the percentage mass concentration of IBDU/CDU in the analysed sample of fertilizer. Following general rules for declaration in regulations to declare the content of the compounds as percen

40、tage mass fraction of nitrogen, calculate the contents, w N(IBDU) /w N(CDU)in percent (g/100 g), according to Formulae (1) and (2): ww (1) ww (2) where ISO 2016 All rights reserved 3 ISO 25705:2016(E)0,322 is the conversion factor for the content of IBDU in the fertilizer into nitrogen content;0,326

41、 is the conversion factor for the content of CDU in the fertilizer into nitrogen content. 6 Method B: Determination of methylene-urea oligomers (MU) NOTE By the condensation of urea and formaldehyde several oligomers like methylenediurea (MDU), dimethylenetriurea (DMTU), trimethylene-tetraurea (TMTU

42、) and higher oligomers are formed. These three molecules are the most soluble in water, the higher compounds are insoluble in hot water, but their nitrogen is available for plants by microbiological decomposition. Also, urea is always a companion of MU oligomers. 6.1 Principle The sample is extracte

43、d with boiling water and analysed using a suitable HPLC system. The methylene-urea soluble oligomers are measured and detected by the HPLC-method. In the HPLC-diagram, methylene-urea oligomers are represented by different peaks: urea, methylene- diurea, dimethylenetriurea; and trimethylene tetraurea

44、 are, in the meantime, the most soluble and important. 6.2 Reagents Use only reagents of recognized analytical grade, unless otherwise specified. 6.2.1 Water, distilled or demineralized water (grade 3 according to ISO 3696). 6.2.2 Acetonitrile, HPLC-grade p.a. should not be used here. It is not comm

45、on. 6.2.3 Urea, reagent grade (or analytical grade), 46,6 % of total nitrogen. 6.2.4 Methylene diurea (MDU), synthesized and purified by a special laboratory, 42,4 % of total nitrogen. 2) 6.2.5 Dimethylenetriurea (DMTU), synthesized and purified by a special laboratory, 41,2 % of total nitrogen. 2)

46、6.2.6 Trimethylene tetraurea (TMTU), synthesized and purified by a special laboratory, 40,6 % of total nitrogen. 6.3 Apparatus 6.3.1 Laboratory equipment and glassware, for preparation of solutions and dilutions. 6.3.2 Analytical balance, capable of weighing to an accuracy of 0,1 mg. 6.3.3 Technical

47、 balance, capable of weighing to an accuracy of 0,01 g. 6.3.4 HPLC-system, equipped with an UV-detector. 6.3.5 Ultrasonic bath. 2) The standard substances MDU and DMTU can be prepared according to the method given in Official Methods of Analysis of AOAC International, AOAC Official Method 983.01, JA

48、OAC 66, 769 (1983).4 ISO 2016 All rights reserved ISO 25705:2016(E) 6.3.6 Magnetic stirrer. 6.3.7 Disposable filt er , 0,45 m. 6.4 Procedure 6.4.1 System parameters of HPLC Analytical/separating column NH 2column, 5 m, 250 mm 4,6 mm 3)A guard-column is recommended. Detection wavelength 195 nm (Diode

49、 Array detector) Eluent acetonitrile/water 85/15 (volume fraction) Flow rate 1 ml/min Temperature 60 C Run time 30 min Injection volume 20 l 6.4.2 Calibration 6.4.2.1 Stock solution of urea, 1 000 mg/kg Weigh (6.3.2) 100/R mg of urea (6.2.3), where R is the purity of urea, to the nearest 0,1 mg and put into a clean and dry 100 ml volumetric flask, weighed ( 6.3.3) before to the nearest 0,01 g. Add 50 ml of water (6.2.1) and disso