1、raising standards worldwideNO COPYING WITHOUT BSI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAWBSI Standards PublicationFertilizers Determination of 3,4-dimethyl-1H-pyrazole phosphate (DMPP) Methodusing high-performance liquidchromatography (HPLC)BS EN 16328:2012National forewordThis British Stand
2、ard is the UK implementation of EN 16328:2012.The UK participation in its preparation was entrusted to Technical CommitteeCII/37, Fertilisers and related chemicals.A list of organizations represented on this committee can be obtained on request to its secretary.This publication does not purport to i
3、nclude all the necessary provisions of a contract. Users are responsible for its correct application. The British Standards Institution 2013.Published by BSI Standards Limited 2013 ISBN 978 0 580 76198 0 ICS 65.080Compliance with a British Standard cannot confer immunity from legal obligations.This
4、British Standard was published under the authority of the Standards Policy and Strategy Committee on 31 January 2013.Amendments issued since publicationDate Text affectedBRITISH STANDARDBS EN 16328:2012EUROPEAN STANDARD NORME EUROPENNE EUROPISCHE NORM EN 16328 December 2012 ICS 65.080 English Versio
5、n Fertilizers - Determination of 3,4-dimethyl-1H-pyrazole phosphate (DMPP) - Method using high-performance liquid chromatography (HPLC) Engrais - Dosage du 3,4-dimthyl-1H-pyrazole phosphate (DMPP) - Mthode par chromatographie liquide haute performance (HPLC) Dngemittel - Bestimmung von 3,4-Dimethyl-
6、1H-pyrazolphosphat (DMPP) - Verfahren mit Hochleistungs-Flssigchromatographie (HPLC) This European Standard was approved by CEN on 27 October 2012. CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this European Standard the status of
7、 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)
8、. 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, C
9、yprus, 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, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and United
10、Kingdom. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN DE NORMALISATION EUROPISCHES KOMITEE FR NORMUNG Management Centre: Avenue Marnix 17, B-1000 Brussels 2012 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Members. Ref. No. EN 16328:2012: EBS
11、 EN 16328:2012EN 16328:2012 (E) 2 Contents Page Foreword 31 Scope 42 Normative references 43 Terms and definitions .44 Principle 45 Reagents .46 Apparatus .47 Sampling .58 Procedure .58.1 Preparation of the test solution 58.2 Preparation of calibration solution 58.3 HPLC conditions 58.4 HPLC determi
12、nation 69 Calculation and expression of the results 610 Precision .610.1 Inter-laboratory test .610.2 Repeatability .610.3 Reproducibility .611 Test report 7Annex A (informative) Results of the inter-laboratory test .8A.1 Inter-laboratory tests .8A.2 Statistical results .8Bibliography 9BS EN 16328:2
13、012EN 16328:2012 (E) 3 Foreword This document (EN 16328:2012) 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 identica
14、l text or by endorsement, at the latest by June 2013, and conflicting national standards shall be withdrawn at the latest by June 2013. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. CEN and/or CENELEC shall not be held responsib
15、le 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. According to the CEN/CENELEC Internal Regulations, the national standards organisations of the following countries are bo
16、und 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, Portug
17、al, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United Kingdom. BS EN 16328:2012EN 16328:2012 (E) 4 1 Scope This European Standard specifies a method for the determination of 3,4-dimethyl-1H-pyrazole phosphate (DMPP, CAS-No: 202842-98-6) in mineral N containing fertilizer
18、s using high-performance liquid chromatography (HPLC). 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
19、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, Fertili
20、zers and liming materials and soil improvers Vocabulary Part 2: Terms relating to fertilizers 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 fertilizer sample is dissolved in or extracted with w
21、ater. The concentration of 3,4-dimethyl-1H-pyrazole phosphate is determined in diluted aqueous solution by high performance liquid chromatography (HPLC) at a reversed phase using a UV detector. 5 Reagents WARNING The described method includes the handling with hazardous materials. Attention must be
22、paid to technical, organizational and personal safety measures. Use only reagents of recognized analytical grade. 5.1 Water, de-ionized. 5.2 Acetonitrile, HPLC grade. 5.3 Sodiumdihydrogenphosphate-monohydrate, NaH2PO4H2O. 5.4 3,4-dimethyl-1H-pyrazole, 99 % purity. 6 Apparatus 6.1 Ultrasonic bath and
23、 magnetic stirrer, respectively. BS EN 16328:2012EN 16328:2012 (E) 5 6.2 Filtration unit, manual or high-pressure filtration with membrane filter 0,45 m or comparable filter. NOTE High pressure filtration is advantageous for difficult samples. 6.3 HPLC unit, for isocratic operation with UV detector.
24、 6.4 Evaluation unit. 6.5 Reverted phase-HPLC separation column, e.g. C18, 5 m, 150 mm 4,6 mm. NOTE With satisfactory separation performance, a shorter separation column can be used. 6.6 Analytical balance, capable of weighing to an accuracy of 0,001 g. 7 Sampling Sampling is not part of the method
25、specified in this document. A recommended sampling method is given in EN 1482-1. Sample preparation shall be carried out in accordance with EN 1482-2. 8 Procedure 8.1 Preparation of the test solution Weigh (6.6) (30 0,001) g of sample (m) into a 1 000 ml volumetric flask. Add approximately 900 ml wa
26、ter (5.1) and dissolve the sample in the ultrasonic bath (6.1) for about 15 min. Fill up the flask with water (5.1) to capacity. Filter part of the sample solution through a membrane filter (6.2) to remove possible conditioners. If necessary, dilute the sample for the calibration range. 8.2 Preparat
27、ion of calibration solution Prepare from at least two weighted samples (stock solutions) made with the present 3,4-dimethyl-1H-pyrazole (5.4) at least four concentrations for calibration. The concentration range should be between 3 mg/l and 100 mg/l. 8.3 HPLC conditions Separation column: filled for
28、 reversed phase-HPLC (6.5); column temperature: room temperature; elution agent: dissolve 1,38 g NaH2PO4H2O (5.3) in 1 l water and add 175 ml acetonitrile (5.2); the elution agent shall be de-gassed, e.g. in an ultrasonic bath (6.1); flow rate: 1,5 ml/min; injection volume: 10 l; wave length: 224 nm
29、; retention time DMPP (at 1,5 ml/min flow rate): approximately 5,3 min. BS EN 16328:2012EN 16328:2012 (E) 6 8.4 HPLC determination To prepare a calibration curve, inject 10 I of each calibration solution three times consecutively. Calculate the compensation curve and correlation coefficient p by lea
30、st-square method. The calibration curve can be used for concentration determination when p 0,99. Inject the test solution two times consecutively. 9 Calculation and expression of the results Carry out the evaluation on the basis of the calibration line over the peak areas. Calculate the content of 3
31、,4-dimethyl-1H-pyrazole phosphate, wDMPP, in mg/kg according to formula (1): 000101,962,194100)(dpkDMPP=maFbAw (1) where Apkis the peak height or peak area; b is the y-axis section of straight calibration line; a is the slope of straight calibration line; Fdis the dilution factor; m is the sample we
32、ight, in milligrams. Calculate the arithmetic mean from both values obtained. Report the result to the nearest 0,001 mg/kg. 10 Precision 10.1 Inter-laboratory test Details of inter laboratory tests performed in 2010 on the precision of the method are summarized in Annex A. The values derived from th
33、is test may not be applicable to concentration ranges and matrices other than those given. 10.2 Repeatability The absolute difference between two independent single test results, obtained using the same method on identical test material in the same laboratory by the same operator using the same equi
34、pment within a short interval of time, will in no more than 5 % of the cases be greater than the repeatability limit r given in Table 1. 10.3 Reproducibility The absolute difference between two single test results, obtained using the same method on identical test material in different laboratories w
35、ith different operators using different equipment, will in no more than 5 % of the cases be greater than the reproducibility limit R given in Table 1. BS EN 16328:2012EN 16328:2012 (E) 7 Table 1 Mean values, repeatability and reproducibility limits for DMPP Sample x r R mg/kg mg/kg mg/kgSample 1 Amm
36、onium sulfate nitrate (ASN) 1 700 90 170 Sample 2 NPK 24+8+7 1 270 70 410 Sample 3 NPK 13+10+20 706 56 210 11 Test report The test report shall contain at least the following information: a) all information necessary for the complete identification of the sample; b) test method used with reference t
37、o this document; c) test results obtained; d) date of sampling and sampling procedure (if known); e) date when the analysis was finished; f) whether the requirement of the repeatability limit has been fulfilled; g) all operating details not specified in this document, or regarded as optional, togeth
38、er with details of any incidents occurred when performing the method, which might have influenced the test result(s). BS EN 16328:2012EN 16328:2012 (E) 8 Annex A (informative) Results of the inter-laboratory test A.1 Inter-laboratory tests An inter-laboratory test has been carried out in 2010 with n
39、ine participating laboratories and three different samples of fertilizers. Repeatability and reproducibility were calculated according to ISO 5725-1 and ISO 5725-2. A.2 Statistical results The statistical results are given in Table A.1. Table A.1 Statistical results for the determination of DMPP usi
40、ng HPLC Parameter Sample 1 Sample 2 Sample 3 Ammonium sulfate nitrate (ASN) NPK 24+8+7 NPK 13+10+20Year of the test 2010 2010 2010 Number of participating laboratories 9 9 9 Number of laboratories after eliminating outliers 7 8 8 Mean value, x , (mg/kg) 1 700 1 270 706 Repeatability standard deviati
41、on sr, (mg/kg) 30 20 20Relative standard deviation of repeatability, RSDr2 1,9 3 Repeatability limit, r, (mg/kg) 90 70 56 Reproducibility standard deviation, sR,(mg/kg) 60 150 75Relative standard deviation of reproducibility, RSDR4 11,6 11 Reproducibility limit, R, (mg/kg) 170 410 210BS EN 16328:201
42、2EN 16328:2012 (E) 9 Bibliography 1 EN 1482-1, Fertilizers and liming materials Sampling and sample preparation Part 1: Sampling 2 EN ISO 3696:1995, Water for analytical laboratory use Specification and test methods (ISO 3696:1987) 3 ISO 5725-1, Accuracy (trueness and precision) of measurement metho
43、ds and results Part 1: General principles and definitions 4 ISO 5725-2, Accuracy (trueness and precision) of measurement methods and results Part 2: Basic method for the determination of repeatability and reproducibility of a standard measurement method 5 Regulation (EC) No 2003/2003 of the European
44、 Parliament and of the Council of 13 October 2003 relating to fertilisers, Official Journal L 304, 21/11/2003 P. 0001-0194 BS EN 16328:2012This page deliberately left blankThis page deliberately left blankBSI is the independent national body responsible for preparing British Standards and other stan
45、dards-related publications, information and services. It presents the UK view on standards in Europe and at the international level. BSI is incorporated by Royal Charter. British Standards and other standardisation products are published by BSI Standards Limited.British Standards Institution (BSI)ra
46、ising standards worldwideBSI389 Chiswick High Road London W4 4AL UKTel +44 (0)20 8996 9001Fax +44 (0)20 8996 Standards and PASs are periodically updated by amendment or revision. Users of British Standards and PASs should make sure that they possess the latest amendments or editions.It is the const
47、ant aim of BSI to improve the quality of our products and services. We would be grateful if anyone finding an inaccuracy or ambiguity while using British Standards would inform the Secretary of the technical committee responsible, the identity of which can be found on the inside frontcover. Similary
48、 for PASs, please notify BSI Customer Services.Tel: +44 (0)20 8996 9001 Fax: +44 (0)20 8996 7001BSI offers BSI Subscribing Members an individual updating service called PLUS which ensures that subscribers automatically receive the latest editions of British Standards and PASs.Tel: +44 (0)20 8996 766
49、9 Fax: +44 (0)20 8996 7001Email: Buying standardsYou may buy PDF and hard copy versions of standards directly using acredit card from the BSI Shop on the website addition all orders for BSI, international and foreign standards publicationscan be addressed to BSI Customer Services.Tel: +44 (0)20 8996 9001 Fax: +44 (0)20 8996 7001Email: In r
