1、BSI Standards PublicationBS ISO 16308:2014Water quality Determinationof glyphosate and AMPA Method using highperformance liquidchromatography (HPLC) withtandem mass spectrometricdetectionBS ISO 16308:2014 BRITISH STANDARDNational forewordThis British Standard is the UK implementation of ISO 16308:20
2、14.The UK participation in its preparation was entrusted to TechnicalCommittee EH/3/2, Physical chemical and biochemical methods.A list of organizations represented on this committee can beobtained on request to its secretary.This publication does not purport to include all the necessaryprovisions o
3、f a contract. Users are responsible for its correctapplication. The British Standards Institution 2014. Published by BSI StandardsLimited 2014ISBN 978 0 580 71830 4ICS 13.060.50Compliance with a British Standard cannot confer immunity fromlegal obligations.This British Standard was published under t
4、he authority of theStandards Policy and Strategy Committee on 30 September 2014.Amendments issued since publicationDate Text affectedBS ISO 16308:2014 ISO 2014Water quality Determination of glyphosate and AMPA Method using high performance liquid chromatography (HPLC) with tandem mass spectrometric
5、detectionQualit de leau Dtermination du glyphosate et de lAMPA Mthode par chromatographie en phase liquide haute performance (CLHP) avec dtection par spectromtrie de masse en tandemINTERNATIONAL STANDARDISO16308First edition2014-09-15Reference numberISO 16308:2014(E)BS ISO 16308:2014ISO 16308:2014(E
6、)ii ISO 2014 All rights reservedCOPYRIGHT PROTECTED DOCUMENT ISO 2014All rights reserved. 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 in
7、tranet, without prior written permission. Permission can be requested from either ISO at the address below or ISOs member body in the country of the requester.ISO copyright officeCase postale 56 CH-1211 Geneva 20Tel. + 41 22 749 01 11Fax + 41 22 749 09 47E-mail copyrightiso.orgWeb www.iso.orgPublish
8、ed in SwitzerlandBS ISO 16308:2014ISO 16308:2014(E) ISO 2014 All rights reserved iiiContents PageForeword ivIntroduction v1 Scope . 12 Normative references 13 Principle 14 Interferences 25 Reagents 26 Apparatus . 47 Sampling 58 Procedure. 58.1 Pre-treatment (Suspended particular matter) 58.2 Chelate
9、 break and derivatization 58.3 Pre-concentration . 68.4 Chromatographic determination. 78.5 Identification and confirmation of the analytes . 88.6 Blank control monitoring. 89 Calibration 89.1 Concentration ranges 89.2 Matrix-matched calibration . 89.3 Internal standard calibration . 910 Expression
10、of results .1011 Test report 10Annex A (informative) Performance data 11Annex B (informative) Examples of chromatographic conditions 14Annex C (informative) Examples of chromatograms 15Annex D (informative) Analysis of gluphosinate .16Annex E (informative) Pre-treatment of hard water samples .20Bibl
11、iography .22BS ISO 16308:2014ISO 16308:2014(E)ForewordISO (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 memb
12、er 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 Ele
13、ctrotechnical Commission (IEC) on all matters of electrotechnical standardization.The procedures used to develop this document and those intended for its further maintenance are described in the ISO/IEC Directives, Part 1. In particular the different approval criteria needed for the different types
14、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 drawn to the possibility that some of the elements of this document may be the subject of patent rights. ISO shall not be he
15、ld 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 on the ISO list of patent declarations received (see www.iso.org/patents). Any trade name used in this document is informati
16、on 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 assessment, as well as information about ISOs adherence to the WTO principles in the Technical Barriers to Trade (TBT) see the fo
17、llowing URL: Foreword - Supplementary informationThe committee responsible for this document is ISO/TC 147, Water quality, Subcommittee SC 2, Physical, chemical and biochemical methods.iv ISO 2014 All rights reservedBS ISO 16308:2014ISO 16308:2014(E)IntroductionGlyphosate N-(phosphonomethyl)glycine
18、is a non-selective broad-spectrum herbicide. The efficiency of this compound makes it a top selling and one of the most widely used herbicides in the world since it entered the market in 1974. Together with its main degradation product, aminomethylphosphonic acid (AMPA), glyphosate is one of the mos
19、t detected substances in water bodies in many developed countries. Note also that AMPA can be released during sewage treatment, e.g. due to breakdown of detergent formulations for textiles.Glyphosate and AMPA belong to the aminophosphonate family and have specific physico-chemical properties that re
20、quire the development of complex analytical methods for analysis and detection. The difficulty in analysis is mainly linked to the high solubility of glyphosate and AMPA and their chelating nature. To solve these problems, their pre-column derivatization with 9-fluorenylmethylchloroformate (FMOC-Cl)
21、 to form less polar derivatives allows a better separation using liquid chromatography.Gluphosinate, another aminophosphonate, is less commonly subject to regulation and can be determined simultaneously, provided it can be demonstrated that there is no interference with the sample under analysis.The
22、re is currently an International Standard for the determination by liquid chromatography and fluorometric detection; however, the determination by HPLCESIMS/MS can be much more specific (unambiguous identification) and more sensitive (limits of quantification of approximately 30 ng/l for both glypho
23、sate and AMPA). This International Standard is based on this analytical technique and is intended for laboratories involved in the regulatory control of the aquatic environment. Many such laboratories are now equipped with this kind of apparatus. ISO 2014 All rights reserved vBS ISO 16308:2014BS ISO
24、 16308:2014Water quality Determination of glyphosate and AMPA Method using high performance liquid chromatography (HPLC) with tandem mass spectrometric detectionWARNING Persons using this International Standard should be familiar with normal laboratory practice. This International Standard does not
25、purport to address all of the safety problems, if any, associated with its use. It is the responsibility of the user to establish appropriate safety and health practices and to ensure compliance with any national regulatory conditions. IMPORTANT It is absolutely essential that tests conducted in acc
26、ordance with this International Standard be carried out by suitably qualified staff.1 ScopeThis International Standard specifies a method for the determination of dissolved fraction of glyphosate and its major metabolite, aminomethylphosphonic acid (AMPA), in drinking water, ground water, and surfac
27、e water at concentrations of 0,03 g/l to 1,5 g/l. It does not apply to marine or salty water. This method can be applicable to other types of waters, provided the method is validated for each case.2 Normative referencesThe following documents, in whole or in part, are normatively referenced in this
28、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, Water for analytical laboratory use Specification and test methodsISO 5667-3, W
29、ater quality Sampling Part 3: Preservation and handling of water samplesISO 8466-1, Water quality Calibration and evaluation of analytical methods and estimation of performance characteristics Part 1: Statistical evaluation of the linear calibration function3 PrincipleGlyphosate and AMPA (dissolved
30、fraction after filtration) are derivatized using 9-fluorenylmethylchloroformate (FMOC-Cl) (5.17) to lower their polarity and increase the retention of compound in a separation on a reverse phase column (e.g. C18) as well as to improve the mass spectrometric detection. If the mass spectrometer has su
31、fficient detection capability, it is possible to omit the solid phase extraction and to analyse the analytes by direct injection (see 8.2.1).The derivatized sample is purified by liquid/liquid extraction and then concentrated by solid phase extraction (SPE).The analysis is performed by high performa
32、nce liquid chromatography coupled with tandem mass spectrometry via an electrospray source (HPLCESIMS/MS), using matrix-matched calibration.INTERNATIONAL STANDARD ISO 16308:2014(E) ISO 2014 All rights reserved 1BS ISO 16308:2014ISO 16308:2014(E)Table 1 Substances addressedName FormulaMolecular mass
33、g/molCAS-RNaGlyphosateN-(phosphonomethyl)glycineC3H8NO5P 169,1 1071836AMPAAminomethylphosphonic acidCH6NO3P 111,0 1066519aCAS-RN Chemical Abstracts Service Registry NumberNOTE Gluphosinate, belonging to the aminophosphonate family, can be determined simultaneously, provided it can be demonstrated th
34、at there is no interference with the sample (matrix) subject to analysis.4 InterferencesThis method is validated for hard water containing up to 3,2 mmol/l of the sum of calcium and magnesium. For waters with a higher calcium and magnesium content, it may be necessary to increase the concentration o
35、f EDTA-Na2(5.16) at the derivatization step (see Annex D).It can prove necessary to include the acidification step described in Annex D even for some water types below 3,2 mmol/l of the sum of calcium and magnesium. The laboratory shall check the necessity of this procedure for its routine samples.T
36、he presence of free chlorine, e.g. in treated waters, can cause losses of glyphosate by oxidation. Consequently sodium thiosulfate shall be used (see Clause 7).5 ReagentsUnless otherwise stated, all reagents and solvents shall be of sufficient purity, e.g. “for trace analysis”.5.1 Deionized water.5.
37、2 Ultra-pure water, complying with grade 1 of ISO 3696.5.3 Nitrogen, N2, purity 99,996 % volume fraction.5.4 Laboratory detergent, alkaline.5.5 Sodium thiosulfate, Na2S2O3.5.6 Acetonitrile, C2H3N, HPLC grade.5.7 Methanol, CH4O, HPLC grade.5.8 Ethanol, C2H6O, 95 %, HPLC grade mass fraction.5.9 Ethyl
38、acetate, C4H8O2, HPLC grade.5.10 Ammonium acetate, C2H7O2N.5.11 Triethylamine, C6H15N.5.12 Ammonium hydroxide, NH4OH,28 % mass fraction.2 ISO 2014 All rights reservedBS ISO 16308:2014ISO 16308:2014(E)5.13 Formic acid, CH2O2.5.14 Hydrochloric acid, HCl, 300 g/l.5.15 Glacial acetic acid, C2H4O2.5.16 E
39、thylenediaminetetraacetic acid (EDTA), disodium salt dihydrate, C10H14N2O8Na22H2O, with a minimum purity of 99 % mass fraction.5.17 9-Fluorenylmethyl chloroformate,(FMOC-Cl), C15H11ClO2, with a minimum purity of 97 % mass fraction.FMOC-Cl is used to prepare the derivatization reagent, FMOC-Cl soluti
40、on, 50 mg/ml, in acetonitrile (5.6). This solution can be stored at 18 C 3 C for 6 months.For direct injection (8.2.1), use a FMOC-Cl solution of 0,5 mg/ml in acetonitrile.5.18 Reference substances, according to Table 1.5.18.1 Glyphosate, N-(phosphonomethyl)glycine, C3H8NO5P, purity 98 % mass fracti
41、on.5.18.2 AMPA, aminomethylphosphonic acid, CH6NO3P, purity 98 % mass fraction.5.18.3 1,2-13C2,15N-labelled glyphosate, surrogate standard, purity 98 % mass fraction.5.18.4 13C,15N-labelled AMPA, surrogate standard, purity 98 % mass fraction.5.19 Calibration solutions.Individual stock solutions of g
42、lyphosate (5.18.1) and AMPA (5.18.2), 100 mg/l, prepared in ultra-pure water (5.2). These solutions can be stored at 4 C 3 C for 1 month.Individual stock solutions of 1,2-13C2,15N-labelled glyphosate (5.18.3) and 13C,15N-labelled AMPA (5.18.4) 100 mg/l, prepared in ultra-pure water (5.2). These solu
43、tions can be stored at 4 C 3 C for 1 month.Multi-substance working solution of surrogates: 1,2-13C2,15N-labelled glyphosate and 13C,15N-labelled AMPA, 20 g/l, prepared in ultra-pure water (5.2). This solution can be stored at 4 C 3 C for 1 month.NOTE Stock and calibration solutions can be stored lon
44、ger, provided the adequate justifications are given regarding stability.5.20 Triethylammonium acetate buffer, 0,1 % triethylamine (5.11) solution adjusted to pH 9,5 with glacial acetic acid (5.15) (mobile phase).5.21 Sodium tetraborate, decahydrate, Na2B4O710H2O.5.22 Borate-Na buffer, 0,05 mol/l; pH
45、 = 9,2.Dissolve 19 0,1 g of sodium tetraborate (5.21), decahydrate in 1 l of water (5.1). This solution can be stored for approximately 1 mo at 4 C 3 C.5.23 Mineral water, containing less than 3,2 mmol/l divalent cations (Mg2+and total Ca2+), for preparing matrix-matched calibration. ISO 2014 All ri
46、ghts reserved 3BS ISO 16308:2014ISO 16308:2014(E)6 ApparatusThe material or any parts that are likely come into contact with the sample shall be free from any residue that could cause unacceptable interference in the blanks.Glass and plastics containers can be used for sampling and for all steps bef
47、ore derivatization. Glass vials (6.10) and glass test tubes (6.11) shall be used after the derivatization step.6.1 Usual laboratory glassware, or apparatus and in particular the following.6.2 Glass, polyethene (PE) or polypropene (PP) bottles, minimum 50 ml, for sampling.6.3 Glass, polyethene (PE) o
48、r polypropene (PP) syringe, 50 ml, for sample filtration.6.4 Single use filter for syringe, diameter 25 mm, with a hydrophilic membrane, 0,45 m, e.g. from regenerated cellulose.6.5 Glass, or single use PE or PP conical bottomed tubes, approximately 50 ml, for derivatization.6.6 Micropipettes, adjust
49、able from 100 l to 500 l.6.7 pH-meter.6.8 SPE cartridges, e.g. Oasis HLB1)Waters, 60 mg, 3 ml, or equivalent.6.9 Centrifugation device, capable of 6 500 m1.6.10 Glass vials, suitable for autosampler, with caps and polytetrafluoroethene (PTFE) or silicon rubber septa.6.11 Glass test tube, 15 ml or smaller.6.12 Reversed phase column, e.g. XBridge C181)column Waters, 50 mm 2,1 mm internal diameter (i.d.) 2,5 m, column with guard column (Waters, 10 mm 2,1 mm i.d. 2,5 m).A
