1、Foods of plant origin - Multimethod for the determination of pesticide residues in vegetable oils by LC-MS/MSBS CEN/TS 17062:2017BSI Standards PublicationWB11885_BSI_StandardCovs_2013_AW.indd 1 15/05/2013 15:06TECHNICAL SPECIFICATION SPCIFICATION TECHNIQUE TECHNISCHE SPEZIFIKATION CEN/TS 17062 Septe
2、mber 2017 ICS 67.050 English Version Foods of plant origin - Multimethod for the determination of pesticide residues in vegetable oils by LC-MS/MS Aliments dorigine vgtale - Multimthode de dtermination de la teneur en rsidus de pesticides dans les huiles vgtales par CL-SM/SM Pflanzliche Lebensmittel
3、 - Multiverfahren zur Bestimmung von Pestizidrckstnden in pflanzlichen len mit LC-MS/MS This Technical Specification (CEN/TS) was approved by CEN on 11 May 2017 for provisional application. The period of validity of this CEN/TS is limited initially to three years. After two years the members of CEN
4、will be requested to submit their comments, particularly on the question whether the CEN/TS can be converted into a European Standard. CEN members are required to announce the existence of this CEN/TS in the same way as for an EN and to make the CEN/TS available promptly at national level in an appr
5、opriate form. It is permissible to keep conflicting national standards in force (in parallel to the CEN/TS) until the final decision about the possible conversion of the CEN/TS into an EN is reached. CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech
6、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 Kingd
7、om. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN DE NORMALISATION EUROPISCHES KOMITEE FR 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 worldwide for CEN national Members. Ref. No. CEN/TS 1706
8、2:2017 ENational forewordThis British Standard is the UK implementation of CEN/TS 17062:2017.The UK participation in its preparation was entrusted to Technical Committee AW/275, Food analysis - Horizontal methods.A list of organizations represented on this committee can be obtained on request to its
9、 secretary.This publication does not purport to include all the necessary provisions of a contract. Users are responsible for its correct application. The British Standards Institution 2017 Published by BSI Standards Limited 2017ISBN 978 0 580 94295 2ICS 67.050Compliance with a British Standard cann
10、ot confer immunity from legal obligations.This British Standard was published under the authority of the Standards Policy and Strategy Committee on 31 October 2017.Amendments/corrigenda issued since publicationDate Text affectedBRITISH STANDARDBS CEN/TS 17062:2017TECHNICAL SPECIFICATION SPCIFICATION
11、 TECHNIQUE TECHNISCHE SPEZIFIKATION CEN/TS 17062 September 2017 ICS 67.050 English Version Foods of plant origin - Multimethod for the determination of pesticide residues in vegetable oils by LC-MS/MS Aliments dorigine vgtale - Multimthode de dtermination de la teneur en rsidus de pesticides dans le
12、s huiles vgtales par CL-SM/SM Pflanzliche Lebensmittel - Multiverfahren zur Bestimmung von Pestizidrckstnden in pflanzlichen len mit LC-MS/MS This Technical Specification (CEN/TS) was approved by CEN on 11 May 2017 for provisional application. The period of validity of this CEN/TS is limited initial
13、ly to three years. After two years the members of CEN will be requested to submit their comments, particularly on the question whether the CEN/TS can be converted into a European Standard. CEN members are required to announce the existence of this CEN/TS in the same way as for an EN and to make the
14、CEN/TS available promptly at national level in an appropriate form. It is permissible to keep conflicting national standards in force (in parallel to the CEN/TS) until the final decision about the possible conversion of the CEN/TS into an EN is reached. CEN members are the national standards bodies
15、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, Sloveni
16、a, Spain, Sweden, Switzerland, Turkey and United Kingdom. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN DE NORMALISATION EUROPISCHES KOMITEE FR 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 wo
17、rldwide for CEN national Members. Ref. No. CEN/TS 17062:2017 EBS CEN/TS 17062:2017CEN/TS 17062:2017 (E) 2 Contents Page European foreword . 3 1 Scope 4 2 Normative references 4 3 Principle . 4 4 Reagents . 4 5 Apparatus . 7 6 Procedure. 7 7 Evaluation of results 9 8 Precision 10 9 Test report 11 Ann
18、ex A (informative) Examples of experimental conditions 12 A.1 HPLC-System 1. 12 A.2 HPLC-System 2. 12 A.3 HPLC-System 3. 13 A.4 HPLC system 4 . 13 Annex B (informative) Precision data and recovery 15 Annex C (informative) Scheme of procedure (for 2 g of sample) . 24 Annex D (informative) Complementa
19、ry information - Recovery studies . 25 Bibliography . 26 BS CEN/TS 17062:2017CEN/TS 17062:2017 (E) 3 European foreword This document (CEN/TS 17062:2017) has been prepared by Technical Committee CEN/TC 275 “Food analysis - Horizontal methods”, the secretariat of which is held by DIN. Attention is dra
20、wn 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. WARNING The application of this Technical Specification may involve hazardous materials, operations and equipment. Th
21、is Technical Specification does not claim to address all the safety problems associated with its use. It is the responsibility of the user of this Technical Specification to establish appropriate safety and health practices and to determine the applicability of regulatory limitations prior to use. A
22、ccording to the CEN-CENELEC Internal Regulations, the national standards organisations of the following countries are bound to announce this Technical Specification: Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, France,
23、 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. BS CEN/TS 17062:2017CEN/TS 17062:2017 (E) 4 1 Scope This Technical Specific
24、ation describes a method for the analysis of pesticide residues in plant oils (fat content 90 %, water content 60 l, 18 C. 5.9 LC-MS/MS system, equipped with electrospray ionization (ESI) interface (see Annex A). 6 Procedure 6.1 Extraction Transfer a representative test portion of 2 g (msample) of t
25、he homogenous sample into a 50 ml centrifuge tube (5.2). Add 10 ml of acetonitrile (4.2 Vex). Close the tube and shake vigorously for 1 min. Centrifuge for 5 min with at least 1 000 g for better separation of the phases. BS CEN/TS 17062:2017CEN/TS 17062:2017 (E) 8 Transfer an aliquot of the acetonit
26、rile phase VAliquot(e.g. 6 ml extract) into a tube with screw cap (5.3). Add a defined volume (VISTD) of the ISTD solution (4.9). The volume corresponds to 1 % of the aliquot volume (e.g. 60 l ISTD solution to 6 ml acetonitrile phase). 6.2 Clean-up 6.2.1 General The two different clean-up methods de
27、scribed in 6.2.2 and 6.2.3 were successfully validated and may be used alternatively. 6.2.2 Clean-up with amino-sorbent and silica-based reversed phase sorbent Transfer an aliquot of 4 ml of the acetonitrile phase (6.1) into a Polypropylene single use tube (5.3) already containing 100 mg of PSA (4.7
28、) and 100 mg of C18 sorbent (4.8). Close the tube, shake vigorously for 30 s and centrifuge (5 min at 1 000 g). Immediately isolate and acidify the clear extract as described in 6.2.4. In case residues with acetic groups (e.g. phenoxy carboxylic acids) shall be determined, a second aliquot of the ce
29、ntrifuged extract from 6.1 is filled into an injection vial and analysed directly with LC-MS(/MS) to avoid losses of acidic groups by PSA clean-up. 25 mg PSA and 25 mg C18 sorbent are needed per ml of extract. 6.2.3 Freezing-out of co-extracted fat and clean-up with amino-sorbent Store an aliquot of
30、 the extract from 6.1 containing the internal standard for at least 1,5 h at - 18 C to freeze out most of the fat in the extract. For separation of the latter filter the extract over cotton wool (4.20). Take 4 ml from the cold and fat separated solution for dispersive SPE. Transfer an aliquot of 4 m
31、l of the acetonitrile phase into a Polypropylene single use tube (5.3) already containing 100 mg of PSA (4.7). Close the tube, shake vigorously for 30 s and centrifuge (5 min at 1 000 g). Immediately isolate and acidify the clear extract as described in 6.2.4. If residues with acetic groups shall be
32、 determined, transfer a second aliquot into an injection vial and analyse directly with LC-MS(/MS) to avoid losses of acidic groups with PSA clean-up. NOTE It is helpful to load the centrifuge tubes with the dispersive SPE sorbents before beginning the extraction procedure needed for one batch of sa
33、mples. 25 mg PSA sorbent are needed per ml of extract. 6.2.4 Extract stabilization Transfer an aliquot of 3 ml of the cleaned-up extract from 6.2.2 or 6.2.3 into a screw cap storage vial (5.3), taking care to avoid sorbent particles of being carried over, and slightly acidify by adding 30 l of a 5 %
34、 formic acid solution in acetonitrile (4.6). Transfer the pH-adjusted extract into auto-sampler vials and use it for liquid chromatographic analysis. Store the residual extract in a refrigerator to be used if necessary. For 1 ml extract 10 l of the formic acid solution (4.6) are necessary. 6.3 Deter
35、mination by liquid chromatography with tandem mass spectrometry (LC-MS/MS) Inject the sample extracts derived from 6.2.2 to 6.2.4 and standard solutions (4.13) into the LC instruments in an appropriate sequence. This may involve bracketing of the sample extracts with the calibration solutions. The m
36、easurement may be performed using various instruments, instrument parameters and columns. Some instrument parameters and columns are listed in Annex A. These conditions have been shown to provide satisfactory results, but are provided as examples, only. BS CEN/TS 17062:2017CEN/TS 17062:2017 (E) 9 Fo
37、r some gradient/column combinations it is necessary to mix the extract with water or the aqueous mobile phase to achieve a sufficient separation of the analytes. NOTE If extracts are diluted with water or aqueous mobile phases it is important to avoid that non-polar parts of the extract precipitate
38、or emulsions occur. This could lead to losses of lipophilic analytes. In this case an injection applying an injector programme can be helpful (see A.4). The chromatographic conditions as outlined in Annex A have been shown to be satisfactory. Suitable experimental conditions of LC-MS/MS measurements
39、 are outlined in CEN/TR 15641 3. Nevertheless, individual tuning of the compounds on the instrument that is used for measurement usually provides better sensitivities. 7 Evaluation of results 7.1 Identification and quantification For the identification of residues in the final extract, use relative
40、retention time ratio against the ISTD (Rt(A)/Rt(ISTD) obtained from the same run. Check positive results by comparing the intensity ratios between the SIM masses (m/z) or SRM transitions of the analyte. The expected intensity ratios can be determined with the standard solutions. If the ratios of the
41、 samples and the standards have a variation of more than 20 %, the rules of EU Quality Control Procedures have to be followed 2. According to these procedures positive results shall be ensured by using additional measures, e.g. additional SIM masses or SRM transitions or other chromatographic condit
42、ions (column, eluents). For calibration and for checking the linearity of detection of each substance, plot the peak area ratio or peak height ratio of pesticide and internal standard calAy / calISTDy (if an internal standard is used) versus the concentration ratio of the analyte against the ISTD (c
43、alA /calISTD ) in the standard solution (4.13). If no internal standards are used, plot the peak areas or peak height calAy against the concentration of the analytecalA . The calibration area shall be adapted to the residue concentration and should not exceed a decimal power. Possibly more calibrati
44、on graphs shall be established using the standard solution. The calibration function is selected according to CEN/TS 17061:2017, 4.2.1. For a first estimation of the residue level or for the verification of absence of residues, solvent based standards (4.13.1) can be used. They can also be used for
45、quantification, if it was shown that no enhancement or suppression of the analyte signal through matrix occurs. If relevant residue levels are observed (e.g. with possible MRL violation) matrix matched standard shall be preferred for exact quantification. 7.2 Calculation of residue concentrations us
46、ing the internal standard The determination of the concentration of the analyte A in the final extract is performed by using the measured peak area ratio or peak height ratio from pesticide and internal standard yA/yISTD in the sample as described in CEN/TS 17061. The mass fraction w of the analyte
47、in the sample, in milligram per kilogram, is calculated with Formula (1): =exASampleVwm(1) BS CEN/TS 17062:2017CEN/TS 17062:2017 (E) 10 where Ais the mass concentration of the analyte in the final extract, in microgram per millilitre; Vexis the volume of acetonitrile used in 6.1, in millilitre; mSam
48、pleis the mass of test portion in 6.1, in gram. 7.3 Calculation of residue concentrations without internal standards Determine the concentration of the analyte A in the final extract by using the measured peak area or peak height from pesticide yAin the sample as described in of CEN/TS 17061:2017, 4
49、.4.2 to 4.4.5. Calculate the mass fraction w of the analyte in the sample by using Formula (1). 7.4 Calculation of residue concentration using the standard additions approach In case of suspected violative residues, or for compounds which are known to be strongly affected by matrix-induced enhancement or suppression phenomena, standard additions are recommended provided that the function between response and concentrations at the concentration range in question is linear. In case of the standard addition to the final extract,
