SIST EN 14185-1:2003
(Main)Non-fatty food - Determination of N-methylcarbamate residues - Part 1: HPLC-method with SPE clean-up
Non-fatty food - Determination of N-methylcarbamate residues - Part 1: HPLC-method with SPE clean-up
This En describes a method for the determination of N-methyl-carbamate residues in foodstuffs.
Fettarme Lebensmittel - Bestimmung von N-Methylcarbamat-Rückständen - Teil 1: HPLC-Verfahren mit Reinigung durch Festphasenextraktion
Aliments non gras - Dosages des N-méthylcarbamates - Partie 1: Méthode CLHP avec purification par SPE
Le présent projet de norme européenne spécifie une méthode par chromatographie liquide haute performance (CLHP) pour la détermination des produits phytosanitaires de la famille des N-méthylcarbamates présents dans les céréales, les fruits et les végétaux.
La méthode a été validée par une étude inter-laboratoires du carbaryl, carbofuran, méthiocarbe, méthomyl, oxamyl et des composés parents du propoxur ainsi que du méthiocarbe sulphoxyde présent dans les poivrons verts et dans les pommes a des niveaux se situant entre 0,08 et 0,9 mg/kg.
On ne dispose d'aucunes données inter-laboratoires concernant l'efficacité de la méthode pour la détermination d'autres métabolites importants ; on sait toutefois que la méthode ne convient pas pour les oximes de l'oxamyl et du méthomyl.
Hrana, ki ne vsebuje maščobe - Določevanje ostankov N-metil karbamata - 1. del: Metoda HPLC s čiščenjem SPE
General Information
Standards Content (Sample)
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.Fettarme Lebensmittel - Bestimmung von N-Methylcarbamat-Rückständen - Teil 1: HPLC-Verfahren mit Reinigung durch FestphasenextraktionAliments non gras - Dosages des N-méthylcarbamates - Partie 1: Méthode CLHP avec purification par SPENon-fatty food - Determination of N-methylcarbamate residues - Part 1: HPLC-method with SPE clean-up67.050Splošne preskusne in analizne metode za živilske proizvodeGeneral methods of tests and analysis for food productsICS:Ta slovenski standard je istoveten z:EN 14185-1:2003SIST EN 14185-1:2003en01-november-2003SIST EN 14185-1:2003SLOVENSKI
STANDARD
SIST EN 14185-1:2003
EUROPEAN STANDARDNORME EUROPÉENNEEUROPÄISCHE NORMEN 14185-1April 2003ICS 67.060; 67.080.01English versionNon-fatty food - Determination of N-methylcarbamate residues -Part 1: HPLC-method with SPE clean-upAliments non gras - Dosages des résidus de N-méthylcarbamates - Partie 1: Méthode par CLHP avecpurification SPEFettarme Lebensmittel - Bestimmung von N-Methylcarbamat-Rückständen - Teil 1: HPLC-Verfahren mitReinigung durch FestphasenextraktionThis European Standard was approved by CEN on 20 February 2003.CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this EuropeanStandard the status of a national standard without any alteration. Up-to-date lists and bibliographical references concerning such nationalstandards may be obtained on application to the Management Centre or to any CEN member.This European Standard exists in three official versions (English, French, German). A version in any other language made by translationunder the responsibility of a CEN member into its own language and notified to the Management Centre has the same status as the officialversions.CEN members are the national standards bodies of Austria, Belgium, Czech Republic, Denmark, Finland, France, Germany, Greece,Hungary, Iceland, Ireland, Italy, Luxembourg, Malta, Netherlands, Norway, Portugal, Slovakia, Spain, Sweden, Switzerland and UnitedKingdom.EUROPEAN COMMITTEE FOR STANDARDIZATIONCOMITÉ EUROPÉEN DE NORMALISATIONEUROPÄISCHES KOMITEE FÜR NORMUNGManagement Centre: rue de Stassart, 36
B-1050 Brussels© 2003 CENAll rights of exploitation in any form and by any means reservedworldwide for CEN national Members.Ref. No. EN 14185-1:2003 ESIST EN 14185-1:2003
EN 14185-1:2003 (E)2ContentspageForeword.31Scope.32Normative references.33Principle.34Reagents.45Apparatus.56Sampling.67Preparation of the samples.68Procedure.79Calculation of the results.810Confirmatory tests.811Precision, Repeatability and Reproducibility.812Test report.9Annex A (informative)
Precision Data.11Annex B (informative)
Alternative HPLC operating conditions.15Bibliography.16SIST EN 14185-1:2003
EN 14185-1:2003 (E)3ForewordThis document (EN 14185-1:2003) has been prepared by Technical Committee CEN/TC 275 "Food analysis -Horizontal methods", 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 identical text orby endorsement, at the latest by October 2003, and conflicting national standards shall be withdrawn at the latestby October 2003.Annexes A and B are informative.According to the CEN/CENELEC Internal Regulations, the national standards organizations of the followingcountries are bound to implement this European Standard: Austria, Belgium, Czech Republic, Denmark, Finland,France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Luxembourg, Malta, Netherlands, Norway, Portugal,Slovakia, Spain, Sweden, Switzerland and the United Kingdom.1 ScopeThis European Standard specifies a high performance liquid chromatographic (HPLC) method for the determinationof N-methylcarbamate pesticides in cereals, fruits and vegetables.The method has been validated by collaborative study for carbaryl, carbofuran, methiocarb, methomyl, oxamyl andpropoxur parent compounds and for methiocarb sulfoxide in green peppers and apples at levels between0,08 mg/kg and 0,9 mg/kg.No collaborative data are available for the performance of the method in the determination of other significantmetabolites although it is known that the method will not work for oxamyl and methomyl oximes.2 Normative referencesThis European Standard incorporates, by dated or undated reference, provisions from other publications. Thesenormative references are cited at the appropriate places in the text, and the publications are listed hereafter. Fordated references, subsequent amendments to or revisions of any of these publications apply to this EuropeanStandard only when incorporated in it by amendment or revision. For undated references the latest edition of thepublication referred to applies (including amendments).EN ISO 3696, Water for analytical laboratory use — Specification and test methods (ISO 3696:1987).3 PrincipleThe sample is homogenized with acetone, dichloromethane and light petroleum and the homogenate is centrifugedto yield two layers of the supernatant. An aliquot portion of the upper layer is evaporated to dryness. Optionally, thisextract may be cleaned up by solid phase extraction (SPE) using a cartridge packed with aminopropyl-bondedsilica. In the extract solution, the N-methylcarbamates are determined by reversed-phase high performance liquidchromatography (HPLC) with post-column hydrolysis. The methylamine formed is allowed to react with o-phthaldialdehyde and 2-mercaptoethanol and the derivatives are detected with a fluorescence detector. For furtherinformation on this method, see [1] to [4].SIST EN 14185-1:2003
EN 14185-1:2003 (E)44 Reagents4.1 GeneralUnless otherwise specified, use reagents of recognized analytical grade, preferably for HPLC and pesticide residueanalysis, and distilled water for cleaning of glassware or water of at least grade 1 as defined in EN ISO 3696.Label all standard containers with name and purity of the pesticides. For the full chemical names and structures,see ISO 1750.4.2 Safety aspects associated with reagentsWARNING — Many pesticides are toxic by various routes of exposure, especially in concentrated form.When working with these pesticides consult safety data sheets of the manufacturer for information.Vapours from some volatile solvents are toxic. Several of these solvents are readily absorbed through skin. Use aneffective fume hood to remove vapours of these solvents as they are set free.4.3Acetone4.4Acetic acid4.5Dichloromethane4.6Light petroleum, boiling range 40 °C to 60 °C4.7Methanol4.8Acetonitrile4.9Solvent mixtureAcetonitrile (4.8) / acetic acid (4.4) 99,9 + 0,1 (V/V)4.10Water, purified by a LC-grade water purification system4.11Sodium acetate4.12o-Phthaldialdehyde4.13Sodium tetraborate, anhydrous4.142-Mercaptoethanol4.15Trimethacarb4.16Mobile phase AAcetonitrile (4.8) / water (4.10) 20 + 80 (V/V), containing 2,5 mmol/l sodium acetate (4.11). Prior to use, filter themobile phase A with gentle suction through a membrane filter (5.10).4.17Mobile phase BMethanol (4.7) / water (4.10) 20 + 80 (V/V), containing 2,5 mmol/l sodium acetate (4.11). Prior to use, filter themobile phase B with gentle suction through a membrane filter (5.10).SIST EN 14185-1:2003
EN 14185-1:2003 (E)54.18Mobile phase CAcetonitrile (4.8) / water (4.10) 60 + 40 (V/V), containing 2,5 mmol/l sodium acetate (4.11). Prior to use, filter themobile phase C with gentle suction through a membrane filter (5.10).4.19OPA reagentIn a 1 000 ml volumetric flask, dissolve 3,8 g of sodium tetraborate (4.13) in approximately 900 ml of water (4.10).Add a solution of 250 mg of o-phthaldialdehyde (4.12) in 10 ml of acetonitrile (4.8). Next, add 0,1 ml of2-mercaptoethanol (4.14) and dilute to 1 000 ml with water.The solution is stable for approximately one week.4.20SPE eluting mixture (optional)Dichloromethane (4.5) / methanol (4.7) 99 + 1 (V/V).4.21Internal standard solution, r = 0,05 µg/ml.Dissolve 10 mg of trimethacarb (4.15) in 10 ml of acetonitrile (4.8) and dilute 100 µl of this solution to 100 ml in avolumetric flask with solvent mixture (4.9) to give dilution A (1 µg/ml). In a second volumetric flask, dilute 5 ml ofdilution A to 100 ml with acetonitrile (4.8) / water (4.10) 20 + 80 (V/V).4.22Standard materialsN-methylcarbamate pesticides such as aldicarb, bendiocarb, bufencarb, butocarboxim, carbanolate, carbaryl,carbofuran, cloethocarb, dioxacarb, dithiocarb, ethidimuron, ethiofencarb, fenobucarb, isoprocarb, methiocarb,methomyl, oxamyl, promecarb, propoxur, thiofanox, the metabolite 3-hydroxy-carbofuran and the sulfoxide andsulfone metabolites of aldicarb, butocarboxim, ethiofencarb, methiocarb and thiofanox.4.23Pesticide stock solutions, r = 0,05 µg/ml.Dissolve 10 mg of a standard material (4.22) in 10 ml of acetonitrile (4.8) and dilute 100 µl of this solution to 100 mlin a volumetric flask with solvent mixture (4.9) to give dilution A (1 µg/ml). In a second volumetric flask, dilute 5 mlof dilution A to 100 ml with acetonitrile (4.8) / water (4.10) 20 + 80 (V/V).4.24Pesticide standard solutionsPrepare appropriate standard solutions by diluting suitable amounts of pesticide stock solutions (4.23) with internalstandard solution (4.21) / water (4.10) 20 + 80 (V/V).4.25Keeper solutionMix 2 g of ethylene glycol with 8 ml of acetone (4.3).5 Apparatus5.1 GeneralGlassware shall be thoroughly cleaned. Hot detergent solution may be used for cleaning, but afterwards theglassware shall be well rinsed with distilled water and acetone before drying.Usual laboratory apparatus is used and, in particular, the following.5.2Chopper5.3Homogenizer or high speed blenderSIST EN 14185-1:2003
EN 14185-1:2003 (E)65.4Centrifuge, provided with polytetrafluoroethylene tubes of capacity 200 ml, and capable of producing arotational speed of at least 4 000 min–1.5.5Water bath, capable of being maintained at 50 °C or 60 °C5.6SPE cartridges, packed with 100 mg aminopropyl-bonded silica, particle size 40 µm (e.g. Bond-ElutÒ1))(optional)5.7Device for eluting SPE cartridges (5.6) with suction (optional)NOTEApparatus for automated SPE elution is commercially available [4].5.8High performance liquid chromatograph, equipped with5.8.1Ternary pumping system with six-port injection valve with a 100 µl sample loop, a post-column system(consisting of a column reactor, a low-dead volume T-piece and a pulse-free reagent pump), a fluorescencedetector and a quantification unit with an integrating system.5.8.2HPLC guard column, stainless steel cartridge, 10 mm long, 4,0 mm inner diameter (e.g. LiChroCARTÒ1),packed with Superspher®1)) 60 RP-8 (particle size 4 µm).5.8.3HPLC analytical column, stainless steel cartridge, 250 mm long, 4,0 mm inner diameter (e.g.LiChroCART®1)), packed with SuperspherÒ1) 60 RP-8 (particle size 4 µm).5.8.4Post-column hydrolysis column, stainless steel, 50 mm long, 4,0 mm inner diameter, packed withAminex®1) A 27 (15 µm).5.9Ultrasonic bath5.10Membrane filters, pore size 0,45 µm6 SamplingPrepare the laboratory sample according to a generally recommended method of sampling to achieve arepresentative part of the product to be analysed.7 Preparation of the samplesWhere possible, carry out the analysis of samples immediately on their arrival in the laboratory. Do not analyse alaboratory sample which is wholly or extensively spoiled.For analysis take only the portion of the laboratory sample to which the maximum residue limit applies. No furtherplant-parts may be removed. A record of the plant-parts which have been removed shall be kept. The sample thusprepared is the test sample.If the sample cannot be analysed immediately, store it at 0 °C to 5 °C for no longer than 3 days before analysis.
1)Bond-Elut® is a trade name of a product supplied by Analytichem International, Harbor City, CA, USA. LiChroCARTÒ andSuperspherÒ are trade names of products supplied by Merck, Darmstadt, Germany. Aminex® is a trade name of a productsupplied by Bio-Rad, Hercules, CA, USA. These informations are given for the convenience of users of this European Standardand do not constitute an endorsement by CEN of the products named. Equivalent products may be used if they can be shown tolead to the same results.SIST EN 14185-1:2003
EN 14185-1:2003 (E)7The reduction of the test sample shall be carried out in such a way that representative portions are obtained (e.g.by division into four and selection of opposite quadrants). When the samples are in small units (e.g. small fruits,legumes, cereals), the test sample shall be thoroughly mixed before weighing out the test portion. When thesamples are in larger units, take wedge-shaped sections (e.g. large fruits and vegetables) or cross sections (e.g.cucumbers) which include the outer surface from each unit.From each test sample, remove those parts which would interfere with the analytical procedure. In the case ofstone fruits, the stones may be removed. Care shall be taken that as little as possible of the remainder such asjuice or flesh is lost. The basis for the calculation of the residue mass fraction is the mass of the original test sample(with stones).If samples have to be stored for more than 3 days, they shall be deep-frozen at –18 °C or lower. To ensure thateven after thawing representative samples can be taken, prepare portions of the product which are each sufficientfor one analysis.Chop the test sample and weigh out test portions of masses of 15 g to an accuracy of ±1 %.8 Pr
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