1、 STD-BSI BS EN 1528-3-ENGL 1997 W 1b2qbbS Ob22317 715 BRITISH STANDARD Fatty food - Determination of pesticides and polychlorinated biphenyls (PCBs) Part 3. Clean-up methods BS EN 1628-3 : 1997 The European Standard EN 15283 : 1996 has the status of a British Standard ICs 67.040 NO COPYING WITHOUT B
2、SI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAW BS EN 15283 : 1997 Committees responsible for this British Standard The preparation of this British standard was entrusted to Technid Panel AWM, Food analysis - Horizontal methods, upon which the foilowing bodies were represented Association of Publ
3、ic Analysts Department of We and indm Food and Drink Fededon Institute of Food Science and Technology Ministry of Agriculture Fisheries and Food (Laboratory of the Government Chemist) Royal Society of Chemistry This British Standard, having been prepared under the direction of the Consumer Products
4、and SeMces Sector Board, was pubiished under the authority of the Standards Board and comes into effect on 15 June 1997 8 BSI 1997 Amendments issued since publication The foiiowing BSI references relate ta the work on this standard Committee reference Awl43 Draft for cornent 94606376 DC ISBN O 580 2
5、7381 4 Contents Committees responsible National foreword Page Inside front cover ii Foreword Text of EN 15283 2 3 O BSI 1997 i STD-BSI BS EN 1528-3-ENGL 1777 Lb24bb7 Ub22322 4OT W BS EN 15283 : 1997 National foreword This British Standard has been prepared by Technical Panel AW/-i3 and is the Englis
6、h language version of EN 15283: 1996 Fatty food - Determinution of pesticides and polychlorinated biphenyls (PCBs) Part 3 : Clean-up methods published by the European Committee for Standardization (CEN). EN 15283 was produced as a result of international discussions in which the United Kingdom took
7、an active part Cross-references Publication referred to EN 15281 : 1996 EN 15282 : 1996 EN 15284 : 1996 Corresponding British Standard BS EN 15281 : 1997 Fatty food - Determination of pesticides and polychlorinated phenyls Part1:Genm.d BS EN 15282 : 1997 Fatty food - Determinution of pesticides and
8、polychLorinated biphenyls Part 2 : Extraction of fat, pesticides and PCBs, and determination of fat content BS EN 15284 : 1997 Fatty food - (K.) CPCBS) Determination of pesticides and polychlornuted biphenyls (PCBs) Part 4: Determination, confimnatol-E/ tests, rn.6scellaneoUs Compliance with a Briti
9、sh Standard does not of itself confer immunity from legal obligations. Summary of pages This document comprises a front cover, an inside front cover, pages i and ii, the EN title page, pages 2 to 16, an inside back cover and a back cover. 11 O BSI 1997 STD*BSI BS EN L528-3-ENGL L777 m Lb24bb9 b22323
10、 34b m EUROPEAN sXmDm EN 152M NORME EUROPENNE EUROP- NOFM November 1996 ICs 67.040 Descriptors: Food products, edible fats, chemical analysis, determination of content, pesticides, polychiorobiphenyl, purim, chromatography English version Fatty food - Determinaiion of pesticides and polychlorinated
11、biphenyls (PCBS) - Part 3 : Clean-up methods Aliments gras - Dosage des pesticides et des polydorobiphnyles (FCB) - Partie 3 : Mthodes de punncation Fettreiche Lebensmial - Bestimmung von den und polychlorierten Biphenylen (PCB) - ?leil 3 : Rehigungwerfahren This European Standard was approved by CE
12、N on 1W10-27. CEN members are bound to comply with the CENKENELEC Internal Regulations which stipulak the conditions for giving this European Sandard the status of a national standard without any aiteration. Up-to-date lists and bibliographicai references concerning such national sandards may be obt
13、ained on application to the Centrai Secretariat or to any CEN member. This European Standard exists in three officiai versions (English, French, German). A version in any other language made by translation under the responsibility of a CEN member into its own language and notified to the Central Sec
14、retariat has the same status as the official versions. CEN members are the national standards bodies of Austria, Belgium, Denmark, Fniand, France, Germany, Greece, Iceland, Ireland, My, Luxembourg, Netherlands, Norway, Portugal, Spain, Sweden, Switzerland and United Kingdom. CEN European Committee f
15、or Standardization Comit Europen de Normalisation Europisches Komitee fiir Normung Central Secretariat: rue de Stassart 36, B-1050 Brussels O 1996 Copyright reserved to CEN members Ref. No. EN 15283 : 1996 E Foreword This European Standard has been prepared by Technical Commit peroxide free. Distil
16、and stabilize with 2,O % of its volume with absolute ethanol. 6.3.6 Eluting mixture A, light petroleum (6.3.2) and diethyl ether (6.3.4) 94 : 6 (VN. 6.3.6 Eluting mixture B, light petroleum (6.3.2) and diethyl ether (6.3.4) 85 : 15 (Vw. 6.3.7 Eluting mixture C, light petroleum (6.3.2) and diethyl et
17、her (6.3.4) 50 : 50 (Vm. 6.3.8 Sodium sulfate, granular, anhydrous. Heat at 500 “C or 550 “C for at least 4 h, ailow to cool and store in a stoppered bottle. 6.3.9 Rmis, 150 pn to 250 pn (60 mesh to 100 mesh). Activate by heating at 650 “C for 4 h, immediately transfer into a bghy stoppered containe
18、r and store in the dark. Before use, heat at 130 “C for at least 5 h and allow to cool in a desiccator. lbt each batch of Florisil* from time to time as follows. Pass 1 ml of a standard niheme solution containing 0,l mg4 each of lindane, heptachlor, aldrin, heptachlor epoxide and dieldrin and 0,3 mg
19、A of endrin through the adsorption column. Elute and concentrate as described in 6.6.3. Determine the recovery by gas chromatography The Florisil is satisfactory if lindane, heptachlor, aldrin and heptachlor epoxide are eluted quantitatively in the eluting mixture A (6.3.6) and dieldrin and endrin a
20、re eluted quantitatively in the eluting mixhire B (6.3.6). 6.3.10 Sodium chloride solution, saturated Before preparing the solution, heat the sodium chloride at 500 “C for at least 4 h. 6.4 Apparatus Usual laboratory apparatus and, in pdcuiar, the following. 6.4.1 Separating funnels, capacity 125 ml
21、 and lo00 ml, with ground stoppers and PTFE stopcocks. 6.4.2 Cbomaographic tube, 25 mm outer diameter (o.d), 50 mm long, with PTFE stopcock and a sintered glass disk or a glass wool plug. 6.4.3 chromatographic tube, 22 mm internai diameter (i.d), 300 mm long, with FTFE stopcock and a sintered giass
22、disk or a glass wool plug. I 1,2,3,4,10, lO-Hexachioro-l,4,4, 5,8,8-hexahydro-l,4-endo-5,Sendodimethanonaphthaiene. O BSI 1997 STD.BSI BS EN 1528-3-ENGL 1997 lb24bb9 0b22327 T9L U Page 5 EN 1628-3 : 1996 6.4.4 Kuderm-Danhh mapomtor, capacity 500 ml, with attached graduated collection tube, or equiva
23、lent. 6.4.6 2-baU micro-Snyer column, or mm-Vigreux column. 6.6 Procedure 6.6.1 Extraction of fat, pesticides und PCBs General methods shall be carried out in accordance with EN 15282 : 1996. 6.6.2 Acetonitrile/l column packing 60 g BioBeads SX3 resin4), pre-swelled overnight in the eluting mixture,
24、 approximately 48 cm bed length. 10.4.2 Roiuqi mapomtor, with evaporation flasks of capacity 500 ml and a water bath capable of being controlled between 20 “C and 50 “C. 10.6 Procedure 10.6.1 Extraction of fat and pesticides 10.6.1.1 Genemc General methods shall be carried out in accordance with EN
25、15282 : 1996. 10.6.1.2 Specu method Place an approximately 40 g fat sample in a giass funnel (8 cm diameter) containing a glass wool plug. Place the funnel in a 250 ml beaker on a hot plate at 110 “C maximum until the fat ceases to drip. Mix thoroughly 10.6.2 Clean-up bu GPC When using a new GPC col
26、umn, must the flow rate of the eluting mixture to 5 ml/mh and check the calibration for quantitative recovery with 2,O g of com oil fortified with relevant compounds. Determine the appropriate dump and collect times for the desired residues. Weigh about 2 g, to the nearest 10 mg, of liquid fat sampl
27、e into a 10 mi volumetric flask, dilute to the mark with eluting mixture and mix thoroughly Centrifuge or filter if particulate matter is visible. Load the 5 mi sample loop using approximately 7 mi of the solution. Elute the GPC column with the eluting mixture, using the dump/collect times determine
28、d beforehand, at a flow rate of 5 ml/min. Collect the eluate in a 250 ml round-bottomed flask and robqy-evaporate just to dryness at a 30 “C maximum bath temperature. lIansfer the residue remaining after evaporation with small aiquot portions of ismtane to a volumetric flask or a graduated test tube
29、, and adjust the solution to a suitable volume, e.g. 5 ml, with isooctane. 10.6 Determination Determination shali be carried out in accordance with clause 7 of EN 1528-1 : 1996 and clause 4 of EN 15284 : 1996. 10.7 Evaluation of results The results shall be evaluated in accordance with clauses 9 to
30、11 of EN 15281 : 1996. 10.8 Test report The resuits of the tests shall be reported in accordance with ciause 12 of EN 15281 : 1996. 3, GPC Autoprep 1001 or 1002 are the trade names of a product supplied by Analyticai Bio Chemistry Laboratories, Inc., Columbia, Mo., USA. This information is given for
31、 the convenience of users of this European Standard and does not constitute an endorsement by CEN of the product named. Equivalent products may be used if they can be shown to lead to the same results. 4, BioBeads S-X3 resin is the Wadename of a product supplied by Bio-Rad Laboratories Richmond, Ca,
32、 USA. This information is given for the convenience of users of this European Standard and does not constitute an endorsement by CEN of the product named. Equivalent products may be used if they can be shown to lead to the same results. O BSI 1997 STD-BSI BS EN L528-3-ENGL 1777 H lb24bb9 Ob22334 121
33、 Page 12 EN 1528-3 : 1996 11 Method G. Gel permeation chromatography (GPC) and column chromatography on partiauly deactivated silica gel (Specht) 7 11.1 Applicability This method has been shown to be applicable for the determination of 24 organochlorine pesticides and metabolites, the PCB indicator
34、congeneB and 13 organophosphorus pesticides, as given in annexA of EN 15281 : 1996. GPC is a very effective technique for separating the residues to be analysed from the surplus of lipids. It covers a particular broad range of compounds. For this reason, the GPC eluate will also contain other pestic
35、ides and contaminants than those mentioned above, e.g. the pyrethroids and chlorinated benzenes or phenols. Moreover, GPC cm be readily automated A silica gel mini-column may be used to provide an additionai clean-up and a fractionation of the residues according to their polarity, thus yieldmg extra
36、 infomation for identincation. 11.2 Principle Extraction of the residues, together with the fat, nom the sample by one of the procedures described in EN 15282 : 1996. Concentration of the extract almost to dryness, redissolving it in cyclohexandethyl acetate. Chromatography using a gel permeation co
37、lumn, with cyclohexandethyl acetate as the eluting solvent. Concentration of the eluate aimost to dryness, and redissolving it in n-hexane. Chromatography on a small column of partially deactivated silica gel, using eluants of increasing polarity. Concentration of the eluates for examinati * -onbyGC
38、. 11.3 Reagents and materials All reagents and materials used shail be suitable for the analysis of residues of pesticides and PCBs and shail be in accordance with clause 4 of EN 1528-1 : 1996. if purScalion is necessary, the procedures given in annex A are appropriate. 11.3.1 Cydokane. 11.3.2 Ethg1
39、 acetate. 11.3.3 GPC eluting mixture, cyclohexane (11.3.1) and ethyl acetate (11.3.2) 1 : 1 (VIV). 11.3.4 Acetone. ll.3.6 ISO-OCW. 11.3.6 n-ne. 11.3.11 Sodium sulfate, granular, anhydrous, heated at 550 “C for at least 2 h. 11.3.12 Silica gel, deactivated with 1,5 % water. Heat silica gel 60 (63 pn
40、to 200 pm 2 70 mesh to 230 mesh), at 130 “C for at least 5 h, allow to cool in a desiccator, and store in a tightly stoppered container in the desiccator. To 98,5 g dried silica gel in a 300 ml Erlenmeyer flask with a ground joint, add 1,5 d water dropwise from a burette, with continuous swirling. I
41、mmediately stopper the flask with a ground stopper and shake vigorously for 5 min until all lumps have disappeared. Next shake for 2 h on a mechanical shaker, and then store in a tightly stoppered container. 11.4 Apparatus Usual laboratory apparatus and, in particulaq the following. 11.4.1 Automated
42、 inshment for GPC, e.g. GPC Autoprep 1001 or 1002, equipped with chromatographic column, 25 mm i.d, 40 cm long, and 23 5 ml sample loops; column packing approximately 50 g BioBead SX3 resin, (38 km up to 75 pn 2 200 mesh to 400 mesh), preswelled overnight in the GPC eluting mixture, approximately 32
43、 cm bed length. 11.4.2 Rotmy mupomw, with evaporation flasks of capacity 500 ml and a water bath capable of being controlled between 20 “C and 50 “C. 11.4.3 Chmtogmphic tube, 7 mm i.d, 23 cm long, with extended outlet. 11.5 Procedure 11.6.1 Extraction of fat, pesticides and PCBs General methods shai
44、l be carried out in accordance with EN 15282 : 1996. 11.6.2 Gel permeation chromatography When using a new GPC column, aust the flow rate of the eluting mixture to 5 dmin and check the calibration with relevant compounds. Determine the appropriate dump and collect times for the desired residues. Dis
45、solve up to 5 g of the fat or oil or of the extracted fat portion in the GPC eluting mixture (11.3.3) in a volumetric flask Ddute to the mark with the same mixture and mix thoroughly Load the 5 ml sample loop using approximately 7 ml of the solution. Elute the GPC column with the GPC eluting mixture
46、, using the dumpkollect times determined beforehand, at a flow rate of 5 mVmin. Collect the eluate in a 250 ml round-bottomed flask and 11.3.7 lbluene. rotary-evaporate it to approximately 1 ml (rotate slowly, and immerse flask only slightly), transfer the concentrate to a ground-stoppered graduaed
47、test tube, rinse with ethyl acetate, and dilute to a suitable 11.3.8 Eluant I, n-hexane (11.3.6) and toluene (11.3.7) 65 : 35 (v,. 11.3.9 Eluant 2, toluene 11.3.n. volume, e.g. 5,O ml, with ethyl acetate. 11.3.10 Eluant 3, toluene (11.3.7) and acetone (11.3.4) 95 : 5 (VIV). NOTE. For the anaipis of
48、organophosphorus pesticides, the GPC eluate can be directly injected for gas chromatographic examination, if a phosphorus selective detector is used. O BSI 1997 STD-BSI BS EN 1528-3-ENGL 1997 W lb2Libb9 b22335 Ob8 Page 13 EN 1528-3 : 1996 11.6.3 Silica gel mini column chromatography Pipette 2,5 ml o
49、f the solution derived from 11.6.2 into a round-bottomed flask with a ground joint, and add 5 mi of iso-octane (11.3.6). Carefully evaporate to 1 ml (on no account to dryness) in a rotary evaporator (rotate slowly and immerse flask only slightly). If the solution still smells of ethyl acetate, again add iso-octane and repeat the evaporation. Pack the chromatographic tube (11.4.3) in the foiowing order: glas wool plug, 1,0 g of deadvahl silica gel (11.3.12) 5 mm to 10 mm layer of sodium sulfate (11.3.11), glass wool plug. Before us
