本 标 准 规定了出口禽肉中二氯二甲D比咤酚残留量检验的抽徉、制样和气相色谱测定方法。 本 标 准适 用于出口鸡肉中二氯二甲毗Rjz酚残留量的检验。

中华人民共和国进出口商品检验行业标准SN/T 0212.2—93
出口禽肉中二氯二甲吡啶酚
残留量检验方法
甲基化-气相色谱法
Method fFor determination of clopidolresidues ir poultry mieat for export-Meihylation-gas chromatography1993-06-04发布
中华人民共和国国家进出口商品检验局1993-08-01实施
中华人民共和国进出口商品检验行业标准出口离肉中二舞二甲啶
酚残留量检验方法
甲基化-气相色谱法
Method far determtnation uf elopidotresldueg io poultry ment far uxportMfethylaliun-gas chromutogrnphy1主器内容与适用范围
SN/T0212.2--93
本样准规定了品口两内中二氛二甲比酚线留量检验的抽样、剧样和气相色诺测是方法求标准适用于尚口将肉中二氯二甲脱晓份戏留些的检验，2抽样和制样
2.1格验批
以个超过2500箱为--个检验批。同一检验批的商品应具有相同特征，如包装，标记.产地.现格和等等，2.2推择数量
批盐（箱》
26-16G
211~500
331--1 95
131~-2 520
2.3、钟挥法
段低推件效(箱）
老2.主规定的拍迁润激随摘取医诺开店，书箱至少取一袋作为原始样品原始样品总母不少于2封方.标明记.及时送实验室，
2.4试摔制爷
每源的样结中取帮分有少克性码岗降，同欧均机政辞，将全帮级降药样品混均用四分法缩分出试件使不少可的孕案人授消资器内.后，规瞬标记2.5试伴保存
试样于-130下终冻仅存。
注，在袖样和制样过，必能止作品受到污象效发生恶留好含量的空化，3测定方法
3. 1方法提要
中华人民共和国国家进出口商品检验1993-05-叫批准1993-08-01实施
SN/T 0212.2-93
用甲醇提取离肉组坝内药二氧二甲吡噻酸，离心后的上清辣经中性氧化短和离子交唤单化。洗脱液经重氮甲烷试剂中尚化、生威的二二年比啶甲醛（3.5-二流4·甲氧基-2.6-甲提吡啶>用配有电子俘炭检测器的气相色谢仪测定，以2，-基多萃为内标进行定垫3.2试剂和材料
除等殊规定外，试烈均为分析纯，水为燃愧衣双相应前去两干水。3. 2. 1 甲醉,
3.2.2中性载化层研用,100~20M月.300灼烧1h.整于下燥器备用。32.3阴离了交换树脂Dovex1-X9,100~290日.孕.或等效品3.2.4Z甲醇落液10.6折V/WV>.3mL冰乙酸落于4汀mL带醇中，3.2.5正已流；加酸货蒸均
3.2.6内标：2+4-二础基激苯，纯度>99%，3.2.提联用的硼基就苯止已烷蒂做，0.12.4-=硝基需举/mL正也烷落液，3-2.8电惠中烷试剂在125mL的蒸瓶中加人35m，1m，乙能和10ml.氢氧化水溶液（68·10L）。放人磁楼，井固定链力微拌器加热上的水浴中。连接漏斗和冷凝爵井率联丙个125m1.维形瓶，链形瓶中各置有10mL乙.胖。冷凝收案管摘入乙使画，链形接收程胃于述溶中。滴理谢斗内盘有21.5g的N-中基-N-亚销基·B-年未破酸胺落于14rI.乙避的波重鼠试剂）。均水浴至！，进行幕留，蒸确瓶内溶液边用邀力扰，达加恶试划，满笼车部游减的时回控制在21m以.上。当增出液近于无包时，降止蒸瘤。将两个雌形接收中的落液台并，并起7心水路上草点铅。将缩出液-重氢甲烷试剂需封，并十一18心保存。保序期一~日.3.2.9款二中吡啶酚标准品，纯度>%。3.2.91二第二带北噬酸标准忙备液：100g/ml。准确弥取15).0mk=氟一甲吡啶酚标准品用甲醇移人！000ml，容些瓶至总体积约500mL，潜解（约需2～3h）。明甲醉定客到刻度，混寸，3.2.9.2中叫标难降恼（1.10Fgml，移液管魅取上述标准延备减10m于100L容量瓶内，月中辟稀辩至刻度。
3.2-5.3中间标难溶（0）1g/mT，用移减智移取中间标谨液（1）10mL于100tnL容量瓶内，用争醇烯样全对。
3.2.9.4标准L作溶流，0.1g/mL。用移使整将延中间标准游液（1）10ml于10）ml.蓉整瓶内，用中醇粥至度。
，3.3按器和放备
3.3.1搅肉机：电。
3.3.2均透机，3900/min
3.3.3两心机：日立2PR52;4r/min或相当的投密3.3.4中性氢化据控：移取24性氧化（3..2）填人100mm×10mm内径载璃层注内。3.3.5附离子定换层折生，将Dowex1-Xs两高子交换谢翡伴以去离于水移入220mm×10mm（内定1层断内，沉降后栏床离为1.5cm，以1JomL1m>1/L的VOH水溶被淋洗树脂，继以去离于水海洗至中生再改\，/.的乙数钠水落浓游院，错以去离了水性洗至中性。波速均为？ml.min.注入甲醇求液：3>50mL备用使用前放出甲醇水溶液使夜%略高于树脂高。3.36试管：带旋益前小#气密性的）.100mm×13mm（内径）。3.3.7指潜举配存电产浮获检测器。3.3.8微注验器：10rl.
3.4测究步题
3.4.1提收
称收约20g试拌（情晚至0.1g）于均质瓶内.加火10CmL单整300/mi质4min后，将均a
SN/T0212.2-93
项版移人高心机，4000r/min离心10min。3.4.2净化
将附离于交换柱置于中作化铝快下，用移滤管移20mL提取的上清按于氧化解住中，恢提取液经中比就化铝和调离子交换佳流入烧怀，用10m早醇清洗中性氧化铝耗的棕些，格宏氢化铝柱烧坏，将25mL的比色管胃阳离子交换性的下方，用两份1UmL的0.6%乙酸中静溶减洗脱二靠二甲吡啶酚,洗提液南堪，1mL/min用甲醇定客并混幻。3.4.3中基化
用移液够取尝容的洗被5mL于试些（3.3.6)内.试管置十70心水浴内·通以氮气施欢干。取出试管加人0.2L甲醉水穿滤（82)，改温使观潜落解，冷至室益，加1mL重微甲烷试.打上施盐，将试普肾于70心水浴中保温反应2min。反应时，试管内笔液面和水浴面持平，取出试懂，冷却至室猛，拧去能益，仅使管腐帮按魅水面，宜至芒醛除，于除去乙醛片的试管内加5ML去离子水和1.IL提取用的销基氧差正己统搭(3.2.),加益，在混知器上混和1min，取出试带4000r/in离心3min，所制旋的上层正己境样液低气相色谱测定。3.4.4忌
3.4.4.斤包避系件
，危诺坡期柱.2m×2nm（内径），填充物为10K（m/m)DC-200硅纳净十心hror0a0rhHP载(0:
b.盆气纯度99.99%.60mLmin
c持温170℃，
d，进样口温度：2;
e，检淡器温度，300C
3.4.4.2色满则定
将样液（如有必要，可多加提取用的确茎盖茉正已烧溶，作进一步蒂择注人气相色谐甚、进样量2L，在上述色游条件下二翁二用吡院甲醛出蜂保留时间纳为2min，2.4二确基出降医舒时河为4inm
3.4.5标准断我的给制
分别取二氟二甲吡啶盼标准工作溶液0,5.1.T2,0.4,1.8.1.6ml.下等试管(2.3.6)内，接上述步聚(3.4.3～3.4.4)进行略去定的洗提液5mI.\，以所母各依度二第二甲吡啶甲色诺雌高机内标择高之比.对底文原标准陷凝的浓度（产g/mL)绘制标准曲线，3.4.6同效系数R
称取一份t.S～20.1g(转确至0.1)不含二期二甲吨定酚的试样，分别如人相当于(.0,0.0)和0.05g/g的二氯二中吡啶龄中间标准落波（)（分别为(1.0.C.2和1mt.)。按3.4.1~5.4.4条操准，并计算出毕与回收系数R，
3.4.7空自试验：除不称取试样外，均接上述测定步峰（3.4.1--3.4.4)进行3.4.日结果计郊知表述
按式计算试样中二需二中吡啶谢的合X-
式中，一—鸡肉中二二用唯啶的含量，mggR—平均网率，
（一-由京准曲线查得股终损离中二案二甲吡院酚的含量·/ml，F最终样渡的体，
SN/T 0212. 2—93
m一最终样液所代的样品量+
二氧二甲吡啶酚含些亦可由色谱效据处理机宜搓求得注，计算结单将空白恼批辞，
4测定低限、同收率
4.1测定低限
本方法测定低限为0.mg/kx，
4.2回收率
回收率实整数据：
二氯二甲此啶略添加浓度在0,U05~~11.025mg/kg范内，回收率为65.4%~86.2%。附加说明：
本标准由中华人民共和国国家连出口商品检验品提出。车标准自中华人民共和国上游，河北，北京、出东逆出口商站检验居负贵起草，本标准主要起毕人朱治平，事华、未坚.沈札虽、离修丽、王风地、张军、刘心。参考文献
A0AC-0fficialMethode ofAnriyss,13th ed,41.C13-41.018.1980.Professianal Standard of the People's Repubtic of Chlnafor Import and Export Commadity InspectionMetbod for determination of ciopidolresidues in poultry meat for exportMethylation-gas chromatngraphy1 Scope aod flald of applicatlanSN/T 0212.2—93
Thie atandard specifies the methods of wampling, aample prepareticn ard detemuinetion by gesckramatography of clopidol residues in poulery meat far expurt.This gtandard sx applicable to the deterrninatiun of clnpilol residwe content in hroilera for expDrt.2 Sanpling and samplc prepuration2.1lnspection loe
The guantity ul an insiuuton lut hnull ruas he rmore than 5DD cartongThe characLeristics of the cargo within the same inspuction lot, such as packing, mark origin,specilication, grade etc., should be the same.2.2Quahtity of the sample lakenNunsber ut cartpus in eaeh iaspeetinn lal1—25
26—10C
101-25c
251—00
5011000
1001—2500
2.3 Sarnpling procedure
Mtiainur numaer of cartoneto be takec1
A number of cartans agccificd in 2. 2 uru tkeu nl ranulortt oud upened ane by one. Frmn ach carton et lease one beg lr:lt lwe laken as a prinary sample. The total weight of all priuary sampiesslumld noc be less rhan 2 kgr which shall be sealed, labeled and sent to laboratory in time.2.4Preparation of tesr sanpleParc of reprasentarive sample is Inken Irorn eh bak of iheprimary sampleand homogenized bygrinling ial h nueat grinder. The hoarogenized sample is thoroughly mixed aud reduced es at leasr Ecc gby guartering. The test batnpte is placed in clean container whick is labcled and sreled.2.5Srortge of the grimpln
The test 5amplea should be storesi unuter-18C.Nare:ln the course of samplipg and xamgle prepararian, precaution musr be token to avoid contaminetion or enyApproved by the Siate Adminisiratia ofImport and Exporl Comaadlty Iaspectlon orthe Peuple's Repubilc of Chiaa on Jun.4:1993mplemeated[romAug.1.1993
SN/T 0212. 2 -93
Eaesors whieh uy cause the chaage of the rexidue content.3Method of determiaatind
3.1Principle
The clapidu! in the hruiler tiaaue Ls extracted with methanol, the extraet is cenerifugalized, andthe supernaiant liyuit! ia cleaned up thrbugh neuiral slumina and anson exehsnge columnx, The cluateis methylated with diazomethane, produeiaz methyl ether of ulojidel(3,5-Jirhloris -4 -metlroxy -2,5lutidine),which is deteered by GC with electras capture dekector. Dinitrochlorobenzene ig used as aninternal atenderd forquumitutive measureraent.3.2Reagenin anil materials.
Unleas otherorise specifiod, the reagen's shoult be of anaiyticul gtade,\water\is dislilrd wateror de-ionised watur.
3.2.1 Me:braoi.Vv99.net
3.2.2 Yeutrul aleminin; For chromatography, 1g0200 mesh, heated at 300 C for 4 h nd coolcd ina desiceatur, ready fnr use.3. 2.3 Anian excharge resin, Dawer 1.X8,1C0-2oa mexh, enhyrinute Eerrs, or equivalent.3. 2. d Methanolic arctic eril xu>lutinn:0. 6% (V/V). add 3 mL pf glacial acerie aeid to 497 mL ofmechanal.
3.2. 5 t-Hexane: Redistilled with alkal.:3.2.6lnternaltandard,2.4-dinicrochiorubenzee,purily>99%3.2.7 n-Hexzane extraetive solution; .I μy tlinicrachlorobenzene/mL n-hexane.3.2.8Diazonethene reageil:Add35ml.of ethaual.10mLol ether and iDml, of KOHsolution(6g/19 int.) into a 125 mI. distilling flask. Place a magnetie bar in the flagk :tnd iummerir ihe flaxk in waler-bath with a magnetit stirrsng device. Attael the drxipping fannrl and the onnderser with theIlark.and corneet the candenser with 1wn 125 mi. enaical flaeka in series,cach contairung 10 ml ofecher urd ler th tips af autlet tubee dip below the surface of the ether. Cool heth rcetiving flaskg inicc-ba:lh. Place in che dropping funnel a sozution of 2l.5 g N mcthyl-N-uicroso-y tujutnesulfutamidefissolveci in 14d mL of ether (diazo zeagcnt). Heut the wuter-ba:h ro ?nC:. Ati tte (nntenis al flaskwhile adding drapwise the diazo rcugent for a μrriud af mire tha 2d nin. Ceaze the distillation henthe diarillare beeomes ncarly colurless. Comline the nntents of two onical flaks. and tedistil it byplacng in the water hath st 7oC. Siore the disrillate (diazomethane reagent) in & sesled conzainerurder-16 C: (:aot exreecink one monch)3.2.9Clapidoi staadard.Puziy>99K3. 2.g, 1 Clopidol siandard stock solutiort: luil μu/L. Accuralely wreigh 1co, 0 mg of clopidol gtau-Gard, rranafer ic into a I L volunetrie fiask with methanol to a toral volume of ea 63 ml, shakp todisgoive clupinta: (ubout 2—a h). Dilute to velume with methanoi and mix welf.3.2.9.2 Intermediate standard solution( 1 ),10 μg/mL. Pipe: 10 tut. uf ibe Etack atandlard soiurioninto 1oo ml vo;umetric flaak uw dilute lu vulume with methanol.3. 2. 9. 3 latcrmeciare atanlard soluziont 1 ): 1 μg/mL. Fipet 10 mL of the inte:mediate standard so-lution (I ) inty a od mf. clumetric ftask and dilute to volume with methanol.3.2.9.4Sinndaed wnrking solution, G,1 g/ml. Pipet 10 ml of thr interandiatn 3. 3.1 Mear grinder; Electtic.SN/T Φ212.2—93
3.3.2Hemogenizer, 3090 r/min.3.3.3 Centrifuge,Model 20 PR-620 Hitachi 4 000 r/min,ar nquivalent.3.3.4Neutral aluiwina column:Add 2galumina(3.2.2) into100 mmX10mm(d) glasg column,3.3.5Anion exchange tegin column.Fil n 25G mm Xlo tnn (id) eolurin witl.DuwexI-Xs rerin,uz-ing de-ionized water to transfer the resin to u heighi of l. s cm(aiter serting down), Elute the columnwith 1oc mL of 1 mol/l. NaOH aislution, rinse with de-ionized water until neutralized, then elule thecplumri with loc ml, oi o, 5 mol/L CHOONa solution, rinse with de iunized water until rrulralizegl.Keep the flow rate always at 2 ml,/min. Add 50 tiL ul methari ayumnus eolution(8 +2) into the col-umr, ready far use.Priur tu r.drzin nff rhe methatol qieoua eolution<8:3),keep the liquid yar-facc sligtly ubove the resin surface.3.3.6Teaz tube, With screw cap.pressure-rengiant and air-tight, 100 mmX1$ mmtid.3.3.7 Gas chrormatograph, With electron ceyture detecior.3. 3.8 Mcrp-syringc: 10 μl.3. 4Prucen
3.4.1Fxtraction
Weigh ca 20 g of the test Numpte3.4.2Cleenup
Place the Bnio ex:lallgr colurlth under the alurmina coluran, pipet 20 mL uf gupernatant extractselujon into the alu;rina column, and let the eolution gass through both columns inr a beaker. Washche col.san ard ite wall with 10 tnL cf rierhano! hy adding it to the elumina wlumn, Rtiove the lumina cnlumn acid the beakes, Place a Bs mL culorirnetric rubr, unrler tlhe anian exchange column and e-lurr she Ji:uidol witt. twu rliony uf J mt.of O.% methaunlie acetic acid soluton with -hn elutionrate of I mi./nrin.Tilutt the eluate to velume with mechanoi, and mix wcll.3.4.3Methylation
Piper 5 ml, of che eluete inso B test icbe (3. 3. 6> and evaporale tn dryusss by placing the tube ina t'C wates-bath with a grtle htreai nf nitrogen. Remove che tube,add o,2 mL of tre:hanoi aque-ous solutinn (s:2), anl warm geatly to redissolve the renidue. oof co roorn temperarurc.Add 1 mlof diazmecaane reagent, close with the acrew cap- and heet for 2 min by plurirg tae the in a 7oCWatez-harh with the weter levei at the saec hcight s ihr noiuihon in the tube. Renove the tuhe andlet ruol to ronru lehupetalure, Takr orr rhe cap and euaporate gently by placing the tube with its lowerend cut:hing the water surfare af the bath (70'a urtil all the ether w expellcd. Add s mL of de-ioz-ized water and 1.C0 mL of n-hexane extzaerive golurion(8.2.7)ti ibur, Iube, Caup ;li ttte arrf nix Ear1 min hy a mixt.Ranrve he tulin ud centrifugaliae for 3 min at 4 Ouo r/min, The upper n-bexaneyyer of ihe ample solutirn ix ugd for GC determinatjon3.4.4 Determination
3.d.4.1c operating coruitioni.G column,Glas,2 rn.X2mmh.Nisrogen,Purityas9.99X,60mL/minc.['olusnn tempetaturerl7oc
SN/T 0212.2—93
d.Injection port tepereture,zooC,e.LDetecturtemperaturr:3(o'C.3.4. 4. 2 Gas chrumatographi: determination: Injwt the test solution (if neceskary, dilute Furtherwilh miore n-besaze extracrive solutian) onto the column of gas chromatogfaph. Injection volumi:2 uL, Retention time under the abave conditious, ca 2 min for methyt ether of clopidol .ca 4 rnin far2.4-dinitrochlorolxenacnc.
3. 4. 5 Preparation of the 6tandard curve, Fipet respectively 0.0. 1,0. 2,0. 4,0. $, 1. 6 tnL of thestandard working solutiotl into & series of test rubes(3. 3- 6y ,proceed by following the bore proce-dures(3. 4. 3-3. 4. 4) with the oniggiun uf the sample rluate. Prepare ihe landard curve hy plottingtlie ratiu of penk hrighta of the etandard and inteenal atandard againsr the concentration (μm/nt,) ofthe standaids.
3. 4. 6 Recorery factor R, Weigh three aamplcs pf 19. 9—z0. 1 g(accuratc tp 0. 1 g) esch al the ho-rnogeaized clopidet.Erer tigsLe at wld sepurelely C, 0, 0, 2 and 1 mL(eyuivalent tu D, O,0. nl aund U, 05pg'g lapidoE) nf mterinediare starudard solution (1 ). Proceed ag in 3. 4. 1-3. 4. 4 and calculate theaverage recuvery factor R.
3 4. 7 Blank rest, Proczed by foilowing thc abvve procedurcs (3. 4. 1—3. 4. 4) with thc omiasion Dfsarpie weighing-
3.4.BCelculation'and expresxiorl af tne resultCalculate the content of the clopidol accarding o the foilowing forrulax
arhrrs
X--Content of elopidol in test gample, mg/kgiRAveragetecovery fcror
c--Cnneeratratwr uf rlopidci in the linal ratnpln olution,rbaruined ftam che atandard curve,pg.mER
--Vulume of the final sample solurion, ml.:mCurrespoauding mass uf ihe serople in the: final test sulution, B-The tontenr of clopidol in the gample can also he caleularerl ly G data ptocegsarNete Ihe teg,xaye valle of the btank teat should he sibtecred from the resul.4Limit ur delermination and recuveryA.1 Lianil af determization
T'he lirit af determinauce af this method is o,wos mg/kg:4.2Recuvery
Acrardiog ta 1he experinenta! data, when the fortifyiag coaceuration ul clopidol is io the rangeof 0, 005 .0.025 tmg/kgthe recovery is 65.4%f:86.2h.Additlnnlexplanutlaas
SN/T0212.2—93
This standard wat proposed by the State Administration ol lmport and Export Commodiry In.spetion of the People's Republic of China.This standard wag dtaftel by the Shainghai, Hehri, Eeifing, Shandong Inuporl and Expnrt Comodtity Trgpection Bureai f the Pepple's Rapublic of China.Principal drelterg ,Zhu Zhiping,Zhujian, Shen Libing+ Ge Xiuli Zhang Jun+ Wang Fenchi,LiuXintong
Refecence:
AOAC-fficial Methods of Analysis.13 th ed,41.013-4l.01 1980.Nate, 'Ths English veraiont, n Lranslalion Irem the Chineda text, in golely for guid anct.
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