SN 1544-2005进出口动物源性食品中玉米赤霉醇残留量的检验方法 高效液相色谱-质谱质谱法

中华人民共和国出入境检验检疫行业标准进出口动物源性食品中玉米赤霉醇残留量的检验方法高效液相色谱-质谱/质谱法2005-02-17发布国家质量监督检验检疫总局I本标准的附录A为资料性附录。本标准由国家认证认可监督管理委员会提出并归口。本标准系首次发布的出入境检验检疫行业标准。1进出口动物源性食品中玉米赤霉醇残留量本标准规定了动物源性食品中玉米赤霉醇残留量检验的抽样、制样和高效液相色谱-质谱/质谱检以不超过2500件为一检验批。抽样数量见表1。表1单位为件最低抽样数126～1005101～250251～500501～10001001～25002.3抽样方法按2.2规定的抽样件数随机抽取，逐件开启。从每件中至少取一袋作为原始样品。如每件中无小包装或有小包装，但每袋小包装质量超过2kg,则可用灭菌刀具从每件中割取不少于500g,混合后置于清洁容器内作为混合原始样品。混合原始样品的总中质量不少于2kg,加封后，标明标记，及时送交从混合原始样品中取出部分有代表性的样品，充分混匀，用四分法缩分至不少于500g,均分成两3.1方法提要匀浆组织样品经葡萄糖苷酸酶水解后，依次用乙醚和氢氧化钠溶液提取玉米赤霉醇。氢氧化钠提取液中和后经C₁₈固相萃取柱净化。甲醇洗脱液经蒸发，定容后供高效液相色谱-质谱/质谱仪测定，外标法定量。23.2.7磷酸：纯度>85%;密度1.7g/mL。3.2.8葡萄糖苷酸酶。酸调节至pH4.8。制成0.10mg/mL的标准储备液。储备液保存于-18℃。称取5g试样(精确至0.01g),置于洁净的50mL具塞离心管中，加入10mL乙酸钠缓冲溶液(3.2.13)和25μL葡萄糖苷酸酶，旋涡混匀，于37℃中保温8h～14h。34玉米赤霉醇添加浓度及其回收率数据：——1pg/kg时，回收率72%;——5μg/kg时，回收率74%;5SN/T1544—2005(资料性附录)0ThisstandardwasproposedbyandisunderthechargeofNationRegulatoryCommissionforCertifi-ThisstandardwasdraftedbyShanghaiEntry-ExitinspectionandQuarantineBureauofthePeople'sRepublicofChina.ThisstandardwasmainlydraftedbyFangXiaoming,ChenJiahua,NiXinluandTangYifeng.Thisstandardispromulgatedforthefirsttime.7productsforimportandexport—HPLC-MS/MSThisstandardspecifianimaloriginalproductsbyhigh-performanceliquidchromatography-tandemmassandspecification,shouldbTable1Minimumnumberofpackagestobetaken126～1005101～250251~500501～10001001～2500samplewasdevidedintwoandhomogenized.Packegeandlabel.8Thetestsampleshouldbestoredat-18℃.AhomogenizedtissuesampleishydrolyzedwithB-glucuronidaseandthehydrolysateisextractedwithethylether.Thesupernatantisevaporatedtodrynessandtheresidueisdissolvedinchloroformandre-extractedwithsodiumhydroxidesoCasolid-phaseextractioncartridge,andanalyzedbyelectrosprayHPLC-MS/MSinnegativeionmode.Reagentswereofanalytical-reagentgradeexceptforthespecificreagents.Thewaterusedwasdoublydistilledordeionizedw3.2.1Acetonitrile:LCgrade.3.2.6Anhydroussodiumacetate.3.2.7Phosphoricacid:purity>85%;3.2.9Zeranol(a-zearalanol):purity97%.withwater.3.2.13Sodiumacetatebuffer,50mmol/L:Take4.1gofanhydroussodiumacetate,dissolveand9diluteto1000mLwithwater.AdjustpHto4.8withglacialaceticacid.3.2.14Zeranolstocksolution,0.10mg/mL:Accuratelyweighproperzeranosolvedwithacetonitrile.Tstocksolution(3.2.14)into100mLvolumetrictively.ThesolutionsanolIntermediatesolutions(3.2.15).Thematrix-matchedstandardsarestoredat-week.3.2.17Cisolid-phaseextractioncartridge,500m3.3.1High-performanceliquidchromatography-electrospraytandemmassspectrometry.3.3.3VacuummanifoldforsolidWeigh5.0g±0.1gofhomogenizedtissuesampleintoa50mLcentrifugetube.Add10mLofsodi-SN/T1544—2005umacetatebuffer(3.2.13)and25μLofβ-glucuronidase,mixwell,andthenincubat(3.2.13)and2mLofmethan200pLofacetonitrileforanalb)Mobilephase:acetonitrile-water(40+60)containing0.025%aceticacid.d)Columntemperature:22℃.a)lonizationmode:negativeelectrospray.b)Conevoltage:50V.c)Capillaryvoltage:3kV.d)Collisionvoltage:20V.e)Analyzervacuum:<3mPa(3×10~5mbar).f)Collisiongas:argon.h)Nebulisergasflow:480L/h.i)Multiplereactionmonitoring(MRM)mode:m/z321→303,m/z321→277.m/forAnalyzematrix-matchedstandards(3.2.16)inorderofincreasingconcentration,obtainingthema-withoutadditionofthesample.Theconcentrationofzeranolinsampleiscalculatedwithchromatographicdataprocessoro…………(1)Where:X—theconcentrationofanalyteofinterestinsample,ug/kg:c~—theconcentrationofanalytefromcalibrationcurve,ng/mL;V--—finalvolumeofsampleextract,mL;m——weightofsample,g.Note:Theblankvalueshouldbesubtracte