SN-T3845-2014出口火鍋底料中多種合成色素的測(cè)定

中華人民共和國(guó)出入境檢驗(yàn)檢疫行業(yè)標(biāo)準(zhǔn)出口火鍋底料中多種合成色素的測(cè)定2014-01-13發(fā)布I請(qǐng)注意本文件的某些內(nèi)容可能涉及專利。本文件的發(fā)布機(jī)構(gòu)不承擔(dān)識(shí)別這些專利的責(zé)任。本標(biāo)準(zhǔn)由國(guó)家認(rèn)證認(rèn)可監(jiān)督管理委員會(huì)提出并歸口1出口火鍋底料中多種合成色素的測(cè)定本標(biāo)準(zhǔn)規(guī)定了出口火鍋底料中多種合成色素的液相色譜檢測(cè)方法。2規(guī)范性引用文件下列文件對(duì)于本文件的應(yīng)用是必不可少的。凡是注日期的引用文件，僅注日期的版本適用于本文件。凡是不注日期的引用文件，其最新版本(包括所有的修改單)適用于本文件。GB/T6682分析實(shí)驗(yàn)室用水規(guī)格和試驗(yàn)方法試樣經(jīng)60℃水浴融化，依次用甲醇-丙酮混合溶液和尿素甲醇溶液(含5%氨水溶液)提取試樣中的合成色素，液液萃取分配后分別經(jīng)聚酰胺樹(shù)脂層析柱和凝膠滲透色譜凈化，液相色譜測(cè)定，外標(biāo)峰面積4.7氨水：濃度為28%。4.10甲醇-丙酮溶液(1+1,體積比);將甲醇(4.1)和丙酮(4.2)等體積混合均勻。4.11乙酸乙酯-環(huán)已烷溶液(1+1,體積比);將乙酸乙酯(4.3)和環(huán)己烷(4.4)等體積混合均勻。4.1280%甲醇溶液(含0.1%磷酸):取80mL甲醇(4.1),加入0.1mL磷酸(4.5),用水稀釋至100mL。4.132mol/L尿素甲醇溶液(含5%氨水溶液):稱取12g尿素(4.6)水(4.7),用甲醇(4.1)稀釋至100mL。2水溶解并稀釋至1000mL。純度均≥90%,各物質(zhì)信息參見(jiàn)附IV、蘇丹橙G和蘇丹紅7B標(biāo)準(zhǔn)貯備液于100mL容量瓶中，用甲醇-丙酮溶液(4.10)稀釋成10pg/mL的混合標(biāo)準(zhǔn)中間液，置于4℃保存。4.23聚酰胺樹(shù)脂層析柱：在層析柱管底部塞入一薄層脫脂棉，稱取1.0g聚酰胺樹(shù)脂(4.22),用10mL5儀器和設(shè)備5.8離心機(jī)：轉(zhuǎn)速不低于5000r/min。6試樣制備與保存3SN/T3845—20147.2.3聚酰胺樹(shù)脂層析柱凈化將水相待凈化液轉(zhuǎn)移至聚酰胺樹(shù)脂層析柱中，以2mL/min～3mL/min流經(jīng)層析柱，同時(shí)用5mL4表1梯度洗脫程序時(shí)間min甲醇%0.01moL/L磷酸鹽緩沖液%007.3.2液相色譜測(cè)定在儀器最佳工作條件下，對(duì)標(biāo)準(zhǔn)工作溶液進(jìn)樣，用標(biāo)準(zhǔn)工作曲線按外標(biāo)法定量。標(biāo)準(zhǔn)工作溶液和樣液中各種合成色素的響應(yīng)值均應(yīng)在儀器的線性范圍內(nèi)。在上述色譜條件下，各目標(biāo)化合物的參考保留時(shí)間參見(jiàn)附錄B中表B.1。標(biāo)準(zhǔn)溶液的液相色譜圖參見(jiàn)附錄C中圖C.1,8空白試驗(yàn)除不加試樣外，均按上述操作步驟進(jìn)行。9結(jié)果計(jì)算和表述用色譜數(shù)據(jù)處理機(jī)或按式(1)計(jì)算試樣中合成色素的含量，計(jì)算結(jié)果須扣除空白值： (1)x;——試樣中待測(cè)物質(zhì)的含量，單位為毫克每千克(mg/kg);c;——從標(biāo)準(zhǔn)曲線上得到的待測(cè)物質(zhì)的溶液濃度，單位為微克每毫升(μg/mL)V——樣液最終定容體積，單位為毫升(mL);m——最終樣液所代表的試樣質(zhì)量，單位為克(g)。10測(cè)定低限和回收率10.1測(cè)定低限本標(biāo)準(zhǔn)對(duì)于火鍋底料中新紅、莧菜紅、胭脂紅、誘惑紅、赤蘚紅、日落黃、酸性紅G、酸性大紅GR、羅丹明B、對(duì)位紅、蘇丹紅I、蘇丹紅Ⅱ、蘇丹紅Ⅲ、蘇丹紅IV、蘇丹橙G和蘇丹紅7B的測(cè)定低限均為mg/kg.回收率本標(biāo)準(zhǔn)的添加水平與回收率范圍參見(jiàn)附錄D中表D.1。5表A.1標(biāo)準(zhǔn)物質(zhì)信息化合物英文名稱CAS號(hào)分子式新紅Ci?H??N?O?S?Na?莧菜紅C??H?N?Na?O??S?胭脂紅誘惑紅AlluraRedACCgH??N?Na?O?S?赤蘚紅日落黃C?H??N?Na?O?S?C??H?N?Na?O?S?對(duì)位紅C?H?N?O?蘇丹紅IC??H??N?O蘇丹紅Ⅱ蘇丹紅ⅢC??H??N?O蘇丹紅IVC??H??N?OC?H?N?O?C??H??N?6(資料性附錄)參考保留時(shí)間表B.1十六種合成色素的參考保留時(shí)間化合物參考保留時(shí)間min化合物參考保留時(shí)間min新紅莧菜紅蘇丹紅I胭脂紅對(duì)位紅日落黃蘇丹紅Ⅱ酸性紅G蘇丹紅Ⅲ誘惑紅蘇丹紅IV赤蘚紅(資料性附錄)標(biāo)準(zhǔn)溶液色譜圖圖C.1十六種合成色素標(biāo)準(zhǔn)溶液的液相色譜圖(0.05mg/L8(資料性附錄)不同添加水平十六種合成色素回收率數(shù)據(jù)化合物添加水平mg/kg回收率范圍%新紅60.5～71.565.7～72.668.4～76.0莧菜紅69.7～81.866.4～80.665.5~79.7胭脂紅69.5～75.573.6～86.870.5～82.3日落黃81.7～90.591.4～98.991.8～96.5赤蘚紅81.0～100.192.9～100.278.3～87.2誘惑紅91.0～100.190.8～99.584.7～94.6酸性紅G80.6～90.686.3～90.282.7～93.681.0～87.180.5～86.780.7～89.072.0～92.081.4～98.078.8～97.19化合物添加水平mg/kg回收率范圍%對(duì)位紅0.0172.0～87.00.0275.9～87.40.0579.4～88.80.0174.1～83.40.0275.9～85.20.0576.5～84.50.0176.7～97.10.0281.7～96.00.0578.0～92.1蘇丹紅Ⅱ0.0169.9～89.10.0277.5～92.00.0581.7～97.0蘇丹紅Ⅲ0.0175.0～97.30.0275.5～92.40.0579.5～93.70.0175.0～87.50.0274.1～87.70.0579.5～92.5蘇丹紅IV0.0169.8～80.70.0269.8～82.10.0582.5～82.4tificationandAccreditationofthePeople'sRepublicofChina.RepublicofChina.ThisstandardwasmainlydraftedbyXiCunxian,TangBobin,ZhangLei,LiXianliang,ZhengXiaoling,DeterminationofmultiplesyntheticdyesinbottommaterialofchafingdishforexportThisstandardspecifiesthemethodsofsamplepreparationandqualifiedandquantifieddeterminationdishforexport.Thisstandardisapplicabletothedeterminationofnewred,carmine,amaranth,allurared,erythrosine,sunsetThefollowingnormativedocumentscntainprovisionwhiprovisionofthispart.Fordaterefeirences,subsequentamgatethepossibilityofapplyingthemostrecenteditionsofthenormativedocumentsindicatedbamide-methanolsolution(contain5%ammoniumhydroxidesolution),respectively,afterliquid-GPC,respectively.TheanalytewasdeterminedbyHPLC,quantifiedSN/T3845—20144.4Cyclohexane.4.7Ammoniumhydroxide:28%4.8Sodiumdihydrogenphosp4.9Disodiumhydrogen4.10Methanol-acetonesolution(1+1,V/V):Equivalentvolumemixtureofmethanol(4.1)andacetone(4.2).4.11Ethylacetate-cyclohexanesolution(1+1,V/V):Equivalentvolumemixtureofethylacetate(4.3)andcyclohexane(4.4)4.1280%methanolsolution(contain0.1%phosphoricacid):Dilute80mLmethanol(4.phosphoricacid(4.5)withwatertothevolumeof100mL4.132mol/LCarbamide-methanolsolution(contain5%ammoniumhydroxidesolution):Dilute12.0gcarbamide(4.6)and5mLammoniumhydroxide(4.7)withmethanol(4.1)tothevolumeof4.1450%Phosphoricacidsolution:Equivalentvolumemixtureofphosphoric(4.5)andwater4.165%Ammoniumhydroxide-methanolsolution:Dilute5mLammoniumhydroxide(4.7)with4.170.01mol/Lphosphatebuffersolution:Accuratelyweight1.41gsodiumdihydrogenphosphateanhydrous(4.8)and1.56disodiumhydrogenphosphateanhydrous(4.9),dissolvedin1000mLwater,andmixtohomogeneity4.19Stockstandardsolutioamaranth,allurared,erythrosine,sunsetyellow,acidredG,acidscarletstockstandardsolutionwas3months.4.21Standardworkingsolution:Accordingtotherequirement,nol-acetonesolution(4.10)justbeforeuse.1.0gpolyamide(4.22)with0.1%Phosphoric4.24Mermbranefilter:0.22μm,Hydrophilicpolyethersu5.2GelpermeationchromatograpSN/T3845—20145.9Chromatographycolumn:1.5cm×10cmAbout500grepresentativfist,thengrindedandblendedbyablendeAccuratelyweigh5.00gofthetestsample(accurateto0.01g)intoa50mLcentrifugetubewithcap,add20mLmethanol-acetonesolution(4.10),heatedin60℃water-bathfor10min,vortexfor2min,thencentrifugeat5000r/minfor5min.Transferthesupernatantto150mLdistillationflask,theresiduewasre-extractedoncewith20mLmethanol-acetonesolution(4.10),andre-extractedwith15mL2mol/LCarbamide-methanolsolution(4.13)fortwice.Aftercentrifuging,combinethesupernatanttothedistillationflask;evaporatetheextractnearlytodrynessonrotaryevaporatorbe-Thedistillationflaskwaswashedwith5mLofethylacetate-cyclohexane(4.11)and5mLofwaterfortwice,respectivly,andthewashingsolutionwastransferredintoa50mLpolypropylenecentrifugetube,vortex,for2minandcentrifuged.Theorganicphasewastransferredtoa10mLcol-orimetrictube,concentratednearlytodrynesswithevaporatorat40℃,dissolvedresiduewith5mLethylacetale-cyclohexane(4.11),cleanupwithGPC.Theaqueousphasewasaddedphosphoricacidsolution(4.14),mixedtocleanupCleanuptheorganicphasesolution(7.2)withGPCaccordingthefollowingconditions:b)Mobilephase:ethylacetate-cyclohexane(4.11).nol(4.12),thendrythecolumnwithair.TheanalytesarCombinedGPCcleanupsolution(7.2.1)and9mLseveraltimes,combinethesolution.Evaporatetheelutesolutionnearlytodryat40flow.Dilutetheresiduetothevolumeof1.0mLofmethanol-acetone(4.10),vortexmixed,filteredthroughhydrophilicpolyethersulfonemembrane,andfinalsolua)LCcolumn:Cg,250mm×4.6(i.d.),5μm,(orotherconformablecolumn),c)Detectionwavelength:programvariablewavelength,0～13.7min,520nm;13.71min~15min.420nm;15.01min～30min,520nm.f)Injectorvolume:10uL.g)Equilibriumtime:5.0min.00Undertheoptimizedinstrumentsworkingcondition,differentconcentrationsolutionsareinjectionintotheinstrumentinseparatsamplesolutionshouldbewithinthelinearrangeoftheinstrumentaldetection.Undertheoptimizedinstrumentsworkingconditiontheret9CalculationandexpressionofresultCalculatingthecontentofanalytesconcentrationaccordingtotheformula(1):c,一theconcentrationofanalytewhichisquantifiedbystandardcalibrationcurV—thefinalvolumeofsamplesolution,mLm—thecorrespondingmassoftestsampleinthefinalsamplesolution,gThelimitofquantificationfornewred,carmine,amaranth,allurared,erythrosine,sunsetyellow,acidAccordingtotheexperimentalCompoundCASNo.MolecularformulaNewRed220658-76-4C?H??N?O?,S?Na?915-67-3C??H??N?Na?O?S?CarmineC??H??O??AlluraRedAC25956-17-6CH??N?Na?O?SErythrosinB568-63-8SunsetYellow2783-94-0C??H?N?Na?O,S?AcidRed13734-67-6CgH??N?Na?OS?BrilliantCrocein5413-75-2C?H??N?Na?O,S?RhodamineB81-88-9C??H??CIN?O?ParaRed6410-10-2C?gH??N?O?842-07-9C?H??N?O3118-97-6CH??N?O85-86-9C?H??N?O85-83-6C??H??N?OSudanOrangeG2051-85-6C??H?N?O?6368-72-5C??H??N?(informative)Compound