SNT 3235-2012 出口動(dòng)物源食品中多類禁用藥物殘留量檢測(cè)方法

出口動(dòng)物源食品中多類禁用藥物殘留量檢測(cè)方法液相色譜-質(zhì)譜/質(zhì)譜法1SN/T3235—2012出口動(dòng)物源食品中多類禁用藥物殘留量檢測(cè)方法液相色譜-質(zhì)譜/質(zhì)譜法相色譜-質(zhì)譜/質(zhì)譜篩查和測(cè)定。多種禁用獸藥標(biāo)準(zhǔn)品信息參見(jiàn)附錄A。晶紫及隱性結(jié)晶紫的篩查和測(cè)定。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3樣品制備與保存從所取全部樣品中取出有代表性樣品可食部分約500g,用組織搗碎機(jī)充分搗碎均勻，裝入潔凈容3.2牛奶從所取全部樣品中取出有代表性樣品500mL,充分混勻，裝入潔凈容器中，密封，并標(biāo)明標(biāo)記，于—18℃以下冷凍存放。2SN/T3235—2012對(duì)于水產(chǎn)品中的禁用藥物，分別以氨化乙腈和酸化乙腈提??；對(duì)于牛奶、肝臟和肌肉中的禁用藥物，用β-葡糖苷醛甙酶-芳基硫酸酯酶在乙酸銨緩沖液中酶解后，分別以氨化乙腈和酸化乙腈提取。提取液以QuEChERS吸附劑凈化后，用高效液相色譜-質(zhì)譜/質(zhì)譜儀測(cè)定，內(nèi)標(biāo)法或外標(biāo)法定5試劑和材料除另有規(guī)定外，所有試劑均為分析純；水為GB/T5.8N-丙基乙二胺吸附劑(PSA),40μm～60μm粒徑范圍，100A平均孔徑。5.10石墨化碳黑吸附劑(GCB),40μm～60μm粒徑范圍，60A平均孔徑。5.11無(wú)水硫酸鈉，分析純，500℃灼燒4h,置于干燥器中備用。5.12無(wú)水硫酸鎂，分析純，500℃灼燒4h,置于干燥器中備用。5.13乙酸銨緩沖溶液：溶解7.7g乙酸銨(5.5)于480mL水中，用冰乙酸(5.3)調(diào)節(jié)溶液pH值到5.2,以水定容至500mL。5.141%氨水-乙腈溶液：準(zhǔn)確吸取40mL25%的氨水(5.4)至1000mL容量瓶，用乙腈(5.1)定容至5.151%乙酸-乙腈溶液：準(zhǔn)確吸取10mL冰乙酸(5.3)至1000mL容量瓶，用乙腈(5.1)定容至刻度，混勻。5.1620%的乙腈-水溶液：準(zhǔn)確吸取10mL乙腈(5.1),加入90mL水，混勻。5.17色譜流動(dòng)相1,0.1%甲酸-水溶液：取1mL甲酸(5.6)用去離子水定容到1000mL,混勻。5.18色譜流動(dòng)相2,0.1%甲酸-甲醇溶液：取1mL甲酸(5.6)用甲醇(5.2)定容到1000mL,混勻。5.19QuEChERS吸附劑1(適用于豬肝和動(dòng)物組織樣品):準(zhǔn)確稱取100mgPSA(5.8)、50mgC?s(5.9)、20mgGCB(5.10)和400mg無(wú)水MgSO?(5.12),儲(chǔ)存于25mL具螺旋蓋聚丙烯離心管中，旋緊5.20QuEChERS吸附劑2(適用于牛奶、水產(chǎn)品樣品):準(zhǔn)確稱取100mgPSA、40mgC?s和600mg無(wú)水MgSO?,儲(chǔ)存于25mL具螺旋蓋聚丙烯離心管中，旋緊管蓋，置于干燥箱內(nèi)備用。羥基苯甲酸內(nèi)酯類、三苯甲烷類以及氯霉素標(biāo)準(zhǔn)物質(zhì)的純度均≥99%,參見(jiàn)附錄A表A.1。唑、D?-結(jié)晶紫、D?-隱性孔雀石綠、D?-己二烯雌酚及D?-氯霉素的標(biāo)準(zhǔn)物質(zhì)的純度均≥95%。5.23標(biāo)準(zhǔn)貯備液：準(zhǔn)確稱取10.0mg按其純度折算為100%質(zhì)量的各個(gè)化合物的標(biāo)準(zhǔn)物質(zhì)(5.21),甲醇溶解定容至10.0mL,溶液濃度相當(dāng)于100.0mg/L,-30℃冷凍保存，有效期為12個(gè)月。5.24標(biāo)準(zhǔn)中間溶液：準(zhǔn)確吸取1.0mL各標(biāo)準(zhǔn)貯備溶液(5.23),以甲醇稀釋并定容于100mL棕色容3SN/T3235—20125.25空白基質(zhì)溶液：選取不含待測(cè)物的樣品，按照5.26基質(zhì)混合標(biāo)準(zhǔn)溶液：根據(jù)需要用空白基質(zhì)溶液(5.25)稀釋標(biāo)準(zhǔn)中間溶液(5.24)成適合濃度的混5.27內(nèi)標(biāo)貯備液：準(zhǔn)確稱取10.0mg按其純度折算為100%質(zhì)量的內(nèi)標(biāo)標(biāo)準(zhǔn)物質(zhì)(5.22),以甲醇溶解5.28內(nèi)標(biāo)中間溶液：準(zhǔn)確吸取1mL各內(nèi)標(biāo)貯備溶液(5.26),以甲醇稀釋并定容于100mL棕色容量5.29內(nèi)標(biāo)工作液：根據(jù)需要用空白基質(zhì)溶液(5.25)稀釋內(nèi)標(biāo)中間溶液(5.28)成合適濃度的標(biāo)準(zhǔn)內(nèi)標(biāo)6儀器和設(shè)備6.9組織搗碎機(jī)。6.10恒溫水浴振蕩器。6.11平板振蕩器。稱取試樣2g(精確到0.01g),置于50mL具螺旋蓋聚丙烯離心管中，準(zhǔn)確加入20μL內(nèi)標(biāo)工作液 (5.29),8mL乙酸銨緩沖液(5.13),在均質(zhì)器中高速均質(zhì)30s,加入30μLβ-葡糖苷醛甙酶-芳基硫酸酯酶(5.7),渦旋混勻30s,37℃振蕩溫育12h。取出樣品靜置，使其溫度降至室溫。加入15mL1%氨水-乙腈溶液(5.14),5.0g無(wú)水硫酸鈉(5.11),渦旋混勻1min后，在4℃、9500r/min離心5min,收集上清液于另一50mL離心管。剩余部分重新加入15mL1%乙酸-乙腈溶液(5.15),旋緊螺旋蓋，室7.1.2水產(chǎn)品(魚)稱取試樣2g(精確到0.01g),置于50mL具螺旋蓋聚丙烯離心管中，準(zhǔn)確加入20μL內(nèi)標(biāo)工作液 (5.29),8mL乙酸銨緩沖液(5.13),高速渦旋混勻30s,室溫靜置30min。加入15mL1%氨水-乙腈溶液(5.14),5.0g無(wú)水硫酸鈉(5.11),渦旋混勻1min后，在4℃、9500r/min離心5min。收集上清4SN/T3235—2012液于另一個(gè)干凈的50mL具螺旋蓋離心管中。剩余混合物中加入15mL1%乙酸-乙腈溶液(5.15),旋緊螺旋蓋，室溫置于水平振蕩器，振蕩提取10min,在4℃、9500r/min下離心5min,吸取上清液層溶一次性全部將QuEChERS吸附劑1加入肝臟或肌肉樣品的提取液中，或?qū)uEChERS吸附劑2加入牛奶、水產(chǎn)品提取液中，旋緊螺旋蓋，高速渦旋1min,在4℃、于95有機(jī)相溶液，在減壓旋轉(zhuǎn)蒸發(fā)裝置上于42℃蒸發(fā)至近干。加入2.0mL的10%乙腈-水溶液(5.16)溶解殘?jiān)?，渦旋混合1min,過(guò)0.22pm濾膜(5.30)。濾液供液相色譜-質(zhì)譜/質(zhì)譜儀測(cè)定。7.3.1液相色譜條件B2:乙腈。梯度洗脫程序見(jiàn)表1。表1液相色譜梯度洗脫程序表正離子模式(ESI+)負(fù)離子模式(ESI-)時(shí)間A1B1時(shí)間A2B2min%%miin%%4.0045.07.3.2質(zhì)譜條件離子源：電噴霧ESI,正/負(fù)離子模式。掃描方式：多反應(yīng)監(jiān)測(cè)MRM(鎖定保留時(shí)間)。霧化氣壓力(GS1)、氣簾氣壓力(CUR)、輔助氣流速(GS2)均為高純氮?dú)饣蚱渌线m氣體；使用前應(yīng)調(diào)節(jié)各氣體流量以及離子源溫度(TEM)使質(zhì)譜靈敏度達(dá)到檢測(cè)要求，質(zhì)譜等詳細(xì)條件參見(jiàn)電噴霧電壓(IS)、碰撞電壓(CE)、去簇電壓(DP)、碰撞室入口電壓(EP)、碰撞室出口電壓(CXP)應(yīng)優(yōu)化至最佳靈敏度。監(jiān)測(cè)離子對(duì)和定量離子等詳細(xì)儀器條件參見(jiàn)表C.1。7.3.3液相色譜-質(zhì)譜/質(zhì)譜測(cè)定根據(jù)試樣中被測(cè)物的藥物含量，選取響應(yīng)值相近的混合標(biāo)準(zhǔn)工作液(5.25)同時(shí)進(jìn)行分析。混合標(biāo)5SN/T3235—2012準(zhǔn)工作液和待測(cè)液中各種藥物的響應(yīng)值均應(yīng)在儀器線性響應(yīng)范圍內(nèi)。如果含量超過(guò)標(biāo)準(zhǔn)曲線范圍，應(yīng)用甲醇：水(1/9,體積比)稀釋到合適濃度后分析。在上述儀器條件下，44種化合物的總離子流色譜圖8結(jié)果計(jì)算和表述8.1定性標(biāo)準(zhǔn)8.1.1保留時(shí)間待測(cè)樣品中化合物色譜峰的保留時(shí)間與標(biāo)準(zhǔn)溶液相比變化范圍應(yīng)在±2.5%之內(nèi)。每種化合物的質(zhì)譜定性離子必須出現(xiàn)，至少應(yīng)包括一個(gè)母離子和兩個(gè)子離子，而且同一檢測(cè)批次，對(duì)同一化合物，樣品中目標(biāo)化合物的兩個(gè)子離子的相對(duì)豐度比與濃度相當(dāng)?shù)臉?biāo)準(zhǔn)溶液相比，其允許偏差不超過(guò)表2規(guī)定的范圍。表2定性時(shí)相對(duì)離子豐度的最大允許偏差>20~~50>10～20允許的相對(duì)偏差/%士20士308.2定量結(jié)果與表述采用標(biāo)準(zhǔn)曲線法定量，按式(1)或儀器數(shù)據(jù)處理系統(tǒng)計(jì)算所有禁用藥物的殘留含量，計(jì)算結(jié)果需扣除空白值。c——待測(cè)組分響應(yīng)值在標(biāo)準(zhǔn)曲線上計(jì)算得到的濃度，單位為微克每升(μg/L);9測(cè)定低限和回收率9.1測(cè)定低限(LOQ)9.2回收率在不同的基質(zhì)中，各化合物添加回收率范圍的實(shí)驗(yàn)數(shù)據(jù)參見(jiàn)表F.1～表F.5。6(資料性附錄)英文名稱中文名稱CAS號(hào)結(jié)構(gòu)式相對(duì)分子質(zhì)量β受體激動(dòng)劑類(8種)克侖特羅C??H?gC?N?O沙丁胺醇C?H?NO?萊克多巴胺C??H??NO?西瑪特羅C?H??N?O特布他林C?2H?9NO?妥布特羅菲諾特羅雄性激素類(10種)去氫睪酮C?9H??O?丙酸睪酮dehydro-17a-methyltestrost去氫甲睪酮methyltestosterone甲睪酮C?0H?0O?睪酮C?9H?8O?丙酸諾龍C??H?0O?表睪酮CgH??O?氟甲睪酮諾龍C?gH??O?nandrolonephenylpropionate苯丙酸諾龍?zhí)瞧べ|(zhì)激素類(7種)倍他米松醋酸可的松C??H??O?丙酸倍氯米松C2?H?9CIO?氟氫可的松氫化可的松C??H??O?曲安西龍曲安奈德雌性激素類(3種)己烯雌酚己烷雌酚CgH??O?己二烯雌酚7SN/T3235—2012表A.1(續(xù))英文名稱中文名稱結(jié)構(gòu)式相對(duì)分子質(zhì)量硝基咪唑類(5種)羥甲基甲硝咪唑C?H?N?O?地美硝唑C?H?N?O?洛硝噠唑metronidazole甲硝唑C?H?N?O?異丙硝唑C?HnN?O?二羥基苯甲酸內(nèi)酯類(6種)α-玉米赤霉醇C??H??Oβ-玉米赤霉醇C?8H??O?α-玉米赤霉烯醇C?8H??O?β-玉米赤霉烯醇C?8H??O?玉米赤霉酮C?gH??O玉米赤霉烯酮C?gH??O?三苯甲烷類(4種)孔雀石綠隱性孔雀石綠結(jié)晶紫隱性結(jié)晶紫鎮(zhèn)靜劑類(2種)氯丙嗪C??H?9CIN?S地西泮C??H??ClN?O氯霉素氯霉素CH??C?2N?O?8SN/T3235—2012(資料性附錄)API5000型質(zhì)譜/質(zhì)譜儀參考質(zhì)譜條件參考質(zhì)譜條件正離子模式(ESI+)負(fù)離子模式(ESI-)離子噴霧電壓4500V—4500V氣簾氣壓力霧化氣壓力輔助氣壓力4.4..bar4.1bar離子源溫度450℃9化合物離子對(duì)Q1/Q3(m/z)保留時(shí)間min去簇電壓V碰撞氣能量碰撞室出口電壓V內(nèi)標(biāo)物ESI+模式丙酸倍氯米松beclomethasone diptopionate521.4/503b521.4/337.24倍他米松betanethasone393.1/373.2b393.1/355.28去氫睪酮boldenone287.6/120.9287.6/134.9氯丙嗪chlorpromazine319.1/86.16D?-氯丙嗪319.1/58.2西瑪特羅cimaterol220/1601b8220/202.16克侖特羅clentuterol276.9/202.8bD?-克侖特羅276.9/258.98醋酸可的松cortisoneacetate403.4/343.2h6403.4/163.24結(jié)晶紫crystalviolet372.2/356.2hD?-結(jié)晶紫372.2/251.2-D?-沙丁胺醇D?-salbutnol243.1/224.9D?-氯丙嗪D?-chlorpromazine325.1/91.8D?-萊克多巴胺D?-ractopamine308.2/290.28D?-隱性結(jié)晶紫D?-leucocrystalviolet380.2/2456D?-孔雀石綠D?-malachitegreen334.1/318SN/T3235—2012化合物離子對(duì)Q1/Q3(m/z)保留時(shí)間去簇電壓V碰撞氣能量碰撞室出口電壓V內(nèi)標(biāo)物D?-克倫特羅D?-clentuterol286/267.94D?-洛硝噠唑D?-ronidazole98D?-結(jié)晶紫378.3/362.2D?-隱性孔雀石綠地西泮6地美硝唑表睪酮289.4/108.9b289.4/96.9非諾特羅304.2/286.2b6氟氫可的松46羥甲基甲硝咪唑46氟甲睪酮?dú)浠傻乃蒱ydrocortisone8363.2/309.06異丙硝唑隱性結(jié)晶紫374.2/239.0D?-隱性結(jié)晶紫374.2/358.1隱性孔雀石綠331.2/315.3bD?-隱性孔雀石綠331.2/239.14去氫甲睪酮methandienone301.4/121.2b4表C.1(續(xù))化合物離子對(duì)Q1/Q3(m/≈)保留時(shí)間min去簇電壓V碰撞氣能量碰撞室出口電壓V內(nèi)標(biāo)物甲睪酮methyltestosterone303.2/97b8303.2/108.9甲硝唑metronidazole4孔雀石綠malachitegreen329.1/313.36D?-孔雀石綠329.1/208.34丙酸諾龍nandrolone331.4/257.4b331.4/275.0諾龍nandrolone275/109b275/257萊克多巴胺ractopamine302.1/164.16Ds-萊克多巴胺302.1/283.94洛硝噠唑ronidazole201/140bD?-洛硝噠唑沙丁胺醇240.1/222D?-沙丁胺醇240.1/148特布他林terbutaline226.1/152.1b4226.1/170丙酸睪酮testosteronepropionate345.4/97.1345.4/109.1睪酮testosterone289.3/96.9b289.3/108.9妥布特羅tobututerol228.2/172t228.2/153.8曲安西龍triamcinolonel395.4/225.1395.4/357.4曲安奈德triamcinoloneacetonide435.2/338.9b435.2/396.9ESI-模式己二烯雌酚dienestrol265.1/92.9bD?-己二烯雌酚265.1/249化合物a離子對(duì)Q1/Q3(m/z)保留時(shí)間min去簇電壓V碰撞氣能量碰撞室出口電壓V內(nèi)標(biāo)物己烯雌酚diethylstilbestrol267/250.9h-115-39-13267/237-115-46-13己烷雌酚hexestrol269/134b-75-22-11269/119--11D8-己二烯雌酚D8-dienestrol275/259-108-36-13D5-氯霉素D5-chloramphenicol326/157.1-51-24-10玉米赤霉烯酮zearalenone317.1/175b-101-32317.1/131-51-40α-玉米赤霉烯醇319.1/301.1b-111319[1/275.2-111429β-玉米赤霉烯醇319.1/301.1)-111-42319.1/275.2—1m-29玉米赤霉酮zearalanone319.1/301.1-111-42319.1/275.2-111-29氯霉素chloramphenicol321/152b-65-25D?-氯霉素321/257-101-42α-玉米赤霉醇321/227.2b-101-29321/303,4-101-42β-玉米赤霉醇321/227.2h-29321/3Q3.4-42供參考，并不涉及商業(yè)目的，鼓勵(lì)標(biāo)準(zhǔn)使用者嘗試不同型號(hào)的儀器。典型譜圖5.5e53.5e52.5e52.0e5·4.8c64.2e6- 2.8e6-2.4c6-ESI+b)圖D.144種藥物標(biāo)準(zhǔn)物質(zhì)的總離子流色譜圖SN/T3235—2012Intensity/Intensity/cps10)西馬特羅16)結(jié)晶紫2)丙酸倍氯米松5)去氫睪酮8)氯丙嗪14)醋酸可的松17)d3-沙丁胺醇3)倍他米松6)去氫蟻酮12)克侖特羅15)結(jié)晶紫18)d5-氧丙嗪21)d5-孔雀石綠圖D.244種藥物及內(nèi)標(biāo)物的重構(gòu)離子色譜圖31)菲諾特羅38)38)HMMHI57)諾龍56)丙酸諾龍57)諾龍56)丙酸諾龍59)萊克多巴胺62)洛硝噠唑SN/T3235—2012SN/T3235—201285)玉米赤霉烯酮90)β-玉米赤霉烯醇93)d5-氯都素圖D.2(續(xù))序號(hào)測(cè)定低限序號(hào)測(cè)定低限1β-受體激動(dòng)劑類西瑪特羅0.5二羥基苯甲酸內(nèi)酯類玉米赤霉烯酮0.52克侖特羅0.5α-玉米赤霉烯醇13菲諾特羅1β-玉米赤霉烯醇14萊克多巴胺0.5玉米赤霉酮15沙丁胺醇0.5α-玉米赤霉醇16特布他林0.5β-玉米赤霉醇17妥布特羅0.5硝基咪唑類地美硝唑0.58雄性激素類去氫睪酮1異丙硝唑0.59表睪酮1甲硝唑0.5去氫甲睪酮1洛硝噠唑0.5諾龍1雌性激素類已二烯雌酚1睪酮1己烯雌酚1糖皮質(zhì)激素類丙酸倍氯米松1己烷雌酚1倍他米松0.5鎮(zhèn)靜劑類氯丙嗪0.5曲安西龍0.5地西泮0.5曲安奈德1氯霉素氯霉素0.05序號(hào)測(cè)定低限ug/kg序號(hào)測(cè)定低限1β-受體激動(dòng)劑類西瑪特羅雄性激素類諾龍12克侖特羅睪酮13萊克多巴胺糖皮質(zhì)激素類倍他米松14沙丁胺醇?xì)浠傻乃?5特布他林曲安西龍16妥布特羅曲安奈德17雄性激素類去氫睪酮二羥基苯甲酸內(nèi)酯類玉米赤霉烯酮8表睪酮α-玉米赤霉烯醇19甲睪酮1β-玉米赤霉烯醇1序號(hào)測(cè)定低限序號(hào)測(cè)定低限二羥基苯甲酸內(nèi)酯類玉米赤霉酮雌性激素類己二烯雌酚1α-玉米赤霉醇1已烯雌酚Iβ-玉米赤霉醇1己烷雌酚1硝基咪唑類洛硝噠唑鎮(zhèn)靜劑類氯丙嗪異丙硝唑地西泮甲硝唑1氯霉素氯霉素洛硝噠唑序號(hào)測(cè)定低限序號(hào)測(cè)定低限1β-受體激動(dòng)劑類西瑪特羅酸內(nèi)酯類玉米赤霉烯酮2克侖特羅α-玉米赤霉烯醇l3菲諾特羅β-玉米赤霉烯醇14萊克多巴胺玉米赤霉酮15沙丁胺醇α-玉米赤霉醇16特布他林β-玉米赤霉醇17妥布特羅硝基咪唑類地美硝唑8雄性激素類去氫睪酮1異丙硝唑9表睪酮1甲硝唑去氫甲睪酮洛硝噠唑諾龍雌性激素類己二烯雌酚1睪酮1己烯雌酚1糖皮質(zhì)激素類丙酸倍氯米松1己烷雌酚l倍他米松鎮(zhèn)靜劑類氯丙嗪曲安西龍地西泮曲安東德1氯霉素氯霉素序號(hào)測(cè)定低限序號(hào)測(cè)定低限1β-受體激動(dòng)劑類西瑪特羅4β-受體激動(dòng)劑類萊克多巴胺2克侖特羅5沙丁胺醇3菲諾特羅16特布他林SN/T3235—2012序號(hào)測(cè)定低限序號(hào)測(cè)定低限7β-受體激動(dòng)劑類妥布特羅二羥基苯甲酸內(nèi)酯類玉米赤霉酮18雄性激素類去氫睪酮1α-玉米赤霉醇19表睪酮1β-玉米赤霉醇去氫甲睪酮1地美硝唑諾龍1-24異丙硝唑硝基咪唑類睪酮1甲硝唑糖皮質(zhì)激素類丙酸倍氯米松1洛硝噠唑倍他米松雌性激素類己二烯雌酚1曲安西龍己烯雌酚1曲安奈德己烷雌酚1二羥基苯甲酸內(nèi)酯類玉米赤霉烯酮鎮(zhèn)靜劑類氯丙嗪α-玉米赤霉烯醇地西泮β-玉米赤霉烯醇1氯霉素氯霉素序號(hào)測(cè)定低限序號(hào)測(cè)定低限1β-受體激動(dòng)劑類西瑪特羅糖皮質(zhì)激素類糖皮質(zhì)激素類丙酸倍氯米松2克侖特羅倍他米松3菲諾特羅醋酸可的松4萊克多巴胺氟氫可的松5沙丁胺醇?xì)浠傻乃?特布他林曲安西龍7妥布特羅曲安奈德8雄性激素類去氫睪酮二羥基苯甲酸內(nèi)酯類玉米赤霉烯酮9表睪酮α-玉米赤霉烯醇氟甲睪酮1β-玉米赤霉烯醇去氫甲睪酮玉米赤霉酮甲睪酮1α-玉米赤霉醇丙酸諾龍β-玉米赤霉醇1諾龍硝基咪唑類地美硝唑丙酸睪酮1異丙硝唑睪酮1甲硝唑22SN/T3235—2012序號(hào)測(cè)定低限序號(hào)測(cè)定低限硝基咪唑類洛硝噠唑鎮(zhèn)靜劑類地西泮羥甲基甲硝咪唑氯霉素氯霉素雌性激素類己二烯雌酚1三苯甲烷類孔雀石綠己烯雌酚1隱性孔雀石綠己烷雌酚1結(jié)晶紫鎮(zhèn)靜劑類氯丙嗪隱性結(jié)晶紫化合物添加濃度回收率范圍%化合物添加濃度回收率范圍%beclomethasonediptopionate70.3～84.1異丙硝唑ipronidazole92～105.2290.5～104.5188.9～104492.25～103.75286.5～92.5倍他米松betanethasone95.6～108.8methandienone183.4～102197.5～113294～104.5293.5～116489.25～108.75去氫睪酮boldenone196～112甲硝唑metronidazole72.4～114288～112190.4～113497.75～114.5278.5～115氯丙嗪chlorpromazine81.6～100.4諾龍nandrolone195.6～114183～111293.5～112.5284～1004106.75～118.5西瑪特羅cimaterol75.6～118ractopamine91.6～103.8178.6～114187.4～103278.5～119291.5～99.5克侖特羅clentuterol98.2～110.6洛硝噠唑ronidazole73.4～103.6192.6～99.9192.1～117293～100.5291.5～118.5地西泮diazepam63.8～99.8沙丁胺醇salbutnol80.8～112.8183.3～113164.3～104272～110265.5～103地美硝唑dimetronidazole67.4～100.4特布他林terbutaline75～107.2169.7～86.8176.8～117276.5～102298～111表睪酮epiandrosterone189.2～111睪酮testosterone184.7～98.32100～111.5291～1094103.5～108.5499～109非諾特羅fenoterol183.1～98妥布特羅tobututerol70～118.2285.2～96.31481.6～99.2299.5～104化合物添加濃度回收率范圍%化合物添加濃度xg/kg回收率范圍%曲安西龍triamcinolonel70～115.8α-玉米赤霉烯醇169.6～99.1197.3～117285～110.5282～113.54100～111.5曲安奈德triamcinoloneacetonide189.5～116β-玉米赤霉烯醇188.9～102297118-524102～114.75491～110.25己二烯雌酚dienestrol1zearalanone172～92.12104～117.5289～112.5490.3～114492.25～117.25己烯雌酚diethylstilbestrol176A～108氯霉素77.6～106266.5～99.294.2～116476.8～106.25chloramphenicol95.5～124.5己烷雌酚hexestrol163.9～91.2α-玉米赤霉醇Q-zeranol191.5～110271.5～1122480~118.75499.5～118.25玉米赤霉烯酮zearalenone72.8～95.6β-玉米赤霉醇189.5～106182.7～1012291～118.5496～114.25化合物添加濃度回收率范圍%化合物添加濃度可收率范圍%倍他米松betanethasone1克侖特羅21498.1～104272.5～78.5去氫睪酮boldenone1地西泮282～97.51482.3～95.52氯丙嗪地美硝唑11287～95.52461.8～74.5西碼特羅表睪酮1182.1～10722化合物添加濃度回收率范圍%化合物添加濃度回收率范圍%氟氫可的松fiudrocortisone176.8～93.1曲安西龍triamcinolonel85.8～102276.4～99.5274.5～98.5485.5～101463.3～96.8異丙硝唑ipronidazole94.2～102曲安奈德triameinolone176.5～93.3183,8~94.9273.5～85acetonide289～98466.5～85.8甲睪酮methyltestosterone88.6～106玉米赤霉烯酮zearalénone182.3～117292～104275～109493.7～12472～117甲硝唑metronidazole187.3～91.9α-玉米赤霉烯醇1285.5～101283.3～118478.3～94.5484～119諾龍nandrolone191～115β-玉米赤霉烯醇179.5～116299.5～105270.5～119 474.5～93.3485～95.5玉米赤霉酮zearalanone99.6～115萊克多巴胺ractopamine188.2～111198.8～108285.1～93.794.5～98.5氯霉素chlofamphenicol181～119洛硝噠唑ronidazole85.1～6.1281～96.5484.8～113290.6～105α-玉米赤霉醇1沙丁胺醇salbutnol76～106283～108471～84170.1～88β-玉米赤霍醇81.5～109280.5～95.5171.2～107特布他林terbutaline96.8～115292～108196.1～118玉米赤霉烯酮zearalenone87.2～112286～104睪酮testosterone175.4～93.769～90α-玉米赤霉烯醇170～103283～992491.5～1184妥布特羅tobututerolβ-玉米赤霉烯醇179.7～126195～119269.3～1052479.1～119化合物添加濃度回收率范圍%化合物添加濃度回收率范圍%beclomethasonedipropionate179.2～98.8methandienone188.1～104287.5～101295.5～106489.8～98.5496.1～98.1倍他米松betanethasone92.4～107甲硝唑metronidazole94.4～102199.1～113192.8～97.522去氫睪酮boldenone179.9～110諾龍nandrolong1294.1～118244氯丙嗪chlorpromazine98.2～116ractopamine97.6～115186.3～111188.6～1032286.5～116西瑪特羅cimaterol85.1～104洛硝噠唑ronidazole86.1～102190.7～117170.2～115297～1132克侖特羅clentuterol86.4～103沙丁胺醇salbutnol86.4～111190.1～107186.5～96.1290～106286.5～92.5地西泮diazepam87.8～99.2特布他林terbutaline84.1～88.681.5～92.7190.9～96.6297.1～110288.5～93.1地美硝唑dimetronidazole91.6～101睪酮testosterone189.7～98.2199.6～10622499.5～109表睪酮epiandrosterone妥布特羅tobututerol89.8～1032190.2～101499.8～1142菲諾特羅fenoterol176.4～93.1曲安西龍triamcinolonel86.3～102284.5～99.590.2～106486～99.52異丙硝唑ipronidazole88.4～105曲安奈德triamcinoloneacetonidel91.9～116194.2～102295.1～106299.5～114485.5～10227SN/T3235—2012化合物添加濃度回收率范圍%化合物回收率范圍%己二烯雌酚dienestrol185.4～112β-玉米赤霉烯醇199.1～116278.1～97.52467.5～107493.2～113己烯雌酚diethylstilbestrol玉米赤霉酮zearalanonel66.8～83.9275.2～119280.5～117489.2～114467.8～100己烷雌酚hexestrol165.1～94.5氯霉素chloramphenicol0.0570.8～88.1276.1～11481.1～97.6470.1～11386.1～99.1玉米赤霉烯酮zearalenone90.2～104α-玉米赤霉醇171.7～99.1178.1～96.8294.5～110296.1～120480.2～116α-玉米赤霉烯醇175.1～87.1β-玉米赤霉醇181.5～101280.5～117293.1～114484.2～108485.2～111化合物添加濃度回收率范圍%化合物添加濃度回收率范圍%丙酸倍氯米松beclomethasone1克侖特羅90～116.62195.1～106483.25～117.752倍他米松betanethasone地西泮98.2～107.41122去氫睪酮boldenone1地美硝唑dimetronidazole82.6～97.82164.7～72.9494.75～113.5260～78.75氯丙嗪71.4～113.8表睪酮1173.6～89.5291.75～108.2524西瑪特羅非諾特羅176.3～89.1180.9～92.6224化合物添加濃度回收率范圍%化合物添加濃度回收率范圍%異丙硝唑ipronidazole91～109.4曲安奈德triamcinoloneacetonide174.3～84.5193.8～107293.5～116285.5～103.75496.75～113methandienone71.2～110dienestrok-189.4～104279.5～100.25283.5～108.54101～113.754100.2～115.75甲硝唑metronidazole88.4～103.8己烯雌酚diethylstilbestrol77.5～1101-79:7～105289.5～108.5260.5～104.75499.75～117.75諾龍nandrolone己烷雌酚hexestroll65～97.5292.4～108263.25～118.254101～116.75474.75～102.25ractopamine99.2～111.8zearalenone91.4～10790～93.995.7～117280～1082104～117.5洛硝噠唑ronidazole85.6～112.8188.5～11984.9～105280.5～99.5284.5～104.25467～79.75沙丁胺醇salbutnol93.2～109.6165.1～105195～105269～106289.5～107467.5～77特布他林terbutaline103～118.2zearalanone83.1～95.299.3～110286.1～106299.5～117492.3～105睪酮testosterone92～118氯霉素chloramphenicol67.2～88291.5～11081.5～95.44100～114.7593.5～110妥布特羅tobututerol92.8～103.887～11796.7～111289～106284～115488～104曲安西龍triamcinolonel93.8～114.6171.8～79.196.2～119265～107.5291.5～114458.2～78.5化合物添加濃度回收率范圍%化合物添加濃度回收率范圍%丙酸倍氯米松beclomethasone diptopionate189.7～103菲諾特羅fenoterol78.2～101281～100.586.1～103480.25～98.75279.2～105倍他米松betanethasone102.2～114.6氟氫可的松fludro-cortisone185.6～112195.1～101273.5～99.5273～96.5482.5～93.5去氫睪酮boldenone93.8～108.2氟甲睪酮fluoxymerostero182.9～1131-80~~101-293～108.5278.5～100.5478.5～94.75氯丙嗪chlorpromazine101.4～109.6羥甲基甲硝咪唑HMMNI93.7～101193.2～104283.5～106287～97480.75～90.75西瑪特羅cimaterol61～94.6氫化可的松hydrocortisone170.9～1101276.5～110.5276.5～90.5486.25～103.5克侖特羅clentuterol99.6～110異丙硝唑ipronidazole106～117.8193.1～101l94.1～118294～99.5286.5～105.5醋酸可的松corisoneacetate180.4～107隱性結(jié)晶紫leucocrystalviolet68.2～110.42182.7～114482.5-95.25290.1～119結(jié)晶紫crystalviolet100～119.8隱性孔雀石綠leucomalachitegreen102～119.8185.3～105182.8～98282～108.5277.5～97地西泮diazepam84～108去氫甲睪酮methandienone93.2～112.2183.5～92.1181～92.7276.5～107283.5～98.5地美硝唑dimetronidazole102～118.6甲睪酮methyltestosterone180.7～104195.5～109267.5～99.5291.5～111484～96.2表睪酮epiandrosterone94.2～110甲硝唑metronidazole70.6～106.6186～113169.6～82.3277～103.5269～95SN/T3235—2012表F.5(續(xù))化合物添加濃度回收率范圍%化合物添加濃度回收率范圍%孔雀石綠84.4～103.2曲安奈德1176.5～95.2283～111.52474.25～85.75丙酸諾龍nandrolone己二烯雌酚1122474.5～108.25諾龍nandrolone己烯雌酚diethylstilbestrol185.1～98.71263.5～108.524萊克多巴胺ractopamine己烷雌酚hexestrol171.8～95.5186.9～94.72287.5～106.54洛硝噠唑ronidazole玉米赤霉烯酮96.8～118.611290.5～99.52沙丁胺醇84.2～117.2α-玉米赤霉烯醇1181.6～91.62279.5～95.5特布他林β-玉米赤霉烯醇88.6～109.8168.2～91.3122丙酸諾龍nandrolone1玉米赤霉酮70.3～99.22176.2～96.34283.1～99.3睪酮1氯霉素281～105.5480.5～90.75妥布特羅α-玉米赤霉醇186.4～94.2186.1～11222曲安西龍1β-玉米赤霉醇12244SN/T3235—2012ThisstandardwasdraftedonthebasisoftherequirementofGB/T1.1—2009ThisstandardwasproposedbyandisunderthechargedofCertificationandAccreditationAdmini-stationofthePeople'sRepublicofChina.SN/T3235—2012Thisstandardspecifiesthemethodsofβ-agonist.androgensazoles,resorcylicacidlactones,triphenylmethanes,sedativesandchloramphenicolresiduesinfood-Thissrandardisadpedtodeterminationandscreeningofcimaterol,cimaterol,fenoterol,ractopamine,salbu-tamol,terbutaline,tulobuterol,boldenone,epiandrosterone,dehydro-17a-methyltestrosterone,nandrolone.zearalenone,α-zearalenol,β-zearalenol,zearalanone,α-zeranol.β-zeranol,dimetronidazole,ipronidazole,metronidazole,ronidazole,dienestroi,diethylstilbestrol,hexestrol,chlorpromazine.mol,terbutaline,tulobuterol,bofdenone,epjándrosterone,methyltestosterone,nandroloneipronidazole,metronidazole,ronicazole,dienestrol,diethylstilbestrol.hexestrol,chlorpromazine,di-ractopamine,salbutamol,terbutaline,tulobuterol,boldenone,epiandrosterone,fluoxymesterone,de-onepropionate,testostrone,beclomethasonedipropionate,betamethasone,cortisoneacetate,fiudro-cortisone,hydrocortisone,triamicinolone,triamcinoloneacetonide,zearalenone,α-zearalenol,βsample.2Normativereferencestuteprovisionsofthisstandard.Fordatedreferences,subsequentamendmentsto,orrevisionsoanyofthesepublicationsdonotapply.Howeverpartiestoagreementsbasedonthisstandardareen-couragedtoinvestigatethepossibilityofapplyingthemostrecenteditionsofthenormativedocu-mentsindicatedbelow.Forundatedreferences,thelatesteditionofthenormativedocumentreferredtoapplies.GB/T6682Waterforanalyticallaboratoryuse—specificationandtestmethods3.1Animalliver,muscleandaquaticproductAbout500grepresentativeediblesamplesshouldbetakenfromallsamples,thengrindedandblendedbyatissueblendertoproducehomogenoussamples,putinsuitablecleancontainers.Afterbeingsealedandlabeled,thesamplesshouldbestoredatbelow-18℃.About500mLrepresentativesamplesshouldbetakenfromallsamples,thenmixedtoproducehomogenoussamples,andputinsuitablecleancontainers.Afterbeingsealedandlabeled,thesamplesshouldbestoredatbelow-18℃4Principle5ReagentsandmaterialsUnlessotherwisespecified,allreagentsshouldbeofanalyticalgrade;“Water”isthefirstgradewaterprescribedbyGB/T6682—19925.1Acetonitrile:HPLCGrade.5.2Methanol:HPLCGrade.5.3Glacialaceticacid:HPLCGrade.5.4Ammonia:HPLCGrade.5.5Ammoniumacetate:HPLCGrade.5.7Glucuronidase/arylsulfatase:Containingglucuronidase134600U/mL,and5.8Primarysecondaryaminesorbent(PSA):Diameterrangeof40μm～60μmandaveragepore5.10Endcapped-C?gsorbent:diameterrangeof-40μm～60μmandaverageporesizeof60A.5.11Anhydroussodiumsulfate:analyticaiGrade.5.12Anhydrousmagnesiumsulfate:analyticalGrade.5.13Acetatebuffer:accuratelyweigh7.7gammoniumacetate(5.5)dissolvedin480mLwateradjustpHvalueto5.2byglacialaceticacid(5.3),dilutedwithwaterto500mL,andmixtohomoge.5.141%Ammonia/acetonitrilesolution:take40mL25%anmonia(5.4)and960mLacetonitrile(5.1),andmixtohomogeneity.5.151%aceticacid/acetonitrlesolution:take10mLaceticacid(5.3)and990mLacetonitrile(5.1)andmixtohomogeneity,5.1620%acetonitrile/watersolution:take200mLacetonitrile(5.1)and800mLwater,andthenmixtohomogeneity.5.170.1%formicacidsolution:dilute1mLformicacidto1000mLbywater,andmixtohomoge-5.180.1%formicacid/methanol:dilute1mLformicacidto1000mLbymethanol(5.2),andmixtohomogeneity.5.19QuEChERSsorbents1(appliedtoanimalliverandmusulesample):accuratelyweigh100mgofPSA(5.8),20mgofGCB(5.9),50mgofC??(5.10)and400mgofanhydrousmagnesi25mLcentrifugetube,andplacein25mLcentrifugetube,screwthecaptightlyforfurtherSN/T3235—20125.20QuEChERSsorbents2(appliedtomilkandaquaticproducts):accuratelyweigh100mgo5.21Standardchemicals:β-agonists,androgens,glucocorticoids,estrogens,nitroimidazoles,Resor-cylicacidlactones,chloramphenicol,Sedatives,malachitegreen,crystalvioletandtheirmetaboliteswereat99%purityorabove.5.22Intemalstandardchemicals:D?-salbutnol,ds-chlorpromazine,ds-ractopamine,dg-1eucocrystalviolet,d?-malachitegreen,dg-ctentuterot,dseronidazote,ds=crystaviolet,D?-leucomalachitegreen,D?-dienestrol,D?-chloramphenicol,DsdienestrolandD?-chloramphenicolwereat95%purityorabove.5.23Standardstocksolution:accuratelyweighconsiderableamountofstandardchemicalcompoundsof(5.21)dissolvedinmethanolanddilutetol0mL,thenmixtohomogeneity.Theco5.24Blanksamplesextracts:banksamplesextractisobtainedbypreparedsamplefolowingwith7.15.25Mixedstandardsolution:acgdratelymeasurequantitysufficientstandardstocksolution(5.23)diluteandatthetemperaturebelow4Cf5.26Mixedstandardworkingsolution:accordingtgpracticelmixedstandardworkingsolutionisdilutedwithblanksampleextract(5.24):5.27Internalstandardstocksolution;accurately/weighconsiclerableamountofstandardchemicalcom-poundsof(5.22)dissolvedinmethanolanddiuteto10m_,5.28Mixedinternalstandardsolution:accuratelymeasurequantitysufficientstandardstocksolutionservedindarkandatthetem5.29Mixedinternalstandardworkingsolution:accordingtopractice,mixedstandardworkingsolutionis5.30Milliporefiliter:0.22μm.7.1.1Milkandanimaltissues(liver,muscle)Accurately2gsampleweighandtransferredintoa50mLpolypropylenecentrifugetubewithscrew-cap,andspikedwith20μLinternalstandardsolution(5.28),andthenwereblendedbyahigh-speedblender.8mLammoniumacetatebuffer(5.13)wasadded,andsampleswerehomogenizedbyvigor-ouslyshakingandvortexfor1min.30.0μLglucuronidase/arylsulfatase(5.9)wasaddedandincuba-tedinthermostaticwater-bathshakerat37℃for12h.Aftercoolingtoroomtemperature,5.0gofanhydroussodiumsulfate(5.11)and15.0mL1%ammonia/acetonitrilesolutionwereadded,andthenhomogenizedfor30secathighspeed,themixturewascentrifugedat4℃for5minat9500r/min.Transferthesupernatanttoanotherclean50mLcentrifugetube.15mLof1%aceticacid/aceSN/T3235—20127.1.2Aquaticproduct(Accurately2.0gsampleweighandtransferredintoa50mLpolypropylenecentrifugetubewithahigh-speedblender.8mLammoniumacetatebuffer(pH5.2)wasadded,andthesamplewerehomogenizedbyvigorouslyshakingandvortexfor1min,andthenwasplacedatroomtemperatureforrTheQuEChERSsorbets1wasaddedintothemilkandanim20%acetonitrile/watersolution(5.16)byvortexingandultrasonic,andthedilutionwasalsofilteredwitha0.22μmfilter(5.30)forfurtheranalysis.Column:C??chroMobilephase:A1:0.1%formicacidsolution;B1:methanol;A2:acetonitrile,B2:water.SN/T3235—2012gradientelutionprocedurelistintable1.Table1—Gradientelutionprocedureforpositiveandnegativemode.Positive(ESI+)Negative(ESI-)TimeminA1%B1%TimeminA2%B2%4.0045.07.3.2MSConditionslonizationmode:ESI;postive/negativeacquirationmode.Determinationmode:MRM(retentionfimescheduled)Resolution:unitresolution(seetableB.1)OtherreferencemasscorditionsarelistedtableC.17.3.3HPLC-MS/MSdeterminationAccordingtotheapproximatecorcentratiorofbanneddrugresiduesinthetestsamplesolution,thestandardworkingsolutionisselectedwithsimilarresponsetothatofsamplesolution.Theresponseofbanneddrugresiduesinthestandardworkingsolútionandsamplésolutionshouldbeinthelinearrangeoftheinstrumentalresponse.Thestandardworkingsdutionshouldbeinjectedrandomlyinbetweentheinjectionsofsamplesolutionofequalvolume.Schedulemulti-reactionmonitor(MRM)chromatogramandretentiontimeof44standardcompoundssolutioncanbefoundFigureD.1Iftheconcentrationexceedstherangeofcalibrationcurve,itisnecessarilytodilutetheanalytetoappropriateconcentrationbymethanol/water(1/9,v/v).Underoptimalconditions,thechromatogramofeachcompoundisgiveninfigureD.2,andtheretentiontimecanreferencetableC.1(Itissuggestedthatthepositiveresultsdetectedbythismethodneedanothermethodtoconfirmandquantifythesubstance).Exceptforspikedsample,thesampleispretreatedaccordingtotheaboveprocedure.SN/T3235—2012Belativeiorintensity/%>20～50Permittedtolerances/%士25formula(1).Where40SN/T3235—20129LimitofdeterminationanThelimitdeterminationofthismethodinmilk,liver,chicken,porkandfishwerelistintableE.9.2RecoveryRecoveryrangesofeachcompoundindifferentmatrixislistedCtableF.1～tableF.5.SN/T3235—2012CompoundCASNoStructuralMolecularweightclenbuterol37148-27-9C??H??C??N,O277.19salbutamol18559-94-9C??H??NO?239.31ractopamine97825-25-7C1gH??NO?301.38cimaterol54239-37-1C??H??N?O219.28terbutaline23031-25-6C??H??NO?225.29tulobuterol41570-61-0Ci?H??CINO227.73fenoterol13392-18-2C??H??NO?303.35Androgens(10)boldenone846-48-0C?gH??O?286.41testosteronepropionate57-85-2C??H??O?344.49dehydro-17a-methyltestrosterone72-63-9C?oH?aO?300.44methyltestosterone58-18-4C??H?oO?302.45testostrone58-22-0C?gH??O?288.43nandrolone17-propionate7207-92-3C??H?oO?330.46epiandrosterone481-30-1C?gH?8O?288.42fluoxymesterone76-43-7CnH?oFO336.44nandrolone434-22-0C??H??O?274.40nandrolonephenylpropionate62-90-8C??H??O?406.56Glucocorticoids(6)betamethasone378-44-9C??H??FO392.45cortisoneacetate50-04-4C??H??O?402.49beclomethasonedipropionate4419-39-0C??H??CIO?408.92hydrocortisone50-23-7C??H??FO?362.47triamicinolone124-94-7C2?H??O?394.44triamcinoloneacetonide76-25-5C??H??FO?434.5Estrogens(3)diethylstilbestrol56-53-1C??H??O?268.35hexestrol84-16-2CgH??O?270.37dienestrol84-17-3C1aH??O?266.3342SN/T3235—2012TableA.1(continued)CompoundMolecularweightNitroimidazoles(5)1-methyl-5-nitro-1H-imidazoleC?H?N?O?dimetronidazoleC?H?N?O?ronidazolemetronidazoleC?H?N?O?C?H?N?O?C?8H??O?C??H?6O?Ci?H??O?CigH?O?C?gH?aO?Triphenylmethanes(4)CH??N?OC??H?6N?C??H?oN?CISedatives(2)C?HgCIN?SC??H??CIN?OChloramphenicol(1)chloramphenicolCnH??C??N?O43ReferencemassspectrometryconditionsforAPl50ReferencemassspectrometryconditionsPositivemode-4500CurtaingaspressureNebulizergaspressureAuxiliarygaspressureTemperatureAnnexC(informativeanne