SN∕T 1594-2019 出口茶叶及代用茶中噻嗪酮残留量的测定(出入境检验检疫).pdf

以正式出版文本为准中华人民共和国出入境检验检疫行业标准SN/T 15942019代替 SN/T 15942005出口茶叶及代用茶中噻嗪酮残留量的测定Determination of buprofezin residue in tea and herbal tea for export2019-12-27 发布2020-07-01 实施ICS 67.050C 53中华人民共和国海关总署发 布以正式出版文本为准以正式出版文本为准SN/T 15942019I前 言本标准依据 GB/T 1.12009 给出的规则起草。本标准代替 SN/T 15942005进出口茶叶中噻嗪酮残留量检验方法 气相色谱法。本标准与 SN/T 15942005 的主要差异如下：修改标准名称；扩大标准适用范围；修改气相色谱法前处理及仪器条件；增加液相色谱-质谱/质谱法。请注意本文件的某些内容可能涉及专利。本文件的发布机构不承担识别这些专利的责任。本标准由中华人民共和国海关总署提出并归口。本标准起草单位：中华人民共和国杭州海关。本标准主要起草人：朱晓雨，黄超群，崔晓美，楼成杰，吴娟，陈丽，蒋沁婷，方群。本标准所代替标准的历次版本发布情况为：SN/T 15942005。以正式出版文本为准以正式出版文本为准1SN/T 15942019出口茶叶及代用茶中噻嗪酮残留量的测定1 范围本标准规定了出口茶叶及代用茶中噻嗪酮残留量测定的液相色谱-质谱/质谱和气相色谱检测方法。本标准适用于出口绿茶、红茶、普洱茶、乌龙茶、牛蒡茶、菊花茶、杜仲茶和麦茶中噻嗪酮残留量的测定。2规范性引用文件下列文件对于本文件的应用是必不可少的。凡是注日期的引用文件，仅所注日期的版本适用于本文件。凡是不注日期的引用文件，其最新版本（包括所有的修改单）适用于本文件。GB/T 6682 分析实验室用水规格和试验方法第一法 液相色谱-质谱/质谱法3方法提要试样中噻嗪酮经丙酮、水提取，用正己烷抽提，经弗罗里硅土柱净化后，液相色谱-质谱/质谱法测定，外标法定量。4试剂和材料除特殊注明外，所用试剂均为色谱纯，所用水为符合 GB/T 6682 规定的一级水。4.1丙酮。4.2正己烷。4.3无水乙醚。4.4甲醇。4.5氯化钠：分析纯。4.6乙酸铵。4.7甲酸。4.8正己烷-乙醚（6+4，V/V）：取 600 mL 正己烷和 400 mL 无水乙醚混合均匀。4.9乙酸铵-甲酸混合溶液：准确称取 0.38 g 乙酸铵用水溶解，加入 1.0 mL 甲酸，用水定容至 1 000 mL。4.10标准物质：噻嗪酮，C16H23N3OS，CAS 登录号 69327-76-0，纯度大于 99.0%。4.11标准储备液：准确称取适量标准物质（4.10），用甲醇溶解并定容，配制成溶液浓度为 200 g/mL的标准储备液，于 0 4 储存，有效期 6 个月。4.12标准中间液：准确移取标准储备液（4.11）0.50 mL，用甲醇稀释并定容至 10 mL，配制成溶液浓度为 10 g/mL 的标准中间液。准确移取 10 g/mL 的标准中间液 1.0 mL，用甲醇稀释并定容至10 mL，配制成溶液浓度为 1.0 g/mL 的标准中间液。以正式出版文本为准2SN/T 159420194.13标准工作液：准确移取 1.0 g/mL 的标准中间液（4.12）1.0 mL，用甲醇稀释并定容至 50 mL，配制成溶液浓度为 20 ng/mL 的标准工作液。分别准确移取 20 ng/mL 的标准工作液 0.25 mL、0.50 mL、1.0 mL、2.5 mL 和 5.0 mL，用甲醇稀释并定容至 10 mL，配制成溶液浓度为 0.50ng/mL、1.0ng/mL、2.0ng/mL、5.0ng/mL 和 10 ng/mL 的标准工作液，于 04 储存，有效期 3 个月。4.14弗罗里硅土固相萃取柱：1 g 或相当者，使用前用 5 mL 正己烷预洗。4.15微孔滤膜：0.45 m，有机相。5仪器和设备5.1液相色谱-质谱/质谱仪：配有电喷雾离子源。5.2分析天平：感量 0.000 1 g 和 0.01 g。5.3涡旋混合器。5.4均质器。5.5离心机：最大转速 4 000 r/min。5.6旋转蒸发仪。5.7固相萃取仪。6试样制备与保存6.1试样制备取代表性样品约 500 g，用粉碎机粉碎并通过 2.0 mm 圆孔筛，混匀，装入洁净的盛样容器中，密封并标明标记。6.2试样保存试样于室温下保存。在制样的操作过程中，应防止样品受到污染或发生残留物含量的变化。7测定步骤7.1提取准确称取 2 g 试样（精确至 0.01 g），置于 50 mL 塑料离心管中，加入 10 mL 水，浸泡 2 h，再加入 20 mL 丙酮，在均质器中均质 2 min，加入 4 g 氯化钠，再加入 15 mL 正己烷，涡旋提取 1 min，以 4 000 r/min 离心 5 min，将上清液收集于浓缩瓶中，残渣再加入 15 mL 正己烷，重复上述操作，合并提取液，在 45 以下水浴减压浓缩至近干，待净化。7.2净化准确移取 10 mL 正己烷溶解残渣。取 1.0 mL 转移至弗罗里硅土固相萃取柱（4.14）内。再用10 mL 正己烷-乙醚混合溶液（4.8）洗脱，收集全部洗脱液，45 以下水浴减压浓缩至干，加入 2.0 mL甲醇溶解定容，经滤膜过滤，供液相色谱-质谱/质谱仪测定。7.3测定7.3.1液相色谱-质谱/质谱条件液相色谱-质谱/质谱条件如下：以正式出版文本为准3SN/T 15942019a）色谱柱：C18，100 mm2.1mm（内径），3.5 m，或相当者；b）流动相：乙酸铵-甲酸混合溶液（4.9）和甲醇，梯度洗脱程序见表 1；表 1液相色谱-质谱/质谱法梯度洗脱程序时间/min乙酸铵-甲酸混合溶液/%甲醇/%0.0070303.0070304.00208011.00208012.00703015.0070307.3.2定量测定按 7.3.1 仪器条件测定标准工作溶液和样液，以外标曲线法计算样液中的噻嗪酮含量。如果样液中噻嗪酮的含量超出标准曲线范围，应用甲醇稀释后再进行分析。在上述条件下，噻嗪酮的参考保留时间约为 10.0 min。标准溶液多反应监测色谱图参见附录 B 中图 B.1。7.3.3定性测定在上述液相色谱-质谱/质谱条件下，如果样液与标准工作液中待测物质色谱峰相对保留时间偏差在 2.5范围内；定性离子对的相对丰度与浓度相当的标准工作溶液的相对丰度一致，相对丰度允许误差不超过表 2 规定的范围，则可判断样品中存在相应的被测物。表 2 相对离子丰度最大容许误差相对离子丰度（基峰）%50 2050 1020 10最大容许误差%202530507.4空白试验除不加试样外，均按上述操作步骤进行。8结果计算和表述用色谱数据处理机或按公式（1）计算试样中噻嗪酮的残留量：1 0001 000c VXm=（1）式中：X 试样中噻嗪酮的残留量，单位为毫克每千克（mg/kg）；c 从标准曲线上得到的待测物质的溶液浓度，单位为微克每毫升（g/mL）；V 样液最终定容体积，单位为毫升（mL）；以正式出版文本为准4SN/T 15942019m 最终样液所代表的试样质量，单位为克（g）；计算结果应扣除空白值。9定量限和回收率9.1定量限本方法对于绿茶、红茶、普洱茶、乌龙茶、牛蒡茶、菊花茶、杜仲茶和麦茶中噻嗪酮的定量限均为 0.005 mg/kg。9.2回收率本方法的添加水平和回收率数据见表 3。表 3 茶叶中噻嗪酮添加水平及回收率数据添加水平mg/kg回收率范围/%绿茶红茶普洱茶乌龙茶牛蒡茶菊花茶杜仲茶麦茶0.005841047486921067610286104901048698841060.01768973978210886101729282101829770980.05809492102801048411076948810480108881040.2091102861078899831059010483103849092107308096749184100859674927690921047798第二法 气相色谱法10方法提要试样中噻嗪酮经丙酮、水提取，用正己烷抽提，经弗罗里硅土柱净化后，用配有氮磷检测器的气相色谱仪测定，外标法定量。11试剂和材料除特殊注明外，所用试剂均为色谱纯，所用水为符合 GB/T 6682 规定的一级水。11.1丙酮。11.2正己烷。11.3无水乙醚。11.4氯化钠：分析纯；11.5正己烷-乙醚（7+3，V/V）：取 700 mL 正己烷和 300 mL 无水乙醚混合均匀。11.6标准物质：噻嗪酮，C16H23N3OS，CAS 登录号 69327-76-0，纯度大于 99.0%。11.7标准储备液：准确称取适量标准物质（11.6），用正己烷溶解并定容，配制成溶液浓度为 200 g/mL 的标准储备液，于 04 储存，有效期 6 个月。11.8标准中间液：准确移取标准储备液（11.7）0.50 mL，用正己烷稀释并定容至 10 mL，配制成以正式出版文本为准5SN/T 15942019溶液浓度为 10 g/mL 的标准中间液。11.9标准工作液：准确移取标准中间液（11.8）1.0 mL，用正己烷稀释并定容至 50 mL，配制成溶液浓度为 0.20 g/mL 的标准工作液。分别准确移取 0.20 g/mL 的标准工作液 0.50 mL、1.0 mL、2.5 mL 和 5.0 mL，用正己烷稀释并定容至 10 mL，配制成溶液浓度为 0.010g/mL、0.020g/mL、0.050g/mL 和 0.10 g/mL 的标准工作液，于 0 4 储存，有效期 3 个月。11.10弗罗里硅土固相萃取柱：5 g 或相当者，使用前用 10 mL 正己烷预洗。12仪器和设备除特殊注明外，所用仪器和设备同 5.25.7。12.1气相色谱仪：配有氮磷检测器。13试样制备与保存试样制备与保存同 6.1 和 6.2。14测定步骤14.1提取提取同 7.1。14.2净化用 2 mL 正己烷溶解残渣，将溶液转移至弗罗里硅土柱（11.10）内，再用 2 mL 正己烷洗涤残渣合并上柱。用 20 mL 正己烷-乙醚混合溶液（11.5）洗脱，收集全部洗脱液，45 以下水浴减压浓缩至干，加入 2.0 mL 正己烷溶解，供气相色谱仪测定。14.3测定14.3.1气相色谱条件气相色谱条件如下：a）色谱柱：SPB-50，30 m0.25 mm0.25 m，或相当者；b）柱温：60 保持 1 min，以 15/min 的速率升温至 150，保持 1 min，随后以 10/min的速率升温至 280，保持 15 min；c）进样口温度：250；d）载气和尾吹气：氮气，纯度 99.999%，流量 3.0 mL/min；尾吹气流量 10 mL/min；空气流量 60 mL/min；氢气流量 3.0 mL/min；e）进样量：2 L；f）进样方式：不分流进样；g）检测器温度：325；h）开阀时间：0.75 min；i）检测器铷珠的电压和灵敏度应调整至最佳状态，为保护铷珠，进样 5 min 内关闭检测器。以正式出版文本为准6SN/T 1594201914.3.2色谱测定按 14.3.1 仪器条件测定标准工作溶液和样液，以外标曲线法计算样液中的噻嗪酮含量。如果样液中噻嗪酮的含量超出标准曲线范围，应用正己烷稀释后再进行分析。在上述条件下，噻嗪酮的参考保留时间约为 21.2 min。标准溶液色谱图参见附录 B 中图 B.2。14.4空白试验除不加试样外，均按上述操作步骤进行。15结果计算和表述同 8。16定量限和回收率16.1定量限本方法对于绿茶、红茶、普洱茶、乌龙茶、牛蒡茶、菊花茶、杜仲茶和麦茶中噻嗪酮的定量限均为 0.01 mg/kg。16.2回收率本方法的添加水平和回收率数据见表 4。表 4 茶叶中噻嗪酮添加水平及回收率数据添加水平mg/kg回收率范围/%绿茶红茶普洱茶乌龙茶牛蒡茶菊花茶杜仲茶麦茶0.01729074938110086978695831018310083980.058694861048498921108010886988810480980.209010486101901038310682100891079210589106307291899591103859887101779673968797以正式出版文本为准7SN/T 15942019附 录 A（资料性附录）a液相色谱-质谱/质谱法仪器参数与监测离子1）A.1 液相色谱-质谱/质谱法仪器参数如下：a）电喷雾电压：3 500 V；b）离子源温度：300 ；c）鞘气压力：50 Arb；d）辅助气压力：20 Arb；e）毛细管温度：270 ；f）碰撞气压力：1.5 mTorr；g）鞘气、辅助气均为高纯氮气；碰撞气为高纯氩气；使用前应调节各参数使质谱灵敏度达到检测要求；h）监测离子对信息、碰撞能量等参数见表 A.1。表 A.1 多反应监测离子对和碰撞能量化合物母离子 m/z子离子 m/z碰撞能量（V）噻嗪酮306.0201.1*10106.032 注：带*为定量离子1）非商业性声明：附录 A 所列参考质谱条件是在 TSQ Quantum ULTRA AM 液质联用仪上完成，此处列出试验用仪器型号仅是为了提供参考，并不涉及商业目的，鼓励标准使用者尝试不同厂家或型号的仪器。以正式出版文本为准8SN/T 15942019附 录 B（资料性附录）气相色谱法和液相色谱-质谱/质谱法标准溶液色谱图图 B.1 噻嗪酮标准溶液的多反应监测（MRM）色谱图（0.50 ng/mL）图 B.2 噻嗪酮标准溶液的气相色谱图（0.01 g/mL）以正式出版文本为准9SN/T 15942019ForewordThis standard was drafted in accordance with the GB/T 1.12009.This standard is the revision of SN/T 15942005Inspection of buprofezin residue in tea for import and export-Gas chromatographic method.The differences between this standard and SN/T 1594-2005 are as follows：Changed the name of standard；Extended the scope；Amended the pretreatment and instrument condition of GC method；Added LC-MS/MS method.Please pay attention that some contents in this standard may refer to patents.The institution doesnt take on the responsibility to identify these patents.This standard is proposed by and is under the charge of General Administration of Customs of the Peoples Republic of China.This standard was drafted by Hangzhou Customs District of the Peoples Republic of China.The main drafters of this standard are Zhu Xiaoyu，Huang Chaoqun，Cui Xiaomei，Lou Chengjie，Wu Juan，Chen Li，Jiang Qinting，Fang Qun.This standard replaces the standard previous published as：SN/T 15942005.以正式出版文本为准以正式出版文本为准11SN/T 15942019Determination of buprofezin residue in tea and herbal tea for export1 ScopeThis standard specifies the determination of buprofezin residue in tea and herbal tea by liquid chromatography-tandem mass spectrum and gas chromatography.This standard is applicable to the determination of buprofezin in green tea，black tea，puer tea，oolong tea，burdock tea，chrysanthemum tea，eucommiae tea and barley tea for export.2 Normative referencesThe following normative documents contain provisions which，through reference in this text，constitute provisions of this standard.For dated references，subsequent amendments to，or revisions of any of these publications do not apply.However parties to agreements based on this standard are encouraged to investigate the possibility of applying the most recent editions of the normative documents indicated below.For undated references，the latest edition of the normative document referred to applies.GB/T 6682 Water for analytical laboratory usespecification and test methodsMethod LC-MS/MS Method3 PrincipleThe test samples are extracted with water-acetone.The extract is partitioned with hexane，cleaned up by florisil solid phase and detected by liquid chromatography-tandem mass spectrometry using external standard method.4 Reagents and materialsUnless otherwise specified，all the reagents used should be chromatographic pure，“water”is distilled water.4.1 Acetone.4.2 Hexane.4.3 Diethyl ether.4.4 Methanol.4.5 Sodium chloride：analytical grade.4.6 Ammonium acetate.4.7 Formic acid.4.8 Hexane-diethyl ether（6+4，V/V）：Volume 600 mL hexane，then add 400 mL diethyl ether，mix them.4.9 Ammonium acetate-formic acid mixed solution：Accurately weigh 0.38 g，of ammonium acetate，dissolved with water，then added 1.0 mL of formic acid and diluted to 1000 mL with water.以正式出版文本为准12SN/T 159420194.10 Standard：Buprofezin，C16H23N3OS，CAS NO.69327-76-0，Purity 99.0%.4.11 Standard stock solution：Accurately weigh appropriate standard（4.10），dissolved with methanol，the concentration of solution is 200 g/mL.Stored under 04 and stable for 6 months.4.12 Standard intermediate solution：Accurately transfer 0.50 mL of standard stock solution，diluted to 10 mL with methanol.The concentration of solution is 10 g/mL.Then accurately transfer 1.0 mL of this solution，diluted to 10 mL with methanol.The concentration of standard intermediate solution is 1.0 g/mL.4.13 Standard work solution：Accurately transfer 1.0 mL of standard intermediate solution，diluted to 50 mL with methanol.The concentration of standard work solution is 20 ng/mL.Accurately transfer 0.25 mL，0.50 mL，1.0 mL，2.5 mL and 5.0 mL of the solution which concentration is 20 ng/mL，diluted to 10 mL with methanol.The concentrations of standard work solutions are 0.50 ng/mL，1.0 ng/mL，2.0 ng/mL，5.0 ng/mL and 10 ng/mL.Stored under 0 4 and stable for 3 months.4.14 Florisil SPE column：1 g，preconditioned with 5 mL of hexane before use.4.15 Millipore filter：0.45 m，organic phase.5 Apparatus and equipment5.1 Liquid chromatography tandem mass spectrometry，equipped with ESI.5.2 Electronic balance：accurate to 0.000 1 g and 0.01 g.5.3 Vortex mixer.5.4 Homogenizer.5.5 Centrifuge（4 000 r/min）.5.6 Rotary vacuum evaporator.5.7 Solid Phase Extraction System.6 Preparation and storage of test sample6.1 Preparation of test sampleTake approximately 500 g of representative sample，smashed thoroughly by a chopper，passed through a 2.0 mm sieve，and put into clean containers，which is sealed and labeled.6.2 Storage of test sampleTest samples should be stored at room temperature.In course of sampling and sample preparation，attention must be taken to avoid contamination or any factors which may cause the change of residue content.7 Procedure7.1 ExtractionWeigh 2 g（accurate to 0.01 g）of the test sample into a 50 mL polypropylene tube，10 mL of water was added.The sampled was soaked for 2 h.Then 20 mL of acetone was added.After homogenized for 2 minutes，4 g sodium chloride and 15 mL of hexane were added.the sample was mixed for 1 minute and centrifuged at 4 000 r/min for 5 minutes.The upper solution was transferred into a concentrate bottle，the residual was extracted with 15 mL of hexane again，the solution was transferred into the same concentrate bottle and condensed to nearly dry，waiting for cleaning-up.以正式出版文本为准13SN/T 159420197.2 Cleaning-upAdd 10 mL of hexane to dissolve the residual.1.0 mL of the solution was transfered to a florisil column（4.14）.After sample loading，buprofezin was eluted by 10 mL of hexane-diethyl ether（4.8）.The eluate was evaporated and redissolved with 2.0 mL of methanol for determination by LC-MS/MS.7.3 Determination7.3.1 LC-MS/MS operating conditionLC-MS/MS operating condition is as follows：a）Column：C18，100 mm 2.1 mm（i.d），3.5 m，or the equivalent；b）Mobile phase：Ammonium acetate-formic acid mixed solution and methanol，for gradient elute condition，see Table 1；Table 1 Gradient elute conditionTime/minAmmonium acetate-formic acid mixed solution/%Methanol/%0.0070303.0070304.00208011.00208012.00703015.007030c）Column temperature：20 ；d）Flow：0.25 mL/min；e）Injection volumn：10 L；f）ion source：ESI；g）Scan mode：positive；h）Detect mode：Multiple reaction monitoring（MRM）；i）Ion-pairs are listed in Table A.1 in Appendix A.7.3.2 Quantitation determinationAnalyze standard working standard solutions and samples with external standard method by LC-MS/MS，using conditions established in section 7.3.1.If the concentration of buprofezin exceeds the linear range of the calibration curve，the final solution should be diluted with methanol and reanalyzed.Under the above conditions，the retention time of chromatographic peak of buprofezin is about 10.0 min.For the MRM chromatogram refer to Fig.B.1 in Appendix B.7.3.3 Qualification confirmationThe relative intensities of the detected ions，expressed as a percentage of the intensity of the most intense ion or transition，shall correspond to those of the calibration standard，either from calibration standard solutions or from spiked samples，at comparable concentrations，measured under the same conditions，以正式出版文本为准14SN/T 15942019within the following tolerances：Table 2 Maximum allowance of relative ion abundanceRelative intensity（base peak）%50 2050 1020 10Maximum permitted tolerances%202530507.4 Blank testThe operation of the blank test is the same as that described in the method of determination，but with omission of sample addition.8 Calculation and expression of the resultCalculate the content of ethoxyquin in the test sample according to the following formula：1 0001 000c VXm=（1）Where：X the content of buprofezin in the test sample，mg/kg；c the concentration of buprofezin in sample solution calculated by standard curve，g/mL；V the final volume of the sample solution，mL；m the corresponding mass of the test sample in the final sample solution，g.The calculated result should be deducted from the blank value.9 Limit of quantitation and recovery9.1 Limit of quantitationThe limits of quantitation for buprofezin in green tea，black tea，puer tea，oolong tea，burdock tea，chrysanthemum tea，eucommiae tea and barley tea are 0.005 mg/kg.9.2 RecoveryThe spike levels and results of recovery test are listed in Table 3.Table 3 Results of recovery testSpike levelmg/kg回收率范围/%Green teaBlack teaPuer teaOolong teaBurdock teaChrysanthemum teaEucommiae teaBarley tea0.005841047486921067610286104901048698841060.01768973978210886101729282101829770980.05809492102801048411076948810480108881040.2091102861078899831059010483103849092107308096749184100859674927690921047798以正式出版文本为准15SN/T 15942019Method GC Method10 PrincipleThe test samples are extracted with water-acetone.The extract is partitioned with hexane，cleaned up by florisil solid phase and detected by gas chromatography with NPD.11 Reagents and materialsUnless otherwise specified，all the reagents used should be chromatographic pure，“water”is distilled water.11.1 Acetone.11.2 Hexane.11.3 Diethyl ether.11.4 Sodium chloride：analytical grade.11.5 Hexane-diethyl ether（7+3，V/V）：Volume 700 mL hexane，then add 300 mL diethyl ether，mix them.11.6 Standard：Buprofezin，C16H23N3OS，CAS NO.69327-76-0，Purity 99.0%.11.7 Standard stock solution：Accurately weigh appropriate standard（11.6），dissolved with hexane，the concentration of solution is 200 g/mL.Stored under 04 and stable for 6 months.11.8