1、华人共和出境I 事噩噩噩墨子了标SNj3699-2013 串串4种自囊相色i曾因留量的测定语/剂残谱gDetermination of four thiazole fungicide residues in food of plant origin for export一直PLC-MSjMSmet匾。d2013-11回06发布2014阳06-01实施。码至J。如o.5Y白白回回飞飞、由D协J1f烧四网运WNICf仙315(m也i.)4l刀693235中华人民共和匾发布圈家庭竞量监督撞验棱楚总爵SN/T 3699一2013萨=1本标准按照GB/T1.1-2009给出的规则编写。请注意本文件的某些
2、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。本标准由国家认证认可监督管理委员会提出并归口。本标准起草单位:中华人民共和国深圳出入境检验检疫局、深圳市检验检疫科学研究皖、中华人民共和国厦门出入境检验检疫局。本标准主要起草人:陈波、靳保辉、肖锋、谢丽琪、陈沛金、徐敦明、周里。I 1 范围出口植物源食晶中4种曝睡类杀菌荆残留量的测定液相色i曾国质谱/质谱法SN/T 3699-2013 本标准规定了植物源性食品中土菌灵、I嚣哇菌胶、辛睦自目和苯I黯硫氧的液相色i普白质谱/质i普测定和确证方法。本标准适用于大豆、玉米、小麦氧的测定。2 规黯性引用文件GB/T 6682 3 方法提要标法定
3、量。4 试剂和材料除另有说明外，所用试剂均4.1 甲醇:高效液相色谱级。4.2 乙睛:高效液相色谱级。4.3 甲苯:高效液相色谱级。4.4 甲酸:高效液相色谱级。4.5 氧化铀。4.6 醋酸镀:高纯试剂，99.999%。、喔瞠菌胶、辛i噩酣和苯I噩硫色i普白质i普/质i普检测和确证，外4.7 乙膳+水溶液(3+2，体积比):量取60mL乙腊(4.2)，加入40mL水，混匀。4.8 乙睛甲苯溶液(3十1，体积比):量取150mL乙腊(4.2)，加入50mL甲苯(4.3)，泪匀。4.9 5 mmol/L醋酸镀水溶液(含0.05%甲酸):称取0.385g醋酸镀(4.的，加入0.5mL甲酸(4.的，用
4、水溶解并定容至1L，混匀。4.10 标准物质:土菌灵、辛隆酣和苯睦硫氧，含量98%以上;睦瞠菌胶被度为10mg/L(相关化学信息见附录A中表A.l)。4.11 标准储备溶液:分别准确称取适量的土菌灵、辛喔酣和苯喔硫氧标准物质(4.10)，用乙腊(4.2)配制成浓度为1000mg/L的标准储备榕液;哇哇菌胶为甲薛溶液配制的10mg/L的标准溶液，一18oc以下避光保存。4.12 混合中间标准溶液:吸取适量的各标准储备溶液(4.11)，用乙蜻(4.2)稀释，配制成5.0mg/L的混SN/T 3699-2013 合中间标准溶液，-18C以下避光保存。4.13 基质混合标准工作溶破:根据样品中哇哇类杀
5、菌剂含量吸取适量的混合中间标准溶液(4.12)，用相应基质的空白样品提取液配成合适坡度的基质混合标准工作溶液，使用前配制。4.14 复合固相萃取柱(石墨化炭黑与氨基复合填料):6mL ,500 mg，或相当者。使用前用5mL乙脂甲苯溶液(4.8)活化，保持柱体湿润。4.15 C18固相萃取柱:6mL, 500 mg，或相当者。使用前用5mL乙腊(4.2)活化，保持柱体湿润。4.16 微孔滤膜:0.22m，有机相型。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(ESD。5.2 固相萃取装置。5.3 氮吹浓缩仪。5.4 均质器:转速不低于10000 r/min, 5.5 涡旋混匀器
6、。5.6 离心机:转速不低于5000 r/min, 5.7 分析天平:感量0.1mg和0.001g。5.8 带刻度聚丙烯离心管:15mL和50mL，具蠢。6 试样制备与保存6.1 通则在制样的操作过程中，应防止样品污染或发生残留物含量的变化。6.2 茶叶、小麦、大豆和玉米取代表性样品约500g，用粉碎机充分粉碎，混匀。试样均分为2份，装入洁净容器，密封，并标明标记。常温下保存。6.3 西瓜、葡萄、菜心和洋葱取代表性样品约500g，将其可食用部分(不可用水洗)切碎后，用捣碎机将样品加工成浆状，混匀。试样均分为2份，装入洁净容器，密封，并标明标记。于一18c以下冷冻存放。7 分析步蘸7.1 提取7
7、.1.1 茶叶、小麦、大豆和玉米称取约5g(精确至0.001g)样品于50mL具塞离心管中，加入15mL乙腊(4.2)，10000 r/min均质提取1min，于5000 r/min离心5min，将乙踌层转移至另一50mL具塞离心管中。残渣再用15mL 乙腊(4.2)重复均质提取l次，合并提取液，定容至30mL。转移10mL乙腊提取液至15mL离心管中，45C吹氮至约2mL，再转移5mL乙睛提取液至向一离心管中，45C吹氮至约2mL，待净化。2 SN/T 3699一20137. 1. 2 西瓜、葡萄、菜心和洋葱称取约10g(精确至0.001g)样品于50mL具塞离心管中，加入20mL乙腊(4.
8、2)，涡旋混匀后，10 000 r/min均质提取1mi丑，加入3g氧化铀(4.日，涡旋氓匀，使氧化铀充分溶解，于5000 r/min离心5囚m。转移上层乙睛提取液10mL至15mL离心管中，45oc吹氮至约2mL，待净化。7.2 净化7.2.1 大豆将待净化液全部转入C18固相萃取柱(4.1日，以约1.5mL/min的流速使样液通过固相萃取柱，再用3mL乙腊(4.2)洗脱并抽干固相萃取柱，收集全部流出液于15mL离心管中，45oc吹氮至近干，用乙腊溶液(4.7)定容至2mL，过微孔滤膜(4.16)，供液相色谱-质谱/质谱仪测定。7.2.2 玉米将待净化液全部转入C18固相萃取柱(4.1日，以
9、约1.5mL/min的流速使样液通过固相萃取柱，再用3mL乙腊(4.2)洗脱并抽干国相萃取柱，收集全部流出液于15mL离心管中，45oc吹氮至近干，用乙腊榕液(4.7)定容至2mL, 4 oc冷藏保存1h后，过微孔滤膜(4.16)，供液相色谱质谱/质谱仪测定。7.2.3 西瓜、南萄、菜心、小麦、茶叶和洋葱将待净化液全部转入复合国相萃取柱(4.14)，以约1.5mL/min的流速使样液通过固相萃取柱，再用5mL乙腊-甲苯溶液(4.8)洗脱并抽干固相萃取柱，收集全部流出液于15mL离心管中，45oc吹氮至近干，加入2mL乙腊(4.2)，涡旋混匀后再吹氮至近干，用乙腊洛液(4.7)定容至2mL，过微
10、孔滤膜(4.16) ，供液相色谱-质谱/质谱仪测定。7.3 甜定7.3.1 液相色谱条件液相色谱条件如下:a) 色谱柱:C18色谱柱，100mmX2.0 mm(内径)，3m，或相当者;b) 柱温:30oC; c) 进样量:10L; d) 流动相、流速及梯度洗脱条件见表1。表1液相色i普梯度洗脱条件日才l司流速5 mmol/L醋酸镀水溶液(含0.05%甲酸)立11口mL/min % 。40 3.0 40 5.0 5 0.30 9.0 5 9.1 40 16.0 40 甲醇% 60 60 95 95 60 60 3 SN/3699-2013 7.3.2 质谱条件质谱条件如下:a) 离子化模式:电喷
11、雾离子源;b) 扫描方式:正离子扫描;c) 检测方式:多反应监测(MRM);d) 分辨率:单位分辨率;其他参考质谱条件参见附录B中表B.L7.3.3 搜相色谱-串联质i普测定 一个川将7.3.3.1 定性测定主每种被测组分选择1个母离子，2个予离手，在一相同实验条件下，注品中待测物质的保留时间，与基:三i质标准溶液的保留时间偏差在:1:2.5%之内;样晶晶中中f各组分定性离手的相X对才丰度与浓度接近的基质1泪昆合标准工作溶液中对应的定性离子的相对丰度进行比较，偏差不1毡主过表2规定的范围，则可判定为样品中/ 存在对应的待测物。 / 表2定性确证时相对离子学酣歇件偏差50% 主二10%士20%士
12、50%图C.LJ 7.3.4 空白实验除不称取样品外，均按上述测定条件和步骤进行。8 结果计算和表述按式(1)计算试样中各喔嗖类杀菌剂的残留量，计算结果应扣除空白值:X;-iXVXn 1 000 m 1 000 . ( 1 ) 式中:Xi一一样品中曝睡类杀菌剂残留，单位为微克每千克(g/kg);Ci一一通过标准工作曲线计算所得进样溶液中哇哇类杀菌剂浓度，单位为微克每升(g/L); V一一样液最终定容体积，单位为毫升(mL);n一一稀释倍数;4 m一一最终样液代表的试样质量，单位为克(g)。9 方法的测定低限和回收率9.1 测定低限本方法的测定低限均为10.0g/峙。9.2 回收率样品的添加浓度
13、及回收率的数据参见附录D中表D.LSN/T 3699-2013 5 SN/3699-2013 附录A(规范性附录)曝瞠类杀菌剂的相关信息表A.l瞌瞌类杀菌剂的梧关信息名称结构图分子式相对分子质量CAS号苯I摩硫银F C,H6N2S3 238.34 21564-17-0 CBenthiazole) N圭子/飞S辛嚓酣气产Cll H19NOS 213.3 26530-20-1 C Octhilinone) 土菌灵，-/0(与2593-15-9 C5日5C13N20S247.5 CEtridiazole) C13 I骤瞠茵胶川t丁台CHH16N60S2 320.42 162650-77-3 CEth
14、aboxam) N 6 参考质谱条件:a) 离子源:电喷雾离子源;b) 扫描方式:正离子扫描;c) 检测方式:多反应监测CMRM);d) 电喷雾电压:5500 V; 巳)气帘气:68.95kPa; f) GSl : 275.8 kPa; g) GS2 :482.65 kPa; h) 脱溶剂气温度:600oC; 附录B(资料性附录)参考质谱条件SN/T 3699-2013 i) 定性离子对、定量离子对、采集时间、去簇电压CDP)、射入电压CEP)、碰撞电压CCE)、碰撞室射出电压CCXP)见表B.l。表13.1嘻瞠类杀菌剂测定的质i普参数参考保留母离子子离子采集时间CE DP EP CXP 化合
15、物日才间Cm/z) Cm/z) eV eV eV eV 盯15红Uil183.1持100 30 60 14 10 嚓瞠茵胶3.1 322.1 200.2 100 30 60 14 10 180.0祷100 9 20 10 10 苯l庭硫氨5.0 239.1 212.0 100 11 20 10 10 101.9铃100 14 100 10 15 辛暖翻8.1 214.1 71.1 100 20 100 10 9 218.9头100 16 35 10 10 土菌灵8.4 246.9 183.0 100 26 35 10 10 注:标头者为定量离子，对于不同质谱仪器，仪器参数可能存在差异，测定前应
16、将质谱参数优化到最佳。非商业性声明:附录B所列参考质谱条件是在API4000型液质联用仪上完成的，此处列出试验用仪器型号仅为提供参考，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。7 SN/3699-2013 附录C(资料性附录)标准多反应监制(MRM)色谱图8.43 8.44 ，咱iM申时闻部】土菌灵2 4 A哇uv nununu nu 可Anuphd M唱迦譬8.06 辛Ifi需2 4 6 6 8 10 12 14 时间/min42 酒也譬14 1. 8 10 12 14 时间/min图C.1浓度为10.0g/L幢幢类杀菌荆标准晶液相色i蕾质谱/质i普多反应监测色谱图8 化添加
17、葡萄西瓜i口L 物水平回收率范围回收率范围名mg/kg % % 称0.01 78.4100.5 70.098.。隆0.1 83.2102.7 80.394.2 膛茵1. 0 88.0107.2 87.7103.7 胶6.0 88.6104.。82.791.5 0.01 92.0112.。76.0108.。苯0.1 85.299.8 83.4109.0 嚓硫氛1.0 90.4109.2 84.3101.3 6.0 88.398.1 80.694.2 0.01 74.0103.。95.0115.0 辛0.1 88.8102.4 77.3102.7 隆酣1. 0 88.4104.0 88.2102.
18、7 6.0 89.6104.6 80.493.4 0.01 79.0103.。68.589.9 土0.1 83.4101.9 79.099.2 茵灵1.0 86.0100.8 76.299.6 6.0 83.8102.4 79.188.9 附录D(资料性附录)回收率表D.1回收率小麦回收率范围% 7L2.;S7.6 83.299.1 85.696.4 76.5 10.56 75941045丈8二6.299.01 非)6.6二也7.4:0.-95.2504094。8斤1:5 飞等v 飞气 .注菜心茶叶回收率范围回收率范围% % 88.0118.。70.590.5 79.8103.0 72.189
19、.6 80;-91.5 80.1-93.8 70.093.8 66.889.2 85.499.8 70.484.7 85.0101.0 80.6100.2 82.3-92.4 86.496.5 61注:主二=86.。73.3 .;93.5 83.9107:0 69.778.8 / 87.0,;,103.5 86.498.7 81.095.5 81.6-99.74 71.099.。70.095.。8219112.。67.883.8 83.5-97.8 82.O/ 92.8 83.8-99.9 80.手-90.6SN/3699-2013 洋葱大豆玉米回收率范围回收率范围回收率范围% % % 603
20、91.5 75.0101.2 61.285.2 85.0104.1 72.592.3 71.482.8 81.1100. 7 87.6107.2 87.0101.5 88.6106.2 87.5-105.3 86.797.5 80.5-106.。64 .4 -85.4 76 .44-104.8 79.9-107.。70.4-88.。68.1-80.5 88.0 110. 7 85.6107.。91.2116.0 90.0105.4 89.5103.9 90.1103.6 70二5104.。67.690.8 63.292.8 77.8100.3 67.389.6 74.582.6 91.2111
21、.2 85.2109.。87.2103.3 96:0106.2 90.5103.5 85.6106.3 71.l104.4 68.4108.。72.096.。76.893.6 68.584.9 74.488.4 80.5101.6 87.6105.0 90.0110.6 94;3109.。84.7100.3 84.6100.6 9 SN/3699-2013 For告wordThis standard is drafted according to GB /T 1.1-2009. Please note that this standard some material may be involv
22、ed in patent , this standard release mecha nism does not assume the responsibility of identifying these patents. 丁hisstandard was proposed byand is under the charge of the Certification and Accreditation Admin istration of the Peoples Republic of China. This standard was drafted by Shen Zhen Entry-E
23、xit Inspection and Quarantine Bureau of the Peoples Republic of China , Shenzhen Academy of Inspection and Quarantine, Xia Men Entr-Exit Inspection and Quarantine Bureau of the People s Republic of China. 丁hemain drafters of this standard are Chen bo , jin baohui , Xiao feng , Xie liqi ,Chen Peijin,
24、 Xu dun ming ,Zhou yu. Note: This English version , a translation from the Chinese text , is solely for guidance. 10 SN/3699-2013 B制告rminationof four thiazol号fungicid告告sidu告sin food of pla白toriginfor告xport-HPLC-MSjMSmethod 1 Scope This standard specifies the methods of determination of 4 kinds of fu
25、ngicide residues bliquid chro matograph-tandem mass spectrometrof ethaboxam , etridiazole, octhilinone and benthiazole. This standard is applicable to the qualitative and quantitative determination of 4 kinds of fungicide residues in wheat, grape , watermelon , flowering cabbage , tea , onion , sobe
26、an and corn. 2 Normative references The following documents are necessary for this standard. For dated reference , only dated editions shall apply to this standard. For undated references , the latest edition of the normative document (including subsequent amendments) is referred to applies. GB /T 6
27、682 Water for analytical laboratory use-Specification and test methods. 3 Principle The residues of fungicide residues in the test sample are extracted with acetonitrile. The extraction is purified on SPE column. The residues are determined bLC-MS /MS, using external standard method. 4 Reagents and
28、materials Unless otherwise specified , all reagents should be of analytical grade. Water is the first grade water prescribed by GB /T 6682. 4.1 Methanol: HPLC grade. 4.2 Acetonitrile: HPLC grade. 4.3 Toluene: HPLC grade. 11 SN/T 3699-2013 4.4 Formic acid: HPLC grade. 4.5 Sodium chloride. 4.6 Ammonia
29、 acetate: 99.999%. 4.7 Acetonitrile + water(3+2, V /V): Mix 60 mL acetonitrile(4.2) with 40 mL water. 4.8 Acetonitrile + toluene (3 + 1, V / V): Mix 150 mL acetonitrile(4.2) with 50 mL toluene. 435mmoi/L ammomacetate idatGrsbiL巾的.05.%formiC:acidj: Weight 0.385 9 ammonia ace tate(4.6) , add 0.5 mL fo
30、rmic aCi9 diluted to 1. L with Water an m?50% :t 20% Under the optimized instrument The response of thiazole be in the linear range of the gicide residues retention times and etridiazole 8.4 min. The mul are shown in Fig.C.1 , annex C. 7.3.4 81ank test The operation of the blank test with omission o
31、f standard addition. 8 Calculation and expression of result (10% 土50%matrix calibration solutions are is , and concentration as x-axis, condition , thiazole fun-atograms of thiazole residues method of determination, but The calculation of thiazole fungicide residues in the sample is according to for
32、mula (1) , the blank value shall be subtracted from the result of calculation: Where: X Ci Vn旦些-m . 1 000 Xj一一-theresidue content of thiazole fungicide residues in the test sample , gjkg; 16 . ( 1 ) SN/T 3699-2013 Cj一一-theconcentration of thiazole fungicide residues calculated by each standard calib
33、ration curve, 9 /L; V一一一thefinal volume of sample solution, mL n-dilution factor; m一-thecorresponding mass of test sample in the final sample solution ,g. 9 Recovery and the limit of 9.1 The limit of determination The limit of determination of this 9.2 Recovery 17 SN/3699-2013 Compound benthiazole o
34、cthilinone etridiazole ethaboxam 18 Annex A CNormative annex) Information of thiazole fungicied residues Table A.1一Informationof thiazole fungicied residues Molecular Molecular Structure formula weight C9Hs N2S3 238.34 可3C H9NOS 213.3 OYNS乓C5H5ClaN20S 247.5 Cl, 自1LmIC4 H,s N4 OS2 320.42 N CAS Number
35、 21564-17-0 26530-20-1 2593-15-9 162650-77 -3 SN/T 3699一2013Annex B (Informative annex) Operating conditions of mass spectrometer Operating conditions of mass spectrometer: a) lon source: ESI; b) Scan mode: Positive ion scan; c) Detetion method: MRM; d) lonspray voltage: 5 500 V; e) Curtain Gas: 68.
36、95 kPa; f) GS1: 275.8 kPa; g) GS2: 482.65 kPa; h) TEM: 600 oC; i) Other mass spectrometer parameters show in table 8.1. Table B.1-Mass spectrometer parameters of thiazole fungicide residues Retention Daughter Dwell Parent ion CE DP EP Compound tme lon time Cm/z) eV eV eV 付1mCm/z) 盯lS183.1於100 30 60
37、14 ethaboxam 3.1 322.1 200.2 100 30 60 14 180.0兴100 9 20 10 benthiazole 5.0 239.1 212.0 100 11 20 10 101.9祷100 14 100 10 octhilinone 8.1 214.1 71.1 100 20 100 10 218.9铃100 16 35 10 etridiazole 8.4 246.9 183.0 100 26 35 10 CXP eV 10 10 10 10 15 9 10 10 Annotation: the symbol 祷represents the quantitat
38、ive ion. Instrument. parameters may be different for different mass spectrometer, so mass spectrum parameters shall be optimized before determination. Non.心omm盯me町rci汩a创Is创ta剖temen川t:the mass p阳ar阳ame创tersin annex A are accomp川li丽shedby AP问14000LC巳-MS/MS，the equipment and its type involved in the st
39、andard method is only for reference and not related to any commercial aim , and the ana Iysts are encouraged to use equipments of different corporation or different type. 19 SN/3699-2013 Annex C (Informative annex) MRM chromatogram of thiazole fungicide 8.43 8.44 夏20j 1 etridi缸。lef2000 I etridiazole
40、 1U、问dkiL且6.518.047 8.91叫13.00 一14.47句。1,f.63 护工La8.1911Q:_飞12巫也巳;.a. 2 4 6 8 :10户1214 2 4 6 8 10 12 14 Time/min 8. 06 、h飞亏4 、主i 5000m.0 h 2 00 4 6 8 10 12 14 3.13 3.13 Time/min 材是的K的口也必4 OOOi I ethaboxam 2 000 龟b9.10 10.30 ATimeA/mm f iIadfl$2二1142 4 8 10 12 14 Time/min S R:l i 耳b1M 1. Oe4 5000.0 0
41、.0 2 4 Fig C.1 standard solution 20 Comound ethaboxam benthiazole octhilinone etridiazole Annex 0 (Informative annex) Recovery SN/3699-2013 Table D.1-Recovery data of different spiked level in different matrix Spiked Grape Wate9r6ml平elonF叫WheaLFlowering T自到Onior Soybean Corn level % ge、% % % % % mg/
42、kg 0.01 78.4-100.5 70.0斗驰。70.095.0 88.0-:118:0 回.3-91.575.0-101.2 61.2-85.2 一一一0.1 83.2-102.7 80.3-94.2 71.2-87.6 9.8103.0 72:.吵89.685.0-104.1 72.5-92.3 71.4-82.8 83.2?-99,1 飞户，、y8io95.o 1.0 88.0-107.2 87.7-103.7 918103J2 / 81.1-100.7 87.6-107.2 87.0-101.5 6.0 88.6-104.0 82.7 -91.5 85.6-96;4 8op9/ 1
43、5 80.1-93.8 88.6-106.2 87.5-105.3 86.7-97.5 0.01 92.0-112.0 76.0?，0可8，016;5105.6 70;0-93.8: 1:6)乒乓80.5-106.0 64.4-85.4 176.44-1归.8国削A?明与75毫7号93.0一均些壁与垒喇些:1型叭企尹些1湖型0.1 85.2-99.8 79.9-107.0 70.4-88.0 68.1-80.5 1.0 90.4-109.2 88.0-110.7 85.6-107.0 91.2-116.0 6.0 88.3-98.1 80.6-94.2 85.6-99.9 82.3，:;:，:
44、92A 、二黯.4-96.590.0-105.4 89.5-103.9 90.1-103.6 0.01 74.0-103.0 95.0115/.0 飞76毫-100.864;486.m 子393.5 70.5-104.0 67.6-90.8 63.2-92.8 0.1 88.8-102.4 77.3-)2 p 74手-91芳83.9;107.0 、:臼部77.8-100.3 67.3-89.6 74.5-82.6 1.0 88.4-104.0 四02.7叫87.010/ 35 82A子飞1i 91.2-111.2 85.2-109.0 87.2-103.3 6.0 89.6-104.6 80.
45、493.4 .86.2-99.0 81.0寸95.581.6:;99.74 96.0-106.2 90.5-103.5 85.6-106.3 0.01 79.0-103.0 68.5-89.9 70.5-96.6 71.099.0 70!095;.0 71.1-104.4 68.4-108.0 72.0-96.0 、户命、0.1 83.4-101.9 79.0-992 74.095.5 82.91-112.0 A37;8妇;76.8-93.6 68.5-84.9 74.4-88.4 1.0 86.0-100.8 76.2与四.680.1 -94.8 83.5-97.8 82;0-92:声80.
46、5-101.6 87.6-105.0 90.0- 110.6 6.0 83.8-102.4 79h833 81.5-96.5 /83.8-99.9 8OA/ 90.6 94.3-109.0 84.7 -100.3 84.6-100.6 21 的FONi的阳军中华人民共和国出入境检验检疫行业标准出口植物源食品中4种瞌瞌类杀菌剂残留髦的那定液相色谱质i普/质谱法SN/T 3699-2013 守令中国标准出版社出版北京市朝阳区和平里西街甲2号(100029)北京市西城区三里何北街16号。00045)总编室:(010)68533533网址中国标准出版社秦皇岛印刷厂印刷争夺开本880X 1230 1/16 印张1.75字数40千字2014年12月第一版2014年12月第一次印刷印数11300 定价27.00元头书号:155066 2-27757 SN/T 3699-2013
