1、 中华人民共和国出入境检验检疫行业标准SN/T 1873-2007 进出口食品中硫丹残留量的检测方法气相色谱一质谱法Determination of endosulfan residues in food for import and export-一GC-MS2007-04-06发布2007-10-16实施中华人民共和国发布国家质量监督检验检疫总局 SN/T 1873-2007 目。吕本标准附录A为资料性附录。本标准由中华人民共和国国家认证认可监督管理委员会提出并归口。本标准由中华人民共和国天津出入境检验检疫局、中华人民共和国广东出入境检验检疫局、中华人民共和国上海出入境检验检疫局负责起草。
2、本标准主要起草人:葛宝坤、王云凤、常春艳、陈捷、陈其勇、刘培、郭德华、焦红、韩丽。本标准系首次发布的检验检疫行业标准。 1 范围进出口食品中硫丹残留量的检测方法气相色谱-质谱法SN/T 1873-2007 本标准规定了食品中硫丹、卢硫丹、硫丹硫酸盐残留量的气相色谱质谱检测方法。本标准适用于鲤鱼、泥锹、鲸鱼、黄鳝、牛肉、大豆、蘑菇、毛豆、丧菜、蒜茎、西红柿、甘蓝、苹果、柑楠、茶叶中硫丹残留量的测定。2 测定方法2.1 方法提要样品经溶剂提取、凝胶层析柱或硅镜吸附剂净化，采用气相色谱质谱测定，外标法定量。2.2 试剂和材料除另有规定外，所有的有机试剂为色谱纯，水为二次蒸馆水。2.2.1 丙酣。2.
3、2.2 乙酸乙醋。2.2.3 环己皖。2.2.4 石油醒:沸程300C600C。2.2.5 元水硫酸铀:分析纯，6500C灼烧4h，在干燥器内冷却至室温，贮于密封瓶中备用。2.2.6 氧化铀:分析纯。2.2.7 洗脱液:乙酸乙醋环己皖(1: 1，体积比)1昆合溶液。2.2.8 凝胶及溶胀:Bio-BeadsS-X3 200目400目或相当者;凝胶的溶胀按每克凝胶加4.6mL乙酸乙醋环己皖(1: 1，体积比)的?昆合溶液浸泡，榕胀6h以上备用。2.2.9 凝胶层析柱:柱长20cm、内径2.0cm具活塞瑛璃层析柱，柱底垫少许瑛璃棉。用洗脱剂乙酸乙醋环己皖(1: 1，体积比)溶胀的凝胶以湿法转入装柱
4、，柱床高约20cm，胶床始终保持在洗脱剂中;洗脱流速约1mL/mi口，上样前用洗脱液以两分之一的洗脱流速淋洗一个柱体积。2.2. 10 固相萃取柱:硅镜吸附剂，1g，或相当者。用前分别用5mL丙酬正己皖(1: 9，体积比)、5 mLlE己皖淋洗活化小柱。2.2.11 硫丹:CAS编码959-98-8，纯度大于99.5%。2.2.12 卢硫丹:CAS编码33213-65-9，纯度大于99.5%。2.2.13 硫丹十卢硫丹混合物:CAS编码115-29-7，纯度(66.1%十33.4%)。2.2.14 硫丹硫酸盐:CAS编码1031-07-8，纯度大于99.5%。2.2.15 硫丹标准储备液:准确
5、称取适量的硫丹标准品，用少量甲苯溶解后，以正己皖稀释成一定浓度的储备液，40C冰箱保存。2.2.16 硫丹标准工作液:取一定量的标准储备液，用正己炕稀释成适当浓度的标准工作液。2.3 仪器和设备2.3. 1 气相色谱质谱仪，配备负化学源(:-.JCl)。2.3.2 旋转蒸发装置。2.3.3 氮吹仪。2.3.4 组织匀浆机。2.3.5 植物粉碎机。2.3.6 振荡器。 SN/T 1873-2007 2.4 测定步骤2.4. 1 样品的制备a) 动物产品:取鲤鱼、泥锹、鲸鱼、黄鳝、牛肉产品取可食部分500g，用组织捣碎机充分捣碎均匀，均分成两份，分别装入洁净容器中，密封、标记、18C冷冻保存。b)
6、 植物产品:取大豆、蘑菇、毛豆、夜菜、蒜苔、西红柿、甘蓝、苹果、柑楠、茶叶500g，组织粉碎机粉碎，分别装入洁净容器中，密封、标记、4C冷藏保存。2.4.2 样品的提取a) 动物产品:称取试样20g(精确到O.Olg)，于100mL具塞三角瓶中，加水6mL(视样品水分含量加水使总水量约20g，7j(产品的肉通常在70%左右，加水6mL即可)，加40mL丙酬，匀浆1min，加氧化纳6g，充分摇匀，再加30mL石油酶，振摇30mi凡取35mL有机层上清液，经无水硫酸铀滤于旋转蒸发瓶中，浓缩至约1mL，加2mL乙酸乙醋环己皖溶液再浓缩，如此重复3次，浓缩至约2mLb) 水果蔬菜:称取试样25g(精确
7、到O.Olg)，于200mL烧杯中，加50mL乙脯，匀浆1min，转移至预先加入6g氯化铀的100mL的具塞量筒中，剧烈振荡2min，静置20min，取10mL上清液于10mL比色管中，在40C水浴中氮吹仪吹干，2mL正己皖、溶解。c) 粮谷、茶叶等:称取粉碎试样10g(精确到0.01g)，于100mL三角瓶中，添加20mL正己:院，振荡30mino过滤，吸取2mL上清液于10mL比色管中。2.4.3 样品的净化a) 对2.4.2.1的提取样品上凝胶层析柱，并以乙酸乙醋环己皖溶液洗脱，弃去omL35 mL流分，收集35mL70 mL流分。将其旋转蒸发浓缩至约2mL，再经凝胶层析柱重复净化一次，
8、收集35mL70 mL流分，蒸发浓缩，用氮吹仪吹除溶剂(40C)，以正己皖定容至5mL，留待GC/MS分析。b) 对2.4.2.2和2.4.2.3的浓缩液全部上固相萃取柱，再用10mL丙酣正己皖洗脱，将盛有淋洗液的离心管置于氮吹仪上，40C水浴氮吹至近干，蔬菜、水果类用正己皖准确定容至5mL , 粮谷类用正己院准确定容至1mL，在旋涡混合器上由匀，移入样品瓶中待测。2.4.4 测定2.4.4. 1 气相色谱质谱条件2 a) 色谱柱:DB-1701柱，30mX 0.25 mmX 0.25m，或相当者;b) 载气:氯气，纯度不低于99.999%，流速:1.3 mL/mi川c) 柱温:40C保持1m
9、i口，以30oC/min速度升至130C，再以50C/min升至260oC，保持1min; d) 进样口温度:250C;巳)进样方式:不分流;f) 开阀时间:0.75min; g) 进样量:2L;h) 电离方式:NCI，30eV; i) 接口温度:280oC; j) 反应气:甲:皖气，CH.1;k) 离子源温度:150oC; 1) 溶剂延迟:10mi川m) 离子检测模式:选择离子监测(SIM)，监测离子及其相对丰度比见表L表1选择离子和相对丰度被测组分选择离子m/z相对丰度/c%)硫丹、卢硫丹374.404.406C定量).408.410 28: 47 : 100: 84: 32 硫丹硫酸盐3
10、84.386C定量).388.39062 : 100 : 72 : 30 SN/T 1873-2007 2.4.4.2 定量测定根据样液中被测硫丹的含量情况，选定浓度相近的标准工作液，其响应值应在方法检测的线性范围内。在上述气相色谱质谱条件下，保留时间分别为硫丹23.7min、硫丹27.7 min、硫丹硫酸盐30.4 min，选择离子色谱图和质谱图见附录A中的图A.1图A.502.4.4.3 定性测定按照色谱质谱条件测定样品和标准工作溶液，如果检测的质量色谱峰的保留时间与标准品一致，相对丰度允许偏差小于20%，则可判断样品中存在对应的被测物。2.4.4.4 空白试验除不加试样外，均按上述步骤进
11、行。2.4.5 结果计算与表述按式(1)计算试样中硫丹的含量(以硫丹、卢硫丹、硫丹硫酸盐的总量计): V -m fku一-Ah A X . ( 1 ) 式中:X 试样中硫丹的含量，单位为毫克每千克(mg/kg); A 试样中硫丹的色谱峰面积;C, 标准工作溶液中硫丹的浓度，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);A 标准工作溶液中硫丹的色谱峰面积;m 最终样液所代表的量，单位为克(g)。2.4.6 测定低限、回收率2.4.6. 1 测定低限本方法的测定低限:动物产品0.004mg/kg、植物产品0.01mg/kg o 2.4.6.2 回收率动物产品、植物产品样
12、品添加回收率见表20表2动物产品、植物产品样品添加回收率样品添加水平/Cmg/kg)回收率范围/c%)样品添加水平/Cmg/kg)回收率范围/C%)0.004 67. 577. 5 0.004 70. 082. 5 鲤鱼0.01 71. 079. 0 给鱼0.01 77.087.5 0.05 70. 088.。0.05 78. 094.。0.004 70. 085.。0.004 75. 087. 5 泥锹0.01 79. 087.。牛肉0.01 81. 088.。0.05 78. 094.。0.05 80. 096.。0.01 70. 080.。0.01 71. 088. 0 大豆0.04 7
13、2. 585. 0 苹果0.04 72. 590. 0 o. 10 80. 090.。o. 10 80. 096.。0.01 76. 087.。0.01 70. 078.。甘蓝0.04 75. 092. 5 蒜苔0.04 70. 082. 5 O. 10 86. 097.。O. 10 79. 089.。0.01 70. 076.。0.01 74. 088.。茶叶0.04 70. 080.。菠菜0.04 72. 590.。O. 10 77. 085.。O. 10 83. 097.。3 SN/T 1873-2007 附录A(资料性附录)标准晶质谱固0609071 2: SIR of 5 Chann
14、cIs CI-l 0609071 100才平7312777 1吐UOI 408 IJ止. 111111111111 20. 00 22. 00 24. 00 26. 00 28. 00 30. 00 32. 00 1: SIR of 5 Channcls Cl TIC 岳08c岳Timc 图A.1-硫丹、p-硫丹、硫丹硫酸盐选择离子色谱图06073且4233 (27. 5且6)100 2: SIR of 5 Channcls Cl-406.00 6. 84c4 % i374. 00 。37岳380 38岳390 39岳400 40岳Dac 410 图A.2-硫丹、p-硫丹选择离子质谱图0610
15、015 209(30.762) 2: SIR of 4 Channcls Cl-386.00 2. 81c4 388.00 390.00 % 。384m 389 390 384 38岳386 387 388 图A.3硫丹硫酸盐的选择离子质谱图0608015 1120(30.559) % n叫1 0邸向，uqdJ/nb mmm 4 oo qd Scan CI-5.07c4 nu nu l /391. 20 422.20 。)ae 340 350 360 370 380 390 400 410 420 430 图A.4硫丹硫酸盐全扫描质i普图4 06071941388(27.486) 99.25
16、1001 100 I岳。200 2岳。nu nu nu 3岳。400 固A.5-硫丹、p-硫丹全扫描质i普图Scan Cl-)ae 4岳。SN/T 1873-2007 5 SN/T 1873-2007 Foreword Annex A of this standard is an information annex. This standard was proposed byand is under the charged of certification and accreditation administa tion of the People s Republic of China. T
17、his standard was drafted by Tianjin Entry-Exit Inspection and Ouarantine Bureau , and Guangdong Entry-Exit Inspection and Ouarantine Bureau and Shanghai Entr-Exit Inspection and Ouarantine Bu reau. The standard was mainly drafted by Ge-bao Kun , Wang-yun Fe吨，Chang一chunYan , Chen-jie ,Chen qi Yong ,
18、Li u Pei , Guo-de Hua , Jiao-hong , Han-l i. This standard is professional standard for entr-exit inspection and quarantine promulgated for the first time. 6 SN/T 1873-2007 1 Scope Determination of endosulfan residues in food for import and export -GC-MS This standard specifies the determination and
19、 confirmation of -endosulfan， -endosulfan and en dosulfan sulfate in food by gas chromatography- mass spectrum. This standard is applicable for the detemination and confirmation of endosulfan in eel , loach , catfish , rice field eel , beef, soybean , mushroom, green soy bean , spinage , garlic, tom
20、ato, cabbage , apple , orange and tea samples. 2 Method of determination 2. 1 Principle Endosulfan in the test sample are extracted with the solvent , then cleaned up with gelatin chromato graphic column. Determination bgas chromatograph-mass spectrum and quanyified busing the external standard. 2.
21、2 Reagents and materials Unless otherwise specified, all the reagent used should be chromatograph pure , and water is dei onized water. 2. 2. 1 Acetone. 2. 2. 2 Ethyl acetate. 2.2.3 Cclhexane. 2.2.4 Petroleum ether: boils the regulation. 2.2.5 Anhdrous sodium sulfate: ignite at 650巳for4 h, and store
22、 in air-tight container. 2.2.6 Sodium chloride Canalytical pure). 2.2.7 Elute solvet: ethyl acetate-cyclohexaneC 1 : 1). 7 SN/T 1873-2007 2.2.8 Gel and swelling: Bio - Beads S-X3 200 mesh-400 mesh or equivalent; The swelling of gel through every 4 9 gel adding 4. 6 mL Ethyl acetate -cclohexane (1 :
23、1) immersion. The gel to be used when swelling 6 h. 2.2.9 Gelatin chromatographic column: the length of the column is 20 cm , Inside diameter 2.0 cm with a piston , and the material quality is glass. Put some glass wool in the base of the column. Put the gelatin soaks with Ethyl acetate-cyclohexane
24、(1 : 1) in the column by aqueous method. The length of the Column bed is nearly 20 cm , gelatin bed throughout in the Ethyl acetate-cyclohexane (1 : 1); The elute flow rate is about 1 mL/min. Elute the column use 1 column volume elute solution at a flow rate of O. 5 mL/min before pour extract soluti
25、on. 2.2.10 SPE cartridge: florisil , 1 g , or equivalent. It was conditioned with 5 mL acetone-n-hexane (10 + 90) followed by 5 mL n-hexan. 2.2. 11亿endosulfan:CAS code 959-98-8 , purity99. 5%. 2.2. 12卢-endosulfan:CAS code 33213-65-9 , purity99.5%. 2.2. 13 The mixture of -endosulfan and -endosulfan:
26、CAS code 115-29-7. 2.2.14 Endosulfan sulfate: CAS code 1031-07-8 , purity99.5%. 2.2. 15 Stock standard solution of endosulfan: accurately weigh adequate amounts of endosulfan standard , dissolution ba little toluene , then dissolved in N - hexane to make up standard stock so lution of the density, s
27、hould be stored at 40C in refrigerator. 2.2. 16 Working standard solution of endosulfan: dilute the stock standard solution with n- hexane to 1.0g/mL as working standard solution. 2.3 Apparatus and equipments 2.3. 1 Gas chromatograph combined with negative chemical ionization mass spectrogram (GC-MS
28、). 2. 3. 2 Rotary vacuum evaporator. 2.3.3 Nitrogen evaperator. 2.3.4 High-speed homogenizer. 2.3.5 Plant disintegrator. 2.3.6 Oscillators. 8 SN/T 1873-2007 2. 4 Procedure 2. 4. 1 Sample preparation a) Creatural sample: Take the edible part of original sample is taken and homogenized, then it is di
29、vided in two angd sealed , labeled for lab test. The test sample is stored in一1S0Crefrigeratory. b) Botanic sample: Take original sample is taken and homogenized , then it is divided in two angd sealed , labeled for lab test. The test sample is stored in 40C refrigeratory. 2.4.2 Sample Extraction a)
30、 Creatural sample: Weight 20 g(accurate to O. 01 g) of the test sample into 100 mL conical flask , accurately adding 6 mL water, adding 40 mL acetone , homogenize 1 min and add 6 9 of sodium chloride, mix well. Adding 30 mL petroleum ether, shake 30 min , Taking the upper liquid 35 mL flask, combine
31、 extracted solution and evaporate to nearly 1 mL, adding 2 mL of ethyl acetate cclohexane (1 :门，combinethrice according to above process , combine the extract solution nearly 2 m L. b) Fruit and vegetable: Weight 25 g(accurate to O. 01 g) of the test sample into 200 mL beaker , ac curately adding 50
32、 mL acetonitrile, homogenize 1 min and transfer into the graduated cylinders with stopper, then add 6 9 of sodium chloride into the graduated cylinders. Shake vigorously 2 min , taking the upper liquid into 10 mL centrifuge tube, below it to dryness with nitrogen flow at 400C water-bath , dissolve w
33、ith 2 mL petroleum ether. c) Grain and tea: Weight 10 g(accurate to 0.01 g) of the test sample into 100 mL conical flask, adding 20 mL n-hexane, shake 30 min , then filter and taking the 2 mL upper liquid into 10 mL centrifuge tube. 2. 4. 3 Sample clean up a) Wash the extract sample solution of 2.4.
34、2. 1 with Ethyl acetate-cyclohexane( 1 : 1) by gelatin chromatographic column , discarding 0.35 mL effluent , collect 35 mL. 70 mL elution , combine the elution and evaporate to nearly 2 mL, clean up twice according to above process. Combine all the elution and evaporate to dryness under nitrogen fl
35、ow , make up to 5 mL with petroleum e ther, The solution is used for GC-MS determination. b) Pour the sample solution of 2.4.2.2 and 2.4.2.3 into the cleanup column , elute with 5 mL ace tone-n-hexane (10 + 90). Repeat again according to above process. Evaporate to dryness under nitrogen flow at 40o
36、C , then make up to 5 mL with n- hexane in fruit and vegetable and make up to 1 mL with n-hexane in grain; mix well. The solution is used for GC-MS determination. 2.4.4 Determination 2.4.4. 1 GC-MS operating conditions 9 SN/T 1873-2007 a) Column: OB-1701 , 30 m X 0.25 mm X 0.25m or equivalent; b) Ca
37、rrier gas: He(purity99.999%) ,flow rate of carrier gas: 1.3 mL/min; c) Temperature program: 400C(keep 1 min) 30oC/min to 1300C 50C/min t02600C(keep 1 min); d) Injection temperature: 250oC; e) Injection mode: splitless; f) Purge time: 0.75 min; g) Injection volume: 2L; h) Electron mode: NCI , 30 eV;
38、i) Interface temperature: 280oC; j) Reaction gas: CH4; k) lon source temperature: 150oC; 1) Solvet delay: 10 min; m) Oetection mode:SIM: Selection ions (m/z) and relative intensity(%) see table1. Table 1-Selected ions and relative intensity Analyte endosu Ifan、卢-endosulfanendosulfan sulfate Selected
39、 ions m/z 374 .404.406(quantitative) .408.410 384.386Cquantitative) .388.390 Relative intensity / (%) 28 : 47 : 100 : 84 : 32 62 : 1 00 : 72 : 30 2.4.4.2 GC-MS determination According to the values of endosulfan in sample , select the standard working solution with similar peak area to that of sampl
40、e solution. The responses of endosulfan in the standard working and the sample solution should be within the liner range of the instrumental detection. Under the above GC MS operating condition , The retention time of for chromatogram of -endosulfan is 23. 7 min , -en dosulfan is 27.7 min and endosu
41、lfan sulfate is 30.4 min , The chromatogram and mass spectrum of the standard , see Figure A. 1 ,._, Figure A. 5. of annex A. 2.4.4.3 Confirmation of GC-MS Under GC-MS conditions, the working solution and solution is injected. If the retention times of sample chromatogram peaks are consistent with t
42、he standards , and the deviation of the abundance ratio between sample and standard within 20% , according to the selected ions and relative intensity to confirmate. 2. 4. 4. 4 Blank test The operation of the blank test is the same as the described in the method of determination, but with the omissi
43、on of sample addition. 2.4.5 Calculation and expression of result Calculation the content of endosulfan residue in the test sample according the formula( 1) , (the result 10 SN/T 1873-2007 is the total of -endosulfan , -endosulfan and endosulfan sulfate) : X=x Cs X V As x m 、，FaEI /h . . . . . . . .
44、 . . . . . . . . . . . . . . . Where , X-the residue content of endosulfan in the test sample, mg/kg; A-the peak area of c in sample solution; cs-the concentration of endosulfan in standard working solution; v- the final volume of the sample solution , mL; As-the peak area of endosulfan in standard
45、working solution; m-Mass of test sample ,g. 2. 4. 6 Limit of determination and recovery 2.4.6. 1 Limit of determination The limit of determination of this method: the creatural sample is 0.004 mg/闸，the Botanic sample is 0.01 mg/kg. 2.4.6.2 Recover According to the experimental data , the correspondi
46、ng recoveries of fortifing concentrations fortif ying concentrations in creatural sample see table 2. Table 2-The corresponding recoveries of fortifying concentrations in creatnral sample and botanic sample sample fortifying level/ recovery / ( % ) sample fortifying level/ recovery / ( % ) (mg/kg) (
47、mg/kg) 0.004 67. 577. 5 0.004 70. 082. 5 eel 0.01 71. 079.。catfish 0.01 77. 087. 5 0.05 70. 088.。0.05 78. 094. 0 0.004 70. 085.。0.004 75. 087. 5 rice field eel 0.01 79. 087.。beef 0.01 81. 088.。0.05 78. 094. 0 0.05 80. 096.。0.01 70. 080.。0.01 71. 088.。soybean 0.04 72. 585.。apple 0.04 72. 590.。0.10 80
48、. 090.。0.10 80. 096.。0.01 76. 087.。0.01 70. 078.。cabbage 0.04 75. 092. 5 garlic 0.04 70. 082. 5 0.10 86. 097.。0.10 79. 089.。0.01 70. O 76.。0.01 74. 088. 0 tea 0.04 70. 080.。splnage 0.04 72. 590.。0.10 77. 085.。0.10 83. 097.。11 SN/T 1873-2007 12 Annex A (informative annex) Chromatogram of the standard 0609071 0609071 100才平7312777 1吐UOI 408 IJ止2: SIR of 5 ChanncIs CI-l
