1、:才己中华人民共和国进出口商品检验行业标准SN 0502-95 出口水产品中毒杀芬残留量检验方法Method for the determination of toxaphene residues in aquatic products for export 1995-11-28发布1996-0个01实施中华人民共和国国家进出口商品检验局发布中华人民共和国进出口商品检验行业标准出口水产品中毒杀芬残留量检验方法SN 0502二95Method for the determination of toxaphene residues in aquatic products for export 主题
2、内容与适用范围本标准规定了出口水产品中毒杀芬咙留量检验的抽样和制样及气相色谱测定方法。本标准适用于出口鲸鱼、扇贝柱中毒杀芬残留量的检验。2 抽样和制样2. 1 检验批以不超过10000件为一检验批。同检验批的商品应具有相同的特征，如包装、标记、产地、规格和等级等。2. 2 抽样数量批量，件最低抽样数，件冷i在品j舌品、盐藏品:S; 150 151 3 200 320110000 王三9091 500 5011 200 1 201 10 000 内JnuqJAE 叶An/UM2. 3 抽样方法接2.2规定的抽样件数随机抽取，逐件开启。每件至少取500g为原始样品，原始样品总量不少于2kg，加封后
3、，标明标记，及时送实验室。2.4 试样制备将抽取的样品去鳞、去骨后，将可食部分充分捣碎，充分由匀，用四分法缩分出不少于1OOOg，作为i式样.装入清洁的容器中，加封后，标明标记。2. 5 试样保存将试样于一18C以下冷冻保存。注:在抽样及制样的操作过程中，必须防止样品受到污染或发生残留物含量的变化。3 测定方法3. 1 方法提要样品中的毒杀芬经用石油酷提取，净化、改缩后，用配有电子俘获检测器的气相色谱仪测定，外标法定量。3. 2 试刑和材料所用水为蒸馆水，所用试剂除特殊规定外均为分析纯。中华人民共和国国家进出口商品检验局1995-11-28批准1996-0个01实施SN 0502-95 3.2
4、.1 元水硫酸纳z经650C灼烧础，置于干燥器内备用。3.2.2 无水硫酸纳柱:25mm(id)X200mm玻璃柱，内装50mm高的无水硫酸纳。3. 2. 3 石油醒:3060C，经全玻璃装置重蒸悟。3. 2. 4 乙酷:经全玻璃装置重蒸馅。3.2.5 乙睛:色谱纯，用石油酷饱和。3.2.6 氧化制榕液:饱和榕液。3. 2. 7 弗罗里硅土:675C灼烧拙，置于干燥器内，用前于130C烘5ho3. 2. 8 弗罗里硅土柱:22mm(id)X 200mm玻璃柱，内装100mm高的活化的弗罗里硅土，顶上加10mm高的无水硫酸纳。3.2.9 毒杀芬标准品:已知纯度(约78%)。3.2.10 毒杀芬标
5、准储备溶液:准确称取适量的毒杀芬标准品，用石油酷配成放度为100g/mL的标准储备榕液，根据需要再配成适当放度的标准工作榕液。3. 3 仪器和设备3.3.1 气相色谱仪:配有电子俘获检测器。3.3.2 微量注射器:10L。3.3.3 高速组织捣碎机。3.3.4 恒温水恪。3. 3. 5 马弗炉。3. 3. 6 烘箱。3. 3. 7 旋转蒸发器或K-D被缩器。3. 3. 8 分被漏斗:1000mL，125mL。3.3.9 刻度试管:10mL，具磨口塞。3.4 测定步骤3.4.1 提取3.4. 1. 1 鱼类称取约50g(精确至O.lg)试样于高速组织捣碎机中，加100g无水硫酸纳，说匀。加150
6、mL石油醋，均质2mino上清液通过铺有两层滤纸的12cm布氏漏斗滤入500mL抽滤瓶中。用2X100mL石油酷提取捣碎机中残渣，每次3min，过滤，合并提取液。转移费渣至布氏漏斗上，以3X50mL石油酷洗涤。全部i虑液过无水硫酸铀柱，收集流出液。用少量石油酷洗柱，合并石油酷流出液。于旋转蒸发器中蒸发至近f，用少量石油酷转移残渣至已知重量的烧杯中，在60C水浴上蒸发，得到脂肪，称量并记录脂肪量。称取3g脂肪于125mL分液漏斗中，加石油酷使之总体积为15mL，加30mL用石油酷饱和的乙睛，慌荡lmn。分层后，放乙睛层至盛有650mL水、40mL饱和氧化纳洛液及100mL石油酷的1L分液漏斗中。
7、用3X30mL石油酷饱和的乙睛掖洗125mL分液漏斗中的液体，合并乙睛层于1L分液漏斗中。放lL分;夜漏斗中的水层至另一lL分掖漏斗中，加100mL石油酷至第二个分被漏斗中，振摇15s，弃去水臣。合并石油醋层，以2X100mL水洗涤。将石油酷层过无水硫酸铀柱、收集于500mLK-D放缩器中，以3X lOmL石油酶洗涤漏斗及柱。合并石油酷洗被于浓缩器中，浓缩至510mL。3. 4.1.2 扇贝柱称取约lOOg(精确至O.lg)试样于高速组织捣碎机中，加200mL乙睛，均质2min，通过铺有滤纸的12cm布氏漏斗洁、入500mL抽滤瓶中。转移滤液至lL分液漏斗中，加lOOmL石油醋，振摇12min
8、，再加lOmL饱和氧化纳液和600mL水，振摇3040s。弃去水层。以2XlOOmL水洗涤。转移溶剂层于lOOmL具塞量筒中，加15g无水硫酸锅，振摇，转至K-D浓缩器上被缩至5lOmLo3.4.2 净比用50mL石油酷预洗弗罗里硅土柱(3.2.肘，将试样提取液移入柱中，以5mL/min速度过柱，用SN 0502-95 200mL乙酷-石油酷(6十94)以5mL/min速度洗脱。收集洗脱液，于旋转蒸发器上放缩至5mL。3. 4.3 测定3.4.3.1 色谱条件a. 色i昔柱:玻璃柱，2mX 2mm(id)，填充物为1.95%OV-210+ 1. 5%OV-17，涂于ChromosorbW I-
9、IP; b. 柱温:195C(lornin)1OC/mi:_225 C (8min); C. 进样口温度:250C; d. 检测器温度:300C; e. 载气:氨气，纯度大于等于99.99%, 30mL/min o 3. 4.3.2 色谱测定分别准确注射1L样液及标准工作搭液于气相、色谱仪中，按3.4.3.1条件进行色谱分析，响应值均!坐在仪器检测的线性范围内。毒杀芬含量采用最后四个峰的峰面积总和进行定量(峰V，W，X，Y)。在上述条件下，峰V、W、X、Y的保留时间分别为14.5min、15.3min、17.0min、18.2min。3. 5 雪白试验除不称取样品外，按上述测定步骤进行。3.6
10、 结果计算和表述用色谱数据处理机或按下式计算试样中毒杀芬残留含量:式中:X一一试样中毒杀芬残留含量，mg/kg;X=A.c.V Z二A. m 11 样1ft中毒杀芬四个峰的峰面积之和，mmz;A一一标准工作陪液中毒杀芬四个峰的峰面积之和，mm2;C 标准工作洛液中毒杀芬的改度，用/mL;V一一最终样液体积，mL;11 最终样液相当的样品重量，go注:空臼值应从计算结果中扣除。4 测定低限、回收率4.1 测定低限本方法的测定低限为0.5mg/kgo4.2 回收率回收率的实验数据:毒杀芬添加浓度在O.55. Omg/kg范围内，回收率为84.5%89. 5%。附加说明:本标准由中华人民共和国国家进
11、出口商品检验局提出。本标准由中华人民共和国烟台进出口商品检验局负责起草。本标准主要起草人李立、穆兆纯、xtJ桂荣、沈志刚、王洪兵。3 Professional Standard of the People s Republic of China f or Import and Export Commodity Inspection SN 0502 95 Method for the determination of toxaphene residues in aquatic products for export 1 Scope and field of application This st
12、andand specifies the methods of sampling , sample preparation and determination by gas chro matography of toxaphene residues in aquatic products. This standard is applicable to the determination of toxaphene residues in spanish mackerel and scallop for export. 2 Sampling and sample preparation 2. 1
13、Inspetion lot The quantity of an inspection lot should not be more than 10000 packages. The characteristics of the cargo within the same inspection lot ,such as packing, mark ,origin ,specifica tion and grade etc. ,should be the same. 2. 2 Quantity of sample taken Number of packages in each inspecti
14、on lot Minimum number of packages to be taken Frozen Live ,salted 主豆150151-3 200 3201-10000 三二9091一-500501-1 200 1 201-10 000 内，JAUJ口口噜i今年2. 3 Sampling procedure A number of packages speciHed in 2. 2 are taken at random and opened one by one. The sample weight takcn as the primary sample from each p
15、ackage should be at least 500g. The total weight of all primary sampl臼shouldnot be less than 2kg. Which shall be sealed , labeled and sent to laberatory in time. 2. 4 Preparation of test sample The combined primary sample is scaled and deboned. The edible portions are blended and reduced by quarteri
16、ng to not less than 1000g as the test sample , which is then placed in a clean container , sealed and la bcled. 2. 5 Storage of test sample The test sample should be stored below -18 C。ote , In the course of sampling and ample preparation.precaution must be taken to avoid the contamination on any Ap
17、proved by the State Administration of Import and Export Commodity Inspection of the People s Republic of China on Nov , 28. 1995 Implemented from Jan , 01. 1996 SN 0502-95 other factors which may cause the change of residue content. 3 Method of determination 3. 1 Principle The toxaphene in the test
18、sample is extracted with petroleum ether. The extract is cleaned up and con centrat时.the toxaphene is determined by GC with electron capture detector , using external standard method. 3. 2 Reagents and materials Wateris distilled water , all reagents shoule be of analytical grade , unless otherwise
19、specified. 3. 2. 1 Anhydrous sodium sulfate: Ignited at 650 C for 4 hours , kept in an air-tight closed container after cooling. 3. 2. 2 Anhydrous sodium sulfate column: 25mm (id) X 200mm glass column, filled with ca 50mm height f anhydrous sodium sulfate. 3. 2. 3 Petroleum ether: Redistilled in all
20、 glass apparatus , collect the fractions between 30 60C. 3. 2. 4 Diethyl ether: Redistilled in all glass apparatus. 3. 2.5 Acetonitrile:Chromatographic grade ,saturated with petroleum ether. 3. 2. 6 Sodium chloride solution: Saturated solution. 3. 2.7 Florisil:Ignite at 675 C for 2h ,and store in a
21、desiccator ,then heat at 130C for 5h before use. 3. 2. 8 Florisil column: 22mm (id) X 200mm glass column , filled with ca 100mm height of florisil. and ca 1 ()mm height of anhydrous sodium sulfate on the top. 3. 2. 9 Toxaphene standard: Known purity(ca 78 %). 3. 2. 10 Toxaphene standard stock soluti
22、on: Accurately weight an adequate amount of toxaphene stan dard . c! issolve in petroleum ether and prepare a solution of 100g/mL as the standard stock solution. Ac cording 10 the requirement , prepare a standard working solution of appropriate concentration. 3. 3 Apparatus and叫uipment3. 3. 1 Gas ch
23、romatograph，巳quippedwith electron capture detector. 3. 3. 2 Micro-syringe : 10L. 3.3.3 High-speed blender. 3. 3. 4 Water-bath , with thermostatic regulator. 3. 3. 5 Muffle furnace. 3. 3. 6 Drying oven. 3.3. 7 Rotary vacuum evaporator or Kuderna-Danish concentrator. 3. 3. 8 Separatory funnel:lOOOmL ,
24、125mL. 3. 3. 9 Graduated cylinder: With ground stopper , 10mL. 3. 4 Procedure 3. 4. 1 Extraction 3.4. 1. 1 Spanish mackerel Weigh ca 50g (accurate to O. 19) of the sample into a high-speed blender. Add 100g of anhydrous sodium sulfate , and blended until sample and sodium sulfate are well mixed . Ad
25、d 150mL of petroleum cther and blend for 2min. Decant the petroleum ether supernatant on a 12cm Buchner funnel with two lay ers of filter paper and filter by suction into a 500mL flask. Extract the residue again with two portions of petroleum ether, each 1 OOmL , blending for 3min each time. Filter
26、and combine the extracts. Transfer the residuc into Buchner funnel and wash with 3 X 50日lLof petroleum ether. Let all the extracts pass through 5 SN 0502-95 the anhydrous sodium sulphate column. Wash the column with petroleum ether. and collect all the petroleum ether effluents in a rotary vacuum ev
27、aporator. Evaporate to near drynees. Transfer the fat into a weighted beaker with small amount of petroleum ether. evaporate off the solvent on a water-bath at 60 C and weigh the fat. Weigh 3g of the fat into a 125mL separatory funnel.add petroleum ether to a total volume of 15mL. and add 30mL of ac
28、etonitrile saturated with petroleum ether. Shake for 1min. Transfer acetonitrile layer in to a 1L separatory funnel containing 650mL of water. 40mL saturated sodium chloride solution and 100mL of petroleum ether. Wash the solution in 125mL separatory funnel with 3 X 30mL of acetonitrile saturated wi
29、th petroleum ether. Combine all the extracts into the 1L separatory funnel. Drain the water layer into an other 1L separatory funnel. Add 100mL of petroleum ether to the second separatory funnel. Shake for 15s. discard the aqueous layer. Combine the petroleum ether extracts and wash with two 100mL-p
30、ortions of wa ter. Transfer petroleum ether extract into anhydrous sodium sulfate column. Collect the petroleum ether ef fluent into Kuderna-Danish concentrator. Rinse the separatory funnel and then the columm with three 10mL portions of petroleum ether. Combine the extract and rinsings. and evapora
31、te to 5-10mL. 3.4.1.2 Scallop Weigh 100g(accurate to o. 19)of the sample into a high-speed blender.add 200mL acetonitrile.blend for 2min. Filter with suction through 12cm Buchner funnel fitted with filter paper into a 500mL flask. Transfer the filtrate into a 1L separatory funnel. Add 100mL of petro
32、leum ether. Shake for 1-2min. Add 10mL of saturated sodium chloride solution and 600mL of water. Mix for 30-40s. Discard the aqueous lay er. Wash the organic layer with two 100mL-portions of water. Transfer organic layer into a 100mL gradu ate with glass-stopper. Add 15g of anhydrous sodium sulphate
33、 and shake vigorously. Concentrate to 5-10mL in Kuderna-Danish concentrator. 3. 4. 2 Cleanup Prewet florisil column(3. 2. 8) with 50mL of petroleum ether. Transfer petroleum ether solution of sample-extract to column. letting it pass through at about 5mL/min. Rinse container (and Na2S04 if present)a
34、nd walls of column with small portions of petroleum ether. Elute column at about 5mL/min with 200mL of ethyl ether-petroleum ether mixture (6+94). Collect the eluate in Kuderna-Danish concentrator and concentrate to 5mL. 3.4. 3 Determination 3.4.3. 1 GC operating condition a. Chromatographic column:
35、Glass column.2mX2mm(id).packed with 1. 95%OV-210十1.5%OV -170n Chromosorb W HP; 1WC/min b. Columntemperature:195C (10min) 225C(8min); c. Injection port temperature: 250 cC ; d. Detector temperature: 300C ; e. Carrier gas:Nitrogen.purity99. 99 % ;30mL/min. 3.4.3.2 GC determination Inject separately ex
36、act 1ILL of the standard working solution and sample solution into gas chromato graph .and carry out the analysis under the conditions indicated in 3.4.3. 1. The responses must be within the linear range of the instrumental detection. Toxaphene content is calculated by the sum of the areas of last f
37、our peaks (peak V W X Y). U nder the above chromatographic condition. the retention times of peak V W. X. Y are respectively about 14. 5min .15. 3min .17. Omin .18. 2min. 3. 5 Blank test 6 SN 0502- 95 The operation of the blank test is the same as that described in the method of determination , but
38、with立out sample addition. 3. 6 Calculation and expression of result Calculate the content of toxaphene in the test sample is carried out by GC data processor or according to follow formula: 可-mc一-bA X where X 一-thecontent of toxaphene in the test sample,mg/kg; A一-thesum of 1ast four peak areas of to
39、xaphene in samp1e solution ,mmz; A,-the sum of 1ast four peak areas of toxaphene in standard working soluton, mm勺c一一theconcentraton of toxaphene in standard working soluton，用/mL;V一-thefina1 vo1umn of test solution , mL; m一一一thecorresponding mass of the test samp1e in the final test solution,g. Iote:
40、 The blank value should be subtracted from the above results of calculation. 4 Limit of determination and recovery 4. 1 Limit of determination The 1imit of determination of ths method is O. 5mg/kg. 4.2 Recovery According to the experimenta1 data, when the fortfed concentration of toxaphene s n the r
41、ange of O.5-5.0mg/悔，therecovery is 84.5%-89.5%. Additional explanations: This standard was proposed by the State Administration of Import and Export Commodity Inspection of the Peop1e s Repub1ic of China. This standard was drafted by the Yantai Import and Export Commodity Inspection Bureau of the Peo p1c s Republic of China. This standard was main1y drafted by Li Li , Mu Zhaochun, Liu Guirong ,Shen Zhigang , Wang Hong bing. Note: This English version.a translation from the chinese text ,is solely for guidance. 7 mm|NOmo z 飞;n中国标准出版社秦皇岛印刷厂印刷1996年7月第一版书号.155066 2-10777 1996年7月第一次印刷中国标准出版社出版
