【商检行业标准(SN)】 进出口疏荣中氟啶脲残留量检测方法 高效液相色谱法

中华人民共和国出入境检验检疫行业标准SN/T 2095—2008
进出口蔬菜中氟啶脲残留量检测方法高效液相色谱法
Determination of chlorfluazuron residues in vegetables for import and export-Highperformanceliquid chromatographicmethod2008-07-17发布
中华人民共和国
国家质量监督检验检疫总局
2009-02-01实施
本标准的附录A是资料性附录。
本标准由国家认证认可监督管理委员会提出并归口。本标准由中华人民共和国浙江出人境检验检疫局负责起草本标准主要起草人：刘海山、陈笑梅、石蕾、池浩超、汤杭燕、杨磊本标准系首次发布的出入境检验检疫行业标准SN/T2095—2008
1范围
进出口蔬菜中氟啶脲残留量检测方法高效液相色谱法
本标准规定了蔬菜中氟啶脲残留量检测的制样和高效液相色谱测定方法。本标准适用于黄瓜、萝下、荷兰豆中氟啶腺残留量的检测。2方法提要
SN/T2095—2008
用乙腈提取试样中的氟啶脲，经弗罗里硅土柱净化，用配有二极管阵列检测器的高效液相色谱仪测定，外标法定量。
3试剂和材料
除另有规定外，试剂均为分析纯，水为重蒸水或去离子水。3.1乙睛：液相色谱级。
3.2正已烷：液相色谱级。
3.3乙醚：液相色谱级。
3.4，氯化钠。
3.5无水硫酸钠：650℃灼烧4h，在干燥器内冷却至室温，贮于密封瓶中备用。3.6洗脱液：正已烷十乙醚（2+8.体积比）。3.7脱脂棉，
3.8弗罗里硅土：100目~200目，粒度：0.075mm~0.15mm650℃灼烧4h，使用前一天130℃活化4h在干燥器内冷却至室温，加1%的水脱活备用。3.9净化柱：200mm×15mm（内径）玻璃柱，底部填约5mm高脱脂棉和20mm高无水硫酸钠（3.5），10g弗罗里硅土，顶端加20mm高无水硫酸钠，使用前用20mL正已烷淋洗。3.10氟啶标准品：纯度大于等于99.0%。3.11氟啶脲标准储备溶液：准确称取适量氟啶脲标准品，用甲醇配制成100ug/mL标准储备液。0℃～4℃储存（有效期6个月）。3.12氟啶脲标准工作溶液：根据需要，将标准储备液（3.12)用甲醇稀释至适当浓度的标准工作溶液4仪器和设备
4.1高效液相色谱仪，配有二极管阵列检测器4.2均质器。
4.3旋转蒸发仪。
4.4混匀器。
4.5离心机（4000r/min）。
5试样制备与保存
从混合原始样品中缩分出1kg，取可食部分·经组织捣碎机捣碎，均分成两份，装人洁净容器内·作为试样，密封，并标明标记。将试样置于一18C以下冷冻保存。在制样操作过程中，应防正样品受到污SN/T2095—2008
染或发生残留物含量的变化。
6测定步骤
6.1提取
称取5g试样（精确至0.01g），置于100mL烧杯中，加人30mL乙晴，10000r/min均质2min。提取液过滤至50mL塑料离心管中，用15mL乙睛分两次洗涤烧杯和均质器，洗涤液与滤液合并。在滤液中加人氯化钠.剧烈振摇后，离心（4000r/min)5min，收集有机相。下层水相再用20mL乙萃取一次，合并有机相，在45℃水浴中用旋转蒸发仪减压浓缩至干。6.2净化
残渣用洗脱液（3.6)溶解洗涤三次，每次5mL。将溶液全部移人净化柱（3.9）中，用100mL洗脱液（3.6)洗脱。收集洗脱液，在35℃水浴中用旋转蒸发仪减压浓缩至干，用流动相（乙睛+水=60十40，体积比）定容至1.0mL，溶液过0.45μm滤膜后供高效液相色谱分析。6.3测定
6.3.1色谱条件
色谱柱：Cl：柱，5um，250mm×4.6mm（内径），或相当者；流动相：梯度洗脱条件见表1；
流速：1.0mL/min；
检测波长：258nm；
进样量：20μL。
表1梯度洗脱条件
时间/min
6.3.2色谱测定
乙腈/%
根据样液中氟啶脲含量，选择浓度相近的标准工作溶液。标准工作溶液和样液中氟脲的响应值均应在仪器检测的线性范围内。对标准工作溶液和样液等体积穿插进样测定。在上述色谱条件下，氟啶脲的保留时间约为17.3min。标准品的色谱图见附录A中图A.1，标准品的紫外光谱图见附录A中图A.2。
6.4空白试验
除不加试样外，均按上述步骤进行结果计算和表述
用色谱数据处理机或按式(1)计算试样中氟啶脲含量，计算结果须扣除空白值。AXcXV
式中：
X试样中氟啶脲的残留量，单位为毫克每千克（mg/kg）；A
样液中氟啶脲的峰面积：
标准工作液中氟啶豚的浓度，单位为微克每毫升（μg/mL）；2
..（1)
样液最终定容体积，单位为毫升（mL）；As——标准工作液中氟啶脲的峰面积；m——最终样液所代表的试样量，单位为克（g）)。测定低限和回收率
测定低限
本方法对蔬菜中氟啶脲的测定低限为0.05mg/kg。8.2
回收率
回收率见表2。
样品基质
液相色谱法测定黄瓜、萝卜和荷兰豆样品中氟啶脲的回收率加浓度bzxZ.net
0.05mg/kg
荷兰豆
86.5%~102.0%
88.7%~104.2%
86.6%~102.1%
0.10mg/kg
87.5%~103.2%
87.5%~102.3%
87.4%~105.7%
0.20mg/kg
86.8%~103.2%
87.6%~102.9%
88.4%~102.8%
SN/T2095—2008
83.3%~106.9%
92.1%~108.5%
82.6%~107.4%
SN/T2095—2008
附录A
（资料性附录）
标准品色谱图和紫外光谱图
氟啶脲标准品的液相色谱图
氟啶脲标准品的紫外光谱图
AnnexAisan informativeannex
Foreword
SN/T2095—2008
This standard is proposed by and is under the charge of Certification and Accreditation administrationofthePeople'sRepublicof China.This standard was drafted by Zhejiang Entry-Exit Inspection and Quarantine Bureau of the People'sRepublic of China
The main drafters of this standard are Liu Haishan,Chen Xiaomei,Shi Lei,Chi Haochao,Tang Hangyan, Yang Lei.
This standard is a professional standard for entry-exit inspection and quarantine promulgated for thefirsttime.
SN/T2095—2008
Determination of chlorfluazuron residues in vegetablesforimport and exportHighperformanceliquidchromatographicmethod
This standard specifies the method of sample preparation and determination of chlorfluazuron resi-dues invegetablesbyhighperformanceliquidchromatography.This standard is applicable to the determination of chlorfluazuron residues in cucumber, radish andgreen soy bean.
Principle
Chlorfluazuronresidue isextractedbyacetonitrilefromthesample.Cleanuptheextractionbypassing througha florisil column.Determine chlorfluazuron by HPLC with diode array detector and quan-titatewithexternal standardmethod.3
Reagents and materials
Unless specified notes,all reagents are analyticallypure,\water\is redistilled or deionized.3.1
Acetonitrile:HPLC grade
n-Hexane:HPLCgrade
Ethyl ether:HPLC grade
Sodium chloride.
Anhydrous sodium sulfate:Lgnite at 65o for 4 h,and keep in a tightly closed container aftercooling.
Eluent:n-Hexane+ethylether(2+8,V/V).Absorbentcotton.
SN/T2095—2008
3.8Florisil:Granulesize0.075mm~0.15mm(100~200mesh).igniteat650℃for4h,andkeepin a tightly closed container,then heat for 4 h at 130 ℃ in an oven,cooling to room temperature indesiccator and adding 1% of water before use.3.9Columnforclean-up:200mmX15mm(i.d.)glasscolumn,packedwith5mmabsorbentcottonat the bottom of the column.fill in 20 mm anhydrous sodium sulfate,10 g florisil,and 20 mm anhydroussodiumsulfateattop.Add20mLn-hexanetowashbeforeuse.3.10
Chlorfluazuronstandard:Purity≥99.0%Chlorfluazuron standard stocksolution:Accurately weigh an appropriate amountof Chlorflua-3.11
zuron standard and dissolve with methanol to prepare a standard stock solution of 1oo μg/mL.Thisstandard stock solution should be stored at o ℃ ~4 ℃. The standard stock solution is stable in sixmonths.
Chlorfluazuron standard working solution:According to the requirement,pipette adequate a-mount of standard stock solution(3.12) and dilute with methanol to prepare standard working solu-tionof suitableconcentration.4
Apparatusandequipment
Highperformance liquid chromatograph,equipped withdiodearraydetector.Highspeedblender
Rotaryvacuumevaporator.
Vortex mixer.
Centrifuge(4000 r/min).
5Preparationand storageoftestsampleThe combined sample is reduced to 1kg,the edible portion is blended,and divided into two equalportions.Each portion is placed in a clear container as the test sample,which is sealed and labled.Thetest sample shouldbe stored below-18 .In the course of samplepreparation,precautionmust be taken to avoid the contamination or any factors which may cause the change of residue con-tent.
SN/T2095—2008
Determinationprocedure
Extraction
Weigh ca 5 g test sample(accurate to 0.01 g) in 100 mL beaker. Add 30 mL acetonitrile and homoge-nize the sample with the blender at 10 000 r/min for2 min.Filtrate the extraction to 50 mL centrifu-gal tube. Wash the beaker and the blender twice with 15 mL acetonitrile aggregately. Filtrate thewashing solution to the centrifugal tube. Add some Sodium chloride into the filtrate and shake thetube vigrously.Centrifugate(4000 r/min)for5minandcollect the organicphase.Add20mLaceto-nitrile to water phase to extrat again. Combine acetonitrile phase and evaporate the organic phase todrynessat45℃.
Cleanup
Dissolve the residue for three times,using 5 mL eluent(3. 6) each time,and transfer all the solutionto column for clean-up(3.9),elute the column with 100 mL eluent(3. 6). Collect all the eluted solu-tion and evaporate the solution to dryness at 35 C. Dissolve the residue with 1. 0 mL mobile phase(acetonitrile+water=60+40.V/V)and filtrate the solutionby0.45μm filterforhigh-performanceliquidchromatographicdetermination.6.3
Determination
HPLCconditions
Chromatographiccolumn：C1.5μm,250mmx4.6mm(i.d.),orequivalent；Mobile phase:for gradient elute condition,see table 1;b)
Mobilephaseflowrate：1.0mL/min;Detectingwavelength：258nm;
Injection volume: 20 μL.
Table1-Gradienteluteconditiontime/min
water/%
acetonitrile/%
6.3.2HPLCdetermination
SN/T2095—2008
According to the approximate concentration of chlorfluazuron in the sample solution,select thestandard working solution with similar concentration to that of sample solution. The responses ofchlorfluazuron in the standard working solution and the sample solution should be in the linear rangeof the instrumental detection.The standard solution should be randomly injected between the injec-tions of sample solution of equal volume.Under the above operating conditions,the retention timeof chlorfluazuron is about 17.3 min. For the chromatogram of the standard,see figure A. 1 in annexA,andfortheUv spectrumofthestandardseefigureA.2inannexA.6.4 Blank test
The operation of the blank test is the same as the method described in the determination procedurebut with the omission of sample addition.7
Calculation and expression of resultThe calculation of result is carried out by data processor or according to formula(1),and the blankvalue shallbe subtracted fromthe resultof calculation.x=
X-the residue content of chlorfluazuron in the test sample,mg/kg;Athepeak areaof chlorfluazuron insamplesolution;cthe concentration of chlorfluazuron in the standard working solution,μg/mL;Vthefinalvolumeof samplesolution,mL;Asthepeakareaof chlorfluazuroninthestandardworkingsolution;m-thecorrespondingmassofthetestsampleinthefinalsamplesolution.g.8Limitofquantification(LOQ)andrecoveryLimitof quantification
The limit of determination of this method is 0. 05 mg/kg8.2
Recovery
Recoveryseetable2
SN/T2095—2008
Sample
cucumber
radish
green soy bean
0.05mg/kg
86.5%~102.0%
88.7%~104.2%
86.6%~102.1%
Table2—Recovery
Fortifying concentrations
0.10mg/kg
87.5%~103.2%
87.5%~102.3%
87.4%~105.7%
0.20mg/kg
86.8%~103.2%
87.6%~102.9%
88.4%~102.8%
83. 3% ~106. 9%
92. 1% ~108.5%
82.6%~107.4%
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