6 3 Vol. 6 No. 3 2015 3 Journal of Food Safety and Quality Mar., 2015 17 何曼莉, 谢建军 *, 陈捷, 卢丽, 陈文锐 (, 510623) 摘要 : 目的 (gel permeation chromatography, GPC) (solid phase extraction, SPE) ( ),,,,,, 17 方法, (1:1, v:v),,, ( µecd ) 结果 17 10~100 µg/kg, R>0.999,, 17 84.6%~115.5%, (RSD, n=6) 3.1%~14.5% (LOQ) 0.31~4.15 µg/kg 结论, 17 关键词 : ; ; ; ; Determination of 17 organochlorine pesticide residues in traditional Chinese medicinal herbs by gel permeation chromatography-solid phase extraction-gas chromatography method HE Man-Li, XIE Jian-Jun *, CHEN Jie, LU Li, CHEN Wen-Rui (Guangdong Entry-Exit Inspection and Quarantine Bureau, Guangzhou 510623, China) ABSTRACT: Objective To establish an analytical method for detecting hexachlorocyclohexane, dichlorodiphenyltrichloroethane, endosulfan, heptachlor, chlordane, aldrin, dieldrin, endrin and their metabolites residues in traditional Chinese medicinal herbs including plumeria, dried date, ginseng, dangshen, radix ginseng rubra, cordate houttuynia and astragalus with gel permeation chromatography-solid phase extraction-gas chromatography method. Methods Traditional Chinese medicinal herbs were crushed and immersed in deionized water, extracted with ethyl acetate and acetone (1:1, v:v), and then purified by gel permeation chromatography and solid phase extraction, and detected by gas chromatography (with μecd detector). Results The linear range of those 17 organo-chlorine pesticides were from 10 to 100 µg/ml with 基金项目 : (201410177-02) Fund: Supported by the Project of General Administration of Quality Supervision, Inspection and Quarantine of the People's Republic of China, Special Found for Public Science and Technology Program (201410177-02) * 通讯作者 :,,, E-mail: jianjunxjj@126.com *Corresponding author: XIE Jian-Jun, Senior Engineer, Guangdong Entry-Exit Inspection and Quarantine Bureau, No.66, Huacheng Road, Tianhe District, Guangzhou 510623, China. E-mail: jianjunxjj@126.com
894 6 the correlation coefficients (R) more than 0.999. The recoveries of the 17 compounds ranged from 84.6% to 115.5% with the relative standard deviation (RSD, n=6) lower than 14.5%, and the limit of quantitation (LOQ) of the method ranged from 0.31 to 4.15 µg/kg. Conclusions The experimental method was simultaneous, accurate, efficient, environmentally friendly, and suitable for the rapid detection of 17 organochlorine pesticides in traditional Chinese medicinal herbs. KEY WORDS: traditional Chinese medicinal herbs; organochlorine pesticides; gas chromatography; gel permeation chromatography; solid phase extraction 1 引言,, [1],, [2], [3,4] 2001 5 127 ( ), 12, (DDT ) 9 [5],,,, [6] [7,8] 300, 2012,, 7 36, 35, 32 3 [9],, 0.05~0.3 mg/kg, 1 mg/kg 3 mg/kg [10],,, ( ) [11] 2010,, GC-NIC-MS,, [12],,, ;,,,, ;,, [13-15],,, 2 材料与方法 2.1 仪器和试剂 (HP6890, ); (AccuPrep TM/AccuVap, J 2 Scientific ) ; (BT124, Srtorius); (3-16PK, Sigma); (Turbovap LV, Caliper life sciences); (NH 2, 200 mg/3 ml, ); (GCB, 250 mg/ 3 ml, Scientific Instrument); (Advantage A10, Milli-Q) : α-bhc(98.0%), β-bhc(98.5%), γ-bhc(98.6%), α- (97.0%), β- (99.5%), (99.5%), (99.6%), (98.0%), (99.0%), (98.0%), (99.0%), (99.0%), Dr. Ehrenstorger ;
3, : - - 17 895 δ-bhc(gerilliant, 99.1%), p,p -DDE(100 µg/ml), o,p -DDT(100 µg/ml), p,p -DDD (100 µg/ml), p,p -DDT(100 µg/ml), ; (, ), (, ); : 2.2 标准工作液的配制 0.01 g α-bhc β-bhc γ-bhc α- β- 10 ml, ( ) 1 mg/ml ( ), 4, 2 2.3 色谱条件 : Agilent HP-1701 (30 m 0.32 mm, 0.25 µm); : 5.0 ml/min 7 min, 2 /min 3.0 ml/min, 5 min; : 80 1 min, 40 /min 180, 3.0 min, 30 /min 250, 10.0 min; : 250 ; : 250 ; : N 2 ( >99.99%); : 1.0 µl (GPC) : : 300 mm 25 mm; : Bio-Beads, S-X3, 38~75 μm; : 45 ; : - (50:50, v:v); : ; : 5 ml/min; : 5.0 ml 2.4 样品处理 2.4.1 提取 2.00 g 50 ml, 10 ml 20 min, : (1:1) 15 ml, 8 g, 10 min, 4500 r/min 5 min, 50 ml, 10 ml : (1:1), 2.4.2 净化 10 ml 0.5 ml, : (1:1) 10 ml, GPC, 10~18 min,, : (8:2) 2 ml; (200 mg/3 ml), 1 g, : (8:2)4 ml, ; GPC, : (8:2) 6 ml 3,, 1 ml GCB (250 mg/3 ml) : (8:2) 4 ml,,, : (8:2)6 ml 3,, 0.5 ml, 1 ml, 0.5 ml, 1 ml, 2.5 标准曲线制作, (α-bch β-bch γ-bch δ-bch p,p -DDE o,p -DDT p,p -DDD p,p -DDT α- β- α- β- ), 10 20 30 40 50 100 ng/ml 3 结果与讨论 3.1 提取溶剂的选择,, 25 ml, 60%~70% 30%~40% 50%~80% 110%~140%,,,, 25 ml : (4:1) : (1:1) : (3:1), 50%~70% 90%~110% 110%~140%, 1,,,,, : (1:1), : (1:1)
896 6 表 1 不同溶剂对 17 种农药的提取效果 Table 1 The extraction of pesticides in different solvents : (4:1) : (1:1) : (3:1) α-666 70.2% 108.1% 139.8% β-666 70.5% 108.4% 140.1% γ-666 70.7% 108.2% 140.4% δ-666 69.1% 108.6% 140.6% p,p -DDE 65.1% 110.3% 138.2% o,p -DDT 63.2% 110.1% 138.7% p,p -DDD 64.5% 110.5% 138.5% p,p -DDT 62.0% 109.5% 138.6% α- 50.2% 90.2% 110.8% β- 50.5% 90.4% 110.1% 50.8% 90.6% 110.6% 52.3% 92.1% 121.7% 52.7% 92.8% 122.8% 51.2% 95.2% 131.8% 52.0% 94.8% 132.3% 51.3% 95.8% 135.7% 53.6% 93.4% 138.5% 3.2 GPC 净化条件的选择,, GPC 17,,, 600~1080 s, 98%~125%, 5 1 1 5 GPC Fig. 1 The elution curves of 5 organochlorine pesticides by GPC 3.3 固相萃取柱 (SPE) 类型及淋洗液量的选择 3.3.1 SPE 类型的选择,, GPC ;,, (GCB SPE, 250 ml), (NH 2 SPE, 200 mg/3 ml), (Florisil SPE, 1.5 g/3 ml), (Al 2 O 3 SPE, 2.0 g/3 ml) 10 ml + (8:2), 50 ng/ml 4 SPE 108%~154% 86%~113% 99%~119% 74%~104% Al 2 O 3 SPE 4 ml : (8:2), 10 ml ( 2,, ), Al 2 O 3 SPE, Al 2 O 3 SPE Fig. 2 2 GCB, NH 2, Florisil, Al 2 O 3 SPE (A:, B: α- ) The elution curves of heptachlor and α-endosulfan with 4 kinds of SPE(A: heptachlor, B: α-endosulfan)
3, : - - 17 897, SPE,, 3 SPE, (NH 2 SPE, 200 mg/3 ml) GCB (GCB SPE, 250 ml),,,, 3, GCB,,,,, (NH 2 SPE, 200 mg/3 ml) GCB(GCB SPE, 250 ml) 3.3.2 SPE 柱淋洗液用量的选择 GCB 17 2 ml : (8:2), 4 (NH 2 SPE, 200 mg/3 ml) GCB(GCB SPE, 250 ml) 6 ml 17,, 6 ml : (8:2) 3.4 线性关系 2.3, (10 20 30 40 50 100 µg/l) α-666 γ-666 β-666 δ-666 α- p,p -DDE o,p -DDT p,p -DDT β- p,p -DDD,, 17 (X) 10~100 ng/ml (Y), 2 3 NH 2 SPE GCB SPE Florisil SPE Fig. 3 Chromatogram of astragalus purified through 3 types of SPE cartridges (NH 2 SPE, GCB SPE, Florisil SPE) combination with each other(a. GCB SPE+ Florisil SPE; B. NH 2 SPE+ Florisil SPE; C. NH 2 SPE+ GCB SPE)
898 6 4 NH 2 SPE GCB SPE Fig. 4 The elution curves of the organochlorine pesticides with the combination of NH 2 SPE and GCB SPE cartridges 3.5 检出限与定量限 2.4, 3 17 (LOD), 10 (LOQ),, 17 0.09~1.25 µg/kg, 0.31~4.15 µg/kg 3 3.6 准确度与精密度 6 ( ), 10.0 20.0 100 µg/kg ( 8, 20, 50 µg/kg) α-666 γ-666 β-666 δ-666 α- p,p -DDE o,p -DDT p,p -DDT β- p,p -DDD,,, 6,, 3 3, 17 84.6%~115.5%; RSD 3.1%~14.5% 17, 5 3.7 实际样品测定 GC-GPC-SPE 20, 17, 26, 6, 3 3 Table 2 表 2 17 种有机氯农药标准品回归曲线汇总表 Summary of the 17 organochlorine pesticides standards regression curves (ng/ml) 1 α-666 Y=929807.17X-158.55 0.9994 10~100 2 γ-666 Y=723741.16X-106.70 0.9994 10~100 3 Y=485701.83X-324.57 0.9994 10~100 4 Y=793413.16X-373.04 0.9995 10~100 5 β-666 Y=329223.00X+65.52 0.9997 10~100 6 δ-666 Y=684993.19X+43.78 0.9991 10~100 7 α- Y=608183.73X+113.96 0.9999 10~100 8 - Y=692509.29X-96.53 0.9996 10~100 9 - Y=663434.97X+23.78 0.9992 10~100 10 p,p -DDE Y=663382.71X+7.54 0.9997 10~100 11 Y=668619.87X-191.53 0.9999 10~100 12 Y=303175.33X-389.22 0.9999 10~100 13 o,p -DDT Y=51287.54X-56.87 0.9994 10~100 14 p,p -DDT Y=780676.65X-22.53 0.9996 10~100 15 β- Y=597867.11X+137.19 0.9999 10~100 16 p,p -DDD Y=28019.90X-28.27 0.9998 10~100 17 Y=455488.08X+65.21 0.9993 10~100
3, : - - 17 899 表 3 17 种化合物加标回收率及相对标准偏差 (RSD) 以及检出限和定量限 Table 3 Recoveries, RSD, LOD and LOQ of the 17 organochlorine pesticides /(µg/kg) /% RSD(n=6)% LOD(µg/kg) LOQ(µg/kg) a-666 8,20,50 89.9,103.9, 91.8 10.8,11.5,7.4 0.09 0.31 r-666 8,20,50 98.6, 93.4, 98.9 11.6, 6.4, 5.6 1.25 4.15 8,20,50 93.8,107.7, 99.9 5.8, 8.4, 8.0 0.14 0.46 8,20,50 102.6,104.1,115.5 12.2, 9.7,14.0 0.33 1.1 β-666 8,20,50 92.2,111.2,100.4 6.5,10.7, 8.0 0.12 0.39 δ-666 8,20,50 107.9,112.4, 90.7 9.1,14.5,13.6 0.31 1.02 a - 8,20,50 89.4,107.9, 89.7 4.5,11.2, 6.7 0.17 0.58-8,20,50 95.0,107.3, 90.2 3.1, 8.4, 9.3 0.19 0.63-8,20,50 98.9,107.8, 92.4 5.3,10.9, 8.8 0.17 0.56 p,p -DDE 8,20,50 84.6, 96.9, 95.7 5.5, 9.3, 8.0 0.19 0.62 8,20,50 99.3, 93.3, 96.2 5.8, 8.0,11.9 0.18 0.61 8,20,50 88.7, 99.3, 96.7 6.3, 7.6, 8.5 0.23 0.77 o,p -DDT 8,20,50 90.7, 93.1, 99.0 7.9,10.3, 6.9 0.76 2.53 p,p -DDT 8,20,50 85.0, 85.2, 96.2 11.1, 7.3,13.9 0.91 3.05 β-- 8,20,50 97.5,106.0,102.3 6.1, 9.6, 7.1 0.30 0.99 p,p -DDD 8,20,50 104.0,103.3, 94.1 4.7, 6.4, 8.2 0.30 1.01 8,20,50 102.0, 96.8, 89.7 5.1,10.5, 8.3 0.95 3.17 5 (A) 17 (10 µg/l,b) (10 µg/l,c) ( 2 ) Fig. 5 The chromatogram of Astragalus purified(a), Standard (10 µg/l, B) of organochlorine pesticides and Astragalus spike recovery(10 µg/l,c)
900 6 4 结语 GC-GPC-SPE 17 (α-666 γ-666 β-666 δ-666 α- p, p -DDE o, p -DDT p, p -DDT β- p, p -DDD ),, 参考文献 [1],. [J]., 2005, 30(6): 405 409. Hao LL, Xue J. Multiresidue analysis of organochlorine pesticides and the application to Chinese herbal medicine [J]. China J Chin Mater Med, 2005, 30(6): 405 409. [2],,. [J]., 2003, 22(5) : 15. Xu L, Liu YX, Bao WK. Advances in study of organochorine pesticides in organisms [J].Sichuan Environ, 2003, 22(5): 15. [3] Pardio V, Martinez D, Flores A, et al. Human health risk of dietary intake of organochlorine pesticide residues in bovine meat and tissues from Veracruz, Mexico [J]. Food Chem, 2012, 135( 3) : 1873 1893. [4],,,. [J]., 2014, 34(3): 549 543. Zeng HH, Qin RQ, Mo LY, et al. Research on tocixity of organochlorine pesticides to human health [J]. J Guilin Univ Technol, 2014, 34(3): 549 543. [5],,,. [J]., 2013, 32(2): 347 353. Li B, Xie QL, Liao T, et al. Contamination characteristics and risk evaluation of organochlorine pesiticide in surface sediments from Zhalong wetland, China [J]. J Agro-Environ Sci, 2013, 32(2): 347 353. [6]. [J]., 2008, 10(3): 3 7. Ran MX. Development status and suggestions of Chinese herbal planting in China [J]. Mod Chin Med, 2008, 10(3): 3 7. [7],,. [J]., 2000, 2(8): 11 13. Chen J, Cheng HZ, Ding WL. Scientific application of pesticides at standardization of Chinese herbal medicine production [J]. Mod Chin Med, 2000, 2(8): 11 13. [8],,. [J]., 2005, 45(5): 302 308. Rong WG, Guo H, Yang H. Current research status in China on pesticide contamination of plant material used in making Chinese herbal medicines [J]. Chin J Pestic, 2005, 45(5): 302-308. [9]. 2012 [EB/OL]. http://www.greenpeace.org/china/zh/publications/ reports/food-agriculture/2013/herbs-rpt/ The World Organization of Greenpeace. Pesticides in Chinese herbal medicines at 2012 the investigation report on pesticide pollution of Chinese herbal medicines [EB/OL]. http://www.greenpeace.org /china/ zh/publications/reports/food-agriculture/2013/herbs-rpt/ [10]. [J]., 2013, 15(8): 633 637. Yang CR. The Current situation and countermeasures of pesticides in TCM herbs [J]. Mod Chin Med, 2013, 15(8): 633 637. [11],. [J]., 2011, 46(5): 376 377. Liang WQ, Pu JB. Determination of organochlorine pesticide residues in Chinese herbal medicines by GC [J]. Zhejiang J Tradit Chin Med, 2011, 46(5): 376 377. [12],,,. GC-NCI-MS[J]., 2009, 15(5): 34 36. Liu L, Guo YZ, Zhang YT, et al. Analysis of organochlorine pesticides in Chinese herbal medicines by GC-NCI-MS [J]. Tianjin Agr Sci, 2009, 15(5): 34-36. [13],,,. [J]., 2011, 30(1): 108 114. Song SL, Rao Z, Ma XD, et al. Extraction techniques of organochlorine pesticides in different matrixes and their development trends [J]. J Instrum Anal, 2011, 30(1): 108 114. [14],,,. 50 [J]., 2011, 46(15): 1204 1209. Huang X, Xue J, Wu XB, et al. Study on analysis method for 50 organochlorine and pyrethroid pesticides pesidues in glycyrrhiza [J]. J Chin Pharm Sci, 2011, 46(15): 1204 1208. [15],,. GC [J]., 2011, 7(10): 42 45. Guan ZM, Guo XL, Lu ZZ. Determination of organochlorine pesticide in Ginseng by gas chromatography [J]. Asia-Pacific Tradit Med, 2011, 7(10): 42 45. ( 责任编辑 : 张宏梁 )
3, : - - 17 901 作者简介 何曼莉, 工程师, 主要研究方向为农药残留分析 E-mail: heml@iqtc.cn 谢建军, 博士, 高级工程师, 主要研究方向为农药残留分析 E-mail: jianjunxjj@126.com 现代发酵工程在食品工业中的应用 专题征稿,,,,,,,,,,,,,, :,,,,,,,, 2015 5,, 2015 4 15 E-mail, 投稿方式 : : www.chinafoodj.com E-mail: tougao@chinafoodj.com 食品安全质量检测学报 编辑部