7 6 Vol. 7 No. 6 2016 6 Journal of Food Safety and Quality Jun., 2016 - - B 1, 薛昆鹏 2 1,, 金小青 2, 余冲 2, 赵岳星 (1., 321004; 2., 321014) 1, 2* 摘要 : 目的 - - B 方法 20% -, Welchrom P-SCX Welchrom, Welch Ultimate XB-C 18 (250 mm 4.6 mm, 5 μm) 75% -, 结果 Welchrom P-SCX,,, Welchrom P-SCX B 1.0~100.0 μg/l (r 2 0.9995), 2 91.4%~106.5%, RSD 2.1%~4.3% 结论 B,, B 关键词 : ; - ; ; B Determination of rhodamine B in pepper products by solid phase extraction high performance liquid chromatography-fluorescence detector XUE Kun-Peng 1, 2, JIN Xiao-Qing 1, 2, YU Chong 2 1, 2*, ZHAO Yue-Xing (1. Zhejiang Normal University Institute of Physical Chemistry, Jinhua 321004, China; 2. Welch Materials (Zhejiang) Co., Ltd., Jinhua 321014, China) ABSTRACT: Objective To establish a method for the determination of the rhodamine B (RB) in pepper products by solid phase extraction high performance liquid chromatography-fluorescence detector (SPE-HPLC-FLD). Methods The sample were extracted by acetonitrile and 20% acetone-n-hexane solution, then purified and concentrated with a strong positive ion exchange SPE column (Welchrom P-SCX) and a neutral alumina SPE column (Welchrom Alumina). The HPLC separation was performed on a Welch Ultimate XB-C 18 HPLC column (250 mm 4.6 mm, 5 μm), eluted with a mobile phase consisting of methanol and water (75:25, V:V). The analyzed sample was detected by fluorescence detector. The content of RB in pepper products was measured using peak area external standard method. Results The results showed that the method of using Welchrom P-SCX SPE column to pre-treat sample had higher efficiency, stability and better repeatability. The calibration curve showed a good linearity in the range of 1.0~100.0 µg/l with correlation coefficient (r 2 ) greater than 0.9995. The recoveries were 91.4%~106.5% at three spiked levels with the relative standard deviations (RSDs) of 2.1%~4.3%. Conclusion This 基金项目 : (2014C01025) Fund: Supported by Major Science and Technology Key Industrial Projects (2014C01025) * 通讯作者 :,, E-mail: eugenezhao@zjnu.com *Corresponding author: ZHAO Yue-Xing, Professor Level Senior Engineer, Zhejiang Normal University Institute of Physical Chemistry, No 688, Yingbin Road, Jinhua 321004, China. E-mail: eugenezhao@zjnu.cn
2376 7 method is sensitive, accurate and precise, which is very in accordance with technical standards of the RB detection. It is suitable for routine detection of the RB in pepper products. KEY WORDS: solid phase extraction; high performance liquid chromatography-fluorescence detector; pepper products; rhodamine B 1 引言 B(rhodamine B) B, [1],,, [2] (IARC) : B, [3], [4], 2008 B, [5,6], B,, B, B C 18 [7,8] [9,10] ( N- ) [11] (GPC) [12] [13,14], C 18,, ; (SPE),, ; SN/T 2430-2010 B GPC,,, B [15], 1 C=N N,,, 1%, Welchrom P-SCX SPE, - (HPLC-FLD) B,,, B,, B 2 材料与方法 2.1 材料 1 B Fig. 1 Structure of rhodamine B 2.1.1 样品来源, 5, 2.1.2 主要仪器与试剂 (Shimadzu LC-20AD, ); ( ); (); XW-80A ( ); 12 ( ) B( 99.0%) Dr. Ehrenstorfer, HPLC, Merck, ( ) Welchrom P-SCX Welchrom, : 0.0100 g B 100 ml, 50% -,, 0.1 mg/ml B 2.2 方法 2.2.1 辣椒制品前处理 (1) ( 2 mm) 1.0 g 25 ml, 10 ml, 10
6, : - - B 2377 min, ; 10 ml 2, 3 50, 6 ml 1%, Welchrom P-SCX SPE, SPE ( 2 mm) 1.0 g 25 ml, 10 ml 20% -, 10 min, 2, 3, Welchrom SPE, SPE (2) 1.0 g 25 ml, 10 ml, 10 min, ; 10 ml 2, 3 50, 6 ml 1%, Welchrom P-SCX SPE, SPE 1.0 g 25 ml, 10 ml 20% -, 10 min, 2, 3, Welchrom SPE, SPE 2.2.2 辣椒制品净化 (1) Welchrom P-SCX SPE 4 : 1 Welchrom P-SCX SPE 5 ml, 5 ml, ; 2 2.2.1 Welchrom P-SCX SPE, ; 3 5 ml, 10 ml, ; 4 10 ml 2% -, ; 5 50, 50% - 1 ml, 0.22 µm PTFE, HPLC-FLD (2) Welchrom SPE 4 : 1 Welchrom SPE 10 ml 20% -, ; 2 2.2.1 ; ; 3 30 ml 20% -, ; 4 10 ml 80% -, ; 5 80% - 10 ml, 0.22 µm PTFE, HPLC-FLD 2.3 色谱条件 : Welch Ultimate XB-C 18 (250 mm 4.6 mm, 5 μm); : : =75: 25; : 1.0 ml/min; : 20 μl; : 30 ºC; λ ex =550 nm, λ em =580 nm 2.4 2 种不同类型 SPE 小柱回收率对比试验 6, 4, 3, 1.0 g, 0.1 ml 100 ng/ml B Welchrom P-SCX Welchrom 2 SPE 2.2.1 2.2.2, 2.3, (RSD), 2 SPE 2.5 定性定量方法,, 100 ng/ml B - : =75: 25 0 0.5 1.0 2.0 5.0 10.0 50.0 80.0 100.0 ng/ml, 2.3, 3, (ng/ml), 3 结果与分析 3.1 提取溶剂的选择 B 3 > > B :,, ;, 1%,, 50, 1%, B, Welchrom P-SCX, 2 : 2a ; 2b (); 2c, 3 2 ; 2d, 3 B 2, Welchrom,,, [16], -, Welchrom,, 3 : 3a ; 3b (); 3c ( ), 3d ()
2378 7 2 Welchrom P-SCX (a: ; b: ; c: ; d: ) Fig. 2 The purification process of Welchrom P-SCX SPE column (a: sample extracting; b: loading; c: washing; d: eluting) 3 Welchrom (a: ; b: ; c: ; d: ) Fig. 3 The purification process of Welchrom Alumina SPE column (a: sample extracting; b: loading; c: washing; d: eluting) 3.2 检测器及检测波长的选择 HPLC B,,,,, B, HPLC-FLD 3.3 2 种 SPE 前处理方法的对比, β- [17],,, SPE B, ; C=N N,, Welchrom P-SCX Welchrom 2 SPE, 1 : Welchrom SPE Welchrom P-SCX SPE, ( 4 ), SPE B,, 4%, SPE, Welchrom P-SCX SPE,,,,, 1, Welchrom P-SCX SPE 3.4 标准曲线线性范围 20 µl,, : B 0.5~100 ng/ml; (Y, X, ng/ml) Y=43631.8 X+23531, 0.9995, 5 B 3.5 方法检测限, 3 (S/N 3), B 0.2 ng/g 3.6 回收率和精密度 9, 3, 1.0 g( 0.001 g),
6, : - - B 2379 Table 1 表 1 Welchrom P-SCX 和 Welchrom 中性氧化铝的净化效果比较 (n=5) Comparison of purification effects between Welchrom P-SCX and Welchrom Alumina SPE column (n=5) SPE (mg/kg) (µg/kg) (%) RSD/% P-SCX - 10.0 101.0 3.6-10.0 83.9 8.4 P-SCX - 10.0 98.4 4.3-10.0 85.2 9.7 3.7 实际样品测定 5 5, 10 B 6, 表 2 罗丹明 B 加标回收率结果 (n=3) Table 2 Recovery results of rhodamine B (n=3) (mg/kg) (µg/kg) (%) RSD/% - 5.0 91.4 3.6-10.0 97.9 2.8 4 Welchrom (a) Welchrom P-SCX (b) Fig. 4 Chromatograms of spike sample for Welchrom Alumina (a) and Welchrom P-SCX (b) SPE column - 50.0 104.0 2.1-5.0 93.2 4.3-10.0 98.7 3.1-50.0 106.5 2.3 5 B Fig. 5 Chromatogram of rhodamine B standard solution 100 ng/ml B 0.05 0.1 0.5 ml, 2.2.1 2.2.2,, 2 2, 91.4%~106.5%, 2.1%~4.3%, 6 Fig. 6 Chromatogram of dry red pepper sample 4 结论 B (1%), Welchrom P-SCX
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