5 5 Vol. 5 No. 5 214 5 Journal of Food Safety and Quality May., 214 22 * 潘小红 (, 411) 摘要 : 目的 (ultra performance liquid chromatography/time-of-flight mass spectrometry, UPLC-Q-TOF-MS-MS) 22 方法, /, ACQUITY TM UPLC HSS T3(5 mm 1.8 mm 1.7 μm),.1-, (ESI),, 22 结果 22, 22, 1 ng, 结论,, 关键词 : ; ; Rapid identification of 22 drugs illegally added into improving sleep health foods by ultra performance liquid chromatography/time-of-flight mass spectrometry method PAN Xiao-Hong* (Hunan Institute for Food and Drug Control, Changsha 411, China) ABSTRACT: Objective To establish a rapid ultra performance liquid chromatography/time-of-flight mass spectrometry method to identify 22 kinds of drugs illegally added into improving sleep health foods. Methods A method of the ultra performance liquid chromatography-mass spectrometry (UPLC-MS/MS) was used to detect the 22 kinds of drugs illegally, samples were extracted with methanol. By the separation column of ACQUITY TM UPLC HSS T3 (5 mm 1.8 mm 1.7 μm), the gradient elution of the mobile phase consisted of.1 formic acid acetate solution and the mass spectrometer conditions (ESI) as operated in positive or negative ion mode and in two scan modes including full scan MS. Results A good resolution of 22 drugs illegally added into the food for improving sleep health was obtained by UPLC-MS/MS, of which the determination limits were all below 1. ng. Conclusion The method which showed sufficiently selective and sensitive could be used to detect illegal drugs in the food for improving sleep health KEY WORDS: ultra performance liquid chromatography/time-of-flight mass spectrometry; improving sleep health foods, illegal adding 基金项目 :,,, E-mail: panxiaohong56781@ 163.com *Corresponding author: PAN Xiao-Hong, Senior Nutritionist, Research Assistant, Major research in the inspection, research and development of food, medicine, health food and cosmetics. Hunan Institute for Food and Drug Control, No.6, Baxi Road, Furong District, Changsha 411, China. E-mail: panxiaohong56781@163.com.
5 : 22 1525,,,, ( ), 213 5 16,, [1],,,, 3, 2924 2124 2132, 15 4 3 [2-4], -,,, (UPLC-Q- TOF-MS-MS), (UPLC), (Q-TOF- MS-MS), [5-7], 22, 1 仪器与试剂 Waters Q-TOF Premier TOF, ESI ; Waters Acquity TM ; R-215 Buchi 22 1, Merck : ( ) 表 1 22 种标准品的详细信息 Table 1 The detailed information of 22 standard substance () 1 8717 99.5 / Dr 2 1219-12 / / () 3 171217-242 / / 4 171248-231 / / 5 171223-242 / / 6 148-241 UV 7 123-21 / 8 187-281 / 9 17125-212 99.9 / 1 171229-232 / 15 3 h 11 171253-241 / 12 167-241 / / 13 122-921 / / 14 171225-232 / / 15 171218-263 / / 16 1543-241 / 17 11774-257 / / 18 1452-231 / 19 171227-232 / / 2 147-266 99.7 15 2 h 21 171222-265 / 22 171221-242 / /
1526 5 2 实验方法 2.1 色谱条件 : ACQUITY TM UPLC HSS T3(5 mm 1.8 mm 1.7 μm), Agilent EC-C 18 (15 mm 2.1 mm, 2.7 μm), Halo TM C 18 (1 mm 2.1 mm 2.7 μm), ACQUITY TM UPLC BEH C 18 (1 mm 2.1 mm 1.7 μm); 3 ; 5 μl;.3 ml/min,.1 (A)- (B), 2.2 质谱条件 ESI ; 3 kv; 3 V; 3 V;.1 V; 35 ; 7 L/h; 13~35 V, () 5~5 (ESI+): 17 (ESI-): 5 2.3 标准品的配制 2.3.1 标准品储备溶液的配制 1 22 1. mg, 5 ml,, 2.3.2 混合标准品溶液的配制,.1~2. μg/ml 22, (ESI+ ESI-) 2.4 供试品的制备 2.4.1 固体试样 1,, (,, )(, ), 5. ml,, 3 min( 25 W, 33 khz),,,,, 1. ml 1 ml,, 2.4.2 液体试样 5~1,,,,, 2.4.1 5. ml 2.5 样品测定 5 μl, 3 结果与分析 3.1 液相色谱条件的优化 4, 4,, ACQUITY TM UPLC HSS T3(5 mm 1.8 mm 1.7 μm), 2924 - -.5 (ESI+) - (ESI-), 2124 2132 -.1.2 mol/l,,, -.1, 2,, 1 1, ESI+ ESI-, 表 2 不同电离模式中流动相梯度洗脱时间表 Table 2 The schedule the gradient elution in different ionization modes /min /(ml min 1 ) A B Curve /min) /(ml min 1 ) A B Curve.25 9 1 / 7.25 7 3 6 1.25 9 1 11 7.1.25 45 55 6 2.25 7 3 6 9.25 45 55 6 3.25 9 1 6 9.1.25 9 1 6 4.25 7 3 6 1.25 9 1 6 :.1 (A)- (B)
5 : 22 1527 3.2 质谱条件的优化.4 μg/ml, ESI,,, 3 V, MS/MS, 2, 3 3.3 22 种化学物质的谱图特性, 22, 3, 4 Fig. 1 1 ( : 1-ESI+, 2-ESI-) Total ion-current grams of standard substance (Note: 1-ESI+, 2-ESI-) Table 3 表 3 22 种改善睡眠类的质谱条件优化结果 Results of best choices of MS conditions of 22 kinds of drugs illegally added improving sleep health foods (component) (Min) (ps eudomolecular ions)/() (collision energy)/ev (fragment ions from full scan MS/MS)/() (ng) 1 2.4 33.33 35 27.15, 181.16, 91.11.22 2 2.62 389.29 13 245.15, 143.16, 99.15.273 3 2.87 3.23 18 283.22, 227.17.13 4 2.77 275.27 15 23.19.52 5 2.99 278.35 2 121.14, 215.25, 58.11.15 6 3.14 356.35 25 192.2, 165.18.22 7 3.16 233.25 13 174.18, 216.21.114 8 4. 326.24 32 291.26, 29.17, 129.13.45 9 5.49 274.16 2 154.13, 13.5.765 1 6.28 36.28 26 236.21, 264.25, 29.19.49 11 6.62 282.22 26 236.21, 27.2, 18.18.12 12 6.77 2.22 26 267.19, 25.18, 138.9.18 13 6.89 287.2 2 241.17, 269.18, 14.11.2 14 7.47 39.24 32 281.21, 25.18, 165.11.45 15 7.64 321.17 2 275.15, 33.15.69 16 7.79 343.21 32 38.23, 239.16, 138.9.419 17 8.13 285.22 32 193.19, 222.23, 154.13.21 18 3.3 183.32 2 14.3, 96.29.4454 19 5.49 231.32 21 188.32, 144.32, 85.21.8386 2 6.97 225.37 24 182.36, 138.36, 85.21.7529 21 7.4 314.28 22 286.27, 278.31, 248.31.463 22 7.74 237.37 25 194.37, 15.37, 85.21.4783
1528 5 Table 4 表 4 22 种标准品特征离子及碎片离子图 Characteristic ion and fragment ions gram of 22 kinds of standard substance (component) (characteristic ion gram) (daughter ion scan gram) 1 2 3 4 5 6 7 8 9 1 11 2.4 33.3.1Da 1.5e7. 2.5 5. 7.5 1. 2.62 389.3.1Da 1.14e6. 2.5 5. 7.5 1. 2.87 3.2.1Da 1.18e7. 2.5 5. 7.5 1. 9-1 S 2.77 5.49 2. 2.5 3. 3.5 4. 4.5 5. 5.5 6. 2.99 278.3.1Da 9.91e6. 2.5 5. 7.5 1. 3.14 356.3.1Da 6.1e6. 2.5 5. 7.5 1. 3.16 233.2.1Da 1.84e6. 2.5 5. 7.5 1. 4. 326.2.1Da 8.66e6. 2.5 5. 7.5 1. 5.47 274.1.1Da 1.39e4. 2.5 5. 7.5 1.. 2.5 5. 7.5 1. 6.28 6.62 36.3.1Da 5.4e6 282.2.1Da 5.48e6. 2.5 5. 7.5 1. HBSHUI5 26 (2.4) Cm (1:4) 1: TOF MSMS 33.32ES+ 181.163 1 2.2e5 27.151 153.1556 137.1374 239.18 255.2253 33.3256 5 1 15 2 25 3 35 HBSHUI5 18 (2.66) Cm (8:24) 2: TOF MSMS 3 245.146 1 247.1424 99.1531 345.2837 143.1628 248.1452 5 1 15 2 25 3 35 4 HBSHUI5 9 (2.863) Cm (8:1) 4: TOF MSMS 3.23ES+ 1 227.1657 283.2227 3.79e5 241.1743 285.2229 3.2337 5 1 15 2 25 3 35 HBSHUI5 12 (2.751) Cm (1:14) 3: TOF MSMS 275.26ES+ 1 23.19 2.8e5 232.1871 275.2651 5 75 1 125 15 175 2 225 25 275 3 HBSHUI5 9 (2.991) Cm (7:11) 5: TOF MSMS 278.34ES+ 1 121.1362 1.5e5 58.178 147.1622 215.2535 26.3279 173.1883 278.347 5 75 1 125 15 175 2 225 25 275 3 132 (2 869) C (13 135) 1TOFMSES HBSHUI5 8 (3.128) Cm (5:11) 192.24 1 6: TOF MSMS 356.34ES+ 1.56e6 165.1797 193.281 356.3482 5 1 15 2 25 3 35 4 HBSHUI5 7 (3.163) Cm (5:9) 174.1848 1 7: TOF MSMS 233.25ES+ 2.83e5 175.1896 233.2468 216.2136 1 12 14 16 18 2 22 24 26 28 3 HBSHUI5 17 (4.946) Cm (1:28) 1 29.172 129.1327 189.1228 292.2597 249.251 326.238 5 1 15 2 25 3 35 8: TOF MSMS 326.24ES+ 291.2552 1.5e6 HBSHUI5 37 (5.52) Cm (24:44) 9: TOF MSMS 274.16ES+ 154.1284 1 6.15e4 156.1241 29.1672 13.47 152.1227 178.1146 274.1613 5 1 15 2 25 3 35 HBSHUI5 34 (6.291) Cm (22:47) 1: TOF MSMS 36.28ES+ 1 236.2135 5.77e5 264.2538 235.237 36.282 5 1 15 2 25 3 35 HBSHUI5 1 (6.634) Cm (4:16) 1 18.177 237.2161 282.2235 5 1 15 2 25 3 35 11: TOF MSMS 282.23ES+ 236.2135 6.32e5
5 : 22 1529 续表 4 (component) (characteristic ion gram) (daughter ion scan gram) 12 13 14 15 16 17 18 19 2 21 6.77 2.2.1Da 6.14e6. 2.5 5. 7.5 1. 6.89. 2.5 5. 7.5 1. 7.47 39.2.1Da 5.64e6. 2.5 5. 7.5 1. 7.64 321.2.1Da 3.8e6. 2.5 5. 7.5 1. 7.79 343.2.1Da 1.24e7. 2.5 5. 7.5 1. 8.13 285.2.1Da 1.8e7. 2.5 5. 7.5 1. 3.3 183.3.1Da 3.3e6. 2.5 5. 7.5 1. 5.49 231.3.1Da 2.9e6. 2.5 5. 7.5 1.. 2.5 5. 7.5 1. 6.74 6.98 7.6 225.4.1Da 3.8e6 314.3.1Da 4.36e6. 2.5 5. 7.5 1. HBSHUI5 13 (6.771) Cm (8:18) 12: TOF MSMS 2.22ES+ 1 267.1865 5.42e5 269.1865 138.893 25.1826 241.1743 2.2167 1 12 14 16 18 2 22 24 26 28 3 32 34 HBSHUI5 12 (6.98) Cm (6:19) 1 231.1859 14.1147 128.19 269.1797 287.14 5 1 15 2 25 3 35 13: TOF MSMS 287.2ES+ 241.1743 9.7e5 HBSHUI5 24 (7.46) Cm (16:29) 14: TOF MSMS 39.23ES+ 1 25.1826 281.289 2.29e5 274.2563 283.254 241.1711 129.1187 165.1112 39.2369 1 12 14 16 18 2 22 24 26 28 3 32 34 HBSHUI5 14 (7.644) Cm (9:17) 1 275.1496 15: TOF MSMS 321.17ES+ 5.22e5 229.1683 33.1522 265.1573 323.1696 2 22 24 26 28 3 32 34 HBSHUI5 14 (7.78) Cm (11:17) 16: TOF MSMS 343.21ES+ 1 38.2288 8.52e5 315.1819 24.1859 239.161 253.1899 279.258 343.2117 2 22 24 26 28 3 32 34 HBSHUI5 2 (8.123) Cm (12:29) 17: TOF MSMS 285.2ES+ 1 193.191 2.17e6 154.1284 222.2267 91.1126 241.1743 285.216 5 1 15 2 25 3 35-2HB32 118 (3.28) Cm (83:161) 1: TOF MSMS 183.3ES- 1 14.363 8.59e4 183.3269 96.2936 141.316 184.33 5 75 1 125 15 175 2 225 25 275 3-2HB32 173 (5.488) Cm (16:184) 1 144.3181 188.3219 232.3385 85.297 145.3194 189.3244 5 75 1 125 15 175 2 225 25 275 3-2HB32 7 (6.973) Cm (4:98) 1 138.3637 182.3681 2: TOF MSMS 231.3ES- 231.3338 3.88e4 136.3462 139.3684 183.3741 226.3765 85.297 5 75 1 125 15 175 2 225 25 275 3 3: TOF MSMS 225.36ES- 2.1e4 225.3816-2HB32 75 (7.41) Cm (7:81) 4: TOF MSMS 314.27ES- 1 278.3177 4.68e6 248.3169 286.2749 314.2942 2 22 24 26 28 3 32 34 36 38 4
153 5 22 7.74 7.49 237.4.1Da 3.42e6. 2.5 5. 7.5 1. -2HB32 6 (7.738) Cm (55:63) 5: TOF MSMS 237.36ES- 1 15.372 194.3684 237.3824 7.45e3 238.3844 85.297 5 1 15 2 25 3 35 4 3, 22, 4, 3.4 检出限,, [8], 22,,, S/N = 3,, 3 3, 22, 1 ng, 3.5 方法的优化及专属性 2.4 2924 2124 2132,, 3,,,,,, 3 22, 2924,,,, 5, 3, 22,, 5,,, S/N 3,, 22,, Table 5 表 5 样品及样品加标的离子图 (ESI-) Ion-current grams of sample and sample with recoveries (ESI-) (component) 22:39:18-Jul-213-1BP2131231-2 5.74 TIC.66 7.367.76 4.51e6 3.4 1.38 4.43 7.2 2. 4. 6. 8. 2:36:4418-Jul-213-1BP213123-2 6.817.31 183.3.1Da 6.15 3.1 3.36 4.3e3 2.21 5.18 7.36 8.72 2. 4. 6. 8. -1BP213123-2 7.6 314.3.1Da.82 7.2 7.16 1.76e3 2.36 3.88 4.98 8.89 2. 4. 6. 8. -1BP213123-2 5.78 TIC.66 7.32 4.88e6 4.37 1.22 6.64 2. 4. 6. 8. 22:39:18-Jul-213-1BP2131231-2 3.4-1BP2131231-2 2. 4. 6. 8. 7.9 2. 4. 6. 8. 183.3.1Da 9.96e5 314.3.1Da 1.7e5-1BP213123-2 1.3 231.3.1Da 1.6 2.17 3.11 4.94 5. 2.54e3 7.7 7.46 8.77 2. 4. 6. 8. -1BP2131231-2 5.5 2. 4. 6. 8. 231.3.1Da 6.e4
5 : 22 1531-1BP213123-2 2.55 6.24 225.4.1Da.692.46 2.61 5.98 7.27.34 3.64e3 8.3 2. 4. 6. 8. -1BP2131231-2 7.1 6.76 2. 4. 6. 8. 225.4.1Da 5.67e5-1BP213123-2 6.96 237.4.1Da 5.85 6.89 7.3 1.5e4 7.7 2. 4. 6. 8. -1BP2131231-2 2. 4. 6. 8. 7.76 237.4.1Da 8.27e5 2 Fig. 2 Ion-current grams of sample 3.6 样品的测定结果, [8 9],,,,,, 2 2,,, 4 讨论 - (UPLC-MS-MS-MS), 1.7 μm, UPLC-Q-TOF/MS : UPLC, 1 min 22, ;,, TOF/MS,,, 22 1 ng, 2924 2124 2132, 15, 4 3,,,, 5 ml, 1,.22 μm,,,, 22,,,, 1 ng,,,,, 1,,,,, 22,,,
1532 5,,,,, 参考文献 [1] [EB/OL].http:// www.sda.gov.cn/ws1/cl1523/8196.html. 213-5-2. The special action of "four not" against health food was comprehensively deployed by Administration[EB/OL]. http://www.sda. gov.cn/ws1/cl1523 /8196.html. 213-5-2. [2] 2924. [Z]. 29-12-4. The approved compensation process and inspecting item of China food and Drug Administration. Serial number 2924. Sedative Chinese patent medicines [Z]. 29-12-4. [3] 2124. [Z]. 212-8-2. The approved compensation process and inspecting item of China food and Drug Administration. Serial number 2124.Sedative Chinese patent medicines and Health food [Z]. 212-8-2. [4] 2132. [Z]. 213-3-26. The approved compensation process and inspecting item of China food and Drug Administration. Serial number 2132.Improving sleep Chinese patent medicines and Health food [Z]. 213-3-26. [5] Li SL, Lai SF, Song JZ, et al. Decocting-induced chemical transformations and global quality of Du-Shen-Tang, the decoction of ginseng evaluated by UPLC-Q-TOF-MS/MS based chemical profiling approach [J]. J Pharm Biomed Anal, 21, 53: 946 7. [6] Li SL, Song JZ, Qiao CF, et al. UPLC-PDA-TOF MS based chemical profiling approach to rapidly evaluate chemical consistency between traditional and dispensing granule decoctions of traditional medicine combinatorial formulae [J]. J Pharm. Biomed Anal, 21, 52: 468 478. [7] Li L, Luo GA, Liang QL, et al. Rapid qualitative and quantitative analyses of Asian ginseng in adulterated American ginseng preparations by UPLC/Q-TOF-MS [J]. J Pharm. Biomed Anal, 21, 52: 66 72. [8],,,. LC-MS/MS 18 [J]., 212, 3 (1): 172 175. Wang ZY, Li D, Wen HM, et al. Screening for 18 mixed illegally cough-relieving chemical composition in health food by LC-MS/MS [J]. Chin Arch Trad Chin Med, 212, 3 (1): 172 175. [9],,,. UPLC-MS/MS 7 [J]., 212, 1 (1): 752 756. Li YB, Meng QY, Zhong QY, et al. Rapid identification of 7 drugs illegally added into weight reducing health foods by UPLC-MS/MS [J]. Cent South Pharm. 212, 1 (1): 752 756. 作者简介 ( 责任编辑 : 张宏梁 ) 潘小红, 高级营养师, 助理研究员, 主要研究方向为食品 药品 保健食品及化妆品的检验及研发 E-mail: panxiaohong56781@163.com