Microsoft Word 李洪燕-二校

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1 6 4 Vol. 6 No Journal of Food Safety and Quality Apr., 2015 李洪燕 1, 李秀英 1, 陈俊禧 2, 刘冬虹 1, 林森煜 1, 卢宇靖 2, 黄金凤 (1., ; 2., ) 1* 摘要 : 目的 - - (ID-UHPLC- MS/MS) 方法, HSS T3 0.1%(v:v) -,, 结果 5~1000 µg/l, (R 2 ) ; 0.02 mg/100 g, 0.07 mg/100 g, %~105%, (n=6) 2.3%~4.4%, (n=3) 2.72% 结论, 关键词 : ; - ; ; Determination of pantothenic acid in multivitamin supplement solution by ultra high performance liquid chromatography-tandem mass spectrometry with isotope dilution LI Hong-Yan 1, LI Xiu-Ying 1, CHEN Jun-Xi 2, LIU Dong-Hong 1, LIN Sen-Yu 1, LU Yu-Jing 2, HUANG Jin-Feng 1* (1. Guangzhou Quality Supervision and Testing Institute, Guangzhou , China; 2. Guangdong University of Technology, Guangzhou, , China) ABSTRACT: Objective To develop a simple and rapid method for determination of pantothenic acid in multivitamin supplement by ultra high performance liquid chromatography - tandem mass spectrometry with isotope dilution (ID-UHPLC-MS/MS). Methods The sample was extracted and purified by perchloric acid solution, and separated on an HSS T3 UPLC column with acetonitrile and 0.1% formic acid solution as mobile phase under gradient elution mode, then detected with tandem mass spectrometry under ESI at positive and multiple reaction monitoring (MRM) mode. Stable isotope was used as an internal standard for quantification. Results Pantothenic acid has a good linear relationship in the range of 5~1000 μg/l, the limit of detection (LOD) and quantitation (LOQ) of the method was 0.02 mg/100 g and 0.07 mg/100 g respectively. The fine recovery (93.4%~105%), the relative standard deviation (2.3%~4.4%, n=6) and inter-day with RSD (2.72%, n=3) of this method were also obtained. Conclusion The proposed method was successfully applied to determine pantothenic acid in multivitamin supplement. Compared with the standard method, the proposed method was sensitive, rapid, and accurate. It provides a rapid approach for the analysis of pantothenic acid in complex matrix multivitamin supplement solution. * 通讯作者 :,,, @qq.com *Corresponding author: HUANG Jin-Feng, Senior Engineer, Guangzhou Quality Supervision and Testing Institute, Guangzhou , China @qq.com

2 4, : KEY WORDS: pantothenic acid; ultra high performance liquid chromatography-tandem mass spectrometry; isotope dilution; multivitamin supplement solution 1 引言 B 5,,, A(Co A) (ACP), ;, ;,, [1-3],,,,,, [4-5] [6-10] - [11] [12] [13] [14-16],, 1 mg/l [6-10],,,,,, [14-16],,,,,,,,,, 2 材料与方法 2.1 主要仪器与试剂 - (Agilent 1290/6490, Agilent ); Acquity HSS T3 ( Waters ); (3K 15, Sigma ); (BSA224S, Sartorius ); (MS3, IKA ) ( 98.0%, Dr Ehrenstorfer GmbH ); -[ 13 C 6, 15 N 2 ]( 99.5%, Iso Sciences ); (, ); (, Merck ); (70%~72%, ); 2.2 标准溶液的配制 (200 μg/ml): 54.5 mg, 250 ml( 0.92), 4 (200 μg/ml): -[ 13 C 6, 15 N 2 ] 10 mg, 50 ml, 4 (0.5 mol/l): 500 ml 300 ml, 50 g,, 1 L, 2.3 样品前处理 1.0 g( g) 10 ml, 5 ml,, 2 min, r/min 2 min, 0.5 ml 50 ml, 25 μl,, 1 min, 0.22 μm, UHPLC-MS/MS 2.4 UHPLC-MS/MS 检测条件 : Waters HSS T3 (2.1 mm 100 mm, 1.8 μm); 0.45 ml/min; 30 ; 2 μl; A: 0.1%(v:v) ; B: 1

1506 6 表 1 流动相梯度洗脱条件 Table 1 Gradient program for PA /min A/% B/% 0.0 92 8 2.2 80 20 2.4 50 50 4.0 50 50 4.1 92 8 7.

5 kv; 150 ; 14 L/min, 250 ; ( )11 L/min; 3000 V 2 3 结果与讨论 3.1 样品处理的优化 1,,, Fig.

8) ( 1 ):, 1250, 68%; 50%(v:v) 50%(v:v),, 3.2 色谱与质谱条件的优化 α, γ- -β, β- -β-,, C 18, HSS T3, HSS T3 [M+H] +,, 0.

3 表 1 流动相梯度洗脱条件 Table 1 Gradient program for PA /min A/% B/% , 86% 75%,,,, ; 0.5 mol/l (ph 3.8),, 10 min,,, 92% 89%, 0.5 mol/l, ; (MRM) ; 1.5 kv; 150 ; 14 L/min, 250 ; ( )11 L/min; 3000 V 2 3 结果与讨论 3.1 样品处理的优化 1,,, Fig. 1 Comparison of various solvent on recoveries,,,,,,,, 50%(v:v) 50%(v:v) 0.5 mol/l (ph 3.8) ( 1 ):, 1250, 68%; 50%(v:v) 50%(v:v),, 3.2 色谱与质谱条件的优化 α, γ- -β, β- -β-,, C 18, HSS T3, HSS T3 [M+H] +,, 0.1%(v:v), -, 2 表 2 泛酸及其同位素内标的定性和定量离子对碰撞池加速电压和碰撞能量 Table 2 Qualitative and quantitative ion pairs, cell accelerator voltages and collision energy for PA and PA-Internal Standard (m/z) (m/z) (V) (V) * [ 13 C 6, 15 N 2 ] * 10 4 : *

4 4, : Fig. 2 2 MRM MRM chromatograms of PA and PA internal standard 3.3 方法的验证线性范围检出限和定量限, ( 100 µg/l ) UHPLC-MS/MS, (A) A i, A A i Y (X, µg/l), 5~1000 µg/l, Y 1.22X , (R 2 ) ( S/N=3) ( S/N=10),,, 0.02 mg/100 g 0.07 mg/100 g 回收率, 40% 80% 130%, 6,, % 105%, 2.3% 4.4% 方法稳定性 1.3, 3, 6,, 4, 2.07%~2.52%, 2.72%, MRM 与国标方法的对比 GB/T , ( 5),,,,,,,,,, UHPLC-MS/MS,

5 Table 3 表 3 泛酸加标回收试验结果 Recoveries and relative standard deviations of PA (mg/100 g) (mg/100 g) (%) (%) (%) Table 4 表 4 样品中泛酸的稳定性试验结果 (n=6) Results of stability experiment of PA in samples (n=6) (mg/100 g) (mg/100 g) (%) (%) day day day MRM Fig. 3 MRM chromatograms of sample Table 5 表 5 本方法与国标法检测结果的比较 Comparison of the proposed method and the standard method (mg/100 g) (mg/100 g) (mg/100 g)

6 4, : 结论 UHPLC-MS/MS,,,, 参考文献 [1],. [J]., 2008, 28(4): Yang YH, Xiao CL. The functions and biosynthesis of pantothenate [J]. Chem Life, 2008, 28(4): [2],,,. [J]., 2014, 29(2): Na HQ, Gao GH, Zhao F, et al. Determination of pantothenic acid in vitamins with minerals tablets by HPLC [J]. Northwest Pharm J, 2014, 29(2): [3],,,. B 6 [J]., 2007, 17(11): , Li YZ, Chen YZ, Li SD, et al. Simultaneous determination of vitamin B 6, nicotinamide calcium and pantothenate by high performance liquid chromatography [J]. Chin J Health Lab Technol, 2007, 17(11): , [4],,,. [J]., 2014, 35(9): Zhang Y, Huo SN, Hu MY, et al. Uncertainty evaluation for pantothenic acid determination in infant formula [J]. Food Res Devel, 2014, 35(9): [5] Tsukatani T, Suenaga H, Ishiyama M, et al. Determination of water-soluble vitamins using a colorimetric microbial viability assay based on the reduction of water-soluble tetrazolium salts [J]. Food Chem, 2011, 127(2): [6] Shibata K, Nakai T, Fukuwatari T. Simultaneous high-performance liquid chromatography determination of coenzyme A, dephospho-coenzyme A, and acetyl-coenzyme A in normal and pantothenic acid-deficient rats [J]. Anal Biochem, 2012, 430(2): [7] Ciulu M, Solinas S, Floris I, et al. RP-HPLC determination of water-soluble vitamins in honey [J]. Talanta, 2011, 83(3): [8] Takahashi K, Fukuwatari T, Shibata K. Fluorometric determination of pantothenic acid in human urine by isocratic reversed-phase ion-pair high-performance liquid chromatography with post-column derivatization [J]. J Chromatogr B, 2009, 877(22): [9] Jin PF, Xia LF, Li Z, et al. Rapid determination of thiamine, riboflavin, niacinamide, pantothenic acid, pyridoxine, folic acid and ascorbic acid in Vitamins with Minerals Tablets by high-performance liquid chromatography with diode array detector [J]. J Pharm Biomed Anal, 2012, 70: [10] Woollard DC, Indyk HE, Christiansen SK. The analysis of pantothenic acid in milk and infant formulas by HPLC [J]. Food Chem, 2000, 69(2): [11] Banno K, Matsuoka M, Horimoto S, et al. Simultaneous determination of pantothenic acid and hopantenic acid in biological samples and natural products by gas chromatography-mass fragmentography [J]. J Chromatogr B: Biomed Sci Appl. 1990, 525: [12],,. β- γ- [J]., 2001, 20(1): Zhang Q, Qiu J, Liao NS. Simultaneous determination of calcium pantothenate and calcium-amino propionate and γ-butyrolactone by high performance capillary zone electrophories [J]. Chin J Anal Lab, 2001, 20(1): [13],,. D- [J]., 2010, 29(4): Cheng J, Xie JC, Su XO. Determination of D-calcium pantothenate in animal feeds by supercritical fluid chromatography [J]. J Instrum Anal, 2010, 29(4): [14] Mittermayr R, Kalman A, Trisconi MJ, et al. Determination of vitamin B 5 in a range of fortified food products by reversed-phase liquid chromatography mass spectrometry with electrospray ionization [J]. J Chromatogr A, 2004, 1032(1-2): 1 6. [15] Chen Z, Chen B, Yao S. High-performance liquid chromatography/electrospray ionization-mass spectrometry for simultaneous determination of taurine and 10 water-soluble vitamins in multivitamin tablets [J]. Anal Chimica Acta, 2006, 569(1 2): [16],,,. - [J]., 2014, 35(8): Qu Y, Cui YJ, Li QX et al. Determination of pantothenic acid in formula milk powder using ultra performance liquid chromatography-isotope dilution mass spectrometry [J]. Food Sci, 2014, 35(8): ( 责任编辑 : 李振飞 )

7 作者简介李洪燕, 工程师, 主要研究方向为食品营养分析检测技术 @qq.com 黄金凤, 高级工程师, 主要研究方向为食品及食品相关产品检测, 仪器分析技术 @qq.com

Microsoft Word 李秀英-二校

Microsoft Word 李秀英-二校 7 5 Vol. 7 No. 5 2016 5 Journal of Food Safety and Quality May, 2016 / - 李秀英 1, 黄嘉乐 1, 邱志超 1, 何雪滢 2, 林森煜 1, 黄宝华 2, 郭新东 (1., 511400; 2., 510006) 1* 摘要 : 目的,, - (UPLC-MS/MS) 方法, 1.5 mol/l, 100 10 min ph,