毛细管电泳法测定化妆品中的烟酸和烟酰胺

2015-06-23 13:56王丽娟刘惠涛
关键词:烟酰胺烟酸超纯水

亓 慧,王丽娟,栾 锋,刘惠涛

(烟台大学化学化工学院,山东烟台264005)

应用技术

毛细管电泳法测定化妆品中的烟酸和烟酰胺

亓 慧,王丽娟,栾 锋,刘惠涛

(烟台大学化学化工学院,山东烟台264005)

建立了一种测定化妆品中烟酸和烟酰胺的毛细管电泳法,成功应用于面霜、乳液和化妆水的检测.样品中的待测物直接用水提取进行检测,以60 cm(有效长度为50.5 cm) ×75 μm的熔融石英毛细管为分离通道,紫外检测波长210 nm,30 mbar压力下进样5 s.电泳分离条件为:10 mmol/L磷酸氢二钠溶液,pH值9.2,运行电压25 kV,温度25℃.2种物质在7 min内实现完全分离.烟酸和烟酰胺分别在1~200 μg/mL和5~500 μg/mL范围内峰面积与浓度呈良好的线性关系,线性相关系数R>0.999.烟酸和烟酰胺的检测限分别为0.35 μg/mL、0.67 μg/mL,加标回收率在97.9%~107.7%,相对标准偏差RSD≤4.5%.该方法具有分离速度快、操作简便、分析时间短、成本低、环境友好等优点.

化妆品;毛细管电泳;烟酸;烟酰胺

烟酸(NA,尼克酸)和烟酰胺(NAM,烟碱酰胺或尼克酰胺)统称为维生素B3且均为水溶性维生素.烟酰胺是一种有效且耐受性良好的抗氧化剂,它有助于改善皮肤表面油脂的分泌、减少皮表水分的流失,还可以抑制黑色素的产生、减少色素沉着[1].研究表明,烟酰胺具有去黄、除皱、祛斑等功效且可以有效改善皮肤状况[2-4].烟酸也因具有美白、除皱、抗炎等功能应用于化妆品中.烟酸和烟酰胺在化妆品中的推荐含量一般为0.5%~2.0%.

烟酸和烟酰胺的检测方法有HPLC[5-6]、CE[7-9]、流动注射法[10]、LC-MS[11-12]、容量分析[13]、微生物分析[14]、离子色谱法[15-16]、荧光分析[17]和König反应[18],主要用于食品、血浆和尿液的分析检测.化妆品中烟酸和烟酰胺的分析方法有HPLC[19-22]和胶束毛细管电泳(MEKC)[23].GB/T 29664—2013[24]中用高效液相色谱法检测化妆品中的烟酸和烟酰胺,但其样品处理方法复杂,萃取后需过固相萃取柱,且其色谱柱为SB-Ap柱.本研究所建立的毛细管电泳法(Capillary Electrophoresis,CE)操作简单,准确度高,是检测化妆品中烟酸和烟酰胺的有效方法.

1 实验

1.1 仪器设备

CAPEL 105型毛细管电泳仪(俄罗斯LUMEX公司);色谱工作站为Chrom&Spec software for chromatography,vertion 1.5x;石英毛细管柱60 cm×75 μm i.d.,有效长度50.5 cm(俄罗斯LUMEX公司);STARTER 3100 pH计(上海奥豪斯仪器有限公司);KQ5200E型超声波清洗器(昆山市超声仪器有限公司).

1.2 试剂与材料

烟酸和烟酰胺标准品(购自于AccuStandard Inc (New Haven,USA));乙腈、甲醇、丙酮均为市售色谱纯;硼砂、氢氧化钠、磷酸、磷酸二氢钠、磷酸氢二钠均为分析纯.不同品牌的化妆品样品(乳液A,乳液B,面霜和化妆水)购自当地超市;实验用水均为超纯水.

1.3 电泳条件

采用压力进样方式,在30 mbar下进样5 s,分离电压25 kV,检测波长210 nm.背景电解质溶液以10 mmol/L的磷酸氢二钠制备.新毛细管使用前用超纯水冲洗10 min,0.5 mol/L氢氧化钠溶液冲洗40 min,超纯水冲洗10 min.每天毛细管首次使用前用超纯水冲洗5 min,0.2 mol/L氢氧化钠溶液冲洗20 min,超纯水冲洗5 min,运行缓冲液冲洗10 min,每两次运行之间用超纯水冲洗1 min,运行缓冲液冲洗10 min.所用溶液均经0.45 μm滤膜过滤.

1.4 标准溶液和样品制备

标准制备:先将烟酸和烟酰胺标准品用超纯水分别配制成1 000 μg/mL的储备液.混合标准使用液由储备液混合并稀释相应的倍数制得.所有标准溶液于避光4℃下可稳定保存一个月.

样品制备:分别称取3.3 g乳液A、0.3 g乳液B和0.3 g面霜于3个100 mL的容量瓶中,用超纯水定容.超声30 min,5 000 r/min离心20 min.取出上清液,经0.45 μm有机膜过滤后分析测定.称取0.3 g化妆水于500 mL容量瓶中,用超纯水定容,0.45 μm有机膜过滤分析测定.

2 结果与讨论

2.1 CE条件的优化

2.1.1 最佳吸收波长的选择在190~400 nm范围内分别对烟酸和烟酰胺的水溶液进行紫外光谱扫描,所得谱图如图1所示.根据扫描结果选择210 nm为检测波长.

图1 烟酸和烟酰胺的紫外吸收光谱图Fig.1 The UV spectra of NA and NAM in water

2.1.2 缓冲溶液种类和pH值的选择采用压力进样的方式,在30 mbar下进样5 s,分离电压为25 kV对缓冲溶液的种类和pH值进行优化.分别选取20 mmol/L的硼砂溶液(pH值9.0~11.0间隔1.0)、磷酸氢二钠溶液(pH值5.0~9.0间隔1.0; 9.0~11.0间隔0.5)和磷酸二氢钠溶液(pH值2.0~5.0间隔1.0)作为背景电解质溶液,结果显示选用磷酸氢二钠缓冲体系可得到较短的分析时间和较好的峰形.经过进一步的实验发现当磷酸氢二钠溶液不调pH值时可得到较好的实验结果,因而选择背景电解质溶液为磷酸氢二钠溶液,不用调节其pH值.

2.1.3 缓冲溶液浓度的选择采用压力进样的方式,在30 mbar下进样5 s,分离电压为25 kV,选取浓度为10、15、20、25、30 mmol/L的磷酸氢二钠为背景电解质溶液,结果显示选用10 mmol/L磷酸氢二钠缓冲体系可得到较短的分析时间和较好的峰形,因而选择磷酸氢二钠缓冲溶液浓度为10 mmol/L.

2.1.4 有机改性剂的选择在缓冲溶液中适当加入有机试剂可以改变背景电解质的组成和极性,降低焦耳热、改善分离效果.本实验对3种有机改性剂进行了考察.固定缓冲溶液磷酸氢二钠的浓度为10 mmol/L,分别加入10%的甲醇、乙醇和乙腈,观察有机试剂对分离效果的影响.结果显示加入乙腈后会对峰形有明显的改善,因而选择加入乙腈有机改性剂.

2.1.5 有机改性剂加入量的选择固定缓冲溶液磷酸氢二钠的浓度为10 mmol/L,分别加入体积分数为5%、8%、10%、15%、20%、25%的乙腈溶液,结果显示当加入8%的乙腈时可明显的改善峰形且有较短的分析时间,因而选择乙腈的加入量为8%.

综上所述,CE分离检测烟酸和烟酰胺的最佳实验条件为:在210 nm检测波长下,缓冲溶液为10 mmol/L的磷酸氢二钠溶液,加入8%的乙腈.采用压力进样方式,在30 mbar下进样5 s,分离电压为25 kV.在最佳条件下烟酸和烟酰胺的混合标准溶液的CE谱图见图2.

2.2 方法验证

2.2.1 线性关系分别配制一系列不同浓度的烟酸和烟酰胺标准溶液(烟酸的浓度分别为1、10、50、100、200 μg/mL,烟酰胺的浓度分别为5、50、100、200、500 μg/mL),在上述电泳条件下进样,结果表明在上述浓度范围内烟酸和烟酰胺的峰面积与浓度具有良好的线性关系,其回归方程和相关系数(R)如表1所示.

图2 最佳条件下标准溶液的电泳谱图Fig.2 Electropherograms of standard mixture

表1 烟酸和烟酰胺的线性回归方程Tab.1 Calibration curves of NA and NAM

2.2.2 精密度连续5 d对化妆品加标样品进行测定,每天测定5次.分别计算日间精密度和日内精密度,结果如表2所示.由表2数据可知,该方法的精密度良好.

2.2.3 检测限和定量限检测限(LOD)和定量限(LQD)分别按公式3σ/s和10σ/s计算,其中σ是空白信号的标准偏差,s是校准曲线的斜率.实验结果如表3所示.

表2 烟酸和烟酰胺精密度实验结果Tab.2 Precision for the analysis of NA and NAM%

表3 烟酸和烟酰胺的检测限和定量限Tab.3 LOD、LOQ data for the analysis of NA and NAM

2.3 样品测定和回收率

将所建立的CE法应用于乳液(A和B)、面霜(C)和化妆水(D)样品分析.样品和样品加标的测定谱图(以面霜为例)如图3所示.

样品的测定结果和添加回收率的实验结果见表4,烟酸的加标回收率为97.9%~107.7%,烟酰胺的加标回收率为99.1%~100.9%

Fig.3 样品和样品加标电泳图Fig.3 The electrophoregrams of the sample and spiked samples

表4 化妆品中烟酸和烟酰胺的添加回收率Tab.4 Recoveries for the determination of NA and NAM in sample

续表

3 结论

本研究建立了一种简单快速分离检测化妆品中烟酸和烟酰胺的毛细管电泳法.样品的前处理方法简单,回收率高.该方法具有分离效率高,灵敏度好,操作简单且环境污染小等优点,是检测化妆品中烟酸和烟酰胺的环境友好型方法.

[1]Ramos-e-Silva M,Celem L R,Ramos-e-Silva S,et al. Anti-aging cosmetics:Facts andcontroversies[J].Clinics in Dermatology,2013,31(6):750-758.

[2]Bissett D,Miyamoto K,Sun P,et al.Topical niacinamide reduces yellowing,wrinkling,red blotchiness,and hyperpigmented spots in aging facial skin[J].International Journal of Cosmetic Science,2004,26(5):231-238.

[3]Otte N,Borelli C,Korting H.Nicotinamide—biologic actions of an emerging cosmetic ingredient[J].International Journal of Cosmetic Science,2005,27(5):255-261.

[4]Bissett D L,Oblong J E,Berge C A.Niacinamide:AB vitamin that improves aging facial skin appearance[J].Dermatologic Surgery,2005,31(s1):860-866.

[5]毛希琴.高效液相色谱法测定化妆品中的烟酸和烟酰胺[J].日用化学工业,2012,42(6):469-472.

[6]Zafra-Gómez A,Garballo A,Morales JC,et al.Simultaneous determination of eight water-soluble vitamins in supplemented foods by liquid chromatography[J].Journal of Agricultural and Food Chemistry,2006,54(13):4531-4536.

[7]Iwaki M,Murakami E,Kikuchi M,et al.Simultaneous determination of nicotinic acidand its metabolites in rat urine by micellar electrokinetic chromatography with photodiode-array detection[J].Journal of Chromatography B:Biomedical Sciences and Applications,1998,716(1):335-342.

[8]Lu Yuanqi,Wu Chunhua,Yuan Zhuobin.Determination of hesperetin,cinnamic acid and nicotinic acid inpropolis with micellar electrokinetic capillary chromatography[J].Fitoterapia,2004,75(3):267-276.

[9]Iwaki M,Murakami E,Kakehi K.Chromatographic and capillary electrophoretic methods for the analysis of nicotinic acid and its metabolites[J].Journal of Chromatography B:Biomedical Sciences and Applications,2000,747(1):229-240.

[10]Hassan R O,Faizullah A T.Determination of nicotinamide by stopped-flow injection method in pharmaceutical formulations[J].Arabian Journal of Chemistry,2013,6(4):393-400.

[11]Catz P,Shinn W,Kapetanovic IM,et al.Simultaneous determination of myristyl nicotinate,nicotinic acid,and nicotinamide in rabbit plasma by liquid chromatography-tandem mass spectrometry using methyl ethyl ketone as a deproteinization solvent[J].Journal of Chromatography B,Analytical Technologies in the Biomedical and Life Sciences,2005,829(1):123-135.

[12]Ma Ming,Luo Xubiao,Chen Bo,et al.Simultaneous determination of water-soluble and fat-soluble synthetic colorants in foodstuff by high-performance liquid chromatography-diode array detection-electrospray mass spectrometry[J].Journal of Chromatography A,2006,1103(1):170-176.

[13]Castanheira I,Batista E,Valente A,et al.Quality assurance of volumetric glassware for the determination of vitamins in food[J].Food Control,2006,17(9):719-726.

[14]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 tetra-zolium salts[J].Food Chemistry,2011,127(2):711-715.

[15]Saccani G,Tanzi E,Mallozzi S,et al.Determination of niacin in fresh and dry cured pork products by ion chromatography:experimental design approach for the optimisationof nicotin-ic acid separation[J].Food Chemistry,2005,92(2):373-379.

[16]Valls F,Sancho M T,Fernández-Muiño M A,et al. Simultaneous determination of nicotinic acid and nicotinamide in cooked sausages[J].Journal of Agricultural and Food Chemistry,2000,48(8):3392-3395.

[17]Krasnova A,Aguilar-Caballos MP,Gómez-Hens A.Selective determination of nicotinic acid and nicotinamide using terbium(III)sensitised luminescence[J].Analytica Chimica Acta,2001,441(2):249-256.

[18]Capella-Peiró M E,Carda-Broch S,Monferrer-Pons L,et al.Micellar liquid chromatographic determination of nicotinic acid and nicotinamide after precolumn König reaction derivatization[J].Analytica Chimica Acta,2004,517(1):81-87.

[19]Muszalska I,Kiaszewicz K,Ksoń D,et al.Determination of nicotinamide(vitamin B3)in cosmetic products using differentialspectrophotometryandliquidchromatography (HPLC)[J].Journal of Analytical Chemistry,2013,68(11): 1007-1013.

[20]Jeon J S,Lee M J,Yoon M H,et al.Determination of arbutin,niacinamide,and adenosine in functional cosmetic products by high-performance liquid chromatography[J].Analytical Letters,2014,47(10):1650-1660.

[21]Yang Yu,Strickland Z,Kapalavavi B,et al.Industrial application of green chromatography-I:Separation and analysis of niacinamide in skincare creams using pure water as the mobile phase[J].Talanta,2011,84(1):169-174.

[22]Lin Chenghui,Wu Hsin lung,Huang Yeoulih.Combining high-performance liquid chromatography with on-line microdialysis sampling for the simultaneous determination of ascorbyl glucoside,kojic acid,and niacinamide in bleaching cosmetics[J].Analytica Chimica Acta,2007,581(1):102-107.

[23]Sun Hongli,Wu Yiwei.Field-amplified sample injection for the determination of iso-nicotinamide and nicotinamide in whitening cosmetics and supplemented foodstuffs by MEKC[J]. Analytical Methods,2013,5(20):5615-5621.

[24]GB/T 29664—2013,化妆品中维生素B3(烟酸、烟酰胺)的测定高效液相色谱法和高效液相色谱串联质谱法[S].

Determination of Nicotinic Acid and Nicotinamide in Cosmetics by Capillary Electrophoresis

QI Hui,WANG Li-juan,LUAN Feng,LIU Hui-tao
(School of Chemistry and Chemical Engineering,Yantai University,Yantai 264005,China)

A simple,rapid and accurate method for the determination of nicotinic acid(NA)and nicotinamide (NAM)in cosmetics by capillary electrophoresis(CE)is developed and successfully applied in creams,lotions,and toning lotions.A fused silica capillary(75 μm×60 cm,with effective length of 50.5 cm)is used,the detection wavelength is 210 nm,the injection pressure is 30 mbr,and the injection time is 5 s.The real samples are extracted with water.After sonication and centrifuging,the extract is analyzed by CE directly.Separation is performed by CE using 10 mmol/L disodium hydrogen phosphate as running buffer,an applied voltage of 25 kV with UV detection at 210 nm.Two targets are separated completely within 7 min.The results show that CE method has a good performance for NA and NAM analysis with linearity(R>0.999),precision(≤4.5%),accuracy (97.9%-107.7%),and enough sensitivity.

cosmetic;capillary electrophoresis;nicotinic acid;nicotinamide

O657

A

(责任编辑 周雪莹)

1004-8820(2015)02-0146-05

10.13951/j.cnki.37-1213/n.2015.02.013

2014-06-19

山东省自然科学基金资助项目(ZR2012BM016).

亓慧(1990-),女,山东莱芜人,硕士研究生.

刘惠涛(liuht-ytu@163.com),教授,博士后,主要研究方向为色谱分析.

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