牛血清白蛋白修饰的有机-硅胶杂化整体柱的制备及电色谱研究
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摘要
以四甲氧基硅烷(Tetramethoxysilane,TMOS)和γ-缩水甘油醚氧基丙基三甲氧基硅烷(γ-Glycidyloxypropyltrimethoxysilane,GPTMS)为反应单体,采用"一步法"制备了有机-硅胶杂化整体柱,通过优化TMOS/GPTMS的含量制备了渗透性和机械强度良好的杂化整体柱,扫描电镜结果显示,毛细管柱内形成了连续的整体结构。采用席夫式碱法将牛血清白蛋白(Bovine serum albumin,BSA)共价键合于柱内,并对制备条件进行了优化,确定8 g/L BSA为最佳反应条件。在毛细管电色谱分离模式下,考察了缓冲液pH值、运行电压、柱温和进样条件对分离的影响,最终选择25 mmol/L Tris缓冲液(pH 7.4)、运行电压10 kV、柱温15℃和进样条件2 kV×3 s为最佳分离条件。此手性柱对色氨酸对映体实现了基线分离,日内、日间及柱间精密度良好。与BSA修饰的硅胶整体柱相比,本研究制备的固载BSA的杂化整体柱具有更优的手性识别能力,并且简化了制备流程,缩短了制备周期。
An epoxy functionalized organic-silica hybrid monolith was successfully synthesized via one-step method with tetramethoxysilane( TMOS) and γ-glycidyloxypropyltrimethoxysilane( GPTMS) as precursors.The volume of reactive monomer was optimized and the capillary monolithic columns with good permeability and mechanical strength were obtained. Scanning electron microscope characterization demonstrated that a structure of continuous bed was formed. Then bovine serum albumin( BSA) was covalently immobilized on capillary monolith inner surface by Schiff base method. The preparation condition was studied and it was found that the appropriate concentration of BSA for preparation of BSA-modified hybrid monoliths was 8.0 g/L.Meanwhile,the influence factors on the sample separation were investigated,including running buffer pH(25 mmol/L Tris,pH 7.4),applied voltage(10 kV),capillary temperature(15℃) and sample condition(2 kV × 3s). The BSA-modified hybrid monolith exhibited good chiral separation ability to( ±)-tryptophan and achieved satisfactory run-to-run,day-to-day and column-to-column repeatability. Compared with BSAimmobilized silica-based monolithic column,this work presented better chiral recognition ability and also simplified preparation procedures and shortened preparation period.
An epoxy functionalized organic-silica hybrid monolith was successfully synthesized via one-step method with tetramethoxysilane( TMOS) and γ-glycidyloxypropyltrimethoxysilane( GPTMS) as precursors.The volume of reactive monomer was optimized and the capillary monolithic columns with good permeability and mechanical strength were obtained. Scanning electron microscope characterization demonstrated that a structure of continuous bed was formed. Then bovine serum albumin( BSA) was covalently immobilized on capillary monolith inner surface by Schiff base method. The preparation condition was studied and it was found that the appropriate concentration of BSA for preparation of BSA-modified hybrid monoliths was 8.0 g/L.Meanwhile,the influence factors on the sample separation were investigated,including running buffer pH(25 mmol/L Tris,pH 7.4),applied voltage(10 kV),capillary temperature(15℃) and sample condition(2 kV × 3s). The BSA-modified hybrid monolith exhibited good chiral separation ability to( ±)-tryptophan and achieved satisfactory run-to-run,day-to-day and column-to-column repeatability. Compared with BSAimmobilized silica-based monolithic column,this work presented better chiral recognition ability and also simplified preparation procedures and shortened preparation period.
引文
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27 XIONG Le-Le,LI Rui-Jun,JI Yi-Bing.Chinese Journal of Chromatography,2017,34(7):712-719熊乐乐,李瑞军,季一兵.色谱.2017,34(7):712-719
28 Hong T T,Zheng Y,Hu W W,Ji Y B.Anal.Biochem.,2014,464:43-50
2 Thurmann S,Lotter C,Heiland J J,Chankvetadze B,Belder D.Anal.Chem.,2015,87(11):5568-5576
3 LI Yang,ZHU Jin-Ke,FENG Hua,LI Lian-Jie,JIANG Deng-Gao.Chinese J.Anal.Chem.,2015,43(8):1162-1168李杨,朱进科,封华,李连杰,蒋登高.分析化学,2015,43(8):1162-1168
4 ZHOU Ren-Dan,LI Lai-Sheng,CHENG Biao-Ping,NIE Gui-Zhen,ZHANG Hong-Fu.Chinese J.Anal.Chem.,2014,42(7):1002-1009周仁丹,李来生,程彪平,聂桂珍,张宏福.分析化学,2014,42(7):1002-1009
5 Aydogan C,Ylmaz F,Cimen D,Uzun L,Denizli A.Electrophoresis,2013,34(13):1908-1914
6 ZHANG Lei,ZHANG Ai-Qin,WANG Man,SHEN Gang-Yi.Chinese J.Anal.Chem.,2017,45(11):1727-1733张蕾,张爱芹,王嫚,申刚义.分析化学,2017,45(11):1727-1733
7 Silva M,Pérez-Quintanilla D,Morante-Zarcero S,Sierra I,Marina M L,Aturki Z,Fanali S.J.Chromatogr.A,2017,1490:166-176
8 Fang G Z,Qian H L,Deng Q L,Ran X Q,Yang Y K,Liu C C,Wang S.RSC Adv.,2014,4(30):15518-15525
9 Zeng J,Liu S Q,Wang M L,Yao S Z,Chen Y Z.Electrophoresis,2017,38(9-10):1325-1333
10 Qiao X Q,Zhang N,Han M M,Li X Y,Qin X Y,Shen S G.J.Sep.Sci.,2017,40(5):1024-1031
11 Yu X H,Ling X,Zou L,Chen Z L.J.Sep.Sci.,2017,40(7):1556-1563
12 Cai J,Zhu G T,He X M,Zhang Z,Wang R Q,Feng Y Q.Talanta,2017,170:252-259
13 Ghanem A,Ahmed M,Ishii H,Ikegami T.Talanta,2015,132:301-314
14 Hsieh M L,Chau L K,Hon Y S.J.Chromatogr.A,2014,1358:208-216
15 Ou J J,Liu Z S,Wang H W,Lin H,Dong J,Zou H F.Electrophoresis,2015,36(1):62-75
16 SHAN Yuan-Hong,SHI Xian-Zhe,DOU A-Bo,QIAO Li-Zhen,LI Hua,XU Guo-Wang.Journal of Instrmental Analysis,2012,31(9):1075-1080单圆鸿,石先哲,窦阿波,乔利珍,李华,许国旺.分析测试学报,2012,31(9):1075-1080
17 Liu S J,Peng J X,Zhang H Y,Li X,Liu Z Y,Kang X H,Wu M H,Wu R A.J.Chromatogr.A,2017,1498:64-71
18 Bai J Y,Liu Z S,Wang H W,You X,Ou J J,Shen Y H,Ye M L.J.Chromatogr.A,2017,1498:37-45
19 Wang Z,Zhao J C,Lian H Z,Chen H Y.Talanta,2015,138:52-58
20 Hayes J D,Malik A.Anal.Chem,2000,72(17):4090-4099
21 Wu M H,Wu R A,Wang F J,Ren L B,Dong J,Liu Z,Zou H F.Anal.Chem.,2009,81(9):3529-3536
22 Xu H R,Xu Z D,Yang L M,Wang Q Q.J.Sep.Sci.,2011,34(16-17):2314-2322
23 Zhao L C,Yang L M,Wang Q Q.J.Chromatogr.A,2016,1446:125-133
24 Hong T T,Yang X,Xu Y J,Ji Y B.Anal.Chim.Acta,2016,931:1-24
25 Li Z,Rodriguez E,Azaria S,Pekarek A,Hage D S.Electrophoresis,2017,38(22-23):2837-2850
26 Lu J Y,Ye F G,Zhang A Z,Wei Z,Peng Y,Zhao S L.J.Sep.Sci.,2011,34(16-17):2329-2336
27 XIONG Le-Le,LI Rui-Jun,JI Yi-Bing.Chinese Journal of Chromatography,2017,34(7):712-719熊乐乐,李瑞军,季一兵.色谱.2017,34(7):712-719
28 Hong T T,Zheng Y,Hu W W,Ji Y B.Anal.Biochem.,2014,464:43-50