基于硅胶整体材料研磨颗粒的多功能聚乙二醇键合固定相的制备及其在高效液相色谱多模式分离中的应用

doi:10.3724/SP.J.1123.2020.03036

摘要/Abstract

摘要：

研究通过对溶胶-凝胶法制备的硅胶整体材料进行研磨、浮选、假晶相转换和水热处理，最终获得了粒径为2~5 μm、孔径为20~60 nm的硅胶颗粒。利用部分含氟的阴离子表面活性剂Capstone FS-66和常用的阳离子表面活性剂十六烷基三甲基溴化铵（CTAB）组成的双胶束模板体系对硅胶基质进行假晶相转换处理；再采用碳酸钠溶液水热处理的方式，进一步扩大孔径。用扫描电镜（SEM）和N₂吸附-解吸等温线测量对扩孔处理前后的硅胶整体材料研磨颗粒进行表征，结果清楚地显示了处理前后的形貌变化和差异。随后将含有长链聚乙二醇（PEG）的硅烷键合到扩孔后的硅胶颗粒表面，分别利用元素分析、红外光谱以及热重分析对固定相进行表征，并对固定相进行色谱性能评价。对键合固定相的元素分析和热重分析数据进行分析表明，硅胶表面键合PEG的含量约为8%。研究揭示了利用假晶相转换法与碳酸钠溶液水热处理和长链PEG硅烷修饰的硅胶整体材料颗粒在尺寸排阻色谱分离蛋白质方面的良好分离效果。同时进一步的高效液相色谱评价结果表明，该键合固定相还可用于疏水作用色谱模式分离核糖核酸酶A和溶菌酶，以及可用于亲水作用色谱模式分离吡啶甲酸、左旋多巴、三聚氰胺和邻苯二酚等极性比较强的化合物。研究显示了PEG键合固定相具有多功能性，及其在多模式高效液相色谱分离中的应用潜力。

关键词: 高效液相色谱, 假晶相转换, 蛋白质分离, 多模式固定相, 硅胶整体材料研磨颗粒

Abstract:

Silica monolith particles with sizes of 2-5 μm and pore sizes of 20-60 nm were obtained by grinding, flotation, pseudomorphic transformation, and hydrothermal treatment of the silica monolith prepared by the sol-gel method. The pseudomorphic transformation was performed with a dual micellar templating system consisting of Capstone FS-66, a partially fluorinated anion surfactant, and cetyltrimethylammonium bromide (CTAB), a commonly used cation surfactant. Hydrothermal treatment with a sodium carbonate solution was adopted to further expand the pore size. Scanning electron microscopy (SEM) images and N₂ adsorption-desorption isotherm measurement results of the silica monolith particles before and after the treatments clearly demonstrated the changes in morphology caused by these treatments. Afterward, a long-chain polyethylene glycol (PEG) containing silane was bonded on the surface of the as-prepared particles, and the resulting products were characterized by elemental analysis and FT-IR spectroscopy analysis, and evaluated by high performance liquid chromatography (HPLC). Elemental analysis and thermogravimetric analysis (TGA) of the bonded stationary phase revealed that the bonding amount of PEG on the silica surface is about 8%. It has been shown that silica monolith particles can be treated and modified for the separation of proteins in size exclusion chromatography mode. It is also demonstrated that the bonded stationary phase can be used for the separation of ribonuclease A and lysozyme in hydrophobic interaction chromatography mode, and for the separation of highly polar compounds (picolinic acid, levodopa, melamine, and catechol) in hydrophilic interaction chromatography mode. The results indicate the versatility of the PEG-bonded stationary phase and its promising application to multi-modal separation in HPLC.

Key words: high performance liquid chromatography (HPLC), pseudomorphic transformation, protein separation, multi-modal stationary phase, silica monolith particles

梁倩, 周玉红, 张之伦, 黄明贤. 基于硅胶整体材料研磨颗粒的多功能聚乙二醇键合固定相的制备及其在高效液相色谱多模式分离中的应用[J]. 色谱, 2020, 38(8): 937-944.

LIANG Qian, ZHOU Yuhong, ZHANG Zhilun, HUANG Mingxian. Preparation of a versatile polyethylene glycol-bonded stationary phase based on silica monolith particles and its application in multi-modal separation in high performance liquid chromatography[J]. Chinese Journal of Chromatography, 2020, 38(8): 937-944.

图/表 9

Fig. 1 SEM images of silica monolith particles (a, b) before and (c, d) after treatment

Fig. 2 Schematic of pore enlargement

Fig. 3 (a) Pore size distribution curves and (b) N2 adsorption-desorption isotherms of the silica particles (after Na2CO3 treatment)

Fig. 4 Schematic procedure for the preparation of PEG-bonded stationary phase

Table 1 Elemental analysis of the product in the preparation of PEG-bonded stationary phase

Product	Contents/ %
Product	C	O	Si
Silica	0	63.87	36.13
PEG-bonded stationary phase	6.49	62.15	31.36

Fig. 5 (a) FT-IR spectra and (b) TGA curves of silica and PEG-bonded stationary phase

Fig. 6 Chromatogram of the standard proteins separated by the PEG-bonded stationary phase using SEC Mobile phase A: 0.3 mol/L NaCl and 100 mmol/L NaH2PO4, (pH 6.8); mobile phase B: H2O; isocratic elution; A-B (75:25, v/v); flow rate: 0.3 mL/min; detector wavelength: 220 nm; column temperature: 25 ℃.

Fig. 7 Chromatogram of the standard proteins separated by the PEG-bonded stationary phase using HIC Mobile phase A: 0.1 mol/L L-arginine and 0.01 mol/L NaH2PO4 (pH 7); mobile phase B: 0.01 mol/L NaH2PO4 and 3 mol/L (NH4)2SO4; gradient elution: 0-5 min: 10%A, 5-35 min: 10%A-100%A; flow rate: 0.3 mL/min; column temperature: 25 ℃; detection wavelength: 280 nm.

Fig. 8 Chromatogram of the four polar compounds PEG-bonded stationary phase (HILIC) and their structural formulas Mobile phase A: acetonitrile; mobile phase B: H2O; gradient elution: A-B (60:40, v/v); column temperature: 25 ℃; flow rate: 0.3 mL/min; detector wavelength: 220 nm.

参考文献

1	Dejaegher B , Vander Heyden Y . J Sep Sci, 2010, 33(6/7): 698
2	Bouvier E S P , Koza S M . TrAC-Trends Anal Chem, 2014, 63: 85
3	Du Y , Cheng L , Chen L , et al. Microporous Mesoporous Mater, 2018, 265: 234
4	Grimalt S , Dehouck P . J Chromatogr A, 2016, 1433: 1
5	Zhang L , Wang S , Qu B , et al. J Pharm Biomed Anal, 2019, 170: 48
6	Guo H , Li X , Frey D D . J Chromatogr A, 2014, 1323: 57
7	Bonfatti V , Giantin M , Rostellato R , et al. Food Chem, 2013, 136(2): 364
8	Xia H , Wan G , Zhao J , et al. J Chromatogr A, 2016, 1471: 138
9	Baca M , De Vos J , Bruylants G , et al. J Chromatogr B Analyt Technol Biomed Life Sci, 2016, 1032: 182
10	Wei Y , Huang X , Liu R , et al. J Sep Sci, 2006, 29(1): 5
11	Zhao K , Yang L , Wang X , et al. Talanta, 2012, 98: 86
12	Rizza M D , Dellavalle P D , Vázquez D , et al. Cereal Chem, 2005, 82(3): 287
13	Hong P , Koza S , Bouvier E S P . J Liq Chromatogr Relat Technol, 2012, 35(20): 2923
14	Fekete S , Beck A , Veuthey J L , et al. J Pharm Biomed Anal, 2014, 101: 161
15	Soyekwo F , Zhang Q , Zhen L , et al. Chem Eng J, 2018, 337: 110
16	Felinger A , Pasti L , Dondi F , et al. Anal Chem, 2005, 77(10): 3138
17	Gey M , Thiem M , Gruel H . Acta Biotechnol, 1989, 9(1): 69
18	Yang Y , Qu Q , Li W , et al. J Sep Sci, 2016, 39(13): 2481
19	Guo Y , Gaiki S . J Chromatogr A, 2011, 1218(35): 5920
20	Buszewski B , Noga S . Anal Bioanal Chem, 2012, 402(1): 231
21	Verzele M , Velde N V D . Chromatographia, 1985, 20(4): 239
22	Blaschke G , Br?ker W , Fraenkel W . Angew Chem-Int Edit, 1986, 25: 830
23	Ali A , Ali F , Cheong W J . J Chromatogr A, 2017, 1525: 79
24	Ali A , Sun G , Kim J S , et al. J Chromatogr A, 2019, 1594: 72
25	Ali F , Cheong W J , Othman Z A , et al. J Chromatogr A, 2013, 1303: 9
26	Sun G , Ali A , Kim Y S , et al. J Sep Sci, 2019, 42(24): 3621
27	Hwang D G , Zaidi S A , Cheong W J . Bull Korean Chem Soc, 2009, 30(12): 3127
28	Ko J H , Baik Y S , Park S T , et al. J Chromatogr A, 2007, 1144(2): 269
29	Jin C Z , Wang J H , Wang Y J , et al. J Sol Gel Sci Technol, 2014, 70(1): 53
30	Kolesnikov A L , Uhlig H , M?llmer J , et al. Micropor Mesopor Mat, 2017, 240: 169
31	Yang W , Zhang C G , Qu H Y , et al. Anal Chim Acta, 2004, 503(2): 163
32	Yoo W C , Stein A . Chem Mat, 2011, 23(7): 1761
33	Li X , Wu D , Wang J , et al. Micropor Mesopor Mat, 2016, 226: 309
34	Galarneau A , Iapichella J , Bonhomme K , et al. Adv Funct Mater, 2006, 16(13): 1657
35	Huang M , Liu L , Wang S , et al. Langmuir, 2017, 33(2): 519
36	Wang Z , Zhang L , Guo B , et al. Chinese Journal of Chromatography, 2019, 37(5): 484
37	Song C , Wang J , Zhao K , et al. Biomed Chromatogr, 2013, 27(12): 1741
38	Hirano A , Arakawa T , Kameda T . J Chromatogr A, 2014, 1338: 58
39	Kanai H , Inouye V , Yazawa L , et al. J Chromatogr Sci, 2005, 43(2): 57

[1]	史洪飞, 徐伯芃, 徐成鑫, 周修齐, 许宏福. 溶剂萃取-高效液相色谱-四极杆飞行时间质谱法快速提取检测干燥恰特草中5种生物碱成分[J]. 色谱, 2023, 41(9): 771-780.
[2]	赵芮, 黄晴雯, 余智颖, 韩铮, 范楷, 赵志辉, 聂冬霞. QuEChERS-超高效液相色谱-串联质谱法同时测定水果中36种真菌毒素[J]. 色谱, 2023, 41(9): 760-770.
[3]	刘宇, 邢家溧, 沈坚, 毕晓丽, 毛玲燕, 徐晓蓉, 张书芬, 娄永江, 吴希, 穆应花. 高效液相色谱-蒸发光散射检测法同时测定固态食品中6种稀有糖[J]. 色谱, 2023, 41(9): 781-788.
[4]	岳超, 赵超群, 毛思浩, 王展华, 施贝, 徐欣丰, 梁晶晶. 通过式固相萃取净化-超高效液相色谱-串联质谱法测定液体乳中10种氨基甲酸酯类农药残留[J]. 色谱, 2023, 41(9): 807-813.
[5]	曹沁玲, 赵小丹, 沈国滨, 汪竹琴, 章弘扬, 张敏, 胡坪. 基于超高效液相色谱-电雾式检测的槐糖脂指纹图谱[J]. 色谱, 2023, 41(8): 722-729.
[6]	钱正明, 吴梦奇, 谭国英, 金李玲, 李宁, 谢菊英. 高效液相色谱紫外等吸收波长法快速测定秦皮中秦皮甲素和秦皮乙素[J]. 色谱, 2023, 41(8): 690-697.
[7]	吴萍萍, 林仁义, 黄丽英. 三相中空纤维液相微萃取-高效液相色谱法测定铁皮石斛和金线莲中3种植物生长调节剂[J]. 色谱, 2023, 41(8): 683-689.
[8]	马超, 倪洪星, 戚羽霖. 超高效液相色谱-傅里叶变换离子回旋共振质谱法解析溶解性有机质的化学多样性[J]. 色谱, 2023, 41(8): 662-672.
[9]	孟二琼, 念琪循, 李峰, 张秋萍, 许茜, 王春民. 磺酸化磁性氮化碳固相萃取-超高液相色谱-串联质谱筛检淡水鱼中孔雀石绿和隐色孔雀石绿[J]. 色谱, 2023, 41(8): 673-682.
[10]	陈东洋, 张昊, 张磊, 王一红, 王小丹, 冯家力, 梁静, 钟旋. 固相萃取-超高效液相色谱-串联质谱法测定发酵食品中的曲酸[J]. 色谱, 2023, 41(7): 632-639.
[11]	王秋旭, 冯启言, 朱雪强. 加速溶剂萃取-固相萃取净化结合超高效液相色谱-串联质谱法测定沉积物中双酚类化合物[J]. 色谱, 2023, 41(7): 582-590.
[12]	杨哲, 吕建霞, 吴一荻, 蒋力维, 李冬梅. 超高效液相色谱法同时测定电子烟油中的5种吲哚/吲唑酰胺类合成大麻素[J]. 色谱, 2023, 41(7): 602-609.
[13]	夏宝林, 汪仕韬, 殷晶晶, 张维益, 杨娜, 刘强, 吴海晶. 自动上样固相萃取-超高效液相色谱-串联质谱法同时测定水中9类43种抗菌药物残留[J]. 色谱, 2023, 41(7): 591-601.
[14]	童学智, 陈东洋, 冯家力, 范翔, 张昊, 杨胜园. QuEChERS-超高效液相色谱-串联质谱法快速测定水产品中5种卤代苯醌[J]. 色谱, 2023, 41(6): 490-496.
[15]	王园媛, 李璐璐, 吕佳, 陈永艳, 张岚. 固相萃取-超高效液相色谱-三重四极杆质谱法测定饮用水中13种卤代苯醌类消毒副产物[J]. 色谱, 2023, 41(6): 482-489.