1μm C18固定相的制备及在手性加压毛细管电色谱中的应用

Preparation of 1 μm non-porous C18 silica gel stationary phase for chiral-pressurized capillary electrochromatography

制备了粒径为1μm的无孔C18固定相,并将其应用在手性加压毛细管电色谱中用以分离对映体。以改良的Stber法先合成粒径为1μm的无孔二氧化硅微球,后采用二次硅烷化方法制备出1μm C18固定相。以加压毛细管电色谱为平台,通过考察影响分离的各因素,确定了最佳拆分条件:乙腈/5 mmol/L pH 4.0乙酸铵缓冲液(20/80,v/v),羧甲基-β-环糊精(CM-β-CD)浓度15 mmol/L,泵流速0.03 mL/min,分离比约300∶1,施加电压2kV。本研究采用的二次硅烷化方法提高了C18的键合量。和毛细管液相色谱相比,4种药物在加压毛细管模式下有更好的分离效果,柱效最高为19万理论塔板数/m。盐酸安非他酮、盐酸克伦特罗、酒石酸美托洛尔、盐酸艾司洛尔对映体的分离度分别为1.55、2.82、1.69、1.70。该研究为手性流动相添加剂法在加压毛细管电色谱中的应用以及微米级填料在手性色谱中的应用提供了新的思路。

Non-porous C18 silica gel stationary phase（1 μm）was prepared and applied to chiral separation in pressurized capillary electrochromatography（pCEC）for the enantioseparation of various basic compounds. The non-porous silica particles（1 μm）were synthesized using modified Stber method.C18 stationary phase（1 μm）was prepared by immobilization of chloro-dimethyl-octadecylsilane. Using carboxymethyl-β-cyclodextrin（CM-β-CD）as the chiral additive,the pCEC conditions including the content of acetonitrile（ACN）,concentration of buffer,pH,the concentration of chiral additive and flow rate as well as applied voltage were investigated to obtain the optimal pCEC conditions for the separation of four basic chiral compounds. The column provided an efficiency of up to 190 000 plates / m.Bupropion hydrochloride,clenbuterol hydrochloride,metoprolol tartrate,and esmolol hydrochloride were baseline separated under the conditions of 5 mmol / L ammonium acetate buffer at pH 4. 0 with20%（v / v）acetonitrile,and 15 mmol / L CM-β-CD as the chiral additive. The applied voltage was 2kV and flow rate was 0. 03 mL / min with splitting ratio of 300 ∶ 1. The resolution were 1. 55,2. 82,1. 69,1. 70 for bupropion hydrochloride,clenbuterol hydrochloride,metoprolol tartrate,esmolol hydrochloride,respectively. The C18 coverage was improved by repeating silylation method. The synthesized1 μm C18 packings have better mechanical strength and longer service life because of the special,nonporous structure. The column used in pCEC mode showed better separation of the racemates and a higher rate compared with those used in the capillary liquid chromatography（cLC）mode. This study provid-ed an alternative way for the method of pCEC enantioseparation with chiral additives in the mobile phase and demonstrated the feasibility of micron particle stationary phase in chiral separation.