Microchip electrophoresis (MCE) is widely applied in food, environment, medicine, and other fields, owing to its high separation efficiency, low consumption of reagents and samples, and ease of integrating multiple operating units. Polymer microchip materials like cycloolefin copolymer (COC) are low-cost and easy to fabricate. However, their practical applications are limited by the non-specific adsorption on channel surface during electrophoresis and the instability of electroosmotic flow. These shortcomings can be solved by COC surface modification. In this study, a static coating and dynamic/static coating combined strategy was used to develop a channel-surface-modified COC microchip. Combined with laser-induced fluorescence (LIF) detection, a MCE-LIF separation and analysis method was developed for detecting functional components in health care products. The separation performance of MCE was improved by the static coating microchannel surface modification method. The static coating was constructed by hydrophobic amino acid adsorption, glutaraldehyde immobilization, and hydrophilic amino acid functionalization on the COC microchannel surface. The separation performance of MCE was improved by microchannel surface modification combined with dynamic/static coating. The static coating was constructed by valine adsorption, carboxyl activation, and ethylenediamine functionalization on the COC microchannel surface. The dynamic coating is automatically formed by introducing a buffer solution containing hydroxypropyl methylcellulose and sodium dodecyl sulfate into the microchannel. The physical and chemical properties of surface-modified microchannels and the factors governing electrophoretic separation were studied. Combined with LIF detection, the MCE-LIF separation and analysis of lysine and γ-aminobutyric acid present in children's health care products, as well as aspartic acid and taurine in sport drinks, were developed. The recoveries of lysine and γ-aminobutyric acid in children's health care products were 84.8%-118%, and the relative standard deviations (RSDs) were less than 7.2% (n=3). The recoveries of aspartic acid and taurine in sport drinks were 97.5%-118%, and the RSDs were less than 6.4% (n=3). The analysis results are consistent with the HPLC results, and the method has potential for application in the separation and analysis of anionic amino acids in health care products.
微芯片电泳(microchip electrophoresis, MCE)分离效率高、试剂和样品消耗量少、易实现多操作单元集成,在食品、环境和药物等领域应用广泛。环烯烃共聚物(cycloolefin copolymer, COC)等聚合物微芯片材料成本低、制作简便,但电泳过程中通道表面易发生非特异性吸附,且电渗流不稳定,限制了其应用。这些不足可通过COC表面改性解决。本文采用静态涂层和动/静态涂层联合策略,研制通道表面改性COC微芯片,结合激光诱导荧光(laser-induced fluorescence, LIF)检测,发展了简单高效的MCE-LIF分离分析方法,用于保健品中功效成分检测。通过负电荷涂层微通道表面改性方法,可以提高MCE对阳离子氨基酸的分离效果,而通过动/静态涂层结合的微通道表面改性方法,可以提高MCE对阴离子氨基酸的分离效果。负电荷涂层由COC微通道表面的疏水氨基酸吸附、戊二醛固定化和亲水氨基酸功能化构建,正电荷涂层由COC微通道表面的缬氨酸吸附、羧基活化和乙二胺功能化构建,动态涂层通过向微通道引入含有羟丙基甲基纤维素与十二烷基硫酸钠的缓冲液自动形成。实验研究了表面改性微通道的理化性质和电泳分离的影响因素,并将所建立的方法用于儿童保健品中赖氨酸和γ-氨基丁酸以及运动饮料中天冬氨酸与牛磺酸MCE-LIF的检测,儿童保健品中赖氨酸和γ-氨基丁酸的加标回收率为84.8%~118%,相对标准偏差(RSD)≤7.2%,运动饮料中天冬氨酸与牛磺酸的加标回收率为97.5%~118%, RSD≤6.4%。分析结果与HPLC方法的测定结果吻合,该法在保健品中氨基酸分离分析中有应用前景。
Keywords: cycloolefin copolymer (COC); electrophoresis; health care products; microchips; surface modification.