Capillary electrochromatography with gradient elution.
Capillary electrochromatography with gradient elution.
复制标题
毛细管电色谱梯度洗脱。
DOI：
10.1021/ac970393t
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发表时间：
1997
影响因子：
7.4
通讯作者：
Horváth,C
中科院分区：
文献类型：
--
作者：
Huber,CG;Choudhary,G;Horváth,C
本文描述了一种具有梯度形成系统的毛细管电色谱仪，并以分离PTH-氨基酸和类固醇激素为例，说明了梯度洗脱在反相电色谱中的应用。梯度形成器采用两个往复式排量泵，以可控的方式控制柱入口处储液器中洗脱液的组成。因此，流经色谱柱并由电动驱动力驱动的流动相的组成可以像在梯度洗脱的高效液相色谱中所惯常的那样，以受控的方式随时间改变。该系统的设计还允许等度洗脱，通过将恒定组成的洗脱液通过柱入口处的空腔，从而连续提供新鲜的缓冲液。洗脱梯度由两台泵和一个10μL混合器产生。从那里，液体以0.1min0.2mLmin的流速通过装有柱进口和电极的17−L腔。流动相的有效总电场强度为500−1500V/cm，流动相通过填充有3.5μm十八烷基二氧化硅颗粒的50μm×20/12 cm毛细管柱。以这种方式产生的梯度分布具有很高的重复性。电雾流的同一天和每天的重现性被发现优于3%。用等度和梯度洗脱的方法证明了毛细管电色谱系统用于分离生物感兴趣物质的复杂混合物。考察了柱温对PTH-氨基酸电动流动速度和保留率的影响。
A capillary electrochromatograph incorporating a gradient-forming system generally employed in HPLC is described, and the use of gradient elution in reversed phase electrochromatography is demonstrated by the separation of PTH-amino acids and steroid hormones. The gradient former employs two reciprocating displacement pumps to control the composition of the eluent in the reservoir at the column inlet with time in a controlled manner. Thus, the composition of the mobile phase flowing through the column and driven by electrosmotic forces can be changed with time in a controlled fashion as customary in HPLC with gradient elution. The design of the system allows also for isocratic elution by pumping the eluent of constant composition through the cavity at the column inlet and thus continuously supplying fresh buffer. The eluent gradient is generated by the two pumps and a 10 μL mixer. From there the liquid passes at a flow rate of 0.1−0.2 mL/min through the 17 μL cavity housing the column inlet and an electrode. The flow of the mobile phase was electrosmotic at an effective overall electric field strength of 500−1500 V/cm through a 50 μm × 20/12 cm capillary column packed with 3.5 μm octadecylated silica particles. Gradient profiles generated in this manner were highly reproducible. The same-day and day-to-day reproducibilities of the electrosmotic flow were found to be better than 3%. The use of the capillary electrochromatographic system was demonstrated with isocratic and gradient elution for the separation of complex mixtures of biologically interesting substances. The influence of the column temperature on the electrosmotic flow velocity and retention of PTH-amino acids was also investigated.