Baltussen Erik, van Dedem Gijs W K
Laboratory of Analytical Biotechnology, Kluyver Institute for Biotechnology, Faculty of Applied Sciences, Delft University of Technology, Delft, The Netherlands.
Electrophoresis. 2002 May;23(9):1224-9. doi: 10.1002/1522-2683(200205)23:9<1224::AID-ELPS1224>3.0.CO;2-S.
At present, the main limitation for the further adoption of capillary electrochromatography (CEC) in the (routine) laboratory is caused by the lack of reproducible and stable columns. The main source of column instability is concentrated in the frits needed to retain the packed bed inside the CEC capillary. The sintering process used to prepare the frits can be rather problematic and irreproducible, particularly for small stationary phase particles and wide column diameters. Since the (surface) composition of the frits is different from the bulk stationary phase packing, different electroosmotic flow (EOF) velocities are generated. This effect is assumed to be primarily responsible for rapid column destruction. In this contribution, a novel approach for the preparation of fritless CEC capillaries is presented and evaluated. Using 5 microm Hypersil ODS particles, separation efficiencies in the range of 130,000-200,000 plates/m were obtained. In a 100 microm inner diameter packed column, electrical currents up to 50 microA could be tolerated without negative effects such as bubble formation. The prepared CEC columns were found to be stable and could easily be operated continuously for several days without column damage. An additional advantage of the proposed tapering approach is that application of pressure on the in- and outlet vial during separation was not required to prevent bubble formation.
目前，毛细管电色谱法（CEC）在（常规）实验室中进一步应用的主要限制因素是缺乏可重现且稳定的色谱柱。色谱柱不稳定的主要原因集中在用于固定CEC毛细管内填充床的烧结垫上。制备烧结垫所采用的烧结过程可能存在相当大的问题且不可重现，特别是对于小粒径的固定相颗粒和大内径的色谱柱而言。由于烧结垫的（表面）组成与整体固定相填料不同，会产生不同的电渗流（EOF）速度。据推测，这种效应是导致色谱柱快速损坏的主要原因。在本论文中，提出并评估了一种制备无烧结垫CEC毛细管的新方法。使用5微米的Hypersil ODS颗粒，获得了130,000 - 200,000理论塔板数/米的分离效率。在一根内径为100微米的填充柱中，能够耐受高达50微安的电流而不会产生诸如气泡形成等负面影响。所制备的CEC色谱柱被发现是稳定的，并且可以轻松地连续运行数天而不会损坏色谱柱。所提出的渐缩方法的另一个优点是，在分离过程中无需对进样瓶和出样瓶施加压力来防止气泡形成。
