The difference between High Performance Capillary Electrophoresis (HPCE) and general electrophoresis is the use of capillary columns. The inner wall of the fused silica capillary is covered with a layer of siloxy (Si-O) anion, which attracts an ionic cation in the solution, thereby forming an anionic double layer on the inner wall of the capillary. The cations enriched in the outer edge diffusion layer are attracted by the electric field cathode to cause the solution to flow toward the cathode. This effect is called electroosmosis. The velocity of the electroosmotic flow, ueo, is proportional to the electric field strength E. The ratio of the electroosmotic velocity μeo to the field strength E is defined as the electroosmotic enthalpy μeo, ie μeo=ueo/E
The electroosmotic twist is proportional to the charge density on the surface of the silicon oxide layer and inversely proportional to the square root of the ionic strength. At low pH, the silicon oxide layer does not form a molecule (Si - OH), thus reducing the surface charge density, so the electroosmotic velocity is reduced. For example, the electroosmotic rate is about 2 mm / s in the sand buffer of pH 9, and the electroosmotic speed is reduced by about an order of magnitude in the medium of pH 3. An important factor affecting electroosmosis is the Joule heat generated by the action of current in the capillary, which can make the temperature at the center of the column = higher than the temperature at the edge, forming a parabolic temperature gradient, the temperature near the tube wall is low, and the center temperature is high. The electroosmotic velocity is not uniform, which causes the zone to widen and the column efficiency to decrease. For this reason, the use of capillary columns that are too long and have an inner diameter of more than 50 μm should be avoided. Care should also be taken to reduce the wall thickness of the capillary, select the appropriate voltage and buffer, and use a good cooling system.
The ion mobility observed in HPCE is the sum of the electrophoretic mobility μep of the ion and the electroosmotic mobility of the solution. Define the apparent degree as μapp, then
Appapp=μep+μeo
According to the above discussion, the positively charged ions have μep>0, μeo>0, so μapp is always a positive sign and the ions move toward the cathode; while the negatively charged ions are repelled by the cathode due to the electrophoretic flow, μep<0 At high pH, ​​if μeo>μep, μapp is still positive, the ions can still move toward the cathode, but at low pH, μapp can be negative and the ions will move in the opposite direction. At this point, the direction of the electric field must be changed to detect the ions to be analyzed. For components whose actual speed is unet (net refers to net speed), the performance intensity can be calculated by:
Where Ld: actual column length from the inlet to the detector; Lt: total column length; V: voltage; t: required analysis time.
Neutral components can be used to actually test the electroosmotic twist. At this time, μep=0, μeo=μapp, μep=μeo is as follows:
The capillary of HPCE is easy to cool, so a high voltage of 20 to 30 kV can be used. Since the inner diameter of the tube is only 25 to 100 μm, no eddy current is diffused, and the mass transfer impedance tends to zero, so that the resolution is high. The electrolyte liquid enters the capillary from the anode end of the capillary, and the separated components can flow into the colorimetric cell of the detector from the cathode end of the capillary. The electrophoresis process and the result analysis are easy to be automated, and have become an extremely efficient analytical instrument.
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