PY5010 Capillary Electrophoresis

https://www.youtube.com/watch?v=3X-ZSdF9hWg

- Electroosmotic flow
- Movement of bulk solution
- Movement of the separation buffer through the silica capillary as a result of
the existence of zeta potential at the solvent/silica interface
- Fused silica capillary
- Inner diameter 10-100 micrometres
- Length 40-100 cm
- Usually coated with thin polyimide coating
- Si-OH silica surface makes the coating polar
- pH dependent silanol groups
- When <3, silanol groups are fully protonated
- When >9 silanol groups are almost fully deprotonated
- -vely charged surface attracts cations causing
drag/friction
- Tightly adsorbed cations cause fixed
layer
- -vely charged surface can still attract cations
from solution causing diffuse layer
- Termed EDL with fixed layer
- In bulk solution, [cations] and [anions] is approximately
the same
- 3-9 silanol groups partially protonated
 - Applying voltage across capillary
- When V applied, cations in diffuse layer will be attracted to -ve
cathode dragging bulk solution with them
- Termed electroosmotic flow
- Imagine the tube is fixed in position and on top of the wall we
have the fixed and diffuse layers and [cations] > [anions] with
bulk solution in the centre
- Applying V, cations will be attracted towards -ve
cathode, dragging bulk solution along with them
- Generally, CE takes on a flat profile rather than a parabolic profile like in
HPLC

- EO flow minimises band broadening, improving peak resolution and

separation efficiency
- Unique flat profile is the main reason for higher separation resolution
and efficiency
- Doesn’t tell us about separation of analytes
- How do we separate anions, cations and neutral species in
CE?
- Electrophoretic mobility
- Electrophoretic mobility
- Separation of charged species in solution
- Applying V to capillary attracts cations to cathode and anions to anode
- Since neutral species aren’t charged this doesn’t affect them
- Directly proportional to ionic charge on analytes
- If same size, cations which are more highly charged will move faster
towards the cathode
- Same for anions/anode
- Frictional retarding force of analyte
- If same charge, bulkier cations move slower due to friction towards the
cathode
- Same for anions/anode
- Affected by viscosity of medium too – more viscous means slower
movement towards anode/cathode
- Electrophoretic mobility is inversely proportional to frictional
retarding force
 - Since frictional retarding factors and electrophoretic mobility are
related, the size referred to above is the Stokes radius
- What is Stokes radius?

- Stokes radius
- Radius of a hard sphere that diffuses at the same rate as that solute
- It is possible that a smaller ion with stronger hydration may have a greater
Stokes radius than a larger ion with weaker hydration

- vtotal = velectrophoretic + velectroosmotic

- Cations – add electrophoretic and electroosmotic velocities
- Anions – electrophoretic migration is in the opposite direction from the
electroosmotic flow
- Rate of EO flow is generally greater than the rate of EP migration of
individual ions
- Therefore in this case after combining EO and EP, both anions
and cations will move towards the cathode with different
migration rates depending on charge and size