Page 68 - Subyek Encyclopedia - Encyclopedia of Separation Science

Sepsci*21*TSK*Venkatachala=BG
I / CHROMATOGRAPHY 63

pressure transducer is installed between the pump plied to the layer as spots or narrow bands using
and sample inlet for programming the inlet pressure low volume dosimeters or spray-on techniques.
or Suid density during the course of a separation. Application volume, method, location and sample
Simultaneous measurement of the column temper- sequence are automated for unattended operation.
ature and pressure control allows constant density or The chromatogram is obtained by manual develop-
density programming under computer control if the ment in a number of development chambers of dif-
appropriate isotherms are known or can be approxi- ferent design, or can be automated such that the
mated. Two pumps are generally used to generate conditioning of the layer, the selected solvents for the
mobile-phase composition gradients comprising development, and the development length are
liquid carbon dioxide and an organic solvent. Loop- preselected and controlled through the use of sensors.
injection valves similar to LC are the most convenient For multiple-development techniques the layer can be
devices for sample introduction. The column oven alternately developed, dried, new solvent introduced
is usually a forced air circulation thermostat similar and the process repeated with changes in the develop-
to those used in GC. The full range of Same- ment length and mobile-phase composition for any or
based detectors used in GC can be used with only all the programmed development steps. Apparatus
slight reoptimization as well as the main optical de- for forced-Sow development is also available and
tectors used in LC, after modiRcation for high pres- resembles a liquid chromatograph with the column
sure operation. A unique feature of the chromato- replaced by the layer sandwiched between a rigid
graph is a restrictor required to maintain constant support and a polymeric membrane in an over-
density along the column and to control the linear pressure development chamber to allow external
velocity of the Suid through the column. OriRce-type pressure to be used to create the desired mobile-phase
restrictors are usually placed between the column and velocity.
detector for Same-based detectors and back-pressure After development the chromatogram is recorded
regulators after the detector Sow cell for optical using scanning or video densitometry. The unique
detectors. feature compared with detection in column
chromatography is that the separation is recorded in
MEKC and CEC space rather than time while in the presence of the
stationary phase. The common forms of detection are
MEKC and CEC employ the same instruments as optical methods based on UV}visible absorption and
used for capillary electrophoresis with the addition of Suorescence. In mechanical scanning the layer is
overpressure capability for the buffer reservoirs moved on a translation stage under a slit projecting
when used for CEC. The separation capillary is ter- the image of the monochromatic light source on the
minated in two buffer reservoirs containing the layer surface and the light reSected from the surface
high voltage electrodes that provide the electric Reld monitored continuously with a photodiode or similar
to generate the Sow of mobile phase. The buffer device. Substances that absorb the light produce
reservoirs can be moved into place pneumatically and a proportional decrease in the intensity of the reSec-
sequenced automatically to introduce a sample vial ted light that can be related to the amount of sample
for sample introduction or a run buffer vial for present (for Suorescence there is a proportionate in-
separation. The column area is thermostated to main- crease in the amount of light emitted at a wavelength
tain a constant temperature. A miniaturized optical that is longer than the absorbed wavelength). Elec-
detector positioned between the buffer reservoirs
tronic scanning is not as well developed but involves
is commonly used for on-column detection. Some
uniformly illuminating the whole layer and imaging
form of interlock mechanism is used to prevent oper-
the plate surface onto a video camera, or similar
ator exposure to the high voltages, up to 30}50 kV,
device, to capture and integrate the static image of the
typically used. A high level of automation is achieved
absorbing zones.
under computer control and unattended operation is
generally possible.
Conclusion
Planar Chromatography
Many of the important developments in chromato-
The total automation of sample application, graphy have already been made, yet the technique
chromatogram development and in situ quantitation continues to evolve by the introduction of new mater-
in planar chromatography has proved difRcult. ials that extend the scope of existing methods and
Instead the individual procedures are automated, re- through Rnding new applications. General applica-
quiring operator intervention to move the layer from tions are dominated by the techniques of gas
one operation to the next. Samples are typically ap- chromatography and column liquid chromatography,