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When an analytes retention factor is less than one, elution is so fast that accurate determination of the retention time is very difficult.High retention factors mean that elution takes a very long time.Ideally, the retention factor for an analyte is between one and five.This entails how well the column is   packed and its kinetic performance.For this reason efficiency can be an enigmatic value since manufacturers may use different methods in determining the efficiency of their columns.The plate model supposes that the chromatographic column is contains a large number of <a href="http://www.targetmol.com/compound/D-Cycloserine">Targetmol's D-Cycloserine</a> Separate layers, called theoretical plates.Separate equilibrations of the sample between the stationary and mobile phase occur in these plates.The analyte moves down the column by transfer of equilibrated mobile phase from one plate to the next.It is important to remember that the plates do not really exist; they are a figment of the imagination that helps us understand the processes at work in the column.For the same reason as inflection method, this measurement is not affected by asymmetry; however, this method is more reproducible from person to person since width at peak height is less prone to be varied.The number of theoretical plates that a real column possesses can be found by examining a chromatographic peak after elution.The resulting band shape of a chromatographic peak is therefore affected by the rate of elution.It is also affected by the different paths available to solute molecules as they travel between particles of stationary phase.A, B, and C are factors which contribute to band broadening.Solute molecules will take different paths through the stationary phase at random.This will cause broadening of the solute band, because different paths are of different lengths.If the velocity of the mobile phase is high then the analyte spends less time on the column, which decreases the effects of longitudinal diffusion.Cm is resistance to mass transfer brought about by the diameter and shape of the particles of stationary phase and the rate of diffusion of a molecule in the mobile phase.Another measure of how well species have been separated is provided by measurement of the resolution.An increase in N, the number of theoretical plates, by lengthening the column leads to an increase in retention time and increased band broadening which may not be desirable.Instead, to increase the number of plates, the height equivalent to a theoretical plate can be reduced by reducing the size of the stationary phase particles.When is close to unity, optimising k and increasing N is not sufficient to give good separation in a reasonable time.The separation is based on adsorption, partition, ionexchange or on combinations of these mechanisms and is carried out by migration in a solvent or a suitable mixture of solvents. It may be necessary to wash the plates prior to separation.This can be done by migration of an appropriate solvent.The plates may also be impregnated by procedures such as development, immersion or spraying.At the time of use, the plates may be activated, if necessary, by heating in an oven at C for min.Chromatographic tank: A chromatographic tank with a flat bottom or twin trough, of inert, transparent material, of a size suitable for the plates used and provided with a tightly fitting lid is required.

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