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Total Volume of Stationary Phase in the Column 0.40ml
Volume of Chrornatographically Available Stationary Phase 0,49ml
Volume of Chrornatographically Unavailable Stationary Phase (by difference)
Total Mobile Phase in the column.
1/By Weighing 2.79m
2/ From the Retention of the alkanes in n-Octane extrapolated
to an alkane carbon number of 3 2.78ml
3/ From the volume fraction average of the isotopes retention
Total Interstitial Volume. Value extrapolated from the retention
volumes of ions of different size. 1.91 ml
Interstitial Moving Phase Volume. From the Retention of 'Silica Smoke' 1.41ml
Interstitial Static Phase Volume. By Difference 0.50ml
Total Pore Volume, By Difference 0.87m)
Pore Volume Containing Components of the Mobile Phase of Different
composition to that of that of the Moving Phase.( by difference) 0.18m 1
Pore Volume Containing Mobile Phase of the Same Composition as the
Moving Phase, (by difference) 0.70ml
It is seen that the distribution of the various chromatographically important volumes within an LC column is neither simple nor obvious, it is also seen that about 70% of the column volume is occupied by mobile phase but only about 50% of that mobile phase is actually moving. Furthermore about 18% of the mobile phase is interstitial but static and about 31% of the mobile phase is contained within the pores and is also static. Just over 6% of the mobile phase in the pores has a different composition to that of the mobile phase proper and thus constitutes a second stationary phase The stationary phase constitutes about 12% of the column volume which is equivalent to about 17% of the mobile phase content of the column.
The values given in table (2) are probably representative of most reverse phase columns but may differ significantly from silica gel columns.
U) H. tngelhardt, H. nul ler and B. Dreyer, Chromatographta, 19( 1984)240.
(2) P L Zhu, Chromatograph/a, 20( 1985)425.
(3) J. H Knox and R. J Kaliszan, J Chromatogr, 349( 1985)211.
(4) R. J. Smith and C. S. Nieass, J. L ip. Chromatogr. ,9( 1986) 1387.
(5) H. Colin, A. Krstulovic, and G. Guiochon, Ana/. Chem ,54(1982)2438.
(6) R. A. D jerki and R. J Laub, J L /ä. Chromatogr. ,10(1987) 1749
(7) E. Grushka, J. Liq Chromatogr, 5( 1982)1392
(8) R. P. W Scott and P Kucera, J Chromatogr. 149(1978)93.
(9) R. M. McCormick and B. L Karger, Ana/. Chem. ,52(1980)2249.
(10) R. P. W. Scott and C. F. Simpson, FaradySymp. Chem Soc. 15(1980)69 (IDA. Alhedai, D. E. Martire and R. P. W. Scott, Ana/yst,\lo\.! 14 (1989)869
(12) J. J. Van Deemter, F. J. Zuiderweg and Ë. Klinkenbcrg,
Chem. Eng. $à ,5( 1956)271.
(13) M. J. E. Golay, in "GasChromatography /958 " (Ed. D.H.Desty), Butterworths, London, (1958)36.
Extensions of the Plate Theory
So far the Plate Theory has been used to determine the equation for the retention volume of a solute, calculate the capacity factor of a solute and identify the dead volume of the column and how it should be calculated. However, the equation for the elution curve of a solute that arises directly from the Plate Theory can do far more than that to explain the characteristics of a chromatogram. The equation will now be used in a variety of ways to expand our knowledge of the chromatographic process.
The Elution Curve of a Finite Charge
In the development of the plate theory and the derivation of the equation for the elution curve of a solute, it was assumed that the initial charge was located in the first plate of the column. In practice, this is difficult to achieve, and any charge will, in fact, occupy a finite column volume and consequently a specific number of the first theoretical plates of the column Consider the situation depicted in figure 1 where the initial charge is distributed over (r) theoretical plates.
The Injection of a Finite Volume of Charge onto an LC Column
Let the mobile phase flow be arrested, and a volume of sample be placed on the column such that it occupies (r) theoretical plates. Now on starting the
COllii'iiM flow, the LOniefitS 01 each pidle wiH ueveiOp dii eiUllOn Lufve, me
points on each curve being one plate volume away from the complimentary points on the adjacent curve. The first peak will result from the elution of
the contents of the (r)'th plate and the last peak from the elution of the contents of the first plate. The peak actually monitored by the detector will be the composite peak formed by the addition of all the individual peaks. It should also be noted that on development, the first peak will pass through all the plates in the column, i.e. (n) plates.However, the sample in plate (r) will only pass through (n-r) plates as it reached the (r)'th plate on injection of the sample. It follows that the contents of plate (r) will start development as though (r) plate volumes of mobile phase had already passed through the column.
The elution curve resulting from that portion of the sample on plate one will be given by the following equation,