The Column Dead Volume

If the column volume is (Vc) and contains a volume of mobile phase (Vm), a volume of stationary phase (Vs) and a volume of support (c.g.,silica) (Vsi), then 

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Recalling the plate theory, it must be emphasized that (Vm) is not the same as (Vm)-(Vm) is the moving phase and a significant amount of (Vm) will be static (e.g., that contained in the pores). It should also be pointed out that the same applies to the volume of stationary phase, (Vs), which is not the same as (Vs), which may include material that is unavailable to the solute due to exclusion. 

the column volume, assuming it has a circular cross-section, will be given by 

The individual volumes, (Vm), (Vs) and (Vsi), can now be apportioned to the different chromatographic activities. 

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Again it is seen that only when second order effects need to be considered does the relationship become more complicated. The dead volume is made up of many components, and they need not be identified and understood, particularly if the thermodynamic properties of a distribution system are to be examined. As a consequence, the subject of the column dead volume and its measurement in chromatography systems will need to be extensively investigated. Initially, however, the retention volume equation will be examined in more detail. 

The idea of the effective plate number was introduced and employed by Purnell [4], Desty [5] and others in the late 1950s. Its conception was evoked as a direct result of the introduction of the capillary column or open tubular column. Even in 1960, the open tubular column could be constructed to produce efficiencies of up to a million theoretical plates [6]. However, it became immediately apparent that these high efficiencies were only obtained for solutes eluted at very low (k ) values and, consequently, very close to the column dead volume. More importantly, on the basis of the performance realized from packed columns, the high efficiencies did not... 

Unfortunately, exclusion chromatography has some inherent disadvantages that make its selection as the separation method of choice a little difficult. Although the separation is based on molecular size, which might be considered an ideal rationale, the total separation must be contained in the pore volume of the stationary phase. That is to say all the solutes must be eluted between the excluded volume and the dead volume, which is approximately half the column dead volume. In a 25 cm long, 4.6 mm i.d. column packed with silica gel, this means that all the solutes must be eluted in about 2 ml of mobile phase. It follows, that to achieve a reasonable separation of a multi-component mixture, the peaks must be very narrow and each occupy only a few microliters of mobile phase. Scott and Kucera (9) constructed a column 14 meters long and 1 mm i.d. packed with 5ja... 

All the solutes in exclusion chromatography are eluted between the interstitial column volume and the column dead volume (i.e. the pore volume). Consequently, the column must be large enough to provide... 

A value for the column dead volume is required in most calculations. It is convenient to have one cosponent of the test mixture as an unretained solute. 

The concept of the effective plate number was introduced and employed in the late nineteen fifties by Purnell (7), Desty (8) and others. Its introduction arose directly as a result of the development of the capillary column, which, even in 1960, could be made to produce efficiencies of up to a million theoretical plates (9). It was noted, however, that these high efficiencies were were only realized for solutes eluted close to the column dead volume, that is, at very low k values. Furthermore, they in no way reflected the increase in resolving power that would be expected from such high efficiencies on the basis of the performance of packed columns. This poor performance, relative to the high efficiencies produced, can be shown theoretically ( and Indeed will be, later in this book) to result from the high phase ratio of capillary columns made at that time. That is the ratio of the mobile phase to the stationary phase in the column. The high phase ratio was... 

It is seen from equation (15) that the maximum overload volume is linearly related to the (k ) value of the first eluted solute of the critical pair, the function (a-1) and the column dead volume, Consequently, the larger the column, either in length and/ or radius, the larger the sample volume can be, This assumes that the column is of such a size, that it can be efficiently packed with practical techniques and that the particle size of the packing is chosen such that the pump pressure available can provide the necessary mobile phase flow-rate. 

Volume of the mobile phase in the column (dead volume)... 

Ion chromatography (1C) allows the separation of substances in the form of ions, chiefly in aqueous solutions. Mobile phases used in the technique contain relatively large amounts of salts stabilizing the pH and determining the sequence of analyte retention. They enable the separation of only cations or only anions the ions that are not separated by a selected phase (cationic or anionic) are eluted in the column dead volume. As the next step, they can be loaded into appropriate ion-exchange columns in the second chromatographic dimension [148]. 

The column dead volume can be defined as the space in the column which is not working for the chromatographic separations (i.e., not occupied by the stationary phase and its support). Its value is generally determined by an elution time or elution volume of a nonretained solute in the chromatographic system. If the column dead volume is measured by a retention time of a nonretained solute, one can refer to this as a column dead time t. ... 

Chromatographic conditions were 25°C, 1.0 ml./min. and 50-100 atm. H20 was used as a measure of the column dead volume. All of the solutes were freshly prepared In the mobile phase. 

Where is the retention volume of the chiral additive, when chromatographed as a solute in a mobile phase containing the additive at a concentration ( ( )). Equation (23) relates the retention volume of the additive (as opposed to the corrected retention volume) to its concentration in the mobile phase and thus, the need to determine the column dead volume is eliminated, and any errors associated with its measurement removed. 

An experimental example of column overload was demonstrated by Scott and Kucera [3] is shown in figure 12.2. The column dead volume was ca 2.5 ml. The chromatographic properties of the three solutes chromatographed on the column at 1 ml/min are shown in table 1. 

Several methods can be used to estimate the amount of surfactant molecules adsorbed on the stationary phase surface in a LC column. The titration, the breakthrough and the stripping methods will be described because they can produce reliable results with common chromatographic equipment. The results obtained with a given method can be used to check and to corroborate the results of another method. Accuracy of the method depends on the exact knowledge of the column dead volume which should be the first parameter to establish. 

The Lewis acid of zirconia can be blocked by other than fluoride. The effectiveness of the blocking of the Lewis acid sites should be related to the strength of the interaction between the Lewis base used and the zirconium ion coordination site. Blackwell and Carr have developed the relative eluotropic strength of a number of Lewis bases in terms of their ability to elute a wide variety of benzoic acid derivatives. Phosphate ranks at the top of the elution series. It brought about elution of the benzoic acid derivatives in the column dead volume. This proves that phosphate binds strongly to the surface of zirconia. The strong affinity of zirconia for phosphate suggested that a phosphate modification of the surface should be a reasonable approach. 

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