Separation time plate number

When only a few solutes are separated, they may occupy only a small portion of the total column volume at any given instant. In such cases, the productivity is improved by cyclic feed injections, timed so that the most strongly retained component from an injection elutes just before the least strongly retained component from the following injection (see Fig. 16-57). For a mixture of two components with k > 1, when the same resolution is maintained between bands of the same injections and bands of successive injections, the cycle time tc and the plate number requirement are ... 

As described above, resolution can be improved by variations in plate number, selectivity or capacity factor. However, when considering the separation of a mixture which contains several components of different retention rates, the adjustment of the capacity factors has a limited influence on resolution. The retention times for the last eluted peaks can be excessive, and in some cases strongly retained sample components would not be eluted at all. 

For coluzms with large plate numbers or operated with a vacuum outlet (as might be the case in GC/MS) such that P 1 the optimum separation time is given by equation (1.59)... 

A typical hplc column (25 cm x 4.6 mm, packed with a 5 jum bonded silica stationary phase) will have an efficiency corresponding to a plate number of 10 000-15 000. For many separations, this efficiency is far more than is needed, as often a plate number of 3000-5000 will give baseline resolution of all solutes. If this is the case, using a conventional column will waste analysis time and sol-... 

The object of a chromatographic separation is to achieve satisfactory resolution of solutes in the minimum time. Resolution is influenced by the capacity factor of the solutes and the selectivity and plate number of the column. 

In the case of more difficult separation problems therparticle diameter is reduced to 5 /im in order to obtain larger plate number or shorter analysis time. If systems with very low difihision odefficients have to be used, the particle dimensions could be farther reduced provided the finer particles are available in narrow enough size diitributioh,... 

Comprehensive 2D HPLC can be also operated under stop-flow mode. In this case, after transferring a desired fraction volume onto the secondary column, the flow of the mobile phase in the first dimension is stopped and the fraction analyzed in the second dimension. When the separation is finished, the mobile-phase flow in the first dimension is switched on and the whole procedure is repeated again for the analysis of all the transferred fractions. The disadvantage of this procedure is the long analysis time, while the advantage can be that the second-dimension column can give higher plate numbers if compared to the continuous approach [23]. 

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