Band broadening plate number

The contribution of the equipment between injection unit and detector cell should be negligable in relation to the column for a sufficient column characterization short connections with narrow capillaries and zero dead volume unions are the precondition for reliable plate numbers. Every end fitting of a column causes additional band broadening. In the past a column type was offered that could be directly combined without any capillary links unfortunately, it has disappeared from the market. 

Efficiency Degree of band broadening for a given retention time. It is expressed as the number of theoretical plates, N, or as the height equivalent to a theoretical plate, HEPT. 

The efficiency, or plate count of a column N is often calculated as 5.54 (tr/a)2, where tr is the retention time of a standard and a is the peak width in time units at half-height.1 2 5 This approach assumes that peaks are Gaussian a number of other methods of plate calculation are in common use. Values measured for column efficiency depend on the standard used for measurement, the method of calculation, and the sources of extra-column band broadening in the test instrument. Therefore, efficiency measurements are used principally to compare the performance of a column over time or to compare the performance of different columns mounted on the same HPLC system. 

CE is a technique with a very high power of resolution. This is attributed to low diffusion and high plate numbers obtained from the absence of band-broadening factors (e.g., eddy diffusion, equilibrium dynamics, etc.) other than diffusion, which is also minimized by short analysis time. 

As readily observed in most chromatograms, peaks tend to be Gaussian in shape and broaden with time, where W, becomes larger with longer This is caused by band-broadening effects inside the column, and is fundamental to all chromatographic processes.The term, plate number (N), is a quantitative measure of the efficiency of the column, and is related to the ratio of the retention time and the standard deviation of... 

Electrochromatography involves the use of a stationary phase hence the concepts of efficiency and band broadening are similar to those that occur in conventional liquid chromatography. The efficiency, expressed in number of theoretical plates N can be obtained directly from an electropherogram using the following equation ... 

Both the number of theoretical plates and the height eqnivalent to a theoretical plate estimate the band broadening occnrring in the separation system, either related to the whole capillary tnbe (AT) or to the portion of the capillary occupied by the analyte (H). The higher is N (or lower is H) the narrower are the recoded peaks in the electropherogram. 

The ideal model and the equilibrium-dispersive model are the two important subclasses of the equilibrium model. The ideal model completely ignores the contribution of kinetics and mobile phase processes to the band broadening. It assumes that thermodynamics is the only factor that influences the evolution of the peak shape. We obtain the mass balance equation of the ideal model if we write > =0 in Equation 10.8, i.e., we assume that the number of theoretical plates is infinity. The ideal model has the advantage of supplying the thermodynamical limit of minimum band broadening under overloaded conditions. 

As long as the sample occupies less than 0.5(n) theoretical plates, there will be no band broadening because of sample size As the total number of theoretical plates increases, within a column, the maximum space (in terms of theoretical plates) that should be occupied by the sample will also increase. However, the percentage of column length available for sample will decrease (because number of theoretical plates per column length increases), as shown in Table 2.3... 

The theoretical plate concept in chromatography is a popular approach to determining column efficiency (relative band broadening in the column). The number of theoretical plates, N, is related to the retention time and to the width of the solute peak by... 

The efficiency of a column is a number that describes peak broadening as a function of retention, and it is described in terms of the number of theoretical plates, N. Two major theories have been developed to describe column efficiency, both of which are used in modern chromatography. The plate theory, proposed by Martin and Synge,31 provides a simple and convenient way to measure column performance and efficiency, whereas the rate theory developed by van Deemter et al.32 provides a means to measure the contributions to band broadening and thereby optimize the efficiency. 

E. Extracolumn Band Broadening or Variance To maximize the effective number of theoretical plates, the contribution of the entire chromatographic system to band broadening (system variance, o-2ys) must be minimized. The system variance may be broken down into contributions from the column variance, a 01, as described above, and extracolumn diffusion and mixing processes, cr2x. As with the case of the column variance, extracolumn variance is an additive property and may be broken down into the major components ... 

In pressure-driven operation, considerable band broadening was observed at high linear velocity, although the separation impedance was much lower than that of a particle-packed column owing to the much lower flow resistance. The separation impedance (E = AP to / r N2 = (AP / N) (to / N) (l/r )) expresses the total column performance in terms of the reciprocal number of theoretical plates per unit time and pressure drop. Because the contributions of the B- and C-terms are expected to be similar for a pressure-driven mode and an electro-driven mode, the difference in performance can be attributed to the greater contribution of the A-terms in Eqn. 5.2 in the pressure-driven mode. The contribution of the A-term is known to be less in CEC than in HPLC [6],... 

A variety of surfaces such as metals, plastics, and glass can retain proteins during a separation process. In CE, a problem is manifested in the adsorption of proteins by fused silica capillaries (see Section IV). This problem is attributed to the adsorption of positively charged sites of proteins on negatively charged sites (silanol groups) on the capillary wall—a process that leads to band broadening and a much lower number of theoretical plates than would be expected on the basis of theory. 

The improvement in the resolution of a separation is a multiple of the square root of the number of plates which the sample encounters. Even if some band broadening occurs because the sample passes repeatedly through the pump, injector, or detector, it is still possible to attain a distinct separation, as demonstrated in the 14-pass recycle shown in Figure 6-11. 

In capillary gel electrophoresis, one of the major contributors to band broadening, besides the injection and detection extra-column effects, is the longitudinal diffusion of the solute molecules in the capillary tube [14], The theoretical plate number (N) is characteristic of column efficiency ... 

If all other causes of band broadening are neglected, the plate number is directly proportional to the electric field E and inversely proportional to the diffusion coefficient D of the solute in the electrolyte. The relationship is given by ... 

Other effects may also contribute to band broadening causing reduced achievable plate counts. Besides the already-mentioned wall adsorption, temperature effects (Joule heating) may reduce plate numbers. Sample application can have a strong influence on plate count, especially when large volumes and/or high sample concentrations are injected. Mobility differences between buffer constituents and analyte ions lead to asymmetric (triangular) peaks caused by electrodispersion, which is extremely noticeable with smaller molecules. Differ-... 

Band broadening within the chromatographic column is described as a function of mobile phase linear velocity ( ) by the well known van Deemter equation [5] that relates the height equivalent to the theoretical plate (HETP) to u the lower the HETP, the higher the plate number per unit length of a column. It follows that the highest efficiency is obtained for the shortest HETP. The van Deemter equation is ... 

The efficiency is the measure of the chromatographic band broadening and the number of the theoretical plates (AO in the column and is usually calculated using the following equation ... 

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