Chromatographic efficiency

The separation ability of a chromatographic column is often measured by the number of theoretical plates, N. This concept comes originally from distillation theory in which the ability to separate volatile compounds by fractional distillation was related to the number of actual plates in the packed distillation column. In chromatography the number of theoretical plates in a column is calculated from the retention time, t, and the average peak width, lu 

Since the shape of chromatographic peaks is essentially the same as a standard error plot in statistics, the peak width at the base may be written in terms of standard deviation. 

A convenient way to measure a from a chromatogram is to apply the relationship that the peak vsddth at one-half the peak height is equal to 2.35 cr. 

Neff tends to be very low when the difference between t and to is sUght and is a better indication of separation power than N. 

In classical theory, N was used as a measure of separation power because N increased with greater column length. Height equivalent of a theoretical plate, H, served as a measure of chromatographic efficiency. H in mm or cm is calculated by dividing the column length, I, by the plate number 

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The purification of value-added pharmaceuticals in the past required multiple chromatographic steps for batch purification processes. The design and optimization of these processes were often cumbersome and the operations were fundamentally complex. Individual batch processes requires optimization between chromatographic efficiency and enantioselectivity, which results in major economic ramifications. An additional problem was the extremely short time for development of the purification process. Commercial constraints demand that the time interval between non-optimized laboratory bench purification and the first process-scale production for clinical trials are kept to a minimum. Therefore, rapid process design and optimization methods based on computer aided simulation of an SMB process will assist at this stage. 

A second simulation study was performed to measure the effect on both extract and raffinate purities of a loss of chromatographic efficiency (Fig. 10.11). 

The graph in Fig. 10.11 shows that the SMB can tolerate a loss of 13 % chromatographic efficiency and still reach a purity of greater than 98 %. The industrial SMB system was designed to operate with 300 theoretical plates without any modification of the operating flowrates. 

The use of both sub- and supercritical fluids as eluents yields mobile phases with increased diffusivity and decreased viscosity relative to liquid eluents [23]. These properties enhance chromatographic efficiency and improve resolution. Higher efficiency in SFC shifts the optimum flowrate to higher values so that analysis time can be reduced without compromising resolution [12]. The low viscosity of the eluent also reduces the pressure-drop across the chromatographic column and facilitates the... 

Other properties of solvents which need to be considered are boiling point, viscosity (lower viscosity generally gives greater chromatographic efficiency), detector compatibility, flammability, and toxicity. Many of the common solvents used in HPLC are flammable and some are toxic and it is therefore advisable for HPLC instrumentation to be used in a well-ventilated laboratory, if possible under an extraction duct or hood. 

Since reproducibility of the flow system is critical to obtaining reproducibility, one approach has been to substitute lower-performance columns (50-to 100-p packings) operated at higher temperatures.1 Often, improvements in detection and data reduction can substitute for resolution. Chemometric principles are a way to sacrifice chromatographic efficiency but still obtain the desired chemical information. An example of how meaningful information can be derived indirectly from chromatographic separation is the use of system or vacancy peaks to monitor chemical reactions such as the titration of aniline and the hydrolysis of aspirin to salicylic acid.18... 

E. Palsso, A. Axelsson and P.-O. Larsson, Theories of chromatographic efficiency applied to expanded beds. J. Chromatogr.A 912 (2001) 235-248. 

A copolymerization approach of 0-9-[2-(methacryloyloxy)ethylcarbamoyl] cinchonine and cinchonidine with methacryl-modified aminopropylsilica particles was utilized by Lee et al. [71] for the immobilization of the cinchona alkaloid-derived selectors onto silica gel. The CSPs synthesized by this copolymerization procedure exhibited merely a moderate enantiomer separation capability and only toward a few racemates (probably because they were based on less stereodifferentiating cinchonine and cinchonidine). Moreover, the chromatographic efficiencies of these polymer-type CSPs were also disappointing. 

It is apparent from early observations [93] that there are at least two different effects exerted by temperature on chromatographic separations. One effect is the influence on the viscosity and on the diffusion coefficient of the solute raising the temperature reduces the viscosity of the mobile phase and also increases the diffusion coefficient of the solute in both the mobile and the stationary phase. This is largely a kinetic effect, which improves the mobile phase mass transfer, and thus the chromatographic efficiency (N). The other completely different temperature effect is the influence on the selectivity factor (a), which usually decreases, as the temperature is increased (thermodynamic effect). This occurs because the partition coefficients and therefore, the Gibbs free energy difference (AG°) of the transfer of the analyte between the stationary and the mobile phase vary with temperature. 

H is the plate height (cm) u is linear velocity (cm/s) dp is particle diameter, and >ni is the diffusion coefficient of analyte (cm /s). By combining the relationships between retention time, U, and retention factor, k tt = to(l + k), the definition of dead time, to, to = L u where L is the length of the column, and H = LIN where N is chromatographic efficiency with Equations 9.2 and 9.3, a relationship (Equation 9.4) for retention time, tt, in terms of diffusion coefficient, efficiency, particle size, and reduced variables (h and v) and retention factor results. Equation 9.4 illustrates that mobile phases with large diffusion coefficients are preferred if short retention times are desired. 

Taguchi et al. [97] and Liang et al. [98,99] reported on the preparation of monolithic carbon columns, which exhibit a hierarchical, fully interconnected porosity. Silica particles (10 pm) have been suspended in an aqueous solution, containing ethanol, FeClj, resorcinol, and formaldehyde. After polymerization, the solid rod was dried, cured, and carbonized by raising temperature to 800°C and finally up to 1250°C. Finally, concentrated FIF was used to remove silica and iron chloride. Even if carbon have been shown to possess a high specihc surface area (up to lllSmVg), their chromatographic efficiency is moderate (FIETP of 72 pm). 

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