Chromatographic columns, performance evaluation

S. Maeno, P. A. Roderiguez, Simple and versatile injection system for capillary gas chromatographic columns. Performance evaluation of a system including mass spectrometric and light-pipe Fourier-Transform infra-red detection. J. Chromatogr. 731(1-2) (1996) 201-215. 

The silanophilic character of 16 reversed-phase high-performance liquid chromatographic columns was evaluated with dimethyl diphenycyclam, a cyclic tetraza macrocycle [101]. The method is rapid, does not require the removal of packing material, and uses a water-miscible solvent. The results demonstrate two points first, cyclic tetraza macrocycles offer substantial benefits over currently used silanophilic agents second, the method can easily differentiate the performance of various columns in terms of their relative hydrophobic and silanophilic contributions to absolute retention. 

Several high performance liquid chromatographic columns were evaluated according to their capabilities to separate and release the MIC-Fluram fluorophor. Reverse phase, normal partition, and amine bonded columns were tested. The reverse phase columns performed the separation and analysis more proficiently than the other columns. A Variarv CH-10 column was the best column from among the several ones that were tested. 

To realistically evaluate the effect of extra-column dispersion on column performance, it is necessary to evaluate the maximum extra-column dispersion that can be tolerated by different types of columns. Such data will indicate the level to which dispersion in the detector and its associated conduits must be constrained to avoid abrogating the chromatographic resolution. 

Retrospective validation uses historical information gathered in actual process runs to evaluate the process. For example, batch records can provide extensive data on column performance and analytical data of fractions and final product can provide valuable information on the efficiency of the chromatographic steps in removing contaminants. Chapman67 cautions that while retrospective validation is a valid and valuable approach, it is not meant to be retroactive — validation must be done before product is released to market. 

When an assay method is performed repeatedly to analyze a high volume of samples, the instability of the calibration curves or an apparent change in response factor often indicates that some conditions of the assay are drifting, are no longer stable, and need to be evaluated. Reasons for the instability of the calibration curves can include variation of the extraction procedures, deterioration of the efficiency of a chromatographic column, or decline of the efficiency of the detection system (28). 

Some people feel that columns cost too much and, therefore, purchase the lowest-price column. This is one strategy for choosing the best column. However, when the nature of a column in a chromatographic system is considered, it makes sense that there should be little to no price sensitivity in the user. For instance, if an instrument sells for 20,000, a typical column sells for 300. Therefore, a column is only 300/ 20,000, or 1.5% of the cost of an instrument installation. This is a minor part even if repeat buying is calculated. Second, keep in mind that without the column performance the entire instrument is worth precisely zero. Third, it is very hard to evaluate how a column will perform without prior experience with it—there is no easy believability to competitive claims that one C 8 column will work as well as, or even the same as, another. In essence, a column is nothing more and nothing less than a user s attempt to solve an analytical problem. Users should not be sensitive to price, they should be concerned with time, effort, and repeatability. Therefore, most individuals should gladly pay for these benefits, because the best column does your separation. 

For analyses on a SP-2331 (or equivalent) GC column the chromatographic resolution is evaluated before the analysis of any calibration standards by the analysis of a commercially available column performance mixture (see Section 5.19) that contains the TCDD isomers that elute most closely with... 

The continuing calibration consists of two parts evaluation of the chromatographic resolution and verification of the RRF values to be used for quantitation. At the beginning of each 12-hour period, the chromatographic resolution is verified in the same fashion as in the initial calibration through the analysis of the CC3 solution on the DB-5 (or equivalent) column or through the analysis of the column performance solution on the SP-2331 (or equivalent) column. 

Evaluation of Chromatographic Performance For information on the evaluation of the performance of chromatographic columns, see under Gas Chromatography, p. 189. 

At present two models are available for description of pore-transport of multicomponent gas mixtures the Mean Transport-Pore Model (MTPM)[4,5] and the Dusty Gas Model (DGM)[6,7]. Both models permit combination of multicomponent transport steps with other rate processes, which proceed simultaneously (catalytic reaction, gas-solid reaction, adsorption, etc). These models are based on the modified Maxwell-Stefan constitutive equation for multicomponent diffusion in pores. One of the experimentally performed transport processes, which can be used for evaluation of transport parameters, is diffusion of simple gases through porous particles packed in a chromatographic column. 

The solvent delivery system is responsible for delivering the pressurized mobile phase with the desired composition and chosen flow rate to the head of the column. To achieve this goal a number of components work together under the supervision of the system computer or microprocessors to achieve the tight specifications typical of a modem liquid chromatograph. Table 5.2 [15-19]. Suitable tests for performance evaluation of solvent delivery systems are briefly described at the end of section 5.2.2. In retention terms a relative standard deviation of better than 0.15% for retention factors under normal operating conditions is expected. 

Having established that a finite volume of sample causes peak dispersion and that it is highly desirable to limit that dispersion to a level that does not impair the performance of the column, the maximum sample volume that can be tolerated can be evaluated by employing the principle of the summation of variances. Let a volume (Vi) be injected onto a column. This sample volume (Vi) will be dispersed on the front of the column in the form of a rectangular distribution. The eluted peak will have an overall variance that consists of that produced by the column and other parts of the mobile phase conduit system plus that due to the dispersion from the finite sample volume. For convenience, the dispersion contributed by parts of the mobile phase system, other than the column (except for that from the finite sample volume), will be considered negligible. In most well-designed chromatographic systems, this will be true, particularly for well-packed GC and LC columns. However, for open tubular columns in GC, and possibly microbore columns in LC, where peak volumes can be extremely small, this may not necessarily be true, and other extra-column dispersion sources may need to be taken into account. It is now possible to apply the principle of the summation of variances to the effect of sample volume. 

High Performance Liquid Chromatography. All separations were performed using an Altex Scientific (1780 Fourth Street, Berkeley, CA 94710) Model 320 Advance Research Chromatograph, consisting of a model lOOA dual piston analytical pump, a Model 153 UV detector, a Model 210 injection valve, and a Model 155 recorder. The columns (600 x7.5mm) evaluated were the Spherogel TSK-SW-2000 and SW-3000 (Altex). Unless otherwise stated, all separations were carried out at 23 - 25 . 

Ascah, T., Kallury, K., Szafranski, C. Corman, S., and Liu, F., Characterization and high performance liquid chromatographic evaluation of a new amide-functionalized reversed-phase column, /. Liq. Chromatogr. Relat. Technol, 19 (17,18), 3049, 1996. 

Viehauer S. et al., 1995. Evaluation and routine application of the novel restricted-access precolumn packing material Alkyl-Diol Silica Coupled-column high-performance liquid chromatographic analysis of the photoreactive drug 8-methoxypsoralen in plasma. J Chromatogr B 666 315. 

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