Column performance criteria

The measurement of column performance criteria to compare columns, column packings, and column loading procedures has long suffered from the non-standardization of test procedures and disagreement as to which of the many performance criteria are most significant. It is the purpose of this paper to point out several practical aspects which must be considered when designing experiments to test column performance. 

These column performance criteria should be specified by the manufacturer and users should measure them whenever a new column is received. These column specification measurements should be kept for future reference and re-checked whenever it is suspected that the column has deteriorated. 

However, it should be pointed out that any given column, operated at a specific flow rate, can exhibit a range of efficiencies depending on the nature of the solute that is chosen for efficiency measurement. Consequently, under exceptional circumstances, the predicted conditions for the separation of the critical pair may not be suitable for another pair and the complete resolution of all solutes may not be obtained. This could occur if the separation ratio of another solute pair, although larger, was very close to that of the critical pair but contains solutes, for example, of widely different molecular weight. However, the possibility of this situation arising, in practice, is extremely remote and will not be considered in this discussion. It follows, that the efficiency required to separate the critical pair, numerically defined, is the first performance criterion... 

The most commonly used criterion for judging column performance is efficiency as measured by the number of theoretical plates or column plate count (N) exhibited by the column during the separation of a test mixture. The larger the number of theoretical plates, the more likely it is that the column will produce the desired separations. However, while popular, N is not a complete performance parameter for making comparisons. For example, N does not take into account particle size as does the reduced plate height, h. Another measurement, hmin, accounts for all of these factors as well as the mobile phase linear velocity and sample diffusion. However, N is the term most commonly recognized as being related to resolution (2), as shown in Equation 1 ... 

However, using peak skew as a performance criterion does suffer from one drawback it is necessary to have a sophisticated computer to make the calculation. Therefore, most users of HPLC columns will have to use hand-calculated values for peak asymmetry measurement (Figure A). However, these hand-calculated values can be correlated to a first approximation to the computer-generated values of peak skew using the graph shown in Figure 5. 

Precision in retention times and peak area or height is a major criterion of a separation system. Retention-time precision is important because Ri is the primary means for peak identification. It is also an important performance criterion and diagnostic for an LC pump and a column. 

Resolution and analysis time being interrelated,. effective plates per second, is sometimes a better criterion for comparing column performance. 

Another criterion for comparing column performance is the column permeability, K°, which is a measure of the facility with which the mobile phase... 

The performance index is a measure of the quality of the separation. Knox (19) was searching for a criterion that is a purer measure of column performance and criticized the performance index for its dependence on the retention factor and the viscosity. Therefore he defined a dimensionless group, the separation impedance ... 

Attempts have been made to combine the different parameters of a separation into a single criterion that describes the overall quality of a column or a separation. We would like to combine a measure of the narrowness of the buds, the analysis time, and the pressure needed to drive the separation. Since we want to maximize efficiency while minimizing analysis time and pressure, a possible measure would be the simple ratio of plate count to the product of analysis time and pressure. However, since we end up with two parameters that we want to minimize in the denominator and only one parameter that we want to maximize in the numerator, we end up with a measure that minimizes plate count To compensate for this, we n to raise the plate count in the numnator to the second power. This measure of overall column performance is the performance index, first developed for gas chromatography by Golay (18) ... 

To reiterate the definition of chromatographic resolution a separation is achieved in a chromatographic system by moving the peaks apart and by constraining the peak dispersion so that the individual peaks can be eluted discretely. Thus, even if the column succeeds in meeting this criterion, the separation can still be destroyed if the peaks are dispersed in parts of the apparatus other than the column. It follows that extra-column dispersion must be controlled and minimized to ensure that the full performance of the column is realized. 

Another aspect of cost reduction would be solvent economy. The need to preferentially select inexpensive solvents and employ the minimum amount of solvent per analysis would be the third performance criteria. Finally, to conserve sample and to have the capability of determining trace contaminants, the fourth criterion would be that the combination of column and detector should provide the maximum possible mass sensitivity and, thus, the minimum amount of sample. The performance criteria are summarized in Table 1. Certain operating limits are inherent in any analytical instrument and these limits will vary with the purpose for which the instrument was designed. For example, the preparative chromatograph will have very different operating characteristics from those of the analytical chromatograph. 

For autosampler precision, 10 consecutive lO-pL injections of an eth-ylparaben solution (20 J,g/mL) are used (Figure 6). A Waters Symmetry column packed with 5- J,m particles is used. The manufacturer s specification for peak area precision at 0.5% RSD is adopted as the acceptance criterion. This stringent precision criterion is required for precise assay testing of drug substances typically specified at 98-102% purity. The linearity test is performed by single injections of 5, 10, 40, and 80 pL of the... 

Given a single multicomponent feed stream of known conditions (i.e. flowrate, composition, temperature and pressure) to be separated into a number of multicomponent products of specified compositions, determine an optimal distillation configuration that performs the desired separation task by allowing the use of nonsharp columns and which satisfies the criterion of minimum total annual cost. 

The excellent reproducibility of the retention times of the late eluting sample components suggest that the performance of the system in rapid analysis is acceptable. Yet, the overall results are believed to be less reproducible than those could be obtained with such columns with an appropriately designed equipment and automated sample injector, which had yielded higher reproducibility of peak height, an essential criterion in quantitative analysis. 

The goal of optimization is safety at maximum profit, but this can only be done if the market value of each product is known. This is not the case when the products of a column are not final products but feed flows to other unit processes. When the product prices are unknown, it is still possible to perform optimization, but the optimization goal changes. The criterion in that case becomes the generation of the required products at minimum operating costs. This can be called an optimum with respect to the column involved, but only a "suboptimum" with respect to the plant of which the column is a part. 

Weyland et al. [560,561] used this method to optimize ternary mobile phase compositions for the separation of sulfonamides by RPLC. They fitted the retention surfaces to a quadratic model similar to eqn.(3.39), and also used a combination of a threshold resolution and minimum analysis time (min tm fl / vmin> 1.25 eqn.4.24) [560]. This criterion may yield a good optimum if the optimization is performed on the final analytical column (see table 4.11). 

III,D,5). In this chapter, the equation is further examined in relation to bed performance, since the turbulence properties of the bed result from interaction between bubbles and the continuous phase. As shown in Fig. 34, the mean slip velocity of bubbles in a fluidized catalyst bed of good fluidity is essentially the same as that for a bubble column when Uq > 20 cm/sec. A criterion under which bubble size approaches a dynamic equilibrium is obviously needed for predicting or evaluating the performance of scaled-up beds. 

High performance liquid chromatography has been used as a criterion of specificity when coupled with a bioassay or radioimmunoassay. With regard to the opioid peptides, brain extracts were chromatographed on an HPLC column that had previously been calibrated with known opioid peptides (Lewis et ai, 1979), and the collected fractions were assayed for activity (Stern et ai, 1980) (Fig. 12). The bioassay provides the quantitation and the HPLC provides the identity of each component. It would be interesting to use HPLC coupled with radioimmunoassay to confirm the identity of the many peptides that have been found in brain by immunohistochemical techniques. 

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