Column Performance In Liquid Chromatography Research Articles

Practical limits to column performance in liquid chromatography - Optimal operations.

Columns of different structures have different potential kinetic performance - the trade-off between separation, time, and pressure. However, the full potential of a structure cannot always be realized in practically existing columns. Each combination of column efficiency, time, and pressure requires certain cross-sectional dimensions of the column flow-through channels. However, there are limits to the narrowest flow-through channels that can be manufactured with current technology. As a result, columns of some structures cannot be optimized for providing the required efficiency in the shortest time. Additionally, the full potential of its structure can be realized only if a column can operate at the highest pressure available from liquid chromatography (LC) equipment, has sufficient loadability, and satisfies other practical requirements. Equations tailored for a systematic approach to evaluation of factors affecting performance of optimized LC columns (effects of column structure, column dimensions, operational conditions, etc.) were developed. Parameters quantifying the performance of a specific column at and below its largest acceptable pressure were identified. New objective performance parameters of columns and their structures were introduced. Among them are the apparent structural quality factor accounting for the effect of insufficiently high pressure acceptable for the column, the dimensionless plate duration - the parameter of a column structure affecting its performance when the pressure is not limited, - and others. Applying the theory developed herein to published data, the performance of several differently structured columns is evaluated, and the factors affecting their comparative performance are discussed. In the final count, not the quality of a column structure, but practical factors such as the narrowest manufacturable flow-through channels can dominate the choice of the kinetically most suitable column for a practical LC analysis.

Characterization of chromatographic peaks using the linearly modified Gaussian model. Comparison with the bi-Gaussian and the Foley and Dorsey approaches

To characterize column performance in liquid chromatography, several parameters must be obtained from experimental data. These parameters can be computed through the numerical integration of the net signal to calculate the moments after subtraction of the baseline. This requires the establishment of the peak integration limits. The whole process introduces significant uncertainty. For this reason, several alternative procedures have been proposed to measure the area, mean time and variance, based on the assumption that the chromatographic peak can be described with a mathematical function. This allows the calculation of the peak position and variance making use of the values of the experimental half-widths. In this work, the linear modified Gaussian model is used to derive several equations for the evaluation of the associated moments. Affordable equations for the calculation of the area, mean time, variance and efficiency are provided, using the half-width values at 10% peak height. The behaviour of experimental peaks obtained under a large variety of experimental conditions is examined to verify the validity of the proposed equations. The values of the peak parameters are compared with those calculated based on the bi-Gaussian model, and the exponentially modified Gaussian model using the equations developed by Foley and Dorsey. The bi-Gaussian model offered the best quantifications for the mean time. The Foley and Dorsey approach gave rather satisfactory results for the area and the best results for the variance and efficiency for tailing peaks of small asymmetry. The LMG approach gave better evaluation of the area for peaks showing small asymmetry, and satisfactory values for the mean time, variance and efficiency in the whole range of asymmetries found in liquid chromatography.

Prediction of Peak Shape and Characterization of Column Performance in Liquid Chromatography as a Function of Flow Rate

Traditionally, column performance in liquid chromatography has been studied using information from the elution of probe compounds at different flow rates through van Deemter plots, which relate the column plate height to the linear mobile phase velocity. A more recent approach to characterize columns is the representation of the peak widths (or the right and left peak half-widths) for a set of compounds versus their retention times, which, for isocratic elution, give rise to almost linear plots. In previous work, these plots have been shown to facilitate the prediction of peak profiles (width and asymmetry) with optimization purposes. In this work, a detailed study on the dependence of the peak widths (or half-widths) on the flow rate is reported. A new approach to quantify the deterioration of column performance for slow and fast flow rates and to characterize chromatographic columns is proposed. The approach makes use of the width (or half-widths) for a set of compounds with similar interaction kinetics and does not require knowledge of the extra-column contributions to the total variance. The chromatographic data of two sets of compounds of different natures (sulfonamides and β-blockers), eluted from Spherisorb and Chromolith columns with acetonitrile-water mixtures, are used to illustrate the approach.

Graphical data representation methods to assess the quality of LC columns.

We discuss the most important plot types for the kinetic performance of liquid chromatography columns and elaborate on how these plots should best be constructed and can be made dimensionless. Distinction is made between plots that are most suited for practitioners (column users) versus those most suited for theoreticians and column manufacturers.

Relationship between trans-column eddy diffusion and retention in liquid chromatography: Theory and experimental evidence

Experimental results demonstrate that trans-column eddy diffusion depends on the retention of compounds. The combination of elution profiles recorded in different points of the exit column cross-section and of the height equivalent to a theoretical plate (HETP) of small molecules clearly show a strong link between retention and column performance in liquid chromatography. These results validate a new model of trans-column eddy diffusion in packed columns. The contribution to the column HETP of the trans-column eddy diffusion term decreases with increasing retention factor from k′ = 0 to k′ = 3 above which it becomes negligible. The best column performance in RPLC is observed for the most retained compounds. This is due to the combination of the lack of a residual trans-column eddy diffusion contribution and the vanishing contribution of the instrument to band broadening.

On the reproducibility of column performance in liquid chromatography and the role of the packing density

The packing behavior of a typical 10 μm C 18 stationary phase was studied in terms of the resultant column efficiency and capacity factor. The column-to-column reproducibility of these parameters under identical packing procedures is assessed. Correlation of these parameters and the column void volume to the column packing density is reported. Two regimes were studied: that of poor-and well-packed columns. For poorly packed columns, the column-to-column variability is high, but a concomitantly poor same-column reproducibility of measurement suggests that little statistically significant difference exists between different columns packed with the same procedure. There is also no statistically significant correlation between the column parameters and the packing densities, however, the poorest columns showed a degradation of performance after the drying procedures used to obtain the column masses. Well-packed columns showed much less degradation upon drying. For the well-packed columns, statistically significant column-to-column differences were observable, mainly due to a high same-column precision of measurement. Analysis of the results suggests that even well-packed columns are not optimally packed and that regions of high and low density coexist along the column. The results are compared to those achieved with semi-preparative columns packed with the same slurry procedure and preparative columns packed under dynamic axial compression. Poor day-to-day, same-column reproducibility (degradation) under ambient conditions was observed in conjunction with a high column-to-column variability for the semi-preparative columns.

Evaluation of Column Performance in Liquid Chromatography

Afin de classer et selectionner une colonne chromatographique et la phase a employer, les proprietes chromatographiques sont etudiees: l'activite du silanol, l'hydrophobicite, l'activite du metal, celui-ci est incorpore sous forme de traces lors de la preparation de la silice. Le type de phase et la dimension des pores sont egalement analyses. Utilisation d'une colonne avec une phase stationnaire C18