Column retention behavior

This column exhibits a retention behavior for PMMA in toluene that is very similar to column No. 4, it is as if they contained identical column packings... 

It is interesting that the retention behavior of lycopene varies dramatically depending on the stationary phase synthesis lycopene usually elutes before a- and P-carotene in monomeric columns, whereas with polymeric C30 columns, lycopene elutes after these carotenes. ... 

Reversed-phase liquid chromatography shape-recognition processes are distinctly limited to describe the enhanced separation of geometric isomers or structurally related compounds that result primarily from the differences between molecular shapes rather than from additional interactions within the stationary-phase and/or silica support. For example, residual silanol activity of the base silica on nonend-capped polymeric Cis phases was found to enhance the separation of the polar carotenoids lutein and zeaxanthin [29]. In contrast, the separations of both the nonpolar carotenoid probes (a- and P-carotene and lycopene) and the SRM 869 column test mixture on endcapped and nonendcapped polymeric Cig phases exhibited no appreciable difference in retention. The nonpolar probes are subject to shape-selective interactions with the alkyl component of the stationary-phase (irrespective of endcapping), whereas the polar carotenoids containing hydroxyl moieties are subject to an additional level of retentive interactions via H-bonding with the surface silanols. Therefore, a direct comparison between the retention behavior of nonpolar and polar carotenoid solutes of similar shape and size that vary by the addition of polar substituents (e.g., dl-trans P-carotene vs. dll-trans P-cryptoxanthin) may not always be appropriate in the context of shape selectivity. 

Fornal et al. [75] determined selectivity differences for bases in RP-HPLC under high pH conditions. They used quantitative structure retention relationships (QSRR) to model retention behavior. They reported that the stability of the columns they used (Waters XTerra MS, Zorbax Extend, Thermo BetaBasic) was limited with... 

Secondly, the pH can have an effect on the selectivity. In pharmaceutical analysis, mainly compounds with either basic or acidic properties have to be analyzed. Since the pH influences the dissociation and charge of these substances, it will affect their retention behavior. Changing the organic modifier does not lead to major selectivity differences, but it can enhance the differences induced by pH and/or stationary phase changes. Also, the column... 

The column should permit the modulation of retention behavior over a very wide range of conditions. This requirement in fact means that the stationary phase is inert, that is it does not facilitate specific interactions with certain molecular functions of solute molecules with the concomitant advantage of a relatively clean and rapid adsorption-desorption kinetics. Preferably then the stationary phase has no functional groups such as fixed charges that would have strong affinity to counterionic solutes and exclude solutes of co-ionic nature. In this regard the properties of well-prepared hydrocarbonaceous bonded phases indeed approach those that we would expect from an ideal phase. 

The packing material first described for direct injection of biological samples was prepared by simply saturating the accessible adsorption sites of a Cis reversed-phase silica with human plasma proteins (105). After saturation, the human plasma proteins were denatured at the external surface, and their native conformation was destroyed. With this treatment, the proteins formed a hydrophilic layer with weak ion-exchange properties, which provided protection from contact with the sample proteins, whereas the alkyl ligands inside the pores remained unchanged and thus served for analyte retention. The retention behavior of the saturated phase did not alter with this treatment, but the efficiency was reduced dramatically. Such protein-coated columns have shown a lifetime of several months (106). 

J. E. Madden, M. J. Shaw, G. W. Dicinoski, and P. R. Haddad, Simulation and Optimization of Retention in Ion Chromatography Using Virtual Column 2 Software, Anal. Chem. 2002, 74, 6023 P. R. Haddad, M. J. Shaw, J. E. Madden, and G. W. Dicinoski, Computer-Based Undergraduate Exercise Using Internet-Accessible Simulation Software for the Study of Retention Behavior and Optimization of Separation Conditions in Ion Chromatography, J. Chem. Ed. 2004, 81, 1293 http //www.virtualcolumn.com. 

Table 3 (73) compares the retention coefficients for synthetic peptides from various sources. To ensure comparability, the data has been standardized with respect to lysine and assigned a value of 100. The table shows that there are discrepancies between the results obtained using different chromatographic systems. Predictions of retention times should therefore be made using chromatographic systems similar to those used to calculate the retention coefficients for the amino acids. Casal et al. (75a) have made a comparative study of the prediction of the retention behavior of small peptides in several columns by using partial least squares and multiple linear regression analysis. 

Retention characteristics and elution order of carotenoid cis-trans isomers with C30-bonded phases are strikingly similar to those obtained with normal-phase systems using calcium hydroxide columns (190). Different carotenoids exhibit varying retention behavior in response to temperature changes for C30 and C34 polymeric stationary phases as compared with a Cl8 polymeric phase (179). These behaviors are believed to be related to conformational changes in the longer stationary phases with temperature. The slot model proposed for the retention of planar... 

Different kinds of HPLC exist. Many kinds of column packings and solvents are available. Retention behavior and resolution are affected by column characteristics (C-loading, chain length, porosity, etc.) and by elution scheme characteristics (mobile phase, pH, organic modifier, etc.). The samples can be separated on the basis of solubility and polarity of the sample components. 

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