Stationary phase analyte interaction with

Applications in this category are those that are generally described as chromatographic in nature. The analyte molecule is eluted through the MIP stationary phase and interacts with the binding sites in a series of discrete transitory interactions. The system is highly dependent upon the chemical environment and these mobile phase effects are discussed in depth below. 

The column is the key component of an LC system, because it is used to effect the separation and holds the stationary phase. The choice of stationary phase is of critical importance, because the stationary phase largely determines what molecular properties the separation will be based on. An appropriate stationary phase should interact with all the analytes but have a wide range of interaction strength. 

Gas-Liquid Chromatography. In gas-liquid chromatography (GLC) the stationary phase is a liquid. GLC capillary columns are coated internally with a liquid (WCOT columns) stationary phase. As discussed above, in GC the interaction of the sample molecules with the mobile phase is very weak. Therefore, the primary means of creating differential adsorption is through the choice of the particular liquid stationary phase to be used. The basic principle is that analytes selectively interact with stationary phases of similar chemical nature. For example, a mixture of nonpolar components of the same chemical type, such as hydrocarbons in most petroleum fractions, often separates well on a column with a nonpolar stationary phase, while samples with polar or polarizable compounds often resolve well on the more polar and/or polarizable stationary phases. Reference 7 is a metabolomics example of capillary GC-MS. 

Retention in RP chromatography is based on the interaction of the hydrophobic part of the analyte with the hydrophobic section of the stationary phase. This interaction can be modulated with the type and the concentration of the organic modiher in the mobile phase. The selectivity is mainly inflnenced by the interaction of the polar fnnctional gronps of the analyte with constituents of the mobile phase (bnffer, salts, etc. in the aqneons part) and with the amonnt and activity of residual surface silanols, which are, of course, also modihed by mobile phase constituents. 

Indicate for each of the chromatographic techniques below the term which best expresses the analyte interaction with the stationary phase. 

Apart from the above-discussed parameters for HPLC optimization of chiral resolution on antibiotic CSPs, some other HPLC conditions may be controlled to improve chiral resolution on these CSPs. The effect of the concentrations of antibiotics (on stationary phase) on enantioresolution varied depending on the type of racemates. The effect of the concentrations of teicoplanin has been studied on the retention (k), enantioselectivity (a), resolution (Rs), and theoretical plate number (N) for five racemates [21]. An increase in the concentration of teicoplanin resulted in an increase of a and Rs values. The most surprising fact is that the theoretical plate number (N) increases with the increase in the concentration of teicoplanin. It may be the result of the resistance of mass transfer resulting from analyte interaction with free silanol and/or the linkage chains (antibiotics linked with silica gel). This would tend to trap an analyte between the silica surface and the bulky chiral selector adhered to it. This is somewhat... 

IC is a liquid chromatography subclass in which analyte separation is based on ion-exchange mechanisms. Analytes interact with both the stationary phase (ion-exchange resin) and the mobile phase. The full details of liquid chromatography, especially IC, will be found in the literature.45-46... 

The most popular classification scheme stems from the manner in which the analyte interacts with the stationary phase. With this approach, chromatography may be divided into five separation mechanisms adsorption, partition, size exclusion, affinity, and ion exchange, as illustrated in Figure 1.1. 

Adsorption chromatography is based on competition for neutral analytes between the liquid mobile phase and a neutral, solid stationary phase. The analytes interact with the stationary phase according to the premise like likes like polar solutes will be retained longest by polar stationary phases, and nonpolar solutes will be retained best by nonpolar stationary phases. In adsorption chromatography the solute molecules are in contact with both the stationary phase and the mobile phase, simultaneously. Under these conditions, the solutes are said to be in an anisotropic environment. 

Specific and essentially stand-alone mode of liquid chromatography is associated with the absence or suppression of any analyte interactions with the stationary phase, which is called size-exclusion chromatography (SEC). In SEC the eluent is selected in such a manner that it will suppress any possible analyte interactions with the surface, and the separation of the analyte molecules in this mode is primarily based on their physical dimensions (size). The larger the analyte molecules, the lower the possibility for them to penetrate into the porous space of the column packing material, and consequently the faster they will move through the column. The schematic of this classification is shown in Figure 1-1. 

The fourth type of HPLC technique, size-exclusion HPLC (see Chapter 6), is based on the absence of any specific analyte interactions with the stationary phase (no force employed in this technique). 

Modern HPLC is a routine tool in any analytical laboratory. Standard HPLC system represents a separation output in the form of chromatogram (typical modern chromatogram is shown in Figure 1-6). Each specific analyte in the chromatogram is represented by a peak. In the absence of the strong specific analyte interactions with the stationary phase and at relatively low analyte concentration, peaks are symmetrical and resemble a typical Gaussian (normal distribution) curve. 

The equilibrium constant, K, thermodynamically could be described as the exponent of the Gibbs free energy of the analyte s competitive interactions with the stationary phase. In hquid chromatography the analyte competes with the eluent for the place on the stationary phase, and resulting energy responsible for the analyte retention is actually the difference between the analyte interaction with the stationary phase and the eluent interactions for the stationary phase as shown in equation (1-5)... 

Equation (1-7) shows that in an ideal case the selectivity of the system is only dependent on the difference in the analytes interaction with the stationary phase. It is important to note that the energetic term responsible for the eluent interactions was canceled out, and this means that the eluent type and the eluent composition in an ideal case does not have any influence on the separation selectivity. In a real situation, eluent type and composition may influence the analyte ionization, solvation, and other secondary equilibria effects that will have effect on the selectivity, but this is only secondary effect. 

In Section 2.1 the main chromatographic descriptors generally used in routine HPLC work were briefly discussed. Retention factor and selectivity are the parameters related to the analyte interaction with the stationary phase and reflect the thermodynamic properties of chromatographic system. Retention factor is calculated using expression (2-1) from the analyte retention time or retention volume and the total volume of the Uquid in the column. Retention... 

TYPES OF ANALYTE INTERACTIONS WITH THE STATIONARY PHASE... 

The LSER theory allows the association of the analyte retention behavior with the energetic components of the analyte interactions with the stationary phase. On the other hand, this theory is based on the assumption of the applicability of equation (2-43) to the analyte retention process, and this equation only allows the existence of one single partitioning retention mechanism. Coexistence of several different retention mechanisms, or the presence of secondary equilibria in a chromatographic system, effectively does not allow the applicability of this theory to the description of the analyte retention [59]. 

In everyday method development practice, it is important to ensure the separation of target compounds, matrix components, and other impurities. The elution of the analyte at the void volume means that it did not interact with the stationary phase and thus could not be separated from other components that do not interact with the surface either. To ensure the analyte interaction with the stationary phase, it is usually recommended to choose chromatographic conditions when any component of interest elutes with at least 1.5 void volume values or even greater. The error of the void volume determination for these purposes could be 20% or even greater (insofar as these void volume values are not used for any calculations but just to estimate where the least retained analye elutes). The use of uracil, thiourea, or allantoin as analytical void volume markers is most common in practical analytical work. 

In Chapters 2, 3, and 4, all aspects of the analyte retention on the HPLC column are discussed. There are many mathematical functions describing retention dependencies versus various parameters (organic composition, temperature, pH, etc.). Most of these dependencies rely on empirical coefficients. Analyte retention is a function of many factors analyte interactions with the stationary and mobile phases analyte structure and chemical properties struc-... 

Solvent competition model for normal-phase liquid chromatography. Like the solvent-interaction model, the solvent-competition model assumes that the stationary phase is covered with a monolayer of molecules of the strongest component of the mobile phase. This model also assumes that the concentration of analyte in the stationary phase is small compared with the concentration of solvent molecules and that solute-solvent interactions in the mobile phase are cancelled by identical interactions in the stationary phase. The competition between the analyte molecules, x, and the mobile phase molecules. A, for the active site or sites on the stationary phase is given by... 

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