Chromatographic systems stationary phase

In a chromatographic separation procedure the parameters of the chromatographic system (stationary phase, flow, temperature, etc.) have to be selected respectively optimized with respect to some criterion (resolution, time, etc.). In gas chromatography retention data series are published and used for the sttidy of solvent/solute interaction, prediction of the retention behaviour, activity coefficients, and other relevant information usable for optimization and classification. Several clKmometrk techniques of data anal s have been employed, e.g. PCA, numerical taxonomic methods, information theory, and j ttern recognition. 

The calculation methods discussed above classify the chromatographic systems (stationary and mobile phases) or solute molecules while simultaneously taking into consideration the retention strength and retention selectivity thus, it cannot be applied when the separation of the strength and selectivity of the effect is required. SPM, another multivariate mathematical-statistical method, overcomes this difficulty. 

Fig. 11.8.1. Reversed phase separation of vitamin A alcohol and esters. Chromatographic conditions stationary phase, Zorbax C-8 (250 x 4.6 mm I.D.) mobile phase, 99.8% water-0.2% perchloric acid (60%), and 99.8% methanol-0.2% perchloric acid flow rate, 1.6 ml/min detection, UV at 254 nm. Peaks 1, vitamin A alcohol 2, impurity 3, vitamin A acetate 4, vitamin A palmitate. Reproduced from Dupont Analytical Systems with permission.

Chromatographic separations are accomplished by continuously passing one sample-free phase, called a mobile phase, over a second sample-free phase that remains fixed, or stationary. The sample is injected, or placed, into the mobile phase. As it moves with the mobile phase, the sample s components partition themselves between the mobile and stationary phases. Those components whose distribution ratio favors the stationary phase require a longer time to pass through the system. Given sufficient time, and sufficient stationary and mobile phase, solutes with similar distribution ratios can be separated. 

SPME has been utilized for deterrnination of pollutants in aqueous solution by the adsorption of analyte onto stationary-phase coated fused-siUca fibers, followed by thermal desorption in the injection system of a capillary gas chromatograph (34). EuU automation can be achieved using an autosampler. Eiber coated with 7- and 100-p.m film thickness and a nitrogen—phosphoms flame thermionic detector were used to evaluate the adsorption and desorption of four j -triazines. The gc peaks resulting from desorption of fibers were shown to be comparable to those obtained using manual injection. 

In the analytical chromatographic process, mixtures are separated either as individual components or as classes of similar materials. The mixture to be separated is first placed in solution, then transferred to the mobile phase to move through the chromatographic system. In some cases, irreversible interaction with the column leaves material permanently attached to the stationary phase. This process has two effects because the material is permanently attached to the stationary phase, it is never detected as leaving the column and the analysis of the mixture is incomplete additionally, the adsorption of material on the stationary phase alters the abiHty of that phase to be used in future experiments. Thus it is extremely important to determine the ultimate fate of known materials when used in a chromatographic system and to develop a feeling for the kinds of materials in an unknown mixture before use of a chromatograph. 

SiHca, alumina, and other metal oxides and salts have been used as the stationary phase in gas—soHd chromatographic systems. The appHcabiHty of these materials is limited by the difficulty of producing a consistent, resiHent, reproducible material. 

In the course of mixture separation, the composition and properties of both mobile phase (MP) and stationary phase (SP) are purposefully altered by means of introduction of some active components into the MP, which are absorbed by it and then sorbed by the SP (e.g. on a silica gel layer). This procedure enables a new principle of control over chromatographic process to be implemented, which enhances the selectivity of separation. As a possible way of controlling the chromatographic system s properties in TLC, the pH of the mobile phase and sorbent surface may be changed by means of partial air replacement by ammonia (a basic gaseous component) or carbon dioxide (an acidic one). 

This type of chromatographic development will only be briefly described as it is rarely used and probably is of academic interest only. This method of development can only be effectively employed in a column distribution system. The sample is fed continuously onto the column, usually as a dilute solution in the mobile phase. This is in contrast to displacement development and elution development, where discrete samples are placed on the system and the separation is subsequently processed. Frontal analysis only separates part of the first compound in a relatively pure state, each subsequent component being mixed with those previously eluted. Consider a three component mixture, containing solutes (A), (B) and (C) as a dilute solution in the mobile phase that is fed continuously onto a column. The first component to elute, (A), will be that solute held least strongly in the stationary phase. Then the... 

In a chromatographic separation, the individual components of a mixture are moved apart in the column due to their different affinities for the stationary phase and, as their dispersion is contained by appropriate system design, the individual solutes can be eluted discretely and resolution is achieved. Chromatography theory has been developed over the last half century, but the two critical theories, the Plate Theory and the Rate Theory, were both well established by 1960. There have been many contributors to chromatography theory over the intervening years but, with the... 

It is clear that the separation ratio is simply the ratio of the distribution coefficients of the two solutes, which only depend on the operating temperature and the nature of the two phases. More importantly, they are independent of the mobile phase flow rate and the phase ratio of the column. This means, for example, that the same separation ratios will be obtained for two solutes chromatographed on either a packed column or a capillary column, providing the temperature is the same and the same phase system is employed. This does, however, assume that there are no exclusion effects from the support or stationary phase. If the support or stationary phase is porous, as, for example, silica gel or silica gel based materials, and a pair of solutes differ in size, then the stationary phase available to one solute may not be available to the other. In which case, unless both stationary phases have exactly the same pore distribution, if separated on another column, the separation ratios may not be the same, even if the same phase system and temperature are employed. This will become more evident when the measurement of dead volume is discussed and the importance of pore distribution is considered. 

Scott and Kucera [4] carried out some experiments that were designed to confirm that the two types of solute/stationary phase interaction, sorption and displacement, did, in fact, occur in chromatographic systems. They dispersed about 10 g of silica gel in a solvent mixture made up of 0.35 %w/v of ethyl acetate in n-heptane. It is seen from the adsorption isotherms shown in Figure 8 that at an ethyl acetate concentration of 0.35%w/v more than 95% of the first layer of ethyl acetate has been formed on the silica gel. In addition, at this solvent composition, very little of the second layer was formed. Consequently, this concentration was chosen to ensure that if significant amounts of ethyl acetate were displaced by the solute, it would be derived from the first layer on the silica and not the less strongly held second layer. 

---