Retention factor columns

Mobile phases with some solvating potential, such as CO2 or ammonia, are necessary in SGC. Even though this technique is performed with ambient outlet pressure, solutes can be separated at lower temperatures than in GC because the average pressure on the column is high enough that solvation occurs. Obviously, solute retention is not constant in the column, and the local values of retention factors increase for all solutes as they near the column outlet. 

According to Equation 3, the resolution of two peaks in column separation is controlled by three major variables retention defined in terms of the retention factor k column efficiency expressed as the number of theoretical plates N and selectivity characterized by the selectivity factor a [48] ... 

In the simplest scheme of 2D HPLC, effluent of the first dimension (lst-D) was directly loaded into an injector loop (500 pL) of the 2nd-D HPLC for 28 s, and 2 s were allowed for injection. This operation was accompanied by the loss of lst-D effluent for 2 s out of 30 s in each cycle. The flow rate of 10 mL/min allowed the elution of solutes having retention factors (k values) up to 8 for the 2nd-D within the 30-s separation window, with f0 of 3.5 s. Figure 7.7 a and b shows the chromatograms for the 1 st-D and the 2nd-D, respectively, obtained for a mixture of hydrocarbons and benzene derivatives. The lst-D chromatogram showed many overlapping peaks. PAHs were eluted as mixtures from the FR column, and some are separated in the 2nd-D. 

Where a, b, and c = van Deemter coefficients, dp = particle size of column, L = column length, Dm = diffusion coefficients of analytes, t = column dead time (depends on flow rate F), tg= gradient time (determines analysis time via tA = tg + t0), Ac = difference in concentrations of the organic modifier at the end and the beginning of the gradient (a continuous linear gradient is assumed), and B = slope of the linear relationship between the logarithm of the retention factor and the solvent composition. 

Ten columns of the 24 available in a cartridge were employed to analyze all compounds in duplicate. Uracil, was employed as a dead volume marker (tO) needed for the evaluation of retention factor [k = (tr - t0)/t0]. Two additional columns were used for simultaneous analysis of the unknown. Values for the log of the capacity factor k were calculated for every compound at each percent organic content of the mobile phase log k = log [(tr - t0)/t0. For each compound, a plot of log k versus percent acetonitrile was used to calculate log k w (log k at 0% acetonitrile). 

In addition to the above strategies, the use of higher column temperatures is another approach that may decrease analysis time and improve sample throughput. The relationship between the chromatographic retention factor, k, and separation temperature is shown in Equation 13.1 ... 

The adjusted retention time provides a measure of the strength of intermolecular interaction between the analyte and the stationary phase, with stronger interactions giving a longer time. The gas hold-up time is derived from the flow rate and the column dimensions and is often measured by injecting a non-retained compound. The retention factor, which represents a ratio of the mass of analyte dissolved in the stationary phase to the mass in the mobile phase, can be calculated from the adjusted retention time and the gas hold-up time. 

Using the retention data and the chromatogram shown in Fig. 14.8, tabulate the following for each peak retention time ( r), adjusted retention time (t K), retention factor (k), partition coefficient (Kc) and number of theoretical plates (N). The column phase ratio was 250 and the gas hold up time ( m) was 0.995 min. 

Table 4. Retention factors k and column efficiencies N for an unretained thiourea and retained compound amylbenzene in columns packed by different methods [53] ...

The resolution can be improved by increasing the column plate number, N, and/ or the separation factor, a (a = the ratio of the retention factors of the two compounds). N is the physical parameter and a is the chemical parameter for the separation. Higher N and a values give a better separation. 

Common standard compounds for reversed phase columns are toluene and naphthalene, which have retention factors, k, of about 3. The eluent modifier is methanol or acetonitrile at a concentration of 50-80%, depending on the hydrophobicity of the stationary phase material. For other stationary phase materials, corresponding analytes, with k = 3-5, can be used. 

The retention factor, k, is the basic value in chromatography, and is related to the void volume (dead volume). The void volume is the space inside the column, where no retention of solutes has occurred and can be measured on a chromatogram, as shown in Figure 1.3. The void volume is about half the total volume of the column when it is packed with porous stationary phase materials. In practice, the effective void experienced by the analyte is smaller because the molecular mass of the analyte is usually much greater than that of the eluent molecule. In a model of porous stationary phase material, the pores can be represented as V-shape valleys (Figure 3.8), where region a is a support, such as... 

The general relationship between the type of solute and its retention can be seen by comparing the retention factors, k, of a set of standard compounds with their octanol-water partition coefficients, i.e. the logP value (listed in Table 4.1), as a measure of their relative solubility in water. The logarithm of the retention factor, log k, of these compounds measured in 50% aqueous acetonitrile on an octadecyl-bonded silica gel column shows a close linear relationship (Figure 4.1). 

Figure 4.1 Retention factors related to log P values. Column, 5/im octadecyl-bonded silica gel (LiChrosorb LC7) 25 cm x 4.1 mm i.d. eluent, 50% aqueous acetonitrile flow rate, 1ml min-1. Compounds , alkanols O, benzoates, O, polycyclic aromatic hydrocarbons, and A, alkylbenzenes.

The maximum retention factor (kQ) is related to the log P value and k and k are the retention factors of the cationic and anionic forms, respectively. The pKa values are known, and the retention factor in a given eluent can therefore be predicted in reversed-phase liquid chromatography using an alkyl-bonded silica gel or polystyrene gel column. The separation conditions can be adjusted according to their logP and pKa values by the selection of a suitable organic modifier concentration and the pH of the eluent.3,4... 

Increasing the column temperature reduces the retention factor. The ion-pair formation is based on a chemical equilibrium therefore, temperature control is important to obtain reproducible results. 

Figure 6.2 Comparison of measured and predicted retention factors of aromatic acids from Rekker s log P values. Column, 10 fim polystyrene gel, Hitachi 3011, 15 cm x 4.6 mm i.d. eluent, 30% aqueous acetonitrile containing 50 mM phosphoric acid at 55 °C. Numbers beside symbols see Table 6.4.

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