Binary Mobile Phases

The first chiral separation using pSFC was published by Caude and co-workers in 1985 [3]. pSFC resembles HPLC. Selectivity in a chromatographic system stems from different interactions of the components of a mixture with the mobile phase and the stationary phase. Characteristics and choice of the stationary phase are described in the method development section. In pSFC, the composition of the mobile phase, especially for chiral separations, is almost always more important than its density for controlling retention and selectivity. Chiral separations are often carried out at T < T-using liquid-modified carbon dioxide. However, a high linear velocity and a low pressure drop typically associated with supercritical fluids are retained with near-critical liquids. Adjusting pressure and temperature can control the density of the subcritical/supercritical mobile phase. Binary or ternary mobile phases are commonly used. Modifiers, such as alcohols, and additives, such as adds and bases, extend the polarity range available to the practitioner. 

Figure 9 Solvent selectivity triangle approach forthe selectivity optimization in HP-RPC. First, three initial experiments (1-3) with three mobile phases (binary mixtures of ACN/water, MeOH/water, and THF/water, respectively) are performed.

Choosing a Mobile Phase Several indices have been developed to assist in selecting a mobile phase, the most useful of which is the polarity index. Table 12.3 provides values for the polarity index, P, of several commonly used mobile phases, in which larger values of P correspond to more polar solvents. Mobile phases of intermediate polarity can be fashioned by mixing together two or more of the mobile phases in Table 12.3. For example, a binary mobile phase made by combining solvents A and B has a polarity index, of... 

Solvent triangle for optimizing reverse-phase HPLC separations. Binary and ternary mixtures contain equal volumes of each of the aqueous mobile phases making up the vertices of the triangle. 

D. E. Martire, Unified Approach to the Theory of Chromatography Incompressible Binary Mobile Phase (Liquid Chromatography) in Theoretical Advancement in Chromatography and Related Separation Techniques (Ed. F. Dondi, G. Guiochon, IGuwer, Academic Publishers, Dordrecht, The Netherlands,(l993)261. 

Concentrations of moderator at or above that which causes the surface of a stationary phase to be completely covered can only govern the interactions that take place in the mobile phase. It follows that retention can be modified by using different mixtures of solvents as the mobile phase, or in GC by using mixed stationary phases. The theory behind solute retention by mixed stationary phases was first examined by Purnell and, at the time, his discoveries were met with considerable criticism and disbelief. Purnell et al. [5], Laub and Purnell [6] and Laub [7], examined the effect of mixed phases on solute retention and concluded that, for a wide range of binary mixtures, the corrected retention volume of a solute was linearly related to the volume fraction of either one of the two phases. This was quite an unexpected relationship, as at that time it was tentatively (although not rationally) assumed that the retention volume would be some form of the exponent of the stationary phase composition. It was also found that certain mixtures did not obey this rule and these will be discussed later. In terms of an expression for solute retention, the results of Purnell and his co-workers can be given as follows,... 

Practically a more convenient way of expressing solute retention in terms of solvent concentration for a binary solvent mixture as the mobile phase is to use the inverse of equation (11), i.e.. 

However, there might be exceptions if the mobile phase consists of a binary mixture of solvents, then a layer of the more polar solvent would be adsorbed on the surface of the silica gel and the mean composition of the solvent in the pores of the silica gel would differ from that of the mobile phase exterior to the pores. Nevertheless, it would still be reasonable to assume that... 

Bonded phases are the most useful types of stationary phase in LC and have a very broad range of application. Of the bonded phases, the reverse phase is by far the most widely used and has been applied successfully to an extensive range of solute types. The reverse phases are commonly used with mobile phases consisting of acetonitrile and water, methanol and water, mixtures of both acetonitrile and methanol and water, and finally under very special circumstances tetrahydrofuran may also be added. Nevertheless, the majority of separations can be accomplished using simple binary mixtures. 

From the general framework of the Snyder and Soczewinski model of the linear adsorption TLC, two very simple relationships were derived, which proved extremely useful for rapid prediction of solute retention in the thin-layer chromatographic systems employing binary mobile phases. One of them (known as the Soczewinski equation) proved successful in the case of the adsorption and the normal phase TLC modes. Another (known as the Snyder equation) proved similarly successful in the case of the reversed-phase TLC mode. 

The following Snyder equation is another simple linear relationship with respeet to cp, whieh links the retention parameter (i.e.. In k) of a given solute with the volume fraetion of the organic modifier in the aqueous binary mobile phase (cp) ... 

Then, the examples from Reference 23, that focus on retention of the selected binary mixtures of the test analytes (one comprising carboxylic acid and ketone and the other made of alcohol and ketone), chromatographed under the deliberately mild working conditions (microcrystalline cellulose was used as adsorbent and either decalin or n-octane as the monocomponent mobile phase) will be discussed. One of the test solutes in each binary mixture (either acid or alcohol) can be viewed as... 

FIGURE 2.18 Comparison of the concentration profiles of 2-phenylbutyric acid (dashed line) and benzophenone (thin solid line) developed as single analytes and as a binary mixture (bold solid line) concentration of 2-phenylbutyric acid in the sample was 1.25 mol 1 and that of benzophenone was 0.10 mol FI Microcrystalline cellulose was used as stationary phase and decalin as mobile phase [26]. 

Equation 4.13 and Equation 4.14 were tested for a series of mobile phases on alumina [29-31] and silica gel [32]. Two eluotropic series of solvent binary mixtures for alumina (a = 0.6) and silica gel (a = 0.7) have been calculated by using Equation 4.13, and the obtained data can be used to establish many such series or series of other selectivities [13,28],... 

Recently, Janjic et al. published some papers [33-36] on the influence of the stationary and mobile phase composition on the solvent strength parameter e° and SP, the system parameter (SP = log xjx, where and denote the mole fractions of the modiher in the stationary and the mobile phase, respectively) in normal phase and reversed-phase column chromatography. They established a linear dependence between SP and the Snyder s solvent strength parameters e° by performing experiments with binary solvent mixtures on silica and alumina layers. 

The separations of some nonionic tensides having biological activity and consisting of ethyleneoxide oligomer mixtures were performed in many different TEC systems (silica and alumina as the stationary phase and single solvent or binary mixtures as the mobile phase). Selectivity was higher on alumina than on the silica layer. Both... 

FIGURE 4.10 Mobile phase selection by microcircular technique, a. Sample of known composition A = nonpolar compound A1 = n-hexane A2 = acetone A3 = n-hexane-acetone, 60-1-40, v/v B = polar compound B1 = methanol B2 = water B3 = methanol-water, 70-1-30, v/v. b. Sample of unknown composition testing with solvents of different Snyder s groups and binary solvent mixture. 

For a binary mobile phase consisting of a mixture of solvents 1 and 2, the value is given by Equation 4.22 [64] ... 

Oscik and Chojnacka [63] use TEC adsorption in the investigation of six aromatic hydrocarbons (naphthalene, diphenyl, anthracene, pyrene, chrysene, and acenaphthene) on silica gel G by elution with different binary mobile phases (trichloroethylene-benzene, carbon tetrachloride-benzene, n-heptane-trichloroethylene. 

Prus and Kowalska [75] dealt with the optimization of separation quality in adsorption TLC with binary mobile phases of alcohol and hydrocarbons. They used the window diagrams to show the relationships between separation selectivity a and the mobile phase eomposition (volume fraction Xj of 2-propanol) that were caleulated on the basis of equations derived using Soezewiriski and Kowalska approaehes for three solute pairs. At the same time, they eompared the efficiency of the three different approaehes for the optimization of separation selectivity in reversed-phase TLC systems, using RP-2 stationary phase and methanol and water as the binary mobile phase. The window diagrams were performed presenting plots of a vs. volume fraetion Xj derived from the retention models of Snyder, Schoen-makers, and Kowalska [76]. 

The most common method for varying the chromatographic selectivity for neutral molecules in RPC is to change the type of organic modifier in the mobile phase. In numerous studies using binary mobile phases, equation (4.15) has been shown to describe reasonably well the variation of solute retention with the volume fraction of organic solvent in an aqueous-organic mobile phase... 

Normal-Phase BPC Vary composition of binary mobile phase linearly to give change in polarity equal to 0.5 P /t. 12-15% B/t,... 

Liquid-Solid Vary composition of binary mobile phase in a concave fashion to give a change in solvent strength equal to 0.02 e /t. (10-18)t, for a 0-100% gradient(a)... 

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