Reversed-phase chromatography solvophobicity

The mechanism of reversed-phase chromatography arises from the tendency of water molecules in the aqueous-organic mobile phase to self-associate by hydrogen bonding. This ordering is perturbed by the presence of nonpolar solute molecules. As a result, solute molecules tend to be excluded from the mobile phase and are bound by the hydrophobic stationary phase. This solvophobic... 

Hydrophobic Effects and Solvophobic Considerations for the Isolation of Peptides by Reversed-Phase Chromatography Methods... 

Offline precolumn derivatization is the most common alternative in this respect it involves separating the esters obtained from the organic acids by reversed-phase chromatography, which amply surpasses solvophobic chromatography (i.e., the use of undissociated acids as such) and allows gradient elution techniques to be applied, thanks to the wider lipophilicity range covered by the derivatized compounds. 

In spite of widespread applications, the exact mechanism of retention in reversed-phase chromatography is still controversial. Various theoretical models of retention for RPC were suggested, such as the model using the Hildebrand solubility parameter theory [32,51-53], or the model supported by the concept of molecular connectivity [54], models based on the solvophobic theory [55,56) or on the molecular statistical theory [57j. Unfortunately, sophisticated models introduce a number of physicochemical constants, which are often not known or are difficult and time-consuming to determine, so that such models are not very suitable for rapid prediction of retention data. 

The cavity model of solvation provides the basis for a number of additional models used to explain retention in reversed-phase chromatography. The main approaches are represented by solvophobic theory [282-286] and lattice theories based on statistical thermodynamics [287-291]. To a lesser extent classical thermodynamics combining partition and displacement models [292] and the phenomenological model of solvent effects [293] have also been used. Compared with the solvation parameter model all these models are mathematically complex, and often require the input of system variables that are either unknown or difficult to calculate, particularly for polar compounds. For this reason, and because of a failure to provide a simple conceptual picture of the retention process in familiar chromatographic terms, these models have largely remained the province of the physical chemist. 

Figure 4,14. Diagram of the thermodynamic cycle used to explain retention in reversed-phase chromatography by solvophobic theory. Na = Avogadro number, AA = reduction of hydrophobic surface area due to the adsorption of the analyte onto the bonded ligand, y = surface tension, = energy correction parameter for the curvature of the cavity, V = molar volume, R = gas constant, T = temperature (K), Pq = atmospheric pressure, AGydw.s.i a complex function of the ionization potential and the Clausius-Moscotti functions of the solute and mobile phase. Subscripts i = ith component (solute or solvent), S = solute, L = bonded phase ligand, SL = solute-ligand complex, R = transfer of analyte from the mobile to the stationary phase (retention), CAV = cavity formation, VDW = van der Waals interactions, ES = electrostatic interactions.

Fig. 7,2. Solvent effects in reversed phase chromatography. The individual terms of the solvophobic equation are plotted as the function of the composition of the water-acetonitrile eluent. The solute is undissociated toluic acid. The data was gathered using an ODS column at ambient temperature. The logarithm of the capacity factor on the ordinate can be considered as a dimensionless energy appropriate at the temperature of the experiment. Reproduced from Horvath and Melander (1978), with permission.

Fig. 3. Retention in reversed-phase chromatography according to the model of interphase and solvophobic effects. Due to the elevated cohesion energy density within the partly aqueous mobile phase, the energy liberated upon closing a cavity therein exceeds the energy required to create a new cavity within the less cohesive alkyl chain interphase.

LC Tan, PW Carr. Revisionist look at solvophobic driving forces in reversed-phase liquid chromatography. II. Partitioning vs. adsorption mechanism in monomeric alkyl bonded phase supports. J Chromat A 775 1-12, 1997. 

The exact mechanism(s) of solute retention in reversed-phase high-performance liquid chromatography (RPLC) is not presently well understood. The lack of a clear understanding of the mechanics of solute retention has led to a myriad of proposals, including the following partition (K21, L6, S16) adsorption (C9, CIO, H3, H15, H16, K13, L3, T2, U2) dispersive interaction (K2) solubility in the mobile phase (L7) solvophobic effects (H26, K6, M5) combined solvophobic and silanophilic interaction (B9, M12, Nl) and a mechanism based upon compulsary absorption (B5). 

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