Principle of Lipophilicity Measurements by RPLC 521 Description of the Method

Lipophilicity of organic compounds can be measured with any standard HPLC equipment. Usually UV detection can be used, but any other detection method is appropriate (e.g., refractive index or electrochemical). The choice of the stationary and mobile phase are discussed in section 5.3 and 5.4. The lipophilicity index measured by RPLC is derived from the capacity factor k (or sometimes written as k') given by k = (tr — to)to (1)

where tx and t0 are the retention times of the solute and of an unretained compound. This latter can be the solvent front, or an inorganic salt such as sodium nitrate or potas-

mobile phase composition ip

Figure 1. Schematic representation of the measurement of a lipophilicity index by RPLC. Isocratic (log k) capacity factors are obtained from the retention time measured at a selected mobile phase composition; polycratic (log kj) capacity factors are calculated from an extrapolation procedure.

sium bichromate. The logarithmic form, log k, can be used as a lipophilicity index. Instead of the isocratic log k values (obtained at a single selected mobile phase concentration), frequently polycratic log kw values are used, which are obtained by extrapolation of isocratic log k values against the mobile phase composition [7-9] (Fig. 1). This procedure should not be confused with the use of a mobile phase gradient, which is a typical HPLC separation technique. The mobile phase in RP-HPLC in the present context consists of a mixture of water and an organic compound, called organic modifier, typically methanol, acetonitrile or tetrahydrofuran. The most common procedure is the linear extrapolation with organic modifier content qr.

The slope S has been studied in more detail over the past few years and encodes to a certain extent hydrogen-bonding capability (see section 5.5.2).

It is often observed that the relationships between log k and cp are not linear, e.g., with protonated bases [9] or when using organic modifiers such as tetrahydrofuran and acetonitrile [10, 11], In such cases two alternatives have been used. Either the linear extrapolation is performed on part of the points, i.e., those with lowest organic modifier concentration, or a quadratic fit is used [12]:

A further alternative is to obtain log kw values from extrapolation from /iT(30) plots [13]. The £x(30) scale is an empirical measure of solvent polarity, and is based on charge transfer absorption measurements of 2,6-diphenyl-4-(2,4,6-triphenyl)-rc-pyridino)phenolate. Finally, the solvophobic theory of Horvath should be mentioned [14, 15], This theory was developed to account for the curvature in log k - q> relationships. In its simplified form this equation is:

where A, B and C are regression coefficients. A is a function of the mobile phase composition, £> is related to the dielectric constant of the mobile phase, and y is the surface tension of the solvent [14,15]. Since these latter two approaches are rarely used and offer no real advantages, we do not discuss the details.

A new chromatographic lipophilicity index cp0 was recently proposed [16]. It is defined at the concentration of organic component in the mobile phase, required for log k = 0, and varies therefore from 0 to 100 %. No advantages in terms of lipophilicity scale are apparent.

As seen in Fig. 2, the slope of the extrapolation of log k versus mobile phase composition qi may vary considerably (see also section 5.5). A proper expression of the differences in lipophilicity is found only at 100 % water composition. Therefore log kw values should be used as RPLC measured lipophilicity parameter [7-9, 17]. However, this view is not shared by others [6, 14,18,19]. Yamagami and colleagues argue that log kw values can predict log P values only for nonhydrogen-bonding or weak hydrogen-accepting substituents, but not for strong hydrogen-accepting substituents. However, such conclusion depends strongly on the type of stationary phase used. Particularly, interaction with nonprotected silanophilic groups may blur the picture (see below).

The intersection point of log k/cp lines is for closely related compounds often well defined, and in the range of cp = 70-100% methanol. However, at present no full theoretical basis for such behavior can be offered. Discussions can be found in [11, 20],

Figure 2. The choice of the RPLC lipophilicity index. Only log kw values have a broad range in lipophilicity values (compare mobile phase composition A, representing log kw values, and B). At certain mobile phase composition several compounds may have similar (at the isoelution point C) or even reversed lipophilicity values (for D) as compared with log /cw values.

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