Introduction a Need for an Accurate Method for Partition Coefficient Measurements

The role of drug lipophilicity in pharmacokinetics and pharmacodynamics is both primordial and ubiquitous, as well documented in the literature [1-3]. While partition coefficients measured in n-octanol/water systems (Poa) have encountered enormous success in the studies of drug lipophilicity and structure-activity relationships, new structural parameters arising from a combination of partition coefficients in different solvent systems such as Alog Poct_a]k (= log Poa - log Paikane) are emerging as structural determinants particularly in blood-brain barrier permeation [4], gastrointestinal absorption [5] and skin penetration [6] of drugs (see Chapter 14). At this point, an essential condition is to determine precisely and accurately partition coefficients in each solvent system to minimize the errors in, for example, Alog Poa-aik■

Thermodynamically, the partition coefficient is defined as a constant relating the activity of a solute in two immiscible phases at equilibrium. A number of experimental models are currently used to simulate partition processes in biological systems and to determine lipophilicity. The "shake-flask" (SF) method using water and a poorly miscible organic solvent is a technique most widely used for measuring partition coefficients [7]. However, the SF method suffers from a number of limitations such as the precision of phase volume ratio, the (im-)purity of the solvent used, the (im-)purity, volatility and adsorption of solutes, and finally the formation of microemulsions induced by vigorous mechanical agitation as previously discussed by Dearden and Bres-nen [8] (see also Fig. 1). Clearly, a more accurate experimental method to determine partition coefficients is called for.

Partition, to be well distinguished from adsorption, chromatography has been explored as an alternative means for measuring lipophilicity. In particular, chromatographic retention parameters obtained by reversed-phase high-performance liquid chromatography (RP-HPLC) [9] (see Chapter 5) have become increasingly popular in replacing the octanol/water partition coefficient measured by the SF method. However, the mechanisms of retention in RP-HPLC are not truly identical, while being similar, to those of partitioning in «-octanol/water systems owing to the restricted mobility of the bonded alkyl chains and the presence of a solid support with a non-negligible proportion of residual silanol groups. Even for a polymeric support grafted with alkyl chains such as octadecylpolyvinyl alcohol, the solute retention behavior in some cases can be very different from the partitioning in octanol/water [T. Ter Laak, unpublished results]. While the RP-HPLC method remains useful, it is difficult - if not unlikely - to derive from it quantitative structural information such as hydrogen-bonding capacity.

In recent years, centrifugal counter-current chromatography, also known as centrifugal partition chromatography (CPC) [10], has been explored as a novel technique for

Figure 1. Problems inherent in partition coefficient measurements using the shake-flask method.

measuring liquid-liquid partition coefficients. This is a unique form of liquid-liquid partition chromatography that is free of a solid support. Thus, the problem of adsorption is avoided and solute retention depends solely on its partition coefficient. Two poorly miscible liquids are used as the stationary and mobile phases in the chromatograph. Centrifugal forces maintain the stationary phase, while the mobile phase is pumped through the system. During the past decade, various types of CPC systems such as the flow-through multilayer coil planet centrifuge, the horizontal flow-through multilayer coil planet centrifuge, the toroidal coil planet centrifuge, and the multichannel cartridges CPC have been used to determine partition coefficients [10].

In this chapter, we review the development and current state-of-the-art of this novel technique in lipophilicity measurements and its application in determining solute structural properties.

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