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coefficients has been attempted using a Corasil I column impregnated with
oc-tanol (287) or a reversed-phase column which had been coated with oc-
tanol (288). In either case, the sample was injected into an aqueous
mobile phase. When the octanol-coated octyldecyl silica support was used,
Wayne R. Melander and Cuba Horvath
logarithm of the relative retention time was highly correlated with the
logarithm of the octanol-water partition coefficients:1
The exact nature of the relationship between chromatographic,retention
and the corresponding log F remains to be fully explicated and clarified.
There is ample basis, however, to expect chromatography to develop into a
powerful tool for extrabiological understanding and prediction of in vivo
effects of pharmaceuticals and toxic agents and for measurement of
biodvnamic hydrophobicity. In a similar fashion, it mav also serve as a
tool tor screening solvent extraction schemes of technological inipoii.
"E iie40. HIM (' a" ai yiy1o1e'e1 (<?'1i||o1? ft"" fM plh"re vhii bo von
Vttlifcttlly U&t) lU tt??U? ttlHtfcttlltlUbtt IH llife phases When the
conventional shake-flask method or its variant is used for the
determination of log P values (290).
O. Measurement of Equilibrium Constants for Association Processes in
Under certain conditions chromatographic retention measurements can be
used to determine the magnitude of equilibrium constants for reversible
associations between a sample component and a complexing agent present in
the eluent. The use of migration rate data to obtain equilibrium
constants for interactions between eluents of the solvent and the solute
is not novel. It has been developed for paper electrophoresis (29/, 292).
Chromatographic measurements discussed here are based on the
evaluation of the effect of the concentration of the complexing agent
(hetaeron) in the eluent on the retention factor of the eluite as!
treated in Section VI. If the stoichiometry of the hetaeron-solute
association process is 1:1. then Eq. (89) of Section VI can be used to
calculate the retention factor. A 'more general expression is available
for the scheme presented in Fig. 59.
In view of the equilibria involved mass balance yields an expression
the retention factor as follows
*. + X k&XJ i ** e =--------fcJ---------
1 + 2 [Xjf n K> i-1 j-1
where Kt is equilibrium constant for the Jth binding, which forms E2CS,
[KfgJ is the concentration of hetaeron in the mobile phase, is the
retention factor of the form of eluite combined with / hetaeron molecules
per molecule and k9 is the retention factor in absence of hetaeron (204).
In many cases it is convenient, or more conventional, to write Eq.
(109) in terms of the corresponding dissociation constants. For the Jth
Reversed-Phasc Chromatography 277
ELj + I. ,--------------> EL.
Fig. 39. A general representation of the binding of n molecules of
hetaeron, L, to an eluite E. Each binding step has an associated
equilibrium constant, K,, for (he formation of species containing /
molecules of hetaeron from the reaction of a species containing i - 1
molecules of hetaeron with hetaeron. As a consequence, the retention
factor can have a complex dependence on hetaeron concentration; the
relationship is given by Eq. (109).
Hum, the dissociation constant, KkJ is related to the association
= 1 /Kj
so that the retention factor can be expressed by
ko + i *.[*"]* n
1 + 2 IXJ n *u j"i i-i
Similar expressions have been obtained for the particular cases of mono-
protic acids and bases, diprotic acids and bases, and zwitterions (207,
208), and in each case the data conformed well to Eq. (111). It has also
been shown Q07) that the acid dissociation constants can be determined by
using reversed phase chromatography. The pAT, values of 10 aromatic acids
calculated from chromatographic data by employing Eq. (91) were
Wayne R< Mdander and Cnba Horvith
Comparison of tbe pff. Values of Organic Adds as Obtained by
Least-Squares Analvnfc of Chromatographic Data and by PMentlaiM#lrtr
Acid P*.. p K.
Chromatography Titration Literature Ref.
Benzoic 3." 3.78 4.19 (/-0)" 293
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