No abstract

Guanabenz (E-2,6-dichlorobenzylideneaminoguanidine acetate, Wy-8678) and its identified metabolites were tested for their ability to displace [3H]clonidine from the cerebral a, -receptors of rat and dog. The Ki' s for inhibition of [3H]clonidine binding to rat membranes were 1.97, 0.96, 14.0, and 108 nM for clonidine, guanabenz, p-hydroxyguanabenz, and the Z-isomer of guanabenz, respectively. The other metabolites at concentrations of 10,000 nM did not displace [3H]clonidine. Scatchard analysis indicated single populations of binding sites with KD' s of 2.37 and 2.39 nM and Bmax's of 5.39 and 5.12 pmollg for rat and dog preparations, respectively. Hill analysis yielded coefficients approximating unity, indicating a lack of cooperativity in binding. The high affinity of guanabenz relative to that of its metabolites for the a,-receptor provided the basis for the development of an assay capable of discriminating guanabenz from its weakly hypotensive (p-hydroxyguanabenz and the Zisomer of guanabenz) and pharmacologically inactive (2-6,dichlorobenzyl alcohol,2,6-dichlorobenzaldehyde, 2,6-dichlorobenzaldehyde azine,2,6-dichlorobenzaldehyde sernicarbizone, creatinine adduct of 2,6-dichlorobenzaldehyde, and an isomer of the creatinine adduct) metabolites. Antihypertensive agents with different mechanisms of action such as indoramin, prazosin, debrisoquine, and bethanidine bound little, if at all, and thus will not interfere with the assay of guanabenz.

Conventional dialysis cells were used in initial attempts to determine the binding characteristics of oxaprozin (4,5-diphenyl-2-oxazolepropionic acid, Wy-21,743). Equilibration required dialysis times up to 22 hours at 37 degrees C resulting in deterioration of plasma proteins, which in turn leads to highly variable binding values. In contrast, dialysis with Dianorm cells requires less than 4 hours to reach equilibrium. The configuration of the cell optimizes the contact between the solutes and the membrane and allows for a more efficient mixing and exchanging of the solute. The percentage of unbound drug was linearly related to total drug in human plasma samples to which oxaprozin in clinically relevant concentrations (55-405 micrograms/ml) had been added. Likewise, a linear relationship between total drug concentration and the percentage unbound was observed in specimens from a pharmacokinetic study in healthy volunteers. Clearance of total oxaprozin from plasma correlated with the percentage unbound drug. Thus the higher clearance observed under steady-state conditions (where concentrations are higher than following single dose administration) was caused by a larger unbound fraction available to the elimination sites.

Twelve normal subjects each received single 300-, 600-, and 1200-mg oral doses of oxaprozin according to a three-period crossover design. Total drug plasma concentrations did not increase in proportion to the dose administered. Total clearance (CIo) and volume of distribution (Vd) increased with dose, though elimination t1 2 remained unchanged. The fraction of unbound oxaprozin in plasma (fup) varied linearly with total plasma concentration: it increased from 0.068 per cent at 10 micrograms/ml to 0.180 per cent at 170 micrograms/ml. A parameter fup was therefore introduced to express the average degree of unbound drug plasma for a given dose, and to allow the calculation of unbound volume of distribution (Vdu) and intrinsic clearance (CIi) as if binding were constant. Even though fup increased with dose, the overall binding in the body (fub approximately 0.52 per cent) was relatively stable. Neither Vdu nor CIi changed with dose; hence, unbound oxaprozin kinetics can be considered to be linear. Protein binding had no effect on unbound oxaprozin plasma levels within the given dose range, and there was a one-to-one proportionality between the dose administered and the unbound drug concentration in plasma.