KINETICS OF CHLORAMBUCIL IN-VITRO - EFFECTS OF FLUID MATRIX, HUMAN GASTRIC-JUICE, PLASMA-PROTEINS AND RED-CELLS

The mechanisms involved in the bioavailability of chlorambucil or 4-[p -(bis[2-hydroxyethyl]amino)phenyl]-butyric acid are poorly understood. The effects of different matrices on the disintegration of chlorambuc il were investigated by HPLC, H-1 NMR, P-31 NMR, and mass spectrometry . Cellular incorporation and protein binding of the drug in vitro was assessed with [H-3]-chlorambucil. Decomposition of chlorambucil and it s major metabolite, phenylacetic acid mustard, to mono- and dihydroxy derivatives, was significantly faster in water than in PBS, (phosphate -buffered saline, pH 7.4). The hydrolysis of chlorambucil was as fast in plasma ultrafiltrate as in PBS; plasma proteins, preferentially alb umin, prevented this disintegration. In phosphate-buffered media, two additional stabile hydrolysis products were found which were character ised as the mono- and bis-phosphates of 4-[p-(bis[2-hydroxyethyl]amino )phenyl]butyric acid, results of the reaction of nucleophilic buffer s pecies with the aziridinium ion intermediates. Chlorambucil bound cova lently to plasma proteins and was incorporated into red cells. These i nteractions are likely to have a significant role in vivo, reducing th e bioavailability of the drug. High H+ concentration associated with h igh chloride concentration in human gastric juice had a stabilizing ef fect on chlorambucil. Incorporation of [H-3]-chlorambucil into red cel ls was inhibited in a concentration-dependent fashion by whole human p lasma as well as by albumin. We conclude that the chemico-biological i nteractions demonstrated in the present investigation provide explanat ions for the remarkable pharmacokinetic differences observed intra- an d inter-individually in the clinical use of chlorambucil. The present information is important, when clinical or in vitro evaluation of effi cacy and bioavailability of chlorambucil is considered. (C) 1997 Elsev ier Science Ireland Ltd.