This work examined the stability of trans alpha-acetoxytamoxifen in Krebs-Henseleit buffer (pH 7.4), in the presence of Sprague-Dawley female rat liver slices, rat liver homogenate and hepatic subcellular fractions, including microsomes and mitochondria, at pH 7.4 and at 37 degrees C over 300 min. The rate of hydrolysis was determined using high-performance liquid chromatography, and degradation profiles were obtained from which the rate and order of degradation were both evaluated. By applying zero-, first-, second- and third-order models of drug disappearance and the generation of by-products, first- and second-order appeared to produce the best fit. trans-alpha-Acetoxytamoxifen degraded rapidly in buffer and more slowly in the biological systems, probably due to the fact that the agent partitions into the hydrophobic component of the biological tissue and hence degrades at a much slower rate. The principal degradation products were trans-alpha-hydroxytamoxifen and, to a lesser extent, cis-alpha-hydroxytamoxifen. Another peak could not be identified. The production of trans-alpha-hydroxytamoxifen was enhanced in the presence of biological enzymes, whereas the concentration of the cis isomer remains relatively constant in buffer only (pH 7.4) and in the presence of biological enzymes. Therefore, the formation of identical adducts with DNA is consistent, because it has been shown that alpha-acetoxytamoxifen breaks down to form alpha-hydroxytamoxifen in vitro. The percentage of trans-alpha-acetoxytamoxifen remaining after 300 min was 40% in mitochondria and 32% in homogenate. The half-life (t1/2) was calculated for each condition by applying zero-, first- and second-order rate kinetics.