The
antitumor agent phyllanthoside is rapidly metabolized in vitro by mouse plasma. This metabolite has now been isolated from mouse plasma and its structural properties and cytotoxicity characterized. The isolated metabolite was estimated to be greater than 98% pure by HPLC analysis. Mass spectral analysis (fast atom bombardment and tandem mass spectrometry) indicated that the metabolite was the aglycone of
phyllanthoside that resulted from the cleavage of the
ester bond linking the aglycone and the
disaccharide moieties of
phyllanthoside. This identification was based on identical collision-induced dissociation spectra of both
phyllanthoside and the metabolite. The aglycone was not formed by mouse plasma that had been boiled, filtered to remove
proteins, or treated with 1.0 mM diisopropyl
fluorophosphate. These results suggest that aglycone formation occurs as a result of plasma
esterase activity. Michaelis-Menten constants, Vmax and Km, for conversion of
phyllanthoside to the aglycone at 22 degrees C were estimated to be 1.1 mmol/ml plasma/min and 2.0 mM, respectively. Concentrations of
phyllanthoside and metabolite required to inhibit cell-colony formation by human A204
rhabdomyosarcoma in vitro were 0.47 nM and 24 microM, respectively. The toxicity of
phyllanthoside, and perhaps its efficacy as an
antitumor agent in mice, may depend on its rate of conversion to the aglycone.