In vitro and in vivo metabolism of pyronaridine characterized by low-energy collision-induced dissociation mass spectrometry with electrospray ionization

The in vitro and in vivo metabolism of pyronaridine, an antimalarial agent, was investigated in rats and humans. In vitro incubation of pyronaridine with rat and human liver microsomes resulted in the formation of 11 metabolites, with pyronaridine quinoneimine (M3) as the major metabolite. The structures of pyronaridine metabolites were characterized on the basis of the product ion mass spectra obtained under low-energy collision-induced dissociation (CID) ion trap mass spectrometry. Both pyronaridine (m/z 518) and M3 (m/z 516) produced the same product ion (m/z 447). These results could be explained by the characteristic neutral loss of a 69 Da fragment from M3 via gamma-H rearrangement and 1,7 sigmatropic shift, whereas the neutral loss of a 71 Da fragment from the pyronaridine occurred by charge site-initiated heterolytic cleavage. These fragmentations were further supported by the tandem mass spectrum of D(3)-pyronaridine. Other metabolites generated in the microsomal incubations were carbonylated, hydroxylated and O-demethylated derivatives. Pyronaridine and its metabolites were detected in both feces and urine after intraperitoneal administration to rats. The in vivo metabolic profile in rats was different from the in vitro profile. M3, a chemically reactive quinonimine, was not detected whereas O-demethylated derivatives (M14, M15, M16, and M19) were identified in fecal and urinary extracts. The role of quinoneimine metabolites in pyronaridine-caused toxicity should be further evaluated, although these metabolites or their conjugates were not detected in urine and feces.