Detecting metabolites of different transition metal-lithospermate B complexes after intravenous injection in rats

Aim: Lithospermate B (LSB) isolated from the traditional Chinese medicine danshen (Salvia miltiorrhiza) is an effective N+/K+-ATPase inhibitor and used to treat congestive heart failure. The inhibition of LSB on Na+/K+-ATPase is potentiated by forming complexes with transition metal ions. Here we investigated the safety and metabolites of different transition metal-LSB complexes in rats. Methods: LSB complexed with six different transition metal ions (Mg2+, Zn2+, Cr3+, Co2+, Ni2+ and Mn2+) were prepared. Adult male SD rats were injected with the different metal-LSB complexes (50 mg/kg, iv), and their bile and blood samples were collected. The metabolites of the metal-LSB complexes in the samples were analyzed using mass spectroscopy. Results: In rats injected with LSB complexed with Mg2+, Zn2+, Cr3+, Ni2+ or Mn2+, LSB and its four putative metabolites were equivalently detected in their bile samples. Mn2+-LSB exhibited distinct metabolite profiles compared with the other four metal-LSB complexes. The four putative metabolites were identified as 3-monomethyl-LSB, 3,3 ''-dimethyl-LSB, 3,3'''-dimethyl-LSB and 3,3 '',3'''-trimethyl-LSB. The tracking of successive bile samples of rats injected with Mg2+-LSB, Zn2+-LSB and Mn2+-LSB concurrently demonstrated that LSB was firstly methylated at position 3, then at position 3 '', and, finally, the 3''' hydroxyl group. All rats injected with Co2+-LSB died. Conclusion: Zn2+-LSB, Cr3+-LSB, Ni2+-LSB or Mn2+-LSB produces identical four methylated metabolites of LSB in rats, and seemed to be as safe as LSB or Mg2+-LSB.