Integrated pharmacokinetics and multidimensional drug-drug interactions assessment of bufadienolides based on UPLC-QqQ-MS/MS

Bufadienolides, the primary active constituents of the traditional Chinese medicine Venenum bufonis, exhibit significant pharmacological activity. However, the potential pharmacokinetic effects of drug-drug interactions among these compounds under combination treatment have not yet been investigated. Thus, this study focused on three representative bufadienolides (resibufogenin, bufalin, and arenobufagin) as the research subjects and assessed the pharmacokinetic interactions among them under co-administration conditions. Firstly, we developed a validated UPLC-QqQ-MS/MS analytical method using digoxigenin as the internal standard (IS) and performed accurate quantification of these compounds in all samples under multiple reaction monitoring (MRM) mode. This analytical method delivered high levels of specificity, recovery, accuracy, precision, and preserved sample stability. Subsequently, pharmacokinetic data for the three compounds under both single-agent and combination treatment conditions were analyzed using non-compartmental modeling. The results demonstrated significant pharmacokinetic variability among different bufadienolide compounds, reflecting distinct structure-pharmacokinetic relationships. Under combination treatment, the pharmacokinetic parameters were significantly altered due to mutual interactions among the bufadienolide compounds. In conclusion, this study has established a drug-drug interaction network for bufadienolides, providing a scientific foundation for the rational clinical application of Venenum Bufonis-based formulations. These findings may facilitate the optimized development of multi-component therapies with cardiotonic pharmacological activity, particularly in the treatment of cardiovascular diseases.