Simultaneous determination of digoxin and digitoxin by derivatization and high performance liquid chromatography

A simple and sensitive liquid chromatographic method has been developed for the determination of digoxin and digitoxin by derivatization. The method i s based on the derivatization of digoxin and/or digitoxin with derivatizing agent, 1-naphthoyl chloride, in pyridine at 50 degrees C. Under the mild c ondition, the digoxin and digitoxin were derivatized into their highly sens itive derivatives. After the derivatization reaction, a dimethylamine-aceto nitrile solution was added to the reaction mixture to eliminate the interfe rence from the excess derivatizing agent. The derivatives obtained were per formed on a reversed-phase C-8, column with 88% acetonitrile as the mobile phase. The parameters affecting the derivatization of digoxin and digitoxin , including reaction temperature, reaction time, and the amount of derivati zing agent, were investigated. The linear ranges of the method for the dete rmination of digoxin and digitoxin were over 1-50 nmoL/mL and 5-50 nmol/mL; the detection limit (signal to noise ratio = 5; injection volume, 20 mu L) were 0.25 nmoL/mL and 0.30 nmol/ml, respectively. Partial application of t he method to the analysis of digoxin in commercial tablets has proven to be satisfactory.