A Simple Dual-Track HPLC-UV Methodology for Monitoring Primary Antiarrhythmic Drugs and Their Active Metabolites in...

Objectives: The aim of the work was to present a method for routine determination of antiarrhythmic drugs, propafenone (PPF), its two metabolites, 5-hydroxypropafenone (5OHPPF) and N-depropylpropafenone (NDPPF), mexiletine (MEX), amiodarone (AD) and desethylamiodarone (DEAD) in serum. Methods: A simple isocratic HPLC-UV system with a manual injector was applied. The separations were performed at ambient temperature on Supelcosil LC-CN column (150 × 4.6 mm, 5 μm). Two analytical procedures (A and B) were used: (A) for AD and (B) for PPF/MEX. The mobile phase for (A) was a mixture of: CH₃OH:CH₃CN:H₂O:0.5M KH₂PO₄ (200:100:194:6 v/v + 0.1 mL 85% H₃PO₄ per 500 mL). The slightly acidified serum sample was extracted with hexane and the analytes were detected at 240 nm. The mobile phase for (B) was a mixture of: CH₃CN:H₂O:0.5M KH₂PO₄ (185:310:5 v/v + 0.1 mL 85% H₃PO₄ per 500 mL). The alkalized serum sample was extracted with diisopropyl ether, then back extracted into 0.01M HCl and finally the analytes were detected at 210 nm. Results: The method was calibrated with adequate selectivity and specificity in the range of 20-4000 ng/mL for AD, DEAD and MEX, 10-4000 ng/mL for PPF and 10-500 ng/mL for 5OHPPF and NDPPF. For all analytes, precision and accuracy fulfilled EMA requirements, i.e., ≤15% (≤20% for LLOQ), ensuring the reliability of the measurements. Conclusions: The method can be suitable for laboratories equipped with basic HPLC apparatus as an economical alternative to the LC-MS/MS technique.