MSACL 2026 Abstract

Development and Validation of a Rapid LC-MS/MS Assay for Serum Amiodarone and Desethylamiodarone

Lori Good
Alberta Precision Laboratories

Lori Good (Presenter) Alberta Precision Laboratories

Presenter Bio: I have worked in the Analytical Toxicology department at Alberta Precision Laboratories for 24 years. During this time, I have gained extensive experience working with both GC-MS and LC-MS/MS technologies. Through hands-on laboratory experience, I have developed strong expertise in method development, instrument troubleshooting, and analytical problem-solving. I currently work as a Lab Scientist, applying this knowledge to support high-quality laboratory testing and operations.

Relevant Financial Disclosures (within past 24 months, reported on May 19, 2026) No relevant financial relationship(s) to disclose. Conflict mitigation NOT REQUIRED.

INTRODUCTION:
Amiodarone is a highly lipophilic antiarrhythmic medication associated with significant toxicity and complex pharmacokinetics, necessitating therapeutic drug monitoring (TDM) to support safe and effective patient management. Desethylamiodarone, the major active metabolite of amiodarone, also contributes to pharmacologic and toxicologic effects. Traditional high-performance liquid chromatography with diode-array detection (HPLC-DAD) methods may be limited by longer analysis times, lower specificity, reduced sensitivity, and complex sample extraction procedures. To improve analytical performance and laboratory workflow, an LC-MS/MS method for the simultaneous quantitation of serum amiodarone and desethylamiodarone was developed and validated.

METHODS:
Sample preparation consisted of protein precipitation of 100 µL serum with acetonitrile. Chromatographic separation was achieved using a Zorbax RRHD Eclipse Plus C18 analytical column with gradient elution consisting of 0.1% formic acid in water and 0.1% formic acid in acetonitrile. Total chromatographic runtime is 4.5 minutes. Detection was performed using tandem mass spectrometry in multiple reaction monitoring mode for the simultaneous analysis of amiodarone, desethylamiodarone, and their corresponding deuterated internal standards. Method validation included assessment of analytical measurement range, precision, carryover, matrix effects, accuracy, and patient correlation with the previous HPLC-DAD method.

RESULTS:
The LC-MS/MS assay demonstrated improved analytical specificity and sensitivity compared with the previous HPLC-DAD method while significantly reducing total sample preparation and analysis time. Protein precipitation provided a simple and efficient extraction approach suitable for routine clinical laboratory implementation. Chromatographic separation of amiodarone and desethylamiodarone was achieved within a 5.5-minute runtime with excellent peak shape and separation from other lipophilic compounds. Method validation, performed in accordance with clinical laboratory guidelines, demonstrated excellent linearity (r² ≥ 0.9990), intra- and inter-assay precision with (CV ≤ 2.18%), minimal carryover (< 5% of the lowest calibrator peak area), and acceptable accuracy across the analytical measurement range. Method comparison studies demonstrated acceptable agreement with the existing HPLC-DAD assay. However, desethylamiodarone demonstrated reduced recovery with the HPLC-DAD method due to analyte loss during the sample drying step, resulting in concentrations up to 20% lower than those obtained by LC-MS/MS. The HPLC-DAD internal standard did not compensate for this loss as it is not as volatile as desethylamiodarone. Elimination of the drying step in the LC-MS/MS method improved both accuracy and precision for desethylamiodarone quantitation.

CONCLUSION:
A rapid and robust LC-MS/MS assay for serum amiodarone and desethylamiodarone was successfully developed for clinical therapeutic drug monitoring. The method provides improved analytical performance over the previous HPLC-DAD assay while utilizing a simple protein precipitation extraction and short chromatographic runtime. Implementation of this assay supports improved laboratory efficiency, enhanced analytical specificity, and improved quantitation of desethylamiodarone for routine monitoring of amiodarone therapy.