Fully Automated Online Sample Preparation AND Quantification of Amiodarone from Whole Blood using CLAM-LC-MS/MS

Importance of the Topic

Amiodarone is an effective antiarrhythmic agent with a narrow therapeutic window and potential for serious adverse effects.
Accurate and rapid quantification in whole blood is essential for therapeutic drug monitoring to optimize dosing and enhance patient safety.

Objectives and Study Overview

This work presents a fully automated workflow integrating online sample preparation with LC-MS/MS for the quantification of amiodarone and its active metabolite desethyl-amiodarone directly from whole blood.
The primary goals were to minimize manual handling, achieve high throughput, maintain analytical robustness, and enable emergency sample prioritization.

Methodology

Whole blood specimens were centrifuged and the resulting serum aliquots were loaded into a CLAM (Clinical Laboratory Automated sample preparation Module).
The CLAM unit performed automated sample cleanup and transferred aliquots online to the LC-MS/MS system.
Sample preparation steps included protein precipitation and online solid-phase extraction using a pre-column.

Instrumentation Used

Key components of the analytical setup included:

• Nexera X2 UPLC system with dual-column configuration (M&N Nucleodur C8 pre-column and Kinetex F5 analytical column).
• Shimadzu CLAM module for automated sample handling.
• Shimadzu LCMS-8050 triple quadrupole mass spectrometer with ESI source operating in positive mode.
• Gradient conditions optimized for rapid separations (<2.5 min duty cycle).
• MRM transitions for amiodarone, desethyl-amiodarone, and deuterated internal standards.

Main Results and Discussion

The system achieved baseline separation and quantification of both analytes in under 2.5 minutes, supporting high sample throughput.
Calibration curves over 0.5–3 ng/µL yielded correlation coefficients (r2) of 0.9994 for both compounds, demonstrating excellent linearity.
The limit of detection was 0.5 ng/µL, with high signal intensities (up to 750 000 counts for desethyl-amiodarone).
Minimal matrix interference and low background noise were observed, attributed to the CLAM cleanup protocol and use of a pre-column.
Emergency samples could be queued and measured with priority, an important feature for clinical decision making.

Benefits and Practical Applications

The fully automated CLAM-LC-MS/MS workflow offers:

• Reduced manual labor and risk of sample handling errors.
• High analytical throughput suitable for busy clinical laboratories.
• Robust quantification with reliable sensitivity and reproducibility.
• Rapid turnaround times, including priority processing for urgent cases.

Future Trends and Opportunities

Potential developments include expanding the panel to multiple therapeutic drugs in a single run, further miniaturization of sample preparation, and integration with laboratory information management systems (LIMS).
Advances in artificial intelligence–driven data processing may enhance result interpretation and workflow efficiency.
Point-of-care adaptations could bring similar automation to decentralized environments.

Conclusion

This study demonstrates that coupling an automated sample preparation module with LC-MS/MS enables fast, accurate, and high-throughput quantification of amiodarone in whole blood.
The approach supports routine therapeutic drug monitoring with minimal hands-on time and reliable analytical performance.

References

No external literature references were provided in the source document.