Quantitation of Tricyclic Antidepressants in Serum by UHPLC-Triple Quadrupole Mass Spectrometry

Significance of the Topic

Tricyclic antidepressant drugs are used to treat major depressive disorders, anxiety, and chronic pain. They feature a narrow therapeutic window, making monitoring of serum concentrations essential to avoid adverse effects ranging from agitation to coma or death. Advanced analytical methods such as UHPLC coupled with triple quadrupole mass spectrometry provide the speed, sensitivity, and robustness required for routine therapeutic drug monitoring in clinical settings.

Study Objectives and Overview

This study aims to develop and validate a straightforward, rapid, and reliable protocol for the simultaneous quantitation of 13 tricyclic antidepressants, their metabolites, and two atypical neuroleptics in human serum. The method utilizes the Bruker Elute UHPLC system combined with the EVOQ LC-TQ Elite MS/MS and a commercial ClinMass TDM sample preparation kit.

Methodology and Instrumentation

Sample preparation involved protein precipitation of 50 µL serum with isotopically labelled internal standards provided by the ClinMass TDM kit. After centrifugation, the supernatant was directly injected into the UHPLC-MS/MS system. Chromatographic separation was achieved in 3.8 minutes using a rapid gradient and a dedicated TDM column.

Used Instrumentation

• Bruker Elute UHPLC with ClinMass TDM MS9030 column and MS9000 kit reagents
• Bruker EVOQ LC-TQ Elite triple quadrupole mass spectrometer with VIP H-ESI source
• ClinMass TDM calibration standards and quality controls

Results and Discussion

The optimized protocol achieved baseline separation of all 15 analytes within 3.8 minutes. Calibration curves constructed at three concentration levels demonstrated excellent linearity (R2 = 0.9985–1.0000). Precision tests yielded relative standard deviations below 6% for all compounds, while accuracy bias remained within ±9%, satisfying common bioanalytical guidelines. Representative MRM trace overlays and the doxepin calibration curve confirmed method robustness and dynamic range.

Practical Contributions and Applications

• Minimal sample volume (50 µL) and simple precipitation workflow increase laboratory throughput.
• Rapid analysis time supports high-volume clinical research and therapeutic drug monitoring operations.
• High sensitivity, precision, and accuracy enable reliable patient sample quantitation across the therapeutic range.

Future Trends and Opportunities

Emerging developments in UHPLC-MS/MS, such as automated sample handling, expanding analyte panels, and integration with data-driven workflows, will further streamline therapeutic drug monitoring. Advances in high-resolution mass spectrometry and AI-based data analysis promise enhanced selectivity and real-time decision support.

Conclusion

The described UHPLC-triple quadrupole MS/MS method combining the Bruker Elute and EVOQ systems with the ClinMass TDM kit achieves fast, precise, and accurate quantitation of tricyclic antidepressants and related compounds in human serum. Its performance and ease of use make it well suited for routine clinical and research laboratories focused on therapeutic drug monitoring.

References

• Martin R., Czentnar Z. Quantitation of Tricyclic Antidepressants in Serum by UHPLC-Triple Quadrupole Mass Spectrometry. Bruker Daltonik GmbH, 2018.