Fast LC-HRAM-MS quantification of 36 antidepressants in human plasma using the Orbitrap Exploris 120 mass spectrometer for clinical research

Importance of Topic

Therapeutic drug monitoring of antidepressants supports optimized dosing and safety by accurately quantifying drug levels in plasma. High-resolution accurate mass (HRAM) LC-MS enables selective and sensitive analysis, increasing confidence in clinical research data.

Objectives and Study Overview

• Develop a single LC-HRAM MS method for quantifying 36 antidepressants in human plasma.
• Implement the method on a Thermo Scientific Orbitrap Exploris 120 MS coupled to a Vanquish Flex UHPLC.
• Validate analytical performance: linearity, accuracy, precision and lower limits of quantification (LLOQ).

Methodology

• Sample preparation by protein precipitation: 50 µL plasma with 100 µL acetonitrile containing deuterated internal standards.
• Chromatography on a Hypersil GOLD Phenyl column (2.1 × 100 mm, 1.9 µm) using a 0.5 mL/min gradient of 5 mM ammonium formate with 0.1% formic acid in water (A) and methanol (B).
• Mass spectrometry in full-scan positive mode (m/z 100–500) at 60,000 resolution using heated electrospray ionization (HESI).
• Data acquisition and quantification performed with Thermo Scientific TraceFinder 5.1 software.

Used Instrumentation

• Thermo Scientific Vanquish Flex Binary UHPLC system
• Thermo Scientific Hypersil GOLD Phenyl analytical column (2.1 × 100 mm, 1.9 µm)
• Thermo Scientific Orbitrap Exploris 120 mass spectrometer with HESI source
• TraceFinder 5.1 data processing software

Main Results and Discussion

• Linearity: calibration curves for all compounds showed R² > 0.993 with 1/x weighting.
• Accuracy of back-calculated calibrators ranged from 92.9 to 106.6% and precision < 7.3% across three levels.
• LLOQs extended below the lowest calibrator, meeting 80-120% accuracy and <20% CV for all analytes.
• Intra-assay precision and accuracy: 85.2–113.9% accuracy, CV < 6.5% (n=5, three days).
• Inter-assay precision and accuracy: 89.2–114.8% accuracy, CV < 6.4% (n=15).

Benefits and Practical Applications

• Rapid analysis: <7 minutes per sample throughput.
• Simple sample preparation with offline protein precipitation.
• High sensitivity and selectivity for comprehensive monitoring of multiple antidepressant classes.
• Supports clinical research and therapeutic drug monitoring workflows.

Future Trends and Potential Applications

• Expansion to include metabolites and additional psychotropic compounds.
• Integration with automated sample preparation and multiplexed assays.
• Application in real-time clinical decision support and personalized medicine.
• Adoption of AI-driven data interpretation to streamline HRAM MS workflows.

Conclusion

A robust LC-HRAM MS method on the Orbitrap Exploris 120 enables fast, accurate quantification of 36 antidepressants in human plasma. The validated workflow meets stringent performance criteria and offers a valuable tool for therapeutic drug monitoring in clinical research.