UV Method for the Determination of Tricyclic Antidepressants’ from Human Plasma Using SOLA CX and Hypersil GOLD HPLC

Importance of the Topic

Accurate measurement of tricyclic antidepressants in human plasma is critical for therapeutic drug monitoring, pharmacokinetic studies, and clinical diagnostics. Challenges include complex biological matrices, low analyte concentrations, and the need for high reproducibility in high-throughput laboratories. The integration of advanced solid-phase extraction with reliable chromatographic separation addresses these demands, improving data quality and workflow efficiency.

Objective and Study Overview

This study demonstrates a UV-based HPLC method for quantifying four tricyclic antidepressants—doxepin, imipramine, amitriptyline, and trimipramine—in human plasma. The primary goals are to evaluate the performance of Thermo Scientific SOLA CX SPE cartridges for clean extraction and to assess separation efficiency and method precision using a Hypersil GOLD column.

Methodology and Used Instrumentation

Sample Pretreatment and SPE Procedure

• Aliquot 121.4 µL of blank human plasma into a tube.
• Add 13.6 µL of analyte standard and 15 µL of internal standard solutions, then dilute with methanol and mix.
• Condition SOLA CX cartridges (10 mg/1 mL) with 500 µL methanol followed by 500 µL water.
• Load 450 µL of a 1:2 mixture of spiked plasma and 100 mM sodium phosphate buffer (pH 6.0).
• Wash with 500 µL water containing 0.1% formic acid and then 500 µL methanol with 0.1% formic acid.
• Elute analytes with 500 µL acetonitrile plus 5% ammonia.
• Dry eluates without heating and reconstitute in 150 µL of 80:20 water/acetonitrile.

Chromatographic Conditions

• Instrument: Thermo Scientific Accela 600 HPLC system.
• Column: Hypersil GOLD, 150 × 2.1 mm, 3 µm particle size.
• Mobile Phase: water+0.1% formic acid and acetonitrile+0.1% formic acid (70:30 v/v).
• Flow Rate: 0.4 mL/min; Column Temperature: 30 °C; Injection Volume: 1 µL.

Main Results and Discussion

All four analytes eluted within eight minutes with sharp, symmetrical peaks. Method precision (%RSD) ranged from 4.0% to 5.1% across compounds, demonstrating excellent reproducibility. Recovery values exceeded 69% for each antidepressant, indicating efficient extraction by SOLA CX cartridges. These performance metrics underscore the suitability of this approach for routine bioanalysis.

Benefits and Practical Applications

This workflow offers several advantages:

• Enhanced reproducibility and peak cleanliness through advanced SPE technology.
• Reduced solvent consumption and lower elution volumes, cutting costs and environmental impact.
• Faster sample preparation and analysis, increasing laboratory throughput.
• Robust performance in complex biological matrices, supporting clinical and pharmaceutical research.

Future Trends and Opportunities

Emerging developments in SPE materials, microflow chromatography, and detector technologies will further improve sensitivity and reduce analysis time. Integration with mass spectrometry can extend quantitation limits and selectivity. Automated, miniaturized sample-prep systems may enable point-of-care testing and expand applications in personalized medicine and therapeutic monitoring.

Conclusion

The combination of SOLA CX SPE cartridges and Hypersil GOLD HPLC achieves reliable extraction and separation of tricyclic antidepressants from human plasma. The method delivers high precision, satisfactory recovery, and efficient solvent use, making it well suited for high-throughput clinical and bioanalytical laboratories.