LC-MS/MS Analysis of Immunosuppressant Drugs in Whole Blood using the Xevo TQ Absolute with the Capitainer® B Device for Clinical Research

Significance of the topic

Monitoring immunosuppressant drugs is critical in clinical research and therapeutic drug management to ensure patient safety and optimize dosing. High-throughput methods capable of delivering accurate results from minimal sample volumes enhance patient compliance, reduce sample handling burdens, and accelerate data generation in both research and routine laboratory settings.

Objectives and study overview

This work presents a rapid LC-MS/MS method for simultaneous quantification of cyclosporine, everolimus, sirolimus, and tacrolimus in human whole blood. Key goals included minimizing sample volume via a Capitainer® B device, reducing analysis time, and achieving robust sensitivity and precision for clinical research applications.

Methodology and instrumentation

Sample preparation involved depositing 30 µL of whole blood onto a Capitainer B device to form a dried 10 µL spot, followed by overnight drying. Extraction was performed using liquid–liquid partitioning (tert-methyl butyl ether) and acidification. Dry extracts were reconstituted in equal proportions of mobile phases.

Chromatography employed a Waters ACQUITY UPLC I-Class PLUS system with an HSS C18 SB column (2.1 × 30 mm, 1.8 µm) at 55 °C, run time of 1.5 minutes per injection, and total cycle time of 2.2 minutes. Mobile phases consisted of water and methanol, each containing 0.05 mM ammonium fluoride. Detection used a Xevo TQ Absolute triple quadrupole mass spectrometer in positive electrospray MRM mode. Data were processed via MassLynx v4.2 with TargetLynx.

Main results and discussion

Signal-to-noise ratios exceeded 10:1 for lowest calibrators across five runs, confirming sufficient sensitivity. Calibration curves over defined ranges yielded linearity with r2 ≥ 0.99. Precision studies (n = 20–25) showed total precision and repeatability ≤ 7.6% RSD. No carryover was observed post high-concentration injections. External Quality Assurance samples demonstrated excellent accuracy compared to all-laboratory trimmed means.

Benefits and practical applications

• High analytical sensitivity from sub-10 µL blood spot enables less invasive sampling.
• Rapid throughput (2.2 minutes per sample) supports large-scale studies.
• Excellent precision and accuracy ensure reliable therapeutic drug monitoring in clinical research.

Future trends and applications

Further integration of microsampling devices with automated extraction systems could enable point-of-care monitoring. Advances in high-resolution mass spectrometry and alternative ionization techniques may expand analyte panels and improve sensitivity. Combining dried blood spot sampling with remote collection platforms can facilitate decentralized clinical trials and therapeutic monitoring.

Conclusion

The described LC-MS/MS method provides a fast, sensitive, and precise approach for simultaneous quantification of four key immunosuppressant drugs from minimal blood volumes. Use of the Capitainer B device for volumetric dried blood spot collection, combined with rapid UPLC and triple quadrupole detection, delivers robust performance suitable for clinical research applications.

Instrumentation used

• Waters ACQUITY UPLC I-Class PLUS system
• Waters ACQUITY UPLC HSS C18 SB column (1.8 µm, 2.1 × 30 mm)
• Xevo TQ Absolute triple quadrupole mass spectrometer
• Capitainer® B microsampling device

Reference

• Deprez S, Van Uytfanghe K, Stove CP. Analytica Chimica Acta. 1242 (2023) 340797.