Simultaneous analysis for Remdesivirand its metabolite [with CLAM-2030]

Importance of the Topic

Rapid and accurate measurement of antiviral agents and their metabolites in biological samples underpins pharmacokinetic studies, therapeutic drug monitoring, and drug development. Simultaneous quantification of remdesivir and its active metabolite GS-441524 enhances our understanding of drug absorption, distribution, metabolism, and excretion profiles in plasma, which is critical for optimizing dosing regimens and ensuring safety in clinical settings.

Study Objectives and Overview

This study aimed to develop and validate a fully automated workflow combining CLAM-2030 sample preparation with Nexera X2 LC and LCMS-8060 triple quadrupole mass spectrometry for simultaneous quantification of remdesivir and GS-441524 in human plasma. Key goals included:

• Establishing robust chromatographic separation and MS detection parameters for both analytes and their stable isotope-labeled internal standards.
• Evaluating method linearity, precision, accuracy, recovery, matrix effects, stability, and carryover.

Methodology and Instrumentation

An automated platform (CLAM-2030) performed protein precipitation using 75% isopropanol and acetonitrile, followed by mixing, filtration, and transfer to an LC vial in approximately 7 minutes. Chromatographic separation employed a Shim-pack Scepter C18 column (50×2.1 mm, 1.9 μm) at 40°C with a water/formic acid and acetonitrile/formic acid gradient (0.4 mL/min) over 4.5 minutes. ESI positive mode MS/MS detection monitored optimized MRM transitions for remdesivir, GS-441524 and their 13C-labeled analogs. Full validation followed regulatory guidelines for bioanalytical methods.

Main Results and Discussion

Calibration curves showed excellent linearity for remdesivir (100–5000 ng/mL, R2 ≥ 0.9986) and GS-441524 (5–250 ng/mL, R2 ≥ 0.9997). Intra-day precision (RSD) ranged 0.9–2.0% for remdesivir and 2.3–3.6% for GS-441524; accuracies were 90.5–106% and 88.5–91.6%, respectively. Inter-day precision and accuracy remained within acceptable limits over three days. Recovery rates exceeded 90% for remdesivir and reached 90.8% for GS-441524, with minimal matrix effects. Carryover after high-concentration injections was below 20% of the lowest level for remdesivir and undetectable for GS-441524. Stability testing indicated potential degradation of remdesivir in refrigerated plasma; care in sample storage is recommended.

Benefits and Practical Applications

• High throughput: Automated sample prep reduces hands-on time and variability.
• Short cycle time: Combined preparation and analysis complete in under 12 minutes per sample.
• Regulatory compliance: Validation metrics meet bioanalytical criteria for precision, accuracy, and stability.
• Clinical utility: Enables reliable pharmacokinetic and therapeutic monitoring studies.

Future Trends and Potential Applications

Further integration of microflow LC and high-resolution MS could enhance sensitivity and reduce solvent consumption. Adaptation to other antiviral candidates and multiplexed panels will support comprehensive profiling of treatment regimens. Advances in on-line sample cleanup and emerging ambient ionization techniques may streamline workflows without chromatographic separation.

Conclusion

The automated CLAM-2030 and LCMS-8060 method delivers rapid, accurate, and robust simultaneous quantification of remdesivir and GS-441524 in plasma. It meets stringent validation criteria and offers a scalable solution for clinical pharmacology and drug development laboratories.

Used Instrumentation

• CLAM-2030 automated sample preparation system
• Nexera X2 UHPLC system
• Shim-pack Scepter C18 column (50×2.1 mm, 1.9 μm)
• LCMS-8060 triple quadrupole mass spectrometer

References

Shimadzu Corporation. Global Application Development Center. IMOTO Eishi. Simultaneous analysis for Remdesivir and its metabolite with CLAM-2030 and LCMS-8060. 2020.