Sample Preparation Method for Determination of Voriconazole in Plasma Using ISOLUTE® SLE+

Significance of the Topic

Voriconazole is a triazole antifungal agent used to treat severe fungal infections under therapeutic drug monitoring (TDM) due to its narrow therapeutic window and potential side effects. Reliable quantification in plasma is crucial for patient safety and efficacy.

Study Objectives and Overview

This work presents a supported liquid extraction (SLE) method using ISOLUTE SLE+ to isolate voriconazole from plasma for both UV and LC-MS/MS analysis. The aim is to simplify sample preparation, reduce matrix effects, and ensure high recovery and sensitivity across clinical concentration ranges.

Methodology and Instrumentation

Sample Preparation Procedure:

• Mix 200 µL plasma with internal standard DDPC (0.5 µg/mL) and 200 µL water.
• Load the solution onto ISOLUTE SLE+ columns (400 µL capacity) and let it equilibrate for 5 min.
• Elute analytes with two 900 µL portions of hexane:diethyl ether (1:1 v/v) under gravity or slight pressure.
• Evaporate under nitrogen and reconstitute in 2 mL water:methanol (1:1 v/v).
• Perform additional dilution (e.g. 10-fold) for LC-MS/MS as required.

Used Instrumentation:

• UHPLC: Shimadzu Nexera LC-30AD with ACQUITY UPLC BEH C18 column (2.1×50 mm, 1.7 µm).
• Mass spectrometer: Shimadzu LCMS-8060 with ESI in positive mode.
• Sample preparation: Biotage ISOLUTE SLE+ columns and 96-well plates.

Key Results and Discussion

• Calibration was linear from 0.1–250 ng/mL with r² = 0.999.
• SRM transitions optimized for voriconazole (350.15→127.10, 350.15→281.10) and DDPC (313.30→284.15).
• Phospholipid removal and matrix interference were effectively eliminated, yielding clean chromatograms at the LLOQ (0.1 µg/mL).
• Recovery rates exceeded 90% at 0.1, 1, and 10 µg/mL, with matrix factors below 9%, demonstrating method robustness.

Benefits and Practical Applications

• High analyte recovery and broad dynamic range suitable for TDM and pharmacokinetic studies.
• No emulsion formation and simplified workflow compared to traditional liquid-liquid extraction.
• Compatibility with both UV and LC-MS/MS detection modes enhances flexibility.

Future Trends and Opportunities

The SLE+ approach can be extended to other antimicrobial agents and small-molecule drugs. Integration with automated high-throughput platforms and coupling to high-resolution mass spectrometry may further enhance sensitivity and selectivity. Emerging sorbent chemistries will likely expand application scope.

Conclusion

The ISOLUTE SLE+ supported liquid extraction protocol offers a streamlined, high-recovery method for voriconazole quantification in plasma. Its versatility for both UV and LC-MS/MS analysis, combined with strong linearity and minimal matrix effects, supports reliable therapeutic monitoring in clinical settings.

Reference

Biotage Application Note AN928, 2019.