Entecavir HPLC-MS/MS quantitation and optimized sample preparation using human plasma: Validation in a...

Area of Science:

• Pharmacology
• Analytical Chemistry
• Biochemistry

Background:

• Entecavir is a crucial antiviral medication for treating chronic hepatitis B.
• Accurate quantification of entecavir in biological matrices is essential for therapeutic drug monitoring and pharmacokinetic studies.
• Existing methods may require improvement in terms of sensitivity, throughput, or specificity.

Purpose of the Study:

• To develop and validate a quantitative high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method for determining entecavir concentrations in human plasma.
• To establish a reliable analytical tool for pharmacokinetic assessments of entecavir.

Main Methods:

• Development of a sensitive HPLC-MS/MS assay utilizing isotope-labeled internal standards.
• Implementation of a high-throughput solid-phase extraction (SPE) for efficient sample preparation.
• Analysis performed using multiple reaction monitoring (MRM) transitions in positive ion mode.

Main Results:

• The method demonstrated a wide linear range (0.025–10 ng/mL) with excellent accuracy (interday/intraday: -3.4% to 5.3%) and precision (≤7.3%).
• Assay validation confirmed acceptable stability, minimal matrix effects, and efficient recovery.
• The validated method was successfully applied to a pharmacokinetic study of entecavir dispersible tablets (0.5 mg oral dose).

Conclusions:

• The developed HPLC-MS/MS method provides a robust and accurate means for quantifying entecavir in human plasma.
• This validated assay is suitable for supporting clinical pharmacokinetic investigations and therapeutic drug monitoring of entecavir.