Development and Validation of a Candidate Reference Measurement Procedure for Plasma Clozapine Quantification Using...

Area of Science:

• Analytical Chemistry
• Clinical Chemistry
• Pharmacology

Background:

• Accurate quantification of clozapine (CLO) in human plasma is crucial for effective therapeutic drug monitoring (TDM).
• Existing methods may lack the specificity and accuracy required for reliable CLO measurements.
• Development of a candidate reference measurement procedure (cRMP) is needed to standardize CLO quantification.

Purpose of the Study:

• To develop and validate a candidate reference measurement procedure (cRMP) for the accurate quantification of clozapine (CLO) in human plasma.
• To establish a reliable method for standardizing therapeutic drug monitoring (TDM) of CLO.
• To improve clinical outcomes and patient safety through precise CLO level determination.

Main Methods:

• Developed a candidate reference measurement procedure (cRMP) utilizing isotope dilution liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS).
• Systematically optimized chromatographic separation and evaluated sample pretreatment methods for clozapine.
• Rigorously validated the analytical performance, including specificity, recovery, precision, linearity, LoQ, LoD, carryover, stability, and method comparison, assessing measurement uncertainties according to GUM.

Main Results:

• The ID-LC-MS/MS method demonstrated excellent linearity (R² = 0.9988) over a wide concentration range (5.65-1693.51 ng/g).
• Achieved high precision with total coefficients of variation (CVs) as low as 0.65% and average recoveries between 97.80% and 99.28%.
• The method exhibited a low limit of quantitation (LoQ) of 2.73 ng/g and a limit of detection (LoD) of 0.91 ng/g, with well-defined measurement uncertainties.

Conclusions:

• The developed cRMP for clozapine quantification meets all criteria for reference measurement procedures, demonstrating excellent specificity, accuracy, precision, and traceability.
• This validated ID-LC-MS/MS method provides a robust tool for accurate clozapine measurement in human plasma.
• The established cRMP has the potential to standardize therapeutic drug monitoring (TDM) for clozapine, ultimately enhancing patient safety and clinical management.