A sensitive LC-MS/MS method for quantifying clofarabine triphosphate concentrations in human peripheral blood...

Area of Science:

• Pharmacology
• Analytical Chemistry
• Biochemistry

Background:

• Clofarabine triphosphate is the active intracellular metabolite of clofarabine.
• Accurate quantification of clofarabine triphosphate is crucial for understanding its pharmacokinetics and efficacy.
• Existing methods may lack the sensitivity or specificity required for cellular analysis.

Purpose of the Study:

• To develop and validate a rapid, sensitive, and selective LC-MS/MS method.
• To quantify clofarabine triphosphate concentrations in human peripheral blood mononuclear cells (PBMCs).
• To assess the stability of clofarabine triphosphate in biological samples.

Main Methods:

• Liquid chromatography-tandem mass spectrometry (LC-MS/MS) with negative electrospray ionization.
• Sample preparation involving Ficoll gradient centrifugation for PBMC isolation.
• Chromatographic separation on a CN column with an isocratic mobile phase.
• Validation across concentration ranges of 1.25-100 ng/10(7) cells.

Main Results:

• The developed LC-MS/MS method demonstrated high sensitivity and selectivity.
• The method exhibited acceptable accuracy and precision within the validated range.
• Clofarabine triphosphate showed stability under various conditions (bench-top, auto-sampler, freeze-thaw).

Conclusions:

• A robust and validated LC-MS/MS method for clofarabine triphosphate quantification in PBMCs has been established.
• This method is suitable for clinical research and therapeutic drug monitoring in cancer patients treated with clofarabine.
• The stability data supports the reliability of measurements in biological matrices.