Determination of topiramate in dried blood spots using single-quadrupole gas chromatography-mass spectrometry after flash methylation with trimethylanilinium hydroxide

- Topiramate can be measured in clinical concentrations in DBS by GC-MS.
- Topiramate is stable in DBS for 21 days at 45 °C, with minimal cost for storage and transport.
- The topiramate assay in DBS has adequate performance characteristics for clinical use.
- Topiramate fraction in plasma increases according to drug concentrations in whole blood.
- The use of whole blood levels of topiramate for patient monitoring should be considered.

Dried blood spots (DBS) sampling obtained from fingerpricks is a promising and patient friendly alternative for obtaining samples for drug quantification, that could be of interest for topiramate (TOP) therapeutic drug monitoring. The aim of this study was to develop and validate a simple and fast GC-MS assay for TOP measurement in dried blood spots (DBS). The method uses a liquid extraction of one 8 mm DBS, followed by a flash methylation with TMAH, and separation in a DB-5 ms capillary column. Total analytical run time was 15 min. Precision assays presented CV% lower than 9.1% and accuracy was in the range of 94.5-115%. TOP was stable at 25 and 45 °C up to 21 days. TOP presents saturable binding to red blood cells, resulting in a fraction in plasma (fp) of 0.09-0.03 at 0.8 μg ml−1 and 0.71-0.45 at 20 μg ml−1 (both at 25-50 Hct% range). The method was applied to DBS samples obtained after phlebotomy and fingerpicks from an adult individual after oral intake of 100 mg TOP (0.25-96 h post dose). Plasma and DBS concentrations were moderately correlated (r = 0.61), with estimated fp values in the range of 0.06-0.18. Translation of TOP DBS to plasma concentrations is challenging due to its concentration-dependent binding to erythrocytes. Thus, the use of whole blood concentrations for patients monitoring should be considered, which favors to the use of DBS in the clinical context.