Determination of Fluconazole in Children in Small Blood Volumes Using Volumetric Absorptive Microsampling (VAMS) and Isocratic High-Performance Liquid Chromatography-Ultraviolet (HPLC-UV) Detection.

: A simple method for quantifying fluconazole in small blood volumes has been developed using volumetric absorptive microsampling (VAMS) technology and isocratic high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection. : For sample collection, Mitra devices are used to keep the sample volume at 10 µL. For the quantitative determination of fluconazole, the Mitra samples are extracted using acetonitrile as the extraction agent, containing 2-(4-chlorophenyl)-1,3-bis(1,2,4-triazol-1-yl)propan-2-ol as the internal standard. A Synergi 4 μm Polar-RP 80 Å (150 × 2 mm) column forms the stationary phase, and a mixture of acetonitrile and phosphate buffer is the mobile phase. The UV detection is set at a wavelength of 210 nm. The therapeutic concentration range of 5 to 160 mg/L is covered, and the linear equation with 1/x weighting is used to determine unknown samples. This method has been validated according to the current EMA and FDA guidelines for bioanalytical methods. : The validation data obtained after analysing whole blood samples (EDTA) showed within- and between-run accuracy between 94.4% and 115% and precision between 0.4% and 9.4%, respectively. A lower limit of quantification (LLOQ) of 5 mg/L was sufficient for therapeutic drug monitoring in paediatric patients receiving fluconazole as antifungal prophylaxis after haematopoietic cell transplantation. : So far, 211 samples from 49 patients were successfully analysed, and concentrations between 5.84 mg/L and 107 mg/L were determined for whole blood Mitra samples. To our knowledge, this is the first application of VAMS technology using simple and cheap HPLC-UV quantification.