Utility of Dried Sample Spot (DSS) for voriconazole TDM

Alicia Gomez Lopez, Macarena Cuadros Nuñez, Ana Maria Donoso, Leticia Bernal Martinez


Background: Variability of the pharmacokinetics of voriconazole among and within individual patients is contributing to recommend TDM for this particular azole antifungal. To extend the possibilities of TDM for patients the Dried Sample Spot (DSS) procedure would be an interesting alternative. DDS would offer many advantages in terms of sampling, transportation, storage and biosafety when compared to classical collection methods for TDM. Basically this method uses very small volume of sample that is deposited in a paper card and dried under standard condition. Therefore, voriconazole is re-solubilized from the paper matrix to be tested using a technical method as much sensitive as possible. The objectives of this study were 1) to develop a dried sample spot (DSS) assays for the quantification of voriconazole in clinical samples using a high-performance liquid chromatographyultraviolet detection (HPLC-UV) procedure developed in our laboratory. 2) to stablish the correlation between paired serum samples and dried spot samples from monitored patients 3) .to stablish the correlation between paired samples from an alternative experimental model of treated larvae of Galleria mellonella

Material/methods: Volumes of 30 µL of samples (control samples prepared in serum and larval haemolymph and clinical samples from patients and from the experimental model) were deposited in the paper cards and dried overnight. Voriconazole was extracted from each card by vortexing and sonication using 500µL of a mixture of methanol and water (60:40). Once centrifuged, all supernatant was evaporated to a final volume of 100µL.Afterwards the resultant extract was transferred to a glass vial for HPLC analysis in a Waters Alliance 2695 HPLC/PDA using a stepwise gradient elution profile

Results: The validation procedure establishes that the method was linear between concentrations ranging from 0.25 and 8 mg/L for both matrices evaluated (serum and haemolymph), being both correlation coefficients (r 2 ) higher than 0.99. Assay reproducibility (%variation coefficient) and accuracy (%relative error) meet acceptance criteria. The concentrations of voriconazole in DSS samples were in good agreement with those in serum or haemolymph. The slope of the regression lines connecting voriconazole concentration based upon the conventional and DSS methods were very close to 1 (0.98-1.10)

Conclusions: The results show that the DSS technique developed is valid and reproducible to study exposure to voriconazole. The ability to handle small sample volumes provides interesting applications of this method to individual studies and experimental models as described herein


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Full conference title: 

26th European Congress of Clinical Microbiology and Infectious Diseases
    • ECCMID 26th (2016)