title: Sampling only ten microliters of whole blood for the quantification of poorly soluble drugs: Itraconazole as case study.
abstract: Nowadays in animal studies, it is important to comply with the so-called Three Rs rule by replacing or reducing the number of tested animals. Volumetric absorptive microsampling (VAMS) can be used to collect small quantities (10 or 20μL) of whole blood, thereby limiting the amount of animals needed. In this study, a quantitative method was developed and subsequently validated for the poorly soluble drug itraconazole (ITZ) using VAMS and ultra-high performance liquid chromatography (UHPLC) coupled to tandem mass spectrometry (MS). A proof of concept study showed that the optimized method is applicable to test the bioavailability of drug formulations containing ITZ. Using VAMS, smaller blood volumes can be taken per sampling point (10-20μL instead of the conventional 0.2-0.5mL) avoiding the sacrifice of animals. Moreover, the same rats can be used to compare different drug formulations which strengthens the validity of the results. In long-term bioavailability studies, it is necessary to guarantee the stability of the tested drugs supported on VAMS devices. In this study, we show that ITZ was only stable for 24h after collection with VAMS, but for at least two weeks by the storage of extracted samples at -80°C.
authors.affiliation: Laboratory of Pharmaceutical Technology and Biopharmacy, Department of Pharmacy, Centre for Interdisciplinary Research on Medicines (CIRM), University of Liège, Quartier Hopital, Avenue Hippocrate 15, 4000, Liège, Belgium.
authors.forename: Justine
authors.initials: J
authors.lastname: Thiry
authors.affiliation: Laboratory of Pharmaceutical Technology and Biopharmacy, Department of Pharmacy, Centre for Interdisciplinary Research on Medicines (CIRM), University of Liège, Quartier Hopital, Avenue Hippocrate 15, 4000, Liège, Belgium.
authors.forename: Brigitte
authors.initials: B
authors.lastname: Evrard
authors.affiliation: Laboratory for the Analysis of Medicines, CIRM, University of Liège, Quartier Hopital, Avenue Hippocrate 15, 4000, Liège, Belgium.
authors.forename: Gwenaël
authors.initials: G
authors.lastname: Nys
authors.affiliation: Laboratory for the Analysis of Medicines, CIRM, University of Liège, Quartier Hopital, Avenue Hippocrate 15, 4000, Liège, Belgium.
authors.forename: Marianne
authors.initials: M
authors.lastname: Fillet
authors.affiliation: Laboratory for the Analysis of Medicines, CIRM, University of Liège, Quartier Hopital, Avenue Hippocrate 15, 4000, Liège, Belgium. Electronic address: miranda.kok@ulg.ac.be.
authors.forename: Miranda G M
authors.initials: MGM
authors.lastname: Kok
mesh_terms: D000818:Animals; D001682:Biological Availability; D002851:Chromatography, High Pressure Liquid; D004339:Drug Compounding; D006207:Half-Life; D017964:Itraconazole / Q000097:blood* / Q000302:isolation & purification / Q000493:pharmacokinetics; D008297:Male; D051381:Rats; D017208:Rats, Wistar; D012997:Solvents / Q000737:chemistry; D053719:Tandem Mass Spectrometry
chemical_list: D012997:Solvents; D017964:Itraconazole
keywords: Hydroxy-itraconazole; Itraconazole; Stability; UHPLC–MS/MS; Validation; Volumetric absorptive microsampling