title: Dried blood spot assay for the quantification of phenytoin using Liquid Chromatography-Mass Spectrometry.
abstract: Phenytoin (PHT) is one of the most commonly used anticonvulsant drugs for the treatment of epilepsy and bipolar disorders. The large amount of plasma required by conventional methods for drug quantification makes mass spectrometry combined with dried blood spot (DBS) sampling crucial for pediatric patients where therapeutic drug monitoring or pharmacokinetic studies may be difficult to realize. DBS represents a new convenient sampling support requiring minimally invasive blood drawing and providing long-term stability of samples and less expensive shipment and storage. The aim of this study was to develop a LC-MS/MS method for the quantification of PHT on DBS. This analytical method was validated and gave good linearity (r(2)=0.999) in the range of 0-100mg/l. LOQ and LOD were 1.0mg/l and 0.3mg/l, respectively. The drug extraction from paper was performed in a few minutes using a mixture composed of organic solvent for 80%. The recovery ranged from 85 to 90%; PHT in DBS showed to be stable at different storage temperatures for one month. A good correlation was also obtained between PHT plasma and DBS concentrations. This method is both precise and accurate and appears to be particularly suitable to monitor treatment with a simple and convenient sample collection procedure.
authors.affiliation: Newborn Screening, Biochemistry and Pharmacology Laboratories, Department of Neurosciences, Meyer University Children's Hospital, Florence, Italy. Electronic address: f.villanelli@meyer.it.
authors.forename: Fabio
authors.initials: F
authors.lastname: Villanelli
authors.affiliation: Newborn Screening, Biochemistry and Pharmacology Laboratories, Department of Neurosciences, Meyer University Children's Hospital, Florence, Italy. Electronic address: e.giocaliere@meyer.it.
authors.forename: Elisa
authors.initials: E
authors.lastname: Giocaliere
authors.affiliation: Newborn Screening, Biochemistry and Pharmacology Laboratories, Department of Neurosciences, Meyer University Children's Hospital, Florence, Italy. Electronic address: s.malvagia@meyer.it.
authors.forename: Sabrina
authors.initials: S
authors.lastname: Malvagia
authors.affiliation: Pediatric Neurology Unit, Meyer University Children's Hospital, Florence, Italy. Electronic address: a.rosati@meyer.it.
authors.forename: Anna
authors.initials: A
authors.lastname: Rosati
authors.affiliation: Newborn Screening, Biochemistry and Pharmacology Laboratories, Department of Neurosciences, Meyer University Children's Hospital, Florence, Italy. Electronic address: giuliaforni1984@libero.it.
authors.forename: Giulia
authors.initials: G
authors.lastname: Forni
authors.affiliation: Newborn Screening, Biochemistry and Pharmacology Laboratories, Department of Neurosciences, Meyer University Children's Hospital, Florence, Italy. Electronic address: s.funghini@meyer.it.
authors.forename: Silvia
authors.initials: S
authors.lastname: Funghini
authors.affiliation: Pharmaceutical Chemistry Department Faculty of Pharmaceutical Sciences and Pharmaceutical Industries, Future University in Egypt, Cairo, Egypt. Electronic address: engyshoukry@fue.edu.eg.
authors.forename: Engy
authors.initials: E
authors.lastname: Shokry
authors.affiliation: Newborn Screening, Biochemistry and Pharmacology Laboratories, Department of Neurosciences, Meyer University Children's Hospital, Florence, Italy. Electronic address: daniela.ombrone@libero.it.
authors.forename: Daniela
authors.initials: D
authors.lastname: Ombrone
authors.affiliation: Newborn Screening, Biochemistry and Pharmacology Laboratories, Department of Neurosciences, Meyer University Children's Hospital, Florence, Italy. Electronic address: m.dellabona@meyer.it.
authors.forename: Maria Luisa
authors.initials: ML
authors.lastname: Della Bona
authors.affiliation: Newborn Screening, Biochemistry and Pharmacology Laboratories, Department of Neurosciences, Meyer University Children's Hospital, Florence, Italy; Department of Neuroscience, Pharmacology and Child Health, University of Florence, Italy; Pediatric Neurology Unit, Meyer University Children's Hospital, Florence, Italy. Electronic address: r.guerrini@meyer.it.
authors.forename: Renzo
authors.initials: R
authors.lastname: Guerrini
authors.affiliation: Newborn Screening, Biochemistry and Pharmacology Laboratories, Department of Neurosciences, Meyer University Children's Hospital, Florence, Italy; Department of Neuroscience, Pharmacology and Child Health, University of Florence, Italy. Electronic address: g.lamarca@meyer.it.
authors.forename: Giancarlo
authors.initials: G
authors.lastname: la Marca
mesh_terms: D002138:Calibration; D002853:Chromatography, Liquid / Q000379:methods*; D059788:Dried Blood Spot Testing / Q000379:methods*; D016903:Drug Monitoring / Q000379:methods*; D004355:Drug Stability; D006801:Humans; D057230:Limit of Detection; D016014:Linear Models; D010672:Phenytoin / Q000097:blood*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D059629:Signal-To-Noise Ratio; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D010672:Phenytoin
keywords: Anticonvulsant drug; Dried blood spot; Epilepsy; Phenytoin; Tandem mass spectrometry; Therapeutic drug monitoring