title: Isotope-dilution gas chromatography-mass spectrometry method for the analysis of hydroxyurea.
abstract: BACKGROUND
  Hydroxyurea is used in the treatment of various malignancies and sickle cell disease. There are limited studies on the pharmacokinetics of hydroxyurea, particularly in pediatric patients. An accurate, precise, and sensitive method is needed to support such studies and to monitor therapeutic adherence. We describe a novel gas chromatography-mass spectrometry (GC-MS) method for the determination of hydroxyurea concentration in plasma using stable labeled hydroxyurea C N2 as an internal standard.
  
  
  METHODS
  The method involved an organic extraction followed by the preparation of trimethylsilyl (TMS) derivatives of hydroxyurea for GC-MS selected ion-monitoring analysis. The following mass-to-charge (m/z) ratio ions for silated hydroxyurea and hydroxyurea C N2 were monitored: hydroxyurea-quantitative ion 277, qualifier ions 292 and 249; hydroxyurea C N2-quantitative ion 280, qualifier ion 295. This method was evaluated for reportable range, accuracy, within-run and between-run imprecisions, and limits of quantification.
  
  
  RESULTS
  The reportable range for the method was 0.1-100 mcg/mL. All results were accurate within an allowable error of 15%. Within-run and between-run imprecisions were <15%. Samples were stable for at least 4 hours at room temperature, 2 months at -20°C, and 6 months at -70°C, and after 3 freeze/thaw cycles. Extraction efficiency for 1-, 5-, 10-, and 50-mcg/mL samples averaged 2.2%, 1.8%, 1.6%, and 1.4%, respectively.
  
  
  CONCLUSIONS
  The isotope-dilution GC-MS method for analysis of hydroxyurea described here is accurate, sensitive, precise, and robust. Its characteristics make the method suitable for supporting pharmacokinetic studies and/or clinical therapeutic monitoring.
authors.affiliation: Departments of *Pathology and Laboratory Medicine, and †Pediatrics; and Divisions of ‡Pediatric Pharmacology and Therapeutic Innovation, and §Hematology/Oncology, University of Missouri-Kansas City School of Medicine, the Children's Mercy Hospital.
authors.forename: Uttam
authors.initials: U
authors.lastname: Garg
authors.forename: David
authors.initials: D
authors.lastname: Scott
authors.forename: Clint
authors.initials: C
authors.lastname: Frazee
authors.forename: Gregory
authors.initials: G
authors.lastname: Kearns
authors.forename: Kathleen
authors.initials: K
authors.lastname: Neville
mesh_terms: D000293:Adolescent; D000986:Antisickling Agents / Q000097:blood / Q000493:pharmacokinetics; D002247:Carbon Isotopes / Q000097:blood; D002648:Child; D016903:Drug Monitoring / Q000379:methods*; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D006918:Hydroxyurea / Q000097:blood* / Q000493:pharmacokinetics*; D007201:Indicator Dilution Techniques; D009587:Nitrogen Isotopes / Q000097:blood
chemical_list: D000986:Antisickling Agents; D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D006918:Hydroxyurea