title: Monitoring tyrosinaemia type I: Blood spot test for nitisinone (NTBC).
abstract: BACKGROUND
  Quantification of nitisinone, 2-(nitro-4-trifluoromethylbenzoyl)1,3-cyclohexanedione (NTBC) has been repeatedly described. Nevertheless monitoring of NTBC has not yet become part of routine therapy surveillance in tyrosinaemia type I (OMIM 276700). We developed a blood spot test to facilitate collection and transport of samples. Furthermore, the test material can be used for determination of other parameters like tyrosine and succinylacetone.
  
  
  METHODS
  For quantification of NTBC in blood spots filter paper discs of 3.2mm diameter were extracted with 150 μL methanol containing mesotrione as internal standard (IS). Analysis was done by UPLC-MS/MS on a Xevo mass spectrometer (ESI+), (MRM). Parent ions were 330.05 for NTBC and 340.05 for IS, daughter ions were m/z 217.95 and m/z 125.95 for NTBC, and m/z 227.95 and m/z 103.95 for IS.
  
  
  RESULTS
  The calibration curve for NTBC in blood spots was linear from 0.1 μmol/L to 100 μmol/L. Recovery exceeded 73.1%, CV intraday and interday were below 9.6%. Instrumental run time was 2.5 min. Sensitivity of the method was 0.1 μmol/L. NTBC concentrations in plasma were higher than in blood spots by a factor of 1.56 ± 0.13.
  
  
  CONCLUSIONS
  As demonstrated in patients with tyrosinaemia type I quantification of NTBC by UPLC-MS/MS in blood spots is feasible and gives valuable information for monitoring NTBC treatment.
authors.affiliation: Screening-Laboratory Hannover, Hannover, Germany. j.sander@metabscreen.de
authors.forename: Johannes
authors.initials: J
authors.lastname: Sander
authors.forename: Nils
authors.initials: N
authors.lastname: Janzen
authors.forename: Michael
authors.initials: M
authors.lastname: Terhardt
authors.forename: Stefanie
authors.initials: S
authors.lastname: Sander
authors.forename: Gülden
authors.initials: G
authors.lastname: Gökcay
authors.forename: Mübeccel
authors.initials: M
authors.lastname: Demirkol
authors.forename: Isıl
authors.initials: I
authors.lastname: Ozer
authors.forename: Michael
authors.initials: M
authors.lastname: Peter
authors.forename: Anibh M
authors.initials: AM
authors.lastname: Das
mesh_terms: D001774:Blood Chemical Analysis / Q000379:methods*; D003512:Cyclohexanones / Q000097:blood* / Q000302:isolation & purification / Q000627:therapeutic use; D005500:Follow-Up Studies; D006801:Humans; D008297:Male; D009579:Nitrobenzoates / Q000097:blood* / Q000302:isolation & purification / Q000627:therapeutic use; D013048:Specimen Handling / Q000379:methods*; D016896:Treatment Outcome; D020176:Tyrosinemias / Q000097:blood* / Q000188:drug therapy
chemical_list: D003512:Cyclohexanones; D009579:Nitrobenzoates; C077073:nitisinone