title: Improved tandem mass spectrometry (MS/MS) derivatized method for the detection of tyrosinemia type I, amino acids and acylcarnitine disorders using a single extraction process.
abstract: BACKGROUND
  Succinylacetone (SUAC), a specific marker for tyrosinemia type I (Tyr I) cannot be detected by the routine LC-MS/MS screening of amino acids (AA) and acylcarnitines (AC) in newborns. The current derivatized methods require double extraction of newborn dried blood spots (DBS); one for AA and AC and the second for SUAC from the blood spot left after the first extraction. We have developed a method in which AA, AC and SUAC are extracted in a single extraction resulting in significant reduction in labor and assay time.
  
  
  METHODS
  The 3.2 mm DBS were extracted by incubating at 45 °C for 45 min with 100 μl of acetonitrile (ACN)-water-formic acid mixture containing hydrazine and stable-isotope labeled internal standards of AA, AC and SUAC. The extract was derivatized with n-butanolic-HCl and analyzed by LC-MS/MS.
  
  
  RESULTS
  The average inter-assay CVs for, AA, AC and SUAC were 10.1, 10.8 and 7.1% respectively. The extraction of analytes with ACN-water mixture showed no significant difference in their recovery compared to commonly used solvent MeOH. The concentration of hydrazine had considerable impact on SUAC extraction.
  
  
  CONCLUSIONS
  We developed a new MS/MS derivatized method to detect AA/AC/SUAC in a single extraction process for screening Tyr I along with disorders of AA and AC.
authors.affiliation: California Department of Public Health, Genetic Disease Laboratory Branch, 850 Marina Bay Pkwy, Richmond, CA 94804, United States.
authors.forename: Kuldeep S
authors.initials: KS
authors.lastname: Dhillon
authors.forename: Ajit S
authors.initials: AS
authors.lastname: Bhandal
authors.forename: Constantino P
authors.initials: CP
authors.lastname: Aznar
authors.forename: Fred W
authors.initials: FW
authors.lastname: Lorey
authors.forename: Partha
authors.initials: P
authors.lastname: Neogi
mesh_terms: D000596:Amino Acids / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification*; D002331:Carnitine / Q000031:analogs & derivatives* / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification; D005591:Chemical Fractionation / Q000379:methods*; D003362:Cost-Benefit Analysis; D006537:Heptanoates / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification; D006801:Humans; D006835:Hydrazones / Q000737:chemistry; D007231:Infant, Newborn; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013997:Time Factors; D020176:Tyrosinemias / Q000175:diagnosis*
chemical_list: D000596:Amino Acids; D006537:Heptanoates; D006835:Hydrazones; C116917:acylcarnitine; C020804:succinylacetone; D002331:Carnitine