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

Clin Chim Acta. 2011 May 12;412(11-12):873-9. doi: 10.1016/j.cca.2010.12.028. Epub 2011 Jan 7.


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.

Conclusion: 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.

MeSH terms

  • Amino Acids / analysis*
  • Amino Acids / chemistry
  • Amino Acids / isolation & purification*
  • Carnitine / analogs & derivatives*
  • Carnitine / analysis
  • Carnitine / chemistry
  • Carnitine / isolation & purification
  • Chemical Fractionation / methods*
  • Cost-Benefit Analysis
  • Heptanoates / analysis
  • Heptanoates / chemistry
  • Heptanoates / isolation & purification
  • Humans
  • Hydrazones / chemistry
  • Infant, Newborn
  • Tandem Mass Spectrometry / methods*
  • Time Factors
  • Tyrosinemias / diagnosis*


  • Amino Acids
  • Heptanoates
  • Hydrazones
  • acylcarnitine
  • succinylacetone
  • Carnitine