Quantitative determination of succinylacetone in dried blood spots for newborn screening of tyrosinemia type I

Mark J. Magera, Nishantha D. Gunawardena, Si Houn Hahn, Silvia Tortorelli, Grant A. Mitchell, Stephen I. Goodman, Piero Rinaldo, Dietrich Matern

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53 Scopus citations


Background: Tyrosinemia type I (TYR 1) is a severe disorder causing early death if left untreated. While tyrosine can be determined in dried blood spots (DBS), it is not a specific marker for TYR 1 and most often associated with benign transient tyrosinemia of the newborn. Succinylacetone (SUAC) is a specific marker for TYR 1 but not detectable by routine newborn screening. We developed a new assay that determines SUAC in DBS by liquid-chromatography tandem mass spectrometry (LC-MS/MS). Methods: Whole blood is eluted from a 3/16-in. DBS by an aqueous solution containing deuterium labeled SUAC as internal standard (IS). SUAC and IS are oximated, then extracted, butylated, and analyzed by LC-MS/MS. Quantitation is from SUAC spiked calibrator DBS over the range 0-200 μM using selected reaction monitoring of transitions m/z 212 to 156 and m/z 214 to 140 for SUAC and IS, respectively. Analysis time is 5 min. To assess the effectiveness of a two-tier screening approach for TYR 1 we applied this assay to our newborn screening program over the last 15 months. Results: The intra-assay precision was determined for three different levels of SUAC (5, 20, and 50 μmol/L) and the CV calculated to be 4.7, 2.6, and 3.1%, respectively (n = 5). Inter-assay precision CVs were 12.7, 8.2, and 7.8%, respectively on the same samples. SUAC levels in DBS from 10 confirmed TYR 1 cases not treated with 2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedione (NTBC) were clearly abnormal (16-150 μmol/L; mean: 61 μmol/L; controls: <5 μmol/L). Over a 15-month period, SUAC was determined in newborn screening samples with elevated tyrosine concentrations when applying different cut off values until it was settled at 150 μmol/L. No case of TYR 1 was detected in 124,780 newborns tested. Conclusion: We have developed a new LC-MS/MS based method for the determination of SUAC in DBS. This assay has the potential to significantly reduce the number of false positive results in newborn screening for TYR 1 and can also be used for the laboratory follow up of patients treated for TYR 1.

Original languageEnglish (US)
Pages (from-to)16-21
Number of pages6
JournalMolecular genetics and metabolism
Issue number1
StatePublished - May 2006


  • Dried blood spot
  • Fumarylacetoacetate hydrolase deficiency
  • Newborn screening
  • Succinylacetone
  • Tandem mass spectrometry
  • Tyrosinemia type I

ASJC Scopus subject areas

  • Endocrinology, Diabetes and Metabolism
  • Biochemistry
  • Molecular Biology
  • Genetics
  • Endocrinology


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