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

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 degrees C for 45 min with 100 mu l 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. Published by Elsevier B.V.