Identification of NTBC metabolites in urine from patients with hereditary tyrosinemia type 1 using two different mass spectrometric platforms: triple stage quadrupole and LTQ-Orbitrap.
The objective of our work was to identify known and unknown metabolites of the drug NTBC (2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedione) in urine from patients during the treatment of hereditary tyrosinemia type 1 (HT-1) disease, a severe inborn error of tyrosine metabolism. Two different mass spectrometric techniques, a triple stage quadrupole and an LTQ-Orbitrap (Fourier transform mass spectrometry (FTMS)), were used for the identification and the structural elucidation of the detected metabolites. Initially, the mass spectrometric (MS) approach consisted of the precursor ion scan detection of the selected product ions, followed by the corresponding collision-induced dissociation (CID) fragmentation analysis (MS(2)) for the targeted selected reaction monitoring (SRM) mode. Subsequently, accurate and high-resolution full scan and MS/MS measurements were performed on the possible metabolites using the LTQ-Orbitrap. Final confirmation of the identified metabolites was achieved by measuring commercially supplied or laboratory-synthesized standards. Altogether six metabolites, including NTBC itself, were extracted, detected and identified. In addition, two new NTBC metabolites were unambiguously identified as amino acid conjugates, namely glycine-NTBC and beta-alanine-NTBC. These identifications were based on their characteristics of chromatographic retention times, protonated molecular ions, elemental compositions, product ions (using CID and higher-energy C-trap dissociation (HCD) techniques) and synthesized references. The applied MS strategy, based on two different MS platforms (LC/MS/MS and FTMS), allowed the rapid identification analysis of the drug metabolites from human extracts and could be used for pharmaceutical research and drug development.
Herebian
Diran
D
Department of General Pediatrics, University Children's Hospital, Heinrich-Heine University, Düsseldorf, Germany. diran.herebian@med.uni-duesseldorf.de
Lamshöft
Marc
M
Mayatepek
Ertan
E
Spiekerkoetter
Ute
U
D003512:Cyclohexanones / Q000737:chemistry / Q000378:metabolism / Q000627:therapeutic use / Q000652:urine*; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008958:Models, Molecular; D008970:Molecular Weight; D009579:Nitrobenzoates / Q000737:chemistry / Q000378:metabolism / Q000627:therapeutic use / Q000652:urine*; D014050:Toluene / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism / Q000652:urine; D020176:Tyrosinemias / Q000188:drug therapy / Q000652:urine*
D003512:Cyclohexanones; D009579:Nitrobenzoates; D014050:Toluene; C077073:nitisinone