title: The metabolism and de-bromination of bromotyrosine in vivo.
abstract: During inflammation, leukocyte-derived eosinophil peroxidase catalyses the formation of hypobromous acid, which can brominate tyrosine residues in proteins to form bromotyrosine. Since eosinophils are involved in the pathogenesis of allergic reactions, such as asthma, urinary bromotyrosine level has been used for the assessment of children with asthma. However, little is known about the metabolism and disposition of bromotyrosine in vivo. The aim of this study was to identify the major urinary metabolites formed during bromotyrosine metabolism and to develop mass spectrometric methods for their quantitation. Deuterium-labeled bromotyrosine was synthesized by deuterium exchange. [D3]bromotyrosine (500 nmole) was injected intraperitoneally into Sprague-Dawley rats and urine was collected for 24h in a metabolic cage. (13)C-labeled derivatives of bromotyrosine and its major urinary metabolite were synthesized and used as internal standards for quantitation. Following solid phase extraction, urine samples were derivatized to the pentafluorobenzyl ester, and analyzed using isotope dilution gas chromatography and negative-ion chemical ionization mass spectrometry. A novel brominated metabolite, 3-bromo-4-hydroxyphenylacetic acid (bromo-HPA), was identified as the major brominated metabolite of bromotyrosine. Bromo-HPA only accounted for 0.43 ± 0.04% of infused [D3]bromotyrosine and 0.12 ± 0.02% of infused [D3]bromotyrosine was excreted in the urine unchanged. However, ~1.3% (6.66 ± 1.33 nmole) of infused [D3]bromotyrosine was excreted in the urine as the de-brominated metabolite, [D3]4-hydroxyphenylacetic acid, which is also a urinary metabolite of tyrosine in mammals. We also tested whether or not iodotyrosine dehalogenase can catalyse de-bromination of bromotyrosine and showed that iodotyrosine dehalogenase is able to de-brominate free bromotyrosine in vitro. We identified bromo-HPA as the main brominated urinary metabolite of bromotyrosine in rats. However, de-halogenation of bromotyrosine is the major metabolic pathway to eliminate free brominated tyrosine in vivo.
authors.affiliation: Division of Medicine, Royal Free Campus, University College London (UCL), Rowland Hill Street, NW3 2PF London, UK. Electronic address: a.r.mani@ucl.ac.uk.
authors.forename: Ali R
authors.initials: AR
authors.lastname: Mani
authors.affiliation: Thyroid Molecular Laboratory, Institute for Medical and Molecular Genetics (INGEMM), La Paz University Hospital, Madrid, Spain.
authors.forename: José C
authors.initials: JC
authors.lastname: Moreno
authors.affiliation: Department of Internal Medicine, Erasmus University Medical Centre, Rotterdam, The Netherlands.
authors.forename: Theo J
authors.initials: TJ
authors.lastname: Visser
authors.affiliation: Division of Medicine, Royal Free Campus, University College London (UCL), Rowland Hill Street, NW3 2PF London, UK.
authors.forename: Kevin P
authors.initials: KP
authors.lastname: Moore
mesh_terms: D000818:Animals; D001966:Bromine / Q000737:chemistry; D003903:Deuterium; D008401:Gas Chromatography-Mass Spectrometry; D008297:Male; D010648:Phenylacetates / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism
chemical_list: D010648:Phenylacetates; C475724:bromotyrosine; C008070:4-hydroxyphenylacetic acid; D014443:Tyrosine; D003903:Deuterium; D001966:Bromine
keywords: 3-Bromo-4-hydroxyphenylacetic acid; 4-Hydroxyphenylacetic acid; Bromotyrosine; De-halogenation; Urine