title: The measurement of MeIQx adducts with mouse haemoglobin in vitro and in vivo: implications for human dosimetry.
abstract: We have investigated covalent binding of radiolabelled [14C]2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) to mouse haemoglobin in vitro and in vivo. Furthermore, we report the development of a capillary column gas chromatography negative ion mass spectrometry (GC-MS) assay capable of detecting MeIQx liberated from haemoglobin after acid or base hydrolysis. Following microsomal activation, the amount of radiolabelled material associated with haemoglobin in vitro increased with incubation time to 0.67 +/- 0.15 nmol/mg haemoglobin at 2 h (initial concentration 0.47 mM [14C]MeIQx, mean +/- SD, n = 6). Hydrolysis of these samples with acid revealed that 47-60% of the radiolabelled material covalently bound to haemoglobin was acid labile. Of this, 7.2-9.8% was recovered as MeIQx as determined by GC-MS. This liberated fraction should reflect the amount of sulphinic acid amide present which is formed when N-hydroxy-MeIQx reacts with sulphydryl-containing amino acids present in haemoglobin. In vivo, no radiolabelled material bound to haemoglobin could be detected in animals treated with the lowest dose of MeIQx (0.2 mg/kg). At higher doses, there was a dose-dependent increase in the covalent binding of radiolabel to haemoglobin (2.0-200 mg/kg). However, the GC-MS assay for hydrolysable adducts of MeIQx yielded detectable quantities of MeIQx (32.2 +/- 17.5 fmol MeIQx/mg haemoglobin) only at the highest dose used. Application of the GC-MS assay to human haemoglobin samples showed that acid-labile adducts of MeIQx, if present, were below the limit of detection of the assay. These results show that levels of sulphinamide adducts of the dietary aromatic amine MeIQx, with haemoglobin, are very low and the implications for future human dosimetry of this carcinogen are discussed.
authors.affiliation: Department of Clinical Pharmacology, Royal Postgraduate Medical School, London, UK.
authors.forename: A M
authors.initials: AM
authors.lastname: Lynch
authors.forename: S
authors.initials: S
authors.lastname: Murray
authors.forename: A R
authors.initials: AR
authors.lastname: Boobis
authors.forename: D S
authors.initials: DS
authors.lastname: Davies
authors.forename: N J
authors.initials: NJ
authors.lastname: Gooderham
mesh_terms: D000818:Animals; D002250:Carbon Radioisotopes; D004784:Environmental Monitoring / Q000379:methods; D008401:Gas Chromatography-Mass Spectrometry; D006454:Hemoglobins / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D066298:In Vitro Techniques; D051379:Mice; D008861:Microsomes / Q000378:metabolism; D009153:Mutagens / Q000378:metabolism*; D011804:Quinolines / Q000378:metabolism; D011810:Quinoxalines / Q000378:metabolism*
chemical_list: D002250:Carbon Radioisotopes; D006454:Hemoglobins; D009153:Mutagens; D011804:Quinolines; D011810:Quinoxalines; C029216:2-amino-3-methylimidazo(4,5-f)quinoline; C036990:2-amino-3,8-dimethylimidazo(4,5-f)quinoxaline