N-terminal adducts of bovine hemoglobin with glutaraldehyde in a hemoglobin-based oxygen carrier.
Hemoglobin-based oxygen carriers (HBOCs) are being developed for the medical field, but because they could increase an athlete's performance, they are misapplied for doping purposes. We previously presented a screening method to detect Oxyglobin (Biopure Corp.) and PolyHeme (Northfield Laboratories Inc.) in serum samples using total acid hydrolysis followed by electrospray mass spectrometry analyses. An alternative mass spectrometric method involving enzymatic hydrolysis is here presented. Digestion of Oxyglobin by endoproteinase Glu-C and LC/MS analyses of the mixture allowed the detection of unique peptidic fragments in comparison with a bovine hemoglobin digest. Tandem mass spectrometry experiments of these peptide ions were performed, and two specific species were actually identified as the N-terminal enzymatic fragment of the beta chain carrying two different modifications. Sequential MS3 experiments using an ion trap mass spectrometer permitted us to locate the chemical modification by the glutaraldehyde on the NH2-terminal group and to propose a structure for the modified peptides. In another set of experiments, screening of these two diagnostic ions into Oxyglobin-spiked serums using precursor ion scan mode in a triple quadrupole instrument allowed the detection of this HBOC with a detection limit of 2 g L(-1).
Gasthuys
Maryline
M
UMR CNRS 7613, University Pierre et Marie Curie, Boite 45, 4 place Jussieu 75252 Paris Cedex 05, France.
Alves
Sandra
S
Tabet
Jean-Claude
JC
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D002853:Chromatography, Liquid; D005976:Glutaral / Q000737:chemistry*; D006454:Hemoglobins / Q000737:chemistry* / Q000378:metabolism; D006868:Hydrolysis; D007477:Ions; D008969:Molecular Sequence Data; D010100:Oxygen / Q000737:chemistry* / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis* / Q000378:metabolism; D012697:Serine Endopeptidases / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization
D006454:Hemoglobins; D007477:Ions; D010446:Peptide Fragments; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D010100:Oxygen; D005976:Glutaral