imidazolone and 2-formyl-5-(hydroxymethyl)pyrrole-1-norleucine

The glycation of BSA leads to protein/peptide modifications that result in the formation of AGEs. AGEs react with the amino groups of N-terminal amino acid residues, particularly arginine and lysine residues. Enhanced AGE formation exists in the blood and tissues of diabetics, as well as in aging and other disorders. The Identification of AGEs is of great importance. Mass spectrometry has been applied to identify and structurally elucidate AGEs. Here, we report on the identification of AGE- peptides and AGE-precursors based on relative mass changes as a result of specific AGE formation. HPLC-ESIMS, ESI-MS/MS, and the Mascot database were used. The relative mass changes due to the specific type of AGE formation were added to the identified peptides followed by a manual search of the glycated samples, which resulted in the identification of seven peptides for the formation of five AGEs, namely CML, pyrraline, imidazolone A, imidazolone B, and AFGP. Four glycated peptides (FPK, ECCDKPLLEK, IETMR, and HLVDEPQNLIK) were identified in the formation of AGE-precursors.

Topics: Amino Acid Sequence; Glycation End Products, Advanced; Glycolipids; Glycosides; Glycosylation; Imidazoles; Lysine; Models, Biological; Norleucine; Pyrroles; Schiff Bases; Serum Albumin, Bovine; Spectrometry, Mass, Electrospray Ionization

Advanced glycation end products (AGEs) are known to be deposited in the target organ of ageing. In addition, the deposition of AGEs accelerate the process of ageing. We investigated the immunohistochemical localization of AGEs in pinguecula, one of the ocular changes related with ageing process.. Surgical specimens of conjunctiva with or without pinguecula were prepared from nine patients, respectively. Immunohistochemical localization of AGEs was investigated using monoclonal antibodies to N(epsilon)-(carboxymethyl)lysine, pentosidine, imidazolone, and pyrraline.. Moderate to strong immunoreactivities to AGEs were detected in the subepithelial amorphous deposits of all the surgical specimens with pinguecula. In contrast, no or weak immunoreactivities to AGEs were detected in the surgical specimens without pinguecula.. Pinguecula is an aggregation of AGEs-modified proteins. The presence of pinguecula would be an index of local irradiation of ultraviolet rays and decreased antioxidant activities.

Topics: Adult; Aged; Aging; Antibodies, Monoclonal; Arginine; Conjunctival Diseases; Female; Glycation End Products, Advanced; Humans; Imidazoles; Immunoenzyme Techniques; Lysine; Male; Middle Aged; Norleucine; Pyrroles

Hemodialysis (HD) patients are frequently in a state of increased oxidative stress, and hyperglycemia appears to be a major factor. We recently found that oxidized human serum albumin (HSA) is a reliable marker of oxidative stress in HD patients. However, the issue of whether oxidized HSA is associated with the progression of oxidative stress in HD patients with or without diabetes is not clear. In the present study, we examined the effect of a qualitative modification of HSA in HD patients with or without diabetes. Blood samples from 10 HD patients with diabetes, 7 HD patients without diabetes, and 10 healthy age-matched controls were examined. The increase in plasma protein carbonyl content and advanced glycation endproducts (AGEs) in HD patients was largely due to an increase in the levels of oxidized HSA. Furthermore, these increases were greatest in HD patients with diabetes. Purified HSA from HD patients (non-DM-HSA) was carbonylated and AGE-modified. The amount of modified HSA was the highest in HD patients with diabetes (DM-HSA). Carboxy methyl lysine (CML)-modified HSA triggered a neutrophil respiratory burst, and this activity was closely correlated with the increase in the CML/HSA ratio. These findings indicate that uremia plays an important role in the progression of oxidative stress in HD patients via an increase in CML-modified HSA. They also indicate that diabetic complications further exacerbate the progression of oxidative stress by further increasing the amount of these modified HSA molecules.

Topics: Adult; Aged; Aged, 80 and over; Arginine; Blood Proteins; Case-Control Studies; Diabetes Mellitus; Female; Glycation End Products, Advanced; Humans; Imidazoles; Kidney Failure, Chronic; Lysine; Male; Middle Aged; Neutrophils; Norleucine; Oxidative Stress; Protein Carbonylation; Pyrroles; Renal Dialysis; Respiratory Burst; Serum Albumin

The levels of plasma 3-deoxyglucosone (3-DG) increase under hyperglycemic conditions and are associated with the pathogenesis of diabetic complications because of the high reactivity of 3-DG with proteins to form advanced glycation end products (AGE). To investigate potential markers for 3-DG-mediated protein modification in vitro and in vivo, we compared the yield of several 3-DG-derived AGE structures by immunochemical analysis and HPLC and measured their localization in human atherosclerotic lesions. When BSA was incubated with 3-DG at 37 degrees C for up to 4 wk, the amounts of N(epsilon)-(carboxymethyl)lysine (CML) and 3-DG-imidazolone steeply increased with incubation time, whereas the levels of pyrraline and pentosidine increased slightly by day 28. In contrast, significant amounts of pyrraline and pentosidine were also observed when BSA was incubated with 3-DG at 60 degrees C to enhance AGE-formation. In atherosclerotic lesions, CML and 3-DG-imidazolone were found intracellularly in the cytoplasm of most foam cells and extracellularly in the atheromatous core. A weak-positive immunoreaction with pyrraline was found in the extracellular matrix and a few foam cells in aortic intima with atherosclerotic lesions. Our results provide the first evidence that CML and 3-DG-imidazolone are major AGE structures in 3-DG-modified proteins, and that 3-DG-imidazolone provides a better marker for protein modification by 3-DG than pyrraline.

Topics: Adult; Aged; Antibodies, Monoclonal; Aorta; Arginine; Arteriosclerosis; Binding, Competitive; Chromatography; Chromatography, High Pressure Liquid; Cytoplasm; Deoxyglucose; Dose-Response Relationship, Drug; Enzyme-Linked Immunosorbent Assay; Female; Foam Cells; Glycation End Products, Advanced; Humans; Imidazoles; Immunochemistry; Immunohistochemistry; Lysine; Male; Middle Aged; Models, Biological; Models, Chemical; Norleucine; Pyrroles; Temperature; Time Factors