Recent studies suggest that advanced glycation endproducts play an important role in cardiovascular complications of ageing, diabetes and end-stage renal failure. Since highly elevated levels of advanced glycation endproducts are present in serum of patients on maintenance haemodialysis, an accurate and rapid assay for their determination would be useful. This would be particularly valuable for monitoring the removal of advanced glycation endproducts by novel dialysis membranes, as well as the effect of new drugs for the inhibition of their formation.Measurement of advanced glycation endproducts in serum was performed by two competitive ELISAs, using a monoclonal antibody directed against imidazolone, an advanced glycation endproduct formed by the reaction of arginine with 3-deoxyglucosone, and a polyclonal antibody directed against keyhole limpet haemocyanin-advanced glycation endproduct, as well as by quantitative fluorescence spectroscopy.Each of the assays showed significant differences between the controls and the maintenance haemodialysis patients. Advanced glycation endproduct levels determined by each of the ELISAs correlated with total and protein-bound fluorescence, but not with each other, suggesting a variable distribution of advanced glycation endproducts on serum proteins among the maintenance haemodialysis patients.
Three oxidized lipid/amino acid reaction (OLAARP): (1-methyl-4-pentyl-1,4-dihydropyridine-3,5-dicarbaldehyde, 1-(5′-amino-1′-carboxypentyl)pyrrole, and N-(carbobenzyloxy)-1(3)-(1′-(formyl(methyl)-hexyl)-L-histidine dehydrate), and two browned proteins (the monomer and the dimmer produced in the reaction between BSA and 4,5(E)-epoxy-2-E)-hepatenal) were prepared and tested for antioxidative activity in a microsomal system in order to investigate the antioxidative function of OLAARP and non-enzymatically browned proteins in biological systems. The microsomal sytem consisted fo freshly prepared trout muscle microsomes, which were oxidized with Cu2+, Fe3+/ascorbate, or Cu2+/H2O2 at 37 °C and in the presence of the compound to be tested as antioxidant. The three OLAARP (tested at 50 μM) and the two browned proteins (tested at 40 μg/mL) efficiently protected against lipid peroxidation, assessed by the formation of thiobarbituric acid-reactive substances, and protein damage, determined by amino acid analysis. These results suggest that the formation of non-enzymatically browned proteins by reaction with lipid oxidation products may constitute an antioxidative defense mechanism, which could play a significant role in vivo.
The objective was to study the heat-induced reaction of each of the monosaccharides glucose, fructose and galactose with casein at 120°C and to build a kinetic model for the reaction. In addition to participating in the Maillard reaction, glucose isomerized into fructose, fructose into glucose and galactose into tagatose, while no mannose, psicose or talose were detected. Furthermore, the sugars degraded into formic acid and other, unidentified, fragments. The reaction products of the Maillard reaction were the corresponding Amadori products of glucose or galactose, but in the case of fructose no Amadori or Heyns product could be detected. The Amadori product was further degraded into formic acid and acetic acid (plus unidentified fragments). Despite the absence of an Amadori or Heyns product in fructose-casein samples, lysine loss was comparable to that of glucose-casein and galactose-casein. Lysine loss was much greater than the concentration of Amadori product, which means that the Amadori product was rather quickly degraded again. Based on these results, a kinetic model was built that could quantitatively predict the behaviour of monosaccharides in the Maillard reaction as well as in the simultaneously occurring sugar isomerization/degradation.
Summary Several Amadori compounds were prepared and solution (NMR) and, in some cases, solid-state structures (X-ray) were determined. Several of the compounds studied contain sugar units which exist predominantly in acyclic forms. Such compounds were used as model Amadori compounds and rates of reactivity with respect to C(1)H2 proton exchange, the ability to generate oxygen free radicals, and the type of dicarbonyl intermediates produced were evaluated. The results of the experiments enable direct, although qualitative, correlations to be made between the population of acyclic forms in solutions of Amadori compounds, the rates of their reversible enolization, and their reactivity in redox and degradation reactions.
3-Deoxyglucosone (3DG) has been reported to be generated from Amadori compounds by non-oxidative reaction in vitro and in vivo . Several non-fluorescent imidazolone compounds were identified as arginine adducts of 3DG. Using a 3DG and N α -benzoylarginine amide (BzArgNH 2 ) reaction system, 2-( N α -benzoyl- N Δ -ornithylamide)-5-(2,3,4,-trihydroxybutyl)-2-imidazoline-4-one (S12), was identified as an intermediate product. A minor product formed from two molecules of 3DG and one molecule of BzArgNH 2 was identified as 2-( N α -benzoyl- N Δ -ornithylamide)-5,6a-di(2,3,4,-trihydroxybutyl)-5,6-dihydioxydehy-drofuro[2,3- d )imidazole (S11). We speculate that S11 is formed by the addition of 3DG to compound S12. S12 was also identified in a 3DG-insulin reaction system by ESI-MS. In further studies, a fluorescent product was purified from a N α -acetyllysine-3DG reaction system and identified as 6,8-di- N α -acetyllysyl-3,3a,8,8a-tetrahydro-3a-hydroxy-2-(1,2-dihydroxyethyl)-5-hydroxymethyl-2 H -furo[3′,2: 4,5]pyrrolo-[2,3- c ]-pyridinium ( N α -acetyllysyl-pyrropyridine) having fluorescence at Ex max : 376nm and Em max : 450nm. Quantification of lysyl-pyrropyridine in glycated lysozyme revealed a significant increase in a 3DG-lysozyme system (74.3mmol/mol lysozyme) when compared with a glucose-lysozyme system (6.3mmol/mol lysozyme) incubated at 50°C for 7 days.
Summary Fifty years ago, skin painting experiments on mice showed that extracts from heated animal muscle caused cancer. Today we know of more than 20 heterocyclic amines (HAs) formed through the Maillard reaction during the cooking of beef, pork, lamb, chicken and fish muscle. These compounds cause mutations in bacteria, mammalian cells and animals, and cancer in animals. Analytical methods today are sufficient to measure these compounds at concentrations of 0.1 ppb and practical enough for examining numerous foods cooked by different methods, and to varying degrees to doneness. There are widespread exposures to low amounts of HAs in the western diet. In general, frying and grilling food well-done result in the highest yield of HAs. Fast-food hamburgers are very low in total heterocyclic amine content, usually below 1 ppb. In contrast, restaurant prepared meats have about ten-fold higher amounts of HAs. For chicken cooked well-done over an open flame, up to 500 ppb of 2-amino-1-methy-6-phenylimidazo[4,5- b ]pyridine (PhIP) has been found. Human risk is based on exposure and potency. Current results suggest that reducing exposure to HAs is feasible and should lower the human risk to those consuming such foods.
