| Maillard or browning reactions between reducing sugars and protein lead to the formation of advanced glycation end-products (AGES) and are thought to contribute to the pathogenesis of diabetic complications. AGE inhibitors such as aminoguanidine (AG) and pyridoxamine (PM) have been shown to limit the formation of AGEs and diabetic nephropathy in animal models of diabetes. PM has also been shown to prevent the chemical modification of protein by advanced lipoxidation end-products (ALEs). We incubated PM with linoleic acid (LA) and arachidonic acid (AA) under oxidizing conditions in order to characterize intermediates trapped by PM during lipid peroxidation reactions. In addition, PM was administered to normal, diabetic, and obese rats to determine if PM trapped intermediates during lipid peroxidation in vivo. We identified twelve PM-adducts formed in incubations of PM with LA and AA; six of these were chemically characterized and subsequently detected in urine of animals treated with PM. The PM-adducts N-formyl-PM (FAPM), N-hexanoyl-PM (HAPM), N-nonanedioyl-PM (NDAPM), N-pentanedioyl-PM (PDAPM), N-pyrrolo-PM (PyPM), and N-(2-formyl)-pyrrolo-PM (FPyPM) were quantified by liquid chromatography-mass spectrometry in urine from PM-treated rats. Levels of PM-adducts in urine of PM-treated, diabetic animals were 5--10 fold higher than in PM-treated, non-diabetic animals. FAPM was the major adduct in urine of non-diabetic and streptozotocin-diabetic Sprague-Dawley rats at 22.9 +/- 0.8 and 132 +/- 12 nmol/24 hr, respectively, whereas PDAPM was the major adduct in urine of Zucker lean, obese, and diabetic-obese rats at 9.1 +/- 0.2, 68.9 +/- 7.0, and 97.0 +/- 4.5 nmol/24 hr, respectively. Treatment with PM also partially corrected dyslipidemia in the diabetic and obese rats. In summary, we have identified specific, lipid-derived products trapped by the AGE/ALE inhibitor PM and excreted in urine as PM amide and amine adducts. We conclude that PM protects against nephropathy in hyperlipidemic and diabetic animals by trapping intermediates formed during lipid peroxidation and by reducing plasma lipid levels. |
