The dose-dependent effect of copper-chelating agents on the kinetics of peroxidation of low-density lipoprotein (LDL)

Copper-induced peroxidation of lipoproteins involves continuous production of free radicals via a redox cycle of copper. Formation of Cu(I) during Cu( II)-induced peroxidation of LDL was previously demonstrated by accumulation of the colored complexes of Cu(I) in the presence of one of the Cu(I)-spec ific chelators bathocuproine (BC) or neocuproine (NC). All the studies cond ucted thus far employed high concentrations of these chelators (chelator/Cu (II) > 10). Under these conditions, at low copper concentrations the chelat ors prolonged the lag preceding oxidation, whereas at high copper concentra tions the chelators shortened the lag. In an attempt to gain understanding of these non-monotonic effects, we have studied systematically the peroxidation of LDL (0.1 muM, 50 mug protein/ml ) at varying concentrations of NC or BC over a wide range of concentrations of the chelators and copper. These studies revealed that: (i) At copper concentrations of 5 muM and below, NC prolonged the lag in a monotonic, dose-dependent fashion typical for other complexing agents. Howe ver, unlike with other chelators, the maximal rate of oxidation was only sl ightly reduced (if at all). (ii) At copper concentrations of 15 mM and above, the addition of about 20 muM NC or BC resulted in prolongation of the lag, but this effect became sm aller at higher concentrations of the chelators, and at yet higher concentr ations the lag became much shorter than that observed in the absence of che lators. Throughout the whole range of NC concentrations, the maximal rate o f peroxidation increased monotonically upon increasing the NC concentration . (iii) Unlike in the absence of chelators, the prooxidative effect of copper did not exhibit saturation with respect to copper, up to copper concentrat ions of 30 muM. Based on these results we conclude that the copper-chelates can partition i nto the hydrophobic core of LDL particles and induce peroxidation by formin g free radicals within the core. This may be significant with respect to th e understanding of the possible mechanisms of peroxidation by chelated tran sition metals in vivo.