LIPID OXIDATION IN UNFRACTIONATED SERUM AND PLASMA

In an attempt to develop an assay for the susceptibility of plasma lip ids to oxidation, we have studied the kinetics of copper-induced oxida tion in diluted serum and plasma prepared with different anticoagulant s (heparin, citrate and EDTA) by monitoring the absorbance of oxidatio n-products at several wavelengths. These studies revealed the complex and interrelated effects of the water-soluble antioxidant ascorbic aci d, citrate and chloride ions on the kinetics of copper-induced oxidati on of plasma lipids. Specifically, the onset of oxidation induced by c opper-citrate chelates is only slightly affected by chloride ions and is accelerated upon increasing the copper concentration. By contrast, in the absence of citrate, the lag preceding oxidation in diluted seru m or plasma (but not the maximal rate of oxidation) depends markedly o n the chloride concentration in the diluting medium. In the absence of Cl-, the lag preceding oxidation is a decreasing saturable function o f copper concentration, whereas in a normal phosphate-buffered saline solution (PBS), the lag shows a biphasic dependence on copper concentr ation such that at copper concentrations above 10-30 mu M (depending: on the extent of plasma dilution), increasing the concentration of cop per results in prolongation of the lag. This dependence of copper-indu ced oxidation on the concentration of copper is not observed for dialy zed serum unless ascorbic acid is added. Our interpretation of these r esults is that water-soluble reductants and chloride ions act synergis tically to stabilize Cu+, on the expense of Cu2+. Quenching of free ra dicals by Cu+ may be responsible for the prolongation of the lag at hi gh copper concentrations, with no reduction of the maximal rate of oxi dation. In spite of the complex dependencies described above. spectrop hotometric monitoring of the kinetics of oxidation of plasma lipids, u nder 'optimized conditions' (50-fold diluted serum, in PBS containing 720 mu M sodium citrate and 100 mu M copper), agrees with independent measurements of the consumption of polyunsaturated fatty acids. Hence, the spectroscopic method may become useful for evaluation of the susc eptibility of plasma lipids to oxidation. This possibility, however, h as yet to be elucidated through investigations of the correlation betw een the susceptibility of serum lipids to copper-induced oxidation in vitro and clinical factors of significance. (C) 1998 Published by Else vier Science Ireland Ltd. All rights reserved.