MICELLAR ACCELERATION OF OXYGEN-DEPENDENT REACTIONS AND ITS POTENTIALUSE IN THE STUDY OF HUMAN LOW-DENSITY-LIPOPROTEIN

The reaction rate between superoxide and nitro-blue tetrazolium (NET) is known to be accelerated/catalysed by micellar systems. Previous rep orts suggest that an accelerated rate of NBT reduction by micellar sys tems may be the result of either the binding of organic substrates suc h as NET to the micellar phase giving a more favourable environment fo r superoxide reduction (an orientation effect), or the electrostatic i nteraction between micelles and superoxide. Here we show, using three different superoxide generating systems, that micelles composed of a n umber of different lipids or human low density lipoprotein (LDL) accel erates the apparent reaction between superoxide and NET. Evidence in f avour of an accelerated production of superoxide as opposed to the acc elerated reduction of NET is provided and we propose that the accelera ted production of superoxide is a consequence of increased oxygen solu bility in the lipid, rather than aqueous, phase. This is supported by: 1. The absence of any spectrophotometric changes due to interaction b etween lipid or LDL and reagents used. 2. The ability of micelles comp osed of a number of different fatty substances, including LDL, to acce lerate superoxide generation, assessed by NET reduction. 3. The behavi our of micelles, which appears to be one of substrate rather than cata lyst, during the acceleration of NET reduction. This is confirmed by t he use of a known micellar catalyst, Triton-X100. This suggests that l ipids contribute to the reaction as a substrate rather than a catalyst . 4. The inability of LDL to accelerate NET reduction by potassium sup eroxide, a reaction which is independent of bimolecular oxygen. 5. The inability of LDL to accelerate NET reduction when added after superox ide generation. 6. Studies that show LDL can enhance an NET-independen t monitor of oxidation, namely the transition metal-catalysed oxidatio n of vitamin C. 7. Estimations of the solubility of oxygen in LDL whic h appear to be consistent with reported physical measurements. Further more, we show that LDL modification can alter LDL-mediated micellar ac celeration of superoxide generation. Extensive oxidation of LDL decrea ses micellar acceleration and minimal oxidation enhances it. We sugges t that LDL micellar acceleration might serve as a novel approach to st udying human LDL. (C) 1997 Elsevier Science B.V.