OXIDATIVE MODIFICATION AND NITRATION OF HUMAN LOW-DENSITY LIPOPROTEINS BY THE REACTION OF HYPOCHLOROUS ACID WITH NITRITE

Hypochlorous acid (HOCl) reacts with nitrite (NO2-) at a molar ratio o f 1:1 yielding an equimolar amount of nitrate. The rate of this reacti on follows the dissociation of hypochlorous acid and decreases with th e increasing of pH from 4 to 10 as assayed by stopped-flow analysis, s uggesting that HOCl, not hypochlorite, is the reactant. The second-ord er rate constant at pH. 7.2, 25 degrees C, was estimated as (7.4 +/- 1 .3) x 10(8) M-1 S-1 a rate considerably higher than that of the Fenton reaction (42 M-1 s(-1)). In human low-density lipoproteins (LDL) the reaction led, to a loss of p-carotene and alpha-tocopherol, The NO2-/H OCl mixture initiated lipid peroxidation in LDL, whereas NO2- or HOCl alone had only little effect, When LDL was added immediately after mix ing of NO2- with HOCl, no loss of antioxidants or accumulation of lipi d peroxidation products was observed, suggesting that a short-lived re active intermediate, previously postulated as nitryl chloride, is the reactive species, The mixture NO2-/ HOCl as well as peroxynitrite led to the formation of S-nitrotyrosine in LDL as assayed using a monoclon al anti-nitrotyrosine antibody. Furthermore, incubation of J774.2 macr ophage-like cells with LDL, pretreated with the NO2-/HOCl mixture, led to increased cellular accumulation of cholesterol. Thus modification of LDL caused by the reaction of nitrite with HOCl contributes to the formation of cholesterol-rich cells, a key feature of the early athero sclerotic lesion. (C) 1997 Academic Press.