Studies on the mechanism of formation of 4-hydroxynonenal during microsomal lipid peroxidation.
The mechanism of the formation of 4-hydroxynonenal through the NADPH-linked microsomal lipid peroxidation was investigated. The results were as follows: 4-hydroxynonenal arises exclusively from arachidonic acid contained in the polar phospholipids, neither arachidonic acid of the neutral lipids nor linoleic acid of the polar or neutral lipids are substrates for 4-hydroxynonenal generation. This finding results from the estimation of the specific radioactivity of 4-hydroxynonenal produced by microsomes prelabelled in vivo with [U-14C]arachidonic acid. Phospholipid-bound 15-hydroperoxyarachidonic acid would have the structural requirements needed for 4-hydroxynonenal (CH3-(CH2)4-CH(OH)-CH=CH-CHO). Microsomes supplemented with 15-hydroperoxyarachidonic acid and NADPH, ADP/iron converted only minimal amounts (0.6 mol%) of 15-hydroperoxyarachidonic acid into 4-hydroxynonenal; similarly, 15-hydroperoxyarachidonic acid incubated at pH 7.4 in the presence of ascorbate/iron yielded only small amounts of 4-hydroxynonenal with a rate orders of magnitude below that observed with microsomes. Phospholipid-bound 15-hydroperoxyarachidonic acid is therefore not a likely intermediate in the reaction pathway leading to 4-hydroxynonenal. The rate of 4-hydroxynonenal formation is highest during the very initial phase of its formation and the onset does not show a lag phase, suggesting a transient intermediate predominantly formed during the early phase of microsomal lipid peroxidation. After 60 min of incubation, 204 nmol polyunsaturated fatty acids (20 nmol 18:2, 143 nmol 20:4, 41 nmol 22:6) were lost per mg microsomal protein and the incubation mixture contained 206 nmol lipid peroxides, 71.6 nmol malonic dialdehyde and 4.6 nmol 4-hydroxynonenal per mg protein. Under artificial conditions (pH 1.0, ascorbate/iron, 20 h of incubation) not comparable to the microsomal peroxidation system, 15-hydroperoxyarachidonic acid can be decomposed in good yields (15 mol%) into 4-hydroxynonenal. Autoxidation of arachidonic acid in the presence of ascorbate/iron gave after 25 h of incubation 2.8 mol% (pH 7.4) and 1.5 mol% (pH 1.0) 4-hydroxynonenal. The most remarkable difference between the non-enzymic system and the enzymic microsomal system is that the latter forms 4-hydroxynonenal at a much higher rate.
Esterbauer
H
H
Benedetti
A
A
Lang
J
J
Fulceri
R
R
Fauler
G
G
Comporti
M
M
D000447:Aldehydes / Q000096:biosynthesis*; D000818:Animals; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000378:metabolism; D005296:Ferrous Compounds / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D015289:Leukotrienes; D008054:Lipid Peroxides / Q000378:metabolism*; D008297:Male; D008315:Malondialdehyde / Q000378:metabolism; D008862:Microsomes, Liver / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D013997:Time Factors
D000447:Aldehydes; D001095:Arachidonic Acids; D005296:Ferrous Compounds; D015289:Leukotrienes; D008054:Lipid Peroxides; D016718:Arachidonic Acid; D008315:Malondialdehyde; D009249:NADP; C025086:15-hydroperoxy-5,8,11,13-eicosatetraenoic acid; C027576:4-hydroxy-2-nonenal