Separation and characterization of the aldehydic products of lipid peroxidation stimulated by ADP-Fe2+ in rat liver microsomes

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Abstract
Methods using TLC and high-pressure liquid chromatography (HPLC) were used to separate the complex variety of substances prossessing a carbonyl function that are produced during lipid peroxidation. The major type of lipid peroxidation studied was the ADP-Fe2+-stimulated peroxidation of rat liver microsomal phospholipids. Preliminary separation of the polar and non-polar products was achieved by TLC; further separation and identification of individual components was performed by HPLC. Estimations were performed on microsomal pellets and the supernatant mixture after incubation of microsomes for 30 min at 37.degree. C. The polar fraction was larger than the non-polar fraction when expressed as nmol of carbonyl groups/g of liver. In the non-polar supernatant fraction the major contributors were n-alkanals (31% of the total), .alpha.-dicarbonyl compounds (22%) and 4-hydroxyalkenals (37%) with the extraction method used. Major individual contributions to the non-polar fraction were propanal, 4-hydroxynoneal, hexanal and oct-2-enal. Other components identified include butanal, pent-2-enal, hex-2-enal, hept-2-enal, 4-hydroxyoctenal and 4-hydroxyundecenal. The polar carbonyl fraction was less complex than the non-polar fraction, although the identities of the individual components have not yet been established. Since these carbonyl compounds do not react significantly in the thiobarbituric acid reaction, which largely demonstrates the presence of malonaldehyde, considerable amounts of biologically reactive carbonyl derivatives may be released in lipid peroxidation and yet not picked up by the thiobarbituric acid reaction.