Direct Measurement of Lipid Peroxidation in Submitochondrial Particles

The susceptibility of the polyunsaturated fatty acid parinaric acid (cis-PnA) to peroxidative damage with concomitant loss of its fluorescent character can be used to detect lipid peroxidation in a direct and sensitive way. The procedure, originally developed to measure peroxidation in lipid vesicle...

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Bibliographic Details
Published in:	Biochemistry (Easton) Vol. 34; no. 39; pp. 12755 - 12760
Main Authors:	de Hingh, Yvette C. M, Meyer, Johnny, Fischer, Johan C, Berger, Ruud, Smeitink, Jan A. M, Op den Kamp, Jos A. F
Format:	Journal Article
Language:	English
Published:	United States American Chemical Society 03-10-1995
Subjects:	Adenosine Diphosphate - analogs & derivatives Adenosine Diphosphate - metabolism Animals Cattle Lipid Peroxidation Male Mitochondria, Heart - metabolism Mitochondria, Liver - metabolism Mitochondria, Muscle - metabolism NAD - metabolism NADP - metabolism Rats Rats, Wistar Spectrometry, Fluorescence Submitochondrial Particles - metabolism
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Summary:	The susceptibility of the polyunsaturated fatty acid parinaric acid (cis-PnA) to peroxidative damage with concomitant loss of its fluorescent character can be used to detect lipid peroxidation in a direct and sensitive way. The procedure, originally developed to measure peroxidation in lipid vesicles and erythrocyte membranes, has been adapted for the study of submitochondrial particles. Optimal conditions for the concentrations of cis-PnA (0.8 mol %), mitochondrial membrane (100 microM membrane phospholipid), and the radical generating system (50 microM NADH and 10 microM:1 mM Fe(III)-ADP) were established. In the absence of peroxidation inducing compounds, a stable fluorescent signal can be detected. Upon addition of NAD(P)H and ADP-Fe(III), lipid peroxidation starts, and the observed fluorescence decrease is a measure of peroxidation. Both NADH and NADPH were able to induce lipid peroxidation in submitochondrial particles in the presence of an iron chelate. The use of NADH resulted in higher rates of peroxidation compared with NADPH at the same concentration. Whereas the rate of NADH-induced lipid peroxidation was maximal at very low NADH concentrations (2.5 microM) and decreased when the concentration became higher, the NADPH-induced lipid peroxidation reaches saturation at 100 microM. NADH-induced lipid peroxidation in submitochondrial particles from different rat tissues (heart, skeletal muscle, and liver) resulted in a clear difference in peroxidation rates. The highest rates were observed in heart submitochondrial particles, while the lowest rates were obtained in submitochondrial particles derived from liver. Skeletal muscle submitochondrial particles showed intermediate rates of lipid peroxidation.
Bibliography:	istex:C15093DC743DC05F464C889D9136BC1654F00EE2 ark:/67375/TPS-WTPC2TP0-L ObjectType-Article-2 SourceType-Scholarly Journals-1 ObjectType-Feature-1 content type line 23 ObjectType-Article-1 ObjectType-Feature-2
ISSN:	0006-2960 1520-4995
DOI:	10.1021/bi00039a035