PEROXIDATION OF ERYTHROCYTES - FTIR SPECTROSCOPY STUDIES OF EXTRACTEDLIPIDS, ISOLATED MEMBRANES, AND INTACT-CELLS

Fourier transform infrared (FTIR) spectroscopy was used to directly mo nitor peroxidative damage to membrane phospholipid acyl chains in eryt hrocyte membranes. Samples were suspended in a mixed (H2O)-H-2/H2O buf fer system, thereby producing a ''spectral window'' in the C-H stretch ing region of the infrared spectrum. A decrease in the number of acyl chain C=C bonds upon erythrocyte peroxidation was quantitated directly from the spectra of isolated membranes. Second-derivative spectroscop y permitted the conformationally sensitive membrane acyl chain methyle ne stretching modes to be separated from the protein (mostly hemoglobi n) vibrations that dominate the spectra of intact cells. The sensitivi ty of these modes in erythrocytes was then determined in a series of t hermotropic experiments. The effect of peroxidation upon the membrane acyl chain conformational order was monitored in isolated membranes an d intact cells. No change in conformational order was detected upon pe roxidation in intact cell and ghost spectra. In contrast, experiments with pure unsaturated phospholipids demonstrated that decreasing the C =C bond population results in increased conformational order. The find ing of identical results for peroxidation versus control samples in is olated membranes (''ghosts'') validates the results observed for intac t cells. FTIR technology permits the direct monitoring of conformation al order in the acyl chains of intact cells. (C) 1995 John Wiley & Son s, Inc.