We used Fourier transform infrared spectroscopy (FT-IR) to quantitatively monitor peroxidative damage in model phospholipids and in human erythrocyte membranes. The (3012-\mathrm{cm}^{-1}) band arising from olefinic (\mathrm{C}-\mathrm{H}) stretching modes diminished in intensity during membrane peroxidation as a result of loss of unsaturation in the phospholipid acyl chains. A precise (correlation coefficient (=0.99) ) linear relationship between the number of (\mathbf{C}=\mathbf{C}) bonds in the chains and the fraction of the total (\mathbf{C}-\mathbf{H}) stretching contour occupied by the olefinic C-H stretch was established for model phospholipids. Intact human erythrocytes and isolated ("ghost") membranes were also peroxidized and the loss of (C=C) bonds was demonstrated by FT-IR. GC studies of erythrocyte membranes confirmed that the primary site of peroxidation occurred at phospholipids containing arachidonate acyl chains. Finally, the formation of a new feature at (1260 \mathrm{~cm}^{-1}) in the FT-IR spectrum was observed in both model compounds and erythrocyte membranes undergoing peroxidative damage. The feature corresponds to a (\mathrm{C}-\mathrm{O}) stretching and/or (\mathrm{O}-\mathrm{H}) inplane bending vibration and is suggestive of the formation of these functional groups during peroxidation. FT-IR provides an accurate means for monitoring the effects of peroxidative damage in human erythrocytes. 1994 Academic Press, Inc.