Lactoperoxidase (LPO) is the enzyme of first-line defense in lung mucous, milk and saliva; oxidizing potential invading bacteria and viruses with peroxide and halide/thiocyanide substrates.The heme is covalently bound by ester groups to heme methylenes at positions 1 and 5, so is both functionally and structurally related to intracellular, mammalian peroxidases.LPO was isolated from cow′s milk, concentrated to over 2 mM heme, reduced with dithionite under CO atm.FTIR spectra were collected at 1 cm-1 resolution in a 0.2 mm ZnS, thermostated cell at 25C, using D2O, 50 mM buffer, 1 mM EDTA from pD 3.0 to 8.8, covering the range of highest enzymic activities.Two CO stretch absorbances were observed in all buffers, with varying area ratios at 1942 and 1955 cm-1 maxima.The spectral bands were separated using a curve resolving program fitting to the Voight approximation; the band widths at half peak maximum were calculated between 11 and 13 cm-1.The stretch maxima are significantly higher energy than observed for plant peroxidases but the peak widths correlate to the Fe(III)/(II) reduction potential at pH 7, generality.MO calculations of a model heme active site address heme-asymmetry, isotopic shifts of vibrational bands and the free electron character of porphyrin electron states.