The active center of methemoglobin Hb(H2O) investigated by Mössbauer and susceptibility experiments.

A fast freezing technique using liquid propane was used to obtain frozen solutions of methemoglobin at pH 7. With this method it was possible to eliminate largely the presence of a low spin component which is usually found in slowly frozen solutions but not present in the sample at room temperature. The magnetic susceptibility and Mössbauer spectra of Hb(H2O) in the temperature range 4.2 K less than or equal to T less than or equal to 250 K have been measured. The data of the high spin compound of Hb(H2O) were evaluated with a Hamiltonian comtaining the Coulomb repulsion of the five 3d-electrons of the Fe3+ ion, a crystal electric field of C2V symmetry and the spin orbit coupling. The term describing the crystal electric field depends on five energy parameters epsilon1, epsilon2, epsilon3, D, and E which are determined by least squares fits to the experimental data. The most relevant parameters epsilon 2 and epsilon 3 which equal the energies of the antibonding single 3d-electron orbitals 3dz2 and 3dx2-y2 with respect to the 3dxy orbital are compared with earlier results of these energies epsilon 2 and epsilon 3 of the high spin compound of Mb(H2O). From this comparison conclusions regarding the different spatial arrangements of Fe3+ in Hb(H2O) and Mb(H2O) are drawn.