Hemin (Fe3+)- and heme (Fe2+)-smectite conjugates as a model of hemoprotein based on spectrophotometry

Hemin (Fe3+) was adsorbed onto synthetic smectite (clay mineral) in acetone to form a hemin-smectite conjugate. The hemin-smectite conjugate became so luble in water to form a transparent colloidal solution with a dark brown c olor. Its absorption spectrum in water showed a sharp Soret band at 398 nm with the molar extinction coefficient as epsilon (398nm) = 11.6 x 10(4) M-1 Cm-1, which is in good agreement with epsilon (398nm) = (12.2 +/- 3) x 10( 4) M-1 cm(-1) of monomeric hematin (1). Hamin (Fe3+)-smectite conjugate had a peroxidase-like activity in the presence of hydrogen peroxide (a hydroge n acceptor) and guaiacol (a hydrogen donor) in aqueous solution and its act ivity was higher than that of hematin. Hemin (Fe3+)-smectite conjugate in w ater was reduced by adding sodium dithionite to form a heme (Fe2+)-smectite conjugate which is also a transparent colloidal solution in water. Its abs orption spectrum in aqueous solution was surprisingly in close agreement wi th that of oxyhemoglobin. Its peak positions of alpha, beta, and Soret band s were located in only a 9-3 nm shift to shorter wavelengths in comparison with those of oxyhemoglobin. Therefore, heme (Fe2+)-smectite conjugate was bound to O-2 to form O-2-heme (Fe2+)-smectite conjugate. The addition of ca rbon monoxide, CO, to O-2-heme (Fe2+)-smectite conjugate caused the formati on of CO-heme (Fe2+)-smectite conjugate with a similar absorption spectrum of carboxyhemoglobin (HbCO) accompanied by shifting 8-10 nm to shorter wave length. Therefore, the transformation of O-2-heme (Fe2+)-smectite conjugate to CO-heme (Fe2+)-smectite conjugate was accompanied by shifting of 7, 4, and 3 nm to shorter wavelengths in the alpha, beta, and Soret bands respect ively, which are similar to the spectral change from oxyhemoglobin to carbo xyhemoglobin. Also the ratio (1:1.6) of the molar extinction coefficient of Soret band of O-2-heme (Fe2+)-smectite conjugate and CO-heme (Fe2+)-smecti te conjugate was surprisingly agreement with ratio (1:1.5) of oxyhemoglobin and carboxyhemoglobin. The phenomenon shown above was unexpectedly found d uring the course of study of bioconjugate of a bioactive substance, hemin ( Fe3+) or heme (Fe2+), and a clay mineral, smectite, in place of the protein of globin in hemoglobin.