Interactions of methemoglobin and green hemoprotein in chemiluminescent gels

Peroxidation of luminol (5-amino-2,3-dihydrophthalazine-1,4-dione) catalyzed by human green hemoprotein (GHP) and bovine methemoglobin (MetHb) in gels composed of cross-linked bovine serum albumin was examined. The chemiluminescence (CL) was followed with a low-light intensity video camera and imaging system attached to a circularly polarized microwave guide (2450 MHz) for heating the samples from 24 degrees C to 37 degrees C. Steady-state CL was maintained in the gels for 10 min. The intensity of the CL varied with temperature. When combined with MetHb in the same gel, GHP inhibited CL of MetHb from 83.6% to 98.2% over a fiftyfold concentration range of GHP. Although MetHb/GHP combination gels were inhibited, they generated a 6.12-fold CL per degree C change compared to a 0.19-fold per degree C change for MetHb gels and a 0.31-fold per degree C change for GHP gels. The data suggest an interaction between GHP and MetHb that inhibits the CL reaction, is not interfered with by large amounts of albumin, and is partially reversed by heating.