Luminescence during the oxidation of hypoxanthine by xanthine oxidase in the presence of dimethylbiacridylium nitrate.

During the course of some studies on xanthme oxidase it was discovered that when the enzyme was incubated with one of its substrates in the presence of either lO , lO’-dimethyl-9,9’-biacridylium nitrate or 5-amino-2,3-dihydrophthalazine-l , 4-dione(luminol)light was produced in considerably greater intensity than would be expected from the rate of production of Hz02 and the known chemiluminescence of these compounds in the presence of HzOz. The emission of light requires the presence of free 02 as is usual in chemiluminescence or bioluminescence. These reactions have been investigated further and there are detailed below several experiments with xanthine oxidase and DBA++nitrate.’ The kinetics of light production with DBA++ differ from those with luminol and from the known kinetics of oxidation of substrates of xanthine oxidase. There appears to be a special relation between purines, especially hypoxanthine, as substrates and the chemiluminescent compound. Although other substrates (DPNH, HCHO, CH&HO) produce light the intensity is very much lower than with hypoxanthine. It appeared during the course of these studies that DBA++ is an electron acceptor with special properties which are of great interest, This compound is a pyridine derivative with many properties similar to diphosphopyridine nucleotide. It forms addition products relatively insoluble in aqueous solvents, with cyanide ion and with hydroxyl ion. It reacts with hydrosulfite with luminescence (2) and we have likewise observed light emission when it reacts with acetone. The analogy with DPN has provided a fruitful field for further investigation of the reactions of DBA++ and these in turn may prove useful in elucidation of some DPN addition reactions. For recent work on DPN addition products and for references see Van Eys (3) and Singer and Kearney (4). The emission of light by DBA++ has been studied by numerous workers since its discovery by Gleu and Petsch (2). A review of the early work has been written by Anderson (5) and some newer work is to be found in the volume on bioluminescence