Coelenterazine analogs as chemiluminescent probe for superoxide anion.

Eleven new coelenterazine analogs containing the 3,7-dihydroimidazo[1,2-alpha]pyrazin-3-one structure were synthesized. The superoxide-triggered chemiluminescence of these compounds was investigated using the hypoxanthine-xanthine oxidase system in comparison with four known compounds. The results showed that an alkyl substitution at the position 5 of the imidazopyrazinone ring causes a drastic decrease in the superoxide-dependent chemiluminescence intensity, whereas a dimethylene bridge added between the position 5 and the phenyl group bound to the position 6 dramatically increases the luminescence intensity, indicating the potential usefulness of this type of compound as a probe for superoxide anion. The luminescence intensity of the bridged analog was 33 times greater than that of MCLA [2-methyl-6-(4-methoxyphenyl)-3, 7-dihydroimidazo[1,2-alpha]pyrazin-3-one], the most sensitive superoxide probe of Cypridina luciferin-type. Two of the analogs synthesized, each with a covalently bound cyclodextrin, had a good solubility in water, an advantage in actual use. Moreover, one of them having a beta-cyclodextrin group showed a unique property; its luminescence was little affected by various substances in the environment.