Abstract
Singlet oxygen (1O2) is one of reactive oxygen species that causes photoinhibition, but, its reliable detection assay is not available. We developed a new detection system using 9-[2-(3-carboxy-9,10-diphenyl)anthril]-6-hydroxy-3H-xanthen-3 (DPAX) (Angew. Chem. Int. Ed. 1999, 38:2899), which enhances fluorescence (Ex. = 505 nm, Em. = 525 nm) when reacting with 1O2 and forming DPAX-endoperoxide. Xe-PAM enabled us to measure the fluorescence under actinic light. The fluorescence was also measured with confocal microscopy. The fluorescence increased when the mixture of DPAX and chloroplasts was exposed to actinic red light, but not under anaerobic condition or in the presence of the electron acceptor, ferricyanide, indicating 1O2 was not produced without O2 or with high electron flux. The results show detecting 1O2 production in plant system is feasible with DPAX. The usage of DPAX with Xe-PAM may be also useful to study the kinetics and to determine the production rate of 1O2.
