A helpful technology - the luminescence detection of singlet oxygen to investigate photodynamic inactivation of bacteria (PDIB)

Photodynamic inactivation of bacteria (PDIB) is considered a new approach for the struggle against multiresistant bacteria. To achieve a sufficient level of bacteria killing, the photosensitizer must attach to and/or penetrate the bacteria and generate a sufficiently high amount of singlet oxygen. To optimize PDIB, the direct detection and quantification of singlet oxygen in bacteria is a helpful tool. Singlet-oxygen luminescence is a very weak signal, in particular in living bacteria. We first performed experiments in aqueous photosensitizer solution to optimize the luminescence system. We eliminated non-singlet-oxygen photons, which is important for the quantification of singlet oxygen and its rise and decay rates. This procedure is even more important when the laser excitation beam is scattered by bacteria (diameter 1 mu m). In suspensions with both Gram-positive and Gram-negative bacteria we then clearly detected singlet oxygen by its luminescence and determined the respective rise and decay times. The decay times should provide an indication of localization of singlet oxygen and hence of the photosensitizer even in small bacteria. [GRAPHICS] The photosensitizer is excited by a laser beam with a Gaussian beam intensity distribution inside a cuvette. This may lead to oxygen depletion at the site of singlet-oxygen generation, in particular when the photosensitizer is inside bacteria.