PHOTODYNAMIC ACTION*

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Abstract
Summary If trypsin is illuminated with visible light in the presence of an appropriate sensitizing dye and oxygen, a loss of enzymatic activity is observed. This phenomenon is called photodynamic inactivation. Quantum requirement studies were made on this system using illumination provided by a projector or high‐pressure mercury arc light equipped with multilayer interference filters. Light energy absorbed was measured using a vacuum thermocouplegalvanometer combination which was calibrated with a standard lamp. Aliquots of the illuminated system were taken at intervals and the tryptic activity remaining was assayed spectrophotometrically using the benzoyl‐arginine ethyl ester (BAEE) technique. BAEE is a synthetic substrate of trypsin. Two sensitizing dyes were used, methylene blue (MeB) and riboflavin‐5′‐phosphate (FMN). The quantum requirement of inactivation with both sensitizers decreases rapidly with increasing dye concentration, reaches a minimum, and then increases. Both sensitizers show increased quantum requirements of inactivation with increasing enzyme concentration. Both show quantum requirements which are independent of light intensity. FMN‐sensitized photodynamic inactivation shows large quantum requirements at acid pH's and low quantum requirements at basic pH's. MeB shows no activity as a sensitizer at acid pH's and high activity at basic pHs. Quantum requirements decrease slightly from 4°C to 33°C. The two dyes show different efficiencies of utilization of trapped light energy under similar conditions, which suggests that different photodynamic mechanisms may be acting in each case.

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