Isolation and initial characterization of 13(2)-hydroxychlorophyll a induced by cyclohexanedione derivatives in tobacco cell suspension cultures

This paper reports that a new photobleaching compound, 2-(2-chloro-5-propoxycarbonylphenyl)aminomethylidene-5-5-dim ethyl- cyclohexane-1,3-dione (RWH-21), stimulates accumulation of 13(2)-hydroxychlorophyll a in cultured tobacco cells. This was shown based on isolation of 13(2)-hydroxychlorophyll a from pigment extracts of cultured tobacco cells by diode-array HPLC and subsequent fast atom bombardment mass spectrometry analysis. 13(2)-Hydroxychlorophyll a rapidly accumulates in tobacco cells both in the light and dark in the presence of RWH-21 (50 microM). Analysis of 13(2)-hydroxychlorophyll a formation in tobacco cells indicates that 13(2)-hydroxychlorophyll a is rapidly accumulated within 20 h incubation time both in the dark and light. Although the amount of 13(2)-hydroxychlorophyll a is continuously increased in the dark, the amount of 13(2)-hydroxychlorophyll a decreased remarkably in the light after 20 h incubation. Analysis of 13(2)-hydroxychlorophyll a formation and lipid peroxidation by determination malondialdehyde in tobacco cells suggests that RWH-21-induced 13(2)-hydroxychlorophyll a has the potential to cause a photodynamic action in cultured tobacco cells.