Toxic mechanism of eucalyptol and β-cyclocitral on Chlamydomonas reinhardtii by inducing programmed cell death

Eucalyptol and β-cyclocitral are 2 main compounds in cyanobacterial volatile organic compounds and can poison other algae. To uncover the toxic mechanism of the 2 compounds, the cell growth, photosynthetic abilities, H₂O₂ production, caspase-like activities, nuclear variation and DNA laddering were investigated in Chlamydomonas reinhardtii treated with eucalyptol and β-cyclocitral. Eucalyptol at ≥ 0.1 mM and β-cyclocitral at ≥ 0.05 mM showed toxic effects on C. reinhardtii cells, and 1.2 mM eucalyptol and 0.4 mM β-cyclocitral killed the whole of the cells during 24 h. During the death process, the photosynthetic pigment gradually degraded, and Fv/Fm gradually declined, indicating that the death is not a necrosis due to the gradual disappearance of the physiological process. In the treatments with 1.2 mM eucalyptol and 0.4 mM β-cyclocitral, H₂O₂ content burst at 10 min and 30 min, respectively. Caspase-9-like and caspase-3-like were activated, and cell nucleuses concentrated firstly and then broke with prolonging the treatment time. Meanwhile, DNA showed laddering after 1 h, and was gradually cleaved by Ca²⁺-dependent endonucleases to mainly about 100-250 bp fragments. These hallmarks indicated that eucalyptol and β-cyclocitral may poison other algal cells by inducing programmed cell death triggered by the increased H₂O₂.