Evaluating the effects of allelochemical ferulic acid on Microcystis aeruginosa by pulse-amplitude-modulated (PAM) fluorometry and flow cytometry

• Ferulic acid exposure inhibited the growth of blue-green algae Microcystis aeruginosa.
• Allelopathic effects were evaluated by PAM and flow cytometry as diagnostic tools.
• Ferulic acid inhibited the photosynthetic activities of M. aeruginosa.
• Ferulic acid inhibited the membrane potential and esterase activity of M. aeruginosa.
• Membrane potential and esterase activity were identified as two sensitive variables.

We investigated the effects of allelochemical ferulic acid (FA) on a series of physiological and biochemical processes of blue-green algae Microcystis aeruginosa, in order to find sensitive diagnostic variables for allelopathic effects. Algal cell density was significantly suppressed by FA (0.31–5.17 mM) only after 48 h exposure. Inhibitions of photosynthetic parameters (Fv/Fm and Fv′/Fm′) occurred more rapidly than cell growth, and the stimulation of non-photochemical quenching was observed as a feed-back mechanisms induced by photosystem II blockage, determining by PAM fluorometry. Inhibitions on esterase activity, membrane potential and integrity, as well as disturbance on cell size, were all detected by flow cytometry with specific fluorescent markers, although exhibiting varied sensitivities. Membrane potential and esterase activity were identified as the most sensitive parameters (with relatively lower EC50 values), and responded more rapidly (significantly inhibited only after 8 h exposure) than photosynthetic parameters and cell growth, thus may be the primary responses of cyanobacteria to FA exposure. The use of PAM fluorometry and flow cytometry for rapid assessment of those sensitive variables may contribute to future mechanistic studies of allolepathic effects on phytoplankton.