Isolation and identification of a novel allelochemical from Ruppia maritima extract against the cyanobacteria Microcystis aeruginosa

Because submerged plants have allelopathic inhibition against the cyanobacterial growth in the nature, the extracts of the submerged plants could be an environment-friendly and efficient alternative for controlling cyanobacterial bloom. A growth inhibitor of Microcystis aeruginosa was identified from the Ruppia maritima extracts by a two-step extraction. In the first step, Ruppia maritima was extracted by methanol, ethyl acetate and petroleum ether, respectively. The petroleum ether extract with the best algal inhibition effect (EC50=92.82 mg L−1) was further isolated, using column chromatography eluted with petroleum ether and ethyl acetate mixtures in volume ratios of 10:1, 7:1, 5:1, 3:1, 1:1 and 0:1 (v:v). Based on the GC–MS analysis results, 2,2’-methylenebis(6-tert-butyl-4-methyl-phenol) (AO 2246) was identified as the most effective compound with an EC50 of 7.80 mg L−1. The further bioassay experiments using Chlorella vulgaris and zebrafish indicated the cyanobacterial-specific allelopathic effect and low acute toxicity of AO 2246. The effect of AO2246 may be also long-acting in the natural environments, as the 28-day illumination of AO2246 had little influence on its cyanobacteria inhibition efficacy.