Isolation and identification of phytotoxic substances from seeds of Coronilla varia L

In previous studies, it was shown that freeze-dried, aqueous leachate from seeds of Coronilla varia L. adversely affected the germination and seedling growth of many plant species. Depending upon the test species, phytotoxic effects were noted at leachate dilutions up to 1:10,000 (w/v), the highest tested. The objective of the present study was to isolate and identify the major phytotoxic compounds. Activated charcoal removed the phytotoxic component but dialysis through a cellulose acetate membrane did not. Using two-dimensional paper chromatography, 28 compounds were found in aqueous and alcoholic extracts of crownvetch seed leachate. The major phytotoxic substances were identified by paper chromatographic techniques and confirmed by co-chromatography with known compounds and, in a few cases, gas chromatography and UV spectra. Most were phenolic compounds such as scopoletin (7-hydroxy-6-methoxycoumarin), umbelliferone (7-hydroxy-coumarin), p-coumaric acid (trans-4-hydroxy-cinnamic acid), o-coumaric acid (trans-2-hydroxycinnamic acid), leuco-anthocyanins (flavan-3-4-diols) and catechin compounds (flavan-3-ols). Other compounds identified were: ferulic acid, cyanidin, delphinidin, glucose, galactose, and several flavonoids.