Partial Purification of a Legume Nodulation Factor Present in Coconut Water

The nodulation of adventitious roots growing from segments of bean hypocotyl tissue was used as a bioassay for the material present in coconut water which stimulated nodulation. The active material in coconut water is acidic, but it was not possible to extract it from an acid solution with organic solvents. A purification of approximately 70-fold (on a dry wt basis) was obtained using activated charcoal, but at least 10 different compounds were present in the active fractions. A purified fraction of coconut water, which is stimulatory to the growth of carrot root explants, was active in the nodulation assay at a concentration of 2 mug/ml. This represents a 4000-fold purification of the diffusible fraction of coconut water. The charcoal fractionation procedure can be applied to the active material present in extracts of bean leaves.