Mushroom based biocatalyst for the synthesis of dehydrodieugenol from clove oil: A potent cytotoxic agent

Dehydrodieugenol, an ortho dimer of eugenol is a widely bioactive natural neolignan. Known chemical or enzymatic methods for its preparation have one or more drawbacks such as hazardous reagents, low efficiency or high cost. In this study, a mushroom catalyzed whole-cell biocatalytic process was developed for the selective oxidative coupling of eugenol in the unfractionated clove oil (2.0 g/L) to produce dehydrodieugenol in its submerged culture. In an optimized condition (yeast malt extract medium, initial pH 7.0, 25-30 degrees C, 130-150 rpm, 6 h), Ganoderma sessile, a polypore mushroom of 'Reishi' group efficiently dimerized eugenol with good to excellent conversion. Preparative scale fermentation generated a metabolite titer of 0.62 g/L in the medium and subsequent purification yielded 38.3% w/w dehydrodieugenol with high purity (>98%). A substrate selectivity study indicated the strain to be highly selective to eugenol for an efficient and productive dimerization. Interestingly, the culture broth lyophilisate performed this coupling successfully even after storage of 3 months, revealing its potential as a ready-to-use storable biocatalyst. Further, cell-based assays found dehydrodieugenol to be 6-16 fold more cytotoxic (IC50 5.2-18.4 mu M) than eugenol against three cancer cell-lines. Thus, the developed bioprocess may have a synthetic utility to convert a natural extract to a promising cytotoxic candidate of natural origin.