Partial purification and characterization of limonoate dehydrogenase from Rhodococcus fascians for the degradation of limonin

An extracellular limonoate dehydrogenase was purified 10-fold from a cell-free extract of <i>Rhodococcus</i> <i>fascians </i>by ammonium sulfate precipitation, dialysis, and ultrafiltration. This purified dehydrogenase catalyzed the<br />conversion of limonoate to 17-dehydrolimonoate. The enzyme showed optimum activity at pH 8.0 and 40oC, with K_m value of 0.9 µM, and requires Zn ions and sulfhydryl groups for catalytic action. The enzyme activity was inhibited by Hg²⁺ and NaN₃ ions. The degradation of limonin (66%) in Kinnow mandarin juice was successfully demonstrated with partially<br />purified limonoate dehydrogenase. With scale-up preparation of limonoate dehydrogenase, a successful debittering operation of fruit juices appears feasible.<br />