d‐DOPA IS UNIDIRECTIONALLY CONVERTED TOl‐DOPA BYd‐AMINO ACID OXIDASE, FOLLOWED BY DOPA TRANSAMINASE

SUMMARYMany studies have shown that administration ofd‐3, 4‐dihydroxyphenylalanine (d‐dopa) produces contralateral rotation in hemi‐parkinsonian animals comparable tol‐dopa, with less potency and slower onset. It was postulated thatd‐dopa was converted tol‐dopa to produce these effects.To investigate the postulated chiral inversion ofd‐dopa tol‐dopa and the related mechanism, an enantiomeric separation method ford‐ andl‐dopa using HPLC was first established. Then, rat kidney homogenates containingd‐dopa orl‐dopa were incubated and subjected to HPLC to detect traces of respective enantiomer generation. The mechanism of the chiral inversion ofd‐dopa was explored by direct measurement of the production ofl‐dopa in kidney homogenates.d‐Dopa incubations containing different concentrations of an inhibitor ofd‐amino acid oxidase (DAAO) and an inhibitor of dopa transaminase were measured forl‐dopa generation using HPLC. The role of DAAO in the chiral inversion ofd‐dopa tol‐dopa was further investigated by using purified DAAO and mutant ddY/DAAO−mouse kidney lacking DAAO activity.In rat kidney homogenate,d‐dopa was, indeed, converted tol‐dopa, whereasl‐dopa was not converted tod‐dopa. Sodium benzoate, a selective inhibitor of DAAO, blockedl‐dopa generation in a concentration‐dependant manner. In contrast with kidney homogenates of wild‐type ddY/DAAO+mice, those of mutant ddY/DAAO−mice lacking DAAO activity did not convertd‐dopa tol‐dopa unless exogenous DAAO protein was added. Conversely, when carbidopa, an inhibitor of dopa transaminase, was added to the homogenates, significant inhibition ofl‐dopa production was noted.These results prove the proposal thatd‐dopa undergoes unidirectional chiral inversion and further suggest thatd‐dopa is first oxidatively deaminated by DAAO to its α‐keto acid and then transaminated by dopa transaminase tol‐dopa.