Fungal biotransformation of synthetic levodopa to stable dopamine in l-ascorbate-mediated aerobic-thermophilic biochemical process

Abstract

In the present study, fungal biotransformation of synthetic levodopa to stable dopamine in an l-ascorbate-mediated thermophilic-aerobic biochemical reaction was investigated. A mutant strain of Aspergillus oryzae EMS-6 was used for the preparation of mycelial biomass. The mutant was previously developed through EMS-induced mutagenesis and repressed against l-cysteine HCl. Growth parameters such as rate of cultivation (48 h), initial pH (6) and incubation temperature (30 °C) supported 18.84 g/l biomass with 23 g/l glucose consumption. Thermophilic behaviour of culture was observed at 25–40 °C. Kinetic variables notably µ = 0.385 /h and Q_s, exhibited consistent growth pattern. Biochemical reactions were performed aerobically using mycelial biomass as the source of enzyme ‘tyrosinase’ in a digital hotplate equipped with magnetic stirrers. The reaction conditions included 5 mg/ml biomass and 2.5 mg/ml levodopa as basal substrate in a thermophilic reaction of 25 min duration acidified with l-ascorbic acid. TLC and HPLC analysis of reaction mixture confirmed the presence of levodopa and dopamine using a CN-9dth (R) column. Activation enthalpy and entropy of dopa decarboxylase (DDC) and its thermal inactivation showed an improved biotransformation of levodopa to dopamine at the optimal temperature (30 °C) as compared to other temperatures being employed. Overall, 3.68 mg/ml dopamine (4.55 mg/ml proteins) synthesis from 2.38 mg/ml levodopa was accomplished. The enhancement in metabolic activity of the mutant strain is ~ 2.75-fold improved when compared to the unoptimized reaction conditions, which is highly significant (HS) indicating an eco-commercially viable (LSD ~ 0.412) bioprocess.