L-DOPA production from tyrosinase immobilized on nylon 6,6

Abstract

The production of L‐DOPA immobilized on chemically modified nylon 6,6 membranes was studied in a batch reactor. Tyrosinase was immobilized on nylon using glutaraldehyde as a crosslinking agent. The effects of membrane pore size and glutaraldehyde concentration upon enzyme uptake and L‐DOPA production were investigated. Enzyme uptake was unaffected by glutaraldehyde concentration; approximately 70% uptake was observed when 25% w/v (group 1), 5% (group 2), and 3% (group 3) glutaraldehyde were used, indicating that glutaraldehyde was in excess. Similarly, uptake was the same for membranes with 0.20 and 10 μm pore sizes. Membranes produced using different levels of glutaraldehyde exhibited dramatically different capacities for L‐DOPA production, despite the fact that enzyme uptake was equivalent. Membranes from groups 2 and 3 (5% and 3% glutaraldehyde) produced L‐DOPA at a rate of 1.70 mg L−1 h−1 over 170 h in a 500‐mL batch reactor. However, no free L‐DOPA was detected when group 1 membranes were used. Experimental evidence suggests that L‐DOPA was produced, but remained bound to these membranes via excess glutaraldehyde left over from the immobilization process. Membrane pore size also effected L‐DOPA production; less production was observed when 10‐μm membranes were used, despite equivalent enzyme uptake. The observed difference in production may be due to differences in the pore density on the two types of membranes which could affect the access of the substrate to the immobilized enzyme. The results of these studies indicate that tyrosinase can be effectively immobilized on nylon 6,6. L‐DOPA production was optimal when 0.20‐μm‐pore‐size membranes were activated with 3–5% glutaraldehyde. Stability studies indicated a 20% reduction in activity over 14 days when the immobilized enzyme was used under turnover conditions. © 1996 John Wiley & Sons, Inc.

Overview on the biotechnological production of l-DOPA

“…It also consecutively oxidizes diphenol to quinone by catecholase activity (Min and Yoo 2009). Accordingly, L-DOPA can be produced by the o-hydroxylation of L-tyrosine by the cresolase activity in tyrosinase (Ates et al 2007;Min et al 2010;Pialis et al 1996;Seetharam and Saville 2002;Xu et al 2012). As well as in biocatalytic conversion, microorganisms with tyrosinase activity have been widely used in microbial fermentation for L-DOPA production using L-tyrosine as the feedstock.…”

“…As a result, 0.143 g of L-DOPA was optimally produced with a productivity of 1.70 mg L −1 h −1 during 170 h in a 500 mL batch reactor using a membrane with a pore size of 0.20 μm and 3 % of glutaraldehyde as the cross linker. In addition, the immobilized tyrosinase retained 80 % of its initial activity over 14 days and thus the tyrosinase-immobilized membrane reactor might be suitable for long-term operation (Pialis et al 1996). Catecholase activity in tyrosinase often hinders the enzymatic L-DOPA synthesis because L-DOPA rapidly oxidizes to the by-product DOPAquinone by the catecholase activity.…”

Biological sources of L-DOPA: An alternative approach

“…It lowers the production cost due to the reusability of the enzymes. The various techniques used for enzyme immobilization include entrapment in polymeric gels, adsorption onto insoluble materials, encapsulation in membranes, cross-linking with bifunctional or multifunctional reagents and linking to an insoluble carrier [16][17][18][19][20][21][22][23]. The substrate used for the synthesis of L-DOPA were catechol, sodium pyruvate and ammonium acetate.…”

Kinetic characterization of tyrosinase containing mushroom (Agaricus bisporus) cells immobilized in silica alginate

“…This reaction pattern has been examined for tyrosinase catalyzed production of various o-diphenols [3,4], among these the antioxidant hydroxytyrosol [5], 3,4-dihydroxyphenylacetic acid [4], a substance with antiproliferative activity against cancer cells [6], and 3,4-dihydroxy-l-phenylalanine (l-DOPA) [7][8][9] which is a prescribed drug for treatment of Parkinson's disease. The formation of the o-quinones in these reaction systems can be reduced by addition of ascorbic acid as a reducing agent [3][4][5][7][8][9].…”

“…The formation of the o-quinones in these reaction systems can be reduced by addition of ascorbic acid as a reducing agent [3][4][5][7][8][9].…”