Poster Presentation 1B-11

Enzymatic Hydrolysis by Immobilized Lipase Applied to a New Prototype Anti-asthma Drug

Juliana Vaz Bevilaqua,¹ Aline Gomes Cunha,³ Lidia M. Lima,² Eliezer J. Barreiro,²

Tito L.M. Alves,¹ Lucia M. C. Paiva³ and Denise M. Guimarães Freire^3*

¹Programa de Engenharia Química/COPPE, UFRJ

²Faculdade de Farmácia, UFRJ

³Instituto de Química, Universidade Federal do Rio de Janeiro

Centro de Tecnologia (CT), sala 549-2, Cidade Universitária

Ilha do Fundão, CEP 21945-970 – Rio de Janeiro, Brazil.

Phone: +55 (21) 2562-7360

Fax: +55 (21) 2562-7266

E-mail: freire@iq.ufrj.br

The last step of the production of 4-(2-(1,3-dioxo-1,2-dihydro-2-isoindoylyl))-phenoxy acetic acid, designed to act as an anti-asthma drug, was performed by enzymatic hydrolysis of the respective ethyl ester. Reactions were performed in a three-phase system with the solid biocatalyst (Lipozyme^® RM IM), the organic solvent phase (ethyl acetate) and the aqueous phase (saturated Na₂CO₃ solution). In order to optimize the reaction conditions, an experimental design optimization procedure was used. The variables studied were the amount of enzyme, the reaction temperature and the volume of the saturated Na₂CO₃ solution. The experimental results were statistically analyzed and the main effects of the variables were adjusted. Dynamic experiments were then performed under optimized reaction conditions for different initial enzyme concentrations (0.5, 0.9 and 1.4 U_H Lipozyme/mL solvent). A semi-empirical non-linear model was proposed to describe the process and the experimental data were used to estimate the model parameters. The optimized reaction conditions found were 20 mg Lipozyme (0.9 U_H/mL_solvent), 5.0 mL Na₂CO_{3 (sat),} a temperature of 40^oC and batch reaction time of 6 hours. Under these optimized conditions i.e., aqueous phase excluded, three other promising substrates as pharmaceuticals building blocks such as 2-{4-[2-(1,3-dioxo-2,3-dihydro-1H-2-isoindolyl) ethyl] phenoxy} ethyl acetate, 2-[2-(1,3-dioxo-2,3-dihydro-1H-2-isoindolyl) ethyl] phenoxy ethyl acetate and 2-[4-(1,3-dioxo-2,3-dihydro-1H-2-isoindolyl) ethyl] phenoxy ethyl acetate were submitted to hydrolysis. The reactions were followed by thin layer chromatography (TLC). The reactions proceeded well in those cases in which para-substituted phenoxy groups were present but not for the orto-substituted substrate. The latter case possibly occurs because there is a steric hindrance, avoiding fitting of the alcoxy-ester group in the hydrophobic pocket of the enzyme. The enzymatic reaction could be used, therefore, to produce the desired 4-substituted compounds.