Biorredução de olefinas ativadas catalisada por leveduras e fungos em meio aquoso ou sistema bifásico

Abstract

As olefinas eletronicamente ativadas com grupos retiradores de elétrons, (4R)-carvona (7), (4S)-carvona (8), (6E)-2-metil-6-[(4-nitrofenil)metileno]-ciclo-hexanona (22), (3E)-3-metil-4-fenil-3-buten-2-ona (23), (2E)-1,3-difenil-2-propen-1-ona (24) e 3-metil-ciclo-hexen-2-ona (25), foram utilizadas como substratos nas reações de biorredução mediada por leveduras e fungos em meio aquoso. Realizou-se a otimização das condições reacionais para a biorredução dos substratos 7, 8 e 22 com Fermento de pão comercial (FP, Saccharomyces cerevisiae), onde avaliou-se o efeito da concentração de levedura e substrato, temperatura, pH e adição de aditivos. A biotransformação de 7, 22, 23, 24, 25 e 8 formou as cetonas saturadas (1R,4R)-dihidrocarvona (10), 2-metil-6-(4-nitrofenilmetil)-ciclo-hexanona (33), (3S)-3-metil-4-fenil-2-butanona (34), 3-difenil-2-propen-1-ona (35) e 3-metil-ciclo-hexanona (36) pela redução da ligação dupla C-C e do álcool saturado (1R,2R,4S)-dihidrocarveol (12) pela redução das ligações dupla C-C e C-O, respectivamente. Na biotransformação de 8, utilizou-se também os fungos isolados da Carapanaúba e as leveduras Cryptococcus flavescens e Spathaspora xylofermentans. Um novo método de extração dos reagentes e produtos do meio aquoso e/ou sistema bifásico foi desenvolvido, baseado em micro extração em fase líquida com fibras ocas de polipropileno (HF-LLLME).<br>

Abstract : In this study, the electronically activated alkenes with electron-withdrawing groups, such as (4R)-carvone (7), (4S)-carvone (8), (6E)-2-methyl-6-[(4-nitrophenyl)metylene]-cyclohexanone (22), (3E)-3-methyl-4-phenyl-3-buten-2-one (23), (2E)-1,3-diphenyl-2-propen-1-one (24) and 3-methyl-cyclohexen-2-one (25) were used as substrates in the bioreduction reaction mediated by yeasts and fungi in aqueous media or in biphasic systems. Firstly, the optimization of experimental condition on the bioreduction of 7 catalyzed by commercial baker#s yeast (BY, Saccharomyces cerevisiae) was carried out, where the effect of biocatalyst system, yeast and substrate concentration, temperature, pH, addition of additives and volumetric percentage of additives were evaluated. The optimum conditions obtained in the reduction of (4R)-carvone mediated FP were when the reaction was carried out in aqueous medium, substrate concentration and yeast of 16.6 mmol L-1 and 100.0 g L-1, temperature 25 °C, pH 7.5, and with the additives DMSO, trehalose and sucrose. Then, and using the best reaction conditions obtained on the biotransformation of 7, the bioreduction of the other electronically activated alkenes was performed. The biotransformation of 7, 23, 24, 25 e 8 formed the corresponding saturated ketones (1R,4R)-dihydrocarvone (10) (conv. 70% and d.e. of 97%), 2-methyl-6-(4-nitrophenylmethyl)-cyclohexanone (33) (conv. 59% and d.e. of 38%), (3S)-3-methyl-4-phenyl-2-butanone (34) (conv. 33% and e.e. of 64%), 3-diphenyl-2-propen-1-one (35) (conv. >99%) by the reduction of C#C double bond, and the saturated alcohol (1R,2R,4S)-dihydrocarveol (12) (conv. 4% and of >99%) by the reduction of C#C and C#O double bond, respectively. As low conversions into 12 and 34 were obtained, the optimization of experimental conditions on the bioreduction of 8 and 23, also was carried out to obtain the product in higher conversions and d.e. or e.e.. The reaction parameters influenced the formation of 10, 12 and 34, however temperature and substrate concentrations were the most outstanding. In the biotransformation of 8, besides using the yeast S. cerevisiae as biocatalyst, the isolated fungis from Aspidosperma excelsum Benth (carapanaúba) (CG 0504 e CF 0305) and the yeast C. flavescens e S. xylofermentan were used. From the obtained results, these fungi and yeasts showed a great potential to be used as biocatalysts. In this study, a new and green methodology for the extraction of reagents and products from the aqueous media or biphasic system based on a liquid-liquid-liquid micro extraction with porous hollow fiber polypropylene membranes (HF-LLLME) is also reported. The main advantages of this methodology in relation to conventional liquid-liquid extraction are the smallest waste generation (organic solvent) and the time required for extraction. In conclusion, the use of microorganisms as biocatalysts in the biotransformation of electronically activated alkenes compounds in aqueous media is a viable alternative to obtain compounds with high optical purity, under mild reaction conditions, low cost reagents and environmental pollution.