Resumo Neste trabalho foi determinada a influência da temperatura, do pH e do tempo de armazenamento na estabilidade enzimática de frutosiltransferase (E.C.2.4.1.9) extracelular de Aspergillus oryzae IPT-301, produzida por fermentação submersa. Os parâmetros termodinâmicos mostraram tendência de aumento da desnaturação da enzima com o aumento da temperatura. A atividade de transfrutosilação máxima foi obtida para a enzima incubada em pH 5,5. Durante o armazenamento a 4 °C, a atividade de transfrutosilação diminuiu, enquanto a atividade hidrolítica aumentou, principalmente nas primeiras nove horas, período em que a enzima apresentou 45,6% de sua atividade inicial de transfructosilação. Esses resultados contribuem para a melhoria das condições de armazenamento, imobilização e utilização das frutosiltransferases solúveis na produção de frutooligossacarídeos.

Abstract In this work, it was determined the influence of temperature, pH and storage time on the enzymatic activity and stability of an extracellular fructosyltransferase (FTase E.C.2.4.1.9) from Aspergillus oryzae IPT-301 produced by submerged fermentation. The thermodynamic parameters showed a tendency for increasing enzyme denaturation with the rise in temperature. The maximum transfructosylation activity was obtained at the incubation pH 5.5. During storage at 4 °C, the transfructosylation activity decreased, whereas the hydrolytic activity increased, especially in the first nine hours, a time in which the enzyme presented 45.6% of its initial transfructosylation activity. These results contributed to the improvement of the conditions of storage, immobilization and use of the soluble fructosyltransferases (FTase) in fructooligosaccharide (FOS) production.

Aspergillus oryzae IPT-301, previously reported as a β-fructofuranosidase producing microorganism, was successfully mutated using UV irradiation at 253.7 nm followed by the screening of survivors resistant to certain stress conditions. Strains were first subjected to the β-fructofuranosidase activity assay using a portion from the colony grown in Petri dish as the enzyme source. Seven mutants with fructofuranosidase activity values relative to the parent culture between 140 -190% were selected from survivors grown at temperature of 40ºC or 0.018% (w/v) sodium dodecyl sulfate concentration. They were cultivated on a rotary shaker to characterize mycelium and extracellular fructosyltransferase activities. Three mutants named IPT-745, IPT-746 and IPT-748 showed the highest amount of mycelium activity whose values increased 1.5 -1.8 fold, compared with the parent strain. It was found that more than 55% of total enzyme activity (mycelium- plus extracellular- activity) from these strains was detected in the mycelium fraction. Only one mutant, IPT-747, exceeded the amount of extracellular enzyme exhibited by the parent strain (1.5 times). This mutant also showed the highest value of total fructosyltransferase activity.