Isolamento, purificação e caracterização da peroxidase de yacon (Smallanthus sonchifolius)
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Data
2006-06-02
Autores
Kamimura, Gengis Kami Ferro [UNESP]
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
Os frutos e vegetais apresentam-se fisiologicamente ativos após a colheita, dessa forma as perdas pós-colheita ocorrem pela influência de diferentes fatores onde se destacam a respiração, a temperatura, a umidade, a concentração de oxigênio e gás carbônico, a produção de etileno e a ação de enzimas endógenas associadas com processos de deterioração. As raízes de yacon têm sido cada vez mais consumidas devido à revelação de qualidades medicinais. A manutenção de sua qualidade in natura é um sério problema nos processos pós-colheita devido a inúmeras reações metabólicas. A peroxidase (POD, E.C.1.11.1.7) é largamente encontrada nos vegetais apresentando importante papel fisiológico/bioquímico embora uma precisa função não tenha ainda sido estabelecida. Sua importância para ciência dos alimentos evidencia-se pelas relações com alterações indesejáveis na qualidade e resistência dos vegetais, tornando interessante sua supressão parcial ou total no pós-colheita. Os objetivos desse trabalho foram isolar, purificar e caracterizar a peroxidase de raízes de yacon. Condições de extração para a POD de yacon foram estabelecidas e a enzima foi isolada por precipitação com sulfato de amônio, eluição em Sephadex G-25 e DEAE-celulose. Somente um pico de atividade foi eluído no processo de purificação com um fator de purificação de 222,33. O peso molecular determinado foi 34.8 kDa com valores de pH e temperatura ótima de 5,5 e 35°C, respectivamente. As constantes Km e Vmax foram de 14,227 mM e 17409 UA/mL, para s-dianisidina, e de 14,434 mM e 14830 UA/mL, para H2O2. Foram testados os efeitos inibidores de sais, quelantes, compostos sulfidrila, ácidos fenólicos e outros. Estudos de inativação térmica da POD foram realizados, nos quais se verificou o efeito protetor da sacarose na inativação enzimática e o efeito promotor de inativação pela...
The fruits and vegetables still continue physiologically active after the crop, in that way the postharvest losses occurs by the influence of different factors, like the respiration, temperature, humidity, concentration of oxygen and carbonic gas, ethylene production and the action of endogenous enzymes associated with deterioration processes. Yacon (Smallanthus sonchifolius) roots has been more and more consumed by the revelation of prebiotic and medicinal properties. The maintenance of its quality in the postharvest is a serious problem due to several metabolic reactions. Peroxidase (POD, E.C.1.11.1.7) is widely found in plants, having physiological/biochemical importance, although a precise function has not still been established. The POD importance for food science is due to its relationships with alteration of the quality and resistance of the vegetables to the postharvest factors. Its total or partial suppression by thermal inactivation or inhibitor compounds may be interesting for postharvest interests. The objectives of this work were to isolate, purify and characterize the POD from yacon roots. Extraction conditions for yacon POD were determined and the enzyme was isolated by precipitation with ammonium sulfate, elution on Sephadex G-25 and DEAE-cellulose. Only one peak of enzyme activity was eluted on the purification process with a 222.33 fold purification factor. The determined molecular weight was 34.8 kDa. The optimum pH and temperature values were pH 5.5 and 35°C, respectively. Km and Vmax for ï-dianisidine was 14.227 mM and 17409 UA/mL, and 14.434 mM and 14830 UA/mL for H2O2, respectively. The effects of metals, chelating agents, sulphidryl, phenolic acids and other compounds as POD inhibitors were evaluated... (Complete abstract, click electronic address below).
The fruits and vegetables still continue physiologically active after the crop, in that way the postharvest losses occurs by the influence of different factors, like the respiration, temperature, humidity, concentration of oxygen and carbonic gas, ethylene production and the action of endogenous enzymes associated with deterioration processes. Yacon (Smallanthus sonchifolius) roots has been more and more consumed by the revelation of prebiotic and medicinal properties. The maintenance of its quality in the postharvest is a serious problem due to several metabolic reactions. Peroxidase (POD, E.C.1.11.1.7) is widely found in plants, having physiological/biochemical importance, although a precise function has not still been established. The POD importance for food science is due to its relationships with alteration of the quality and resistance of the vegetables to the postharvest factors. Its total or partial suppression by thermal inactivation or inhibitor compounds may be interesting for postharvest interests. The objectives of this work were to isolate, purify and characterize the POD from yacon roots. Extraction conditions for yacon POD were determined and the enzyme was isolated by precipitation with ammonium sulfate, elution on Sephadex G-25 and DEAE-cellulose. Only one peak of enzyme activity was eluted on the purification process with a 222.33 fold purification factor. The determined molecular weight was 34.8 kDa. The optimum pH and temperature values were pH 5.5 and 35°C, respectively. Km and Vmax for ï-dianisidine was 14.227 mM and 17409 UA/mL, and 14.434 mM and 14830 UA/mL for H2O2, respectively. The effects of metals, chelating agents, sulphidryl, phenolic acids and other compounds as POD inhibitors were evaluated... (Complete abstract, click electronic address below).
Descrição
Palavras-chave
Yacon, Peroxidase, Cinética enzimática, Inativação térmica, Thermal inactivation
Como citar
KAMIMURA, Gengis Kami Ferro. Isolamento, purificação e caracterização da peroxidase de yacon (Smallanthus sonchifolius). 2006. 100 f. Dissertação (mestrado) - Universidade Estadual Paulista, Faculdade de Ciências Farmacêuticas, 2006.