Embalagem a vácuo: efeito no escurecimento e endurecimento do feijão durante o armazenamento / Vacuum package: its effect on the hardening and darkening of the beans during storage

A influência do uso de embalagem a vácuo no controle do escurecimento e endurecimento do tegumento foi avaliada em feijões da cv. 'Pérola'. Os feijões após secos foram submetidos a diferentes embalagens: polietileno seladas a vácuo (espessura de 80µm), polietileno seladas sem vácuo (espessura de 80µm) e embalagem comercial (espessura de 20µm), armazenados por oito meses em condições ambiente. Foram realizadas análises de umidade, tempo de cozimento, cor, atividade enzimática e compostos fenólicos, logo após a secagem e em intervalos de dois meses até o oitavo mês. O tempo de cozimento foi maior para os grãos acondicionados em embalagem comercial do que nas outras embalagens, ao final do armazenamento. Os grãos embalados em sacos de polietileno selados a vácuo apresentaram menor queda nos valores de L*. As amostras da embalagem selada a vácuo apresentaram atividade da Peroxidase e Polifenoloxidase inferior aos grãos da embalagem comercial. O teor de fenólicos totais não apresentou diferença significativa. O uso da embalagem selada a vácuo foi eficiente em retardar o aumento no tempo de cozimento, o escurecimento do tegumento, a atividade das enzimas Polifenoloxidase e Peroxidase da cultivar 'Pérola', durante o armazenamento por oito meses em condição ambiente.

The influence of using vacuum packaging to control browning and hardening of the seed coat was evaluated in beans cv. 'Pérola'. The beans after dried, were subjected to different packaging: vacuum sealed polyethylene (thickness: 80μm), no vacuum sealed polyethylene (thickness: 80μm) and commercial packaging (thickness: 20μm), stored for eight months under ambient conditions. Analyses of moisture, cooking time, color, enzymatic activity and phenolic compounds, shortly after drying and at intervals of two months until the eighth month. Cooking time was longer for the beans packed in commercial packaging, than in the other containers at the end of storage. The grains packed in polyethylene bags sealed under vacuum showed less decline in the values of L *. The samples will vacuum sealed package showed peroxidase activity and polyphenoloxidase below the grain of commercial packaging. The total phenolic content was not significantly different. The use of vacuum sealed package will was effective in slowing the increase in cooking time, the darkening of the integument, the activity of the enzymes polyphenoloxidase and peroxidase of bean cv. 'Perola' during storage for eight months at ambient condition.

Effects of oligosaccharides from endophytic Fusarium oxysporum Dzf17 on activities of defense-related enzymes in Dioscorea zingiberensis suspension cell and seedling cultures

Background Three oligosaccharides (EOS, WOS and SOS) were respectively prepared from the corresponding polysaccharides, namely exopolysaccharide (EPS), water-extracted mycelial polysaccharide (WPS) and sodium hydroxide-extracted mycelial polysaccharides (SPS) from the endophytic fungus Fusarium oxysporum Dzf17. In this study, the effects of EOS, WOS and SOS on the activities of the defense-related enzymes, namely phenylalanine ammonia lyase (PAL), polyphenoloxidase (PPO) and peroxidase (POD) in its host plant Dioscorea zingiberensis cultures were investigated. Results For the suspension cell cultures of D. zingiberensis, the highest PAL activity was induced by 0.5 mg/mL of WOS at 48 h after treatment, which was 4.55-fold as that of control. Both PPO and POD activities were increased to the maximum values by 0.25 mg/mL of WOS at 48 h after treatment, which were respectively 3.74 and 3.45-fold as those of control. For the seedling cultures, the highest PAL activity was elicited by 2.5 mg/mL of EOS at 48 h after treatment, which was 3.62-fold as that of control. Both PPO and POD reached their maximum values treated with 2.5 mg/mL of WOS at 48 h after treatment, which were 4.61 and 4.19-fold as those of control, separately. Conclusions Both EOS and WOS significantly increased the activities of PAL, PPO and POD in the suspension cell and seedling cultures of D. zingiberensis. The results suggested that the oligosaccharides from the endophytic fungus F. oxysporum Dzf17 may be related to the activation and enhancement of the defensive mechanisms of D. zingiberensis suspension cell and seedling cultures.

Avaliação do tempo de secagem e da atividade de óxido-redutases de yacon (Smallanthus sonchifolius) sob tratamento químico / Drying evaluation time and yacon (Smallanthus sonchifolius) enzymatic activity inhibition under chemical treatment

Evidências científicas reconhecem o yacon (Smallanthus sonchifolius) como fonte promissora de frutanos, tais como inulina e frutooligossacarídeos (FOS). No entanto, a presença de compostos fenólicos torna-o suscetível à reação de escurecimento enzimático. Este trabalho teve como objetivo avaliar o emprego de agentes químicos no processamento de yacon, para obtenção de farinha, a fim de inibir o escurecimento enzimático do produto e favorecer o tempo de secagem. Amostras de yacon sem inibição química, de yacon submetido à solução de cloreto de cálcio 1,0g 100g-1 por 30 minutos e de yacon submetido à solução de metabissulfito de potássio 0,5g 100g-1 por 5 minutos foram secas a 55°C, em estufa ventilada, e o teor de água e curvas de secagem foram determinados. As atividades das enzimas peroxidase (POD) e polifenoloxidase (PPO) foram determinadas antes e após a secagem como um possível marcador bioquímico do escurecimento enzimático desse tubérculo. No parâmetro umidade, os três tratamentos apresentaram-se semelhantes; porém, o tratamento 2 (cloreto de cálcio) reduziu a umidade no menor tempo. Antes e após o tratamento térmico, a atividade enzimática foi maior no tratamento 3 (metabissulfito de potássio). As atividades enzimáticas não foram inativadas completamente pela ação térmica. Para a obtenção da farinha de yacon, o tratamento com cloreto de cálcio 1,0g 100g-1 por 30 minutos foi o que apresentou melhor resultado. Apesar de não inibir totalmente a atividade das enzimas POD e PPO foi o melhor tratamento de inativação, propiciando menor tempo de secagem e melhor firmeza da matéria-prima e facilitando o processamento para obtenção da farinha.

Scientific evidences acknowledged the yacon (Smallanthus sonchifolius) like a promise source of fructans, as inulin and fructooligosaccharides - FOS, meanwhile, the presence of phenolic coumpounds became this susceptible to enzimatic browning reaction. This project aimed to evaluate the use of chemical agents in yacon processing to obtain flour in a way that inhibits enzymatic darkening of the product besides determining the enzymatic activity in these treatments. Samples of yacon without chemical inhibitions, yacon treated with 1.0g 100g-1 calcium chloride for 30 seconds and yacon treated with 0.5g 100g-1 potassium metabisulfite for 5 minutes were dried at 55oC in a ventilated greenhouse and the proportions of humidity and drying curves were determined. The peroxidase activities and polyphenol oxidase enzymes were checked before and after being dried with an enzymatic darkening possible biochemist marker of this tubercle. Regarding humidity Parameter all the three treatment were equivalent, but treatment 2 (calcium chloride) reduced the humidity in lower time. Before and after the thermal treatment the enzymatic activity was higher in treatment 3 (potassium metabisulfite). The thermal action did not inhibit completely polyphenol oxidase and peroxidase. The treatment with calcium chloride at 1.0g 100g-1 for 30 minutes to obtain yacon flour was the one with better result despite the fact that it did not inhibit completely peroxidase and polyphenol oxidase enzymes activity, offering a better drying time better material of row firmness , facilitating the process for obtaining the meal.

Some Biochemical properties of polyphenoloxidase from spearmint (Mentha arvensis)

Polyphenoloxidase (PPO; EC 1.14.18.1) extracted from Mentha arvensis leaves was isolated by (NH4)2SO4 precipitation and extensive dialysis. Its optimum pH and temperature varied with the substrate. The PPO showed activity with various diphenols. Km values were found 0.825, 0.928 and 7.41mM for caffeic acid, 4-methylcatechol and catechol, respectively. On heat-inactivation, half of the activity was lost after 60 and 15 sec at 70 and 75ºC, respectively. Measuring of residual activity showed a stabilizing effect of sucrose at various temperatures with activation energy (Ea) for inactivation increasing with sucrose concentration from 0 to 40 percent (w/w). Ea values of 78.13; 80.37; 82.79 and 81.00 kJ/Mol were found for 0, 15; 30 and 40 percent sucrose, respectively. PPO was inhibited by ascorbic, benzoic, cinnamic, ferulic, p-coumaric, protocatechuich acids, sodium metabisulfite, pyrogallol and resorcinol. The Ki values showed that ascorbic acid was the most effective inhibitor. The type inhibition was determined for each inhibitor.

Polifenoloxidase (PPO, EC 1.14.18.1) extraída de folhas de Mentha arvensis foi isolada por precipitação com (NH4)2SO4 e diálise extensiva. Seu pH e temperatura ótimos variaram com o substrato. A PPO apresentou atividade com vários difenóis. Valores de Km foram 0,825; 0,928 e 7,41 mM para ácido caféico, 4-metilcatecol e catecol, respectivamente. Na inativação térmica, 50 por cento da enzima foi inativada após 60 e 15 segundos a 70 e 75ºC, respectivamente. A medida de atividade residual mostrou um efeito estabilizante de sacarose a várias temperaturas e uma energia de ativação (Ea) para inativação aumentando com a concentração de sacarose de 0 a 40 por cento (p/p). Valores de energias de ativação de 78,13; 80,37; 82,79 and 81,00 kJ/Mol foram encontradas para 0, 15, 30 e 40 por cento de sacarose, respectivamente. A PPO foi inibida pelos ácidos ascórbico, benzóico, cinamico, ferulico, p-cumárico, protocatéquico, além de metabisulfito de sódio, resorcinol e pirogalol. Os valores de Ki mostram que o ácido ascórbico foi o mais efetivo inibidor. O tipo de inibição foi determinado para cada inibidor.

Influência do processamento por via úmida e tipos de secagem sobre a composição, físico química e química do café (Coffea arabica L) / Influence of the wet processing and drying types on chemical and physicochemical composition of coffee (Coffea arabica L. )

O café (Coffea arabica L.) é um importante produto de exportação brasileira, por fazer parte do hábito alimentar da população de diversos países. Sua produção vem passando por transformações tecnológicas que têm como objetivo agregar valores qualitativos ao produto destinado tanto para mercado interno como externo. A exportação do café e de produtos alimentícios deve se adequar aos programas de qualidade estabelecidos por acordos políticos internacionais. Assim sendo, objetivou-se, nesse estudo, verificar a influência do método de preparo via úmida com tipos distintos de secagens, na obtenção do café cereja descascado, sobre a composição, físico-química e química do café. Houve influência da forma, preparo e tipo de secagem sobre as principais características estudadas. O café cereja descascado apresentou diferenças nos principais indicadores químicos físico-químicos e com uma superioridade para esse método de preparo com secagem exclusiva no terreiro, em diversos aspectos. Houve redução na condutividade elétrica, lixiviação de potássio e aumento da atividade da polifenoloxidase.

Coffee (Coffea arabica L.) is an important Brazilian export product, being part of the eating habit in several countries. Its production has experienced technological transformations, with the aim at adding quality values to the product destined to internal as well as to external market. The export of coffee and food products must adjust to the quality programs established by international political agreements. In this context, this work was carried out to verify the influence of the wet processing using different drying types on the obtentainment of the husked coffee cherry on the chemical and physicochemical composition of the coffee. There was influence of the preparation and the drying type methods on the main studied characteristics. The peeled red cherry coffee presented differences in the main chemical and physicochemical indicators, with a superiority in the preparation method with exclusive drying on the patio, in several aspects. Reduction in the electric conductivity and potassium leaching was observed as well as increase in the polyphenoloxidase activity.

INHIBICIÓN DE LESIONES POR FRÍO DE PITAYA AMARILLA (Acanthocereus pitajaya) A TRAVÉS DEL CHOQUE TÉRMICO: CATALASA, PEROXIDASA Y POLIFENOLOXIDASA / Inhibition of Chillyng Injury in Pitaya Amarilla (Acanthocereus pitajaya) Fruit by Heat Shock: Catalese, Peroxidase and Polyphenoloxidase

En los ensayos de almacenamiento de pitaya a temperatura ambiente de Bogotá (18 ºC) se encontró que esta fruta tiene un comportamiento climatérico con un máximo en la respiración luego de tres días de iniciado el almacenamiento. En el máximo climatérico la actividad de catalasa fue máxima, en tanto que en la etapa de senescencia las actividades de peroxidasa y polifenoloxidasa exhibieron valores máximos. El choque térmico inhibió las lesiones por frío, vistas en los frutos refrigerados a 2 °C, este choque incrementó la actividad de catalasa y peroxidasa y disminuyó la actividad de polifenoloxidasa, respecto a los frutos refrigerados sin tratamiento de choque térmico. Los resultados muestran que la catalasa está en relación directa con la vida útil del fruto, mientras que polifenoloxidasa guarda estrecha relación con el deterioro. La peroxidasa manifiesta su acción antioxidante con la generación de pardeamiento, en frutos almacenados a temperatura ambiente, si bien en los tratados con choque térmico, su acción antioxidante no va de la mano con el incremento en el pardeamiento, por lo que en este caso, su expresión fue favorable. Los resultados encontrados se constituyen en un aporte en la búsqueda de técnicas que permitan mayores tiempos de vida en anaquel de los frutos.

In the storage of yellow pitaya fruit at room temperature in Bogotá (18 ºC), it was found that this fruit has a climacteric behavior with a maximum in the respiration after 3 days of its storage. In the climacteric the activity of catalase was higher, while in the senescence stage the activities of peroxidase and polyphenoloxidase exhibited maximum values. The heat shock inhibited the chilling injury, shows in the fruits refrigerated at 2 °C, this heat shock increased the activity of catalase and peroxidase and it delayed the activity of polyphenoloxidase, regarding the fruits refrigerated without treatment of heat shock. The results show that catalase is in direct relationship with the useful life of the fruit, while polyphenoloxidase keeps it narrows relationship with the deterioration. In fruits stored at room temperature, peroxidase exhibited its antioxidant action with browning generation, although in the treaties with heat shock, their antioxidant action doesn’t go of the hand with increment in the browning, for that in this case, its expression was favorable. The opposing results are constituted in a contribution in the search of techniques that allow bigger times of life in shelf of the fruits.

Cinética de inativação da polifenoloxidase e peroxidase de abacate (Persea americana Mill.) / Kinetic of inactivation of polyphenoloxidase and peroxidase of avocado (Persea americana Mill.)

Extratos enzimáticos foram preparados a partir da polpa de abacate (Persea americana Mill.) dos cultivares Quintal, Fortuna e Choquete, em dois estádios de maturação (verde e maduro). A polpa de abacate (150,00g) foi homogeneizada em liquidificador, com 300 mL de solução tampão fosfato de sódio 100mM em pH 7,4 para polifenoloxidase (PPO) e pH 6,0 para peroxidase (POD). A peroxidase ionicamente ligada foi extraída usando solução de NaCl 1,0M, em tampão fosfato de sódio 100mM pH 6,0. Para o estudo da estabilidade térmica, os extratos foram submetidos a temperaturas de 60, 65, 70, 75 e 80ºC e por períodos que variaram de 1 à 10 minutos e a atividade enzimática foi determinada por espectrofotometria (lambda= 395nm para PPO e lambda= 460nm para POD). Pode-se observar que o declínio rápido da atividade das enzimas era maior nos primeiros quatro minutos e após esse período, com o aumento da temperatura e do tempo, a atividade continuou diminuindo, porém de forma mais lenta. A energia de ativação na faixa de temperatura estudada para as porções termolábil e termorresistente, apresentou valores dentro da faixa 12-100 kcal/mol.

Enzymatic extracts were prepared from the avocado pulp (Persea americana Mill.), using the cultivars Quintal, Fortuna and Choquete, in two different maturation stages (green and mature avocados). The avocado pulp (150,00g) was homogenized in blender with 300 mL of sodium phosphate buffer 100mM at pH 7.4 for polyphenoloxidase (PPO) and pH 6.0 for peroxidase (POD). The peroxidase ionically bound was extracted using NaCl solution 1.0M in sodium phosphate buffer 100mM pH 6.0. For the study of the thermal stability the extracts were submitted at temperatures of 60, 65, 70, 75 and 80ºC for periods that had a variation from 1 to 10 minutes and the enzymatic activity was determined using a spectrophotometer (lambda= 395nm for PPO and lambda= 460nm for POD). It was possible to observe a fast decline of the enzyme activity in the first four minutes, after this period, with the increase of the temperature and time, the activity continued decreasing, however in a slower way. The activation energy in the studied temperature showed for the portions heat labile and heat resistant presented values inside 12-100 kcal/mol.

INDUCCIÓN DE POLIFENOLOXIDASA EN FRUTOS DE LULO (Solanum quitoense) COMO RESPUESTA A LA INFECCIÓN CON Colletotrichum acutatum / Polyphenol Oxidase Induction in Lulo Fruits (Solanum quitoense) Infected by Colletotrichum acutatum

Se evaluó la actividad polifenoloxidasa (PFO) en corteza de frutos de lulo con el fin de determinar su participación en respuestas hacia el patógeno Colletotrichum acutatum, causante de la antracnosis. Se estudiaron condiciones para la adecuada extracción de esta enzima, encontrándose que con buffer fosfatos 100 mM pH 7, 1% SDS y 1% PVPP se logran las mayores actividades. Se determinaron como mejores parámetros para medir la actividad de la enzima extraída, sustrato catecol 40 mM, pH 7,0, 23 °C y 30 µL de extracto. Para determinar su posible inducción en la interacción con el patógeno, se realizó un ensayo in vivo usando frutos verdes, pintones y maduros, inoculados con el hongo o con agua estéril. A nueve tiempos postinoculación se determinó la actividad PFO encontrándose que hay una respuesta diferencial con el tiempo y la madurez de los frutos y por efecto del patógeno. Se obtuvo aumento de actividad en lulos verdes a 48, 96 y 144 horas postinoculación (hpi) y en maduros a la mayoría de los tiempos evaluados, siendo éste estado en el que se presentó la respuesta más notable de inducción. En pintones aumentó solo a 72 y 144 hpi. Los mayores valores se registraron en general para frutos en estado verde. Los frutos respondieron al estrés ocasionado por la herida activando también esta enzima. La inducción de actividad se presentó a tiempos más rápidos en los frutos menos afectados por la enfermedad (verdes y maduros), por lo que se puede postular una relación positiva entre inducción de PFO y respuesta de tolerancia a la antracnosis.

Polyphenol oxidase (PFO) activity induction was evaluated in lulo fruits to determine the role of this enzyme in resistance responses towards the pathogen Colletotrichum acutatum which causes anthracnose disease. We studied the experimental conditions to obtain the enzyme, using lulo peel, and found that extraction with phosphates buffer 100 mM pH 7, 1% SDS y 1% PVPP showed higher activities. The adequate parameters for activity measurement was also evaluated and established as cathecol 40 mM, pH 7,0, 23 °C and 30 µL of extract. Then, we performed an in vivo assay using lulo fruits in three maturity stages, green, semimature and mature, which were inoculated with the fungus or with sterile water. Enzymatic induction of this protein was studied at nine postinoculation times, and it was found that the induction was differential according to the time, the maturity stage, and as consequence of pathogen presence. The PFO activity increased in green fruits at 48, 96 y 144 (hours postinoculation (hpi), and in mature lulos for the majority of times studied, with the most significant induction response at this stage. In semimature lulo, the induction was observed only at 72 and 144 hpi. The highest nominal value of activity was found in green fruits. Fruits responded to incision with enzyme activation. The increase in the activity of the enzyme was faster in the fruits with the minor anthracnose symptoms than the ones that were more affected. Therefore, it is possible to postulate a positive relation between PFO induction and tolerance to anthracnose symptoms.

CATALASA, PEROXIDASA Y POLIFENOLOXIDASA DE PITAHAYA AMARILLA (Acanthocereus pitajaya) / CATALASE, PEROXIDASE AND POLYPHENOLOXIDASE FROM PITAHAYA AMARILLA FRUITS (Acanthocereus pitajaya)

Las enzimas catalasa (CAT), peroxidasa (POD) y polifenoloxidasa (PFO) fueron extraídas de la corteza de frutos de pitaya amarilla (Acanthocereus pitajaya) y caracterizadas parcialmente. Para CAT se halló que su actividad fue máxima a pH entre 6,8 y 7,5 y temperatura entre 30 a 50 °C y un K M de 442 mM con H2O2 como sustrato. Para POD se encontró un pH de máxima actividad entre 5,0 a 5,5, temperatura de máxima actividad entre 20 a 25 °C y valores de K M de 10,6 mM para guayacol y 5,1 mM para H2O2. Para PFO las actividades máximas se obtuvieron a pH 7,0 y a temperaturas entre 30 a 40 °C; para esta enzima se obtuvo un K M de 5,5 mM con L-DOPA como sustrato. Las características encontradas para POD y PFO indican que estas enzimas pueden jugar un papel importante en el pardeamiento de la corteza de pitaya amarilla. Además, se evidenció el papel complementario que tienen CAT y POD ante diversas concentraciones celulares de H2O2.

Catalase (CAT), peroxidase (POD) and polyphenoloxidase (PPO) were extracted from the peel of pitaya amarilla (Acanthocereus pitajaya) and were partially characterized. CAT had maximum activities at pH between 6.8 and 7.5 and at temperatures in the range 30 to 50 °C, its K M value is 442 mM for H2O2.The pH for maximum activity of POD was from 5.0 to 5.5 and its temperatures between 20 and 25 °C; POD had a K M values of 10.6 mM and 5.1 for guaiacol and H2O2, respectively. PPO exhibited its maximum activity at pH 7.0 and at temperatures between 30 and 40 °C; PPO had a K M value of 5.5 mM for L-DOPA. Our results indicate that both POD and PPO play an important role in the browning of the pitaya amarilla peel; on the other hand, it was observed that CAT and POD must have a complementary role in the H2O2 degradation in the cells.

Purification and properties of polyphenoloxidase of mango peel (Mangifera indica).

Polyphenoloxidase from mango (Mangifera indica) peel was purified to homogeneity by ammonium sulphate fractionation, chromatography on DEAE-Sephadex and gel filtration of Sephadex G-200. The enzyme had an apparent molecular weight of 136,000. Its pH and temperature optimum were 5.4 and 50°C, respectively. The enzyme possessed catecholase activity and was specific to o-dihydroxy phenols. The enzyme also exhibited peroxidase activity. Some non-oxidizable phenolic compounds inhibited the enzyme competitively. High inhibitory effects were also shown by some metal chelators and reducing agents, Mango peel polyphenol oxidase when immobilized onto DEAE Sephadex showed slightly higher Km for catechol and lower pH and temperature optima.