ABSTRACT
Mn-peroxidase (MnP), a biotechnologically important enzyme was purified for the first time from a plant source Musa paradisiaca (banana) stem, which is an agro-waste easily available after harvest of banana fruits. MnP was earlier purified only from the fungal sources. The enzyme was purified from stem juice by ultrafiltration and anion-exchange column chromatography on diethylamino ethylcellulose with 8-fold purification and purification yield of 65%. The enzyme gave a single protein band in SDS-PAGE corresponding to molecular mass 43 kDa. The Native-PAGE of the enzyme also gave a single protein band, confirming the purity of the enzyme. The UV/VIS spectrum of the purified enzyme differed from the other heme peroxidases, as the Soret band was shifted towards lower wavelength and the enzyme had an intense absorption band around 250 nm. The Km values using MnSO4 and H2O2 as the substrates of the purified enzyme were 21.0 and 9.5 μM, respectively. The calculated kcat value of the purified enzyme using Mn(II) as the substrate in 50 mM lactate buffer (pH 4.5) at 25°C was 6.7s-1, giving a kcat/Km value of 0.32 μM-1s-1. The kcat value for the MnP-catalyzed reaction was found to be dependent of the Mn(III) chelator molecules malonate, lactate and oxalate, indicating that the enzyme oxidized chelated Mn(II) to Mn(III). The pH and temperature optima of the enzyme were 4.5 and 25°C, respectively. The enzyme in combination with H2O2 liberated bromine and iodine in presence of KBr and KI respectively. All these enzymatic characteristics were similar to those of fungal MnP. The enzyme has the potential as a green brominating and iodinating agent in combination with KBr/KI and H2O2.
Subject(s)
Catalysis , Chromatography, DEAE-Cellulose , Enzyme Stability , Halogenation , Hydrogen-Ion Concentration , Kinetics , Molecular Weight , Musa/enzymology , Oxidation-Reduction , Peroxidases/chemistry , Peroxidases/isolation & purification , Peroxidases/pharmacokinetics , Plant Extracts/isolation & purification , Plant Proteins/chemistry , Plant Proteins/isolation & purification , Plant Proteins/pharmacokinetics , Plant Stems/enzymology , Spectrophotometry, Ultraviolet , Substrate Specificity , Temperature , UltrafiltrationABSTRACT
La extrusión se emplea para producir expandidos (botanas o snacks), crujientes y quebradizos. El impacto nutricional de este tipo de proceso no ha sido suficientemente estudiado, por ello se evaluó la biodisponibilidad in vitro e in vivo de la proteína y el almidón en mezclas de harinas de maíz (Zea mays) (M) y de frijol lima (Phaseolus lunatus) (F), tanto crudas como extrudidas. Las mezclas de harinas 75M/25F y 50M/50F (p/p) y fueron procesadas en un extrusor Brabender a 160°C, 100 rpm, y 15,5% de humedad. La composición proximal indicó un aumento del tenor de proteína y cenizas y una disminución de la grasa en los productos extrudidos. La digestibilidad in vitro de la proteína fue mayor en los extrudidos (82%) que en las harinas crudas (77%). Los contenidos de almidón potencialmente disponible y almidón resistente total disminuyeron con la extrusión. Los ensayos in vitro indican que la extrusión mejora la digestibilidad de la proteína y el almidón en las mezclas estudiadas. La biodisponibilidad in vivo se evaluó utilizando gorgojos de arroz (Sithophilus oryzae) como modelo biológico. Los biomarcadores más descriptivos de los cambios sugeridos por las pruebas in vivo fueron: el contenido de proteínas corporales, que incrementó por efecto de la extrusión, y la actividad de la alfa-amilasa intestinal, que disminuyó a consecuencia del procesamiento. Se concluye que la extrusión mejora apreciablemente la calidad nutricional de mezclas de maíz y frijol de lima.
Effect of extrusion on protein and starch bioavailability in corn and lima bean flour blends. Extrusion is used to produce crunchy expanded foods, such as snacks. The nutritional impact of this process has not been studied sufficiently. In this study, in vitro and in vivo protein and starch bioavailability was evaluated in both raw and extruded corn (Zea mays)(C) and lima bean (Phaseolus lunatus)(B) flour blends, prepared in 75C/25B and 50C/ 50B (p/p) proportions. These were processed with a Brabender extruder at 160°C, 100 rpm and 15.5% moisture content. Proximate composition showed that in the extruded products protein and ash contents increased whereas the fat level decreased. In vitro protein digestibility was higher in the extrudates (82%) than in the raw flours (77%). Potentially available starch and resistant starch contents decreased with extrusion. The in vitro assays indicated that extrusion improved protein and starch availability in the studied blends. In vivo bioavailability was evaluated using the rice weevil (Sithophilus oryzae) as a biological model. The most descriptive biomarkers of the changes suggested by the in vivo tests were body protein content (increased by extrusion) and intestinal alpha-amylase activity (decreased by processing). Overall, results suggest that extrusion notably increases the nutritional quality of corn and lima bean flour blends.
Subject(s)
Animals , Flour/analysis , Food Handling/methods , Phaseolus/chemistry , Plant Proteins/analysis , Starch/analysis , Zea mays/chemistry , Biological Availability , Chemistry, Physical , Models, Biological , Plant Proteins/pharmacokinetics , Starch/pharmacokinetics , Weevils/chemistry , Weevils/enzymologyABSTRACT
Histological studies of the spleen and thymus of rats fed raw black beans (Phaseolus vulgaris) show an atrophy of both lymphoid organs. Decrease in relative thymus weight was most marked. All histological organization of this organ appeared altered. An evident decrease in cell number was also observed in both organs. Proliferative response of splenic cells stimulated in vitro with Concanavalin A was increased as compared to that from animals fed the control diet. It is likely that histological changes observed in the spleen and the thymus are due mainly to a protein caloric deficiency, although the possibility that toxic factors present in the raw diet have an effect on the immune system of the rat can not be overruled