Effect of apotransferrin, lactoferrin and ovotransferrin on the hydroxyl radical mediated degradation of beta-glucan.

Beta-glucan is a polysaccharide widely accepted and used as a functional ingredient due to its positive effects on human health. However, beta-glucan is readily degraded in aqueous systems in presence of a hydroxyl radical generating system such as ascorbic/iron(II). In the present study, we tested whether iron binding proteins; apotransferrin, lactoferrin and ovotransferrin; could prevent the hydroxyl radical mediated degradation of beta-glucan. The radical formation was investigated by ESR spectroscopy and the polysaccharide degradation was monitored by the viscosity loss of the solutions. Apo-transferrin increased the formation of hydroxyl radicals and this related with a faster degradation of beta-glucan. Lactoferrin did not have any effect on the ascorbate induced degradation of beta-glucan, whereas ovotransferrin completely inhibited the hydroxyl radical generation by a system containing ascorbic acid and iron(II). However, the presence of ovotransferrin in beta-glucan decreased the viscosity of the solution, which was accompanied by the apparition of a precipitate, indicating a potential interaction between the protein and beta-glucan. FT-IR analyses indicate the presence of beta-glucan and ovotransferrin in both precipitate and supernatant, as well as the occurrence of interactions between the two compounds. This study reveals that ovotransferrin is a promising candidate for inhibiting the formation of ascorbate/iron(II) induced hydroxyl radicals in beta-glucan solutions.

Iron(II) binding by cereal beta-glucan.

Beta-glucan is a dietary fiber, which possesses several health benefits, such as cholesterol lowering, however this fiber is easily degraded in the presence of Fenton reagents. In the present study, the iron binding capacity of oat beta-glucan and barley beta-glucan was evaluated by investigating the kinetics of the Fenton reaction at pH 2.7 and 4.7 using stopped flow spectroscopy. The rate constant of the Fenton reaction is not affected by the presence of the beta-glucans in a solution pH 2.7, hence none of the beta-glucans bind iron(II) at this pH. However, at pH 4.7, the kinetics of the Fenton reaction vary between acetate buffer (k=2.8×10(2)M(-1)s(-1)), barley beta-glucan (k=2.2×10(2)M(-1)s(-1)) and oat beta-glucan (k=1.2×10(2)M(-1)s(-1)), which demonstrates that barley beta-glucan and oat beta-glucan form complexes with iron(II). Moreover, oat beta-glucan shows a stronger affinity for iron(II) than barley beta-glucan, and may thereby reduce the formation of hydroxyl radicals and diminish the rate of viscosity loss of the oat beta-glucan solution, as shown by the ESR and rheological data. The results presented in this study suggest that cereal beta-glucans can potentially reduce the bioavailability of iron.

Modulating the structural properties of ß-D-glucan degradation products by alternative reaction pathways.

The aim of the present study was to compare the degradation of ß-D-glucan induced by hydroxyl radical to the degradation induced by heat treatment. ß-D-Glucan was quickly and widely degraded by the action of hydroxyl radicals produced by a Fenton system at 85 °C, while thermal hydrolysis at 85 °C induced slow ß-D-glucan depolymerization. The hydroxyl radical-induced degradation of ß-D-glucan was accompanied by the formation of peroxyl radicals and new oxidized functional groups (i.e. lactones, carboxylic acids, ketones and aldehydes), as detected by ESR and NMR, respectively. In contrast, no changes in the monomer chemical structure of ß-D-glucan were observed upon thermal hydrolysis. Therefore, different mechanisms are proposed for the oxidative cleavage of ß-D-glucan, which are initiated by the presence of an unpaired electron on the anomeric carbon.

Reactive oxygen species responsible for beta-glucan degradation.

The presence of iron(II) in beta-glucan in solution causes the formation of hydroxyl radical, which further oxidises the polysaccharide. This degradation can be enhanced by the presence of a reducing agent, such as ascorbic acid. In this study we investigated the effect the iron(II) concentration on the hydroxyl radical-mediated degradation of beta-glucan and identified the intermediate species involved in the formation of hydroxyl radicals. An increase in the iron(II) concentration did not have a significant effect on the degradation in the presence of a reducing agent (ascorbic acid), while in the mere presence of iron(II) it accelerates the degradation. The addition of catalase and superoxide dismutase (SOD) prevented the hydroxyl radical driven-degradation of beta-glucan induced by iron(II) or ascorbic acid/iron(II), demonstrating the involvement of both superoxide and hydrogen peroxide in the hydroxyl radical formation. SOD, which catalyses the dismutation of superoxide into hydrogen peroxide, should have stimulated the formation of radicals, since these radicals are generated from the reaction between hydrogen peroxide and iron(II). In the present study, we hypothesise the mechanism of the inhibition of beta-glucan degradation by superoxide dismutase.

Indirect, non-competitive amperometric immunoassay for accurate quantification of calpastatin, a meat tenderness marker, in bovine muscle.

A method is described for quantification of the beef tenderness marker, calpastatin, in meat samples by amperometric detection. Using a novel bovine recombinant partial calpastatin protein as standard antigen a low detection limit of 0.2 ng/mL was achieved. The influence of the complex matrix was minimised by heat pretreatment and dilution of the samples prior to detection of calpastatin. The relative error between the direct sample measurement and standard addition methods was 5.89%, confirming the accuracy of the developed amperometric immunoassay.