Paolo Spettoli

Biodegradation of phenols by laccase immobilised in a membrane reactor

Abstract The results of a study of immobilisation of a commercial laccase onto a spiral-wound asymmetric polyethersulphone membrane are described. The immobilised enzyme system displayed a promising half-life of more than 150 h for oxidation of syringaldazine. The laccase membrane reactor was applied to the biodegradation of a model phenol solution containing 18 phenolic substrates, including chlorophenols, cresols and methoxyphenols. It has been confirmed that the type and/or the position of substituent group affect the level of substrate oxidation.

Enhanced microbial cell lysis by the use of lysozyme immobilized on different carriers

Abstract Lysozyme was immobilized on chitosan and silica gel by adsorption, on cross-linked polystyrene divinylbenzene matrix (Deacidite KMP) by ionic binding and on non-porous glass beads by covalent attachment to improve the lytic activity towards complex substrates such as Micrococcus luteus . The lysozyme immobilization yields decreased on decreasing the amount of enzyme loaded, whereas the activation yields (defined as percentage ratio of immobilized active enzyme to immobilized enzyme) increased. Chitosan, silica gel, Deacidite KMP and non-porous glass bead preparations showed activation yields of 10%, 13·20%, 26·67% and 12·77%, respectively.

Production of a bacteriocin active on lactate-fermenting clostridia by Lactococcus lactis subsp. lactis immobilized in coated alginate beads

We report the isolation and immobilization of a nisinogenic strain (NZ1) of Lactococcus lactis subsp. lactis , active on gas-forming lactate-fermenting clostridia responsible for late blowing of Asiago and Montasio cheeses. The bacteriocin (nisin) produced by strain NZ1 is pronase-sensitive and is released in culture media during the growth phase. Using the sensitive indicator strain Lactobacillus delbrueckii subsp. bulgaricus NCDO 1489, a rapid microtitre plate based assay was developed for quantitative determination of the bacteriocin produced by NZ1 cells, either free or immobilized in gel beads. Scanning electron microscopy of cells immobilized in calcium alginate coated beads and viable counts of the surrounding medium showed that no cell leakage occurred during a 24 h assay. The bacteriocin released from immobilized cells reached, after 5 and 24 h, concentrations comparable to that of the free cell system after 3–4 h incubation in culture media.

Immobilized Laccase for Must and Wine Processinga

Whereas the use of enzyme preparations in the food industry is a wellestablished, rapidly expanding technology, their utilization in enology is less frequent. The “classic” wine industry is in fact still largely based on traditional methods. Moreover, musts and wines represent a hostile environment for purified enzymes, i.e., low pH, high ethanol concentration, presence of sulfur dioxide and tannins. White and ‘‘rosk” wines are known to be more sensitive than red wines to browning and to the development of madeirized flavor, which are mainly due to the presence of polyphenols. Aiming at the replacement of traditional stabilization treatments (i-e., sulfur dioxide, fining agents) with mild technologies to prevent madeirization, unconventional systems like the use of oxidative enzymes (chiefly laccase) have been proposed.’,* However, the residual enzyme activity in wine during storage or, in the case of musts, its inactivation by heat treatment could lead to negative effects, i.e., color browning. Therefore, the use of immobilized oxidative enzymes that can be easily recovered from the reaction mixture and reused could be exploited. Studies have been reportedly carried out on fungal laccase immobilized by covalent attachment to CNBr-activated Sepharose 4B and by adsorption to Concanavalin A-Sepharose? covalently bound to a diatomaceous silica product (R-650 Celite): immobilized on porous glass beads5s6 to detoxify the environment polluted with aromatic and phenolic compounds of agricultural or industrial origin. Taking into account our previous trials on the approach to the use of immobilized oxidative enzymes in white wine stabilization,7vu in the present work we report the results on a commercial laccase (p-diphenol: oxygen oxidoreductase; EC 1.10.3.2) immobilized by adsorption on silica gel carrier followed by treatment with glutaraldehyde and by covalent binding on polymer carrier VA-Hydroxy Biosinth, and CNBractivated Sepharose 4B, respectively. In addition, the possibility of an enzymatic removal of phenolics was tested in model solution, in must, and in wine.

Detection of Saccharomyces cerevisiae carboxylesterase activity after native and sodium dodecyl sulfate electrophoresis by using fluorescein diacetate as substrate.

A simple method for the visualisation of wine yeast esterase (carboxylesterase EC 3.1.1.1) activity on electrophoretic gels was developed, using the fluorescent substrate fluorescein diacetate. The zymogram system allows a sensitive detection of esterase bands in only 5 min of incubation of both native and sodium dodecyl sulfate gels.

Chemical parameters, biologically active polyphenols and sensory characteristics of some Italian organic wines

Abstract This paper describes 23 commercial Italian wines, made from organic grapes, by their chemical parameters, biologically active polyphenol concentrations and sensory analysis. Our data show inconsistent differences in the trans-resveratrol and p-coumaric acid contents in comparison with non-organic wine, with the exception of quercetin concentrations in Chianti samples. Furthermore, the chemical analyses and sensory data of organic wines did not significantly differ from those previously cited. In general the organic wines display satisfactory sensory properties such as flavour intensity, body and a general acceptance.

Chemical composition of papaya seeds

Seeds of papaya cultivated in Somalia, which accounted for about 16% of the fresh fruit weight, were divided into sarcotesta and endosperm. Sarcotesta showed higher percentages of ash, crude protein, and crude fiber than did endosperm, but was lacking in fat. In contrast, endosperm contained 60% fat. Oil extract showed very high levels of oleic and palmitic acids. The essential amino acid profiles of endosperm and sarcotesta protein were determined and compared with the provisional FAO essential amino acid profile. The results indicated that the endosperm protein was a good potential source of supplemental protein.

Chemical and enzymatic changes in strongly damaged beets

Abstract Damage caused by skips during loading and unloading may influence beet quality during the first few hours following the damage. Sucrose content decreases sharply during the first 24 h and (together with acid and neutral invertase) sucrose synthetase seems to be metabolically active in sucrose hydrolysis after harvest. These data are important since, following factory processing, losses could be considerable by the end of the campaign.

Affinity chromatography of malic enzyme from grape berries

Abstract NADP-dependent malic enzyme from grape berries is associated with NAD-dependent malate dehydrogenase. A two step procedure, involving affinity chromatography on 2′,5′-ADP-Sepharose 4B, followed by gel- permeation on Bio-Gel A- 1.5 m, was used to separate malic enzyme from malate dehydrogenase and other proteins. The yield was ca 60% Malic enzyme and malate dehydrogenase migrated respectively as three bands and one band during disc electrophoresis in polyacrylamide gel. The MW resulting from gel-permeation was 220 000 for malic enzyme and 53 000 for malate dehydrogenase.

Activity and multiplicity of Glutamic Dehydrogenase in sugar-beet

Sixty roots of a diploid and sixty of a tetraploid sugar-beet variety were individually characterized for the fresh weight, sugar content, Glutamic Dehydrogenase (GDH) activity and multiplicity as revealed by the electrofocusing technique. Tetraploid roots showed significantly higher fresh weight (P<0.01), lower sugar content (P<0.01) and lower GDH activity (P<0.05) in comparison with diploids. Within each ploidy status families with similar GDH isoenzyme profile showed a negative correlation between GDH activity and sugar content in the case of diploids while in tetraploid such a correlation was absent. GDH multiplicity failed to show any significant correlation with sugar content in both diploid and tetraploid varieties.