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Abstract: The citrus processing industry aims to maintain turbidity and attractive colour of the juice obtained from Sicilian blood oranges. Nevertheless, the presence of pectinesterase (PE, E.C. 3.1.1.11) causes the loss of these peculiar characteristics, due to precipitation of colloids and very fine pieces of pulp in suspension, with negative effects on colour and clarification of the juices. This study compares total PE activity of Sicilian blood oranges (Sanguinello, Moro, Tarocco) with the blonde cultivar Navel, checking enzyme stability with various pasteurisation times and temperatures conditions. Decimal reduction time and temperature (D and z) as well as the kinetic constant (k) were established to optimise and increase the shelf-life of the pasteurised juice. Finally, a heat treatment (85°C×3min) of both microbiological and enzymatic efficacy has been developed that does not compromise anthocyanin stability; this treatment could be used by the citrus fruit processing industry as a valid alternative in the production of blood orange juices. [Copyright &y& Elsevier]

A comparison has been made of the main physicochemical and kinetic parameters of the β-glucosidase (βG, EC 3.2.1.21) of a commercial preparation from Aspergillus niger chosen for its high activity and that of 463 yeast strains, isolated and screened from Sicilian musts and wines. A higher percentage of βG-positive strains (30%) was found on Yeast Carbon Base + lysine compared to 18% on Sabouraud + ethanol + SO2 and just 8% on Sabouraud medium. The positive strains with the highest βG activity were subsequently identified and compared with the purified commercial preparation. Optimum pH values were found to be 4.5 for A. niger βG, with relative activity of about 87% at pH 3.5, that is close to that of wine, whilst amongst the yeasts, AL 41 was the strain with optimum pH closest to that of wine. Optimum temperatures were found to be about 62 and 20–25 °C for A. niger and yeast βG, respectively. Within the latter temperature range, that is potentially the most suitable for the use of the enzymes in enology, mould βG relative activity was found to be 40–50%. Enzyme stability in wine model solution (at 20 °C) was 12 days for mould activity and at least 35–45 for that from yeasts. Finally, the Vmax values were ranged from 1.75 to 6.94 U mg-l of protein for the yeasts, while the Km values were 0.61–2.53 mM. The behaviour of βG from A. niger could not be explained by the Michaelis–Menten equation. The case was ascribable to the typical behaviour of an unproductive bond with substrate concentration exceeding saturation point and incorrect positioning at the enzyme’s active site. The enzyme from mould would be useful during the final stages of alcoholic fermentation, whilst some yeast strains (AL 112 and AL 41) could be directly added during fermentation in order to enhance wine aroma. [Copyright &y& Elsevier]

A remarkable variability in the β-glucosidase activity was observed between 10 wild strains of Oenococcus oeni isolated from wines of the Valpolicella area. The activity occurred in both whole and sonicated cells (intercellular and parietal activities) but not in the supernatants of the cultures. The properties of β-glucosidase produced by DSTVR 7 and 10 strains include its activation in the presence of ethanol. These strains retained the enzyme activity at oenological conditions (pH value, temperature and ethanol) and showed good malolactic performance when inoculated in wine. None of the tested strains possessed anthocyanase activity. [Copyright &y& Elsevier]

Pectin esterase (PE, E.C. 3.1.1.11) is implicated in the cloud loss in citrus juices. This study describes a specific method of extracting pectin esterase from blood oranges and developing, contemporarily, an enzyme assay suitable for the pH of such juices. The PE showed an optimum extraction at pH 7.0, without addition of surfactants. The enzyme assay was carried out at pH 3.6, a condition typical of the orange juices, increasing sensitivity in determining PE compared to other methods proposed up until now. [Copyright &y& Elsevier]

Three hundred sixty-one yeast strains (80 of which ascribable to Saccharomyces cerevisiae) were isolated from Sicilian musts and wines with the purpose of looking for β-glucosidase (βG, EC 3.2.1.21) activity. Of these, the AL 41 strain had highest endogenous βG activity and was identified as belonging to the species S. cerevisiae by biochemical and molecular methods. This enzyme was subsequently characterized. It had optimum effect at pH 3.5–4.0, whilst optimum temperature was 20 °C, compatible with typical wine-cellar conditions; it was not inhibited by ethanol, at concentrations of 12–14%, or fructose and glucose. The βG was also characterised in terms of the kinetic parameters Km (2.55 mM) and Vmax (1.71 U mg−1 of protein). Finally, it remained stable for at least 35 days in model solutions of must and wine. [Copyright &y& Elsevier]

The AL 112 strain, isolated from 361 yeast strains in Sicilian musts and wines, has been identified by biochemical and molecular methods as belonging to Pichia anomala, and your endogenous β-glucosidase (βG, EC 3.2.1.21) subsequently characterised. This strain not only has extremely high specific productivity of βG, but above all shows arabinosidase (Ara, EC 3.2.1.55) activity, essential for aroma enhancement of wine. βG from Al 112 is activated by ethanol at the concentrations typically found in wine; it is not inhibited by fructose, whilst glucose, a non-competitive inhibitor, despite lowering activity, actually protects the enzyme from factors that could damage it. It has an optimum temperature of 20 °C, compatible with typical cellar conditions, and stability in model must-wine and wine solutions ≥40 days. [Copyright &y& Elsevier]