Synergistic Effect of Laccase and Sugar Beet Pectin on the Properties of Concentrated Protein Emulsions and Its Application in Concentrated Coconut Milk.

Concentrated coconut milk (CCM), a raw material from coconut products, is extremely unstable because of its high oil content (>30%). In this study, three model emulsions-primary emulsions stabilized by coconut proteins only, secondary emulsions stabilized by the conjugation of sugar beet pectin (SBP) and coconut protein, and laccase-treated secondary emulsions-were prepared to investigate the effects of different factors (coconut proteins, coconut proteins + SBP, laccase-treated emulsions) on the stability of model emulsions and the application of this method to real CCM. The stability of the emulsions was evaluated based on their interfacial tension, zeta potential, particle size distribution, rheological properties, and the assembly formation of SBP and coconut protein at the oil⁻water interface. Results showed that addition of SBP or laccase can increase the viscosity and reduce the interfacial tension of the emulsion, and the effect was concentration dependent. Zeta potential of the emulsion decreased with the increase of protein (from -16 to -32 mV) and addition of SBP (from -32 to -46 mV), and it was reduced when laccase was added (from -9.5 to -6.0 mV). The secondary emulsion exhibited the narrowest particle size distribution (from 0.1 to 20 µm); however, laccase-catalyzed secondary emulsions showed the best storage stability and no layering when the laccase content reached 10 U/100 g. Confocal laser scanning microscopy (CLSM) revealed that protein was adsorbed on the oil⁻water interface and SBP distributed in the continuous phase could undergo oxidative crosslinking by laccase. These results show that the stability of the concentrated emulsion can be effectively improved by adding SBP and laccase.

A laccase with antiproliferative activity against tumor cells from an edible mushroom, white common Agrocybe cylindracea.

A laccase, with HIV-1 reverse transcriptase inhibitory activity (IC(50)=12.7 µM) and antiproliferative activity against HepG2 cells (IC(50)=5.6 µM) and MCF7 cells (IC(50)=6.5 µM), was purified from fresh fruiting bodies of the edible white common Agrocybe cylindracea mushroom. The laccase, which had a novel N-terminal sequence, displayed a molecular mass of 58 kDa within the range reported for most other mushroom laccases. The purification protocol entailed ion exchange chromatography on DEAE-cellulose, SP-Sepharose, and Q-Sepharose and gel filtration on Superdex 75. The laccase was adsorbed on DEAE-cellulose and Q-Sepharose, but unadsorbed on SP-Sepharose. Its optimum pH was pH 3-4 and its optimum temperature was 50°C. The activity of the isolated laccase differed from one substrate to another. The ranking was ABTS>N,N-dimethyl-1,4-phenylenediamine>hydroquinone>catechol>2-methylcatechol>pyrogallol.

Inhibition of cytokine-induced ß cell apoptosis via laccase and its therapeutic advantages for insulin-dependent diabetes mellitus, type 1 diabetes.

In pancreatic islets, free radical formation produced upon exposure to proinflammatory cytokines mediates ß cell destruction, which ultimately leads to type 1 diabetes (T1D). In this study, we examined whether laccase, a family of the blue copper protein, can be successfully used to prevent ß cells from cytokine-mediated apoptosis. Non-obese diabetic (NOD) mice were used for these experiments. In parallel, the RINm5f ß cell line was employed as a model system for in vitro experiments. The results demonstrated that laccase effectively scavenged peroxinitrite, which can be formed by nitric oxide, and upregulated the expression of antioxidant enzymes, such as manganese superoxide dismutase (MnSOD) and catalase. Interestingly, laccase balanced pro- (Bax) and anti-apoptotic (Bcl-2) proteins in terms of both the mRNA and protein levels with a downregulation of cytochrome c protein in RINm5f cells. In addition, laccase maintained blood glucose concentrations at a normal level with a simultaneous increase in plasma insulin levels during the spontaneous induction of diabetes in NOD mice. In conclusion, the antioxidant potentials of laccase in scavenging free radicals and upregulation of antioxidant enzymes may exert its pro-survival effect by counteracting the increased intracellular oxidative stress, and, consequently, by inhibiting apoptosis induced by cytokine-mediated activation during the course of T1D.