Double-emulsion technique applied to laccase immobilization on polymeric nanoparticles

One primary drawback of enzyme catalysis at industrial scale is the short term service life of the enzymes. Enzymes lose their activity due to oxidation or other processes which results in less stability and a shorter lifetime thereby rendering them less efficient. An effective way to increase the stability, longevity and reusability of the enzymes is to attach them to nanoparticles by applying the double-emulsion technique. In this work the polymer Eudragit® L 100-55 sensitive to pH was used to prepare laccase polymeric nanoparticles by the double-emulsion solvent evaporation approach. The size and morphology of the nanoparticles obtained was evaluated by Scanning Electron Microscope and particle size distribution was assessed by Photon Correlation Spectroscopy. Encapsulation efficiency and zeta potential were calculated. The effect of pH on laccase activity and stability was compared between free laccase and the immobilized one. Their stability was also determined in a sequential assay involving acidic pHs up to alkaline ones. The nanoparticles had a spherical shape with a mean size of 289 nm, zeta potential of -22.7 mV at pH 7.0, and load efficiency of 87%. The optimum pH of both free and immobilized laccases was 3.0, being the nanoparticles more stable at acidic pHs (2.0-4.0). Howevwe, this last kept 80% of enzyme activity at pH 2.0 approx., after 24 h. The polymer Eudragit® L 100-55 also conferred them resistance towards the pHs usually found at the gastrointestinal tract. These results suggest the potential use of nanoparticles as adjuvants in animal feed, serving as carriers for oral laccase delivery.