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In this study, ethylcellulose/hydroxypropylmethylcellulose/ethylcellulose (EC/HPMC/EC) three-layer films including Nisaplin® or nisin and lecithins were formulated. Lecithins were used as plasticizers to ensure cohesion between hydrophobic ethylcellulose and hydrophilic HPMC layers. It was observed that the introduction of pure nisin or its non-pure commercial form Nisaplin® into films didn't significantly alter their mechanical and optical properties. Additionally, these nisin or Nisaplin-loaded multilayer films showed significant antimicrobial activity. The comparison of inhibition diameters obtained with EC/HPMC film used as control and EC/HPMC/EC films demonstrated that the three-layer films delayed nisin desorption. This was confirmed by the kinetics of nisin release in a (0.8% w/v) NaCl solution at 28°C: nisin from two-layer EC/HPMC films totally desorbed within 0.5h, while the three-layer films allowed to …