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BACKGROUND The development of bioactive compound delivery systems using protein‐based nanoparticles as carriers is a rapidly growing field of study. α‐Lactalbumin (α‐La) nanoparticles are adequate for this task because they are nonantigenic, biodegradable, and easily modifiable via the surface physical adsorption of other molecules. Here, the effect of poly‐L‐lysine (PLL) coating of α‐La nanoparticles on the stability against enzymatic degradation in the gastrointestinal tract was investigated. RESULTS α‐La nanoparticles were prepared by desolvation with acetone. It was observed that the pH of the initial protein suspension did not have a significant effect on the size, polydispersity index (PDI), or ζ‐potential of the nanoparticles, so the subsequent experiments were performed at pH 9, where 151.2 nm nanoparticles with a PDI of 0.072 and a ζ‐potential of −29.7 mV were obtained. The nanoparticles coating was performed with two MW‐range PLL at three concentrations. Coating efficiency increased as the polymer concentration increased and this was independent of the PLL MW. The best coating efficiency (90–95%) was obtained in the case of the 1 mg mL−1 PLL coating. Most PLL‐coated nanoparticles showed a good stability to the conditions of in vitro simulated gastrointestinal digestion compared with the uncoated nanoparticles. CONCLUSION It can be concluded that the acetone desolvation method at pH 9 is adequate to obtain α‐La nanoparticles with good characteristics. The process of PLL coating of nanoparticles showed an increase in efficiency with the increase in concentration of the polymer. The resulting coated nanoparticles had enhanced resistance to simulated gastrointestinal conditions. © 2021 Society of Chemical Industry (SCI).