Ensuring the longevity of wooden constructions depends heavily on the preservation process. However, several traditional preservation methods involving fossil-based compounds have become outdated because they pose a significant risk to the environment and to human health. Therefore, the use of bio-based and bioactive solutions, such as essential oils, has emerged as a more sustainable alternative in protecting wood from biotic attacks. The entrapment of essential oils in polymeric carrier matrices provides protection against oxidation and subsequent degradation or rapid evaporation, which implies the loss of their biocidal effect. In this work, lignin as well as PLGA nanoparticles containing the essential oils from two different thyme species (Thymus capitatus and T. vulgaris) were applied on beech wood samples using spray coating. The prepared coatings were investigated using FTIR imaging, SEM, as well as LSM analysis. Release experiments were conducted to investigate the release behavior of the essential oils from their respective lignin and PLGA carrier materials. The study found that lignin nanoparticles were more effective at trapping and retaining essential oils than PLGA nanoparticles, despite having larger average particle diameters and a more uneven particle size distribution. An analysis of the lignin coatings showed that they formed a uniform layer that covered most of the surface pores. PLGA nanoparticles formed a film-like layer on the cell walls, and after leaching, larger areas of native wood were evident on the wood samples treated with PLGA NPs compared to the ones coated with lignin NPs. The loading capacity and efficiency varied with the type of essential oil, while the release behaviors were similar between the two essential oil types applied in this study.