*3.2. Homogeneity Evaluation*

The homogeneity of the performed treatment was investigated by ATR-FTIR analysis in five points on each side of tissue samples. This analysis was carried out only on tissues treated with the highest concentration of the active molecules to have enough intense IR signals to be detected.

Figures 6 and 7 show the spectra acquired for the tissues treated, respectively, with chitin and polyphenols. The spectrum of the untreated paper tissue was also considered as a reference, in order to select the bands that could reveal the presence of the active

compounds in the coating. The intense signals appearing at 1650 cm-1 and 1615 cm-1 (related to amide groups) were, respectively, used to identify the presence of antioxidant polyphenols and chitin on the surface of the tissues. Similar intensities were obtained for this band in all five points analysed of each side, denoting a good homogeneity of the treatment. The comparison between the top and bottom (opposite side) parts of each tissue evidenced the prevalence of the active molecules on the top (application side), but they were detectable also on the bottom (opposite side). This difference was more evident in the chitin-treated tissue than in the polyphenols one, indicating that this latter had a higher penetration in the tissue. This phenomenon can be attributed to the similar molecular structure of chitin and cellulose (they both are polysaccharides) compared to polyphenols, to the lowest molecular weight of polyphenols with respect to the chitin, and to the fact that ChNFs were dispersed as a solid suspension in water, while polyphenols were at least partially dissolved. Hence, ChNFs were applied mainly on the application side where nanofibrils tend to be deposited forming a superficial tissue.

**Figure 6.** Spectra of the top and bottom part of chitin-treated tissues compared with IR spectrum of pure chitin (red). Capital letter on each spectrum indicate the point where it was acquired according to the relative picture on its left.

**Figure 7.** Spectra of the top and bottom part of antioxidant-treated tissues compared with IR spectrum of pure antioxidant extract (red). Capital letter on each spectrum indicate the point where it was acquired according to the relative picture on its left.
