2.2.2. Water Solubility

The highest water solubility was observed in pure collagen films (32.14%) and decreased significantly (*p* < 0.05) with the incorporation of increased chitosan percentage (24.55% and 17.64% for the C50 and C75 films respectively; Table 3).

**Table 3.** Color properties and solubility of collagen films where L\*: luminance/brightness, a\*: red/green, b\*: yellow/blue and ΔE\*: total difference in colour.


All values are mean ± standard deviation; a–d different superscripts in the same column indicate significant differences (*p* < 0.05). CO: pure collagen film; CH: pure chitosan film; C50: Collagen-chitosan film 50%:50%, C75: Collagen-chitosan film 25%:75%.

Actually, the film's resistance to water is owed to the hydrophobic nature of chitosan molecule and to the covalent bond "amide bond" which has the effect of reducing the polarity of the films [54].

Indeed, an edible film must have both good resistance to water in order to preserve the integrity of the product [69] and a good ability to dissolve when ingested by the consumer and degrade naturally if it released into the environment [70].

However, the increase of water resistance of composite films could not be perceived as an advantage since high solubility cannot shield the product from humidity and water loss [71].

#### 2.2.3. Optical Properties—Colour, Opacity and Light Transmittance of the Films

The film colour is a key element in the consumer's appreciation of the product since this parameter has a direct influence on the product appearance, especially when the film is to be used for packaging. For the various elaborated films, the highest (*p* < 0.05) L\*-value (Lightness) and a\*-value (redness/greenness) and the lowest b\*-value (yellowness) and ΔE\* (colour difference) were recorded in pure collagen films (Table 2). Thus, incorporation of chitosan induced a significant decrease in the lightness (*p* < 0.05), particularly in the films with the highest concentration of chitosan C75, making them more yellowish. This may be due to the reaction of Maillard which took place between the carbonyls groups of chitosan and collagen amino groups [72].

The highest b\*-value was observed in pure chitosan film (CH) and as described by Kurek et al. [73], this parameter (b\*-value) defines the natural colour of chitosan, Yellow, which is related to the presence of β-1-4 linked 2-amino-2-deoxy-D-glucopyranose repeating units [74].

In addition to the colour of the film, transparency is also a very influential parameter in relation to the acceptability of the product. Generally, a clear film is more attractive clearly displaying the contents of the product. For all films, the light transmission in UV-Visible range was negligible at 200 nm, regardless of types and concentrations of chitosan. Collagen film exhibit the highest transmission at 280 nm but after the addition of chitosan, transmission decreased from 72.6 % for collagen pure film to 63.3% and 50.6% for composite films C50 and C75 respectively (*p* < 0.05).

The results show that when increasing the concentration of chitosan, composite films have better UV barrier properties. For instance, it makes these collagen-chitosan films usable as preventive materials against loss of nutrients and discoloration caused by the lipid oxidation [75]. The transmission of visible light at 400–800 nm, was superior to 80% in pure collagen film (CO), and was significantly higher (*p* < 0.05) than that of the collagen-chitosan composite films (Figure 7).

**Figure 7.** Optical transmission spectra of collagen films. CO: pure collagen film; CH: pure chitosan film; C50: Collagen-chitosan film 50%:50%, C75: Collagen-chitosan film 25%:75%.
