# Shellfish Chitosan Potential in Wine Clarification
## 3 Results and Discussion
### Chitosan Viscosity  
As is known, viscosity reflects molecular characteristics of chitosan, namely the molar mass and the surface charge [23; 27]. The molecular weight was calculated as reported by [24] and is reported in Table 1. An evident lower molecular weight was registered for MC, probably depending on the enzymatic treatment necessary to reduce the glucan content on the polysaccharide extract from fungi. Regarding SC, the calculated molecular weights were generally in agreement with those, when available, declared by the suppliers, except for the sample A9. As previously mentioned, DDA also reveals a strict correlation with chitosan viscosity, in addition to the distribution of charges that could play an important role, modifying conformational behavior of chitosan. [PERSON] and colleagues [28] suggested that animal and fugal chitosan could differ for CTS charge distribution. Hence, studied samples were evaluated for the viscosities expressed when dissolved in four different acids, namely acetic, malic, succinic, and hydrochloric acid. Previous experimental studies explored the effect that the dissolution acid could have on chitosan viscosity [19; 29]; however, that work did not compare several chitosans or chitosans of different origin. Statistical analyses revealed a significant difference among acid and categories, together with their interaction, as evidenced by Figure 4. The data highlight a different trend between animal and fungal chitosan upon the acid change. Moreover, shell-derived CTS manifested higher variability than the MC, which, as expected from their low calculated molecular weights, actually did not differ in viscosity from the respective controls. Figure 5 represents the time requested for the solutions to throw the glass capillary, which means that high values correspond to high viscosities. Figure 5 is focused on the animal chitosan behavior because this category evidenced the major variability. In all of the cases, CTSs revealed the highest density when dissolved in acetic acid. Figure 4: Chitosan viscosity. Mean and standard deviations (three replications for each sample) are expressed. Black bars: fungal-derived chitosan, light grey bars: animal-derived chitosan, white bars: corresponding acid solution (control). AA: acetic acid, MA: malic acid, SA: succinic acid, HCl: hydrochloric acid. Capital letters represent statistically significant differences among dissolving acids (\(p<0.05\), no letter means the absence of significance). Following [PERSON] and colleagues [23], the constant a used for the calculation of intrinsic viscosity n is described by the equation:  
\[\text{a}=[\text{DA}/(\text{pH}\ \mu)]\]  
where DA is the degree of acetylation, pH is the pH, and (\(\mu\)) is the solution ionic strength, demonstrating a direct relationship between the dissolution media and the chitosan viscosity. The differences recorded from the comparison of chitosan viscosities among the four dissolving acids confirmed this interaction between CTS and the dissolution system. As a matter of fact, while deacetylation could be influenced by the chitosan manufacturing as reported by [PERSON] and colleagues [30], and therefore could explain the differences among chitosan, it should be assumed as constant when comparing the same chitosan sample dissolved in different acids. As explained by [PERSON] et al. [23], low pH should lead to a higher degree of expansion of chitosan due to electrostatic repulsions, reducing the mobility of its structure, causing an increase in the viscosity. Unexpectedly, for each CTS, viscosities decreased following the pH lowering order (2.8, 2.6, 2.3, and 0.6 for acetic, succininic, malic, and HCl, respectively). However, it was also demonstrated that the intrinsic viscosity decreases with the increase of the ionic strength, as the chain became more flexible and compact with a reduction of the repulsive potential owing to the masking effect of anions [31].