## 4 Functional Properties of Chitosan and Derivatives  
### Sedimentation and Flocculation in the Wine Industry  
Chitin and CS have been allowed by the Codex Alimentarius since 2003 as coaguling/clarifying agents for fruit juices and nects. Fungal CS extracted from _Aspergillus niger_ is the only type of CS allowed in winemaking, since 2009, as specified by the Oenological Codex (OIV-OENO 368-2009). The process from which CS is obtained from chitin in fungi is protected by a patent [23] and its origin is guaranteed according to OIV-OENO 368-2009 by the three following properties: _(i)_ residual glucans have to be lower than 2%; _(ii)_ viscosity in 1% acetic acid has to be higher than 15 Cps; and _(iii)_ the settled density has to be lower than 0.7 g/cm\({}^{3}\). CS is a flexible polymer with several functional groups (amine, N-acetamide, and hydroxyl groups, as seen in the previous sections), which makes it a very reactive molecule in wine. Therefore, it has numerous potential applications in oenology, \begin{table}
\begin{tabular}{c c c c c} \hline \hline
**Type of Method** & \
\begin{tabular}{c} **Depolymerization** \\ **Methods** \\ \end{tabular} & **Conditions** & **Mw(1) DP(2)** & **References** \\ \hline \multirow{8}{*}{**PHYSICAL**} & High pressure & 1500 bars 1\% CS in 1\% & \\  & homogenization & acetic acid & 30 kDa & [68] \\ \cline{2-5}  & Sonication &
\begin{tabular}{c} Sonication at 35.2 W/cm\({}^{2}\), \\ 30 min \\ \end{tabular} & 140–143 kDa & [26] \\ \cline{2-5}  & Gamma radiations & 2\% CS in 2\% acetic acid, 200 KGy & 3–5 kDa & [60] \\ \cline{2-5}  & & 1\% CS, 0.1\% Tween 80 & \\  & & irradiation 50 kGy & 75–77 kDa & [61] \\ \cline{2-5}  & Autoclave & 1\% CS, 1\% acetic acid, 121 \({}^{\circ}\)C, 60 min, 1 bar & 313 kDa & [62] \\ \hline \multirow{8}{*}{**CHEMICAL**} & & 0.5 M HCl, 1\% CS, 30 h, 65 \({}^{\circ}\)C & - & [56] \\ \cline{2-5}  & Acid hydrolysis & 2\% CS, 1.8 M HCl reflux 100 \({}^{\circ}\)C, 2 h & DP \textless{} 40 & [70] \\ \cline{2-5}  & & 0.976\% CS, 50 mM HCl, 3.89 mM HNO3, 35 \({}^{\circ}\)C, 30 min & \textless{} 16 kDa & [57] \\ \cline{2-5}  & & 1\% CS in HCl 1.8 M, 100 \({}^{\circ}\)C, 2 h & DP \textgreater{} 6 & [57] \\ \cline{2-5}  & Free radical & 2\% CS, 2\% acetic acid, 1.5\% & H\({}_{2}\)O\({}_{2}\) (final) pH 3.0, 6 h & 9.9 kDa & [58] \\ \cline{2-5}  & methods & 1.5\% CS in 2\% acetic acid & 1.74 kDa & [59] \\ \cline{2-5}  & & solution, 1.08 g KPS, 70 \({}^{\circ}\)C & & \\ \hline \multirow{8}{*}{**ENZYMATIC**} & & Chitosanse from & & \\  & Specific enzymes & _Aspergillus_ sp. UU 15.5\% & DP \textless{} 10 & [64] \\ \cline{2-5}  & & CS solution 45–50 \({}^{\circ}\)C, 68 h & & \\ \cline{2-5}  & & Chitinase from _Aeronomas_ & DP 1 to 5 & [63] \\ \cline{2-5}  & & _h hydrophilic_ & & \\ \cline{2-5}  & Non-specific & 1\% CS in 100 mM sodium & & \\  & enzymes & acetate pH 4 with 1:100 & 9–13 kDa & [65] \\ \cline{2-5}  & Non-specific & Pepsin ratio, 2 h & & \\  & enzymes & 4\% CS 1\% acetic acid 50 \({}^{\circ}\)C & & \\  & & E/S protease ratio 1:20 & DP 1 to 8 & [67] \\ \cline{2-5}  & & 4.5\% CS in 0.5 M acetic & & \\  & acid bicarbonate pH 5.6, & DP 3 to 8 & [66] \\ \cline{2-5}  & cellulase, 50 \({}^{\circ}\)C, 14 h & & & \\ \hline \hline \end{tabular} \({}^{(1)}\) Mw: Molecular Weight and \({}^{(2)}\) DP: Degree of Polymerization.  
\end{table} 
Table 1: Methods reported for producing LMW chitosan or COS.  
and is allowed for fining must or wines (OIV-OENO 336A-2009 and 337A-2009) up to a maximal dose of 100 g/hL, but also for treating wines to remove the following contaminants (OIV-OENO 338A-2009): (_i_) ochratoxine A (up to a treatment limit of 500 g/hL) and (_ii_) iron, lead, cadmium, and copper (maximum dose: 100 g/hL), and finally to reduce the main wine spoilage yeast populations, _Brettanomyces_ (maximum dose: 10 g/hL) [72]. Even though most CS is soluble in most organic acid solutions [73], it is not entirely soluble in wine. The sediment formed after CS treatment should be removed by tracking.