# Chitin and Chitosan in the Alcoholic and Non-Alcoholic Beverage Industry: An Overview
## 4 Chitosan Immobilization
### Chitosan Enzyme Immobilization
[152] immobilized lysozyme from hen egg white (HEWL) on chitosan beads to develop a system for the continuous, and food-grade enzymatic lysis of lactic acid bacteria (_Oenococcus oeni_) in white and red wines, thus limiting the use of sulfur dioxide required to control malolactic fermentation (MLF). HEWL appeared more effective in the immobilized than in the free form, suggesting that covalent immobilization renders the enzyme less sensitive to the inhibitory effect of wine flavans. Another use is to protect consumers from undesirable fermentation by-products, such as higher alcohols (ethanol, 1-propanol, isobutanol, 1-butanol, isoamyl alcohol, and 1-hexanol) [153]. [PERSON] et al. [153] used apple crude enzymes immobilized in sodium-alginate and chitosan to decrease the higher alcohols content of Chinese liquors with 45-56% of ethanol. Significant degradation rates of higher alcohols were observed at different degrees. [PERSON] et al. [154] immobilized triethylene tetramine-modified water-insoluble corn flour, in magnetic chitosan resin (TETA-WICF/MCR) aiming to eliminate the mycotoxin Pattulin (biosynthesized by certain fungi that contaminate agricultural commodities) from apple juice to improve biocatalytic features. Such is the case of pectinases immobilized in chitosan. Pectinase has become more stable, robust, and can be recoverable [155; 156]. In the work of [PERSON] et al. [157], a composite system, based on high amounts of montmorillonite and low amounts of chitosan, was proposed as potential alternatives to the free nano clay particles, commonly used as carriers for enzyme covalent immobilization. The chitosan was used to make the clay's appropriate supports for the protease bromelain and helped to maintain the clay particles together in a unique structure, acting as a binder. The same authors [158] also performed the immobilization of two enzymes (bromelain and a commercial pectinase-Pectinase(r) BE XXL) on chitosan beads and studied their application in pomergnate juice clarification. The enzymes were combined in a multi-enzymatic system and used in a fluidized bed reactor. At the end of the storage period, this technique allowed a significant reduction of haze-active molecules. Besides the good properties achieved with enzymes chitosan immobilization, some cutbacks may occur. The low mechanical strength and crosslinking of chitosan sometimes require the use of agents to form more stable networks [159]. Glutaldehyde is the compound most widely used for this purpose [160], yet this agent must be avoided in food and beverage products due to its cytotoxic effects at concentrations higher than 0.5 ppm [161]. Therefore, some studies have aimed to find natural agents with lower toxicity. One of those recently studied, is genipin (methyl (1S,2R,6S)-2-hydroxy-9-(hydroxymethyl))-3-oxabicyclo[4.3.0] nona-4,8-diene-5-carboxylate), a compound obtained from the fruits of Genipa americana and Gardenia jasminoides Ellis. [PERSON] et al. [159] evaluated the immobilization of the enzyme \(\beta\)-galactosidasease, produced by _[PERSON]_ lactis, in genipin-activated chitosan support aiming to apply the enzymatic derivatives in the hydrolysis of ultra-high-temperature (UHT) diluted milk in lower temperatures. The immobilization proved to be a viable strategy for the biocatalysts' process. Genipin was efficient as an activating and crosslinking agent of the chitosan support, making the immobilized enzyme more stable and enabling mechanical resistance for the enzymatic derivatives.