The aimed of this work, was improved the stability of the chitosanase enzyme by immobilization with different natural carriers to enhance the economic of industrial biocatalytic process. It was used in preparation of biological active chitooligosaccharides. Pepper chitosanase extracted from (Capsiuum annuum) leaves was immobilized by four different immobilization methods with chitosan, DEAE-cellulose and sodium alginate. The resulted immobilized chitosanases were compared with respect to their immobilization efficiency, reusability and storage stability. Immobilization of chitosanase improved their enzymatic activity. They were 105, 83.8 and 65.2 U/g for immobilized chitosanase with entrapment, covalent and ionic binding methods, respectively. Immobilized chitosanase by covalent and ionic binding exhibited good reusability more than by entrapment method. Generally, the immobilized chitosanases showed better storage stability than that of the free one. The amount chitooligosaccharides produced by using immobilized chitosanase by ionic binding, covalent binding and entrapment methods was higher than that by adsorption method and also more than produced by free one. From a economical point of view, good reusability and storage stability are of the most important feature for the industrial application as biocatalytic.