Partial Characterization of Chitosanase from Digestive Tract of Channastriata

Introduction

Chitin is the second largest polysaccharide after cellulose which has the chemical formula of poly (2-acetamide-2-dioksi-β-D-Glucose) with β- glycosidic (1.4) linkage connecting each unit. The chemical structure of chitin is similar to cellulose, only differentiated by a group attached to the atom C2. If the cellulose cluster bound to the atom C2 is OH, then the chitin-bound is acetamide group¹. Chitosan is a deacetylated product of chitin which is found in many insects, crustaceans and fungi². Chitin is a fibrous material and very insoluble, while chitosan is slightly soluble in water and is much more tractable material with a broad range of applications³.

Chitosan is also found in nature; found in the insect cuticle⁴ and the cell walls of fungi class Zygomycetes, in chlorophycean algae Chlorella sp.⁵. This chitosan naturally synthesized by the joint action of chitin synthetase and chitin deacetylase, as shown for Mucorrouxii and Colletotrichum lindemuthianum⁶.

Materials and Methods

Chitosanase Activity

Chitosanase assay was conducted according to Yoon et al.⁷, with modification. One (1) unit of chitosanase activity was defined as the amount of the enzyme which produces 1 μmol of reducing sugar (glucosamine) per minute.

Effcet of pH Enzyme Activity

Optimum pH was determined by assaying in buffer with pHvalues of 6-9 using citric acid buffer (pH 3-4), citric-phosphate buffer (pH 4-6), phosphate buffer (pH 6-8) and boric buffer (pH 8-9), in thepresence of soluble chitosan substrate.

Effcet of Temperature on Enzyme Activity

Optimum temperature was measured at 30, 40, 50, 60, 70 and 80 °C at pH 7 by using soluble chitosan as the substrate.

Result and Discussion

Chitosanase Activity

The results of the chitosanase activity fromdigestive tract of Snakehead fish (Channastriata) are presented in Figure 1.

Figure 1: Chitosanase activity from digestive tract of snakehead fish (Channastriata) Click here to View figure

 

Fig1 shows two different chitosanase activity of Channastriata (domesticated and non domesticated fish). Enzymes isolated from both stomach and intestine of domesticated fish had higher activity in the amount of 0.0395 U/mL and 0.0296 U/mL respectively.Chitosanaseisolated from the stomach of domesticated and non domesticated samplesconsiderably showed higher activity than the same enzyme isolated from the intestine.

Effect of pH on Chitosanase Activity

Effects of pH on the activity of chitosanase isolated from digestive tract of Channastriataare presented in Figure 2 and 3.

Figure 2: Effects of pH on the activity of chitosanase isolated from digestive tract of Channastriata (a= domesticated; b= non domesticated) Click here to View figure

 

Optimumchitosana sea ctivities from the stomach and the intestine were at pH 7 (domesticated) and pH 6 (non domesticated).The results were higher than pH 5.8 of Bacillus cereus⁸, pH 5.0 of Streptomyces cyaneogriseus⁹and pH 6 of Paenibacillus ehimensis¹⁰.

Figure 3: Effect of pH on Chitosanase Activity from the Intestine (a= domesticated; b= non domesticated). Click here to View figure

 

Effect of Temperature on Chitosanase Activity

Temperature ranged between 30 ^oC and 80 ^oC were used to study the effect of pH on chitosanase activity. Fig 4 showed the effect of temperature on the activity of chitosanaseisolated from digestive tract of Channastriata.

Figure 4: Effect of Temperature on Chitosanase Activity (a= the stomach; b=the Intestine)  Click here to View figure

 

The optimum temperature of 70°C was recorded for the chitosanase(the stomach and the intestine) in this study. The result was higher than the 54°C optimum temperature reported for Bacillus cereus⁸, 50 ^oCforBacillus megaterium¹¹, and Gongronella sp.¹².

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