Optimization Of Fermentation Conditons Of Penicillium Oxalicum, And Characterization Of Chitosanase

In recent years, the activity of microorganism chitosanase was low in the domestic studies. There were few strains having a good capability of chitosanase-producing, and a valuable application of industrialisation. And that's why the price of chitosanase was high. In order to explore the resources of sea, and get a new enzyme source which have a good potential application of industrialization, this research abtained a new strain. This study included optimization of fermentation condition, purification and characterization of chitosanase. This research provided direct theory and experiment foundation on industrialized production of chitosanase.Research results were as follows:⑴A marine chitosanase-producing strain was isolated from whelk and shell by the seaside, and identification by Colony morphology, electron microscope and rDNA - ITS sequence analysis. And named Penicillium oxalicum PCS-7. This was the first time reported Penicillium oxalicum possessed the ability of production of chitosanase.⑵The optimal composition of oligotrophic medium for Penicillium oxalicum. PCS-7 producing chitosanase was as follows(g/L): bacteriological peptone 9 g, glucose 10 g. The results showed that Penicillium oxalicum. could directly make use of seawater without other mineral nutrition in fermentation medium. In this way, we could save the inorganic salt-adding and the consumption of freshwater in the fermentation process and reduce the cost of fermentation medium. In the purpose of fermentation chitosanase in industrial, we studied different kinds of cheap medium for production of chitosanase, and we found the soybean powder was the best, and the medium of soybean powder was as follow (g/L seawater): soybean powder 25 g, and the optimal enzyme activity was 8.5 U/mL. This medium was insufficient 5% of the cost of original.⑶After fermentation in the medium of soybean powder with seawater, we could obtained the pure-chitosanase by utilize the refrigeration centrifuge. The molecular weight was estimated to 42,000 on SDS-PAGE.⑷The enzymatic peoperties were investigated: The optimal temperature for hydrolysis of the chitosanase is 50℃, and the optimal pH is 6.0. The enzyme activity was significant increased by Mg²⁺,Mn²⁺,Ca²⁺. The chitosanase have a great stability of low temperature and a remarkable of toleration of acid. Vmax=0.05 mg/(mL·min),Km=9.02 mg/mL。...