Studies On β-mannanase Activities Produced By Aspergillus Niger SL-8 With Apple Pomace Under Solid Fermentations

With the development of apple processing industry, the pomace recycling is vitally important. In this study,β-mannanase activities produced by Aspergillus niger with apple pomace under solid fermentation were systemically studied. The results are as follows:1. To enhance the enzyme activity of A. niger MQ-1, ultraviolet and microwave mutagenesis was adopted. Though different screening methods, one strain (SL-8) with the highest enzyme activity of 420 U/g was obtained and its activity was 2.01 times than its parent strain.2. The optimal fermentation conditions for producingβ-mannanase by strain A. niger SL-8 were determined and the results are as follow. The mixture of apple pomace and cottonseed powder (1:1, w/w) with 59% (w/w) initial moisture, supplemented with 1.995% (w/w) urea, 0.0955% KH₂PO₄, 0.2%CaCl₂ and 0.1%MgCl₂ (w/w),was proved to be the optimum medium. When the test fungi were inoculated in the optimized medium and incubated at 30℃for 48h, the activity ofβ-mannase increased by 28.3% than that in the basal medium and achieved 539 u/g, which was as high as the enzyme activity obtained when the bran was used as the raw material.3. The effects of the optimal temperature, pH, thermal stability, stability of acid and alkali, common metal ions to enzyme were researched. The results showed that theβ-mannanase was acidic enzymes and the optimal pH was 5.0. The enzyme remained above 85% of the initial activity after incubated at pH3.5-6.0. The optimal reaction temperature were 50℃. Thermal stabilities ofβ-mannanase were high. It remained above 80% of initial activity afterβ-mannanase incubated for 5 h at 50℃. The Km and Vmax of theβ-mannanase were obtained, which were 0.83 g/L and 166.67μmol/min, respectively. The activity of the mannanase was inhibited greatly by Cu²⁺(91%) and was activated by Fe²⁺, Fe³⁺ and Mg²⁺, especially Fe²⁺(127%). The activity of mannanase was inhibited by 0.5 mmol/L Ca²⁺; however it was activated at 1.0 mmol/L Ca²⁺.4. The study in enhancing thermostabilization of theβ-mananase by protective agent showed that glycerol was the best proteetive substance, followed by mannitol, trehalose and sorbitol. Gelatin and glucose demonstrated slightly protective ability. The best formula of different protectors optimized by orthogonal experiment design consist of glycerol 20%, trehalose 2mmol/L and sorbitol 1g/L. The relative activity ofβ-mannanase preparation containing multiplex protectors was increased by 5.8%-18.8% at 30-60℃, and the multiplex protectors may increase the optimal reaction temperature from 50℃to about 60℃due to their good protection effect.