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|標題:||Optimization of the cultivation conditions on the chitinase production from Paenibacillus sp.|
|摘要:||A bacterial strain capable of secreting high level of chitinase was isolated from chitin-enriched soil. The isolated bacterium identified as Paenibacillus sp. was grown in a medium consisting of 1% chitin powder, 0.1% MgSO4‧7H2O, 0.2% (NH4)2SO4, 0.1% K2HPO4, 0.2% CaCO3, 0.1% NaCl, 0.1% trace salt, at 28℃ for 2 days to study its chitinase production. The optimum pH and temperature ranges, pH and thermal stability ranges of the enzyme were 6.5 to 8.0, 50 to 60℃ respectively, 5.0 to 7.0 and 30 to 60℃ respectively for colloidal chitin. It was observed that using crab shell chitin powder (CSCP) as the sole carbon source resulted in a significant increase of enzyme production as compared with chitin powder.
In the process of using Paenibacillus sp. for chitinase production, the statistical design method, RSM (Response Surface Methodology) was used to determine the optimal conditions including medium compositions, initial pH, temperature of fermentation in shake flask. The obtained optimal concentrations of the medium components were CSCP, 5.54%;peptone, 0.185%;(NH4)2SO4, 0.161%. The optimal values of initial pH and temperature were 8.5 and 34.3℃, respectively. When cultivated in these optimum conditions, the enzyme activity increased 49.3% as compared with the original conditions.
The chitinase fermentation process utilizing these optimum conditions was operated in a 5-L stirred tank bioreactor. The influence of the variables, such as agitation speed, air flow rate and pH values on the fermentation were also evaluated and discussed. The variations of chitinase activity, reducing sugar, air concentration and pH, during batch fermentation are also recorded. It is found that the optimal values of air flow rate, agition speed were 3 vvm and 200rpm respectively with the uncontrolled pH yielded the maximal enzyme production of 11.8U/ml.|
本研究自土壤中分離具有分解幾丁質能力之菌株Paenibacillus sp.，以含有幾丁質粉末(chitin powder) 1%、peptone 0.1％、含水硫酸鎂(MgSO4‧7H2O) 0.1％、硫酸銨((NH4)2SO4)0.2%、磷酸氫二鉀(K2HPO4) 0.1％、碳酸鈣(CaCO3) 0.2%、氯化鈉(NaCl) 0.1％、Trace salt 0.1%的培養基，在28℃、200rpm下，振盪培養兩天，進行幾丁質酵素的液態培養醱酵。結果顯示，所產酵素，對於膠態幾丁質受質的分解（colloidal chitin）具有顯著性的活性。其最適反應pH和溫度範圍，分別為6.5~8.0和50~60℃；而pH和溫度的穩定性範圍，則分別為5.0~7.0和30~60℃。在誘導生產幾丁質酵素能力試驗得知，以蟹殼粉末幾丁質(crab shell chitin powder, CSCP)作為主要碳源較優於幾丁質粉末(chitin powder)。 利用“回應曲面法(Response Surface Methodology, RSM)”尋求三角燒瓶液態培養產生最適生產Paenibacillus sp.幾丁質酵素之最適生產條件(包括培養液組成、初始pH與醱酵溫度)結果，為CSCP 5.54%、peptone 0.18%、(NH4)2SO4 0.16%。而培養液最適初始pH與醱酵溫度，則分別為8.5及34.3℃，製得酵素最佳之活性為8.01U/ml。(比先前尚未最適化時的培養條件下，所得幾丁質酵素活性，提高49.3%。) 以上述RSM所得到的最適培養條件，應用於5-L攪拌式醱酵槽培養。探討攪拌轉速、通氣量及起始pH等因數，對液體培養幾丁質酵素活性、還原醣濃度、溶氧量和pH的變化之影響。結果顯示，以設定通氣量3 vvm、攪拌轉速200rpm，培養基酸鹼值設定不做任何調控時，所得酵素的活性為11.8U/ml最高。
|Appears in Collections:||化學工程學系所|
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