本研究利用本土菌株(strain C1)所分泌之幾丁質酶來分解幾丁質以生產N-乙醯葡萄醣胺，並藉由實驗設計法求得幾丁質酶活性之最適化反應條件以利於N-乙醯葡萄醣胺之生產。實驗結果發現幾丁質酶活性與細胞生長之趨勢正相關，但葡萄醣胺之生產則主要發生在菌株C1不再生長之時，顯示菌株C1會利用葡萄醣胺作為營養源。然而當利用菌株C1醱酵上清液(含幾丁質酶)進行幾丁質之水解，其葡萄醣胺產物的濃度上升達8500 mg/L，而幾丁質酶的活性也有所提高，其總活性與比活性分別為1.12 OD550/ml/h和1.14 OD550/mg protein/h。本研究亦以反應曲面法求得幾丁質酶最佳活性之溫度及pH分別為45℃及7.0。另外於酵素動力學方面以菌株C1醱酵上清液進行不同濃度CM-chitin-RBV (carboxymethyl-chitin- Remazol Brilliant Violet 5R)之水解反應，發現其幾丁質酶活性與基質濃度之關係可以Michaelis-Menten動力模式來描述，並求得其Km及vmax值分別為2.26 g/L及3.16 OD550/mg protein/h，而以不同濃度之幾丁質粉末進行酵素動力學分析，同樣符合Michaelis-Menten動力學模式，其Km及vmax值分別為0.156 g/L及222.9 mg NAG/mg protein/h。該幾丁質酶在4oC冷藏保存兩個月後，仍能維持原始活性，顯示極佳之穩定性。該酵素經SDS-PAGE分析後，估計其分子量約為39 kDa。

In this study, an indigenous isolate strain C1 secreting chitinase was used to hydrolyze chitin to produce N-acetyl-D-glucosamine (NAG), which is a derivative of glucosamine. Results of batch cultures show that chitinase activity positively correlated with cell growth, whereas NAG production mainly occurred when cell growth ceased. This indicates that strain C1 utilized NAG for growth. However, using chitinase-containing fermentation supernatant to hydrolyze chintin resulted in relatively higher NAG production of 8500 mg/L. Meanwhile, the total and specific chintinase activity also slightly increased to 1.12 OD550/ml/h and 1.14 OD550/mg protein/h. Statistical methodology, namely response surface methodology (RSM), was applied to identify optimal pH and tempertature achieving the best chitinase activity and NAG production. The results show that the optimal condition for chitinase activity was pH 7.0 and 45oC. Enzyme kinetics study shows that when using CM-chitin-RBV as the substrate, the Michaelis constant and maximum rate were 2.26 g/L and 3.16 OD550/mg protein/h, respectively. When using chitin powder as the substrate, Michaelis constant and maximum rate were 0.156 g/L and 222.9 OD550/mg protein/h, respectively. The concentrated chitinase maintained its original activity when it was stored at 4oC for 60 days. Results of SDS-PAGE show that the chitinase produced from strain C1 may have a molecular weight of 39 kDa.

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