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題名 | 固定化金屬親和膜固定盤尼西林醯胺酵素之動力學參數研究=Study the Kinetic Parameters for the Immobilization of Enzymes on the Immobilized Metal Affinity Membrane |
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作者 | 曾慧娟; 陳志義; | 書刊名 | 桃園創新學報 |
卷期 | 34 2014.12[民103.12] |
頁次 | 頁93-107 |
分類號 | 460.02 |
關鍵詞 | 盤尼西林醯胺酵素; 固定化金屬親和膜; 最適化條件; 動力學參數; Penicillin G acylase; The regenerated cellulose membrane; Metal affinity immobilization; Chemical reactions; Optimum conditions; The kinetic parameters; |
語文 | 中文(Chinese) |
中文摘要 | 工業應用將盤尼西林醯胺酵素(penicillin G acylase;PGA) 固定在生物載體上當作生物觸媒,它與自由 PGA相比較,具有高活性和高穩定性,於生化反應可長期使用特性。首先利用一連串化學反應,再生纖維膜(the regenerated cellulose membrane; RCM) 與 20 ml 1.4 M NaOH, 5 ml epichlorohydrin (EPI), 於 24 ℃, 150 rpm反應 14h;接著,浸泡在 25 ml 1M iminodiacetic acid (IDA) 於 24 ℃,120 rpm,反應 12 h;將與 25ml 25mM銅離子配位鍵結,形成固定化金屬親和膜 (the immobilized metal affinity membrane ; IMAM)。在此最適化條件下,每片金屬親和膜螯合銅離子 75.6 ± 0.3 μmol與 PGA活性 1.8 U。經實驗發現固定化 PGA膜 (Immobilized PGA membrane ;IPM),可容許更寬廣的 pH值範圍和較高的熱穩定性。此外,經過 40天儲存 16次重複使用,仍可保留 99%的殘餘活性;同時,測得固定化 PGA膜之 Vmax(9.62 U/g-membrane)、Km(16.83 mM)、活化能 Ea(4.2 kcal/gmol)等動力學參數值。 |
英文摘要 | The industrial application of penicillin G acylase (PGA) requires efficient methods for immobilizing the enzyme, which yields a biocatalyst with high activity and stability when compared to free PGA. This study constructs the immobilized metal affinity membrane (IMAM) via a series of chemical reactions to sequentially couple 20ml 1.4 M NaOH, 5 ml epichlorohydrin (EPI), 24 ℃, 150 rpm, then immersed in 25ml 1M iminodiacetic acid (IDA), 24 ℃,120 rpm,12 h, and 10ml 100mM copper ions on the regenerated cellulose membrane. The condition of PGA immobilized to IMAM was examined. Under the optimal condition, a copper ion of 75.6 ± 0.3 μmol/disc with a PGA activity of 1.8 U/disc was obtained. The immobilized PGA membrane (IPM) could achieve a broader pH tolerance and higher heat stability. In addition, 99% of the residual activity could be retained via a 16-times repeated use within 40 days of storage. This study also determines the kinetic parameters Vmax(9.62 U/g-membrane), Km(16.83 mM), and the energy of activation Ea(4.2 kcal/gmol) for the immobilized PGA membrane. |
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