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研究生： 蔡忠廷
Tsai chung-ting
論文名稱： 以酵素法進行麴酸酯生產之製程開發
Process development of kojic acid ester production by using enzyme
指導教授： 吳文騰
Wu Wen-Teng
口試委員:
學位類別： 碩士
Master
系所名稱： 工學院 - 化學工程學系
Department of Chemical Engineering
論文出版年： 2005
畢業學年度： 93
語文別： 中文
論文頁數： 62
中文關鍵詞： 麴酸 、麴狻酯 、酯化反應 、脂肪酸分解酶 、尿素包埋法
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    • 麴酸為絲狀真菌所代謝的有機酸，其具有特殊的生物性質，所以常被應用在許多用途上，例如止痛劑、食品添加劑。近年來更發現具有抑制黑色素生成能力，而被大量使用於美白保養品中。但是，麴酸是一種水溶性物質，而且對光和熱效應敏感，因此影響其安定性，這些原因都造成麴酸在應用上的限制。有鑒於麴酸廣泛的應用性，如何增加麴酸的脂溶性與安定性成為一個重要的課題。

    藉由脂肪酸與麴酸進行酯化反應產生的麴酸酯，已被證實是一種有效改質麴酸安定性並提高麴酸脂溶性之方法。本實驗利用酵素法進行麴酸酯的生產，此外，酵素反應具有較高選擇性與反應條件較溫和的優點，有利於反應後麴酸酯純化分離。本研究目標產物為麴酸棕櫚酯，利用固定化酵素Novozyme 435 ( Candida antartica , type B lipase ) 做為催化劑，並探討最佳酯化反應條件與麴酸酯純化方法。

    本研究在酯化反應條件的研究上，探討了Lipase種類、反應溫度、麴酸/棕櫚酸比例、和分子篩添加量對酯化反應的影響。最佳反應結果顯示1 mmole 麴酸與2 mmole棕櫚酸於15 ml 的丙酮溶液中，添加1.5 g Novozyme 435以及4 g分子篩於40℃中，反應30小時，其轉化率可達92%。

    在麴酸棕櫚酯的純化研究中，發現透過尿素包埋法來進行純化，其最佳純化結果顯示2 g的初步純化物與4 g尿素溶於40 ml甲醇溶液中，能得到純0.88的麴酸棕櫚酯。

    目錄 摘要-----------------------------------------------------------------------------------I 謝誌--------------------------------------------------------------------------------III 目錄--------------------------------------------------------------------------------V 圖目錄----------------------------------------------------------------------------IV 表目錄------------------------------------------------------------------------------XI 第一章 緒論 1.1 前言------------------------------------------------------------------------------------1 1.2 研究目標與方法---------------------------------------------------------------------2 第二章 文獻回顧 2.1 麴酸簡介------------------------------------------------------------------------------3 2.1.1 麴酸基本性質與量測方法-------------------------------------------------3 2.1.2 麴酸之生產-------------------------------------------------------------------3 2.1.3 麴酸之應用-------------------------------------------------------------------5 2.1.4 麴酸與市面上美白原料之美白活性比較-------------------------------7 2.1.5 麴酸的安全性----------------------------------------------------------------8 2.1.6 麴酸衍生物之應用----------------------------------------------------------8 2.1.7 麴酸在商業應用上之問題與改善方法---------------------------------10 2.2 麴酸酯之生產-----------------------------------------------------------------------11 2.2.1 化學法與酵素法生產麴酸酯之優劣處比較---------------------------11 2.2.2 脂肪分解酶之簡介---------------------------------------------------------13 2.2.3 脂肪分解酶於酯化反應之應用------------------------------------------15 2.2.4 溫度影響---------------------------------------------------------------------16 2.2.5 有機溶劑之影響------------------------------------------------------------19 2.2.6 麴酸與脂肪酸比例之影響------------------------------------------------19 2.2.7 反應系統中水分的影響---------------------------------------------------19 2.2.8 脂肪分解酵素之回收與再活化------------------------------------------23 2.3 麴酸酯之純化-----------------------------------------------------------------------23 2.3.1 尿素包埋法------------------------------------------------------------------23 2.4.2 液相層析法------------------------------------------------------------------24 第三章 實驗材料與方法 3.1 實驗材料-----------------------------------------------------------------------------25 3.1.1 實驗藥品---------------------------------------------------------------------25 3.1.2 實驗器材---------------------------------------------------------------------26 3.2 反應條件探討-----------------------------------------------------------------------26 3.2.1 酵素活性篩選---------------------------------------------------------------26 3.2.2 麴酸/脂肪酸比實驗--------------------------------------------------------26 3.2.3 溫度實驗---------------------------------------------------------------------27 3.2.4 分子篩添加量實驗---------------------------------------------------------27 3.2.5 酵素添加量實驗------------------------------------------------------------27 3.2.6 Novozyme 435 使用安定性實驗----------------------------------------28 3.3 麴酸棕櫚酯之分析-----------------------------------------------------------------28 3.4 麴酸棕櫚酯之純化實驗-----------------------------------------------------------29 3.4.1 初步純化物製備實驗------------------------------------------------------29 3.4.2 液相層析法純化麴酸酯之實驗------------------------------------------29 3.4.2.1 沖提液組成之探討---------------------------------------------------29 3.4.2.2 液相層析法分離麴酸棕櫚酯與棕櫚酸之探討------------------29 3.4.3尿素包埋法純化麴酸酯之實驗-------------------------------------------30 第四章 實驗結果與討論 4.1 麴酸棕櫚酯分析方法之建立-----------------------------------------------------33 4.1.1 酵素法是否生產麴酸棕櫚酯之確認------------------------------------33 4.1.2 麴酸棕櫚酯標準品之純度分析------------------------------------------35 4.1.3 麴酸棕櫚酯HPLC分析檢量線之建立---------------------------------37 4.2 麴酸棕櫚酯生產條件之探討-----------------------------------------------------38 4.2.1 脂肪分解酵素之篩選------------------------------------------------------38 4.2.2 麴酸/棕櫚酸比例之影響--------------------------------------------------39 4.2.3 溫度影響---------------------------------------------------------------------41 4.2.4 添加分子篩對麴酸棕櫚酯轉化率之影響------------------------------42 4.2.5 酵素添加量之影響---------------------------------------------------------44 4.2.6 Novozyme 435 使用安定性之探討--------------------------------------45 4.3 麴酸棕櫚酯之純化-----------------------------------------------------------------46 4.3.1 初步純化物之製備與純化步驟之檢視------------------------------46 4.3.2 液相層析法純化麴酸酯---------------------------------------------------48 4.3.2.1 不同組成沖提液效果之比較---------------------------------------48 4.3.2.2 麴酸棕櫚酯與棕櫚酸分離之研究---------------------------------49 4.3.3 尿素包埋法純化麴酸棕櫚酯---------------------------------------------50 4.3.3.1甲醇溶液量對純化麴酸酯之影響----------------------------------52 4.3.3.2尿素添加量對純化麴酸棕櫚酯之影響----------------------------53 第五章 結論與未來展望 5.1 結論-----------------------------------------------------------------------------------54 5.2 未來展望-----------------------------------------------------------------------------56 參考文獻---------------------------------------------------------------------------58

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