以微生物生產葡萄糖胺之動力學研究

Production Kinetic of Glucosamine by Microorganism

謝睿文(Jui-Wen Hsieh)

指導教授 ： 吳和生 ； 王孝憲

元智大學/工程學院/化學工程與材料科學學系/碩士(2006年)

摘要

葡萄糖胺(glucosamine)為一種單胺醣類物質，是構成幾丁質(chitin)和幾丁聚醣(chitosan)之單糖；為人類關節間潤滑結締組織之主要成分，具修復、補充、滋潤、再造受損的軟骨，可治療退化性關節炎。本研究主要是利用微生物醱酵生產葡萄糖胺，討論製備葡萄糖胺的可行性以及分離純化葡萄糖胺。使用四種菌株: Rhizopus oligosorus BCRC 31996; Monascus purpures BCRC 31499; Monascus pilosus BCRC31527; Aspergillus sp. BCRC31742，實驗主要分成三部份討論：(1)分析純化，分析方式為利用1-naphthyl isothiocyanate為衍生試劑，衍生反應在50°C下反應1小時，以吡啶為反應溶劑，衍生物利用逆向液相層析分離管柱分離，以230 nm 紫外光檢出器偵測生產之葡萄糖胺含量，其精密度可達1.69%、準確度可達2.73%； (2)以搖瓶培養探討培養條件，如菌種種類、培養基、酸鹼度、碳氮比等等，菌種產量最高為Aspergillus sp. BCRC31742培養於葡萄糖胺(glucose)和蛋白質(peptone)複合培養基（3430 mg/L），在培養條件下以酸鹼度影響最大；(3)以4公升醱酵槽進行培養，探討變因有酸鹼度、轉速、碳氮比，實驗中使用Aspergillus sp. BCRC31742培養於葡萄糖胺和蛋白質複合培養基中，可獲得葡萄糖胺濃度為2310 mg/L、生物量為10.1g/L、含量為229mg/g biomass、產率為92.4mg/g carbon source、生產力13.8mg/L×h；(4)純化方式使用Aspergillus sp. BCRC31742醱酵所得之菌塊經6N HCl在100oC下反應24小時，以NaOH中和至pH 7，使用活性碳除色，再由減壓濃縮機濃縮成葡萄糖胺鹽酸鹽濃縮液，經烘箱烘乾後，其純度由原來2.29 %提升至13.4 %。

關鍵字

鹽酸化葡萄糖胺 ； 微生物發酵 ； 動力學策略 ； 產率 ； 生產力

並列摘要

Glucosamine is an amino-monosaccharide and one of the basic constituents of Chitin and Chitosan. Glucosamine is the major component of human inter-articular lubricant connective tissue. It can repair and rebuild the damaged cartilage and it also can cure the disease of osteoarthritis. The objective of this work is to study the parameters affecting the production of glucosamine hydrochloride by microbial fermentation and to obtain the method of separation and purification of glucosamine hydrochloride.This study uses four funguses Rhizopus oligosorus BCRC 31996, Monascus purpures BCRC 31499, Monascus pilosus BCRC31527 and Aspergillus sp. BCRC31742 to product glucosamine hydrochloride by using fermentation.Three parts were discussed in this study.First is the kinetic and strategy by flask culture which conditions include kinds of fungus, mediums, pH value, and carbon and nitrogen source.The experimental result shows that the glucosamine concentration had an optimum value and was 3428mg/L by using Aspergillus sp. BCRC31742 culture in GP medium, the pH controlled an important rate in culture. Second, the kinetics and strategoy by fermenter culture that the factors were pH value, incubation time and carbon and nitrogen source in the 4L fermentor fermentation.The result shown the glucosamine concentration was 2311mg/L; biomass, 10.1g/L; content, 229(mg/g biomass); yield, 92.4mg/g of carbon source; productivity, 13.8mg/L×h-1 that Aspergillus sp. BCRC31742 was incubated in GP medium. Third is the part of analysis and purification of glucosamine. The glucosamine was analyzed with 1-naphthyl isothiocyanate (NITC) as derivatizing agent. The reaction was carried out in pyridine at 50oC for 1 h. The derivative was analyzed of High Performance Liquid Chromatography. The precision of glucosamine is below to 1.69% and the accuracy is to 2.73%. Purify method is the dry cell reacting with 6N HCl at 100oC for 24 h, then neutralization with NaOH to pH 7 to obtain glucosamine hydrochloride aqueous solution. The purity of glucosamine is from 2.29% to 13.4% after the purification process containing depigmentation and condensing by Rotary Evaporator.

並列關鍵字

Glucosamine hydrochloride ； microbial fermentation ； kinetics and strategy ； yield ； productivity

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