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標題: | 由Bacillus subtilis var. natto BCRC 80517生物轉換發酵液回收金雀異黃酮磷酸酯衍生物之研究 Integrated Process for Genistein 7-O-Phosphate Recovery from the Harvested Culture Broth of Bacillus subtilis var. natto BCRC 80517 with Genistein |
作者: | Wen-Yun Luo 羅文韵 |
指導教授: | 蘇南維(Nan-Wei Su) |
關鍵字: | Bacillus subtilis var. natto,金雀異黃酮,生物轉換,異黃酮磷酸酯衍生物,固定床吸附,膨脹床吸附, Bacillus subtilis var. natto,genistein,biotransformation,isoflavone phosphate conjugate,fixed bed process,expanded bed process, |
出版年 : | 2015 |
學位: | 碩士 |
摘要: | 存在於黃豆中的大豆異黃酮為黃豆之二級代謝物,可與人體雌激素受體結合,具有雌激素活性,被認為是植物雌激素。依化學結構可分成四大類: malonyl-glucosides、acetyl-glucosides、glucosides和aglycones。其中,aglycones被認為生理活性最佳,但卻因水溶性差造成生物可利用率低。本研究室先前篩選出Bacillus subtilis var. natto BCRC 80517菌株可將金雀異黃酮(genistein)轉換成金雀異黃酮磷酸酯衍生物(genistein 7-O-phosphate, G7P)。本研究探討之目的是從發酵液中回收金雀異黃酮磷酸酯之程序。 本研究分別探討利用固定床及膨脹床兩種吸附方式回收金雀異黃酮磷酸酯衍生物。在固定床部分,發酵液經除菌處理後,以HP20、XAD16、SP700和SP850四種大孔樹脂進行吸附與脫附的篩選。比較四者的脫附比例和購置成本,選用 HP20作為吸附劑,接著透過吸附貫穿曲線探討pH值和進樣濃度對於大孔樹脂吸附能力的影響,結果顯示發酵液調至pH 1、進樣濃度0.72 mg G7P/mL於5g HP20填充下為較佳之吸附條件。探討不同濃度之NaCl溶液對於金雀異黃酮磷酸酯衍生物之清洗效果,結果顯示以1% NaCl溶液(pH 1)作為清洗液最適當。最後以30%乙醇脫附目標物,脫附液經濃縮、凍乾後,產物純度約為67%,回收率約為70%。在膨脹床部分,發酵液未經除菌處理,以HP20為吸附劑,並沿用固定床吸附之結果加以修正,探討不同流速對於吸附能力的影響,結果顯示0.88 cm/min為較佳之條件。探討不同濃度之乙醇溶劑對脫附產物回收率和純度的結果顯示,經由30%乙醇脫附之金雀異黃酮磷酸酯產物的純度約為65%,回收率可達90%。 Genistein, one of the primary bioactive agents in soybeans, has a number of pharmacological and biological activities; however, low water solubility and poor bioavailability limit its use. Previous study revealed a water-soluble phosphate conjugate of genistein, genistein 7-O-phosphate (G7P), generated by biotransformation of Bacillus subtilis var. natto BCRC 80517 with genistein. This study aimed to develop a feasible and promising process for recovering G7P from the fermentation broth of biotransformation. Two techniques with macroporous resin adsorption including packed bed adsorption and expanded bed adsorption were used in this work. For the processing of packed bed adsorption, factors including pHs, various solvents for extraction, the selection of adsorbents were investigated. The results showed that the pH of the harvested broth was adjusted to 1, and then subjected to HP-20 resin, the adsorption capacity of G7P by resin was approximately 46 mg/g. After extensively washing with pH 1, 1% aqueous NaCl solution, the G7P concentrate could be obtained from the 30% ethanol eluate with 70% recovery and approximately 67% in purity. For the procedure of expanded bed adsorption, the same adsorbent HP20 was employed for G7P recovery. The harvested broth was pumped through the expanded bed at flow rate of 10 mL/min to capture G7P without prior microbial biomass separation. With the optimal conditions for the process of expanded bed adsorption chromatography, the recovery of G7P was up to 90% and the product of G7P concentrate was approximately in 65% (w/w) purity. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/53143 |
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顯示於系所單位: | 生化科技學系 |
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