起始pH與通氣量對深層培養豬苓（Polyporusumbellatus）菌絲體生長及多醣體生產之影響

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Title:	起始pH與通氣量對深層培養豬苓（Polyporusumbellatus）菌絲體生長及多醣體生產之影響
Other Titles:	Effects of initial pH and aeration rate on the mycelial growth and polysaccharide production in submerged culture of Polyporus umbellatus
Authors:	李婉瑜 Lee, Wan-Yu
Contributors:	顏文義 Yen, Wen-Yi 東海大學食品科學系
Keywords:	豬苓,多醣體 Polyporus umbellatus, polysaccharides
Date:	2008
Issue Date:	2011-03-11T06:07:35Z (UTC)
Abstract:	豬苓（Polyporus umbellatus）為一種常用的傳統中藥。其菌核應用於治療水腫且利尿。從豬苓分離出的水溶性多醣主要結構是以β-1,3-D-葡聚糖為主鏈具有β-1,6鍵結的支鏈。豬苓多醣被證實具有抗菌、抗病毒、免疫調節和抗腫瘤等功效。由於野生豬苓生長緩慢且日益匱乏，因此以人工培養方式針對豬苓生物活性代謝物質的生產與應用?得加以探討。本研究以深層培養法最適化豬苓之培養，並擴大至發酵槽探討通氣量對豬苓菌絲體生長及多醣體生成之影響。??試驗結果顯示，豬苓菌絲乾重（MDW）與胞內多醣（IPS）最大產量皆在培養第18天可得，胞外多醣（EPS）則於第16天達最大產量。30 g/l glucose和5 g/l corn steep powder分別為豬苓培養之最佳碳、氮源組合與濃度。以不同起始pH培養豬苓，最適合菌絲體生長繁殖的pH為pH 7，可得到最大MDW為7.55 mg/ml。隨著pH值降低則MDW遞減，pH 4之菌絲乾重最低。IPS產量在pH 4.80（未調整pH）至pH 7的範圍內沒有明顯差異，最大產量可達0.097 mg/ml。未調整培養基起始pH（pH 4.80）培養豬苓最適合EPS生產，可得到最高EPS產量為0.588 mg/ml，其分子量分布於341 kDa至432 kDa。然而以pH 4.80（未調整pH）至pH 6培養豬苓在菌絲體生長、胞內和胞外多醣產量方面皆無明顯差異，並與pH 7培養者差異不大。發酵槽操作之通氣量愈高則菌絲體生長愈佳。以通氣量2 vvm培養豬苓於第18天達到最大MDW和IPS產量，分別為7.25 mg/ml和0.100 mg/ml，intra- β-1,3-glucan含量為1.372 μg/ml LE，分子量為206 kDa。以通氣量1.5 vvm培養時，則於第16天達EPS最大產量為0.600 mg/ml，exo-β-1,3-glucan含量為2.773 μg/ml LE。與單一通氣量相較，兩段式通氣量調控（A2S）培養豬苓之最大MDW和多醣產量生成時間較早。A2S之最大MDW和IPS產量於培養第16天可得，培養第14天則達到最大EPS產量。且A2S可同時得到相對高產量的MDW、IPS和EPS。兩段式調控通氣量培養豬苓之最大MDW和IPS產量分別為7.14 mg/ml和0.076 mg/ml，intra-β-1,3-glucan含量為1.221 μg/ml LE，分子量為331 kDa至398 kDa。最大EPS產量則為0.583 mg/ml，exo-β-1,3-glucan含量為1.193 μg/ml LE，分子量為20.2 kDa。 Polyporus umbellatus is a widely used traditional Chinese medicinal fungus. Its sclerotium can be used for treating edema and diuresis. The water-soluble polysaccarides isolated from P. umbellatus possess anti-bacteria, anti-virus, immunomodulatory and anti-tumor activities. P. umbellatus in the wild is growing slowly and has been depleted. Research on the methods of artificial cultivation for production of the biomass and the bioactive metabolites by P. umbellatus is necessary. This study was conducted to optimize the growth conditions in submerged culture. Effect of different aeration rate in the fermentor was also investigated.The effects of differential combination with carbon and nitrogen sources of culture media on mycelial growth and polysaccharide biosynthesis were evaluated in shake flask culture. Glucose and corn steep powder（CSP） combination was the favorable carbon and nitrogen source for the culture media of P. umbellatus. The hightest mycelial dry weight（CDW） and intra-polysaccharides（IPS） production, 5.65 mg/ml and 0.0457 mg/ml, were obtained respectively. The maximum production of exopolysaccharides（EPS） was 0.356 mg/ml, with a relative high molecular weight of 432 kDa was obtained. The hightest production of CDW（4.65 mg/ml）, IPS（0.036 mg/ml） and EPS（0.315 mg/ml） were achieved with concentration of 30 g/l in the glucose medium. The enhancement of CDW was obtained when the glucose concentration was increasing to 40 g/l, but the EPS production was not significant as compared to 30 g/l. CSP in 5 g/l was the suitable nitrogen source concentration. The optimal initial pH of growth medium was pH 7, the highest CDW（7.55 mg/ml） was obtained. The mycelial yield was diminished by decreasing pH, initial pH 4 was unsuitable for cultivaton. The productions of IPS were not significant among unadjest pH, pH 5, pH 6 and pH 7 of the medium, the maximum IPS content was reached to 0.097 mg/ml. The highest production of EPS（0.588 mg/ml） was obtained in unadjest pH culture. The optimal initial pH for mycelial growth and polysaccharides production was the unadjest pH of the culture medium.In fermentor studies, the high aeration rate was favorable for mycelial growth. After 18 days of cultivation, the highest CDW（7.25 mg/ml）and IPS（0.100 mg/ml）production were abtained at 2 vvm with a molecular weight of 206 kDa and intra-β-1,3-glucan content（1.372 μg/ml LE）. The maximum production of EPS（0.600 mg/ml）and exo-β-1,3-glucan（2.773 μg/ml LE） were obtained after 16 days in 1.5 vvm culture. A two-stage aeration rate process was evaluated. The highest production of CDW, IPS and EPS obtained from two-stage fermentation was two days earlier than any other single-control. The maximum CDW（7.14 mg/ml）, IPS（0.076 mg/ml）and intra-β-1,3-glucan content（1.221 μg/ml LE）were abtained after 16 days of cultivation. The molecular weight of IPS was distributed on 331 to 398 kDa. The highest production of EPS（0.583 mg/ml） with the molecular weight of 20.2 kDa and exo-β-1,3-glucan（1.193 μg/ml LE） were obtained after 14 days of cultivation.
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