초록

Ⅳ. 연구개발결과
□ 감귤부산물을 기질로 단백다당체 고생산성 균주 선별
□ 감귤부산물을 기질로 단백다당체 고생산성 균주 개발
□ 감귤부산물 기질을 이용한 소규모 배양을 통한 생산균주의 배양 생리적 특성 연구
□ 감귤부산물의 신속하고 효율적인 전처리 공정 개발
□ 감귤부산물 첨가 배지 최적화
□ 발효기 배양을 통한 대규모 생산조건 확립
□ 파일롯 규모의 단백다당체 생산공정 확립
□ 생산된 단백다당체 특성 평가 완료
□ 공정 통합 및 경제성 평가 완료

Abstract ▼

Strain improvement and fermentation physiology investigation in the flask cultures were undertaken in suspended cultures of Sparassis crispa mycelia using by-products of mandarin industry as a carbon source for enhanced production of protein-bound polysaccharides(soluble β-D-glucan), a powerful immu

Strain improvement and fermentation physiology investigation in the flask cultures were undertaken in suspended cultures of Sparassis crispa mycelia using by-products of mandarin industry as a carbon source for enhanced production of protein-bound polysaccharides(soluble β-D-glucan), a powerful immuno-stimulating agent. Intensive strain improvement program was carried out by obtaining large amounts of protoplasts for the isolation of single cell colonies. Rapid and large screening of high-yielding producers was possible because large numbers of protoplasts (104~105 protoplasts/ml) formed using the banding filtration method with the cell wall-disrupting enzymes could be regenerated in relatively high regeneration frequency(98%) in the newly developed regeneration medium(RPA). It was demonstrated that the strains showing high performances in the protoplast regeneration and solid growth medium were able to produce 25~30%(w/w) of β-D-glucan and 7~10 g/L of biomass in stable manners in suspended shake-flask cultures of S.crispa mycelia. In addition, cell mass increase was observed to be the most important in order to enhance β-D-glucan productivity during the course of strain improvement program, since the amount of β-D-glucan extracted from the cell wall of S.crispa mycelia was almost constant on the unit biomass basis. Therefore we investigated the fermentation physiology, the several factors affecting cell growth extent in the suspended cultures of mycelial fungal cells using mandarin peels as a carbon source. As a results, The cell growth in production medium including mandarin peels was the highest at the initial pH 5.5 and 24℃ of the culture temperature in the shake flask cultures of S.crispa mycelia. But, still the productivity is not enough for mass production of protein-bound polysaccharides in industry scale. In this case, For the success of mass production of protein-bound polysaccharides in the bioreactor cultures of S.crispa mycelia using by-products of mandarin industry as a carbon source, we have to develop the pre-treated process of mandarin peels and the optimized production medium.
Fermentation physiology were investigated in submerged mycelial cultures of Sparasis crispa using by-products of mandarin industry as a carbon source for enhanced production of water-soluble protein-bound polysaccharides(β-D-glucan), a clinically important agent due to its strong immunostimulating activities. Attempts to enhance cell growth were carried out in both shake flask and bioreactor cultures of Sparassis crispa mycelia, since increment in cell mass turned out to be the most important factor for the enhancement in β-D-glucan productivity (i.e. constant amount of β-D-glucan per unit cell mass). Firstly, production medium was optimized using powerful statistical methods such as Plackett-Burman design, fractional factorial design(FFD), steepest ascent method(SAM) followed by response surface method(RSM) with central composite design. About 2-fold higher amount of cell mass was obtained in the production cultures, when an optimized production medium was utilized.
Notably, it was found that the kinds of growth medium in the growth cultures also had a strong influence on the mycelial growth in the production cultures. When QG growth medium was used in the growth cultures, maximum cell mass was approximately 40% higher compared to the control growth medium, reaching almost 20 g/L in the production cultures performed with CM solution added production medium.
The relationship between the fermentation parameters pH, temperature and the concentrations of feeding solution(a mixture of mandarin peels, glucose and yeast extract) and the protein-bound polysaccharides(soluble β-D-glucan) of Sparassis crispa in different fed-batch methods are discussed. Different fed-batch methods including constant feed rate fed-batch, constant glucose concentration fed-batch, exponential fed-batch, pH-control exponential fed-batch, pulse fed-batch and pH-control pulse fed-batch are compared.
pH-control pulse fed-batch was an effective method for the production of protein-bound polysaccharides(cell mass), the concentration of which reached 25g/l, 30% higher than in the other fermentations.

목차 Contents

• 표지 ... 1
• 제출문 ... 2
• 요약문 ... 3
• SUMMARY ... 5
• 목차 ... 7
• 제 1 장 서 론 ... 8
• 1절. 연구개발의 필요성 ... 8
• 제 2 장 국내외 기술개발 현황 ... 11
• 1절. 국내 연구 현황 ... 11
• 2절. 국외 연구 현황 ... 12
• 제 3 장 연구개발수행 내용 및 결과 ... 13
• 1절. 감귤부산물을 기질로 단백다당체 고생산성 균주 선별 ... 13
• 2절. 감귤부산물을 기질로 단백다당체 고생산성 균주 개발 ... 14
• 3절. 소규모 배양을 통한 생산균주의 배양생리적 특성 연구 ... 15
• 4절. 감귤부산물의 신속하고 효율적인 전처리 공정 개발 ... 17
• 5절. 감귤부산물 첨가 배지 최적화 ... 24
• 6절. 발효기 배양을 통한 꽃송이 버섯 유래의 단백다당체 고생산 조건 확립 ... 38
• 7절. 파일롯 규모의 단백다당체 생산공정 확립 ... 50
• 8절. 생산된 단백다당체 특성 평가 ... 63
• 9절. 공정 통합 및 경제성 평가 ... 77
• 제 4 장 연구개발목표 달성도 및 대외 기여도 ... 82
• 1절 : 목표대비 달성도 ... 82
• 2절 : 정량적 성과(논문게재, 특허출원, 기타) ... 85
• 제 5 장 연구개발결과의 활용계획 ... 87
• 제 6 장 연구개발과정에서 수집한 해외과학기술정보 ... 88
• 제 7 장 기타 중요 변동사항 ... 89
• 제 8 장 국가과학기술종합정보시스템에 등록한 연구장비 현황 ... 90
• 제 9 장 참고문헌 ... 91
• 끝페이지 ... 94