보고서 정보
주관연구기관 |
국립농업과학원 National Institute of Agricultural Sciences |
보고서유형 | 최종보고서 |
발행국가 | 대한민국 |
언어 |
한국어
|
발행년월 | 2015-02 |
주관부처 |
농촌진흥청 Rural Development Administration(RDA) |
등록번호 |
TRKO201500010398 |
DB 구축일자 |
2015-07-11
|
초록
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Ⅳ. 연구개발결과
□ 장류 진균의 다양성 확보, 특성 평가 및 자원화
○ 장류 진균(사상균, 효모)의 분포상 조사 및 다양성 확보
- 된장 초기에는 사상균 Mucor, Scopulariopsis, Aspergillus, Penicillium등이 관찰되었으나 후기(9월)에는 Lichtheimia, Monascus, Eurotium 등이 관찰됨
- 된장에서 Debaryomyces hansenii, Meyerozyma guillermondii, Rhodotorula mucilaginosa 등, 간장에서 Ca
Ⅳ. 연구개발결과
□ 장류 진균의 다양성 확보, 특성 평가 및 자원화
○ 장류 진균(사상균, 효모)의 분포상 조사 및 다양성 확보
- 된장 초기에는 사상균 Mucor, Scopulariopsis, Aspergillus, Penicillium등이 관찰되었으나 후기(9월)에는 Lichtheimia, Monascus, Eurotium 등이 관찰됨
- 된장에서 Debaryomyces hansenii, Meyerozyma guillermondii, Rhodotorula mucilaginosa 등, 간장에서 Candida versatalis, Debaryomyces hansenii, Rhodotorula mucilaginosa, Wickerhamomyces anomalus 등 6속 18종 220균주 효모 수집
○ 메주 사상균의 특성평가
- 메주에서 분리한 26속 101종 1479균주 중에서 배지상에서 Amylase, Protease 우수 생성 116 균주를 선발
- 다시 콩발효물에서 효소 우수생성 16 균주 선발
- 선발된 16 균주로 콩발효물을 제조하고 NMR 분석을 통하여 생성 유기물을 조사하여 장류 발효 특성이 우수한 Aspergillus candidus M2079 등 7 균주에 대하여 특허 출원
○ 메주 사상균의 유래 조사
- 메주에서 주로 분리되는 곰팡이 중에서 Penicillium polonicum, P. solitum은 볏짚에서, Mucor circinelloides, M. racemosus, Lichtheimia ramosa, Scopulariopsis brevicaulis는 공기에서 Aspergillus oryzae, Eurotium repens, E. chevalieri 는 볏짚, 공기, 콩 등에 존재.
- 볏짚에 높은 빈도로 존재하는A. oryzae, E. repens, E. chevalieri, P. polonicum, P. solitum는 볏짚에서 메주에 전이된 후에 메주에서 발생하면 발효실에 상존할 가능성이 높음. 따라서 매년 사용하는 발효실은 볏짚을 사용하지 않더라도 주요 균은 메주에서 발생. 볏짚은 메주에 유용한 곰팡이를 공급 하지만 때로는 유해한 곰팡이를 공급할 수도 있으므로 적절한 활용이 필요
□ 농업 환경과 청국장 유래 세균의 다양성 확보 및 특성 평가
○ 청국장 제조용 발효콩으로부터 세균 수집 : 총 224균주
- 청국장 발효콩 시료 15점으로부터 균주 분리 : 224 균주
- 분리 균주의 16S rRNA 염기서열 분석 : Bacillus subtilis 등 Bacillus 속 171균주, nterococcus, Lactococcus 등의 유산균울 포함한 균주 53균주
Bacillus 속 균주들의 계통수 작성
- Pyrosequecing을 통한 발효콩 시료의 분석 : Firmicutes가 96∼99%로 대부분 관찰됨
○ 청국장 유래 균주 선발 및 선발균주의 특성 검정 : Bacillus 속 8균주 선발
- 16S rRNA, RAPD, γ-Polyglutamate을 이용한 1차 균주 선발(88균주)
: 각 분석별 다양한 특성을 가지는 균주 88균주 1차 선발
- 다양한 특성 검정으로 2차 균주 선발(47균주)
효소활성, 혈전용해능, 독소유전자 유무, 항균력 검정, 바이오제닉 아민 생성틍,
바이오제닉 분해능으로 47 균주 2차 선발
- 선발균주에 의한 발효콩의 특성 분석(최종 8균주 선발)
관능 분석, 휘발성 아민 분석, metabolite 분석으로 최종 8균주 선발
○ 수집 세균의 분류학적 특성 구명: Compostimonas suwonensis 등 12속 28종 분석
- 농업환경 유래 신속 및 신종 보고: Compostimonas suwonensis gen.. nov., sp. nov. 등 신속 12속, 신종 28종 보고
□ 야생 버섯 자원의 다양성 확보 및 특성 평가
○ 야생버섯의 분포 조사 및 종다양성 연구
- 강릉 등 214개 지역에서 야생버섯류 32목 92과 279속 654종 4,052표본 수집, 동정
- 한국미기록종 및 희귀종버섯 : Melanophyllum eyrei 등 15종 동정
- 월드컵공원의 종 모니터링 결과
평화공원에서 자생하는 버섯류 10목 46과 32속 64종 105표본
하늘공원의 버섯류 : 8목 25과 31속 35종 43표본
노을공원의 버섯류 5목 13과 15속 16종 22표본
난지천공원의 버섯류 5목 9과 13속 14종 15표본 수집 동정함
- 농과원 농업생물부 생태원 버섯류 종다양성 조사 결과 경단버섯류 등 15목 33과 55속 77종 107표본 동정
- 분자유전학적 특성평가(ITS) : ASIS 23101 Amanita melleiceps 등 80표본 분석
○ 아속· 종별 분류법 및 한국자생버섯의 분류검색표 작성
- 송이과 속, 종검색표 작성 및 특징 기술: 낭상체버섯속, 솔밭버섯속, 뽕나무버섯속, 겨자버섯속, 패랭이버섯속
○ 국가버섯표준목록집 작성 : 느타리 등 국내기록종 1,901종 작성
- 버섯이름의 원기재문 작성 : 1,901종
- 문헌조사 : 한국버섯이름통일안, 1978, 한국균학회지)등 328편
○ 자생버섯류의 확증표본제작
- 당해연도 수집 표본 1,200표본 중 700여 표본 제작
- 확증표본자료의 통합DB구축 : 표본 5,000점 및 이미지자료 슬라이드 10,000장
(‘13년 생명자원 통합DB구축 사업(BRIS)으로 농업유전자원종합관리시스템과 연계)
Abstract
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Microorganisms play very important roles in the field of agriculture and food, and Rural Development Administration established a microbial resources center, Korean Agricultural Culture Collection (KACC). KACC mange agricultural and food related microorganisms and supply them to institute, universit
Microorganisms play very important roles in the field of agriculture and food, and Rural Development Administration established a microbial resources center, Korean Agricultural Culture Collection (KACC). KACC mange agricultural and food related microorganisms and supply them to institute, university and company for research, education and industrialization. In this project, we aimed to collect and characterize indigenous microorganisms from agricultural and food environments, and we deposit them in KACC. The project is composed of three parts, the first is collection and characterization of fungi from traditional Korean Jang, the second is collection and characterization of bacteria from Cheonggukjang and agricultural environments, and the last is collection and characterization of wild mushrooms in Korea.
Dominant genera on Meju are Mucor, Lichtheimia, Rhizopus in Zygomaycoa and Aspergillus, Penicillium, Eurotium, Scopulariopsis, Cladosporium in Ascomycota. Meju submerge in salt water and become to Doenjang. In the early stage of Doenjang, the predominant genera were Mucor, Scopulariopsis, Aspergillus, Penicillium but in the later stage, predominant genera were Lichtheimia, Monascus, Eurotium which are xerophilics. Dominant yeast species in Doenjang were Debaryomyces hansenii, Meyerozyma guillermondii, Rhodotorula mucilaginosa.
In order to develop new starters for standardization and industrialization of Meju production, we examined extracellular enzyme activities of Meju fungi. We selected 116 strains with high amylase and protease activity on media, and additionally we chose 16 strains which show high extracellular enzyme activities on boiled soybeans. We made fermented soybean products with the fungi and examined quality properties of the soybean products by 1H-NMR analysis. Strains of Aspergillus candidus, Aspergillus oryzae, Lichtheimia ramosa, Scopulariopsis brevicaulis, Mcucor circinelloides, Penicillium polonicum, Penicillium roqueforti , Penicillium solitum, Rhizomucor pusillus and Rhizopus oryzae produced larger amount of glutamate and glycine which taste umami, than Aspergillys oryzae strain which is usually used in Jang industry in Korea. We also examined growth of the selected fungi on various temperatures and media in order to optimize conditions for industrialization. Seven of them were applied for patent.
In the study on the origin of the fungi on Meju, the mycobiota of the soybeans, rice straws and the air in J ang factories were determined. Aspergillus oryzae, Eurotium repens and E. chevalieri were frequently isolated from the soybeans, rice straws and air of Jang factories. Penicillium polonicum and P. solitum were frequently isolated from the rice straws and were rarely detected on the soybeans and in the air of Jang factories. Therefore, the rice straws could act as an important provider of fungi on Meju and it has been observed that many fungi are transferred from rice straw to Meju. However, the rice straws also provide Meju with fungi that is not wanted in Meju fermentations. It could be a good method to only use rice straws during the high temperature fermentation process, in order not to be contaminated with unwanted fungi.
Cheonggukjang is a unique sauce made of soybeans in Korea. It is fermented naturally using Bacillus in the air and straw. One of strong points of Cheonggukjang is that the fermentation period is shorter than other sauces. Bacillus which makes the Cheonggukjang is an aerobic gram-positive and widely distributed in nature such as soil. Due to easy growth at medium, Bacillus is generally applied in industry, medicine and ecology areas. The objects of this study are not only to identify Bacillus strains isolated from Cheonggukjang, a Korean traditional fermented food, and analyze their functional properties, but also to select Bacillus strains producing the high-quality Cheonggukjang.
We collected fermented soybeans of tasty and famous fifteen Cheonggukjang all over the country. Colonies different in morphology were isolated from fifteen Cheonggukjang samples all over the country, and 16S rRNA sequences were analyzed for their taxonomic identification. A total of 224 strains were isolated from the Cheonggukjang samples. They were mainly genus Bacillus and lactic acid bacteria. Pyrosequencing was performed to determine the microbial community of six whole fermented soybeans samples. The result of phylum level analysis showed a high percentage on Firmicutes phylum including Bacillus. In Bacillus genus level, these results of distribution patterns were similar with those of the strains isolated in the plates. First, on the basis of DNA fingerprinting, 16S rRNA sequence analysis and γ-PGA production pattern, 78 strains were selected. Selected 78 strains showed positive activities of amylase, cellulase, protease and lipase, and 38 strains have positive fibrinolytic activity. To ensure the safety of isolated strainsfrom the samples of Cheonggukjang which are manufactured by traditional method of production, strains were tested for the presence of seven toxin genes of Bacillus cereus strains which has close phylogenetic relationship of the Bacillus subtilis strains. All 78 strains showed negative results. In order to select strains secreting antimicrobial substances which inhibits pathogenic fungi or bacteria, antibiotic test was performed. 50 strains produced antibiotic substance. The biogenic amines produced during the fermentation of Cheongukjang cause allergy and food poisoning. Among the strains tested, 49 strains did not produce or merely produce both histamine and tyramine which are the most representative biogenic amines, and 37 strains could degrade the biogenic amines. 44 strains including 5 reference strains were selected in the these steps. In this biogenic amine degradation analysis by HPLC, strain 2RL2-1 showd approximately 47% tyramine degradation activity. Fermentation property, sensory test and volatile basic nitrogen production were tested for checking that the strains are actually superior to fermentation capability. Eight strains was selected to make high-quality Cheonggukjang. Metabolic profiles by 1H-NMR for each Cheonggukjangs were analyzed.
To the country of mushrooms biodiversity research from 2014 to 2012 were studied conservation forest area of the 21 National Parks and Provincial Park. Were identified and referred specimens. In the 241 area ASIS25666 Phellinus lonicerina 4149, including collected 5 phylum 10, class 6 subclass, 32 order, 92 families 279 genera and 654 species. Examine the mushrooms of the distribution ratio of the total collection of fungi species present results showed the highest number of occurrences Amanitaceae to 490 samples. Russulaceae of 400 specimens of fungi Agaricaceae 378 specimens, Boletaceae the 306 samples, Tricholomataceae was identified as 229 samples. Among the rare species is Psathyrella pervelatoid, Clitocybe alboinfundibuliforme, Volvariella koreana including five species have been confirmed. Domestic unrecorded and new candidate species Melanophyllum haematospermum (Bull.) Kreisel, including 10 species have been identified. In addition, survey results for each life aspects exogenous Mycorrhizal 1,745 specimens (42%) were wood rot fungi is 1393 samples (33%), compost live parts are 922 samples (22%), insect parasites of 46 samples (1%), parasites are 37 were sampled. In order to take advantage of the development of bioactive materials collected samples as the basis for edible mushrooms, medicinal, and determine whether a poisonous mushroom. Edible mushrooms such clusters were 1,102 specimens (26%), medicinal mushrooms such bulrocho 716 samples (17%) were identified as poisonous Amanita virosa, including 853 samples (20%).
The insect ecology garden, it is where you have to observe the change of artificially insects by planting the 128 species of insect host plant and rape, such as more than 100 kinds of plants such as Fupumiru pepper. For observation of insect ecology garden of fallen leaves and mushrooms that occur in compost by examining the 23 times until July 2014 before moving to a new front of the earth in August 2012, such as Dumontinia tuberosa collected 154 specimens 19 neck 38 and I have identified 118 species in 60. Appearance of mushrooms, but perennial mushroom and is one year, such as Coriolus versicolor the configuration of the organization does not disappear easily to become from the binding hyphae and skeletal fungus company has remained until the following year, can be observed throughout the year . However, most of the mushrooms, by the influence of temperature and humidity, occurred in March to October. Most advent paper many time was a 45 specimens was in July.
This survey was conducted in order to clarify the species diversity and distribution of mushrooms at Worldcup Park in Seoul, from May to November 2010 and 2014. In the survey, a total of 98 taxa, 98 species, 86 genera, 38 families, 12 orders, three classes, and two phyla were identified. A total of 250 specimens of Mushrooms were collected from Pyonghwa Park and 63species, 71 genera, 36 families, 13 orders, three classes, and two phyla were identified. Coprinus atramentarius and Psathyrella candolleana were high in frequency, as ten times and nine times, respectively. In Noeul Park, 102 specimens were collected, 41species, 42 genera, 19 families, 19 orders, two classes, and two phyla were identified. In Noeul Park mostly layed with lawn, Agrocybe semiorbicularis and Agaricus campestris were high in frequency, as ten times and nine times, respectively. There were no mushrooms in succession during three years. In Haneul Park, a total of 128 specimens of mushrooms were collected. 49 species, 57 genera, 30 families, 11 orders, three classes, and two phyla were identified. Agrocybe semiorbicularis and Phallus rugulosus were high in frequency, as four times. In Nanjicheon Park, a total of 68 specimens of mushrooms were collected. 31 species, 34 genera, 30 families, nine orders, one class, and one phylum were identified. Coprinus atramentarius and Psathyrella velutina were high in frequency, as three times. Investigation on the diversity of mushrooms at the Worldcup Park, piled up with the city's waste, will be of the great significance in the aspect of the ecological change. Local indigenous mushrooms wrote a classification taxonomy and search key of the subgenus and species native to Korea and mushrooms. Voucher specimens production and preservation of indigenous fungi are HCCN25302 Leucopaxillus sp. Including 3,200 specimens were produced. All specimens examined in this work are preserved in HCCN (Herbarium Conservation Center of National Academy of Agricultural Science).
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