KR101825957B1 - Saccharomyces cerevisiae Y297 with adaptability at low temperature and manufacturing method of fermented alchoholic beverage using the same - Google Patents
Saccharomyces cerevisiae Y297 with adaptability at low temperature and manufacturing method of fermented alchoholic beverage using the same Download PDFInfo
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Abstract
본 발명은 기탁번호 KACC93244P로 기탁된 신규한 사카로마이세스 세레비지(Saccharomyces cerevisiae)에 Y297 및 이를 이용하여 제조된 발효주에 관한 것으로, 구체적으로 500종의 발효 균주 중 15℃의 저온 조건에서 생장이 뛰어난 효모, 사카로마이세스 세레비지에 Y297을 선발하였고, 상기 균주는 저온 내성 및 알코올 생성능이 우수함을 확인하였으며, 효모의 향기성분 생산과 관련된 효소 생성능을 비교하여 Y297 효모의 glucosidase, esterase의 활성이 상업효모 대비 2~5배 우수함을 확인하였으며, 발효주의 품질 특성 확인결과, 상업효모에 비해 산도가 낮고, 알코올 함량이 높으며, 유기산 함량이 높고, 유리아미노산 함량 높음을 확인하여, 본 발명의 저온 적응성 Y297 효모는 감칠맛과 단맛, 향기가 우수한 고품질의 발효주 생산에 유용하게 이용될 수 있다.The present invention relates to a new yeast Saccharomyces cerevisiae Y297 deposited with the deposit number KACC93244P and a fermentation liquor produced using the same. Specifically, among 500 fermentation strains, growth is carried out at a low temperature of 15 ° C Y297 was selected as an excellent yeast, Saccharomyces cerevisiae. The strain was found to have excellent low-temperature tolerance and alcohol-producing ability, and the activity of glucosidase and esterase of Y297 yeast was compared with that of yeast It was confirmed that the quality of the fermented wine was 2 to 5 times better than that of the commercial yeast. As a result of confirming the quality characteristics of the fermented wine, the acidity was low, the alcohol content was high, the organic acid content was high and the free amino acid content was high. Y297 yeast can be used to produce high quality fermented beverage with excellent flavor, sweetness and aroma.
Description
본 발명은 기탁번호 KACC93244P로 기탁된 신규한 사카로마이세스 세레비지에(Saccharomyces cerevisiae) Y297 균주, 상기 균주 또는 이의 배양액을 포함하는 생균제 조성물, 이를 이용한 발효주 및 이의 제조방법에 관한 것이다.The present invention relates to a novel Saccharomyces cerevisiae strain Y297 deposited with the deposit number KACC93244P, a strain or a culture thereof, a fermentation liquor using the same and a method for producing the same.
우리나라의 전통주인 발효주는 미생물에 의해 탄수화물이 분해되면서 알코올을 비롯한 여러 가지 성분이 생기는 일종의 발효 음료이며, 발효원으로는 전분을 주성분으로 하는 곡류, 감자류와 당분을 주성분으로 하는 과일, 당밀 등이 이용된다.A traditional fermented beverage in Korea is a type of fermented beverage that contains various components such as alcohol as a result of decomposition of carbohydrates by microorganisms. Examples of fermentation sources are cereals based on starch, fruits such as potatoes and sugar-based fruits, molasses do.
발효주로서의 약주 및 탁주는 전통적으로 곡류와 누룩을 사용하여 병행복발효로 제조되며, 양조 후에 술덧을 체로 걸러서 외관이 백탁한 것은 막걸리 또는 탁주라 하고, 술덧에 용수를 받아서 맑은 액만 취한 것을 약주라 한다.Yakju and Takju as fermented beans are traditionally made by using the cereals and yeast, and they are made from jade fermentation. After brewing, the sake is sieved and the appearance is opaque is called makgeolli or takju. .
일반적으로 담금 후, 누룩 중의 미생물에 의한 효소작용으로 원료성분이 분해, 발효되어 알코올 농도가 12-15%가 되며, 이것을 거를 때 물을 첨가하여 알코올 농도 6-8%의 탁주를 제조한다. 탁주는 비타민 B군을 비롯한 필수아미노산인 lysine, leucine, glutamic acid, proline 및 glutathion을 함유하여 영양가가 풍부하며, 산미에 관여하는 유기산은 갈증을 해소하고 신진대사를 원활히 하는데 효과가 있다.Generally, after immersion, the raw material component is decomposed and fermented by an enzyme action by the microorganism in the yeast, so that the alcohol concentration becomes 12-15%, and water is added thereto to prepare Takju with an alcohol concentration of 6-8%. Takju contains nutrients such as lysine, leucine, glutamic acid, proline and glutathion which are essential amino acids including vitamin B group. Organic acid which is involved in acidity is effective in relieving thirst and facilitating metabolism.
약주를 제조하는 과정에서 효모는 알코올을 생성하는 발효과정에 직접적으로 관여할 뿐 아니라, 다양한 대사산물과 휘발성 향미성분을 생산하는 등의 약주의 품질을 형성하는 중요한 역학을 한다(Baek SY., 등, Microbiol Biotechnol lett, 2015, 43, 1-9, Lee SR., 등, 1985, Ewha women's university press, p224-294, Shin KR., 등, 1999, J Korean Soc Food Sci Nutr, 28, 794-800). 그러나, 약주 제조의 주요한 효모 중 하나인 사카로마이세스 세레비지에(Saccharomyces cerevisiae)의 경우, 단백질 전사, 세포막의 유동성, 효소 활성과 같은 특성도 저온의 영향을 받아, 균 생장 정도가 저하되거나 이상발효를 일으키는 등의 좋지 못한 영향을 주기도 한다. 따라서, 저온 적응성이 뛰어난 효모를 이용하여 발효를 할 수 있다면 좀 더 좋은 품질의 약주를 생산할 수 있을 것이다.In the process of manufacturing Yakju, yeast is important not only directly involved in the fermentation process that produces alcohol but also forming the quality of Yakju, such as producing various metabolites and volatile flavor components (Baek SY, et al. , Microbiol Biotechnol Lett, 2015, 43, 1-9, Lee SR., Et al., 1985, Ewha women's university press, p224-294, Shin KR, et al., 1999, J Korean Soc Food Sci Nutr, 28, 794-800 ). However, in the case of Saccharomyces cerevisiae , which is one of the major yeasts in the manufacture of Yakju, the properties such as protein transcription, cell membrane fluidity, and enzyme activity are also affected by low temperatures, It can also cause bad effects such as fermentation. Therefore, if fermentation can be performed using a yeast having excellent adaptability to low temperature, it will be possible to produce a better quality of the Yakju.
약주 제조 과정에서 온도는 술의 품질을 좌우하는 휘발성 향미 성분을 생산하는데 있어서 중요한 인자로 알려져 있고, 와인 제조의 경우 저온발효(10~15℃)는 primary aroma의 손실을 방지할 수 있고, secondary aroma(ehyl 과 acertate esters 등)의 합성을 증가시키는 것으로 알려져 있다(Beltran G., 등, 2007, World j Microbiol Biotechnol, 23, 809-815, Chiva R., 등, 2012, FEMS Yeast Res, 831-843, Torija MJ., 등, 2003, Int J Food Microbiol, 80, 47-53). 또한, 10~12℃의 온도에서의 발효는 ester의 생산을 75%이상 증가시킨다는 결과도 알려져 있다(Engan S., 1977, 16th European brewery convention congress, Amsterdam, Netherlands, 591-607).The temperature during the fermentation process is known to be an important factor in producing the volatile flavor components that influence the quality of the wine. In the case of wine production, low temperature fermentation (10 ~ 15 ℃) (Beltran G., et al., 2007, World J Microbiol Biotechnol, 23, 809-815, Chiva R., et al., 2012, FEMS Yeast Res, 831-843 , Torija MJ., Et al., 2003, Int J Food Microbiol, 80, 47-53). It is also known that fermentation at temperatures between 10 and 12 ° C increases ester production by more than 75% (Engan S., 1977, 16 th European brewery convention congress, Amsterdam, Netherlands, 591-607).
유기산과 알코올이 결합하여 만들어지는 ester는 다양한 음식의 풍미 성분으로 알려져 있으며, 발효주에서는 향기 성분의 주요 지표로서 사용되고 있다. 와인의 경우, ester의 생산은 Esterase(EC 3.1.1.1)에 의한 가수분해 반응과 alcohol acetyltransferase(EC 2.3.1.84)를 포함한 합성반응의 상호작용에 의해 이루어진다. 사카로마이세스 속 균주의 경우 ester의 합성과 가수분해에 관련된 유전자를 갖고 있음이 여러연구를 통해 보고되고 있다.Ester, which is formed by the combination of organic acid and alcohol, is known as a flavor ingredient of various foods, and is used as a main index of flavor ingredient in fermented beverages. In the case of wines, the production of esters is achieved by the interaction of hydrolysis with esterase (EC 3.1.1.1) and synthetic reactions involving alcohol acetyltransferase (EC 2.3.1.84). Several studies have reported that the Saccharomyces sp. Strain has genes involved in ester synthesis and hydrolysis.
전통주 제조와 관련된 선행기술로써, 대한민국 공개특허 10-2004-0117468에는 신규 젖산균 및 이를 이용한 발효주의 제조방법이 개시되어 있고, 10-2010-0138957에는 저온 내성을 갖는 효모 변이체를 발굴함에 대하여 개시하고 있으나, 저온 조건에서 생장이 뛰어난 효모, 및 이를 이용한 발효주에 대해서는 알려진 바가 없다.Korean Patent Laid-open Publication No. 10-2004-0117468 discloses a novel lactic acid bacterium and a method for producing a fermented liquor using the lactic acid bacterium as a prior art related to the manufacture of a traditional liquor, and 10-2010-0138957 discloses discovery of a yeast mutant having a low temperature tolerance , A yeast excellent in growth under a low temperature condition, and a fermentation product using the yeast are not known.
이에, 상기 문제점을 해결하고자, 본 발명자들은 500종의 발효 균주 중에서 15℃의 저온 조건에서 생장이 뛰어난 효모, 사카로마이세스 세레비지에 Y297을 선발하였고, 상기 균주는 저온 내성 및 알코올 생성능이 우수함을 확인하였으며, 효모의 향기성분 생산과 관련된 효소 생성능을 비교하여 Y297 효모의 glucosidase, esterase 의 활성이 상업효모 대비 2~5배 우수함을 확인하였으며, Y297 효모를 이용하여 제조된 발효주의 품질 특성 확인결과, 상업효모에 비해 산도가 낮고, 알코올 함량, 유기산 함량, 및 유리아미노산 함량이 높음을 확인하여, 본 발명의 저온 적응성 Y297 효모는 감칠맛과 단맛, 향기가 우수한 고품질의 발효주 생산에 유용하게 이용될 수 있음을 확인함으로써, 본 발명을 완성하였다.In order to solve the above problems, the present inventors selected Y297 as a yeast, Saccharomyces cerevisiae, which is excellent in growth at a low temperature of 15 ° C among 500 fermenting strains. The strain has excellent low-temperature tolerance and alcohol producing ability And the activity of glucosidase and esterase of Y297 yeast was found to be 2 to 5 times better than that of commercial yeast. The results of the confirmation of the quality characteristics of Y297 yeast , Low acidity, high alcohol content, organic acid content, and free amino acid content, compared with commercial yeast. Thus, the low temperature adaptable Y297 yeast of the present invention can be used for producing high quality fermented liquor with excellent flavor, sweetness and aroma The present invention has been completed.
본 발명의 목적은 기탁번호 KACC93244P로 기탁된 신규한 사카로마이세스 세레비지에(Saccharomyces cerevisiae) Y297 균주, 상기 균주 또는 이의 배양액을 포함하는 생균제 조성물, 및 이를 이용한 발효주 및 이의 제조방법에 관한 것이다.An object of the present invention is a novel Saccharomyces cerevisiae Y297 strain deposited with Accession No. KACC93244P, a prophylactic composition comprising the strain or a culture thereof, a fermentation liquor using the same and a method for producing the same.
상기 목적을 달성하기 위하여, 본 발명은 In order to achieve the above object,
KACC93244P로 기탁된 신규한 사카로마이세스 세레비지애(Sacharomyces cerevisiae) Y297균주를 제공한다.A novel strain of Sacharomyces cerevisiae Y297 deposited with KACC93244P is provided.
또한, 상기 균주 또는 이의 배양액을 포함하는 생균제 조성물을 제공한다. The present invention also provides a probiotic composition comprising the strain or a culture thereof.
또한, 본 발명은 In addition,
1) 제 1항의 균주를 곡류에 접종하는 단계; 및1) Inoculating the cereal of claim 1 into cereals; And
2) 상기 단계 1)의 곡류를 발효시키는 단계를 포함하는 발효주 제조방법을 제공한다.2) fermenting the cereal in the step 1).
아울러 본 발명은, 상기 발효주 제조방법으로 제조된 발효주를 제공한다. In addition, the present invention provides a fermented beverage produced by the method for producing a fermented beverage.
본 발명에서 500종의 발효 균주 중 15℃의 저온 조건에서 생장이 뛰어난 효모, 사카로마이세스 세레비지에 Y297을 선발하였고, 상기 균주는 저온 내성 및 알코올 생성능이 우수함을 확인하였으며, 효모의 향기성분 생산과 관련된 효소 생성능을 비교하여 Y297 효모의 glucosidase, esterase의 활성이 상업효모 대비 2~5배 우수함을 확인하였으며, Y297 효모를 이용하여 제조된 발효주의 품질 특성 확인결과, 상업효모에 비해 산도가 낮고, 알코올 함량, 유기산 함량, 및 유리아미노산 함량 높음을 확인하여, 본 발명의 저온 적응성 Y297 효모는 감칠맛과 단맛, 향기가 우수한 고품질의 발효주 생산에 유용하게 이용될 수 있다.In the present invention, among the 500 fermenting strains, Y297 was selected as a yeast, Saccharomyces cerevisiae, which is excellent in growth at a low temperature of 15 ° C. The strain was found to have excellent low-temperature resistance and alcohol-producing ability, The activity of glucosidase and esterase of Y297 yeast was found to be 2 ~ 5 times better than that of commercial yeast. The results of the quality characteristics of Y297 yeast were found to be lower than that of commercial yeast , Alcohol content, organic acid content, and free amino acid content, the low temperature adaptable Y297 yeast of the present invention can be usefully used for producing high quality fermented liquor having excellent flavor, sweetness and aroma.
도 1은 Y297 효모를 주사전자현미경으로 관찰한 도이다.
도 2는 Y297 효모의 동정 및 계통도를 나타낸 도이다.
도 3은 저온에서 Y297의 효소활성을 측정하여 나타낸 도이다.
도 4는 발효온도에 따른 저온 적응성 Y297 효모로 제조된 발효주의 향기패턴을 나타낸 도이다.
도 5는 상업 효모 또는 Y297 효모를 이용하여 제조된 발효주의 향기성분을 나타낸 크로마토그램이다.Fig. 1 is a view of Y297 yeast observed with a scanning electron microscope.
Fig. 2 is a diagram showing the identification and flow diagram of Y297 yeast.
FIG. 3 is a graph showing the enzyme activity of Y297 measured at low temperature.
FIG. 4 is a view showing a flavor pattern of a fermentation broth prepared from a low temperature adaptable Y297 yeast according to a fermentation temperature. FIG.
FIG. 5 is a chromatogram showing the aroma component of the fermented wine produced using commercial yeast or Y297 yeast.
이하, 본 발명을 상세히 설명한다.Hereinafter, the present invention will be described in detail.
본 발명은 KACC93244P로 기탁된 신규한 사카로마이세스 세레비지애(Sacharomyces cerevisiae) Y297 균주를 제공한다. The present invention provides a novel strain of Sacharomyces cerevisiae Y297 deposited with KACC93244P.
상기 균주는 서열번호 1의 26s rDNA의 염기서열을 갖는 것이 바람직하고, 10 내지 20℃의 저온생장에 내성이 있는 것이 바람직하다. Preferably, the strain has a nucleotide sequence of 26s rDNA of SEQ ID NO: 1 and is resistant to low-temperature growth at 10 to 20 ° C.
본 발명의 구체적인 실시예에서, 482종의 발효균를 15℃의 저온 조건에서 배양하여 대조구에 비해 생균수가 높은 효모로써, Y297 효모를 선발하였고(표 1 참조), 계통 분석을 통해 상기 균주가 사카로마이세스 세레비지에 계통임을 확인하였으며, 이를 사카로마이세스 세레비지에 Y297로 명명하고(도 2 참조), 상기 균주를 2015년 11월 10일자로 기탁기관인 국립농업과학원에 기탁하였다(기탁번호: KACC93244P).In a specific example of the present invention, 482 kinds of fermenting bacteria were cultured at a low temperature of 15 ° C to select Y297 yeast as a yeast having a higher viable cell count than the control (see Table 1) (See Fig. 2). The strain was deposited on Nov. 10, 2015 with the National Institute of Agricultural Science and Technology (National Academy of Agricultural Science) (Accession No .: KACC93244P ).
또한, 상기 균주의 특성을 분석한 결과, 저온내성 및 알코올 생성능이 우수함을 확인하였고(표 2참조), 효모의 향기성분 생산과 관련된 효소 생성능을 비교하여 Y297효모의 glucosidase 및 esterase활성이 상업효모 대비 2~5배 우수함을 확인하였다(도 3참조). As a result of analyzing the characteristics of the above strains, it was confirmed that the low temperature resistance and alcohol producing ability were excellent (see Table 2), and the enzyme production ability related to the production of aroma components of yeast was compared to determine the glucosidase and esterase activities of Y297 yeast 2 to 5 times higher (see Fig. 3).
따라서, 본 발명의 균주는 저온내성 및 알코올 생성능이 우수한 특징을 갖는 신규한 사카로마이세스 세레비지에 Y297 균주(KACC93244P)임을 확인하였다.Therefore, it was confirmed that the strain of the present invention is a strain Y297 (KACC93244P) in a novel saccharomyces cerevisiae which is characterized by excellent low-temperature resistance and alcohol-producing ability.
또한, 본 발명은 상기 균주 또는 이의 배양액을 포함하는 생균제 조성물을 제공한다. In addition, the present invention provides a biocidal composition comprising the strain or a culture thereof.
상기 생균제 조성물은 통상적인 약학적 담체 및 부형제를 추가로 포함할 수 있으며, 이러한 조성물은 통상적인 생균제 조성물 제조방법에 따라 동결건조 또는 열건조 될 수 있고, 캡슐화된 형태 또는 배양 현탁액이나 건조 분말 형태일 수 있다.The biocide composition may further comprise conventional pharmaceutical carriers and excipients. The composition may be freeze-dried or heat-dried according to a conventional method for producing a biocidal composition, and may be in an encapsulated form or in a culture suspension or a dry powder form. .
또한 본 발명은, Further, according to the present invention,
1) 제 1항의 균주를 곡류에 접종하는 단계; 및1) Inoculating the cereal of claim 1 into cereals; And
2) 상기 단계 1)의 곡류를 발효시키는 단계를 포함하는 발효주 제조방법을 제공한다.2) fermenting the cereal in the step 1).
상기 방법에 있어서, 상기 단계 1)의 곡류는 쌀, 보리, 밀, 또는 고구마인 것이 바람직하나, 이에 한정되지 않는다.In the above method, the cereal in step 1) is preferably rice, barley, wheat, or sweet potato, but is not limited thereto.
아울러 본 발명은, 상기 발효주 제조방법으로 제조된 발효주를 제공한다.In addition, the present invention provides a fermented beverage produced by the method for producing a fermented beverage.
본 발명의 구체적인 실시예에서, 상기 Y297 효모로 발효한 발효주의 품질 특성을 확인한 결과, 상업효모를 이용하여 발효한 발효주보다 pH가 0.3~0.5% 정도 높고, 산도는 0.05~0.15% 정도 낮았으며, 저온발효 후의 알코올 함량이 1~3% 높아, 저온에서도 안정적으로 발효가 일어남을 확인하였다(표 3 내지 표 5 참조). 또한, 유기산 함량이 높고, 필수아미노산 및 비필수아미노산의 함량이 높음을 확인하였으며, 특히 감칠맛과 단맛을 내는 아미노산인 aspartic acid, glutamic acid 및 alanine의 함량이 높은것을 확인하였다(표 6 및 표 7 참조). 또한, 저온적응성 효모 Y297로 발효한 발효주의 향기패턴 및 향기 성분이 대조구와 다르게 나타났으며(도 4, 도 5, 및 표 8 참조), 관능평가 결과, 기호도, 맛, 및 색상 평가에서 높은 점수를 받았다(표 9 참조). In a specific example of the present invention, the quality characteristics of the fermentation broth fermented with the yeast Y297 yeast were examined. As a result, the pH was 0.3 to 0.5% higher and the acidity was lower by 0.05 to 0.15% than the fermented broth fermented with commercial yeast. It was confirmed that the alcohol content after the low temperature fermentation was 1 to 3% higher, and the fermentation stably occurred even at a low temperature (see Tables 3 to 5). In addition, it was confirmed that the content of essential amino acids and non-essential amino acids was high, and the contents of aspartic acid, glutamic acid and alanine, which are rich in the flavor and sweet taste, were high (see Table 6 and Table 7) ). In addition, the aroma pattern and aroma component of the fermented fermented product fermented with the low temperature adaptable yeast Y297 were different from those of the control (see Figs. 4, 5 and 8). As a result of the sensory evaluation, (See Table 9).
따라서, 본 발명의 저온적응성 Y297 효모로 발효한 발효주는 기존의 발효주와 다른 감칠맛과 단맛, 그리고 다른 향기를 가진 고품질의 발효주임을 확인하였다. Therefore, it was confirmed that the fermentation broth fermented by the low temperature adaptable Y297 yeast of the present invention is a high quality fermentation broth having a different flavor, sweetness, and other aroma than the conventional fermentation broth.
이하, 본 발명을 실시예 및 실험예에 의해서 상세히 설명한다.Hereinafter, the present invention will be described in detail with reference to Examples and Experimental Examples.
단, 하기 실시예 및 실험예는 본 발명을 예시하는 것일 뿐, 본 발명의 내용이 하기 실시예 및 실험예에 의해서 한정되는 것은 아니다.However, the following Examples and Experimental Examples are merely illustrative of the present invention, and the content of the present invention is not limited by the following Examples and Experimental Examples.
<실시예 1> 저온(15℃) 조건에서 생장 및 알코올 생성능 우수 효모 선발<Example 1> Selection of yeast having excellent growth and alcohol production ability at a low temperature (15 ° C)
저온 조건에서의 생장 및 알코올 생성능이 우수한 효모를 선발하기 위하여, 전국에서 수집한 메주, 누룩 등의 발효식품 50 여종으로부터 분리한 482개의 균주를 사용하였다. 비교 균주로는 시판 효모인 Fermivin(S. cerevisiae, Oenobrands, France; Con 1)와 La parisienne (S. cerevisiae, DB ingredients, UK; Con 2) 사용하였다. 실험에 사용된 효모는 이스트 추출물(yeast extract) 1%, 펩톤(peptone) 2%, 덱스트로스(dextros) 2% (Difico, Becton Dickinson and company, spark, USA)를 함유하는 액체배지(YPD)에서 배양한 후, 글리세롤을 20%(v/v)로 첨가하여 -80℃에서 보관하여 실험에 사용하였다. 482 strains isolated from 50 kinds of fermented foods such as meju and leek were collected from the whole country in order to select yeast with excellent growth and alcohol production ability at low temperature. Fermivin ( S. cerevisiae , Oenobrands, France; Con 1) and La parisienne ( S. cerevisiae , DB ingredients, UK; Con 2) were used as comparative strains. The yeasts used in the experiment were liquid medium (YPD) containing 1% of yeast extract, 2% of peptone, 2% of dextrose (Difico, Becton Dickinson and Company, After incubation, glycerol was added at 20% (v / v) and stored at -80 ° C for use in the experiment.
그런 다음, YPD 한천 배지에 도말하여 2일간 배양한 후 순수분리하였다. 저온에서 생육이 우수한 효모를 선발하기 위해 진탕배양기(shaking incubator)에서 15℃에서 2일간 배양한 후 분광 광도계(spectrophotometer)를 이용하여 600nm로 흡광도를 측정하여, 저온(15℃)에서의 생균수를 확인한 결과, 표 1에 나타난 바와 같이, Y297 효모는 대조구보다 높은 생균수를 나타내어 저온에서 생육이 뛰어남을 확인하였고(표 1), 이를 통해 저온에서의 생육이 우수한 균주로써 사카로마이세스 세레비지에 Y297 효모를 최종 선발하였다. Then, the cells were plated on a YPD agar medium and cultured for 2 days, followed by pure separation. In order to select yeasts with good growth at low temperatures, the cells were incubated at 15 ° C for 2 days in a shaking incubator, and the absorbance was measured at 600 nm using a spectrophotometer. The number of viable cells at low temperature (15 ° C) As a result, as shown in Table 1, Y297 yeast showed higher viable cell count than the control, confirming excellent growth at low temperature (Table 1). As a result, it was confirmed that the strain Y297 Y297 yeast was finally selected.
또한, 저온(15℃) 알코올 발효 후의 알코올 생성 정도를 측정한 결과, 본 발명의 Y297 효모는 대조구 수준의 알코올 생성능을 나타냄을 확인하였다(표 2).Further, the degree of alcohol production after low-temperature (15 ° C) alcohol fermentation was measured. As a result, it was confirmed that Y297 yeast of the present invention had an alcohol-producing ability at the control level (Table 2).
<실시예 2> Y297 효모의 동정Example 2 Identification of Y297 Yeast
분리된 Y297 효모를 동정하기 위해 26S rDNA로 유전자의 D1/D2 부위 염기서열을 분석하였다.To identify the isolated Y297 yeast, the D1 / D2 region of the gene was analyzed with 26S rDNA.
구체적으로, NL1(5'-GCATATCAATAAGCGGAGGAAAAG-3'; 서열번호 2) 및 NL4(5'-GGTCCGTGTTTCAAGACGG-3'; 서열번호 3)의 프라이머를 사용하여 26S rDNA로 유전자의 D1/D2 부위 단편을 증폭 후, (주)ZENOTECH사에 의뢰하여 염기서열을 분석하였다. 염기서열은 NCBI(National center for biotechnology information) 인터넷 사이트의 BLAST 기능을 사용하여 유전자의 상동성을 비교하였으며, DNASTAR pro software를 사용하여 분석하였고, 효모의 계통분석도 작성은 MEGA6.0의 kimura2-parameter method를 이용하여 작성하였다.Specifically, the D1 / D2 region fragment of the gene was amplified with 26S rDNA using a primer of NL1 (5'-GCATATCAATAAGCGGAGGAAAAG-3 '; SEQ ID NO: 2) and NL4 (5'-GGTCCGTGTTTCAAGACGG-3' , And the nucleotide sequence was analyzed by ZENOTECH Co., Ltd. The homology of the genes was compared using the BLAST function of the National Center for Biotechnology Information (NCBI) website, and DNASTAR pro software was used to analyze the sequence. The gene analysis of the yeast was performed using the kimura2-parameter of MEGA6.0 method.
그 결과, 도 1에 나타난 바와 같이, 주사전자현미경(scanning electron Microscope, SEM)을 이용하여 Y297 효모의 형태를 확인한 결과, 타원 형태로 약 2~5 ㎛의 크기임을 확인하였다(도 1). 또한, 염기서열 분석에 의한 Y297 효모를 동정하기 위하여, Y297 효모의 26S rRNA (D1/D2) 부위를 염기서열 분석 후 표준 균주와 비교하여 계통도를 작성한 결과, Y297 효모는 사카로마이세스 세레비지에(Saccharomyces cerevisiae)로 동정되었다(도 2). As a result, as shown in FIG. 1, the shape of Y297 yeast was confirmed using a scanning electron microscope (SEM), and it was confirmed that the size of the Y297 yeast was about 2 to 5 μm in an elliptical shape (FIG. 1). In order to identify the Y297 yeast by the nucleotide sequence analysis, the 26S rRNA (D1 / D2) region of Y297 yeast was analyzed by base sequence analysis and compared with the standard strain. As a result, the Y297 yeast was identified as Saccharomyces cerevisiae ( Saccharomyces cerevisiae) (Fig. 2).
<실험예 1> 향기성분 생산과 관련된 효소 활성 조사<Experimental Example 1> Enzyme activity related to perfume ingredient production
효모가 생성하는 효소의 활성 검정 및 비교를 위하여, 저온(15℃)에서 효모의 발효 과정 중 향기 관련 물질 생산에 영향을 주는 효소인 β-glucosidase, esterase의 활성을 분석하기 위해 효모의 세포질 단백질을 추출하여 효소의 활성 확인하였다.To investigate the activity of β-glucosidase and esterase, enzymes affecting the production of aroma-related substances during yeast fermentation process at low temperature (15 ℃), yeast cytoplasmic proteins And the activity of the enzyme was confirmed.
구체적으로, 효모의 효소활성을 확인하기 위해 yeast protein extraction buffer kit(GE healthcare, St Louis, USA)를 사용하여 단백질을 추출하였다. β-glucosidase(EC 3.2.1.21.)의 활성은 Greover 등(Biochem Biophys Acta, 1997, 482, 98-108)의 방법을 적용하여 실험하였다. 10 mM의 기질 ρ-nitrophenyl-β-D-glucopyranoside(Sigma-Aldrich Co., St Louise, USA)을 100 ㎕ 넣고, 100 mM citrate phosphate buffer(pH 4.0)을 890 ㎕ 넣은 후, 효모 추출액을 넣고, 45℃에서 반응시킨 후, 0.5 M sodium carbonate(Sigma-Aldrich Co)로 반응을 멈춘 후 원심분리한 상등액을 분광 광도계에서 흡광도 405 nm로 활성을 확인하였다.Specifically, the protein was extracted using a yeast protein extraction buffer kit (GE healthcare, St Louis, USA) to confirm the yeast enzyme activity. The activity of β-glucosidase (EC 3.2.1.21.) was tested by the method of Greover et al. (Biochem Biophys Acta, 1997, 482, 98-108). 100 μl of 10 mM ρ-nitrophenyl-β-D-glucopyranoside (Sigma-Aldrich Co., St Louise, USA) was added and 890 μl of 100 mM citrate phosphate buffer (pH 4.0) was added. After incubation at 45 ° C, the reaction was stopped with 0.5 M sodium carbonate (Sigma-Aldrich Co), and the supernatant was centrifuged and its activity was confirmed to be 405 nm by a spectrophotometer.
또한, esterasse(EC 3.1.1.6)의 활성은 Frank 등(Appl Environ Microbiol, 2006, 72, 7140-7147)의 방법을 변형하여 수행하였고, 효모 추출액(32 ㎕)에 4.5 M의 ρ-nitrophenyl acetate(Sigma-Aldrich)가 포함된 0.1 M tris-HCL buffer(pH 7.0)을 넣고 35℃에서 반응시켜 효소활성을 확인하였다. 또한, 5 M ρ-nitrophenyl-β-butyric acid(Sigma-Aldrich)를 기질로 하여, 0.5 M sodium phosphate buffer(pH 7.0)에서 40℃에서 반응시킨 후, 405 nm에서 활성을 측정하였다. 효모는 YPD(25% glucose) 배지에서 5일간 120 rpm으로 배양하였고, 분당 생성되는 1μmol의 ρ-nitrophenol을 1 unit으로 정의하여 계산하였다.In addition, the activity of esterase (EC 3.1.1.6) was modified by the method of Frank et al. (Appl Environ Microbiol, 2006, 72, 7140-7147) and a 4.5 M ρ-nitrophenyl acetate Sigma-Aldrich) and incubated at 35 ° C to confirm enzyme activity. The activity was measured at 405 nm in 0.5 M sodium phosphate buffer (pH 7.0) at 40 ° C using 5 M ρ-nitrophenyl-β-butyric acid as a substrate. Yeast cells were cultured in YPD (25% glucose) medium at 120 rpm for 5 days, and 1 μmol of ρ-nitrophenol produced per minute was defined as 1 unit.
그 결과, 도 3에 나타난 바와 같이, Y297 효모의 β-glucosidase의 활성은 대조구(상업효모)보다 약 2.5~5.0배 높은 활성을 나타내었고, esterase의 활성은 ρ-nitrophenyl acetate를 기질로 사용하였을 경우엔 대조구(상업효모, Con 2)에 비해 약 2배 높은 활성을 나타내었으며, ρ-nitrophenyl-β-butyric acid를 기질로 사용하였을 때는 대조구(상업효모)보다 약 2~5배 높은 활성을 나타냄을 확인하였다(도 3). As a result, as shown in FIG. 3, the activity of β-glucosidase of Y297 yeast was about 2.5-5.0 times higher than that of control (commercial yeast). The activity of esterase was found to be higher when ρ-nitrophenyl acetate was used as a substrate (Commercial yeast, Con 2). When ρ-nitrophenyl-β-butyric acid was used as a substrate, the activity was about 2-5 times higher than that of the control (commercial yeast) (Fig. 3).
<실험예 2> 저온적응성 Y297 효모로 발효한 발효주의 품질 특성 확인<Experimental Example 2> Determination of quality characteristics of fermented fermented soybeans with low temperature adaptive Y297 yeast
<2-1> Y297 효모를 이용한 발효주의 제조<2-1> Preparation of Fermented Juice Using Y297 Yeast
멥쌀 1 Kg을 흐르는 물에 씻어준 후 15~20℃에서 2~3시간 침지 후 물 빼기를 1시간한 후 고두밥으로 찌어준다. 고두밥의 열을 식힌 후 물 1.2 L에 당화효소 2 g을 넣어주고 2일 동안 배양된 Y297를 접종하여 10~20일간 15~25℃에서 발효하여 약주를 제조한다.Wash 1 Kg of rice with flowing water, immerse at 15 ~ 20 ℃ for 2 ~ 3 hours, drain for 1 hour, and cook with high rice. After cooling the heat of rice cakes, 2 g of saccharogenic enzyme is added to 1.2 L of water, Y297 cultured for 2 days is inoculated, and fermented at 15 to 25 ° C for 10 to 20 days to prepare a Yakju.
<2-2> 저온적응성 Y297 효모로 발효한 발효주의 pH 확인<2-2> Determination of the pH of the fermented fermented by low temperature adaptive Y297 yeast
Y297효모로 발효한 발효주의 pH를 확인하기 위하여, Y297효모를 이용하여 제조된 약주 시료 10 ml를 취하여 pH meter (FE20, Mettler Toledo, Switzerland)로 측정하였다.To determine the pH of the fermented yeast fermented with Y297 yeast, 10 ml of the Yakju sample prepared with Y297 yeast was measured with a pH meter (FE20, Mettler Toledo, Switzerland).
Y297 효모로 발효한 발효주 및 대조구(상업효모)를 이용한 발효주의 pH를 측정한 결과, Y297 효모를 이용한 발효주의 pH가 대조구의 발효주보다 0.3~0.5% 정도 높은 것으로 나타났다(표 3).The pH of the Y297 yeast fermented and fermented by using the control (commercial yeast) was 0.3 ~ 0.5% higher than that of the control (Table 3).
<2-3> 저온적응성 Y297 효모로 발효한 발효주의 총산도 확인<2-3> Determination of total acidity of fermented soybean fermented with low temperature adaptive Y297 yeast
Y297 효모로 발효한 발효주의 총 산도를 확인하기 위하여, Y297효모를 이용하여 제조된 약주의 10 ml에 페놀프탈레인 용액(0.5%)을 1~2방울 첨가한 후 뷰렛에 들어있는 0.1N NaOH 용액으로 적색이 될 때까지 적정한다. 소비된 적정량에 환산계수(초산)으로 계산하여 총산량을 계산한다.To determine the total acidity of the fermented yeast fermented with Y297 yeast, 1 ~ 2 drops of phenolphthalein solution (0.5%) was added to 10 ml of Yakju prepared using Y297 yeast, and then 0.1N NaOH solution in burette To the point where it is necessary. Calculate the total amount by calculating the conversion factor (acetic acid) to the appropriate amount consumed.
Y297 효모로 발효한 발효주의 총산도는 대조구보다 0.05~0.15% 낮은 것으로 나타났으며, 일반적인 발효주의 산도 범위는 0.2~0.5%로 이 범위에 속하면서, 대조구보다 산도가 낮음을 확인하였다(표 4).The total acidity of Y297 yeast fermented soybean was 0.05 ~ 0.15% lower than that of the control. The acidity range of general fermented soybean was 0.2 ~ 0.5%, which was lower than that of the control (Table 4) .
<2-4> 저온적응성 Y297 효모로 발효한 발효주의 알코올 함량 확인<2-4> Determination of Alcohol Content in Fermented Juice Fermented with Low Temperature Adaptive Y297 Yeast
Y297 효모로 발효한 발효주의 알코올 함량을 확인하기 위하여, Y297효모를 이용하여 제조한 약주 100 ml에 증류수 100 ml를 첨가한 후 500 ml 삼각플라스크에 옮긴 후 소포제와 액화효소를 각각 0.5 ml를 넣고 냉각기가 연결한 후 증류한다. 메스실린더에 유액 70 ml를 채운 후 증류수 30 ml를 넣어주고 15℃에서 주정계를 이용하여 측정한다. To determine the alcohol content of Y297 fermented yeast, 100 ml of distilled water was added to 100 ml of Yakju prepared with Y297 yeast, transferred to a 500 ml Erlenmeyer flask, 0.5 ml of defoamer and liquefying enzyme were added, And then distilled. Fill the measuring cylinder with 70 ml of emulsion, add 30 ml of distilled water, and measure at 15 ° C using an umbrella.
Y297 효모로 발효한 발효주의 알코올 함량은 대조구 보다 1~3% 정도 높은 편으로, 특히, 저온발효 후의 알코올 함량에서도 대조구보다 높은 알코올 함량을 보여 저온에서 발효하였을 때, 알코올 발효가 안정적으로 일어남을 확인하였다(표 5).The alcohol content of fermented Y297 fermented yeast was 1 ~ 3% higher than that of the control. Especially, alcohol content after low temperature fermentation showed higher alcohol content than that of the control, indicating that alcohol fermentation stably occurs at low temperature (Table 5).
<2-5> 저온적응성 Y297 효모로 발효한 발효주의 유기산 함량 분석<2-5> Analysis of Organic Acid Content of Fermented Soybean Fermented by Low Temperature Adaptive Y297 Yeast
Y297 효모로 발효한 발효주의 유기산 함량을 측정하기 위하여, Y297효모를 이용하여 제조된 약주 10 ml를 5,000 rpm에서 원심분리한 후, 상등액 2 ml를 0.45 um membrane filter(Millipore co., Bedford, MA, USA)로 여과한 후 Sep-pak C18 cartridge(Waters Oasis, Milford, MA, USA)로 색소를 제거하였다. 시료는 HPLC(Chromaster 5000, Hitachi, Ltd., Tokyo, Japan)로 분석하였으며 ODS-100W(4.6 × 250mm) column을 사용하여 분석하였다. 표준물질은 oxalic acid, citric acid, tartaric acid, malic acid, succinic acid, fumaric acid, lactic acid, formic acid, acetic acid를 이용하여 표준곡선을 만든 후 정량하였다. In order to determine the content of organic acid in the fermented yeast fermented with Y297 yeast, 10 ml of Yakju prepared with Y297 yeast was centrifuged at 5,000 rpm and 2 ml of the supernatant was transferred to 0.45 um membrane filter (Millipore co., Bedford, MA, USA) and then discarded with Sep-pak C18 cartridge (Waters Oasis, Milford, Mass., USA). The samples were analyzed by HPLC (Chromaster 5000, Hitachi, Ltd., Tokyo, Japan) and analyzed using ODS-100W (4.6 × 250 mm) column. Standard curves were prepared using oxalic acid, citric acid, tartaric acid, malic acid, succinic acid, fumaric acid, lactic acid, formic acid and acetic acid.
Y297 효모로 발효한 발효주의 유기산 함량을 분석한 결과, 대조구에 비해 유기산 함량이 높게 검출됨을 확인하였다(표 6). Y297 fermented yeast fermented soybeans showed higher content of organic acids than the control (Table 6).
N.D: 측정 불가.N.D: Measurement not possible.
<2-6> 저온적응성 Y297 효모로 발효한 발효주의 유리아미노산 분석<2-6> Free amino acid analysis of fermented soybean fermented with low temperature adaptive Y297 yeast
Y297 효모로 발효한 발효주의 유리아미노산 함량을 측정하기 위하여, 5,000 rpm에서 원심분리한 후, 상등액 2 ml를 0.45 um membrane filter(Millipore co., Bedford, MA, USA)로 여과한 후 Sep-pak C18 cartridge(Waters Oasis, Milford, MA, USA)로 색소를 제거하였다. 시료 100 ul에 Ninhydrin 반응액 1 ml과 완충액(PF-1) 1 ml을 첨가한 후 3분간 100에서 가열한다. 반응 후 아미노산 분석기 (L-8900 Amino Acid Analyzer, HITACHI, Ltd., Tokyo, Japan)를 이용하여 분석하였다. 시료 20 ul를 HPLC Packed Column Column (4.6 × 60 mm) Ion Exchange column (HITACHI, Ltd., Tokyo, Japan)과 UV-Detector(570 nm, 440 nm)를 사용하여 분석하였다.To determine the free amino acid content of Y297 fermented yeast, the supernatant was filtered through a 0.45 um membrane filter (Millipore co., Bedford, MA, USA) and centrifuged at 5,000 rpm. Sep-pak C18 cartridge (Waters Oasis, Milford, MA, USA). Add 1 ml of Ninhydrin Reaction Solution and 1 ml of Buffer (PF-1) to 100 μl of the sample, and heat at 100 ° C for 3 minutes. After the reaction, it was analyzed using an amino acid analyzer (L-8900 Amino Acid Analyzer, HITACHI, Ltd., Tokyo, Japan). 20 μl of the sample was analyzed using a HPLC packed column (4.6 × 60 mm) Ion exchange column (HITACHI, Ltd., Tokyo, Japan) and a UV-detector (570 nm, 440 nm).
Y297 효모로 발효한 발효주의 유리아미노산을 분석한 결과, 표 7에 나타난 바와 같이, 필수 아미노산 및 비 필수아미노산의 함량이 대조구보다 높은 수치를 나타내었고, 이는 술의 기호에 영향을 줄 것으로 예상된다. 또한, 감칠맛과 단맛을 내는 아미노산인 aspartic acid, glutamic acid, alanine의 함량이 높아, 발효주의 단맛과 감칠맛이 높을 것으로 기대된다. As shown in Table 7, the content of essential amino acid and non-essential amino acid was higher than that of the control, and it is expected that this will affect the taste of the wine. In addition, it is expected that the content of aspartic acid, glutamic acid and alanine, which are rich in amino acids such as richness and sweetness, is high and sweetness and richness of fermented beverage are high.
<2-7> 저온적응성 Y297 효모로 발효한 발효주의 향기패턴 분석<2-7> Analysis of aroma pattern of fermented fermented soybeans with low temperature adaptive Y297 yeast
발효온도(15, 20, 25℃)를 달리하여 저온적응성 Y297 효모로 제조한 발효주의 향기 패턴을 전자코를 이용하여 분석한 결과, 15, 20℃의 저온에서 발효한 경우 대조구와 다른 향기 패턴을 나타내었다(도 4).The fermentation patterns of fermented soybeans prepared with low temperature adaptive Y297 yeast were different according to fermentation temperature (15, 20, 25 ℃). When fermented at low temperature of 15 and 20 ℃, (Fig. 4).
또한, 저온적응성 Y297 효모로 제조한 발효주의 향기 성분을 한국기초과학지원연구원에 의뢰하여 GC-TOF/MS로 분석하였고, 샘플의 정량 분석을 위해 octanoic acid를 내부 표준물질로 사용하였다. In addition, fragrance components of the fermented juice made from low-temperature adaptive Y297 yeast were analyzed by GC-TOF / MS, and octanoic acid was used as an internal standard for quantitative analysis of the samples.
그 결과, Y297 효모로 제조한 발효주는 대조구로 발효한 발효주와 다른 향기 성분을 나타냄을 확인하였다(도 5, 표 8).As a result, it was confirmed that the fermented yeast prepared from Y297 yeast exhibited different fragrance components from the fermented fermented fermented soybean (FIG. 5, Table 8).
<실험예 3> 저온적응성 Y297 효모로 발효한 발효주의 관능 특성<Experimental Example 3> Sensory properties of fermented soybean fermented with low temperature adaptive Y297 yeast
멥쌀 1 kg을 고두밥으로 찐 후 물 1.2 L에 당화수소 2 g을 넣어주고 2일 동안 배양된 선발효모 Y297을 접종하여 20일간 발효하여 발효주를 제조한 후, 국립농업과학원 농식품자원부 종합실험실에서 13명 인원의 관능검사를 통해 저온적응성 Y297효모의 기호도를 조사하였다. 그 결과, 통계적으로 유의한 결과를 얻지는 못하였으나, 수치상의 결과로 Y297 효모를 이용한 발효주가 기호도와 맛, 그리고 색에서 가장 좋은 평가를 받음을 확인하였다(표 9).1 kg of rice was steamed, then 2 g of hydrogen peroxide was added to 1.2 L of water, followed by inoculation with a selective yeast Y297 cultured for 2 days. The fermented soybean was fermented for 20 days, and then 13 strains were obtained from the National Institute of Agricultural Science and Technology The preference of low temperature adaptive Y297 yeast was investigated by sensory evaluation. As a result, although statistically significant results were not obtained, it was confirmed that Y297 yeast was best evaluated in taste, taste and color (Table 9).
<110> Rural Development Administration <120> Saccharomyces cerevisiae Y297 with adaptability at low temperature and manufacturing method of fermented alchoholic beverage using the same <160> 3 <170> KoPatentIn 3.0 <210> 1 <211> 1253 <212> DNA <213> Saccharomyces cerevisiae <400> 1 atcatcgaat ctttgaacgc acattgcgcc ccttggtatt ccagggggca tgcctgtttg 60 agcgtcattt ccttctcaaa cattctgttt ggtagtgagt gatactcttt ggagttaact 120 tgaaattgct ggccttttca ttggatgttt ttttttccaa agagaggttt ctctgcgtgc 180 ttgaggtata atgcaagtac ggtcgtttta ggttttacca actgcggcta atctttttta 240 tactgagcgt attggaacgt tatcgataag aagagagcgt ctaggcgaac aatgttctta 300 aagtttgacc tcaaatcagg taggagtacc cgctgaactt aagcatatca ataagcggag 360 gaaaagaaac caaccgggat tgccttagta acggcgagtg aagcggcaaa agctcaaatt 420 tgaaatctgg taccttcggt gcccgagttg taatttggag agggcaactt tggggccgtt 480 ccttgtctat gttccttgga acaggacgtc atagagggtg agaatcccgt gtggcgagga 540 gtgcggttct ttgtaaagtg ccttcgaaga gtcgagttgt ttgggaatgc agctctaagt 600 gggtggtaaa ttccatctaa agctaaatat tggcgagaga ccgatagcga acaagtacag 660 tgatggaaag atgaaaagaa ctttgaaaag agagtgaaaa agtacgtgaa attgttgaaa 720 gggaagggca tttgatcaga catggtgttt tgtgccctct gctccttgtg ggtaggggaa 780 tctcgcattt cactgggcca gcatcagttt tggtggcagg ataaatccat aggaatgtag 840 cttgcctcgg taagtattat agcctgtggg aatactgcca gctgggactg aggactgcga 900 cgtaagtcaa ggatgctggc ataatggtta tatgccgccc gtcttgaaac acggaccaag 960 gagtctaacg tctatgcgag tgtttgggtg taaaacccat acgcgtaatg aaagtgaacg 1020 taggttgggg cctcgcaaga ggtgcacaat cgaccgatcc tgatgtcttc ggatggattt 1080 gagtaagagc atagctgttg ggacccgaaa gatggtgaac tatgcctgaa tagggtgaag 1140 ccagaggaaa ctctggtgga ggctcgtagc ggttctgacg tgcaaatcga tcgtcgaatt 1200 tgggtatagg ggcgaaagac taatcgaacc atctagtagc tggttcctgc cga 1253 <210> 2 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> artificial sequence <400> 2 gcatatcaat aagcggagga aaag 24 <210> 3 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> artificial sequence <400> 3 ggtccgtgtt tcaagacgg 19 <110> Rural Development Administration <120> Saccharomyces cerevisiae Y297 with adaptability at low temperature and manufacturing method of fermented alchoholic beverage using the same <160> 3 <170> KoPatentin 3.0 <210> 1 <211> 1253 <212> DNA <213> Saccharomyces cerevisiae <400> 1 atcatcgaat ctttgaacgc acattgcgcc ccttggtatt ccagggggca tgcctgtttg 60 agcgtcattt ccttctcaaa cattctgttt ggtagtgagt gatactcttt ggagttaact 120 tgaaattgct ggccttttca ttggatgttt ttttttccaa agagaggttt ctctgcgtgc 180 ttgaggtata atgcaagtac ggtcgtttta ggttttacca actgcggcta atctttttta 240 tactgagcgt attggaacgt tatcgataag aagagagcgt ctaggcgaac aatgttctta 300 aagtttgacc tcaaatcagg taggagtacc cgctgaactt aagcatatca ataagcggag 360 gaaaagaaac caaccgggat tgccttagta acggcgagtg aagcggcaaa agctcaaatt 420 tgaaatctgg taccttcggt gcccgagttg taatttggag agggcaactt tggggccgtt 480 ccttgtctat gttccttgga acaggacgtc atagagggtg agaatcccgt gtggcgagga 540 gtgcggttct ttgtaaagtg ccttcgaaga gtcgagttgt ttgggaatgc agctctaagt 600 gggtggtaaa ttccatctaa agctaaatat tggcgagaga ccgatagcga acaagtacag 660 tgatggaaag atgaaaagaa ctttgaaaag agagtgaaaa agtacgtgaa attgttgaaa 720 gggaagggca tttgatcaga catggtgttt tgtgccctct gctccttgtg ggtaggggaa 780 tctcgcattt cactgggcca gcatcagttt tggtggcagg ataaatccat aggaatgtag 840 cttgcctcgg taagtattat agcctgtggg aatactgcca gctgggactg aggactgcga 900 cgtaagtcaa ggatgctggc ataatggtta tatgccgccc gtcttgaaac acggaccaag 960 gagtctaacg tctatgcgag tgtttgggtg taaaacccat acgcgtaatg aaagtgaacg 1020 taggttgggg cctcgcaaga ggtgcacaat cgaccgatcc tgatgtcttc ggatggattt 1080 gagtaagagc atagctgttg ggacccgaaa gatggtgaac tatgcctgaa tagggtgaag 1140 ccagaggaaa ctctggtgga ggctcgtagc ggttctgacg tgcaaatcga tcgtcgaatt 1200 tgggtatagg ggcgaaagac taatcgaacc atctagtagc tggttcctgc cga 1253 <210> 2 <211> 24 <212> DNA <213> Artificial Sequence <220> Artificial sequence <400> 2 gcatatcaat aagcggagga aaag 24 <210> 3 <211> 19 <212> DNA <213> Artificial Sequence <220> Artificial sequence <400> 3 ggtccgtgtt tcaagacgg 19
Claims (7)
A novel strain of Sacharomyces cerevisiae Y297 having the nucleotide sequence of 26s rDNA of SEQ ID NO: 1 deposited with KACC93244P.
The novel strain of Saccharomyces cerevisiae Y297 according to claim 1, wherein the strain is resistant to low-temperature growth at 10 to 20 占 폚.
A prophylactic composition comprising the strain of claim 1 or a culture thereof.
2) 상기 단계 1)의 곡류를 발효시키는 단계를 포함하는 발효주 제조 방법.
1) Inoculating the cereal of claim 1 into cereals; And
2) fermenting the cereal of step 1) above.
6. The method of claim 5, wherein the cereal in step (1) is rice, barley, wheat, or sweet potato.
A fermented beverage produced by the method of claim 5.
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