KR101480308B1 - Brewing yeast saccharomyces cerevisiae 192-4 and brewed alcohol made therewith - Google Patents
Brewing yeast saccharomyces cerevisiae 192-4 and brewed alcohol made therewith Download PDFInfo
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Abstract
본 발명은, 사카로마이세스 세레비지애(Saccharomyces cerevisiae) 192-4(KCCM11397P), 사카로마이세스 세레비지애 192-4(KCCM11397P)를 이용하여 제조된 발효주, 및 사카로마이세스 세레비지애 192-4(KCCM11397P)를 이용하여 발효시키는 것을 특징으로 하는 발효주의 제조 방법에 관한 것이다.The present invention, in three Levy saccharide My process jiae (Saccharomyces (KCCM11397P), Saccharomyces cerevisiae 192-4 (KCCM11397P), Saccharomyces cerevisiae 192-4 (KCCM11397P), and Saccharomyces cerevisiae 192-4 To a method for producing a fermented beverage.
Description
본 발명은 발효주의 제조에 이용되는 신규한 사카로마이세스 세레비지애(Saccharomyces cerevisiae) 균주, 이를 이용하여 제조된 발효주, 및 이를 이용하여 발효주를 제조하는 방법에 관한 것이다.The present invention relates to novel strains of Saccharomyces cerevisiae cerevisiae strain, a fermentation broth manufactured using the same, and a method for producing a fermented broth using the same.
술은 자연적으로 발생되어 지역, 민족, 기후, 풍토 및 문화적 차이에 따라 여러 형태의 개성 있는 형태로 발전되어 왔다. 세균, 효모 등의 존재가 발견되기 이전에는 현대적인 발효 기술이 없었으므로, 누룩 등에 존재하는 자생 효모에 의하여 발효주를 제조하였다. 그러나 자생 효모들을 이용하면 우수하고 일정한 발효능을 유지하기가 어려운 문제점이 있었다. 이 문제점을 해결하기 위해, 근대에 들어서는 자생 효모들 중에서 우수한 특성을 갖는 효모를 선별, 분리하여 발효를 수행함으로써, 우수하고 일정한 발효능을 유지하여 우수한 발효주를 생산하기 시작했다.Drinking has been naturally occurring and has evolved into a variety of unique forms depending on region, ethnicity, climate, climate and cultural differences. Since there was no modern fermentation technology before the existence of bacteria, yeast, etc., The fermented bean was prepared by the native yeast present in the yeast. However, there is a problem in that it is difficult to maintain good and constant efficacy by using native yeasts. In order to solve this problem, yeast having excellent characteristics among the native yeasts in modern times was selected and separated to carry out fermentation, and excellent and constant efficacy was maintained to produce excellent fermented beverage.
현재 맥주, 청주 또는 포도주 등 흔히 접할 수 있는 대표적인 발효주들은 모두 엄선된 발효 효모에 의해 발효된 것이며, 새로운 발효 효모를 탐색하는 작업이 계속적으로 이루어지고 있다. 이러한 탐색을 위해서는 실험실에서 선별된 균주들을 일일이 발효시킨 후 각각의 발효능 등의 특성을 측정해야 하므로, 많은 시간과 비용을 필요로 한다. 현재 발효주에 관하여 여러 연구가 진행되고 있으며, 한국등록특허 제10-0526260호, 한국등록특허 제10-0424043호, 한국공개특허 제10-2012-0056423호 등에 관련 내용이 개시되어 있다.Current representative fermented beverages, such as beer, sake, or wine, are all fermented by selected fermenting yeast, and the search for new fermenting yeast continues. In order to perform such a search, it is necessary to ferment the strains selected in the laboratory and to measure the characteristics of each efficacy, so that it takes a lot of time and cost. Various studies have been conducted on the present fermentation stock, and related contents are disclosed in Korean Patent No. 10-0526260, Korean Patent No. 10-0424043, Korean Patent Publication No. 10-2012-0056423, and the like.
한편, 멜리비오스(melibiose)는 식물계에 널리 분포하는 6-O-α-D-갈락토피라노실-D-글루코오스 구조의 환원성 이당류인데, 사카로마이세스 세레비지애는 멜리비오스를 이용하지 못하는 것으로 알려져 있다.On the other hand, melibiose is a reducing disaccharide of 6- O- alpha -D-galactopyranosyl-D-glucose structure widely distributed in the vegetable field. Saccharomyces cerevisiae does not utilize melibiose It is known.
본 발명이 이루고자 하는 기술적 과제는 기존에 사카로마이세스 세레비지애 균주가 이용하지 못하는 것으로 알려진 멜리비오스를 이용할 수 있는 내알코올성이 우수한 신규 효모 균주를 발견하고, 이러한 신규 효모 균주를 이용하여 우수한 향미를 나타내고, 관능 특성이 뛰어난, 고 알코올 생성 발효주를 제공하는 것을 목적으로 한다.Disclosure of Invention Technical Problem [8] The present invention provides a new yeast strain having excellent alcohol resistance, which can utilize melibiose, which is known to be unusable by Saccharomyces cerevisiae strains, And a high alcohol-producing fermented product having excellent sensory characteristics.
이에 본 발명자들은 상기와 같은 목적을 달성하기 위하여 예의 연구를 거듭한 결과, MEL-gen을 갖고 있고, 그 결과 균체 외 효소 α-갈락토시다아제(멜리비아제)를 생성함으로써 멜리비오스를 이용할 수 있는 우수한 발효용 효모 균주를 발견하여 본 발명을 완성하기에 이르렀다.Accordingly, the present inventors have conducted intensive studies to achieve the above object, and as a result, they have found that MEL-gene can be used as a result of producing an extracellular enzyme α-galactosidase (melibiase) And found the yeast strains for fermentation, thus leading to the completion of the present invention.
본 발명에 따른 사카로마이세스 세레비지애 192-4는 사카로마이세스 세레비지애 균주가 이용하지 못하는 것으로 알려진 D-멜리비오스를 이용하여 발효할 수 있고, D-멜리비오스를 동화하여 균체 증식을 할 수 있으며, 또한 에탄올 내성이 높다.The saccharomyces cerevisiae 192-4 according to the present invention can be fermented using D-melibiose, which is known to be unable to utilize Saccharomyces cerevisiae strain, and is capable of fermenting D- And also has high ethanol tolerance.
도 1은 본 발명의 사카로마이세스 세레비지애 192-4와 동일한 종의 공지 균주간의 18S rRNA 염기 서열을 비교한 것이다. 도면 중 CBS4054는 사카로마이세스 세레비지애 부분 18S rRNA 유전자, ITS1, 5.8S rRNA 유전자, ITS2 및 부분 26S rRNA 유전자, 기준주 CBS4054를 나타낸다.
도 2는 본 발명의 사카로마이세스 세레비지애 192-4의 에탄올 내성을 측정한 그래프이다.1 compares 18S rRNA nucleotide sequences between known strains of the same species as Saccharomyces cerevisiae 192-4 of the present invention. In the figure, CBS4054 represents Saccharomyces cerevisiae partial 18S rRNA gene, ITS1, 5.8S rRNA gene, ITS2 and partial 26S rRNA gene, reference strain CBS4054.
2 is a graph showing the ethanol resistance of Saccharomyces cerevisiae 192-4 of the present invention.
이하, 본 발명을 상세히 설명하기로 한다. 다만, 본 발명은 다양한 형태로 변경되어 구현될 수 있으며, 여기에서 설명하는 구현예에 한정되는 것은 아니다.Hereinafter, the present invention will be described in detail. However, it should be understood that the present invention may be embodied in many other specific forms without departing from the spirit or essential characteristics thereof.
본 발명은 새롭게 분리 동정된 사카로마이세스 세레비지애 192-4를 제공한다.The present invention provides a newly isolated Saccharomyces cerevisiae 192-4.
본 발명자들은 다양한 누룩으로부터 수많은 효모 균주를 분리한 후 당 이용능, 알코올 내성, 관능 검사 결과 등을 종합적으로 평가하여 최적의 효모를 선별하였으며, 선별된 효모를 동정하여 사카로마이세스 세레비지애임을 확인하였다. 이를 192-4 균주라고 명명하고, 2013년 3월 5일자로 한국미생물보존센터에 기탁번호 KCCM11397P로 기탁하였다.After isolating a large number of yeast strains from a variety of yeast, the present inventors evaluated the yeast activity, alcohol tolerance, sensory evaluation results, and the like to select an optimal yeast. The yeast was selected and identified, and the yeast was identified as Saccharomyces cerevisiae Respectively. This strain was named as the strain 192-4 and deposited on May 5, 2013 with the deposit number KCCM11397P at the Korean Society for Microbiological Research.
본 발명에 따른 발효 균주 192-4는 전통 누룩에서 분리된 야생 양조 효모로서 발효능이 우수하며, 이화학적, 유전학적 특성에 대한 분석을 통해 동정한 결과 포도주 효모, 맥주 효모 및 일본의 청주 효모와는 다른 신 균주임을 확인하였다. 또한, 본 발명의 균주 192-4는 사카로마이세스 세레비지애 균주가 이용하지 못하는 것으로 알려진 D-멜리비오스를 이용하여 발효할 수 있고, D-멜리비오스를 동화하여 균체 증식을 할 수 있으며, 또한 에탄올 내성이 높음을 확인하였다.The fermentation strain 192-4 according to the present invention is a wild brewer's yeast isolated from a traditional yeast, and has excellent pharmacological activity. As a result of the analysis on the physicochemical and genetic characteristics, the yeast, brewer's yeast, Were found to be other new strains. In addition, the strain 192-4 of the present invention can be fermented using D-melibiose, which is known to be unavailable to Saccharomyces cerevisiae, and can be used to assimilate D-melibiose to grow cells, Also, it was confirmed that ethanol tolerance was high.
본 발명에 따른 효모 균주는 통상적인 사카로마이세스 속 효모의 배양법에 의해 대량으로 배양할 수 있다. 배양 배지로는 탄소원, 질소원, 비타민 및 미네랄로 구성된 배지를 사용할 수 있다. 예를 들어, YM 아가(yeast malt extract agar) 배지, PDA(potato dextrose agar) 배지에서 25℃ 내지 30℃, pH 5.6±0.2, 36 시간 내지 72 시간, 호기적인 조건 하에서 배양하여 4℃에서 보관하면서 계대하여 사용할 수 있다.The yeast strain according to the present invention can be cultured in a large amount by a conventional culture method of Saccharomyces sp. As the culture medium, a medium composed of carbon source, nitrogen source, vitamins and minerals can be used. For example, the cells are cultured in yeast malt extract agar medium, PDA (potato dextrose agar) medium at 25 ° C to 30 ° C, pH 5.6 ± 0.2, 36 hours to 72 hours under aerobic conditions, Can be used.
본 발명은 사카로마이세스 세레비지애 192-4(KCCM11397P)를 이용하여 제조된 발효주에 관한 것이고, 또한 본 발명은 사카로마이세스 세레비지애 192-4(KCCM11397P)를 이용하여 발효시키는 것을 특징으로 하는 발효주의 제조 방법에 관한 것이다.The present invention relates to a fermented milk produced using Saccharomyces cerevisiae 192-4 (KCCM11397P), and the present invention is characterized in that fermentation is carried out using Saccharomyces cerevisiae 192-4 (KCCM11397P) To a method for producing a fermented beverage.
즉, 본 발명은 192-4 균주를 이용하여 제조된 약주, 탁주, 막걸리, 청주, 포도주, 과실주 등의 발효주를 제공하며, 또한 상기 192-4 균주를 이용하여 발효주를 제조하는 방법을 제공한다.That is, the present invention provides a fermentation stock such as a fermented soybean meal, a mangolli, sake wine, wine, fruit wine, etc. manufactured using the strain 192-4, and also provides a method for producing a fermented soybean using the strain 192-4.
일 구현예에서, 본 발명에 따른 발효주는 (a) 전분질 원료(예를 들어, 찹쌀), 누룩 및 본 발명의 균주에 물을 첨가하여 밑술을 제조하는 단계; (b) 상기 (a) 단계에서 수득한 밑술에 증자한 전분질 원료, 누룩 및 물을 추가로 첨가하여 발효시키는 담금 단계; 및 (c) 상기 (b) 단계에서 수득한 발효액을 여과하는 단계를 포함하는 방법에 의해 제조될 수 있다.In one embodiment, the fermentation liquor according to the present invention comprises the steps of: (a) adding water to a starch raw material (for example, glutinous rice), yeast, and the strain of the present invention to prepare a slurry; (b) a fermentation step in which fermented starch material, koji and water are added to the ginseng obtained in step (a) and fermented; And (c) filtering the fermentation liquid obtained in the step (b).
상기 (a) 또는 (b) 단계 이후에 동충하초, 적하수오, 발효 홍삼, 대추, 감초 등의 식물 약재, 매실, 포도 등의 과실 등 여러 가지 성분이 첨가될 수도 있다.After step (a) or (b), various components such as a plant medicine such as cordyceps, red pepper, fermented red ginseng, jujube, licorice, fruit of plum, grape may be added.
일 구현예에서, 상기 밑술을 제조하는 단계 (a)는 5℃ 내지 30℃에서 2 일 내지 20 일간 발효시켜 수행될 수 있으며, 상기 담금 단계 (b)는 5℃ 내지 30℃에서 2 일 내지 30 일간 수행될 수 있다.In one embodiment, step (a) of making the veneer may be performed by fermentation at 5 ° C to 30 ° C for 2 days to 20 days, and the dipping step (b) Day.
하기의 실시예를 통하여 본 발명을 보다 상세하게 설명한다. 그러나 하기 실시예는 본 발명의 내용을 구체화하기 위한 것일 뿐 실시예에 의해 본 발명이 한정되는 것은 아니다.The present invention will be described in more detail with reference to the following examples. However, the following examples are for the purpose of illustrating the present invention and are not intended to limit the scope of the present invention.
[실시예][Example]
효모의 선별Selection of Yeast
균주 1차 선별Selection of strain first
전국에서 수집한 누룩 300 점으로부터 분리한 효모 1,000 여 균주를 대상으로 하였다. 알코올 생산 능력은 입국 당화액 10 ㎖를 듀람관을 포함한 시험관에 넣고 25℃에서 3일 동안 발효 정도를 관찰하였다. 입국 당화액은 입국과 물을 1 : 3 (w/v)으로 혼합하여 62℃에서 5 시간 동안 교반하여 제조하고, 6,000 rpm에서 20 분간 원심분리한 후 14˚brix로 희석하여 사용하였다. 효모는 PDA 배지에서 29℃의 온도에서 24 시간 동안 2차 계대 배양한 후, 멸균한 0.85 % NaCl에 현탁하여 2×107/㎖ 농도로 접종하여 사용하였다. 이렇게 접종한 균체의 CO2 가스 생성 속도를 비교 및 측정하였으며, 가스가 생성된 것으로 인정된 것을 알코올 발효성이 있는 것으로 판정하였다. 향 생성 능력은 7 일 동안 발효한 후 관능적으로 비교하였다. A total of 1,000 strains of yeast isolates from 300 Korean leeks were collected. The alcohol production capacity was determined by inserting 10 ㎖ of immigrated saccharide into a test tube containing a duroman and observing the fermentation degree at 25 캜 for 3 days. The immigrated glycation solution was prepared by mixing 1: 3 (w / v) of water and immersion for 5 hours at 62 ° C, centrifuging at 6,000 rpm for 20 minutes and diluting to 14 ° brix. Yeast cells were cultured in a PDA medium at 29 ° C for 24 hours and then suspended in sterile 0.85% NaCl at a concentration of 2 × 10 7 / ml. The CO 2 gas production rate of the inoculated cells was compared and measured, and it was judged that the gas production was recognized as alcohol fermentation. The fragrance production ability was sensed after fermentation for 7 days.
균주 2차 선별Secondary selection of strains
1차 선별된 206 균주를 대상으로 하였다. 300 ㎖ 용량으로 입국과 물을 1 : 3 (w/v)으로 혼합하여 25℃에서 7 일 동안 발효시켜 알코올 함량, pH, 산도, 고형분 함량 및 향 생성 능력을 비교하였다. The first 206 strains were selected. The ethanol content, pH, acidity, solids content and flavor - producing ability were compared by mixing the water and the water at a ratio of 1: 3 (w / v) at 300 ㎖.
균주 3차 선별Selection of strain third
2차 선별된 25 균주를 대상으로 1,500 ㎖ 용량으로 발효주를 제조하였다. 발효제로는 sp90의 입국을 사용하였다. 멥쌀, 발효제, 효모(0.02%) 및 물을 넣고 25℃에서 2 일 동안 발효하여 밑술을 제조하고, 멥쌀과 물을 추가로 첨가하여 덧술을 제조한 후, 5 일 동안 발효하였으며 급수율은 200 %로 하였다. 제조한 발효주의 알코올 함량, 관능 특성, 향기 성분 등을 분석하였다. 이러한 방법으로 제조한 발효주의 알코올 함량 및 이화학 분석, 유기산 함량, 관능 특성과 향기 성분 분석, 및 미생물 동정 결과를 바탕으로 알코올을 18 %(v/v) 이상 생성하는 고 알코올 생성 균주 사카로마이세스 세레비지애 192-4(기탁번호: KCCM11397P)를 선발하였다.Twenty - five strains were selected and the fermentation broth was prepared in a volume of 1,500 ㎖. The entry of sp90 was used as the fermenting agent. The extracts were prepared by adding rice, fermentation broth, yeast (0.02%) and water, fermented at 25 ℃ for 2 days, added with rice and water, and fermented for 5 days. The yield was 200% Respectively. The alcohol content, sensory characteristics, and aroma components of the fermented soybeans were analyzed. Based on the alcohol content, physicochemical analysis, organic acid content, sensory characteristics, aroma component analysis, and microbial identification result of the fermented wine produced by this method, a high alcohol producing strain Saccharomyces producing alcohol with 18% (v / v) Serrabiasi 192-4 (Accession No .: KCCM11397P) was selected.
효모 균주의 동정Identification of yeast strains
미생물 동정은 다음과 같이 수행하였다. DNA 추출(99℃, 10 분) 후 ITS1(5'-TCCGTAGGTGAACCTGCGG-3'), ITS4(5'-TCCTCCGCTTATTGATATGC-3') 프라이머를 사용하여 PCR을 수행하였다. 증폭된 DNA를 정제 후, ABI PRISM® BigDyeTM Terminator Cycle Sequencing Kit를 사용하고, PCR과 동일한 프라이머를 이용하여 18S rRNA 염기서열을 시퀀싱(ABI PRISM® 3730XL DNA Analyzer, Applied Biosystems, Foster City, CA, USA)하였다. 분석된 염기 서열을 BLAST(http://www.ncbi.nlm.nih.gov) 프로그램을 이용하여 동정하였다.Identification of microorganisms was carried out as follows. PCR was performed using ITS1 (5'-TCCGTAGGTGAACCTGCGG-3 ') and ITS4 (5'-TCCTCCGCTTATTGATATGC-3') primers after DNA extraction (99 ° C, 10 min). After the amplified DNA was purified, the 18S rRNA sequence was sequenced using the ABI PRISM ® BigDye ™ Terminator Cycle Sequencing Kit and the same primers as the PCR (ABI PRISM ® 3730XL DNA Analyzer, Applied Biosystems, Foster City, CA, USA ). The analyzed nucleotide sequence was identified using BLAST ( http://www.ncbi.nlm.nih.gov ) program.
분석 결과, 본 발명에 따른 효모 균주 192-4는 사카로마이세스 세레비지애(Gen bank accession number AB533543)와 18S rRNA 서열이 99 %의 상동성을 나타내어, 사카로마이세스 세레비지애 균주임을 확인하였다.As a result of the analysis, the yeast strain 192-4 according to the present invention showed 99% homology with the 18S rRNA sequence with the genbank accession number AB533543, confirming that it was Saccharomyces cerevisiae strain Respectively.
또한, NCBI 등록된 기준주(Type strain) 사카로마이세스 세레비지애와 균주 192-4의 염기 서열을 비교하여, 본 발명에 따른 균주가 새롭게 동정된 균주임을 확인하였다. 본 발명의 사카로마이세스 세레비지애 192-4와 동일한 종의 공지 균주간의 18S rRNA 염기 서열을 비교하여 도 1에 나타내었다. 도면 중 CBS4054는 사카로마이세스 세레비지애 부분 18S rRNA 유전자, ITS1, 5.8S rRNA 유전자, ITS2 및 부분 26S rRNA 유전자, 기준주 CBS4054를 나타낸다.In addition, the nucleotide sequences of Saccharomyces cerevisiae and strain 192-4 of the NCBI registered type strain were compared to confirm that the strain according to the present invention was a newly identified strain. The 18S rRNA nucleotide sequences of known strains of the same species as Saccharomyces cerevisiae 192-4 of the present invention were compared and shown in Fig. In the figure, CBS4054 represents Saccharomyces cerevisiae partial 18S rRNA gene, ITS1, 5.8S rRNA gene, ITS2 and partial 26S rRNA gene, reference strain CBS4054.
또한, 192-4 균주의 18S rRNA 서열을 서열 번호 1에 나타내었다. In addition, the 18S rRNA sequence of the strain 192-4 is shown in SEQ ID NO: 1.
발효주의 제조Manufacture of fermented beverages
선별된 야생 효모 사카로마이세스 세레비지애 192-4를 이용하여 발효주를 제조하였다. 전체 술 양의 36 중량%로 밑술(효모는 전체 술 양의 0.02 중량%; 발효제 입국 : 물 = 38 : 62)을 제조하여 25℃에서 2 일간 배양하였다. 제조된 밑술에 원료 곡물인 쌀을 증자(멥쌀 : 물 = 32 : 68)하여 전체 술 양의 64 중량%로 덧술을 만들어 25℃에서 5 일간 발효하여 발효주를 제조하였다. The fermented juice was prepared using the selected wild yeast Saccharomyces cerevisiae 192-4. (Yeast is 0.02 wt% of the total sake; Accession: Water = 38: 62), which is 36 wt% of the total sake amount, and cultured at 25 ° C for 2 days. The fermented soybean was fermented at 25 ℃ for 5 days to make 64% by weight of whole rice wine.
하기 표 1에 나타낸 바와 같이, 동일한 기원으로부터 분리한 다른 효모 Yaa-1 및 Yaa-2를 이용하여 발효주를 제조하였을 때와 비교할 때, 본 발명의 효모 균주 192-4를 이용한 발효주의 알코올 함량은 18.1%로서 상대적으로 고알코올을 생성하였고, 고형분 함량은 9.5 % 이었음을 확인할 수 있었다.As shown in the following Table 1, the alcohol content of the fermentation broth using the yeast strain 192-4 of the present invention was 18.1% as compared with when the fermentation broth was prepared using other yeasts Yaa-1 and Yaa-2 isolated from the same origin %, And the solid content was 9.5%.
당 Party 이용능Usability 확인 Confirm
분석 방법Analysis method
효모 균주의 탄소 이용능에 관한 측정은 서로 다른 종류의 95 개 탄소원으로 코팅된 96-웰 마이크로플레이트(BIOLOG, CA, USA)를 이용하여 MicroLogTM System(Release 4.3, BIOLOG, CA, USA)으로 분석하였다. 효모 균체는 BUY (BIOLOG, CA, USA) 배지에서 48 시간 동안 배양하여 멸균 증류수에 현탁하여 탁도계(BIOLOG)로 47%T로 투과율을 맞추어 YT 마이크로플레이트에 접종한 후(웰 당 100 ㎕), 26℃에서 48 시간, 72 시간 배양하여 Microstation reader(BIOLOG)를 이용하여 590 nm에서의 광학 밀도(optical density)를 확인하였다. 산화-발효 시험은 효모 균체의 미토콘드리아 내의 호흡 관여 탈수소효소에 의해 환원되어 보라색을 띄는 테트라졸륨을 사용하여 흡광도를 측정하였다. 동화(assimilation) 시험은 탄소원을 이용한 효모 균체의 증식에 의한 탁도(turbidity)를 측정하였다.Measurements of the carbon availability of the yeast strains were analyzed with a MicroLog ™ System (Release 4.3, BIOLOG, CA, USA) using 96-well microplates coated with 95 different carbon sources (BIOLOG, CA, USA) . Yeast cells were cultured in BUY (BIOLOG, CA, USA) medium for 48 hours, suspended in sterilized distilled water, inoculated into YT microplate (100 μL per well) with a turbidity meter (BIOLOG) And the optical density at 590 nm was confirmed using a Microstation reader (BIOLOG). The oxidation - fermentation test was carried out by measuring the absorbance of purple tetrazolium which was reduced by respiration - dehydrogenase in the mitochondria of yeast cells. The assimilation test measured the turbidity due to the growth of yeast cells using a carbon source.
산화-발효 시험Oxidation-fermentation test
하기 표 2에 나타난 바와 같이, 본 발명의 효모 균주 192-4는 사카로마이세스 세레비지애가 발효하지 못하는 것으로 알려진 D-멜리비오스를 이용하여 발효함을 확인하였다.As shown in the following Table 2, yeast strain 192-4 of the present invention was fermented using D-melibiose known to be unable to ferment saccharomyces cerevisiae.
* 는 약한 양성(weak positive) 반응을 나타낸다.
- 는 음성(negative) 반응을 나타낸다.+ Indicates a positive reaction.
* Indicates a weak positive response.
- indicates a negative reaction.
동화 시험Assimilation test
하기 표 3에 나타난 바와 같이, 본 발명의 효모 균주 192-4는 사카로마이세스 세레비지애가 동화하지 못하는 것으로 알려진 D-멜리비오스를 동화하여 균체 증식을 하였음을 확인하였다.As shown in the following Table 3, it was confirmed that the yeast strain 192-4 of the present invention assimilates D-melibiose known to be unable to assimilate saccharomyces cerevisiae to perform cell proliferation.
* 는 약한 양성(weak positive) 반응을 나타낸다.
- 는 음성(negative) 반응을 나타낸다.+ Indicates a positive reaction.
* Indicates a weak positive response.
- indicates a negative reaction.
이상의 결과를 통해, 효모 192-4가 식물계에 널리 존재하는 멜리비오스를 이용하여 산업적으로 이용 가능한 바이오에탄올을 생성할 수 있음을 확인하였다.From the above results, it was confirmed that yeast 192-4 can produce industrially usable bioethanol using melibiose, which exists widely in plants.
에탄올 내성Ethanol tolerance
에탄올 내성 측정은 YPD(2 % 글루코오스, 0.5 % 효모 추출액, 1 % 박토-펩톤; Sigma-Aldrich, St. Louis, MO, USA) 액체 배지에 효모 균체 접종 직후, 무수 에탄올을 0, 8, 12, 16 %(v/v)가 되도록 각각 첨가하여 20℃에서 72 시간 동안 배양한 후 660 nm에서의 흡광도를 측정하여 비교하였다. ATCC24858은 에탄올 내성이 높은 것으로 보고된 사카로마이세스 세레비지애 균주이다.Ethanol tolerance was measured by adding anhydrous ethanol at 0, 8, 12, and 12 h immediately after inoculation with yeast cells in a liquid medium of YPD (2% glucose, 0.5% yeast extract, 1% bacto-peptone; Sigma-Aldrich, St. Louis, MO, USA) 16% (v / v), respectively. After incubation at 20 ° C for 72 hours, the absorbance at 660 nm was measured and compared. ATCC 24858 is Saccharomyces cerevisiae which is reported to be highly ethanol resistant.
도 2에 나타난 바와 같이, 본 발명의 효모 균주 192-4의 에탄올 내성은 16 % 에탄올 함량에서도 ATCC24858 균주 보다 생존율이 더 높음을 확인하였다.As shown in FIG. 2, the ethanol tolerance of the yeast strain 192-4 of the present invention was higher than that of the ATCC 24858 strain even at the content of 16% ethanol.
관능 검사Sensory test
선발된 균주로 제조한 발효주를 시료로 사용하였다. 9점 척도(1점: 대단히 싫다, 5점: 좋지도 싫지도 않다, 9점: 대단히 좋다)에 의해 향 및 전체적인 기호도를 평가하였고, 시료는 상온에서 난수표로 표기되어 유리컵에 제시되었으며 무작위로 제시된 시료에 대해 평가하였다. 그 결과를 하기 표 4에 나타내었다.The selected strains were used as the selected strains. The fragrance and overall likelihood were evaluated by a 9-point scale (1 point: very disliked, 5 points: not good or not, 9 points: very good), and the samples were presented in glass cups at random The samples were evaluated. The results are shown in Table 4 below.
* 는 p < 0.05 의 유의 수준을 나타낸다.The mean value with superscript letters (a, b) represents a significant difference of 5% level as determined by Duncan's multiple test.
* Represents a significant level of p < 0.05.
상기 표 4에 나타나는 바와 같이, 본 발명에 따른 효모 균주 192-4로 제조된 발효주는 전반적인 기호도가 높음을 알 수 있었고, 신 향, 상큼한 향, 신 맛, 떫은 맛과 쓴 맛이 어우러져 전체적 기호도 9 점 평점법에서 5 점을 나타내었다.As shown in Table 4, the fermented product of the yeast strain 192-4 according to the present invention was found to have a high overall acceptability, and it was found that the overall acceptability of the fermented product was high with the combination of new flavor, refreshing flavor, fresh taste, And 5 points in the point scale method.
<110> KOREA FOOD RESEARCH INSTITUTE <120> BREWING YEAST SACCHAROMYCES CEREVISIAE 192-4 AND BREWED ALCOHOL MADE THEREWITH <130> IPDC-49785 <160> 1 <170> KopatentIn 2.0 <210> 1 <211> 639 <212> DNA <213> 18S rRNA of Saccharomyces cerevisiae 192-4 (KCCM11397P) <400> 1 gttggtaaaa cctaaaacga ccgtacttgc attatacctc aagcacgcag agaaacctct 60 ctttggaaaa aaaacatcca atgaaaaggc cagcaatttc aagttaactc caaagagtat 120 cactcactac caaacagaat gtttgagaag gaaatgacgc tcaaacaggc atgccccctg 180 gaataccaag gggcgcaatg tgcgttcaaa gattcgatga ttcacggaat tctgcaattc 240 acattacgta tcgcatttcg ctgcgttctt catcgatgcg agaaccaaga gatccgttgt 300 tgaaagtttt taatatttta aaatttccag ttacgaaaat tcttgttttt gacaaaaatt 360 taatgaataa ataaaattgt ttgtgtttgt tacctctggg ccccgattgc tcgaatgccc 420 aaagaaaaag ttgcaaagat atgaaaactc cacagtgtgt tgtattgaaa cggttttaat 480 tgtcctataa caaaagcaca gaaatctctc accgtttgga atagcaagaa agaaacttac 540 aggcctagca aaaccgcgca cttaagcgca ggcccggctg gactctccat ctcttgtctt 600 cttgcccagt aaaagctctc atgctcttgc caaaacaaa 639 <110> KOREA FOOD RESEARCH INSTITUTE <120> BREWING YEAST SACCHAROMYCES CEREVISIAE 192-4 AND BREWED ALCOHOL MADE THEREWITH <130> IPDC-49785 <160> 1 <170> Kopatentin 2.0 <210> 1 <211> 639 <212> DNA 18S rRNA of Saccharomyces cerevisiae 192-4 (KCCM11397P) <400> 1 gttggtaaaa cctaaaacga ccgtacttgc attatacctc aagcacgcag agaaacctct 60 ctttggaaaa aaaacatcca atgaaaaggc cagcaatttc aagttaactc caaagagtat 120 cactcactac caaacagaat gtttgagaag gaaatgacgc tcaaacaggc atgccccctg 180 gaataccaag gggcgcaatg tgcgttcaaa gattcgatga ttcacggaat tctgcaattc 240 acattacgta tcgcatttcg ctgcgttctt catcgatgcg agaaccaaga gatccgttgt 300 tgaaagtttt taatatttta aaatttccag ttacgaaaat tcttgttttt gacaaaaatt 360 taatgaataa ataaaattgt ttgtgtttgt tacctctggg ccccgattgc tcgaatgccc 420 aaagaaaaag ttgcaaagat atgaaaactc cacagtgtgt tgtattgaaa cggttttaat 480 tgtcctataa caaaagcaca gaaatctctc accgtttgga atagcaagaa agaaacttac 540 aggcctagca aaaccgcgca cttaagcgca ggcccggctg gactctccat ctcttgtctt 600 cttgcccagt aaaagctctc atgctcttgc caaaacaaa 639
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| KR101825957B1 (en) * | 2016-06-20 | 2018-02-07 | 대한민국 | Saccharomyces cerevisiae Y297 with adaptability at low temperature and manufacturing method of fermented alchoholic beverage using the same |
| KR20220073466A (en) | 2020-11-26 | 2022-06-03 | 한국식품연구원 | Yeast saccharomyces cerevisiae gnpea4, manufacturing method of distilled spirit with phehylethyl alcohol and phenethyl acetate flavour compounds using it, and distilled spirit manufactured therefrom |
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| KR100396959B1 (en) | 2000-11-10 | 2003-09-02 | 주식회사 두산 | Yeast new strain, Saccharomyces cerevisiae SC-55(No.KFCC-11224), method for isolating it and method for preparing fermentated liquor of plum using it |
| KR101135096B1 (en) | 2009-10-12 | 2012-04-16 | 영농조합법인 오가식품 | Manufacturing method of fermented alcoholic drink from Acanthopanax cortex |
| KR101166489B1 (en) | 2010-11-29 | 2012-07-19 | 한국식품연구원 | Brewing yeast Saccharomyces cerevisiae 183-2 and brewed alcohol made therewith |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100396959B1 (en) | 2000-11-10 | 2003-09-02 | 주식회사 두산 | Yeast new strain, Saccharomyces cerevisiae SC-55(No.KFCC-11224), method for isolating it and method for preparing fermentated liquor of plum using it |
| KR101135096B1 (en) | 2009-10-12 | 2012-04-16 | 영농조합법인 오가식품 | Manufacturing method of fermented alcoholic drink from Acanthopanax cortex |
| KR101166489B1 (en) | 2010-11-29 | 2012-07-19 | 한국식품연구원 | Brewing yeast Saccharomyces cerevisiae 183-2 and brewed alcohol made therewith |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101777555B1 (en) * | 2015-09-11 | 2017-09-13 | 대한민국 | Distilled liquor using Saccharomyces cerevisiae N9 and method for preparation thereof |
| KR101825957B1 (en) * | 2016-06-20 | 2018-02-07 | 대한민국 | Saccharomyces cerevisiae Y297 with adaptability at low temperature and manufacturing method of fermented alchoholic beverage using the same |
| KR20220073466A (en) | 2020-11-26 | 2022-06-03 | 한국식품연구원 | Yeast saccharomyces cerevisiae gnpea4, manufacturing method of distilled spirit with phehylethyl alcohol and phenethyl acetate flavour compounds using it, and distilled spirit manufactured therefrom |
| KR102411443B1 (en) | 2020-11-26 | 2022-06-22 | 한국식품연구원 | Yeast saccharomyces cerevisiae gnpea4, manufacturing method of distilled spirit with phehylethyl alcohol and phenethyl acetate flavour compounds using it, and distilled spirit manufactured therefrom |
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| KR20140120512A (en) | 2014-10-14 |
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