Screening of the Fine Brewer′s Yeast Strain JW1-3 and Its Pilot Fermentation

啤酒酵母菌种的优劣在啤酒酿造生产中起着至关重要的作用.为了获得优良的啤酒酿造酵母菌株,试验用半导体激光诱变处理啤酒酵母(Saccharomyces cerevisiae)菌株XB05,在含双乙酰的麦芽汁琼脂平板上分离抗双乙酰的突变株,然后用三角瓶在低温下发酵,以发酵液的发酵度、双乙酰、乙醛和总高级醇的含量为筛选指标,结果得到一株发酵特性优良的菌株JW1-3.菌株JW1-3以9°Bx麦芽汁为培养基,用500 L发酵罐在12℃下发酵13 d,发酵液的发酵度为67.8%,发酵液中的双乙酰、乙醛和总高级醇的含量分别为0.039 4、4.81和60.2 mg/L.菌株JW1-3的主要发酵特性优良且稳定,啤酒口感良好.该菌株在啤酒生产中具有很好的应用前景.Beer is one of the most popular drink at present,and the yeast strain plays a key role on the beer brewing.In order to obtain the fine brewer′s yeast strain,Saccharomyces cerevisiae strain XB05 was radiated by semiconductor laser(wavelength:630 nm;power:1 w) for 10,20 and 30 min.The mono-colonies growing on wort plate agar containing diacetyl 135 μg/mL were isolated.The anti-diacetyl mutants were fermented in 500 mL flask with 300 mL wort at 12℃ for 8 d and selected with the fermented degree and the content of diacetyl,acetaldehyde and total higher alcohols of the fermented liquid as the main target.As a result,the fine strain JW1-3 was obtained.Under the conditions of 500 L fermentor with 400 L 9°Bx wort at 12℃ for 13 d,the fermented degree of mutant JW1-3 was 67.8%,the content of diacetyl,acetaldehyde and total higher alcohols in the fermented liquid were 0.039 4,4.81 and 60.2 mg/L,respectively.The fermented properties of the mutant JW1-3 were relatively stable.The sensory evaluation of the beer produced by the mutant strain JW1-3 was nice.Strain JW1-3 is of well applied prospect on the beer brewing.福建雪津啤酒有限公司资

Screening of the Brewer′s Yeast Strain JY2-2 by Laser Mutagenesis and Its Pilot Fermentation

用He-Ne激光辐照诱变啤酒酵母(Saccharomyces cerevisiae)XB05,分离抗双乙酰的菌株,通过三角瓶发酵筛选,得到几株优良的菌株,其中菌株JY2-2用500 mL三角瓶装300 mL 12.5°Bx麦芽汁在12℃下发酵8 d,发酵液的发酵度为66%,双乙酰、乙醛和总高级醇的含量分别为0.079 0、7.43和87.22 mg/L.以12°Bx麦芽汁为培养基用500 L发酵罐发酵,发酵7 d发酵液的发酵度为68.8%,双乙酰、乙醛和总高级醇的含量分别为0.034 4、5.12和70.6 mg/L.啤酒口感纯正清爽,突变株JY2-2的主要发酵特性稳定,具有很好的应用前景.Several fine brewer′s yeast strains were obtained from the original strain XB05 after mutagenesis with laser and screening.Among the mutants,fermented degree of mutant JY2-2 was 66%,the content of diacetyl,acetaldehyde and total higher alcohols were respectively 0.079 0,7.43,87.22 mg/L under the fermentation conditions of 500 mL flask with 300 mL 12.5°Bx wort at 12℃ for 8 days.The fermentation degree of mutant JY2-2 was 68.8%,the content of diacetyl,acetaldehyde and total higher alcohols in the fermented liquid were respectively 0.034 4,70.6,5.12 mg/L under the conditions of 500 L fermentor with 400L 12°Bx wort at 12℃ for 7 days.The sensory evaluation of the beer produced by the mutant strain JY2-2 was nice.The fermentated properties of the mutant JY2-2 were relatively stable.JY2-2 is of well applied prospect on the beer brewing.福建雪津啤酒有限公司资

The Selection of Low Diacetyl Brewer's Yeast Strain ZT21-9-2 and Its ILV2 Gene Analysis

为了获得低产双乙酰的菌株,从保藏的啤酒工业酵母菌种中分离甲磺隆(SM)抗性自发突变株,经低温发酵测定发酵液的发酵度和双乙酰等啤酒风味物质的含量,结果得到ZT21-9-2等5株优良的SM抗性株.其中SM抗性株ZT21-9-2用500 L发酵罐发酵8 d,发酵液的发酵度为68.4%,发酵液的双乙酰、总高级醇和乙酸的含量分别为0.036 3、79.53和74.90 mg/L,该菌株具有很好的应用前景.为了探讨致使乙酰乳酸合成酶(ALS)编码基因ILV2突变而使ALS结构改变的关键碱基,为构建低产双乙酰的啤酒酵母工程菌提供依据,作者比较了SM抗性株ZT21-9-2及其亲株EY2-1的ILV2基因的DNA序列,结果发现SM抗性株ZT21-9-2共有6个碱基发生了转换,1个碱基发生了颠换.In order to gain brewer's yeast strains of low diacetyl yield,sulfometuron methyl(SM)-resistant spontaneous mutants were isolated from brewer's yeast strains kept in our laboratory.After low temperature fermentation,five SM-resistant strains with good fermentation characteristics were screened out by the determination of fermentation degree and the flavor elements contents such as diacetyl,total high alcohols and acetate in the fermented liquid.ZT21-9-2 is one of the strains with good application prospects.Following 8 days' fermentation in the 500 L fermentor at 12℃,the fermentation degree of mutant ZT21-9-2 was 68.4%,the contents of diacetyl,total high alcohols and acetate in the fermented liquid were respectively 0.036 3,79.53 and 74.90 mg/L.To find out the key mutation zone of the ILV2 gene which could lead to the disfunction of acetolactate synthase(ALS),and to provide the mechanism for the construction of low diacetyl brewer's yeast,the DNA sequences of the ILV2 gene encoding ALS of SM-resistant mutant ZT21-9-2 and its parent strain EY2-1 were compared,and six bases transition and one base transversion in the ILV2 gene of strain ZT21-9-2 were found.英博雪津啤酒有限公司资

Screening of the Fine Brewer’s Yeast Strain UN_(41)

以啤酒酵母(Saccharomyces cerevisiae) W作为出发菌株,用亚硝酸和紫外线连续诱变,在含双乙酰 150μg/mL(亚硝酸诱变)或200μg/mL(紫外线诱变)的麦芽汁琼脂平板上分离抗双乙酰的菌株,通过三角瓶发酵筛选得到一株优良的啤酒酵母菌株UN41.该菌株在500 mL三角瓶装300 mL 12°Bx麦芽汁的培养基中,12℃下发酵8 d,发酵液的发酵度为65.0%;双乙酰、高级醇和乙醛的含量分别为0.076 8 mg/L、92.15 mg/L和9.33 mg/L,比出发菌株W的降低了37.6%、19.9%和27.6%.The objective of this work was to obtain a fine brewer’s yeast strain by mutating Saccharomyces cerevisiae W.After continual mutagenesis with nitrous acid (HNO_2) and ultraviolet rays (UV) and isolating the mono-colonies growing on 6°Bx wort plate agar containing diacetyl 150 μg/mL (as treated with HNO_2) or 200 μg/mL (as treated with UV),49 mutants was selected.Fermentation degree and diacetyl,aldehyde and total higher alcohols content were taken as the main target,the fine strain UN_(41) was obtained from strain Saccharomyces cerevisiae W.In the fermented liquid of strain UN_(41) under the fermentation conditions of 500 mL flask with 300 mL 12°Bx wort at 12℃ for 8 days,degree of fermentation was 65.0%,the content of diacetyl,higher alcohols and acetaldehyde were 0.076 8 mg/L,92.15 mg/L and 9.33 mg/L,37.6%,19.9% and 27.6% lower than that of the original strain W,respectively. The strain UN_(41) is of potentially applicable to beer brewing

Study on Optimization of Flocculence and Fermentation Characteristics of Brewer's Yeast by Protoplast Fusion

以发酵度较高、凝絮性较差的啤酒酵母菌株H38的原生质体为受体,以凝絮性较强、发酵度较低的啤酒酵母菌株N1热灭活的原生质体为供体进行融合.用正交试验法分别优化两亲株的原生质体制备和再生的条件以及原生质体融合的条件.用制霉菌素抗性为遗传标记选择融合子.融合子初筛时以凝絮性为指标,复筛时以发酵度和发酵液中的主要风味物质的含量为指标.对得到的融合子进行了异核体和遗传稳定性的检验,结果得到两株凝絮性以及发酵特性较好的菌株HN31和HN40.The objective of this work was to optimize the flocculence and fermentation characters of brewer's yeast by protoplast fusion technology.Saccharomyces cerevisiae H38 and N1 were used as the parent strains for protoplast fusion.The strain H38 was a non-flocculence strain with a relatively high fermentation degree and the strain N1 was a strain with a relatively low fermentation degree and the relatively strong flocculence.The formational and regenerative conditions of strain H38 and N1 protoplast were optimized with the orthogonal test method.The protoplast fusants were constructed with H38 protoplasts as receptors and heat-inactivated strain N1 protoplasts as donors.The fusion conditions between strain H38 and N1 protoplast were optimized with the orthogonal test method.Anti-mycostatin character were used as genetic mark to select the fusion strains.Fusants were selected with flocculation ability,fermentation degree and content of diacetyl,acetaldehyde,total higher alcohols in the fermented liquid as screening criteria.As a result,two fine fusants with the relatively strong floccutence,high fermentation degree and low content of dicetyl,acetaldehyde and total alcohole in the fermented liquid were obtained.福建雪津啤酒有限公司资

Studies on fermentative characters of a brewer,syeast protoplast fusant GR8

研究了啤酒酵母原生质体融合株GR8的主要发酵特性。以12°Bx麦芽汁为三角瓶发酵培养基,在12℃下发酵8d,融合株GR8的发酵度为66.5%,凝絮性(本斯值)为3.0mL;发酵液中的双乙酰、乙醛、高级醇和乙酸乙酯等风味物质的含量分别为0.0583mg/L、9.66mg/L、91.20mg/L和22.8mg/L。从主要发酵特性的指标和口感品评以及遗传稳定性的结果表明,融合株GR8是一株具有工业应用前景的啤酒酿造酵母菌株。The fermentative characters of the brewer,s yeast protoplast fusant GR8 were studied. The results showed that flocculence ( Burns value )of the strain GR8 was 3.0mL. Under the fermentative conditions of 500mL flask with 300mL 12oBx wort at 12oC for 8 days,the real degree of fermentation of the strain GR8 was 66.5%, and content of diacetyl, acetaldehyde, acetic ether and total higher alcohols in the fermented liquid of the fusant GR8 were 0.0583mg/L, 9.66mg/L, 22.8mg/L and 91.20mg/L, respectively. The fermentative characters of the strain GR8 was stable during continued transfer of culture. The results showed that the strain GR8 has a good prospect in the application for beer brewing

Pilot Beer Brewing Test of Protoplast Fusant HN31-6 of Saccharomyces cerevisiae

为了考察啤酒酵母融合株HN31-6的啤酒酿造生产性能,以12.5°Bx麦芽汁为培养基,用500 L发酵罐在12℃下发酵,发酵期间每天取样测定发酵液的发酵度和酒精度以及发酵液的双乙酰、乙醛、高级醇和酯类等挥发性物质的含量.结果表明:在上述条件下,发酵14 d,发酵液的发酵度和酒精度分别为69.9%和4.46%;双乙酰和乙醛的含量分别为0.036 4 mg/L和4.53 mg/L;异戊醇、异丁醇和正丙醇的含量分别为46.47、8.34和9.82 mg/L,总高级醇的含量为64.6mg/L;甲酸乙酯、乙酸乙酯、乙酸异丁酯、乙酸异戊酯和己酸乙酯的含量分别为0.132、21.92、0.037、2.22和0.293 mg/L,总酯为24.6 mg/L;该菌株的发酵度以及发酵液的酒精、双乙酰、乙醛、总高级醇和一些酯类等的含量均达到国家优质啤酒的标准,啤酒口感良好.该融合株在啤酒酿造生产中具有很好的应用前景.In order to inspect the characters of beer brewing of protoplast fusant HN31-6 of Saccharomyces cerevisiae,the fermentation test was carried out under the conditions of 500 L fermentor with 400 L 12.5° Bx wort as the medium at 12℃ for 14 days.The fermentative degree of the fermented liquid and the contents of diacetyl etc volatility substances were analyzed during the fermentation.The results showed that the fermentative degree and alcohol degree of the fermented liquid of fusant HN31-6 under the above-mentioned conditions were 69.9% and 4.46%,respectively.The contents of diacetyl and acetaldehyde in the fermented liquid were(0.036 4) mg/L and 4.53 mg/L,respectively.The contents of isopentitol,isobutylalcohol and n-propyl-alcohol were 46.47,8.34 and 9.82 mg/L,respectively,and the content of total higher alcohols was 64.6 mg/L in the fermented liquid.The contents of methylacetate,ethyl acetate,isobutylacetate,isoamylacetate and ethylcaproate were 0.132,21.92,0.037,2.22 and 0.293 mg/L,respectively;and total ester was 24.6 mg/L.The fermentative degree of the fermented liquid and contents of diacetyl,acetaldehyde and total higher alcohols and some esters in the fermented liquid all achieved the standards of high quality beer.The sensory evaluation of the beer produced by fusant HN31-6 was nice.The fusant HN31-6 has a good prospect in the application for the beer brewing福建雪津啤酒有限公司资

Construction of the Fine Brewer's Yeast Strains DR9-2 by Fusion of Two Inactivated Parents' Protoplast and Study of its Fermentation Characters

啤酒酵母菌株JW1-1发酵液中的双乙酰、总高级醇的含量较低,但发酵度较低,发酵液中的乙醛含量较高;啤酒酵母菌株NW7-45发酵度较高,发酵液中的乙醛含量较低,但发酵液中的双乙酰、总高级醇的含量较高。为了得到优良的菌株,以紫外线灭活的菌株JW1-1原生质体和热灭活的菌株NW7-45原生质体为亲本进行融合。经三角瓶发酵筛选,得到较优良的融合株DR9-2和DR9-24。其中融合株DR9-2以12°Bx麦芽汁为培养基,用500L的发酵罐在12℃下发酵,发酵11d发酵液的发酵度为69.5%,发酵液中的双乙酰、乙醛和总高级醇的含量分别为0.0124mg/L、7.70mg/L和61.88mg/L。从融合株DR9-2的主要发酵特性和啤酒口感品评的结果表明,该菌株在啤酒酿造工业中具有应用前景。Sacchararomyces cerevisiae JW1-1 was a strain with the relatively low content of diacetyl and total higher alcohols but relatively high content of acetaldehyde in the fermented liquid and relatively low fermented degree, while Sacchararomyces cerevisiae NW7-45 was a strain with relatively high fermented degree and relatively low content of acetaldehyde but relatively high content of diacetyl and total higher alcohols in the fermented liquid. .In order to construct the fine brewer's yeast strains, the fusion of UV-inactivated strain JW1-1 protoplast and heat-inactivated strain NW7-45 protoplast was carried out. The fine fusants DR9-2 was obtained after selecting by flask fermentation. The content of diacetyl, acetaldehyde, total high alcohols of DR9-2 in the fermented liquid were 0.0124mg/L,7.70mg/L and 61.88mg/L, respectively, and the fermentation degree was 69.5% in 500L fermentor using 12°Bx wort as the medium at 12℃ for 11 days. The main fermentative property and the sensory evaluation of the beer of DR9-2 indicated that strain DR9-2 is of well applied prospect on the beer brewing

Construction of a fine brewer's yeast protoplast fusant GR_5 and its fermentation characteristics

以发酵度较高的非絮凝性的啤酒酵母菌株X6和发酵度较低、絮凝性较强的啤酒酵母菌株N1为亲本进行原生质体融合。用亚硝酸诱变原养型的菌株X6,经筛选得到一株需酪素水解物的营养缺陷型菌株X6~20。采用正交试验法分别优化菌株X6~20和N1的原生质体形成和再生的条件。用X6~20菌株的原生质体作为受体和热灭活的N1菌株原生质体作为供体进行融合。融合株经三角瓶发酵筛选,得到一株较优良的融合株GR5。该融合株的絮凝性较强(本斯值为2.7),以12°Bx麦芽汁为培养基,用500 L的发酵罐在12℃下发酵,发酵至第8 d菌株GR5的发酵度为69.2%,发酵液中的双乙酰含量为0.0498 mg/L、乙醛含量为6.34 mg/L,总高级醇含量为74.4mg/L。融合株GR5具有双亲的优点,发酵的啤酒风味较好,是一株具有工业应用前景的啤酒酿造酵母菌株。Saccharomyces cerevisiae X_6 and N_1 strains were used as the parent strains for protoplast fusion.The strain X_6 was a non-flocculence strain with higher fermentation degree and the strain N_1 was a strain with lower fermentation(degree) and stronger flocculence.The auxotrophic mutant X_(6～20) required casein hydrolysate was derived from the(prototrophic) strain X_6 by treatment of nitrous acid(HNO_2) and nutritional identification.The formation and regeneration conditions of strain X_(6～20) and N_1 protoplast were optimized by orthogonal test method.The protoplast fusants were(constructed) with auxotrophic mutant X_(6～20) protoplasts for receptors and heat-inactivated strain N_1 protoplasts for(suppliers).The fusants obtained were fermented and selected.As a result a fine fusant strain GR_5 was obtained.The flocculence of fusant strain GR_5 was stronger with 2.7 mL of burns value.The content of diacetyl,acetaldehyde,total higher(alcohols) in the(fermented) liquid were 0.0498 mg/L,6.34 mg/L,74.4 mg/L,respectively,and the fermentation degree was(69.2 %) in 500 L fermentor using 12°Bx wort as the medimum at 12℃ for 8 days.The sensory evaluation of the beer produced by the fusant strain GR_5 was nice.The results showed that the fusant strain GR_5 with the fine properties from its parent strains and well(applied) prospect in the beer brewing