CN101148637A - Hydrolyzate rich in biologically active peptides for promoting beer yeast fermentation and its preparation method and application - Google Patents
Hydrolyzate rich in biologically active peptides for promoting beer yeast fermentation and its preparation method and application Download PDFInfo
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- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 38
- 238000000855 fermentation Methods 0.000 title claims abstract description 30
- 230000004151 fermentation Effects 0.000 title claims abstract description 30
- 240000004808 Saccharomyces cerevisiae Species 0.000 title claims abstract description 22
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 title claims abstract description 22
- 102000004196 processed proteins & peptides Human genes 0.000 title claims abstract description 22
- 238000002360 preparation method Methods 0.000 title claims abstract description 11
- 230000001737 promoting effect Effects 0.000 title claims abstract description 11
- 241000186226 Corynebacterium glutamicum Species 0.000 claims abstract description 33
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- 229940062043 nitrogen 50 % Drugs 0.000 claims 1
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- 239000002994 raw material Substances 0.000 abstract 1
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 9
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Abstract
本发明公开了一种富含促进啤酒酵母发酵生物活性肽的水解物及其制备方法和应用，水解物是以谷氨酸棒状杆菌为原料制备得到，所述生物活性肽的分子量集中在500Da～2000Da，生物活性肽占总氮的50％～60％质量。该水解物应用于啤酒生产，可显著缩短发酵时间，提高发酵液中乙醇含量，降低发酵液中残糖含量。The invention discloses a hydrolyzate rich in biologically active peptides for promoting beer yeast fermentation and its preparation method and application. The hydrolyzate is prepared from Corynebacterium glutamicum as a raw material, and the molecular weight of the biologically active peptides is concentrated in the range of 500Da to 2000Da, bioactive peptides account for 50% to 60% of the total nitrogen by mass. The hydrolyzate is applied to beer production, which can significantly shorten the fermentation time, increase the ethanol content in the fermentation liquid, and reduce the residual sugar content in the fermentation liquid.
Description
技术领域 technical field
本发明涉及食品生物技术领域，具体涉及一种富含促进啤酒酵母发酵生物活性肽的水解物，其制备方法及应用。The invention relates to the field of food biotechnology, in particular to a hydrolyzate rich in biologically active peptides for promoting beer yeast fermentation, its preparation method and application.
背景技术 Background technique
谷氨酸棒状杆菌，革兰氏阳性菌，严格好氧，不产生孢子，是谷氨酸发酵后经超滤分离所得废弃物的主要成分。谷氨酸棒状杆菌含有丰富的蛋白质、核酸、碳水化合物、灰分、维生素以及微生物生长必需的微量元素，以干基计，其蛋白质含量在70％以上。谷氨酸棒状杆菌蛋白质的氨基酸组成具明显的特色，其中谷氨酸含量占总蛋白的30.60％，天冬氨酸占8.47％，丙氨酸占7.45％，甘氨酸占3.92％，，共计50.44％，这四种氨基酸均为酵母细胞非常容易吸收的有机氮源。然而，由于谷氨酸棒状杆菌细胞壁较厚，破壁困难，开发利用难度较大，目前谷氨酸棒杆菌主要用于肥料生产，效益和生物利用价值都较低。Corynebacterium glutamicum, a Gram-positive bacterium, is strictly aerobic and does not produce spores. It is the main component of the waste obtained by ultrafiltration after glutamic acid fermentation. Corynebacterium glutamicum is rich in protein, nucleic acid, carbohydrate, ash, vitamins and trace elements necessary for microbial growth, and its protein content is more than 70% on a dry basis. The amino acid composition of Corynebacterium glutamicum protein has obvious characteristics, among which glutamic acid accounts for 30.60% of the total protein, aspartic acid accounts for 8.47%, alanine accounts for 7.45%, glycine accounts for 3.92%, and a total of 50.44%. , these four amino acids are organic nitrogen sources that yeast cells can easily absorb. However, due to the thick cell wall of Corynebacterium glutamicum, it is difficult to break the wall, and it is difficult to develop and utilize it. At present, Corynebacterium glutamicum is mainly used for fertilizer production, and its benefit and biological utilization value are low.
常规啤酒发酵过程比较长，同时乙醇产量低，发酵液中残糖含量较高，影响了生产厂家的生产效率和产品的质量。寻找一种成本较低的物质和方法来促进啤酒酵母发酵十分迫切。The conventional beer fermentation process is relatively long, while the ethanol yield is low, and the residual sugar content in the fermentation liquid is high, which affects the production efficiency of the manufacturer and the quality of the product. It is very urgent to find a material and method with lower cost to promote the fermentation of brewer's yeast.
发明内容 Contents of the invention
本发明的目的是提供一种以谷氨酸棒状杆菌为原料制备得到促进啤酒酵母发酵的生物活性肽。The object of the present invention is to provide a biologically active peptide prepared from Corynebacterium glutamicum for promoting the fermentation of brewer's yeast.
本发明的另一个目的是提供所述生物活性肽的制备方法。Another object of the present invention is to provide a preparation method of the biologically active peptide.
本发明还有一个目的是提供所述生物活性肽的应用。Another object of the present invention is to provide the application of said biologically active peptide.
本发明的技术方案是提供一种促进啤酒酵母发酵的生物活性肽，以谷氨酸棒状杆菌为原料制备得到，所述生物活性肽的分子量主要集中在500Da～2000Da，肽占总氮的50％～60％质量。The technical solution of the present invention is to provide a biologically active peptide that promotes the fermentation of beer yeast, which is prepared from Corynebacterium glutamicum. The molecular weight of the biologically active peptide is mainly concentrated in the range of 500Da to 2000Da, and the peptide accounts for 50% of the total nitrogen. ~60% mass.
本发明提供了所述促进啤酒酵母发酵的生物活性肽的制备方法，是采用两种内切蛋白酶限制性水解谷氨酸棒状杆菌菌体得到。The invention provides the preparation method of the bioactive peptide for promoting the fermentation of brewer's yeast, which is obtained by restrictively hydrolyzing Corynebacterium glutamicum cells with two kinds of endoproteases.
所述两种内切蛋白酶为木瓜蛋白酶或碱生蛋白酶与胰凝乳蛋白酶或胰蛋白酶，选用食品级蛋白酶。The two endoproteases are papain or alkaline protease and chymotrypsin or trypsin, and food-grade protease is selected for use.
所述促进啤酒酵母发酵的生物活性肽的制备方法包括以下步骤：The preparation method of the bioactive peptide for promoting the fermentation of brewer's yeast comprises the following steps:
(1)将谷氨酸棒状杆菌与水以1∶1质量比例混合，搅拌均匀制备谷氨酸棒状杆菌液；(1) mixing Corynebacterium glutamicum and water in a mass ratio of 1:1, stirring evenly to prepare Corynebacterium glutamicum liquid;
(2)采用高压均质机对谷氨酸棒状杆菌液进行均质预处理，以改变谷氨酸棒状杆菌细胞壁的通透性；(2) Carry out homogeneous pretreatment to the Corynebacterium glutamicum liquid by using a high-pressure homogenizer to change the permeability of the Corynebacterium glutamicum cell wall;
(3)在55℃～65℃，pH6.0～7.5条件下先采用木瓜蛋白酶或碱性蛋白酶(AlcalaseTM)进行水解，水解度达到5％～6％后，添加胰蛋白酶或胰凝乳蛋白酶进行水解；(3) Under the conditions of 55°C-65°C and pH 6.0-7.5, first use papain or alkaline protease (Alcalase TM ) for hydrolysis, and after the degree of hydrolysis reaches 5%-6%, add trypsin or chymotrypsin carry out hydrolysis;
(4)当水解度达到10～12％之间时，灭酶终止水解；(4) When the degree of hydrolysis reaches between 10% and 12%, the hydrolysis is terminated by inactivating the enzyme;
(5)将水解液离心除去谷氨酸棒状杆菌细胞壁；(5) Centrifuging the hydrolyzate to remove the cell wall of Corynebacterium glutamicum;
(6)将水解液过5000Da的超滤膜除去大分子量多肽和蛋白质，干燥得富含促进啤酒酵母发酵生物活性肽的水解物。(6) Pass the hydrolyzate through a 5000Da ultrafiltration membrane to remove large molecular weight polypeptides and proteins, and dry to obtain a hydrolyzate rich in bioactive peptides that promote brewer's yeast fermentation.
其中，步骤(2)所述的均质处理条件为30MPa～45MPa，均质两次；(4)所述灭酶条件为85～90℃温度，时间为15min～20min，所述水解度的测定采用pH-Stat法；步骤(5)所述离心条件是6000r/m，时间为15min。Wherein, the homogenization treatment condition described in step (2) is 30MPa～45MPa, homogenize twice; (4) the described enzymatic deactivation condition is 85～90 ℃ temperature, time is 15min～20min, the determination of the hydrolysis degree The pH-Stat method is adopted; the centrifugal condition in step (5) is 6000r/m, and the time is 15min.
本发明提供了所述促进啤酒酵母发酵的生物活性肽的应用，以0.1％～0.3％质量的浓度添加到酵母发酵液中，应用于啤酒生产。The invention provides the application of the biologically active peptide for promoting the fermentation of beer yeast, which is added to the yeast fermentation liquid at a concentration of 0.1% to 0.3% by mass and applied to beer production.
本发明与现有技术相比具有如下优点：Compared with the prior art, the present invention has the following advantages:
1、为谷氨酸发酵后超滤分离后所得废弃物谷氨酸棒状杆菌提供了新的、市场前景广泛的应用。1. It provides a new application with wide market prospects for the waste Corynebacterium glutamicum obtained after ultrafiltration separation after glutamic acid fermentation.
2、提供了一种新的富含生物活性肽的水解物，可显著缩短酵母发酵时间，提高乙醇产量，降低发酵液中残糖含量，从而大大提高啤酒生产厂家的生产效率和经济效益。2. Provide a new hydrolyzate rich in biologically active peptides, which can significantly shorten the fermentation time of yeast, increase ethanol production, and reduce the residual sugar content in the fermentation liquid, thereby greatly improving the production efficiency and economic benefits of beer manufacturers.
具体实施方式 Detailed ways
实施例1Example 1
(1)将谷氨酸棒状杆菌与水各1kg，即以1∶1重量比例混合，搅拌均匀制备谷氨酸棒状杆菌液；(1) each 1kg of Corynebacterium glutamicum and water are mixed in a 1:1 weight ratio, and stirred evenly to prepare Corynebacterium glutamicum liquid;
(2)采用高压均质机对谷氨酸棒状杆菌进行均质处理，30MPa均质处理2次；(2) Adopt high-pressure homogenizer to carry out homogenization treatment to Corynebacterium glutamicum, 30MPa homogenization treatment 2 times;
(3)在55℃，用氢氧化钠溶液调pH至6.0后，先采用木瓜蛋白酶进行水解，用pH-Stat法控制水解度达到5.0％后，添加胰凝乳蛋白酶进行水解，采用pH-Stat法控制水解度至10.2％，得酶解液；(3) At 55°C, adjust the pH to 6.0 with sodium hydroxide solution, first use papain to hydrolyze, and use the pH-Stat method to control the degree of hydrolysis to 5.0%, then add chymotrypsin to hydrolyze, and use pH-Stat method to control the degree of hydrolysis to 10.2%, to obtain the enzymatic solution;
(4)将酶解液在85℃保持18min，使蛋白酶失活；(4) Keep the enzymatic solution at 85°C for 18 minutes to inactivate the protease;
(5)将酶解液6000r/m离心15min，除去谷氨酸棒状杆菌细胞壁；(5) Centrifuge the enzymatic solution at 6000r/m for 15min to remove the cell wall of Corynebacterium glutamicum;
(6)将水解液过5000Da的超滤膜除去大分子量多肽和蛋白质，获得富含促进酵母菌发酵生物活性肽的酶解物。生物活性肽分子量范围为500Da～2000Da，生物活性肽占总氮的54.3％。所述生物活性肽促进酵母菌发酵机能与其它发酵氮源实验数据比较见表1。(6) Pass the hydrolyzate through a 5000Da ultrafiltration membrane to remove large molecular weight polypeptides and proteins, and obtain an enzymatic hydrolyzate rich in bioactive peptides that promote yeast fermentation. The molecular weight range of the biologically active peptide is 500Da-2000Da, and the biologically active peptide accounts for 54.3% of the total nitrogen. Table 1 compares the experimental data of the bioactive peptide promoting yeast fermentation with other fermentative nitrogen sources.
表1Table 1
实施例2Example 2
(1)将谷氨酸棒状杆菌与水各1kg，混合，搅拌均匀制备谷氨酸棒状杆菌液；(1) each 1kg of Corynebacterium glutamicum and water are mixed, stirred evenly to prepare Corynebacterium glutamicum liquid;
(2)采用高压均质机对谷氨酸棒状杆菌液在35MPa下均质处理2次；(2) Adopting a high-pressure homogenizer to homogenize the Corynebacterium glutamicum liquid at 35MPa for 2 times;
(3)在60℃，采用氢氧化钠溶液调pH至6.8，先采用碱性蛋白酶进行水解，当水解度达到5.5％后(pH-Stat法)，添加胰蛋白酶进行水解，采用pH-Stat法控制水解度至11.2％，得酶解液；(3) At 60°C, use sodium hydroxide solution to adjust the pH to 6.8, first use alkaline protease for hydrolysis, and when the degree of hydrolysis reaches 5.5% (pH-Stat method), add trypsin for hydrolysis, use pH-Stat method Control the degree of hydrolysis to 11.2% to obtain the enzymatic solution;
(4)将(3)酶解液在88℃保持15min，使蛋白酶失活；(4) Keep the enzymatic hydrolysis solution of (3) at 88°C for 15 minutes to inactivate the protease;
(5)将酶解液6000r/m离心操作15min，除去谷氨酸棒状杆菌细胞壁；(5) Centrifuge the enzymatic solution at 6000r/m for 15min to remove the cell wall of Corynebacterium glutamicum;
(6)将水解液过5000Da的超滤膜除去大分子量多肽和蛋白质，获得富含促进酵母菌发酵生物活性肽的水解物。经凝胶过滤色谱法测定得出生物活性肽分子量范围在500Da-2000Da，生物活性肽占总氮的56.3％。(6) Pass the hydrolyzate through a 5000Da ultrafiltration membrane to remove large molecular weight polypeptides and proteins, and obtain a hydrolyzate rich in bioactive peptides that promote yeast fermentation. The molecular weight range of the biologically active peptide is 500Da-2000Da determined by gel filtration chromatography, and the biologically active peptide accounts for 56.3% of the total nitrogen.
实施例3Example 3
(1)将谷氨酸棒状杆菌与水以1∶1质量比混合均匀制备谷氨酸棒状杆菌液；(1) mixing Corynebacterium glutamicum and water with a mass ratio of 1:1 to prepare Corynebacterium glutamicum liquid;
(2)采用高压均质机对谷氨酸棒状杆菌液进行均质处理，45MPa条件下均质处理2次；(2) Adopt high-pressure homogenizer to carry out homogenization treatment to Corynebacterium glutamicum liquid, under the condition of 45MPa homogenization treatment 2 times;
(3)在65℃，采用氢氧化钠溶液调pH至7.5，先采用木瓜蛋白酶进行水解，用pH-Stat法控制水解度达到5.8％后，添加胰蛋白酶进行水解，采用pH-Stat法控制水解度至11.8％；(3) At 65°C, use sodium hydroxide solution to adjust the pH to 7.5, first use papain to hydrolyze, use the pH-Stat method to control the degree of hydrolysis to 5.8%, add trypsin for hydrolysis, and use the pH-Stat method to control the hydrolysis to 11.8%;
(4)将(3)酶解液在90℃保持15min，使蛋白酶失活；(4) Keep the enzymatic hydrolysis solution of (3) at 90°C for 15 minutes to inactivate the protease;
(5)将水解液在6000r/m离心操作15min，除去谷氨酸棒状杆菌细胞壁；(5) The hydrolyzate was centrifuged at 6000r/m for 15min to remove the cell wall of Corynebacterium glutamicum;
(6)将水解液过5000Da的超滤膜除去大分子量多肽和蛋白质，获得富含促进酵母菌发酵生物活性肽的水解物。经凝胶过滤色谱测定水解物中生物活性肽分子量集中在500-2000Da，生物活性肽占总氮的58.2％。(6) Pass the hydrolyzate through a 5000Da ultrafiltration membrane to remove large molecular weight polypeptides and proteins, and obtain a hydrolyzate rich in bioactive peptides that promote yeast fermentation. The molecular weight of biologically active peptides in the hydrolyzate was determined by gel filtration chromatography to be concentrated at 500-2000Da, and the biologically active peptides accounted for 58.2% of the total nitrogen.
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| CN2007100296759A Expired - Fee Related CN101148637B (en) | 2007-08-10 | 2007-08-10 | Hydrolysate enriched with biological bioactive peptide for accelerating beer yeast fermenting and its preparation method and application |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102550802A (en) * | 2011-12-19 | 2012-07-11 | 成都宏安生物科技有限公司 | Method for extracting polypeptide and amino acid from waste beer yeast |
| CN102925521A (en) * | 2012-10-31 | 2013-02-13 | 华南理工大学 | Soybean peptide capable of shortening ultrahigh-concentration beer fermentation period, preparation method and application thereof |
| CN107384658A (en) * | 2017-08-08 | 2017-11-24 | 王启含 | A kind of active peptide beer |
| CN108041339A (en) * | 2017-11-15 | 2018-05-18 | 兰溪市酉泽饲料技术服务有限公司 | Add the prawn children phase feed of Sage extract |
| CN108157667A (en) * | 2017-11-15 | 2018-06-15 | 兰溪市酉泽饲料技术服务有限公司 | Add the preparation method of the shallow sea fish meal of ocean rhodotorula extract |
| CN113862322A (en) * | 2021-11-15 | 2021-12-31 | 唐山拓普生物科技有限公司 | Preparation method and application of beer yeast polypeptide with high antioxidant activity |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| FR2410476B1 (en) * | 1977-11-30 | 1980-08-22 | Inst Nat Sante Rech Med | APPLICATION OF PEPTIDOGLYCANS EXTRACTED FROM BACTERIA, PARTICULARLY BACILLUS AS MEDICAMENTS, ESPECIALLY STIMULATING ANTI-TUMOR IMMUNITY |
| CN1187947A (en) * | 1997-06-10 | 1998-07-22 | 李红光 | Production of yeast fodder by concentrated treatment of ethyl alcohol or aginomoto waste water |
| CN1152043C (en) * | 2001-02-27 | 2004-06-02 | 武汉生科生物股份有限公司 | Nucleotide extracting process from waste gourmet powder material |
| CN1300327C (en) * | 2005-04-22 | 2007-02-14 | 广东省微生物研究所 | Process for preparing peptidoglycan |
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2007
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102550802A (en) * | 2011-12-19 | 2012-07-11 | 成都宏安生物科技有限公司 | Method for extracting polypeptide and amino acid from waste beer yeast |
| CN102550802B (en) * | 2011-12-19 | 2013-09-18 | 成都宏安生物科技有限公司 | Method for extracting polypeptide and amino acid from waste beer yeast |
| CN102925521A (en) * | 2012-10-31 | 2013-02-13 | 华南理工大学 | Soybean peptide capable of shortening ultrahigh-concentration beer fermentation period, preparation method and application thereof |
| CN107384658A (en) * | 2017-08-08 | 2017-11-24 | 王启含 | A kind of active peptide beer |
| CN108041339A (en) * | 2017-11-15 | 2018-05-18 | 兰溪市酉泽饲料技术服务有限公司 | Add the prawn children phase feed of Sage extract |
| CN108157667A (en) * | 2017-11-15 | 2018-06-15 | 兰溪市酉泽饲料技术服务有限公司 | Add the preparation method of the shallow sea fish meal of ocean rhodotorula extract |
| CN113862322A (en) * | 2021-11-15 | 2021-12-31 | 唐山拓普生物科技有限公司 | Preparation method and application of beer yeast polypeptide with high antioxidant activity |
| CN113862322B (en) * | 2021-11-15 | 2024-05-14 | 唐山拓普生物科技有限公司 | Preparation method and application of beer yeast polypeptide with high antioxidant activity |
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| CN101148637B (en) | 2011-05-11 |
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