CN107904058B - Preparation method of beer fermented by mixing aroma-producing yeast and saccharomyces cerevisiae - Google Patents
Preparation method of beer fermented by mixing aroma-producing yeast and saccharomyces cerevisiae Download PDFInfo
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- CN107904058B CN107904058B CN201711435833.0A CN201711435833A CN107904058B CN 107904058 B CN107904058 B CN 107904058B CN 201711435833 A CN201711435833 A CN 201711435833A CN 107904058 B CN107904058 B CN 107904058B
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12C—BEER; PREPARATION OF BEER BY FERMENTATION; PREPARATION OF MALT FOR MAKING BEER; PREPARATION OF HOPS FOR MAKING BEER
- C12C12/00—Processes specially adapted for making special kinds of beer
- C12C12/002—Processes specially adapted for making special kinds of beer using special microorganisms
- C12C12/006—Yeasts
Abstract
The invention discloses a preparation method of beer by mixed fermentation of aroma-producing yeast and saccharomyces cerevisiae, and relates to the technical field of beer brewing. Comprises the following steps: (a) inoculating saccharomyces cerevisiae and pichia pastoris on the inclined plane into 12-DEG P whole wheat juice, performing step-by-step propagation, and collecting yeast paste; (b) pulverizing fructus Hordei Germinatus, saccharifying, filtering, boiling, adding flos Lupuli and precipitate, and making into 12 ° P whole wheat juice; (c) quickly cooling the wort to 10 ℃, inoculating saccharomyces cerevisiae mud (3.0-6.0g/L) and pichia pastoris mud (0.01-1.0 g/L), and carrying out main fermentation for 7 days at the temperature of 8-12 ℃; performing after-fermentation at 0-4 deg.C for 5-7 days; filtering to obtain beer. The pichia pastoris and the saccharomyces cerevisiae with the flavor of increasing the beer aroma are mixed and fermented to brew the beer, so that the ester aroma substance content in the beer is increased, and the beer obtained by fermentation has excellent quality, rich ester aroma, harmonious flavor and mellow wine body.
Description
Technical Field
The invention relates to the technical field of beer brewing, in particular to a preparation method of beer by mixed fermentation of aroma-producing yeast and saccharomyces cerevisiae.
Background
The beer is a low-concentration alcoholic beverage containing carbon dioxide gas, which is brewed by using barley, hop and water as main raw materials and fermenting the raw materials at low temperature through beer yeast. Beer is beverage wine with complex components, and the flavor components of the beer are mostly generated in the brewing process (wort saccharification and fermentation) except for brewing raw materials (malt, hops and the like), wherein the performance of yeast and related process conditions play an important role in the generation of the flavor components. The main flavors of beer include: higher alcohols, esters, organic acids, aldehydes, etc., and these metabolites are mainly produced by the metabolism of brewer's yeast. At present, most of yeasts used for producing beer are Saccharomyces cerevisiae (Saccharomyces cerevisiae), single application of the yeast for fermentation often causes monotonous flavor and flat taste of products, and meanwhile, a large amount of auxiliary materials (rice, starch and syrup) are added in the beer brewing process, and the beer flavor is also slight due to a high-concentration beer brewing technology. At present, the method for improving the beer flavor is mainly realized by breeding beer yeast and adjusting the fermentation process. However, the effect of these methods on the improvement of beer flavor is not significant. Therefore, the invention aims to prepare the beer with rich alcohol and ester fragrance by a method of mixed fermentation of aroma-producing yeast and saccharomyces cerevisiae.
Disclosure of Invention
The invention utilizes the non-saccharomyces yeast with the flavor of beer aroma and the saccharomyces cerevisiae with high fermentation efficiency to carry out mixed fermentation to brew the beer, thereby not only increasing the content of ester aroma substances in the beer, but also ensuring that the yeast for beer fermentation has higher fermentation efficiency. Yeasts that impart bouquet in alcoholic fermentation are called aroma-producing yeasts. The Pichia pastoris (Pichia fabianii) can generate a certain amount of esters, higher alcohols and lower fatty acids in the fermentation process, so that the beer has rich alcohol and ester flavors, the content of volatile flavor substances can be increased when the Pichia pastoris is mixed with the saccharomyces cerevisiae for fermentation, and the beer obtained by fermentation has excellent quality, rich ester flavor, harmonious flavor and mellow wine body.
Discloses a preparation method of beer by mixed fermentation of aroma-producing yeast and saccharomyces cerevisiae, relating to the technical field of beer brewing. Comprises the following steps:
(a) pulverizing fructus Hordei Germinatus, saccharifying, filtering, boiling, adding flos Lupuli and precipitate, and making into 12 ° P whole wheat juice; respectively inoculating saccharomyces cerevisiae and aroma-producing yeast into 10-12 DEG P whole wheat juice for stepwise propagation, wherein the bacterial load reaches 5-10g/L, and respectively centrifuging to obtain yeast paste;
(b) quickly cooling 12 DEG P whole wheat juice to 10 ℃, inoculating 3.0-6.0g/L saccharomyces cerevisiae mud and 0.01-1.0 g/L aroma-producing yeast mud, performing main fermentation and after-fermentation, and filtering to obtain the beer.
In the case of the above-described embodiment, the aroma-producing yeast is Pichia pastoris (Pichia fabianii).
In the technical scheme, the main fermentation is carried out for 7 days at the temperature of 8-12 ℃; and (4) storing for 5-7 days at 0-4 ℃ for after-fermentation.
In the technical scheme, when the concentration of the wort of the main fermentation is reduced to 3.5-4.0 DEG P, the tank is sealed and the pressure is increased, and the pressure is kept at 0.11-0.13 MPa; and (3) performing after-fermentation, reducing the temperature of the fermentation tank to 0-4 ℃ within 24 hours, and storing for 5-7 days, wherein the fermentation end time is that the content of diacetyl is reduced to 0.1 mg/L.
In the technical scheme, the saccharomyces cerevisiae paste is inoculated into 10-12 DEG P whole wheat juice for fermentation according to the inoculation amount of 4.0 g/L.
In the technical scheme, the pichia pastoris is inoculated into 10-12 DEG P whole wheat juice for fermentation according to the inoculation amount of 0.2 g/L.
In the technical scheme, the saccharomyces cerevisiae is prepared by inoculating saccharomyces cerevisiae on an inclined plane into whole wheat juice at 12 degrees P, performing shake culture at 28 ℃ for 24 hours, inoculating the saccharomyces cerevisiae into a triangular flask filled with 50mL of whole wheat juice according to the inoculation amount of 5%, performing shake culture at 20 ℃ for 36 hours at 150rpm, transferring the whole wheat juice into 300mL of fresh 10-12 degrees P, performing shake culture at 16 ℃/100rpm for 72 hours, centrifuging the whole wheat juice for 10 minutes at 15 ℃/6000rpm, and collecting yeast paste.
In the technical scheme, the preparation method of the aroma-producing yeast comprises the steps of inoculating the aroma-producing yeast on the inclined plane into 10-12 DEG P whole wheat juice, carrying out shake cultivation at 28 ℃ for 24h, inoculating the aroma-producing yeast into a triangular flask filled with 50mL of whole wheat juice according to the inoculation amount of 5%, carrying out shake cultivation at 20 ℃ for 36h, transferring the aroma-producing yeast into 300mL of fresh 10-12 DEG P whole wheat juice, carrying out shake cultivation at 16 ℃/100rpm for 72h, centrifuging the aroma-producing yeast for 10min at 15 ℃/6000rpm, and collecting yeast paste.
In the above technical solution, the whole wheat juice preparation method comprises: crushing malt, and uniformly mixing the crushed malt with water according to the material-water ratio of 1: 3; placing in a saccharifying pot, and saccharifying by saccharifying process of keeping temperature at 45 deg.C for 30min, heating to 63 deg.C, keeping temperature for 60min, heating to 72 deg.C, keeping temperature for 10min, heating to 78 deg.C, and keeping temperature for 10 min; and (3) after the obtained mash passes through a filter tank, adding hops with the addition of 1.5g/L, adding into the boiled wheat juice in batches, boiling the wheat juice for 90min, and removing precipitates in a rotary sedimentation pot to obtain the 12-DEG P whole wheat juice.
The beer is brewed by mixed fermentation of pichia pastoris and saccharomyces cerevisiae with flavor for increasing beer aroma, and the imported Australian malt and the Saflager produced in France are preferably used in the inventionTMSaccharomyces cerevisiae was fermented under S-189. The Pichia pastoris (Pichia fabianii CICC 1040) can be purchased from the China center for Industrial culture of microorganisms (CICC). The content of ester flavor substances in the beer is increased, the beer obtained by fermentation has excellent quality, rich ester flavor, harmonious flavor and mellow wine body.
Advantageous effects
The preparation method of the beer by mixed fermentation of the aroma-producing yeast and the saccharomyces cerevisiae, which is provided by the invention, aims to solve the problems of monotonous flavor and flat taste of the following brewed beer, and the pichia pastoris and the saccharomyces cerevisiae which have the flavor of increasing the aroma of the beer are used for mixed fermentation to brew the beer, so that the content of ester aroma substances in the beer is increased, and the yeast for beer fermentation is ensured to have higher fermentation efficiency. The total ester content is improved by 20 percent through the mixed fermentation of the pichia pastoris and the saccharomyces cerevisiae, the preparation method is simple, and the rich ester aroma, the harmonious flavor and the mellow wine body of the beer are ensured.
Detailed Description
The following non-limiting examples will allow one of ordinary skill in the art to more fully understand the present invention, but are not intended to limit the invention in any way. The present invention will be described in further detail with reference to specific embodiments.
The raw materials of malt, hop and bottom fermentation Saccharomyces cerevisiae for producing beer of the present invention can be purchased from related beer production material companies, and the present invention preferably uses imported Australian malt and Saflager produced in FranceTMSaccharomyces cerevisiae was fermented under S-189. The Pichia pastoris (Pichia fabianii CICC 1040) can be purchased from the China center for Industrial culture of microorganisms (CICC).
EXAMPLE 1 beer of Mixed fermentation of aroma-producing Yeast and Saccharomyces cerevisiae
(1) Inoculating saccharomyces cerevisiae and pichia pastoris on the inclined plane into 12 DEG P whole wheat juice, performing shake culture at 28 ℃ for 24h, inoculating the mixture into a triangular flask filled with 50mL of whole wheat juice according to the inoculation amount of 5%, performing shake culture at 20 ℃ for 36h by using a shaking table (150rpm), transferring the mixture into 300mL of fresh whole wheat juice, performing shake culture at 16 ℃/100rpm for 72h, centrifuging at 15 ℃/6000rpm for 10min, and collecting yeast paste.
(2) After being crushed into 20 kg, the malt is evenly mixed with water according to the material-water ratio of 1: 3. Placing in a saccharifying pot, and saccharifying by saccharifying process of keeping temperature at 45 deg.C for 30min, heating to 63 deg.C, keeping temperature for 60min, heating to 72 deg.C, keeping temperature for 10min, heating to 78 deg.C, and keeping temperature for 10 min; filtering the obtained mash, adding flos Lupuli (the addition amount of flos Lupuli is 1.5g/L, and adding into boiled wheat juice in batches), boiling wheat juice for 90min, removing precipitate in rotary sedimentation pot to obtain 12 ° P whole wheat juice.
(3) Quickly cooling the wort to 10 ℃, adding the cooled wort into a 50L fermentation tank, inoculating saccharomyces cerevisiae mud (4.0g/L) and pichia pastoris mud (0.2g/L), carrying out main fermentation for 7 days at 10 ℃, sealing the tank and boosting the pressure when the concentration of the wort in the main fermentation is reduced to 3.5-4.0 DEG P, and keeping the pressure at 0.11-0.13 MPa; carrying out after-fermentation, cooling the temperature of the fermentation tank to 4 ℃ within 24 hours, and storing for 7 days; and (4) after the fermentation is finished, reducing the content of diacetyl to 0.1mg/L, and filtering to obtain the beer.
EXAMPLE 2 beer of Mixed fermentation of aroma-producing Yeast and Saccharomyces cerevisiae
(1) Inoculating the saccharomyces cerevisiae and the pichia pastoris on the inclined plane into 12 DEG P whole wheat juice, performing shake culture at 28 ℃ for 24h, inoculating the mixture into a triangular flask filled with 50mL of whole wheat juice according to the inoculation amount of 5%, performing shake culture at 20 ℃ for 36h, transferring the mixture into 300mL of fresh 12 DEG P whole wheat juice, performing shake culture at 18 ℃/110rpm for 65h, centrifuging at 15 ℃/6000rpm for 10min, and collecting yeast paste.
(2) After being crushed into 20 kg, the malt is evenly mixed with water according to the material-water ratio of 1: 3. Placing in a saccharifying pot, and saccharifying by saccharifying process of keeping temperature at 45 deg.C for 30min, heating to 63 deg.C, keeping temperature for 60min, heating to 72 deg.C, keeping temperature for 10min, heating to 78 deg.C, and keeping temperature for 10 min; filtering the obtained mash, adding flos Lupuli (the addition amount of flos Lupuli is 1.6g/L, and adding into boiled wheat juice in batches), boiling wheat juice for 90min, removing precipitate in rotary sedimentation pot to obtain 12 ° P whole wheat juice.
(3) Quickly cooling the wort to 10 ℃, adding the cooled wort into a 50L fermentation tank, inoculating saccharomyces cerevisiae mud (3.8g/L) and pichia pastoris mud (0.4g/L), carrying out main fermentation for 7 days at 10 ℃, sealing the tank and boosting the pressure when the concentration of the wort subjected to the main fermentation is reduced to 3.5-4.0 DEG P, and keeping the pressure at 0.11-0.13 MPa; carrying out after-fermentation, cooling the temperature of the fermentation tank to 4 ℃ within 24 hours, and storing for 7 days; and (4) after the fermentation is finished, reducing the content of diacetyl to 0.1mg/L, and filtering to obtain the beer.
EXAMPLE 3 beer of Mixed fermentation of aroma-producing Yeast and Saccharomyces cerevisiae
(1) Inoculating the saccharomyces cerevisiae and the pichia pastoris on the inclined plane into 12 DEG P whole wheat juice, performing shake culture at 28 ℃ for 24h, inoculating the mixture into a triangular flask filled with 50mL of whole wheat juice according to the inoculation amount of 5%, performing shake culture at 20 ℃ for 36h, transferring the mixture into 300mL of fresh 12 DEG P whole wheat juice, performing shake culture at 18 ℃/120rpm for 68h, centrifuging at 15 ℃/6000rpm for 10min, and collecting yeast paste.
(2) After being crushed into 20 kg, the malt is evenly mixed with water according to the material-water ratio of 1: 3. Placing in a saccharifying pot, and saccharifying by saccharifying process of keeping temperature at 45 deg.C for 30min, heating to 63 deg.C, keeping temperature for 60min, heating to 72 deg.C, keeping temperature for 10min, heating to 78 deg.C, and keeping temperature for 10 min; filtering the obtained mash, adding flos Lupuli (the addition amount of flos Lupuli is 1.6g/L, and adding into boiled wheat juice in batches), boiling wheat juice for 90min, removing precipitate in rotary sedimentation pot to obtain 12 ° P whole wheat juice.
(3) Quickly cooling the wort to 10 ℃, adding the cooled wort into a 50L fermentation tank, inoculating saccharomyces cerevisiae mud (4.2g/L) and pichia pastoris mud (0.6g/L), carrying out main fermentation for 7 days at 10 ℃, sealing the tank and boosting the pressure when the concentration of the wort in the main fermentation is reduced to 3.5-4.0 DEG P, and keeping the pressure at 0.11-0.13 MPa; carrying out after-fermentation, cooling the temperature of the fermentation tank to 4 ℃ within 24 hours, and storing for 7 days; and (4) after the fermentation is finished, reducing the content of diacetyl to 0.1mg/L, and filtering to obtain the beer.
EXAMPLE 4 beer brewed conventionally (control)
(1) The activated yeast paste was prepared as described in example 1 (1).
(2) Whole wheat juice was obtained as described in example 1 (2).
(3) Rapidly cooling the wort to 10 deg.C, adding into 50L fermentation tank, inoculating Saccharomyces cerevisiae paste (4.0g/L), performing primary fermentation at 10 deg.C for 7 days, performing after-fermentation, cooling the fermentation tank to 4 deg.C within 24 hr, and storing for 7 days; filtering to obtain beer.
The checking method comprises the following steps:
and (3) determining the content of ethanol: the gas chromatograph method is used for measuring, when a sample enters a chromatographic column in the gas chromatograph, the ethanol is separated from other components due to different adsorption coefficients in gas and solid, the ethanol is identified by a hydrogen flame ionization detector, and the ethanol is compared with a standard sample, qualitative according to retention time and quantitative by an internal standard method.
Volatile flavor substance determination: the beer sample is injected into a sampling bottle to make the gas phase and the liquid phase reach an equilibrium state, the gas phase is extracted to a gas chromatograph equipped with an FID detector, and the internal standard sample is used for quantification.
The experimental method comprises the following steps:
(1) sample preparation: for the standard samples (isoamyl alcohol 40.15mg/L, isobutyl alcohol 7.1mg/L, n-propanol 8.15mg/L, ethyl acetate 7.18mg/L, isoamyl acetate 0.87mg/L, ethyl hexanoate 0.084mg/L, ethyl formate 0.075mg/L and isobutyl acetate 0.085mg/L), 5mL of each of beer samples stored at 0 ℃ for 12 hours or more was taken and transferred to 20mL sampling bottles, and 0.1mL of n-butanol was added as an internal standard substance and sealed with aluminum stoppers.
(2) The determination method comprises the following steps: sample inlet temperature: 250 ℃; detector temperature: 260 ℃; column temperature: keeping the temperature at 43 ℃ for 2 minutes, heating up at 10 ℃/minute, keeping the temperature at 90 ℃ for 1 minute, and heating up to 200 ℃ at 30 ℃/minute; carrier gas: nitrogen (10 mL/min); and recording the chromatogram, and comparing with a standard curve to obtain the content of each component in the sample.
In the beer produced in examples 1-2, the main physical and chemical indexes of the beer are shown in Table 1.
TABLE 1 physical and chemical indexes of mixed fermented beer
Note: esters include ethyl acetate, isoamyl acetate, ethyl hexanoate, ethyl formate and isobutyl hexanoate; the higher alcohols include isobutanol and isoamyl alcohol.
Claims (4)
1. A preparation method of beer by mixed fermentation of aroma-producing yeast and saccharomyces cerevisiae is characterized in that:
(a) pulverizing fructus Hordei Germinatus, saccharifying, filtering, boiling, adding flos Lupuli and precipitate, and making into 12 ° P whole wheat juice; respectively inoculating saccharomyces cerevisiae and aroma-producing yeast into 10-12 DEG P whole wheat juice for stepwise propagation, wherein the bacterial load reaches 5-10g/L, and respectively centrifuging to obtain yeast paste;
(b) rapidly cooling 12 ° P whole wheat juice to 10 deg.C, inoculating 3.0-6.0g/L Saccharomyces cerevisiae paste and 0.01-1.0 g/L aroma-producing yeast paste, performing main fermentation and after fermentation, and filtering to obtain beer;
the aroma-producing yeast is Pichia pastoris (Pichia fabianii);
the main fermentation is carried out for 7 days at the temperature of 8-12 ℃; storing for 5-7 days at 0-4 ℃;
the preparation method of the saccharomyces cerevisiae comprises the steps of inoculating saccharomyces cerevisiae on an inclined plane into 12 DEG P whole wheat juice, performing shake culture at 28 ℃ for 24 hours, inoculating the saccharomyces cerevisiae into a triangular flask filled with 50mL of whole wheat juice according to the inoculation amount of 5%, performing shake culture at 20 ℃ for 36 hours at 150rpm, transferring the whole wheat juice into 300mL of fresh 10-12 DEG P whole wheat juice, performing shake culture at 16 ℃/100rpm for 72 hours, centrifuging at 15 ℃/6000rpm for 10 minutes, and collecting yeast paste;
the preparation method of the aroma-producing yeast comprises the steps of inoculating the aroma-producing yeast on an inclined plane into 10-12 DEG P whole wheat juice, performing shake cultivation for 24 hours at 28 ℃, inoculating the aroma-producing yeast into a triangular flask filled with 50mL of whole wheat juice according to the inoculation amount of 5%, performing shake cultivation for 36 hours at 20 ℃ and 150rpm, transferring the aroma-producing yeast into 300mL of fresh 10-12 DEG P whole wheat juice, performing shake cultivation for 72 hours at 16 ℃/100rpm, centrifuging for 10min at 15 ℃/6000rpm, and collecting yeast paste;
the main fermentation is that when the wort concentration is reduced to 3.5-4.0 DEG P, the tank is sealed and the pressure is increased, and the pressure is kept at 0.11-0.13 MPa; and (3) performing after-fermentation, reducing the temperature of the fermentation tank to 0-4 ℃ within 24 hours, and storing for 5-7 days, wherein the fermentation end time is that the content of diacetyl is reduced to 0.1 mg/L.
2. The method for preparing beer by mixed fermentation of aroma-producing yeast and saccharomyces cerevisiae according to claim 1, wherein the method comprises the following steps: and inoculating the saccharomyces cerevisiae mud into 10-12-DEG P whole wheat juice according to the inoculation amount of 4.0g/L for fermentation.
3. The method for preparing beer by mixed fermentation of aroma-producing yeast and saccharomyces cerevisiae according to claim 1, wherein the method comprises the following steps: and inoculating the pichia pastoris sludge into 10-12-DEG P whole wheat juice according to the inoculation amount of 0.2g/L for fermentation.
4. The method for preparing beer by mixed fermentation of aroma-producing yeast and saccharomyces cerevisiae according to claim 1, wherein the method comprises the following steps: the preparation method of the whole wheat juice comprises the following steps: crushing malt, and uniformly mixing the crushed malt with water according to the material-water ratio of 1: 3; placing in a saccharifying pot, and saccharifying by saccharifying process of keeping temperature at 45 deg.C for 30min, heating to 63 deg.C, keeping temperature for 60min, heating to 72 deg.C, keeping temperature for 10min, heating to 78 deg.C, and keeping temperature for 10 min; and (3) after the obtained mash passes through a filter tank, adding hops with the addition of 1.5g/L, adding into the boiled wheat juice in batches, boiling the wheat juice for 90min, and removing precipitates in a rotary sedimentation pot to obtain the 12-DEG P whole wheat juice.
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| CN108410605A (en) * | 2018-05-11 | 2018-08-17 | 大连工业大学 | A kind of preparation method of the refreshing beer of acid |
| CN110628538A (en) * | 2019-10-23 | 2019-12-31 | 北部湾大学 | Method for making high-alcohol-content sweet beer with flavor of noble fermented bean curd wine |
| CN111117820A (en) * | 2019-12-31 | 2020-05-08 | 云南鸿禾酒业有限公司 | Method for preparing beer with pineapple flavor by using pichia pastoris strain |
| CN113416659B (en) * | 2021-07-01 | 2022-12-23 | 中国农业大学 | Application of candida glabrata capable of producing fruity flavor and commercial yeast in beer brewing through sequential inoculation fermentation |
| CN114540138B (en) * | 2022-03-25 | 2023-09-22 | 北京燕京啤酒股份有限公司 | Method for producing novel fruity acid beer by double yeast mixed fermentation wort |
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