CN112760257B - Lactobacillus plantarum strain YC21 and application thereof - Google Patents
Lactobacillus plantarum strain YC21 and application thereof Download PDFInfo
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- CN112760257B CN112760257B CN202110107456.8A CN202110107456A CN112760257B CN 112760257 B CN112760257 B CN 112760257B CN 202110107456 A CN202110107456 A CN 202110107456A CN 112760257 B CN112760257 B CN 112760257B
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12G—WINE; PREPARATION THEREOF; ALCOHOLIC BEVERAGES; PREPARATION OF ALCOHOLIC BEVERAGES NOT PROVIDED FOR IN SUBCLASSES C12C OR C12H
- C12G1/00—Preparation of wine or sparkling wine
- C12G1/02—Preparation of must from grapes; Must treatment and fermentation
- C12G1/0203—Preparation of must from grapes; Must treatment and fermentation by microbiological or enzymatic treatment
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
Abstract
The invention relates to a Lactobacillus plantarum strain YC21 and application thereof, belonging to the technical field of microorganisms. The preservation number of the Lactobacillus plantarum strain YC21 is CCTCC M2021080. The yield of ethyl acetate generated by fermenting grape juice with the strain is up to 179.88 mu g/L, and the yield of 1-pentanol is up to 167.49 mu g/L.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and particularly relates to a Lactobacillus plantarum strain YC21 and application thereof.
Background
Lactobacillus plantarum (Lactobacillus plantarum) is one of lactic acid bacteria, the optimal growth temperature is 30-35, the Lactobacillus plantarum is anaerobic or facultative anaerobic, the strain is a straight or bent rod, the strain is single or sometimes paired or chained, the optimal pH is about 6.5, and the Lactobacillus plantarum belongs to homofermentation lactic acid bacteria. The biological characteristics are as follows: the number of the terminal rectus is usually 0.9-1.2 vtm × 3.0-8.0 μm, single, paired or short chain. Usually lacking flagella, but able to move. Gram positive, no sporulation. Facultative anaerobe, surface colony diameter about 3mm, convex, round, smooth surface, fine, white, occasionally light yellow or dark yellow. Belongs to chemoheterotrophic bacteria, needs a nutrient-rich culture medium for growth, needs calcium pantothenate and nicotinic acid, but does not need thiamine, pyridoxal or pyridoxamine, folic acid and vitamin B12. Can ferment pentose or gluconate, and the final product contains lactic acid more than 85%. Nitrate is generally not reduced, gelatin is not liquefied, and both catalase and oxidase are negative. Can produce DL-lactic acid, has the activity of fructose-1, 6-diphosphate aldolase and hexose monophosphate pathways, can grow in gluconate, and produces C02. Fermentation of 1 molecule of ribose or other pentose sugar produces 1 molecule of lactic acid and 1 molecule of acetic acid. Can grow at 15 ℃, and the optimal growth temperature is 30-35 ℃ generally.
The industrial application of lactobacillus plantarum reported at present is common in food fermentation, such as fermented milk production, and also can be applied to heavy metal degradation, and the lactobacillus produced in the propagation process can be used as a natural preservative, so the lactobacillus can also be used in the production and cultivation fields.
However, the lactobacillus plantarum strain capable of producing aroma substances such as 2-heptanol and ethyl acetate with high yield has not been reported in the field.
Disclosure of Invention
Based on the blank in the field, the lactobacillus plantarum strain capable of fermenting and highly producing aroma substances such as 2-heptanol, ethyl acetate and the like by taking grape juice as a substrate is obtained through screening.
The technical scheme of the invention is as follows:
lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080.
Application of Lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080 in producing aroma substances.
The aromatic substance is selected from the group consisting of 2-heptanol, 1-pentanol, isobutyraldehyde, isovaleraldehyde, ethyl acetate, 4-methyl-1-pentanol, n-octanol, n-heptanol, 1-octen-3-ol, linalool, damascenone, hexyl acetate, isoamyl acetate, ethyl decanoate and ethyl hexanoate.
The aroma substances are selected from: 2-heptanol, 1-pentanol, isobutyraldehyde, isovaleraldehyde and ethyl acetate.
The Lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080 is applied to brewing.
The wine-making leavening agent is characterized in that fermentation active substances of the wine-making leavening agent comprise Lactobacillus plantarum strain YC21 with the collection number of CCTCC M2021080.
The fermentation active substance also comprises saccharomyces cerevisiae.
The brewing leaven is a microbial inoculum;
preferably, the brewing leavening agent also comprises auxiliary materials acceptable for microbial inoculum;
preferably, the auxiliary material acceptable by the microbial inoculum comprises a culture medium substance of the microbial inoculum; media materials for the bacteria include, but are not limited to: starch, sucrose, peptone and water.
A method for brewing wine is characterized in that Lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080 is adopted for fermentation brewing.
The fermentation refers to the fermentation by using the Lactobacillus plantarum strain YC21 and using grape or grape juice as a substrate;
preferably, the Lactobacillus plantarum strain YC21 is added to the substrate at a concentration of 10 6 -10 7 CFU/ml, preferably 10 6 CFU/ml。
The invention obtains a strain through screening, and the aroma substance component identification experiment shows that the strain can produce a series of aroma substances such as ethyl acetate, 4-methyl-1-pentanol and the like with high yield, and compared with a commercial saccharomyces cerevisiae strain F33, the strain can also produce the aroma substances which can not be generated by F33, such as: 2-heptanol, 1-pentanol, isobutyraldehyde, isovaleraldehyde and ethyl acetate, wherein the yield of the ethyl acetate is 179.88 mu g/L, and the yield of the 1-pentanol is 167.49 mu g/L. The strain is identified to be a Lactobacillus plantarum strain, and the applicant names the strain YC21 and sends the strain for preservation.
The Lactobacillus plantarum strain YC21 of the present invention has the following preservation information:
naming: YC21
And (4) classification name: lactobacillus plantarum
The name of Latin is: lactobacillus plantarum
The preservation number is as follows: CCTCC M2021080
The preservation organization: china center for type culture Collection
And (4) storage address: eight-way 299 # in Wuchang area of Wuhan city, Hubei province
The preservation date is as follows: 1 month and 15 days 2021
Detailed Description
The following detailed description of the present invention will be made with reference to specific examples, but the scope of the present invention is not limited thereto.
Sources and documentations of biological materials
The sources and origins of the grape varieties used in experimental example 1 are shown in table 1 below:
TABLE 1
| Grape variety | Producing area | Source |
| Cabernet Sauvignon (Cabernet Sauvignon) bead | Ningxia Helan mountain east foot producing area | Collecting |
| Meile wine | Ningxia Helan mountain east foot producing area | Collecting |
| Xila | Ningxia Helan mountain east foot producing area | Collecting |
| Snake dragon bead | Ningxia Helan mountain east foot producing area | Collection of |
| Xiaoweierduo | Ningxia Helan mountain eastern foot producing area | Collection of |
| Noble incense | Ningxia Helan mountain east foot producing area | Collecting |
| Weidai Er | Ningxia Helan mountain eastern foot producing area | Collecting |
| Rivastigmine | Ningxia Helan mountain east foot producing area | Collection of |
| Chardonnay | Ningxia Helan mountain east foot producing area | Collecting |
The grape varieties in table 1 above are all known and public grape varieties, and are also commercially available.
The grape variety used in experimental example 2 was a vicat ice grape, purchased from ningxia burger drunk american border winery ltd;
commercial strain Saccharomyces cerevisiae F33 was purchased from Lafford (Laffort) France.
Group 1 example, Strain YC21 of the present invention
The embodiment of the group provides a Lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080.
EXAMPLE 2 group, use of the Strain YC21 according to the invention
The embodiment of the group provides application of Lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080 in producing aroma substances.
In some embodiments, the aroma substance is selected from the group consisting of 2-heptanol, 1-pentanol, isobutyraldehyde, isovaleraldehyde, ethyl acetate, 4-methyl-1-pentanol, n-octanol, n-heptanol, 1-octen-3-ol, linalool, damascenone, hexyl acetate, isoamyl acetate, ethyl decanoate, ethyl hexanoate.
In other embodiments, the aroma is selected from the group consisting of: 2-heptanol, 1-pentanol, isobutyraldehyde, isovaleraldehyde and ethyl acetate.
Group 3 examples, brewing applications of Strain YC21 of the invention
The embodiment of the group provides application of Lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080 in brewing.
Group 4 examples of the fermentation agents for brewing wine of the present invention
The embodiment of the group provides a wine brewing leavening agent. All embodiments of this group share the following common features: the fermentation active substances of the brewing leaven comprise Lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080.
In a further embodiment, the fermentation active further comprises saccharomyces cerevisiae. The YC21 strain of the present invention can be used in combination with Saccharomyces cerevisiae, which is currently available in the art, for fermenting Saccharomyces cerevisiae, by a person skilled in the art according to the teachings of the present invention.
In a specific embodiment, the brewing leavening agent is a microbial inoculum;
in a preferred embodiment, the brewing leavening agent also comprises auxiliary materials acceptable for microbial inoculum;
in other preferred embodiments, the microbial inoculum acceptable adjuvant comprises a medium material of the microbial inoculum; media materials for the bacteria include, but are not limited to: starch, sucrose, peptone and water.
Example 5 set of methods for brewing wine according to the invention
The present set of embodiments provide a wine brewing method. All embodiments of this group share the following common features: fermenting and brewing by adopting Lactobacillus plantarum YC21 with the preservation number of CCTCC M2021080.
In a specific embodiment, the fermentation refers to fermentation using the Lactobacillus plantarum strain YC21 with grape or grape juice as substrate.
In a preferred embodiment, the Lactobacillus plantarum strain YC21 is added to the substrate in a concentration of 10 6 -10 7 CFU/ml, preferably 10 6 CFU/ml。
Experimental example 1 screening Process of Lactobacillus plantarum Strain of the present invention
Collecting different varieties of wine grapes (shown in table 1) at different production places of eastern foot of Ningxia Helan mountain, removing rotten, mildewed and damaged fruit grains and sundries, weighing 10.0g of uniform and complete grape fruit grains, adding the uniform and complete grape fruit grains into 90mL of sterile YPD liquid culture medium, oscillating for 10min to prepare bacterial suspension, coating the bacterial suspension with proper dilution on YPD solid culture medium added with 100mg/L chloramphenicol by adopting a gradient dilution method, culturing at 28 ℃ for 2 d-3 d, and carrying out streak purification on the YPD solid culture medium for multiple times according to colony morphology to obtain a purified strain. After the purified strain is activated in YPD culture medium for 24h, 1mL of bacterial liquid is taken to extract yeast genome according to instructions of a Biospin fungal genome DNA extraction kit, primers NL-1(5'-GCATATCAATAAGCGGAGGAAAAG-3') and NL-4(5'-GGTCCGTGTTTCAAGACGG-3') are used for PCR amplification, a gene sequence of a 26S rDNA fragment is obtained through sequencing, known standard strain information with high homology is obtained through BLAST comparison of NCBI, species identification is carried out on the strain with similarity of more than 99%, and finally strain species information is obtained.
In this experimental example, a total of 18 Lactobacillus plantarum strains were obtained. 18 strains of Lactobacillus plantarum were each added at an initial concentration of 10 6 The CFU/mL is inoculated into sterilized Vidal grape juice (100 ℃, 10min), fermented for 18 days at 18 ℃ +/-2 ℃, the aroma content is measured, and finally the Lactobacillus plantarum strain YC21 with the best aroma production effect is obtained and is preserved, and the preservation information is as follows:
naming: YC21
And (4) classification name: lactobacillus plantarum
The name of Latin is: lactobacillus plantarum
The preservation number is as follows: CCTCC M2021080
The preservation organization: china center for type culture Collection
The preservation date is as follows: year 2021, month 15.
Experimental example 2, data on aroma-producing substances produced by Lactobacillus plantarum strain YC21 according to the present invention
Grape variety: vidal iced grape
The grape juice production process comprises the following steps: pressing the harvested ice grape with ice by air bag press, and adding sulfur dioxide (50mg/L K) 2 S 2 O 5 ) And 20mg/L of pectinase (more than or equal to 500U/mg), inhibits bacterial diseases and improves the juice yield.
The grape juice fermentation conditions are as follows: the steeped grape juice is respectively added at an initial concentration of 10 6 And (3) inoculating the CFU/mL into F33 saccharomyces cerevisiae and Lactobacillus plantarum strain YC21, fermenting at the temperature of 18 +/-2 ℃, and stopping fermentation when the weight loss of the grape juice is not changed for three consecutive days. All wine samples were centrifuged at 7500rpm for 8 minutes and the supernatant was stored at 4 ℃.
The detection method of each aroma substance comprises the following steps: headspace-solid phase microextraction-gas mass spectrometry (HS-SPME-GC/MS) was used. An 8mL sample of wine was accurately weighed into a headspace bottle containing 1.5g NaCl, while 394.08. mu.g/L4-methyl-1-pentanol (internal standard) was capped and sealed. Inserting CAR/DVB/PDMS extraction fiber, adsorbing at 45 deg.C for 30min, desorbing at 250 deg.C for 3min, and performing GC-MS analysis. A chromatographic column: InertCap WAX polar chromatography column (60m × 0.25mm, 0.25 μm); the temperature rising procedure is as follows: maintaining at 40 deg.C for 5min, heating to 120 deg.C at 3 deg.C/min, heating to 230 deg.C at 8 deg.C/min, and maintaining for 10 min; the flow rate of the carrier gas (He) was 0.8mL/min, and it was not branched. Electron bombardment ion source; electron energy 70 eV; the transmission line temperature was 275 ℃; the ion source temperature is 230 ℃; the activation voltage is 1.5V; the filament flow is 0.25 mA; the mass scanning range m/z is 33-450. Compound quantitative analysis was performed using external standard quantitation method.
Taking the same batch of grape juice produced by the same variety of grapes as a substrate, respectively carrying out fermentation brewing treatment on the equal amount of grape juice by using a commercial strain Saccharomyces cerevisiae F33 and a Lactobacillus plantarum strain YC21 of the invention under the same fermentation conditions, and detecting the content of each aroma substance (unit: mu g/L, meaning: the content of the aroma substance in each liter of grape wine) by a headspace-solid phase microextraction-gas mass spectrometry method to obtain the following table 2:
TABLE 2
The aroma threshold in table 1 above refers to the lower limit of the minimum concentration at which a human can smell the substance.
SEQUENCE LISTING
<110> Ningxia agricultural product quality standard and detection technology research institute (Ningxia agricultural product quality monitoring center)
<120> Lactobacillus plantarum strain YC21 for high-yield aroma substance and application thereof
<130> P201027/NNZ
<160> 2
<170> PatentIn version 3.5
<210> 1
<211> 24
<212> DNA
<213> Artificial Sequence
<220>
<223> primer NL-1
<400> 1
gcatatcaat aagcggagga aaag 24
<210> 2
<211> 19
<212> DNA
<213> Artificial Sequence
<220>
<223> primer NL-4
<400> 2
ggtccgtgtt tcaagacgg 19
Claims (14)
1. A Lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080.
2. Application of Lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080 in producing aroma substances.
3. Use according to claim 2, characterized in that the aroma substances are selected from the group consisting of 2-heptanol, 1-pentanol, isobutyraldehyde, isovaleraldehyde, ethyl acetate, 4-methyl-1-pentanol, n-octanol, n-heptanol, 1-octen-3-ol, linalool, damascenone, hexyl acetate, isoamyl acetate, ethyl decanoate, ethyl hexanoate.
4. Use according to claim 2 or 3, wherein the aroma substances are selected from: 2-heptanol, 1-pentanol, isobutyraldehyde, isovaleraldehyde and ethyl acetate.
5. The lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080 is applied to brewing.
6. A wine brewing leaven is characterized in that fermentation active substances comprise a lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080.
7. The leavening agent of claim 6, wherein the fermentation active further comprises Saccharomyces cerevisiae.
8. The wine brewing starter culture according to claim 6 or 7, wherein the wine brewing starter culture is a microbial inoculum.
9. The wine brewing starter culture according to claim 6 or 7, wherein the wine brewing starter culture further comprises auxiliary materials acceptable for microbial inoculum.
10. The wine brewing starter culture according to claim 9, wherein the auxiliary materials acceptable for the microbial inoculum comprise a culture medium material of the microbial inoculum; the culture medium material of the bacteria comprises: starch, sucrose, peptone and water.
11. A method for brewing wine is characterized in that lactobacillus plantarum strain YC21 with the preservation number of CCTCC M2021080 is adopted for fermentation brewing.
12. A wine brewing process according to claim 11, wherein said fermentation is carried out using said lactobacillus plantarum strain YC21, grape or grape juice as substrate.
13. A wine brewing method according to claim 12, wherein lactobacillus plantarum strain YC21 was added to the substrate at a concentration of 10 6 -10 7 CFU/ml。
14. A wine brewing process according to claim 13, wherein lactobacillus plantarum strain YC21 is added to the substrate at a concentration of 10 6 CFU/ml。
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| CN115191485A (en) * | 2022-07-04 | 2022-10-18 | 微康益生菌(苏州)股份有限公司 | Application of lactobacillus plantarum Lp90 in preparation of high-yield ethyl acetate product and yogurt starter culture containing lactobacillus plantarum Lp90 |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104388329A (en) * | 2014-09-15 | 2015-03-04 | 中国食品发酵工业研究院 | Application of lactobacillus plantarum in degradation of biogenic amines in wine making |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104388329A (en) * | 2014-09-15 | 2015-03-04 | 中国食品发酵工业研究院 | Application of lactobacillus plantarum in degradation of biogenic amines in wine making |
Non-Patent Citations (2)
| Title |
|---|
| Characterization of γ-aminobutyric acid (GABA)-producing Saccharomyces cerevisiae and coculture with Lactobacillus plantarum for mulberry beverage brewing;Qing Zhang;《J Biosci Bioeng》;20191031;第129卷(第4期);第447-453页 * |
| 葡萄酒中植物乳杆菌苹果酸-乳酸发酵潜能评价;卜潇;《中国农业科学》;20171231;第50卷(第5期);摘要，表2 * |
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