Phytobacterium plantarum and application thereof

Abstract

The invention relates to the field of biological fermentation, in particular to plant lactobacillus and application thereof. The invention provides lactobacillus plantarum, and the preservation number of the lactobacillus plantarum is CGMCC (China General Microbiological Culture Collection Center) No.33376. The lactobacillus plantarum has the advantages that the preservation number is CGMCC No.33376; the plant lactobacillus C10 provided by the invention has the advantages that the plant lactobacillus C10 can tolerate a wine environment with relatively high stress and has high beta-glycosidase activity, and can be applied to grape wine malolactic fermentation, so that the aroma composition of cabernet sauvignon grape wine is improved, the fruit aroma of the grape wine is improved, and the sensory attribute of the grape wine is further improved.

Description

Technical Field

[0001] The invention relates to the field of biological fermentation, in particular to lactobacillus plantarum and applications thereof. Background Art

[0002] Winemaking involves the joint action of multiple microorganisms, the two most important of which are yeast and lactic acid bacteria. First, yeast-led alcoholic fermentation converts sugar into alcohol to produce wine. After alcoholic fermentation, lactic acid bacteria lead malolactic fermentation, converting L-malic acid in wine into L-lactic acid, which is mostly carried out in red wine and some high-acid white wines. After malolactic fermentation, the acidity of the wine decreases and the biological stability of the wine body increases. At the same time, the enzyme metabolism of lactic acid bacteria changes the composition of the aroma compounds of the wine.

[0003] Lactobacillus plantarum is a common wine lactic acid bacterium that can grow in high pH wine and lead malic acid-lactic acid fermentation. Many studies have shown that Lactobacillus plantarum can also survive in the wine environment and complete malic acid-lactic acid fermentation. It consumes sugars without producing acetic acid during fermentation and has the characteristics of homotypic fermentation, which makes it have good application prospects in wine fermentation. Lactobacillus plantarum also has complex enzyme system characteristics, such as β-glycosidase, esterase, and volatile phenol-related enzyme system, which may have a positive effect on the sensory characteristics of wine.

[0004] At present, many single bacteria or compound Lactobacillus plantarum fermentation agents have been commercialized abroad. These fermentation agents are widely used in many fields such as food, beverages and fermentation engineering, especially in the production of dairy products. However, China is still relatively backward in the research and development of lactic acid bacteria preparations, and the application research of Lactobacillus plantarum is mostly concentrated in the traditional dairy industry. With the continuous deepening of research on wine lactic acid bacteria, Lactobacillus plantarum, as a kind of lactic acid fermentation agent with good malic acid and lactic acid fermentation ability and complex enzyme system characteristics, has also received attention for its potential in wine. Summary of the invention

[0005] In view of this, the present invention provides Lactobacillus plantarum and its application. The Lactobacillus plantarum C10 provided by the present invention has tolerance to a highly stressful wine environment and high β-glycosidase activity, and can be applied in wine malolactic fermentation to improve the aroma composition of Cabernet Sauvignon wine, enhance the fruity aroma of wine, and thus enhance the sensory attributes of wine.

[0006] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0007] The invention provides Lactobacillus plantarum, and its preservation number is CGMCC No.33376.

[0008] The invention also provides a mixed strain, comprising: the above-mentioned Lactobacillus plantarum and other strains.

[0009] In some embodiments of the present invention, in the above-mentioned mixed strains, the other strains include: lactic acid bacteria.

[0010] In some embodiments of the present invention, in the above-mentioned mixed strain, the lactic acid bacteria is: Lalvin VP41.

[0011] The present invention also provides a microbial agent, comprising: the above-mentioned Lactobacillus plantarum and / or the above-mentioned mixed strain.

[0012] The present invention also provides application of the Lactobacillus plantarum, the mixed strain and / or the microbial agent in fruit wine fermentation.

[0013] In some embodiments of the present invention, in the above application, the fruit wine includes: grape wine.

[0014] In some embodiments of the present invention, in the above application, the wine includes: Cabernet Sauvignon wine.

[0015] The present invention also provides the use of the Lactobacillus plantarum, the mixed strain and / or the microbial agent in improving the aroma and / or taste of fruit wine.

[0016] In some embodiments of the present invention, in the above application, the aroma includes: sweet aroma and / or fruity aroma.

[0017] In some embodiments of the present invention, in the above application, improving the taste of fruit wine comprises:

[0018] (I), reducing the content of reducing sugars in fruit wine; and / or

[0019] (II), reducing the alcohol content of the wine; and / or

[0020] (III) reducing the content of L-malic acid in fruit wine;

[0021] The method of improving the aroma of fruit wine comprises increasing the content of one or more of alcohol substances, ester substances and fatty acid substances.

[0022] In some embodiments of the present invention, in the above application, the fruit wine includes: grape wine.

[0023] In some embodiments of the present invention, in the above application, the wine includes: Cabernet Sauvignon wine.

[0024] In some embodiments of the present invention, in the above application, the alcohol substance includes: one or more of 1-propanol, isobutanol, 1-butanol, isopentanol, phenylethyl alcohol, 2,3-butanediol, linalool, and citronellol.

[0025] In some embodiments of the present invention, in the above application, the ester substance includes: one or more of ethyl lactate, ethyl 2-hydroxy-4-methylvalerate, ethyl isopentyl succinate, ethyl palmitate, diethyl succinate, isopentyl lactate, acetate, fatty acid ethyl esters, isobutyl acetate, isopentyl acetate, hexyl acetate, phenylethyl acetate, ethyl acetate, ethyl isobutyrate, ethyl butyrate, ethyl 2-methylbutyrate, ethyl 3-methylbutyrate and ethyl hexanoate.

[0026] In some embodiments of the present invention, in the above application, the fatty acid substance includes: one or more of butyric acid, isovaleric acid, caproic acid and octanoic acid.

[0027] The present invention also provides a method for fermenting fruit wine, which comprises mixing the Lactobacillus plantarum, the mixed strain and / or the microbial agent with raw materials for fermentation to obtain the fruit wine.

[0028] The beneficial effects of the present invention include:

[0029] (1) Fermentation characteristics. Lactobacillus plantarum C10 can tolerate a high alcohol content and low pH simulated wine environment and also has a high β-glycosidase activity, which indicates that it has a high tolerance and the potential to change the aroma of wine during the malolactic fermentation of wine.

[0030] (2) Improve aroma. Compared with commercial lactic acid bacteria, Lactobacillus plantarum C10 inoculation and fermentation greatly increases the content of higher alcohols and esters, which improves the aroma quality of wine after malolactic fermentation to a certain extent. Mixed inoculation and fermentation of commercial lactic acid bacteria and Lactobacillus plantarum C10 can increase the content of acetate in wine. Whether Lactobacillus plantarum C10 is used alone or in combination to initiate malolactic fermentation, it can contribute rich fruity aroma characteristics to wine.

[0031] (3) Sensory flavor. Lactobacillus plantarum C10 has good strain activity in Cabernet Sauvignon wine and can successfully complete malolactic fermentation, reducing acidity and thus softening the taste. Lactobacillus plantarum C10 single-strain fermentation can increase the sensory attributes of red berries in wine, and Lactobacillus plantarum C10 and mixed-strain fermentation can increase the sensory attributes of fresh fruity aroma in wine.

[0032] In summary, the Lactobacillus plantarum C10 provided by the present invention can be applied in the actual wine fermentation process to improve the flavor and sensory attributes of wine, and provide a basis for the malolactic acid fermentation agent of Lactobacillus plantarum for Chinese native wine.

[0033] Biological Deposit Description

[0034] Biological material: C10; classification name: Lactiplantibacillus plantarum; deposited in the General Microbiology Center of China Culture Collection Administration on January 10, 2025; address: Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing; deposit number: CGMCC No.33376. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art are briefly introduced below.

[0036] Figure 1 The colony morphology of Lactobacillus plantarum C10 of the present invention on MRS solid culture medium is shown;

[0037] Figure 2 The strain activity of Lactobacillus plantarum C10 and commercial lactic acid bacteria VP41 in a simulated wine environment is shown;

[0038] Figure 3 The β-glycosidase activities of Lactobacillus plantarum C10 and commercial lactic acid bacteria VP41 are shown;

[0039] Figure 4 The figures show the activity and L-malic acid consumption of Lactobacillus plantarum C10, commercial lactic acid bacteria VP41 and their mixed fermentation strains; wherein: a shows the dynamic consumption of L-malic acid; b shows the dynamic changes of strain activity;

[0040] Figure 5 The results of sensory evaluation of wines after malolactic fermentation of Lactobacillus plantarum C10, commercial lactic acid bacteria VP41 and their mixed fermentation in wine are shown. DETAILED DESCRIPTION

[0041] The invention discloses plant lactobacillus and application thereof.

[0042] It should be understood that the expression "one or more of..." includes each of the items recited after the expression individually and in various different combinations of two or more of the recited items, unless otherwise understood from the context and usage. The expression "and / or" in combination with three or more recited items should be understood to have the same meaning, unless otherwise understood from the context.

[0043] The use of the terms "comprising", "having" or "containing", including their grammatical synonyms, should generally be understood as open and non-restrictive, for example not excluding other unrecited elements or steps, unless otherwise specifically stated or otherwise understood from the context.

[0044] It should be understood that the order of steps or the order in which certain actions are performed is not important as long as the present invention remains operable. In addition, two or more steps or actions may be performed simultaneously.

[0045] The use of any and all examples or exemplary language, such as "for example" or "including", herein is intended only to better illustrate the invention and does not limit the scope of the invention unless otherwise claimed. No language in this specification should be construed as indicating that any non-claimed element is essential to the practice of the invention.

[0046] In addition, the numerical ranges and parameters used to define the present invention are approximate values, and the relevant values ​​in the specific embodiments have been presented as accurately as possible. However, any numerical value inherently inevitably contains standard deviations due to individual test methods. Therefore, unless otherwise expressly stated, it should be understood that all ranges, quantities, values ​​and percentages used in this disclosure are modified by "about". Here, "about" generally means that the actual value is within plus or minus 10%, 5%, 1% or 0.5% of a specific value or range.

[0047] The invention provides a Lactobacillus plantarum C10, which is deposited in the General Microbiological Center of China Microbiological Culture Collection Administration, and the deposit number is CGMCC No.33376.

[0048] The Lactobacillus plantarum C10 of the invention is separated from wine produced by natural apple-lactation fermentation and is identified as Lactobacillus plantarum by adopting a 16S rDNA genome sequencing method.

[0049] The colony characteristics of the plant lactobacillus C10 of the present invention are round, 1-2 mm in diameter, white and shiny, opaque, smooth and waxy in surface, and protruding on the surface of the solid culture medium with smooth edges.

[0050] The plant lactobacillus C10 can maintain a certain growth activity in simulated wine with an alcohol content of 14% (v / v) and a pH of 3.4.

[0051] The plant lactobacillus C10 has strong beta-glycosidase activity.

[0052] The present invention also provides an application of Lactobacillus plantarum C10 in improving the aroma of wine.

[0053] The wine is Cabernet Sauvignon wine; it is fermented by single bacteria Lactobacillus plantarum C10, and mixed bacteria fermentation of Lactobacillus plantarum C10 and commercial lactic acid bacteria;

[0054] The applications can change the aroma composition of Cabernet Sauvignon wine and improve the aroma and sensory flavor of Cabernet Sauvignon wine.

[0055] In this application, Lactobacillus plantarum C10 can significantly increase the content of higher alcohols and ester aroma substances in Cabernet Sauvignon wine, and increase the sensory flavor of fresh fruity aroma; mixed inoculation of Lactobacillus plantarum C10 and commercial lactic acid bacteria significantly increases the content of ester compounds, especially acetate and fatty acid ethyl esters, and increases the sensory flavor of red berries in the wine.

[0056] In Examples 1 to 3 and the Effect Examples of the present invention, all the raw materials and reagents used can be purchased from the market.

[0057] The present invention will be further described below in conjunction with embodiments:

[0058] Example 1 Isolation, purification and identification of Lactobacillus plantarum C10 strain

[0059] For the wine provided by Huailai, Hebei, samples were taken in the middle of natural fermentation and diluted and spread on MRS agar medium (with 100 mg / L cycloheximide), cultured in a 37°C constant temperature incubator for 48 to 72 hours, and colonies with different morphologies were picked and streaked for pure culture to obtain isolated strains. The identification of lactic acid bacteria was carried out by 16S rDNA genome sequencing, and the strain genomic DNA extraction method was as follows: a single colony obtained by streaking pure culture was picked and centrifuged at 12000r / min for 1 minute, the supernatant was discarded, and then the strain genomic DNA was extracted using a bacterial DNA genome extraction kit according to the instructions. The 16S rDNA gene PCR amplification method is as follows: the extracted strain DNA is amplified in a PCR amplification instrument using forward primer 27F (5'-AGAGTTTGATCCTG GCTCAG-3' (as shown in SEQ ID NO: 1)) and reverse primer 1492R (5'-GGTTACCT TGTTACGACTT-3' (as shown in SEQ ID NO: 2)) to the 16S rDNA gene region of the strain DNA. The PCR amplification system and program are shown in Table 1. The amplified product is detected by 1% agarose gel electrophoresis and gel imaging instrument, and a band of about 1500bp is observed, indicating that the PCR is successful, and it is sent to Shanghai Biotech for unidirectional sequencing. The BLAST program is used to compare the obtained strain 16S rDNA gene sequence in the GenBank database with the known bacterial gene sequence. The isolated strain has a homology similarity with plant lactobacillus greater than 99%, and is confirmed to be plant lactobacillus, named Lactiplantibacillus plantarum C10. Colony characteristics of Lactobacillus plantarum C10 Figure 1 As shown, it is round, 1 to 2 mm in diameter, white and shiny, opaque, with a smooth and waxy surface. It is raised on the surface of the solid culture medium and has smooth edges.

[0060] The successfully identified Lactobacillus plantarum C10 was streaked and cultured to obtain a single colony, which was cultured in MRS broth for about 24 hours until the mid-logarithmic phase. 50% glycerol was used as a cryoprotectant, and the culture medium was mixed with 50% glycerol (1:1) and added to a preservation tube, which was stored at -80°C.

[0061] The plant lactobacillus C10 obtained by the present invention was sent to the General Microbiological Center of China Microbiological Culture Collection Administration for preservation, and the preservation number was CGMCC No.33376.

[0062] Table 1 PCR amplification system and procedure

[0063]

[0064] Example 2 Preliminary detection experiment of the characteristics of Lactobacillus plantarum C10 strain

[0065] The plant lactobacillus C10 of the present invention can tolerate a simulated wine environment with high alcohol content and low pH, and has high β-glycosidase activity. The control group strain of this experiment is the commercial lactic acid bacteria VP41 already available in the laboratory.

[0066] 1. Activation of Lactobacillus plantarum C10: Streak the bacterial solution preserved with 50% glycerol at -80°C on MRS agar medium, incubate at 37°C for 48h to obtain a single colony, pick a single colony with an inoculating loop and place it in MRS broth medium, incubate at 37°C for 12h to obtain an inoculated bacterial solution, and take the bacterial solution for subculture to the logarithmic growth phase for preparation of inoculation.

[0067] 2. Screening of tolerance to brewing conditions: After the strains were activated, they were each 7 The inoculum of 50 cfu / mL was inoculated into 100 mL of simulated wine solution containing 16% alcohol and pH 3.4, and the viable count was determined at 30°C for 20 days to determine the strain's tolerance in the simulated wine environment. The experiment was repeated three times.

[0068] The simulated wine was configured as follows: alcohol content 16% (v / v), 5 g / L tartaric acid, 3.5 g / L L-malic acid, 0.6 g / L acetic acid, 2 g / L D-glucose, 2 g / L D-fructose, 0.2 g / L sodium chloride, 1 g / L ammonium sulfate, 2 g / L dipotassium hydrogen phosphate, 0.056 g / L magnesium sulfate, 0.05 g / L manganese sulfate, and 2 g / L yeast extract.

[0069] 3. β-Glucosidase activity:

[0070] Bacterial cells were harvested from 1 mL of culture, washed with 150 mM NaCl, resuspended in 500 μL of 125 mM citrate-phosphate buffer (pH 5.0) containing 5 mM p-nitrophenyl-β-D-pyranoglucoside (p-NPG), and incubated at 25°C for two hours. The sample was diluted with 1 mL of 0.2 M carbonate buffer (pH 10.2), clarified by centrifugation, and the absorbance of the supernatant at a wavelength of 400 nm was measured. Based on this measurement, the concentration of released p-nitrophenol (p-NP) was calculated using a calibration curve prepared with a standard p-nitrophenol solution. Two blank controls were prepared for each test, in which the bacterial cell suspension or substrate solution was replaced with buffer.

[0071] The experimental results are as follows Figure 2 As shown in the figure, in the simulated wine environment, the strain viability of commercial lactic acid bacteria VP41 and Lactobacillus plantarum C10 is better than that of commercial lactic acid bacteria. The β-glycosidase activity of Lactobacillus plantarum C10 is higher than that of commercial lactic acid bacteria.

[0072] Example 3 Pilot fermentation experiment of Lactobacillus plantarum C10 strain

[0073] In order to evaluate the fermentation and aroma production ability of Lactobacillus plantarum C10, a fermentation experiment was conducted using Cabernet Sauvignon wine inoculated with Lactobacillus plantarum C10. The control group of this experiment was a commercial lactic acid bacteria (Lalvin VP41, Lallemand, Petaluma, CA, USA). Cabernet Sauvignon wine was obtained from the Collaborative Innovation Center for Wine Industry Technology in the Eastern Foothills of Helan Mountain, Ningxia.

[0074] Lactobacillus plantarum C10 and commercial lactic acid bacteria VP41 were added at 10 7 cfu / mL was inoculated into a 20L stainless steel fermentation tank filled with Cabernet Sauvignon wine and fermented in the full tank. There were three inoculation methods: commercial lactic acid bacteria fermentation alone (VP41), Lactobacillus plantarum C10 fermentation alone (C10), and commercial lactic acid bacteria VP41 and Lactobacillus plantarum C10 mixed inoculation fermentation (VP41+C10). The inoculation amount of both strains was 10 7 During the malic acid fermentation, samples were taken regularly to detect the malic acid rate and strain growth. When the L-malic acid content dropped below 0.2 g / L, 40 mg / L SO2 was added to terminate the fermentation. The fermentation time required for each inoculation method was about 50 days.

[0075] At the end of fermentation, samples were taken for volatile aroma determination, physical and chemical index testing, color parameters and sensory evaluation, and analyzed according to the following methods.

[0076] 1. Determination of viable bacteria count during fermentation: Use hot culture medium pouring method to determine viable bacteria count. Take wine sample 10 -9 -10 -1 After CFU / mL gradient dilution, add it to the culture dish, pour in hot MRS culture medium and shake well. After the culture medium cools and solidifies, place it in a constant temperature incubator at 37℃ and culture it upside down for 48 hours, then calculate the number of colonies.

[0077] 2. Determination of L-malic acid during fermentation: Y15 detects the content of L-malic acid during the fermentation process. Take the wine and put it into the sample cup and directly load it. It is equipped with L-MALIC ACID kit.

[0078] 3. Determination of basic physical and chemical indicators of wine: FOSS Winescan (FT 120, Foss Elec tric, Denmark) to test the basic physical and chemical indicators of wine (reducing sugar, alcohol content, volatile acid, pH, total acid, lactic acid, tartaric acid, citric acid). The wine sample was centrifuged at 8000rpm for 10min and the supernatant was taken for loading onto the machine.

[0079] 4. Determination of volatile flavor compounds: The headspace solid phase microextraction combined with gas chromatography and mass spectrometry (HS-SPME-GC-MS) method was used to determine the volatile aroma substances such as higher alcohols, esters, fatty acids, terpenes and norisoprene in wine. Take 5mL of wine sample, 1.5g of sodium chloride, and 10μL of internal standard (4-methyl-2-pentanol, 1.018g / L) and add them to a 20mL sample bottle, seal it with a polytetrafluoroethylene cap and prepare it for the machine. Use CTC CombiPAL autosampler (CTC An alytics, Zwingen, Switzerland) to complete HS-SPME pretreatment. After the wine sample was equilibrated at 40℃ for 30min, it was adsorbed on a 2cm polydimethylsiloxane / carbon mesh / divinylbenzene (DVB / CAR / PDMS) SPME extraction head (50 / 30μm, Supelco, US) for 30min.

[0080] GC-MS analysis was performed using an Agilent 7890B-5975C (Agilent Technologies, Santa Clara, CA, USA) and an HP-INNOWAX column (60m×0.25mm×0.25μm, J&W Scientific, CA, USA). The injection port temperature was 250°C, the injection split ratio was 5:1, and the thermal desorption was 8min. The carrier gas was high-purity helium with a flow rate of 1mL / min. The column heating mode was: 50°C for 1min, 3°C / min to 220°C for 5min. The mass spectrometer interface temperature was 250°C, the electron impact ion source (EI), the ion energy was 70eV, the ion source temperature was 230°C, the quadrupole temperature was 150°C, and the mass scan range was 29-350u.

[0081] The compounds were qualitatively identified by calculating the retention index (RI) using the automatic mass spectrometry deconvolution system (AMDIS) and comparing the mass spectra with the NIST 11 commercial spectral library. The peak areas were integrated using Masshunter software (MS quantitative analysis), and the peak area ratio of each substance to the corresponding internal standard was used as the basis for quantification. Quantification was performed based on the standard curve of the corresponding compound standard.

[0082] 5. CIELAB color parameter determination: The color of wine was determined using a spectrophotometer (CM-3700A, Konica Minolta, Tokyo, Japan). The wine sample was filtered through a 0.45 μm polyethersulfone membrane (J inteng Experimental Equipment Co., Ltd, Tianjin, China) and tested in a 2 mm cuvette. The L *value, a * Value, b * The blank control was distilled water.

[0083] 6. Sensory analysis: The aroma profile of wine samples was evaluated using the quantitative descriptive analysis (QDA) method. The sensory panel included 16 tasters (8 males, 8 females, aged 22 to 30 years old). All tasters were from the Grape and Wine Research Center of China Agricultural University, and had undergone systematic sensory training and had experience in wine tasting. All tasters were trained before the formal sensory analysis.

[0084] Determine aroma descriptors. First, a team of expert reviewers with rich sensory experience (3 men and 1 woman, aged 24-32 years) pre-tasted the collected wine samples and recorded the aroma descriptors of each sample. After discussion, 7 aroma descriptors were finally determined to summarize the aroma profile of all wine samples, including green, red berries, black berries, fresh fruits, dried fruits, cheese and overall aroma intensity.

[0085] Sensory training. Before the formal experiment, each taster of the sensory panel received sensory training. Sensory training included familiarizing the panel members with the standard reference materials corresponding to the descriptors and reaching a consensus on the score of each aroma intensity. The intensity of each aroma descriptor was measured on a 10-point scale. Finally, the data of each evaluator was evaluated and corrected using Panel Check software, and the data of evaluators with poor reproducibility for the same sample were eliminated.

[0086] Effect example

[0087] 1. Activity and L-malic acid consumption of Lactobacillus plantarum C10, commercial lactic acid bacteria VP41 and their mixed fermentation strains Figure 4 As shown. Plant Lactobacillus C10 can complete the malolactic fermentation in wine, which requires a certain adaptation period, and the activity of the strain remains increased in the later stage of fermentation. The consumption trend of L-malic acid in wine fermented with mixed bacteria of Plant Lactobacillus C10 and VP41 is similar to that of wine fermented with VP41 alone.

[0088] 2. The physical and chemical indicators of wine fermented by Lactobacillus plantarum C10, VP41 and their mixed fermentation are shown in Table 2. As can be seen from the data in Table 2, the basic physical and chemical indicators of each fermentation experimental group are within the control range of the national standard and meet the requirements of various indicators of wine. Compared with VP41 fermentation alone, the reducing sugar of Lactobacillus plantarum C10 fermentation alone is significantly reduced, and the lactic acid and pH are significantly increased, indicating that Lactobacillus plantarum C10 has a stronger acid-reducing ability. The alcohol content and tartaric acid of Lactobacillus plantarum C10 are significantly reduced.

[0089] Table 2 Physical and chemical indicators of Lactobacillus plantarum C10, VP41 and their mixed fermentation in wine after apple-milk fermentation

[0090]

[0091] Note: VP41 is the abbreviation of commercial lactic acid bacteria, C10 is the abbreviation of Lactobacillus plantarum C10, and VP41+C10 is the abbreviation of mixed fermentation of commercial lactic acid bacteria and Lactobacillus plantarum C10. The data in the table are mean ± standard deviation; Duncan test was used for significant variance analysis, and different letters after the compound content indicate significant differences between different treatment groups (p<0.05).

[0092] 3. The volatile compounds of wine after apple-lactation fermentation of Lactobacillus plantarum C10, commercial lactic acid bacteria VP41 and their mixed fermentation are shown in Table 3.

[0093] As shown in Table 3, compared with commercial lactic acid bacteria, the content of higher alcohols, esters, and fatty acid compounds in wine fermented by Lactobacillus plantarum C10 increased significantly. Higher alcohols can contribute to the sensory flavor of sweetness in wine. The content of 1-propanol, isobutanol, 1-butanol, isopentanol, phenylethyl alcohol, and 2,3-butanediol in wine fermented by Lactobacillus plantarum C10 increased significantly and exceeded their threshold. Among them, 1-propanol contributes to fruity aroma and butter; phenylethyl alcohol contributes to fruity aroma, sweet aroma, and honey; 2,3-butanediol contributes to fruity aroma, sweetness, butter and other flavors. The fruity aroma in wine is mainly contributed by esters. Compared with commercial lactic acid bacteria, the ester substances in Lactobacillus plantarum C10 wine increased significantly, including ethyl lactate (fruity aroma, butter), ethyl 2-hydroxy-4-methylvalerate, ethyl isopentanyl succinate, ethyl palmitate, diethyl succinate (fruity, melon), and isopentanyl lactate (creamy). After Lactobacillus plantarum C10 starts the apple-milk fermentation, ethyl lactate and isoamyl lactate accumulate in large quantities, contributing fruity and buttery flavors to the wine. Lactobacillus plantarum C10 can also increase the content of linalool and citronellol, which contribute floral and sweet flavors to the wine. Compared with the commercial lactic acid bacteria VP41, the fatty acid substances in Lactobacillus plantarum C10 wine increased significantly, among which butyric acid, isovaleric acid, caproic acid, and caprylic acid exceeded their sensory thresholds, which can add cheese and fat flavors to the wine, making the wine flavor more complex.

[0094] Compared with commercial lactic acid bacteria fermentation, the mixed fermentation of C10 and VP41 has a significant increase in the overall content of higher alcohols and esters. In the wine fermented by mixed bacteria, the content of acetate is significantly higher than that of the other two inoculation methods, such as isobutyl acetate, isoamyl acetate, hexyl acetate, phenylethyl acetate, ethyl acetate, etc. Among them, isoamyl acetate is far higher than its threshold, contributing banana and fresh fruit flavors; ethyl acetate contributes fruity, pineapple and sweet aromas to the wine. This shows that mixed fermentation has stronger metabolic activity in the production of acetate. There is also a significant increase trend in short-chain fatty acid ethyl esters in ester compounds, such as ethyl isobutyrate (caramel, butter, fruity, cream, sweet aroma), ethyl butyrate, ethyl 2-methylbutyrate (caramel, butter, fruity, cream, sweet aroma), ethyl 3-methylbutyrate (fruity aroma), and ethyl hexanoate (fruity aroma).

[0095] Table 3 Volatile compounds of Lactobacillus plantarum C10, commercial lactic acid bacteria and their mixed strains after apple-lactation fermentation in wine

[0096]

[0097]

[0098]

[0099] Note: VP41 is the abbreviation of commercial lactic acid bacteria, C10 is the abbreviation of Lactobacillus plantarum C10, and VP41+C10 is the abbreviation of mixed fermentation of commercial lactic acid bacteria and Lactobacillus plantarum C10. Compounds with OAV values ​​greater than 1 are marked as ** , while compounds with OAV values ​​between 0.1 and 1 are labeled * ; The data in the table are mean ± standard deviation; Duncan test was used to perform significant variance analysis, and different letters after the content of the same compound indicate significant differences among different treatment groups (p<0.05).

[0100] 4. The color parameters of wines after malolactic fermentation of Lactobacillus plantarum C10, commercial lactic acid bacteria VP41 and the two mixed fermentations are shown in Table 4. Compared with commercial lactic acid bacteria VP41, the color parameters of Lactobacillus plantarum C10 fermented with single bacteria were * 、a * The value increases slightly, the wine color deepens, the red hue increases, b * The value decreased and the yellow hue decreased. * The value increases, the wine color becomes lighter, a * As the value decreases, the red tint decreases.

[0101] Table 4 Color parameters of Lactobacillus plantarum C10, commercial lactic acid bacteria and their mixed strains after apple-lactation fermentation in wine

[0102] VP41 C10 VP41+C10 L* <![CDATA[42.03±0.21 b ]]> <![CDATA[42.19±0.18 b ]]> <![CDATA[45.94±1.50 a ]]> a* <![CDATA[46.12±0.07 a ]]> <![CDATA[46.27±0.06 a ]]> <![CDATA[44.37±0.41 b ]]> b* <![CDATA[16.13±0.04 ab ]]> <![CDATA[15.04±0.47 b ]]> <![CDATA[16.25±0.89 a ]]>

[0103] 5. Sensory evaluation results of wine after Lactobacillus plantarum C10, commercial lactic acid bacteria VP41 and their mixed fermentation in wine Figure 5 As shown in the figure, wine fermented with Lactobacillus plantarum C10 alone can reduce the taste of butter and cream and increase the fresh fruity aroma. Lactobacillus plantarum C10 and mixed bacteria fermentation can increase the flavor of red berries.

[0104] In summary, Lactobacillus plantarum C10 can initiate malolactic fermentation in wine, increase the fruity aroma of wine in both aroma analysis and sensory analysis, and improve the flavor quality of wine.

[0105] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. Lactobacillus plantarum, characterized in that Its deposit number is: CGMCC No.33376.

2. A mixed strain, characterized in that: include: Lactobacillus plantarum and other strains as claimed in claim 1.

3. The mixed strain according to claim 2, characterized in that The other strains include: lactic acid bacteria.

4. A microbial agent, characterized in that: include: The Lactobacillus plantarum according to claim 1 and / or the mixed strain according to claim 2 or 3.

5. Use of the plant lactobacillus (Lactiplantibacillus plantarum) according to claim 1, the mixed strain according to claim 2 or 3 and / or the microbial agent according to claim 4 in fruit wine fermentation.

6. The use according to claim 5, characterized in that The fruit wine includes: grape wine.

7. Use of the plant lactobacillus plantarum according to claim 1, the mixed strain according to claim 2 or 3 and / or the microbial agent according to claim 4 in improving the aroma and / or taste of fruit wine.

8. The use according to claim 7, characterized in that The improved taste of fruit wine comprises: (I), reducing the content of reducing sugars in fruit wine; and / or (II), reducing the alcohol content of the wine; and / or (III) reducing the content of L-malic acid in fruit wine; The method of improving the aroma of fruit wine comprises increasing the content of one or more of alcohol substances, ester substances and fatty acid substances.

9. A method for fermenting fruit wine, characterized in that: The fruit wine is obtained by mixing and fermenting the Lactobacillus plantarum according to claim 1, the mixed strain according to claim 2 or 3 and / or the microbial agent according to claim 4 with raw materials.

10. Fruit wine obtained by the fermentation method as claimed in claim 9.

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