CN110760471B - Acetobacter pasteurianus, microbial agent and application thereof, and vinegar preparation method - Google Patents
Acetobacter pasteurianus, microbial agent and application thereof, and vinegar preparation method Download PDFInfo
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Abstract
The invention discloses acetobacter pasteurianus, which is preserved in the common microorganism center of China general microbiological culture Collection center, wherein the preservation date is 5 and 13 days in 2019, and the preservation number is CGMCC No. 17802. The invention also discloses a microbial agent containing the acetobacter pasteurianus. The invention also discloses application of the acetobacter pasteurianus and the microbial agent in vinegar brewing and a preparation method of vinegar. When the acetobacter pasteurianus is applied to brewing solid vinegar, the acid production rate is high, the acid production is high, the heat extraction is fast, and the overall flavor of the obtained vinegar is good; when the acetobacter pasteurianus is applied to brewing liquid vinegar, the yield of acid is high, particularly, the lactic acid and ester production performance are excellent, and the obtained vinegar has good integral flavor and high quality.
Description
Technical Field
The invention relates to Acetobacter pasteurianus HSCY1085(Acetobacter pasteurianus) and application thereof, and also relates to a microbial agent containing the Acetobacter pasteurianus and a preparation method of vinegar.
Background
The edible vinegar mainly comprises aromatic vinegar and mature vinegar obtained by solid fermentation and rice vinegar and fruit vinegar obtained by liquid fermentation. The traditional solid state fermentation environment is an open type fermentation environment, the flora structure is complex, the microorganisms are various and difficult to regulate, and when the microbial flora structure is analyzed, the acetic acid bacteria occupy an important position in the acetic acid fermentation process. When the acetic acid bacteria are applied to the solid-state fermentation process, the heat extraction speed, acid production and the like are main concerns, but the single-strain inoculation fermentation result is not ideal, so that the excavation of the excellent acetic acid bacteria is a long-term work, and the excellent acetic acid bacteria are precious strain resources.
The liquid vinegar such as rice vinegar is prepared by fermenting and brewing grains such as millet, sorghum, sticky rice, barley, corn, sweet potato, vinasse and the like, and the liquid vinegar needs high acid production during fermentation. At present, rice vinegar on the market is produced by adopting a traditional industrialized process, the organic acid content is low, the acetic acid content is high, the whole flavor is stimulated and not soft, even the flavor is uncoordinated, and the taste is influenced.
In general, acetic acid bacteria isolated from a solid brewing environment can be applied to solid fermentation, but are not well adapted to the harsh environment of liquid fermentation, so that the application of acetic acid bacteria is limited. In addition, some protective agents are usually added and freeze-drying methods are used when preparing the microbial agents in solid form, so that the processing cost is high.
Disclosure of Invention
The purpose of the invention is as follows: the invention aims at providing a acetobacter pasteurianus which has high acid production and improves the overall flavor and quality of vinegar, the invention aims at providing a microbial agent containing the acetobacter pasteurianus, the invention aims at providing an application of the acetobacter pasteurianus and the microbial agent in vinegar brewing, and the invention aims at providing a vinegar preparation method.
The technical scheme is as follows: the acetobacter pasteurianus is preserved in the common microorganism center of China Committee for culture Collection of microorganisms, the preservation date is 5 months and 13 days in 2019, and the preservation number is CGMCC No. 17802.
The microbial agent comprises the acetobacter pasteurianus. The preparation method comprises the following steps:
(1) inoculating into liquid fermentation tank with 10% inoculum size, culturing for 24 hr, and detecting viable count to 107Taking out the culture solution when the concentration is CFU/mL;
(2) centrifuging at 7000rpm for 8min, collecting thallus, and eluting the obtained bacterial sludge with 11% skim milk;
(3) sterilizing the bacterial suspension and bran (sterilized at 105 ℃ for 3h, then cooling to 40 ℃ for later use) by a ratio of 1: 1 (mass/mass), drying at 40 ℃ for 2h, drying at 34 ℃ for 12h to complete drying, placing in a sealing bag, pumping out the gas in the bag, sealing, and storing at 4 ℃ for later use.
The application of the acetobacter pasteurianus in vinegar brewing is solid vinegar brewing or liquid vinegar brewing.
The microbial agent is applied to vinegar brewing, and the vinegar brewing is solid vinegar brewing.
Preferably, the vinegar is one of Zhenjiang aromatic vinegar, rice vinegar or mulberry vinegar.
The vinegar brewing method of the present invention is characterized by comprising the steps of:
(1) preparing a strain seed solution using the Acetobacter pasteurianus of claim 1;
(2) inoculating and fermenting the strain seed liquid prepared in the step (1).
Preferably, the preparation of the strain seed solution in the step (1) comprises the preparation of a primary seed solution and a secondary seed solution.
Preferably, the preparation of the primary seed solution comprises the following steps: culturing the liquid activated pasteurella vinegar bacillus culture solution at the rotating speed of 180 rpm-220 rpm at the temperature of 30-35 ℃ for 20-36 h according to the inoculation amount of 10-20%.
Preferably, the preparation of the secondary seed liquid comprises the following steps: culturing the secondary seed liquid at 30-35 ℃ for 20-24 h according to the inoculation amount of 10-20 percent, wherein the viable count reaches 107CFU/mL。
The vinegar brewing method comprises the following steps of (2) when Zhenjiang aromatic vinegar is prepared, carrying out enzymolysis, sterilization, inoculation and solid layered fermentation according to a Zhenjiang aromatic vinegar brewing process until the fermentation is finished; when rice vinegar is prepared, inoculating the acetobacter pasteurianus strain seed liquid to carry out acetic fermentation to obtain the vinegar; and (2) when the mulberry vinegar is prepared, adding the mulberry wine into a fermentation tank, adjusting the overall alcoholic strength, inoculating the seed liquid of the acetobacter pasteurianus strain, preserving heat, stirring and ventilating, and fermenting in a fermentation mode of cutting vinegar until the end.
The acetobacter pasteurianus is separated from the traditional solid vinegar brewing environment, not only can adapt to the solid vinegar brewing environment, but also can adapt to the liquid vinegar brewing environment well, so that the acetobacter pasteurianus can be applied to solid vinegar brewing and liquid vinegar brewing. In addition, the acetobacter pasteurianus has strong tolerance capability, and the microbial agent can be prepared by a drying mode, so that the processing cost can be reduced.
Has the advantages that: compared with the prior art, the invention has the following remarkable advantages:
(1) the acetobacter pasteurianus can adapt to the environments of solid vinegar brewing and liquid vinegar brewing, when the acetobacter pasteurianus is applied to the solid vinegar brewing, the acid production rate is high, the acid production is high, the heat extraction is fast, and the overall flavor of the obtained vinegar is good; when the method is applied to brewing liquid vinegar, the yield of acid is high, particularly, the lactic acid and ester are better, and the obtained vinegar has good integral flavor and high quality.
(2) The acetobacter pasteurianus has strong tolerance capability, can be prepared by a drying mode, can greatly save the cost and is convenient to store and apply.
(3) The microbial agent disclosed by the invention can adapt to the brewing environment of solid vinegar.
(4) The vinegar brewing method can endow vinegar with abundant flavor substances.
Drawings
FIG. 1 shows the colony morphology of Acetobacter pasteurianus HSCY1085 according to the present invention;
FIG. 2 is a temperature change chart during fermentation of Zhenjiang aromatic vinegar;
FIG. 3 is a diagram showing the total acid change during fermentation of Zhenjiang aromatic vinegar.
Detailed Description
The technical solution of the present invention is further illustrated by the following examples.
Example 1: isolation and identification of Strain Acetobacter pasteurianus HSCY1085
Wherein calcium carbonate, glucose, agar powder, absolute ethyl alcohol, acetic acid and sodium hydroxide are all purchased from chemical reagents of national medicine group, Inc.; yeast extracts were purchased from OXOID, United kingdom.
1. Strain isolation
Taking 10g of Zhenjiang aromatic vinegar fermented grains, adding into 90mL of sterilized physiological saline, shaking uniformly in a shaking table, then adding 100 mu L of sample into 900 mu L of physiological saline, uniformly mixing in a vortex oscillator, and then carrying out gradient dilution. Mixing, sequentially spreading on solid culture medium (containing glucose 20g, yeast extract 10g, and agar powder 15g per 1L, sterilizing at 121 deg.C for 20min, cooling, adding 3% ethanol), and culturing at 30 deg.C for 3 days. Observing whether a transparent ring exists on the flat plate, and picking corresponding strains.
2. Bacterial strain rescreening
The primarily screened strains are inoculated on a re-screening solid plate (20 g of glucose, 10g of yeast extract, 30mL of acetic acid and 30mL of ethanol, 15g/L of agar powder is added), and after 3 days of culture at 30 ℃, the strains with the largest transparent circles are selected.
The screened strains are inoculated into a re-screening liquid culture medium (20 g of glucose, 10g of yeast extract, 30mL of acetic acid, 50mL of ethanol and 1L of distilled water), cultured at the rotating speed of 200rpm at the temperature of 30 ℃ for 20h, the content of total acid (calculated by acetic acid) is titrated by sodium hydroxide, the measurement is carried out once every 24h, and the change condition of acid production is recorded.
After two rounds of re-screening, a Acetobacter pasteurianus HSCY1085 strain with excellent acid production performance is finally obtained, and the colony morphology of the Acetobacter pasteurianus HSCY1085 strain is shown in figure 1.
3. Identification of strains
The measured 16S rDNA sequence is compared and analyzed in an NCBI database, and the strain is named as Acetobacter pasteurianus HSCY1085(Acetobacter pasteurianus) by combining physiological and biochemical characteristics, and the 16S rDNA sequence is shown in SEQ ID NO. 1.
The strain is preserved in the common microorganism center of China Committee for culture Collection of microorganisms, the preservation place is the microorganism research institute of China academy of sciences No. 3 of Beijing, facing the sunny region, the preservation date is 2019, 5 and 13 days, the preservation number is CGMCC No.17802, and the strain is classified and named Acetobacter passaturia Nus.
Example 2: activation of Acetobacter pasteurianus HSCY1085 and preparation of seed liquid
(1) Bacterial activation and propagation
The strain preserved in the glycerol tube is activated by liquid, namely 1mL of the strain is respectively taken and transferred into a test tube (containing 5mL of culture medium), and the strain is cultured for 24h at the temperature of 30 ℃ and the rotating speed of 180 rpm.
(2) Preparation of first-order seed liquid
10mL of the culture solution is taken, namely 10% of the inoculum size is transferred into a 250mL triangular flask (the liquid loading volume is 100mL), the culture solution is cultured for 20 hours at 30 ℃ and 220rpm by a shaking table, and first-stage seed solution is prepared.
(3) Preparation of Secondary seed liquid
Inoculating the first-stage seed solution into 2L triangular flask (liquid content 1L) at 10%, culturing at 30 deg.C for 20 hr with viable count of 8.9 × 107CFU/mL, the fermentation broth can be used for subsequent experiments.
Example 3: activation of Acetobacter pasteurianus HSCY1085 and preparation of seed liquid
(1) Bacterial activation and expanding culture
The strain preserved in the glycerol tube is activated by liquid, namely 1mL of the strain is respectively taken and transferred into a test tube (containing 5mL of culture medium), and the strain is cultured for 24 hours at 30 ℃ and 180rpm by a shaking table.
(2) Preparation of first-order seed liquid
20mL of the culture solution is taken, namely 20% of the inoculum size is transferred into a 250mL triangular flask (the liquid loading volume is 100mL), the culture solution is cultured for 36 hours at 35 ℃ and 180rpm by a shaking table, and first-stage seed solution is prepared.
(3) Preparation of Secondary seed liquid
Inoculating the first-stage seed solution into 2L triangular flask (liquid content 1L) at 20%, culturing at 35 deg.C for 24 hr with viable count of 5.4 × 107CFU/mL, the fermentation broth can be used for subsequent experiments.
Example 4: application of acetobacter pasteurianus HSCY1085 in Zhenjiang aromatic vinegar brewing
The embodiment provides the application of the pure bacteria in Zhenjiang aromatic vinegar brewing.
1. Test group
(1) Selection and preparation of raw materials
Removing impurities from fresh glutinous rice by winnowing, sieving, selecting full glutinous rice, adding water with the same mass, soaking at room temperature overnight, steaming in a steam box until the cooked rice is soft and glutinous, watering with cold water to cool the cooked rice to room temperature, pulverizing, and grinding into thick liquid.
(2) Enzymolysis
Adding amylase into glutinous rice for enzymolysis for 30min, adding diastase, performing enzymolysis at 60 deg.C for 6 hr, and transferring into fermentation tank.
(3) Sterilizing, inoculating and fermenting
Sterilizing the fermentation tank at high temperature, cooling to about 30 deg.C, adding yeast and Daqu, and fermenting at 28 deg.C for about 7 days.
(4) And (3) taking 3 350kg of large jars in the step (3), adding 90kg of fermented glutinous rice into each jar, adding 30.6kg of bran and 14.4kg of large bran, and uniformly mixing the fermented glutinous rice and grains (namely vinegar grains). Taking 1L of secondary seed liquid of the acetobacter pasteurianus prepared by the method in the embodiment 2 of the invention, uniformly scattering the secondary seed liquid on the surface of the vinegar culture, and finally covering a small amount of bran on the top for heat preservation.
(5) And (3) turning the fermented grains layer by layer according to the Zhenjiang aromatic vinegar brewing process, and ending the fermentation when the total acid is not increased after the fermented grains are fermented.
2. Control group
And (3) taking 3 350kg of large jars in the step (3), adding 90kg of fermented glutinous rice into each jar, adding 30.6kg of bran and 14.4kg of large bran, and uniformly mixing the fermented glutinous rice and grains (namely vinegar grains). Taking Shanghai brewing 1.01 (purchased from CICC, numbered CICC 20001), activating and culturing according to the method of the embodiment 2 of the invention, adjusting the concentration of the seed solution to be consistent with the secondary seed solution of the test group Acetobacter pasteurianus after the culture is finished, uniformly spreading the seed solution on the surface of the vinegar culture, covering a small amount of bran on the top, and preserving the heat, wherein the subsequent steps are the same as the step (5) above.
As shown in FIG. 2, the heat-extracting speed of the Acetobacter pasteurianus HSCY1085 of the present invention was faster than that of Shanghai-Niang 1.01 in the first two days, the temperature of the product was rapidly increased to 40 ℃ or higher, and then the temperature was at a relatively high level in a fluctuating state, as compared with the control group.
As shown in FIG. 3, compared with the control group, the acid production capacity of the Acetobacter pasteurianus HSCY1085 of the present invention is better than that of Shanghai brewing 1.01 during the whole fermentation process, wherein the amount of the Acetobacter pasteurianus HSCY1085 of the present invention can reach 8.12g/100mL, and the amount of the Shanghai brewing 1.01 is 6.52g/100 mL.
The finished vinegar prepared by the test group and the control group is subjected to sensory evaluation, the finished vinegar is developed from four dimensions of color, body form, fragrance and taste, scores are respectively given, and the sensory evaluation result is shown in table 1.
TABLE 1 sensory evaluation results of finished Vinegar
| Group of | Color and luster (10 minutes) | Posture (10 minutes) | Fragrance (10 points) | Taste (10 points) |
| Test group | 9.2 | 9 | 9.4 | 9.5 |
| Control group | 8.8 | 8.5 | 8.1 | 7.9 |
Sensory results show that the values of the test group in four dimensions of color, shape, aroma and taste are higher than those of the control group, and the overall flavor of the finished vinegar prepared by the test group is superior to that of the control group.
Example 5: application of acetobacter pasteurianus HSCY1085 in rice vinegar brewing
1. Test group
Preparing rice wine and rice vinegar:
(1) selection and preparation of raw materials
Removing impurities from fresh rice by winnowing, sieving, selecting full-grain rice, adding water with the same mass, soaking at room temperature overnight, placing in a steam box, steaming until the rice is soft and glutinous, then pouring cold water to cool the rice to room temperature, and then crushing and grinding into thick liquid.
(2) Enzymolysis
Adding amylase into the cooked rice for enzymolysis for 30min, adding diastase, and performing enzymolysis at 60 deg.C for 6 h.
(3) Sterilizing, inoculating and fermenting
Sterilizing the fermentation tank at high temperature, cooling to about 30 deg.C, adding yeast and Daqu, fermenting at 28 deg.C for 7 days, and adjusting alcohol content to 8% vol after fermentation. The secondary seed liquid of Acetobacter pasteurianus prepared by the method described in the example 3 of the present invention was added to the diluted fermentation supernatant (1L of alcohol in each pot, 2L of distilled water) at a volume ratio of 10% to perform acetic fermentation at 30 ℃ to obtain raw vinegar.
(4) Filtering, clarifying, sterilizing, etc. to obtain rice vinegar.
2. Control group
Taking Shanghai Niang 1.01, activating and culturing according to the method of the invention in the embodiment 3, adjusting the concentration of the seed liquid to be consistent with the secondary seed liquid of the test group Acetobacter pasteurianus after the culture is finished, adding the seed liquid into the fermentation liquid in the step (3), performing acetic fermentation at 30 ℃ to obtain the original vinegar, and performing the subsequent steps as in the step (4).
The vinegar was prepared by the above method, and the acidity of the test group was 12.5g/100mL, and the acidity of the control group was 7.1g/100 mL. Firstly, the composition of organic acid for preparing the rice vinegar is evaluated, as shown in table 2, the content of acetic acid in a test group is obviously higher than that of a control group, and simultaneously, the content of lactic acid is also higher than that of the control group, which shows that the acetic acid producing capacity and the lactic acid producing capacity of the acetobacter pasteurianus HSCY1085 are stronger, so that the overall taste of the rice vinegar is softer.
Secondly, the volatile flavor substance composition of the prepared rice vinegar is evaluated, as shown in table 3, the acetobacter pasteurianus can produce various esters, alcohols, aldehydes and the like, wherein the contents of dimethyl succinate and beta-phenylethyl alcohol in the test group are obviously higher than those in the control group, and the two substances respectively have fruity fragrance and flowery fragrance, so that the rice vinegar has pleasant smell, and the quality of the rice vinegar prepared by the test group is better than that of the control group.
Finally, the antioxidant capacity of the prepared rice vinegar is evaluated, and as shown in Table 4, the total phenol content and the total flavone content of the rice vinegar prepared by the acetobacter pasteurianus are obviously higher than those of a control group, so that the antioxidant capacity of the prepared vinegar is stronger.
In conclusion, the overall quality of the rice vinegar prepared by the test group is better than that of the control group.
TABLE 2 composition and content of organic acids in rice vinegar
TABLE 3 composition of volatile flavor components in rice vinegar and their ratio (%)
| Volatile substance | Control group | Test group |
| Acetaldehyde | 3.03 | 2.66 |
| Acetic acid methyl ester | 2.15 | 1.81 |
| Isobutanol | 2.56 | 1.27 |
| Acetic acid isoamyl ester | 3.34 | 2.18 |
| Furfural | 4.08 | 4.19 |
| Lactic acid ethyl ester | 2.99 | 3.02 |
| Beta-phenylethyl alcohol | 30.31 | 32.81 |
| Ethyl acetate | 2.54 | 2.10 |
| Succinic acid dimethyl ester | 36.19 | 38.23 |
| Succinic acid diethyl ester | 8.79 | 6.58 |
| Tyrosol | 4.02 | 5.15 |
TABLE 4 index (mg/L) relating to oxidation resistance of rice vinegar
Example 6: application of acetobacter pasteurianus HSCY1085 in brewing of mulberry vinegar
1. Test group
(1) Sterilizing a fermentation tank, diluting the concentrated mulberry juice with purified water, pouring the diluted mulberry juice into the fermentation tank, filling the fermentation tank with 60% of liquid, inoculating yeast, fermenting at 28 ℃, stopping fermentation until the residual sugar content is not changed any more, and refrigerating the fermented mulberry wine for later use.
(2) Selecting a 5L fermentation tank, adding 2L of distilled water and 1L of mulberry wine (the alcoholic strength is 8-12% vol), and adjusting the overall alcoholic strength to be about 3%. Inoculating the secondary seed liquid of acetobacter pasteurianus prepared by the method in the embodiment 2 of the invention according to the inoculation amount of 10-15%, keeping the temperature at 28-32 ℃, stirring at 125-135 rpm, and ventilating at 0.6-0.8 vvm, discharging 1L fermentation liquid by adopting a fermentation mode of cutting vinegar when fermenting until the total acid is 5g/100mL, and simultaneously supplementing 1L mulberry wine with the alcoholic strength of 8-12 vol%. A certain amount of fermentation liquor is discharged every time, the rest is used as next seed liquor, 1L of mulberry wine (containing 0.6 thousandth of nutrient salt and 1.5 thousandth of glucose) with a certain amount of alcoholic strength of 8-12% vol is added, and by analogy, the total acidity is divided and supplemented once every time the total acidity is increased by 1.5-2 degrees.
2. Control group
Taking Shanghai brewing 1.01, activating and culturing according to the method of the invention in the embodiment 2, adjusting the concentration of the seed liquid to be consistent with the secondary seed liquid of the test group Acetobacter pasteurianus after the culture is finished, adding the seed liquid into the fermentation liquid in the step (2), and performing subsequent fermentation according to the method in the step (2).
TABLE 5 composition and content of organic acids in Mulberry vinegar
| Name of organic acid | Control group (g/L) | Test set (g/L) |
| Tartaric acid | 0.88 | 0.67 |
| Pyruvic acid | 0.64 | 0.85 |
| Ketoglutaric acid | 0.22 | 0.41 |
| Lactic acid | 7.90 | 10.93 |
| Acetic acid | 33.87 | 35.36 |
| Citric acid | 0.49 | 0.78 |
| Pyroglutamic acid | 2.52 | 2.94 |
| Succinic acid | 3.29 | 3.16 |
TABLE 6 Mulberry vinegar antioxidant capacity related index (mg/L)
Firstly, the composition of organic acid for preparing the mulberry vinegar is evaluated, and as shown in table 5, the lactic acid content of a test group is obviously higher than that of a control group, so that the taste is softer; secondly, the antioxidant capacity of the prepared mulberry vinegar is evaluated, as shown in table 6, the contents of total flavones and total phenols in the test group are higher than those in the control group, and therefore the prepared mulberry vinegar has better antioxidant capacity.
In conclusion, the overall quality of the mulberry vinegar prepared by the test group is better than that of the control group.
Example 7: acetobacter pasteurianus HSCY1085 microbial inoculum and application thereof in solid fermentation of vinegar
The embodiment provides a microbial agent and application thereof in solid-state fermented vinegar.
1. Preparation of Acetobacter pasteurianus HSCY1085 microbial inoculum
(1) Inoculating 10% second-stage seed solution (fermentation tank liquid loading amount is about 25L) into 50L liquid fermentation tank, culturing for 24 hr, and detecting viable bacteria count to 107The culture broth was removed at CFU/mL.
(2) The thalli is collected after centrifugation at 7000rpm for 8min, and the obtained bacterial sludge is eluted by 11% skim milk.
(3) Sterilizing the bacterial suspension and bran (sterilized at 105 ℃ for 3h, then cooling to 40 ℃ for later use) by a ratio of 1: 1 (mass/mass), drying at 40 ℃ for 2h, drying at 34 ℃ for 12h until completely dried, placing in a sealing bag, removing gas in the bag, sealing, storing at 4 ℃ for later use, and sampling before sealing to count colonies.
(4) Comparative resistance test
The Shanghai brewing is activated and cultured according to the method of the invention, the seed liquid obtained by culturing is prepared according to the steps (1) to (3) of the control group bacterium agent, and the samples are taken for colony counting before sealing.
2. Application of acetobacter pasteurianus HSCY1085 microbial inoculum in solid state fermentation of vinegar
(1) Test group
The preliminary preparation is carried out according to the steps (1) to (3) in the embodiment 4, bran, chaff and fermented glutinous rice (the total amount of water is 350kg) are added into a cylinder (the materials are added according to the proportion in the step (4) in the embodiment 4), the mixture is uniformly mixed, the microbial inoculum of the invention with the total amount of the extracted materials being 3 percent (g/kg) is stirred and activated for at least 15min in warm water (35-42 ℃), the activated microbial inoculum suspension is uniformly scattered on the upper surface of the materials and is fully mixed and contacted with the materials, the mixture is piled on the surface, bran is covered, the temperature is kept and the heat is raised, when the temperature is raised to be higher than 38 ℃, the fermented grains can be turned and taken, then, the fermented grains are manually turned once a day, and the mixture enters the acetic acid fermentation stage.
(2) Control group
The difference from the test group is that the fermentation agent adopts Shanghai 1.01, the activation and the culture are carried out according to the method of the embodiment 2 of the invention, and the concentration of the seed liquid is adjusted to be consistent with the microbial agent suspension of the test group after the culture is finished.
The number of live bacteria of the Acetobacter pasteurianus HSCY1085 after drying can reach 107CFU/g, survival rate of about 40%, survival rate of control group of about 15%, and viable count of 106CFU/g shows that the temperature tolerance of the acetobacter pasteurianus HSCY1085 is stronger than that of a control group, and the microbial agent can well preserve the activity of strains; in addition, when the microbial agent is applied to solid state fermentation vinegar, the temperature of unstrained spirits can be raised to more than 40 ℃ about 1.5 days after inoculation, which shows that the microbial agent can be well adapted to the environment of the solid state fermentation vinegar.
In conclusion, the microbial agent disclosed by the invention can well preserve the activity of strains and can be well adapted to the environment of solid-state fermentation vinegar.
Sequence listing
<120> Acetobacter pasteurianus, microbial agent and application thereof, and vinegar preparation method
<140> 2019112918017
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gtacagtcag atgtgaaatc cccgggctta acctgggagc tgcatttgat acgtgcagac 540
tagagtgtga gagagggttg tggaattccc agtgtagagg tgaaattcgt agatattggg 600
aagaacaccg gtggcgaagg cggcaacctg gctcattact gacgctgagg cgcgaaagcg 660
tggggagcaa acaggattag ataccctggt agtccacgct gtaaacgatg tgtgctagat 720
gttgggtgac ttagtcattc agtgtcgcag ttaacgcgtt aagcacaccg cctggggagt 780
acggccgcaa ggttgaaact caaaggaatt gacgggggcc cgcacaagcg gtggagcatg 840
tggtttaatt cgaagcaacg cgcagaacct taccagggct tgaatgtaga ggctgcaagc 900
agagatgttt gtttcccgca agggacctct aacacaggtg ctgcatggct gtcgtcagct 960
cgtgtcgtga gatgttgggt taagtcccgc aacgagcgca acccctatct ttagttgcca 1020
tcaggttggg ctgggcactc tagagagact gccggtgaca agccggagga aggtggggat 1080
gacgtcaagt cctcatggcc cttatgtcct gggctacaca cgtgctacaa tggcggtgac 1140
agtgggaagc taggtggtga caccatgctg atctctaaaa gccgtctcag ttcggattgc 1200
actctgcaac tcgagtgcat gaaggtggaa tcgctagtaa tcgcggatca gcatgccgcg 1260
gtgaatacgt tcccgggcct tgtacacacc gcccgtcaca ccatgggagt tggtttgacc 1320
ttaagccggt gagcgaaccg caaggacgca gc 1352
Claims (10)
1. Acetobacter pasteurianus strain (A), (B), (C)Acetobacter pasteurianus) The culture medium is characterized in that the acetobacter pasteurianus HSCY1085 is preserved in China general microbiological culture Collection center (CGMCC), the preservation date is 5 months and 13 days in 2019, and the preservation number is CGMCC No. 17802.
2. A microbial agent, comprising the acetobacter pasteurii of claim 1.
3. Use of Acetobacter pasteurianus in vinegar brewing according to claim 1, wherein the vinegar brewing is solid vinegar brewing or liquid vinegar brewing.
4. Use of the microbial inoculum according to claim 2 in vinegar brewing, wherein the vinegar brewing is solid vinegar brewing.
5. The use of claim 3, wherein the vinegar is one of Zhenjiang aromatic vinegar, rice vinegar or mulberry vinegar.
6. A vinegar brewing method is characterized by comprising the following steps:
(1) preparing a strain seed solution using the Acetobacter pasteurianus of claim 1;
(2) inoculating and fermenting the strain seed liquid prepared in the step (1).
7. The vinegar brewing method according to claim 6, wherein the preparation of the strain seed solution in the step (1) includes preparation of a primary seed solution and a secondary seed solution.
8. The vinegar brewing method according to claim 7, wherein the preparation of the primary seed solution comprises the steps of: culturing the liquid activated pasteurella vinegar bacillus culture solution at the temperature of 30-35 ℃ and the rotating speed of 180-220 rpm for 20-36 h according to the inoculation amount of 10-20%.
9. The vinegar brewing method according to claim 7, wherein the preparation of the secondary seed solution comprises the steps of: culturing the secondary seed liquid at 30-35 ℃ for 20-24 h according to the inoculation amount of 10-20%, wherein the viable count reaches 107CFU/mL。
10. The vinegar brewing method according to claim 6, wherein in the step (2), when preparing Zhenjiang aromatic vinegar, the fermentation is completed by performing enzymolysis, sterilization, inoculation and solid layered fermentation according to the Zhenjiang aromatic vinegar brewing process; when rice vinegar is prepared, inoculating the acetobacter pasteurianus strain seed liquid to carry out acetic fermentation to obtain the vinegar; and (2) when the mulberry vinegar is prepared, adding the mulberry wine into a fermentation tank, adjusting the overall alcoholic strength, inoculating the seed liquid of the acetobacter pasteurianus strain, preserving heat, stirring and ventilating, and fermenting in a fermentation mode of cutting vinegar until the end.
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| CN112410162B (en) * | 2020-11-09 | 2023-01-13 | 河北科技师范学院 | Hawthorn polyphenol fermented fruit vinegar beverage and preparation process thereof |
| CN113699069B (en) * | 2021-08-30 | 2023-07-21 | 江苏恒顺醋业股份有限公司 | Bacterial strain HSCY2073, separation screening thereof and application thereof in improving flavor quality of vinegar |
| CN115074266B (en) * | 2022-05-09 | 2023-09-22 | 江苏恒顺醋业股份有限公司 | Stress-resistant acetic acid bacteria and application thereof in vinegar brewing |
| CN116286559B (en) * | 2023-05-10 | 2023-09-05 | 阳西美味鲜食品有限公司 | High-yield high-temperature-resistant acetobacter pasteurii M112 and application thereof |
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