CN114507585A - Fermentation column capable of recovering lactic acid bacteria in fermentation liquor, fermentation tank and fermentation method - Google Patents
Fermentation column capable of recovering lactic acid bacteria in fermentation liquor, fermentation tank and fermentation method Download PDFInfo
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- 238000000855 fermentation Methods 0.000 title claims abstract description 378
- 230000004151 fermentation Effects 0.000 title claims abstract description 372
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 title claims abstract description 86
- 241000894006 Bacteria Species 0.000 title claims abstract description 51
- 239000004310 lactic acid Substances 0.000 title claims abstract description 43
- 235000014655 lactic acid Nutrition 0.000 title claims abstract description 43
- 238000000034 method Methods 0.000 title claims abstract description 31
- AFSDNFLWKVMVRB-UHFFFAOYSA-N Ellagic acid Chemical compound OC1=C(O)C(OC2=O)=C3C4=C2C=C(O)C(O)=C4OC(=O)C3=C1 AFSDNFLWKVMVRB-UHFFFAOYSA-N 0.000 claims abstract description 91
- 235000013525 pomegranate juice Nutrition 0.000 claims abstract description 91
- 238000000502 dialysis Methods 0.000 claims abstract description 56
- 238000007789 sealing Methods 0.000 claims abstract description 48
- 239000012528 membrane Substances 0.000 claims abstract description 34
- 238000009423 ventilation Methods 0.000 claims abstract description 20
- 235000013305 food Nutrition 0.000 claims abstract description 6
- 240000006024 Lactobacillus plantarum Species 0.000 claims description 39
- 235000013965 Lactobacillus plantarum Nutrition 0.000 claims description 34
- 229940072205 lactobacillus plantarum Drugs 0.000 claims description 34
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 32
- 239000007788 liquid Substances 0.000 claims description 31
- 230000001954 sterilising effect Effects 0.000 claims description 23
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 23
- 238000004659 sterilization and disinfection Methods 0.000 claims description 22
- 241000219991 Lythraceae Species 0.000 claims description 21
- 235000014360 Punica granatum Nutrition 0.000 claims description 21
- 239000000047 product Substances 0.000 claims description 17
- 239000008223 sterile water Substances 0.000 claims description 17
- 239000001963 growth medium Substances 0.000 claims description 15
- 238000005406 washing Methods 0.000 claims description 14
- 229910052701 rubidium Inorganic materials 0.000 claims description 13
- IGLNJRXAVVLDKE-UHFFFAOYSA-N rubidium atom Chemical compound [Rb] IGLNJRXAVVLDKE-UHFFFAOYSA-N 0.000 claims description 13
- 238000004140 cleaning Methods 0.000 claims description 10
- 239000000463 material Substances 0.000 claims description 10
- 238000011049 filling Methods 0.000 claims description 8
- 229910001220 stainless steel Inorganic materials 0.000 claims description 8
- 239000010935 stainless steel Substances 0.000 claims description 8
- CDBYLPFSWZWCQE-UHFFFAOYSA-L sodium carbonate Substances [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 claims description 7
- 229920001817 Agar Polymers 0.000 claims description 6
- 238000009631 Broth culture Methods 0.000 claims description 6
- 239000008272 agar Substances 0.000 claims description 6
- 239000012153 distilled water Substances 0.000 claims description 6
- 238000011156 evaluation Methods 0.000 claims description 6
- 229910000029 sodium carbonate Inorganic materials 0.000 claims description 6
- 239000006228 supernatant Substances 0.000 claims description 6
- 238000012371 Aseptic Filling Methods 0.000 claims description 5
- 241001052560 Thallis Species 0.000 claims description 5
- 238000009924 canning Methods 0.000 claims description 5
- 238000001914 filtration Methods 0.000 claims description 5
- 238000010438 heat treatment Methods 0.000 claims description 5
- 239000002609 medium Substances 0.000 claims description 5
- 238000003860 storage Methods 0.000 claims description 5
- 238000005303 weighing Methods 0.000 claims description 5
- 230000001580 bacterial effect Effects 0.000 claims description 4
- 230000001953 sensory effect Effects 0.000 claims description 4
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 3
- 239000000741 silica gel Substances 0.000 claims description 3
- 229910002027 silica gel Inorganic materials 0.000 claims description 3
- 238000012258 culturing Methods 0.000 claims description 2
- 238000007599 discharging Methods 0.000 claims 1
- 239000013049 sediment Substances 0.000 claims 1
- 230000000813 microbial effect Effects 0.000 abstract description 2
- 238000004519 manufacturing process Methods 0.000 description 14
- 241000186660 Lactobacillus Species 0.000 description 10
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- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M23/00—Constructional details, e.g. recesses, hinges
- C12M23/02—Form or structure of the vessel
- C12M23/14—Bags
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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Abstract
The invention belongs to the technical field of food microbial fermentation, and particularly relates to a fermentation column capable of recovering lactic acid bacteria in fermentation liquor, a fermentation tank and a fermentation method thereof. The technical scheme of the invention is as follows: the fermentation column is a hollow cylindrical structure, the top of the fermentation column is provided with an opening, the bottom of the fermentation column is provided with a closed end, and the surface of the cylindrical structure is fully distributed with vent holes; the inside of the semipermeable membrane is wrapped with strain fermentation liquor, the aperture of the semipermeable membrane limits the free inlet and outlet of lactic acid bacteria, and the semipermeable membrane is placed in the hollow structure of the fermentation column after wrapping the strain fermentation liquor; the top opening of the fermentation column is blocked by the sealing plug, the middle part of the sealing plug is provided with a ventilation channel, and a one-way sealing structure is arranged in the ventilation channel. Adding lactic acid bacteria into the dialysis bag, immersing part of the dialysis bag into the pomegranate juice, and fixing the dialysis bag in a certain way to enable the dialysis bag to be suspended in the pomegranate juice for fermentation. After fermentation, the dialysis bag in the fermentation tank can be transferred into a new fermentation tank, so that the repeated utilization of the lactic acid bacteria is realized.
Description
The technical field is as follows:
the invention belongs to the technical field of food microbial fermentation, and particularly relates to a fermentation column capable of recovering lactic acid bacteria in fermentation liquor, a fermentation tank and a fermentation method thereof.
Technical background:
the pomegranate juice lactic acid bacteria beverage has unique flavor and health care effect. At present, the fermentation process of the pomegranate juice lactic acid bacteria beverage is mainly a traditional strain direct-vat-set fermentation method, namely, lactic acid bacteria is added into pomegranate juice according to a certain proportion for anaerobic fermentation.
The strain direct-vat-set fermentation method has certain defects in production, the added strains cannot be recycled, but the lactic acid strains are expensive, so the production cost of the pomegranate juice lactic acid bacteria fermented beverage is high.
At present, few fermentation equipment for recovering strains are available, and the fermentation equipment is suitable for pomegranate juice fermentation liquor.
The invention content is as follows:
the invention aims to provide a method for recovering lactic acid bacteria strains, which can be repeatedly used for a plurality of times and reduce the production cost of pomegranate juice lactic acid bacteria fermented beverage.
In order to achieve the purpose, the technical scheme of the invention is as follows:
the invention relates to a fermentation column capable of recovering lactic acid bacteria in fermentation liquor, which comprises,
the fermentation column is of a hollow cylindrical structure, the top of the fermentation column is provided with an opening, the bottom of the fermentation column is provided with a closed end, and the surface of the cylindrical structure is fully distributed with vent holes;
the inner part of the semipermeable membrane is wrapped with strain fermentation liquor, the aperture of the semipermeable membrane limits the free inlet and outlet of lactic acid bacteria, and the semipermeable membrane is placed in the hollow structure of the fermentation column after wrapping the strain fermentation liquor;
the sealing plug blocks the top opening of the fermentation column, a ventilation channel is arranged in the middle of the sealing plug, a one-way sealing structure is arranged in the ventilation channel, and the ventilation channel is communicated with the semipermeable membrane to wrap the inner structure and the external natural air.
Preferably, the semi-permeable membrane is bag-shaped and is made of a dialysis membrane material, the top opening of the semi-permeable membrane is outwards sleeved on the opening edge of the fermentation column, and the closing plug is plugged into the top opening of the semi-permeable membrane.
Further preferred, the closing plug is for having elastic food grade silica gel material, and the ventilation channel is connected with one-way seal structure, and one-way seal structure is the U-shaped pipe, and the air inlet and the gas outlet of the UNICOM ventilation channel of U-shaped pipe have been irritated to the U-shaped intraductal confining liquid that has irritated, and fermentation gas diffuses to outside natural air in from the pellicle of bag form, otherwise sealed.
Further preferably, the fermentation column is made of food-grade stainless steel, and the fixing piece is arranged on the column body and is fixedly used for fixing the column body and the fermentation equipment for placing the fermentation column.
The invention relates to a fermentation tank with a fermentation column, which is a tank body made of food-grade stainless steel, wherein the fermentation column is placed at an opening at the top, a fixing structure is arranged in the fermentation column and the tank body, the fermentation column is fixed in the tank body by the fixing structure without moving, and pomegranate juice is in the tank body.
Preferably, the fixed structure is a rubidium magnet, a group of rubidium magnets is arranged at the bottom end inside the fermentation column and at the bottom inside the fermentation tank respectively, the fermentation column is placed in the fermentation tank, then the two groups of rubidium magnets attract and fix, the fermentation column is fixed in the fermentation tank, a sealing cover is arranged above a sealing plug at the top of the fermentation column, the sealing cover completely seals the top opening of the fermentation tank, and the top opening of the sealing cover is connected with the U-shaped pipe.
The method for fermenting pomegranate juice by using the fermentation tank comprises the following steps of (1) cultivating plant lactobacillus strains,
selecting a single colony which is full in growth on a sterile table, culturing the single colony in a sterile broth culture medium at 37 ℃ for 48 h, standing for anaerobic culture, removing the upper clear liquid of the culture solution on the sterile table after the culture is finished, centrifugally enriching the lower thallus precipitate of the culture solution, re-suspending and washing the thallus by using sterile water, centrifugally collecting, and centrifugally concentrating under the centrifugal condition of 3000 rpm at room temperature.min-1And 5 min, repeatedly washing the bacteria for three times, removing the components of the culture medium, and finally weighing the lactobacillus plantarum bacteria of which the supernatant is removed to obtain a large number of lactobacillus plantarum strains;
(2) the fermentation tank is sterilized, and then the fermentation tank is sterilized,
cleaning the interior of the fermentation tank, pouring edible alcohol into the fermentation tank for sterilization, pumping out the alcohol in the fermentation tank after sterilization, pouring sterile water for washing for a plurality of times until no alcohol taste exists in the fermentation tank, and ending the sterilization and cleaning process in the fermentation tank;
(3) pomegranate juice canning
Peeling off fresh pomegranate seeds, squeezing pomegranate juice from the pomegranate seeds by using clean gauze, ensuring that the pomegranate seeds cannot be damaged, and using food-grade Na2CO3Adjusting the pH value of the pomegranate juice to be 7.5-8.0, then heating the pomegranate juice to slightly boil, skimming froth, putting the pomegranate juice into a fermentation tank while the pomegranate juice is hot, reserving one tenth of space in the fermentation tank, then properly sealing the fermentation tank, and carrying out the next step of operation when the pomegranate juice is cooled to 37 ℃;
(4) starting fermentation
Adding centrifuged lactobacillus plantarum thalli, wherein the addition amount of the thalli is 1 per mill of the weight of fermentation liquor, then adding sterile water with the volume of one third of that of a dialysis bag, fixing the dialysis bag inside a fermentation column, and finally slowly placing the fermentation column into a fermentation tank, wherein redundant gas in the dialysis bag can be slowly discharged through a ventilation pipeline of the fermentation column at the moment, the liquid level of the fermentation tank cannot exceed the lower part of the ventilation pipeline of the fermentation column, and finally assembling the rest parts of the fermentation tank, including a sealing cover and a U-shaped pipe connected with the ventilation pipeline, wherein an appropriate volume of inert liquid needs to be added into the pipe, so that the pomegranate juice lactobacillus plantarum fermentation is formally started.
(5) Controlling fermentation, wherein in the fermentation process, the fermentation temperature is controlled at 18-20 ℃, the fermentation is carried out in a dark place and in an anaerobic fermentation mode, the fermentation time in a tank is 20 days, and after the fermentation is finished, the pH value, the sweetness, the number, the color and the sensory evaluation of lactic acid bacteria of fermentation liquids of different fermentation tanks are detected to obtain pomegranate juice fermentation liquid;
(6) product filling and storage
After fermentation, transferring the product into an aseptic container, storing in a dark and sealed condition at 4 ℃, and carrying out aseptic filling to obtain the product;
(7) the lactobacillus in the fermentation column is recovered and reused, and when the pomegranate juice in the fermentation tank is fermented, the fermentation column can be partially disassembled to another pomegranate juice fermentation tank for continuous fermentation, so that the lactobacillus is recovered and reused.
Preferably, the pomegranate juice fermentation method comprises cutting a semi-transparent MD77 dialysis bag in a fermentation column, cutting a MD77 dialysis bag with a bottom seal, cutting to a length consistent with the height of the fermentation column, boiling in distilled water for 1 hr for sufficiently sterilizing, and taking out under aseptic condition for reuse.
As a preferred method for pomegranate juice fermentation, step 1) is to cultivate lactobacillus plantarum strain, pick lactobacillus plantarum on a sterile table with an inoculating loop, put into a bacterial incubator for inverted culture, and streak on MRS agar medium plate. And (3) performing standing anaerobic culture at 37 ℃ for 48 h by using a parafilm sealing plate, and then picking a single colony which grows fully into a sterile MRS broth culture medium for culture.
The preferable step 2) as the method for pomegranate juice fermentation is sterilization of the fermentation tank,
pouring edible alcohol into the fermentation tank, wherein the alcohol concentration is 75%, the alcohol liquid level is required to completely submerge the tank opening, simultaneously sealing the tank opening, standing for 72 h at room temperature, completing the sterilization process of the fermentation tank, extracting the alcohol in the fermentation tank, pouring sterile water for washing for a plurality of times until no alcohol taste exists in the tank, taking out the fermentation column structure and placing in a sterile environment for later use.
The invention relates to a fermentation column, a fermentation tank and a fermentation method for recyclable lactic acid bacteria. Along with the fermentation, the pomegranate juice slowly permeates into the dialysis bag, then the lactic acid bacteria carry out anaerobic fermentation to the effective component in the pomegranate juice in the dialysis bag, and the fermentation product passes through the dialysis bag through the free diffusion mode and gets into outside zymotic fluid, but the lactic acid bacteria can not freely get into the dialysis bag. The method confines the lactic acid bacteria species within the dialysis bag. And (3) when the fermentation is finished, adding lactic acid bacteria into the dialysis bag with one sealed end, immersing part of the dialysis bag into the pomegranate juice, and fixing the dialysis bag in a certain mode to suspend the dialysis bag in the pomegranate juice. The dialysis bag in the fermentation tank can be transferred into a new fermentation tank, so that the recycling of the lactic acid bacteria is realized, and the production cost is reduced.
Drawings
FIG. 1 is a structural view of a fermentation column of the present invention,
FIG. 2 is a schematic view of a fermenter according to the present invention.
Reference numerals: 1. fermentation column, 2, vent hole, 3, semipermeable membrane, 4, strain fermentation liquor, 5, sealing plug, 6, vent channel, 7, U-shaped pipe, 8, sealing liquid, 9, fermentation tank, 10 rubidium magnet in the fermentation column, 11, sealing cover, 12, air filter opening, 13 and rubidium magnet in the tank.
Detailed Description
The following is a detailed description of the present invention:
the fermentation column capable of recovering the lactic acid bacteria in the fermentation liquid has the structure shown in figure 1 and comprises the fermentation column 1, a semi-permeable membrane 3 and a closing plug 5.
As shown in fig. 1, the fermentation column 1 is a hollow cylindrical structure, the top of the fermentation column is provided with an opening, the bottom of the fermentation column is provided with a closed end, and the surface of the cylindrical structure is fully distributed with vent holes 2; the semi-permeable membrane 3 is internally wrapped with a strain fermentation liquid 4, the aperture of the semi-permeable membrane 3 limits the free inlet and outlet of lactic acid bacteria, the semi-permeable membrane 3 is wrapped with the strain fermentation liquid 4 and then placed in the hollow structure of the fermentation column 1, and the opening is upward; the top opening of the fermentation column 1 is blocked by the closing plug 5, the middle part of the closing plug 5 is provided with a ventilation channel 6, the ventilation channel 6 is provided with a one-way sealing structure, and the ventilation channel 6 is communicated with the semipermeable membrane 3 to wrap the inner structure and the external natural air.
The semi-permeable membrane 3 is bag-shaped and is made of a dialysis membrane material, the top opening of the semi-permeable membrane is outwards sleeved on the opening edge of the fermentation column 1, and the closing plug 5 is plugged into the top opening of the semi-permeable membrane 3. The semipermeable membrane can isolate lactobacillus from fermentation liquid during pomegranate juice fermentation, and can recover lactobacillus after fermentation,
the sealing plug 5 is made of elastic food-grade silica gel, the ventilation channel 6 is connected with a one-way sealing structure, the one-way sealing structure is a U-shaped tube 7, one end of the U-shaped tube 7 is connected with an air outlet of the ventilation channel 6, the other end of the U-shaped tube 7 is provided with an air filtering port 12, sealing liquid 8 is filled in the U-shaped tube 7, and fermentation gas is diffused into external natural air from the bag-shaped semipermeable membrane 3, otherwise, the sealing structure is sealed.
The plastics piston can firmly fix the dialysis bag for possessing certain elasticity, and the central part of plastics piston is ventilated 6 and is stainless steel's diameter 2 mm's tubule way, and the fermentation gas in the dialysis bag can be through the quick discharge of tubule way for the inside and outside atmospheric pressure of fermentation post is balanced, avoids the interior gas output of dialysis bag too much and leads to the dialysis bag flatulence.
The fermentation column 1 is made of food-grade stainless steel materials, a fixing part is arranged on the column body, and the fixing part is fixedly used for fixing the column body and fermentation equipment for placing the fermentation column.
The fermentation tank 9 of the fermentation column of the invention has the structure shown in figure 2, the fermentation tank 9 is a tank body made of food-grade stainless steel, the fermentation column is placed at the top opening, the fermentation column and the tank body are internally provided with a fixing structure, the fermentation column is fixed in the tank body by the fixing structure and does not move, and pomegranate juice is in the tank body.
Fixed knot constructs for rubidium magnet 10, and the inside bottom of fermentation post 1 and the bottom in the fermentation cylinder 9 are equipped with a set of rubidium magnet 10 respectively, and two sets of rubidium magnet 10 attract behind fermentation post 1 puts into fermentation cylinder 9 fixedly, fix fermentation post 1 in fermentation cylinder 9, and the closing plug 5 top at fermentation top is equipped with sealed lid 11, and sealed lid 11 is sealed the open-top of fermentation cylinder 9 completely, and the open-top of sealed lid 11 is connected the U-shaped pipe 7. The U-shaped pipe device can ensure the one-way flow of gas and prevent suck-back, and the air filter port can ensure that external microorganisms cannot enter the U-shaped pipe device, thereby ensuring the safety of the whole fermentation device.
The fermentation column is connected with the fermentation tank through a rubidium magnet, and when the pomegranate juice in the fermentation tank is fermented, the fermentation column can be detached into another pomegranate juice fermentation tank for continuous fermentation. The outside of the rubidium magnet is wrapped by a layer of food-grade stainless steel material, so that the fermentation liquor environment can be prevented from being polluted to the maximum extent.
The method for fermenting pomegranate juice by using the fermentation main body and the fermentation tank comprises three specific operation embodiments,
example 1
Production of materials:
fresh mature sweet pomegranate, gauze, lactobacillus plantarum strain: (Lactobacillus plantarum) (Dalian Suzhou core Biotechnology Co., Ltd.), MD77 dialysis bag (MW: 8000-MRS agar medium (Kyoto Kyork Microscience Co., Ltd.), sterile distilled water, edible alcohol, and food-grade Na2CO3。
The production device comprises:
such as the fermentation column of figure 1 and the fermenter of figure 2.
The production scheme is as follows:
(1) cultivating plant lactobacillus strain
On a sterile table, lactobacillus plantarum was picked with an inoculating loop and streaked on MRS agar medium plates. Placing the culture medium into a bacteria incubator for inverted culture by using a parafilm sealing plate under the culture condition of 37 ℃ and 48 h, and standing for anaerobic culture.
And (3) on a sterile table, picking the plump single colony to be cultured in a sterile MRS broth culture medium under the culture condition of standing anaerobic culture at 37 ℃ for 48 h. After the culture, removing the upper clear liquid of the culture solution on a sterile table, centrifuging to enrich the thallus precipitate at the lower part of the culture solution, using sterile water to resuspend and wash the thallus, centrifuging and collecting at room temperature and 3000 rpm.min-1And 5 min. And repeatedly washing the bacteria for three times, removing the components of the culture medium as much as possible, and finally weighing the lactobacillus plantarum bacteria of which the supernatant is removed to finally obtain a large number of lactobacillus plantarum strains.
(2) Sterilization of fermentation tank
Cleaning and assembling the components in the fermentation tank (at the moment, the semi-permeable membrane cannot be fixed in the fermentation tank), pouring edible alcohol (with alcohol concentration of 75%) into the fermentation tank, and sealing the tank opening while the alcohol liquid level completely submerges the tank opening. Standing for 72 h at room temperature to finish the sterilization process of the fermentation tank. Pumping out alcohol from the fermentation tank, pouring sterile water for washing for several times until no alcohol smell exists in the fermentation tank, taking out the fermentation column structure, placing the fermentation column structure in a sterile environment for later use, and ending the sterilization and cleaning process in the fermentation tank.
(3) Pomegranate juice canning
Peeling off fresh pomegranate seeds, squeezing pomegranate juice in the pomegranate seeds by using clean gauze, and simultaneously ensuring that the pomegranate seeds cannot be damaged, otherwise, the outflow of the seed contents can cause the pomegranate juice fermentation process to generate peculiar smell. Using food-grade Na2CO3Adjusting the pH value of the pomegranate juice to be 7.5-8.0, then heating the pomegranate juice to slightly boil, skimming froth, putting the pomegranate juice into a fermentation tank while the pomegranate juice is hot, reserving one tenth of space in the fermentation tank, then properly sealing the fermentation tank, and carrying out the next step of operation when the pomegranate juice is cooled to 37 ℃.
(5) Starting fermentation
Directly adding lactobacillus plantarum thallus with the mass of 1 per mill of the fermentation liquid into pomegranate juice in a fermentation tank, uniformly stirring, and immediately assembling the fermentation tank for fermentation (a fermentation column device does not participate in the assembly process), wherein the fermentation mode is direct-vat lactic acid bacteria fermentation.
Cutting MD77 dialysis bag with bottom sealed, cutting to length consistent with height of fermentation column, and boiling in distilled water for 1 hr for sufficient sterilization. Taking out under aseptic condition, adding centrifuged Lactobacillus plantarum thallus, wherein the addition amount of the thallus is 1 per mill of the weight of the fermentation liquid. Sterile water was then added to the dialysis bag in one third of its volume and the bag was fixed inside the column. Finally, the fermentation column is slowly placed into the fermentation tank, redundant gas in the dialysis bag can be slowly discharged through the fermentation column vent pipe, and the liquid level of the fermentation tank cannot submerge the lower part of the fermentation column vent pipe. And finally, assembling the rest parts of the fermentation tank, including a sealing cover, a U-shaped pipe (inert liquid with a proper volume needs to be added in the pipe), and an air filtering hole. So that the pomegranate juice lactobacillus plantarum fermentation is formally started.
In the fermentation process, the fermentation temperature is controlled at 18-20 ℃, light is avoided, anaerobic fermentation is carried out, and the fermentation time in a tank is 20 days. And (4) detecting the pH value, sweetness, lactobacillus quantity, color and sense evaluation of fermentation liquor of different fermentation tanks after the fermentation is finished, and comparing and analyzing the pH value, sweetness, lactobacillus quantity, color and sense evaluation with relevant index data of the direct vat lactobacillus fermentation liquor.
(5) Product filling and storage
After fermentation, transferring the product into an aseptic container, storing in a dark and sealed condition at 4 ℃, and carrying out aseptic filling to obtain the finished product.
When the pomegranate juice fermentation in the fermentation tank is finished, the fermentation column can be disassembled to another pomegranate juice fermentation tank for continuous fermentation.
Example 2
Production of materials:
fresh pomegranate fruit, sour pomegranate fruit, gauze, lactobacillus plantarum strain: (Lactobacillus plantarum) (Dalian Suiya Biotechnology Co., Ltd.), MD77 dialysis bag (MW: 8000-2CO3。
The production device comprises:
such as the fermentation column of figure 1 and the fermenter of figure 2.
The production scheme is as follows:
(1) cultivation of Lactobacillus plantarum strains
On a sterile table, plant lactobacillus thalli are picked up by using an inoculating loop and streaked on an MRS agar culture medium plate. Placing the culture medium into a bacteria incubator for inverted culture by using a parafilm sealing plate under the culture condition of 37 ℃ and 48 h, and standing for anaerobic culture.
And (3) on a sterile table, picking the plump single colony to be cultured in a sterile MRS broth culture medium under the culture condition of standing anaerobic culture at 37 ℃ for 48 h. After the culture, removing the upper clear liquid of the culture solution on a sterile table, centrifuging to enrich the thallus precipitate at the lower part of the culture solution, using sterile water to resuspend and wash the thallus, centrifuging and collecting at room temperature and 3000 rpm.min-1And 5 min. And repeatedly washing the bacteria for three times, removing the components of the culture medium as much as possible, and finally weighing the lactobacillus plantarum bacteria of which the supernatant is removed to finally obtain a large number of lactobacillus plantarum strains.
(2) Sterilization of fermentation tank
Cleaning and assembling the components in the fermentation tank (at the moment, the semi-permeable membrane cannot be fixed in the fermentation tank), pouring edible alcohol (with alcohol concentration of 75%) into the fermentation tank, and sealing the tank opening while the alcohol liquid level completely submerges the tank opening. Standing for 72 h at room temperature to finish the sterilization process of the fermentation tank. Pumping out alcohol from the fermentation tank, pouring sterile water for washing for several times until no alcohol smell exists in the fermentation tank, taking out the fermentation column structure, placing the fermentation column structure in a sterile environment for later use, and ending the sterilization and cleaning process in the fermentation tank.
(3) Pomegranate juice canning
The method comprises the steps of respectively peeling off fresh Dahongpao sweet pomegranate and sour pomegranate seeds, respectively squeezing pomegranate juice in the pomegranate seeds by using clean gauze, and simultaneously ensuring that the pomegranate seeds cannot be damaged, otherwise, the outflow of seed contents can cause peculiar smell in the pomegranate juice fermentation process. Using food-grade Na2CO3Adjusting the pH value of the pomegranate juice to be 7.5-8.0, then heating the pomegranate juice to slightly boil, skimming floating foam, respectively filling the sweet pomegranate juice and the sour pomegranate juice into a fermentation tank while the pomegranate juice is hot, reserving one tenth of space in the fermentation tank, then properly sealing the fermentation tank, and carrying out next operation when the cold energy of the pomegranate juice reaches 37 ℃.
(4) Starting fermentation
Cutting MD77 dialysis bag with bottom sealed, cutting to length consistent with height of fermentation column, and boiling in distilled water for 1 hr for sufficient sterilization. Taking out under aseptic condition, adding centrifuged Lactobacillus plantarum thallus with the addition amount of 1 ‰ of the fermentation broth. Sterile water was then added to the dialysis bag in one third of its volume and the bag was fixed inside the column. Finally, the fermentation column is slowly placed into the fermentation tank, redundant gas in the dialysis bag can be slowly discharged through the fermentation column vent pipe, and the liquid level of the fermentation tank cannot submerge the lower part of the fermentation column vent pipe. And finally, assembling the rest parts of the fermentation tank, including a sealing cover, a U-shaped pipe (inert liquid with a proper volume needs to be added in the pipe), and an air filtering hole. So that the pomegranate juice lactobacillus plantarum fermentation is formally started. In the fermentation process, the fermentation temperature is controlled at 18-20 ℃, the fermentation is carried out in a dark place and under anaerobic condition, and the fermentation time in the tank is 20 days. And after the fermentation is finished, respectively detecting the pH value, sweetness and lactic acid bacteria quantity of the fermentation liquor of the sweet pomegranate juice and the sour pomegranate juice, and simultaneously carrying out corresponding sensory evaluation. So as to obtain the pomegranate juice variety most suitable for lactobacillus plantarum fermentation.
(5) Product filling and storage
After fermentation, transferring the product into an aseptic container, storing in a dark and sealed condition at 4 ℃, and carrying out aseptic filling to obtain the finished product.
When the pomegranate juice fermentation in the fermentation tank is finished, the fermentation column can be disassembled to another pomegranate juice fermentation tank for continuous fermentation.
Example 3
Production of materials:
fresh sweet pomegranate (selected from Dahongpao pomegranate variety), gauze, Lactobacillus plantarum strain: (A)Lactobacillus plantarum) (Dalian Suiya Biotechnology Co., Ltd.), MD77 dialysis bag (MW: 8000-2CO3。
The production device comprises: such as the fermentation column of figure 1 and the fermenter of figure 2.
The production scheme is as follows:
(1) cultivation of Lactobacillus plantarum strains
On a sterile table, lactobacillus plantarum strains were picked with an inoculating loop and streaked on a MRS agar medium plate. Placing the culture medium into a bacteria incubator for inverted culture by using a parafilm sealing plate under the culture condition of 37 ℃ and 48 h, and standing for anaerobic culture. And (3) on a sterile table, picking the plump single colony to be cultured in a sterile MRS broth culture medium under the culture condition of standing anaerobic culture at 37 ℃ for 48 h. After the culture, removing the upper clear liquid of the culture solution on a sterile table, centrifuging to enrich the thallus precipitate at the lower part of the culture solution, using sterile water to resuspend and wash the thallus, centrifuging and collecting at room temperature and 3000 rpm.min-1And 5 min. And repeatedly washing the bacteria for three times, removing the components of the culture medium as much as possible, and finally weighing the lactobacillus plantarum bacteria of which the supernatant is removed to finally obtain a large number of lactobacillus plantarum strains.
(2) Sterilization of fermentation tank
Cleaning and assembling the components in the fermentation tank (at the moment, the semi-permeable membrane cannot be fixed in the fermentation tank), pouring edible alcohol (with alcohol concentration of 75%) into the fermentation tank, and sealing the tank opening while the alcohol liquid level completely submerges the tank opening. Standing for 72 h at room temperature to finish the sterilization process of the fermentation tank. Pumping out alcohol from the fermentation tank, pouring sterile water for washing for several times until no alcohol smell exists in the fermentation tank, taking out the fermentation column structure, placing the fermentation column structure in a sterile environment for later use, and ending the sterilization and cleaning process in the fermentation tank.
(3) Pomegranate juice canning
The fresh pomegranate seeds are stripped, the pomegranate juice in the pomegranate seeds is squeezed by using clean gauze, meanwhile, the pomegranate seeds are ensured not to be damaged, otherwise, the outflow of the seed contents can cause the peculiar smell generated in the pomegranate juice fermentation process. Using food-grade Na2CO3Adjusting the pH value of the pomegranate juice to be 7.5-8.0, then heating the pomegranate juice to slightly boil, skimming floating foam, filling the pomegranate juice into a fermentation tank while the pomegranate juice is hot, filling the three fermentation tanks, reserving one tenth of space in the fermentation tank, then properly sealing the fermentation tank, and carrying out the next operation when the cold energy of the pomegranate juice reaches 37 ℃.
(4) Starting fermentation
Cutting MD77 dialysis bag with bottom sealed, cutting to length consistent with height of fermentation column, and boiling in distilled water for 1 hr for sufficient sterilization. Taking out under aseptic condition, adding centrifuged Lactobacillus plantarum thallus with the addition amount of 2 per mill of the fermentation broth. Sterile water was then added to the dialysis bag in one third of its volume and the bag was fixed inside the column. Finally, the fermentation column is slowly placed into the fermentation tank, redundant gas in the dialysis bag can be slowly discharged through the fermentation column vent pipe at the moment, and the liquid level of the fermentation tank cannot exceed the lower part of the fermentation column vent pipe. And finally, assembling the rest parts of the fermentation tank, including a sealing cover, a U-shaped pipe (inert liquid with a proper volume needs to be added in the pipe), and an air filtering hole. So that the pomegranate juice lactobacillus plantarum fermentation is formally started. In the fermentation process, the fermentation temperature is controlled at 18-20 ℃, the fermentation is carried out in a dark place and under anaerobic condition, and the fermentation time in the tank is 20 days. And (3) respectively detecting the pH value, sweetness and lactic acid bacteria quantity of fermentation liquor of different lactobacillus plantarum inoculation amounts after fermentation is finished, and simultaneously carrying out corresponding sensory evaluation. So as to obtain the best lactobacillus plantarum inoculation amount.
(5) Product filling and storage
After fermentation, transferring the product into an aseptic container, storing in a dark and sealed condition at 4 ℃, and carrying out aseptic filling to obtain the finished product.
When the pomegranate juice fermentation in the fermentation tank is finished, the fermentation column can be disassembled to another pomegranate juice fermentation tank for continuous fermentation.
The invention relates to a fermentation column and a fermentation tank of pomegranate juice lactic acid bacteria capable of recovering lactic acid bacteria, wherein the lactic acid bacteria is added into a dialysis bag with one end sealed, part of the dialysis bag is immersed into pomegranate juice, and the dialysis bag is fixed in a certain way and is suspended in the pomegranate juice. Along with the fermentation, the pomegranate juice slowly permeates into the dialysis bag, then the lactic acid bacteria carry out anaerobic fermentation to the effective component in the pomegranate juice in the dialysis bag, and the fermentation product passes through the dialysis bag through the free diffusion mode and gets into outside zymotic fluid, but the lactic acid bacteria can not freely get into the dialysis bag. The method confines the lactic acid bacteria species within the dialysis bag. When the fermentation is finished, the dialysis bag in the fermentation tank can be switched into a new fermentation tank, so that the repeated utilization of the lactic acid bacteria is realized, and the production cost is reduced.
Claims (10)
1. A fermentation column capable of recovering lactic acid bacteria in fermentation liquor is characterized by comprising,
the fermentation column (1) is of a hollow cylindrical structure, the top of the fermentation column is provided with an opening, the bottom of the fermentation column is provided with a closed end, and the surface of the cylindrical structure is fully distributed with vent holes (2);
the inner part of the semipermeable membrane (3) is wrapped with strain fermentation liquor (4), the aperture of the semipermeable membrane (3) limits the free access of lactic acid bacteria, and the semipermeable membrane (3) is wrapped with the strain fermentation liquor (4) and then placed in the hollow structure of the fermentation column (1);
the top opening of the fermentation column (1) is blocked by the sealing plug (5), the middle part of the sealing plug (5) is provided with a ventilation channel (6), the ventilation channel (6) is provided with a one-way sealing structure, and the ventilation channel (6) is communicated with the semipermeable membrane (3) to wrap the inner structure and the external natural air.
2. A fermentation column capable of recovering lactic acid bacteria in a fermentation broth according to claim 1, wherein the semi-permeable membrane (3) is in the form of a bag made of a dialysis membrane material, the top opening is a type of material that is outwardly fitted over the opening edge of the fermentation column (1), and the closing plug (5) is inserted into the top opening of the semi-permeable membrane (3).
3. The fermentation column capable of recovering lactic acid bacteria in fermentation liquid according to claim 2, wherein the sealing plug (5) is made of elastic food grade silica gel, the ventilation channel (6) is connected with a one-way sealing structure, the one-way sealing structure is a U-shaped tube (7), one end of the U-shaped tube (7) is connected with the air outlet of the ventilation channel (6), the other end of the U-shaped tube is provided with an air filtering port (12), the U-shaped tube (7) is filled with sealing liquid (8), and the fermentation gas diffuses from the bag-shaped semipermeable membrane (3) to the external natural air, otherwise, the fermentation column is sealed.
4. The fermentation column capable of recovering lactic acid bacteria in fermentation broth according to claim 2, wherein the fermentation column (1) is made of food grade stainless steel, and the column body is provided with a fixing member for fixing the column body to the fermentation equipment on which the fermentation column is placed.
5. A fermentation tank (9) provided with a fermentation column according to claim 3, characterized in that the fermentation tank (9) is a tank made of food grade stainless steel, the top opening is used for placing the fermentation column, and the fermentation column and the tank are internally provided with a fixing structure which fixes the fermentation column in the tank without moving, and the tank is internally provided with pomegranate juice.
6. The fermentation tank (9) according to claim 5, characterized in that the fixed structure is a rubidium magnet (10), the bottom end inside the fermentation column (1) and the bottom inside the fermentation tank (9) are respectively provided with a group of rubidium magnets (10), the two groups of rubidium magnets (10) attract and fix the fermentation column (1) after the fermentation column (1) is placed in the fermentation tank (9), the fermentation column (1) is fixed in the fermentation tank (9), a sealing cover (11) is arranged above the sealing plug (5) at the top of the fermentation column, the sealing cover (11) completely seals the top opening of the fermentation tank (9), and the top opening of the sealing cover (11) is connected with the U-shaped pipe (7).
7. A method of pomegranate juice fermentation in the fermenter of claim 5, comprising the steps of (1) cultivating the Lactobacillus plantarum strain,
on a sterile platform, picking and culturing the plump single bacterial colony in a sterile broth culture medium under the conditions of 37 ℃ and 48 h, standing for anaerobic culture, discarding the supernatant of the culture solution on the sterile platform after the culture is finished, centrifugally enriching the bacterial sediment at the lower part of the culture solution, re-suspending and washing the bacterial with sterile water, centrifugally collecting, and centrifugally collecting under the centrifugal conditions of room temperature and 3000 rpm.min-1And 5 min, repeatedly washing the bacteria for three times, removing the components of the culture medium, and finally weighing the lactobacillus plantarum bacteria of which the supernatant is removed to obtain a large number of lactobacillus plantarum strains;
(2) the fermentation tank (9) is sterilized,
cleaning the interior of the fermentation tank (9), pouring edible alcohol into the fermentation tank (9) for sterilization, extracting the alcohol in the fermentation tank (9) after sterilization, pouring sterile water for washing for a plurality of times until no alcohol taste exists in the tank, and ending the sterilization and cleaning process in the fermentation tank (9);
(3) pomegranate juice canning
Peeling off fresh pomegranate seeds, squeezing pomegranate juice from the pomegranate seeds by using clean gauze, ensuring that the pomegranate seeds cannot be damaged, and using food-grade Na2CO3Adjusting the pH value of the pomegranate juice to be 7.5-8.0, then heating the pomegranate juice to slightly boil, skimming froth, putting the pomegranate juice into a fermentation tank (9) while the pomegranate juice is hot, reserving one tenth of space in the fermentation tank (9), then properly sealing the fermentation tank (9), and carrying out the next operation when the pomegranate juice is cooled to 37 ℃;
starting fermentation
Adding the centrifuged lactobacillus plantarum thalli, wherein the addition amount of the thalli is 1 per mill of the weight of fermentation liquor, then adding sterile water with one third of the volume of a dialysis bag, fixing the dialysis bag in a fermentation column, finally slowly putting the fermentation column into a fermentation tank (9), slowly discharging redundant gas in the dialysis bag through a fermentation column vent pipe at the moment, ensuring that the liquid level of the fermentation tank (9) cannot exceed the lower part of the fermentation column vent pipe, finally assembling the rest parts of the fermentation tank (9), including a sealing cover (11) and a U-shaped pipe (7) connected with the vent pipe, and adding an appropriate volume of inert liquid into the pipe, so that the pomegranate juice lactobacillus plantarum fermentation is formally started;
(5) controlling fermentation, wherein in the fermentation process, the fermentation temperature is controlled at 18-20 ℃, the fermentation is carried out in a dark place and in an anaerobic fermentation mode, the fermentation time in the tank is 20 days, and after the fermentation is finished, the pH value, the sweetness, the quantity and the color of lactic acid bacteria and sensory evaluation of fermentation liquor of different fermentation tanks (9) are detected to obtain pomegranate juice fermentation liquor;
(6) product filling and storage
After fermentation, transferring the product into an aseptic container, storing in a dark and sealed condition at 4 ℃, and carrying out aseptic filling to obtain the product;
(7) the lactic acid bacteria in the fermentation column are recovered and reused, when the pomegranate juice in the fermentation tank (9) is fermented, the fermentation column can be partially disassembled into another pomegranate juice fermentation tank (9) to continue fermentation, and the purpose of recovering and reusing the lactic acid bacteria is achieved.
8. The method of claim 7, wherein the fermentation column is translucent and is a MD77 dialysis bag, the MD77 dialysis bag is cut and sealed at the bottom, the length of the cut MD77 dialysis bag is identical to the height of the fermentation column, the MD77 dialysis bag is placed in distilled water and boiled for 1 hour for sufficient sterilization, and the MD77 dialysis bag is taken out under aseptic conditions and is reused.
9. The method for fermenting pomegranate juice according to claim 7, wherein the lactobacillus plantarum strain is cultured in step 1), lactobacillus plantarum is picked up by using an inoculating loop on a sterile table, placed in a bacteria incubator for inverted culture, streaked on a MRS agar medium plate, and subjected to anaerobic culture by using a parafilm seal plate under a standing condition of 37 ℃ for 48 hours, and then a single colony which grows well is picked up and cultured in sterile MRS broth.
10. The method of fermentation of pomegranate juice according to claim 7, wherein the fermentation tank (9) is sterilized in step 2), edible alcohol with an alcohol concentration of 75% is poured into the fermentation tank (9) and the alcohol level is completely submerged in the tank opening, the tank opening is sealed and allowed to stand at room temperature for 72 hours, the sterilization process of the fermentation tank (9) is completed, the alcohol in the fermentation tank (9) is extracted, sterile water is poured into the tank for washing several times until there is no alcohol taste in the tank, and the fermentation column structure is taken out and placed in a sterile environment for use.
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