CN112795483A - Preparation method of lactobacillus fermentation lysate and cracking device - Google Patents
Preparation method of lactobacillus fermentation lysate and cracking device Download PDFInfo
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Abstract
The invention discloses a preparation method of lactobacillus fermentation lysate, which comprises the steps of putting a fermented lactobacillus culture solution into a high-pressure resistant container, introducing carbon dioxide and applying pressure to ensure that the carbon dioxide fully permeates into lactobacillus thallus, then conveying the lactobacillus culture solution into a pressure reduction container to quickly reduce the pressure, quickly expanding the carbon dioxide in the thallus, and bursting the thallus to release lactobacillus thallus content. The lactobacillus fermentation cracking liquid obtained by inputting the fermented lactobacillus culture liquid into the high-pressure tank, introducing carbon dioxide and applying pressure has the characteristics of high cracking rate, high inhibition rate, high polypeptide content and the like, compared with the traditional lactobacillus secretion and high-temperature expansion cracking liquid, the cracked thallus content is pollution-free and has higher biological activity content, so that the lactobacillus fermentation cracking liquid can be widely applied to skin care products, and has the effects of nourishing, whitening, resisting oxidation, removing wrinkles and the like.
Description
Technical Field
The invention belongs to the technical field of cosmetics, relates to a fermentation lysate, and particularly relates to a lactobacillus fermentation lysate prepared by a lactobacillus gas expansion pyrolysis method and a pyrolysis device thereof.
Background
Any bacteria that produce lactic acid from the fermentation of glucose or lactose are collectively referred to as lactic acid bacteria. This is a rather bulky group of bacteria, which can be currently divided into at least 18 genera, and a total of over 200. Most of them are, except for a very small number, flora which are indispensable to the human body and have important physiological functions, and are widely present in the intestinal tract of the human body. At present, as proved by biologists at home and abroad, the lactobacillus in the intestinal tract has a very close relationship with the health and longevity. Lactic acid bacteria can produce a large amount of small molecular peptides, organic acids and other substances with anti-inflammatory, whitening and moisturizing effects on skin. Usually, such substances are mostly preserved in the lactic acid bacteria and a small part is secreted out of the bacteria by secretion. The beneficial components in the lactobacillus can be released by crushing the lactobacillus, so that the yield of beneficial substances of the lactobacillus is greatly improved.
The cell body is broken, usually by using lysate, and is mainly used for eukaryotic cells without cell walls, such as yeast. Eukaryotic cell lysate generally consists of detergents, surfactants and salts, cell membranes are damaged, cell contents are released, but the obtained cell lysate contains lysate which has a damage or inhibition effect on cell active substances and influences the use effect; for lactic acid bacteria with cell walls, it is difficult to destroy the cells with the lysis solution under the protection of peptidoglycan cell walls. In the prior art, the lactobacillus thallus content can be obtained by swelling and demulsifying the lactobacillus thallus by adopting a spray high-temperature expansion method. However, high temperatures can destroy temperature-sensitive active ingredients, and although protective agents are added in the method, the protective agents can only protect a few active ingredients, and the protective effect is still limited. At the same time, the protective agent also inhibits the activity of the active.
Disclosure of Invention
Compared with the traditional lactic acid bacteria secretion and high-temperature swelling lysate, the cracked thallus content is pollution-free, has higher bioactivity content, can be widely used in skin care products, and has the effects of nourishing, whitening, resisting oxidation, removing wrinkles and the like.
In order to achieve the purpose, the technical scheme of the invention is as follows: provides a preparation method of lactobacillus fermentation lysate, wherein: putting the fermented lactobacillus culture solution into a high-pressure resistant container, introducing carbon dioxide and applying pressure to ensure that the carbon dioxide fully permeates into lactobacillus thallus, then conveying the lactobacillus thallus into a pressure reduction container to quickly reduce the pressure, quickly expanding the carbon dioxide in the thallus, and bursting the thallus, thereby releasing the content of the lactobacillus thallus.
Further, the pressure applied by introducing carbon dioxide is 0.5-100 MPa.
Further, the pressure applied by introducing carbon dioxide is 3-10 MPa.
Furthermore, the lactobacillus fermentation raw material contains pearl powder.
Further, the lactic acid bacteria culture solution is prepared by fermenting xylo-oligosaccharide, pearl powder, glucose, lactic acid bacteria, milk powder and water.
Further, the temperature of the high-pressure resistant container is adjusted to 0-5 ℃, the high-pressure resistant container is sealed and then pressurized, carbon dioxide is introduced, the pressure is increased to 0.5-100 MPa, the pressure and the temperature are stably kept for 5-10 hours, then the temperature is increased to 15-30 ℃, a discharge pipe at the bottom of the high-pressure resistant container is opened, and the fermented product in the high-pressure resistant container rapidly enters the pressure reduction container under pressure.
Further, the carbon dioxide gas may be changed to nitrogen, air, oxygen, methane, or the like.
Furthermore, the lactic acid bacteria are also suitable for the lysis of yeast, other eukaryotic and prokaryotic bacteria, biological cells and the like.
The invention also aims to provide a cracking device for preparing the lactobacillus fermentation lysate, which comprises a fermentation tank, a high-pressure resistant container and a pressure reduction container, wherein a stirrer is arranged in the fermentation tank, one end of the fermentation tank is connected with a constant temperature machine, the bottom of the fermentation tank is connected with the high-pressure resistant container through a discharge pipe, a temperature regulating device and a pressurizing device are arranged on the high-pressure resistant container, and a pipeline which can be filled with gas is arranged at the top of the high-pressure resistant container; the bottom of the high pressure resistant container is connected with the pressure reducing container through a discharge pipe, a centrifugal separator is arranged in the pressure reducing container, and a discharge hole is formed in the bottom of the pressure reducing container.
Further, the high-pressure resistant container is a high-pressure tank, and the pressure reduction container is a cracking tank.
Furthermore, an exhaust port is arranged at the top of the cracking tank, and a cyclone is arranged at the exhaust port.
According to the preparation method of the lactobacillus fermentation lysate, the lactobacillus fermentation lysate obtained by introducing carbon dioxide into the high-pressure tank and applying pressure has the characteristics of high lysis rate, high inhibition rate, high polypeptide content and the like, compared with the traditional lactobacillus secretion and high-temperature expansion lysate, the cracked thallus content is pollution-free, has higher biological activity content, can be widely applied to skin care products, and has the effects of nourishing, whitening, resisting oxidation, removing wrinkles and the like.
Drawings
FIG. 1 is a schematic view of the cracking apparatus of the present invention;
in the figure: 1. a thermostat; 2. a fermentation tank; 3. a cover; 4. a blender; 5. a pipeline into which gas can be introduced; 6. a cover; 7. a high-pressure tank; 8. a discharge port; 9. a pyrolysis tank; 10. a cyclone; 11. and (7) an exhaust port.
Detailed Description
Example 1
Culturing lactic acid bacteria: 3kg of xylo-oligosaccharide, 5kg of pearl powder, 6kg of glucose, 0.5kg of lactic acid bacteria (dry weight), 6kg of milk powder and 79.5kg of water. As shown in FIG. 1, the lid 3 was opened, the fermentation material was put into a fermenter 2 containing 200 liters of water, stirred uniformly by a stirrer 4, and kept at a constant temperature of 25 ℃ for 10 hours by a thermostat 1 to obtain 98.7kg of a fermented product. The fermented product enters a 200L high-pressure tank 7 through a discharge pipe, the high-pressure tank 7 is sealed through a cover 6, a temperature adjusting device and a pressurizing device are further arranged (both the temperature adjusting device and the pressurizing device are in the prior art), a pipeline 5 capable of introducing gas is arranged at the top of the high-pressure tank 7, and the pipeline 5 can introduce gas; the bottom of the high-pressure tank 7 is provided with a discharge pipe which is communicated with a cracking tank 9, the cracking tank 9 is provided with a centrifugal separator, the top of the cracking tank 9 is provided with an exhaust port 11, the exhaust port 11 is provided with a cyclone machine 10, and the bottom of the cracking tank 9 is provided with a discharge port 8.
The temperature of the high-pressure tank 7 is adjusted to 5 ℃, the high-pressure tank 7 is sealed and then pressurized, carbon dioxide is introduced, the pressure is increased to 5MPa, the pressure and the temperature are stably kept for 5h, then the temperature is increased to 25 ℃, a bottom discharge pipe is opened, and the fermented materials in the high-pressure tank 7 rapidly enter the cracking tank 9 under the pressure compression. The pressure difference between the cracking tank 9 and the high-pressure tank 7 is kept to be not less than 5MPa by pressurizing the high-pressure tank 7, and the material discharge speed in the high-pressure tank 7 is not lower than 1L/min. The cracking tank 9 is communicated with the outside through the exhaust port 11 and is not sealed, so that pressure is not formed; when the lactobacillus material in the high-pressure tank 7 is pressed by pressure and quickly enters the cracking tank 9 (pressure reducing container), the lactobacillus can be quickly cracked under the condition of quick pressure reduction, and the content of the lactobacillus can be released. The content is filtered and centrifuged to remove solid impurities, and 86.1kg of a mixture of lactobacillus fermentation liquor and lactobacillus lysate, which is lactobacillus fermentation lysate for short, can be obtained.
Example 2
Culturing lactic acid bacteria: 3kg of xylo-oligosaccharide, 5kg of pearl powder, 6kg of glucose, 0.5kg of lactic acid bacteria (dry weight), 6kg of milk powder and 79.5kg of water. As shown in FIG. 1, the lid 3 was opened, the fermentation material was put into a fermenter 2 containing 200 liters of water, stirred uniformly by a stirrer 4, and kept at a constant temperature of 25 ℃ for 10 hours by a thermostat 1 to obtain 98.7kg of a fermented product. The fermented product enters a 200L high-pressure tank 7 through a discharge pipe, the high-pressure tank 7 is sealed through a cover 6, a temperature adjusting device and a pressurizing device are further arranged (both the temperature adjusting device and the pressurizing device are in the prior art), a pipeline 5 capable of introducing gas is arranged at the top of the high-pressure tank 7, and the pipeline 5 can introduce gas; the bottom of the high-pressure tank 7 is provided with a discharge pipe which is communicated with a cracking tank 9, the cracking tank 9 is provided with a centrifugal separator, the top of the cracking tank 9 is provided with an exhaust port 11, the exhaust port 11 is provided with a cyclone machine 10, and the bottom of the cracking tank 9 is provided with a discharge port 8.
The temperature of the high-pressure tank 7 is adjusted to 3 ℃, the high-pressure tank 7 is sealed and then pressurized, carbon dioxide is introduced, the pressure is increased to 3MPa, the pressure and the temperature are stably kept for 5h, then the temperature is increased to 28 ℃, a bottom discharge pipe is opened, and the fermented materials in the high-pressure tank 7 rapidly enter the cracking tank 9 under the pressure compression. The pressure difference between the cracking tank 9 and the high-pressure tank 7 is kept to be not less than 3MPa by pressurizing the high-pressure tank 7, and the material discharge speed in the high-pressure tank 7 is not lower than 1L/min. The cracking tank 9 is communicated with the outside through the exhaust port 11 and is not sealed, so that pressure is not formed; when the lactobacillus material in the high-pressure tank 7 is pressed by pressure and quickly enters the cracking tank 9 (pressure reducing container), the lactobacillus can be quickly cracked under the condition of quick pressure reduction, and the content of the lactobacillus can be released. The content is filtered and centrifuged to remove solid impurities, and 83.5kg of a mixture of lactobacillus fermentation liquor and lactobacillus lysate, which is lactobacillus fermentation lysate for short, can be obtained.
Example 3
Culturing lactic acid bacteria: 3kg of xylo-oligosaccharide, 5kg of pearl powder, 6kg of glucose, 0.5kg of lactic acid bacteria (dry weight), 6kg of milk powder and 79.5kg of water. As shown in FIG. 1, the lid 3 was opened, the fermentation material was put into a fermenter 2 containing 200 liters of water, stirred uniformly by a stirrer 4, and kept at a constant temperature of 25 ℃ for 10 hours by a thermostat 1 to obtain 98.7kg of a fermented product. The fermented product enters a 200L high-pressure tank 7 through a discharge pipe, the high-pressure tank 7 is sealed through a cover 6, a temperature adjusting device and a pressurizing device are further arranged (both the temperature adjusting device and the pressurizing device are in the prior art), a pipeline 5 capable of introducing gas is arranged at the top of the high-pressure tank 7, and the pipeline 5 can introduce gas; the bottom of the high-pressure tank 7 is provided with a discharge pipe which is communicated with a cracking tank 9, the cracking tank 9 is provided with a centrifugal separator, the top of the cracking tank 9 is provided with an exhaust port 11, the exhaust port 11 is provided with a cyclone machine 10, and the bottom of the cracking tank 9 is provided with a discharge port 8.
The temperature of the high-pressure tank 7 is adjusted to 1 ℃, the high-pressure tank 7 is sealed and then pressurized, carbon dioxide is introduced, the pressure is increased to 10MPa, the pressure and the temperature are stably kept for 5h, then the temperature is increased to 30 ℃, a bottom discharge pipe is opened, and the fermented materials in the high-pressure tank 7 rapidly enter the cracking tank 9 under the pressure compression. The pressure difference between the cracking tank 9 and the high-pressure tank 7 is kept to be not less than 10MPa by pressurizing the high-pressure tank 7, and the material discharge speed in the high-pressure tank 7 is not lower than 1L/min. The cracking tank 9 is communicated with the outside through the exhaust port 11 and is not sealed, so that pressure is not formed; when the lactobacillus material in the high-pressure tank 7 is pressed by pressure and quickly enters the cracking tank 9 (pressure reducing container), the lactobacillus can be quickly cracked under the condition of quick pressure reduction, and the content of the lactobacillus can be released. The content is filtered and centrifuged to remove solid impurities, and 87.8kg of a mixture of the lactobacillus fermentation liquor and the lactobacillus lysate, which is called lactobacillus fermentation lysate for short, can be obtained.
Example 4
Culturing lactic acid bacteria: 3kg of xylo-oligosaccharide, 5kg of pearl powder, 6kg of glucose, 0.5kg of lactic acid bacteria (dry weight), 6kg of milk powder and 79.5kg of water. As shown in FIG. 1, the lid 3 was opened, the fermentation material was put into a fermenter 2 containing 200 liters of water, stirred uniformly by a stirrer 4, and kept at a constant temperature of 25 ℃ for 10 hours by a thermostat 1 to obtain 98.7kg of a fermented product. The fermented product enters a 200L high-pressure tank 7 through a discharge pipe, the high-pressure tank 7 is sealed through a cover 6, a temperature adjusting device and a pressurizing device are further arranged (both the temperature adjusting device and the pressurizing device are in the prior art), a pipeline 5 capable of introducing gas is arranged at the top of the high-pressure tank 7, and the pipeline 5 can introduce gas; the bottom of the high-pressure tank 7 is provided with a discharge pipe which is communicated with a cracking tank 9, the cracking tank 9 is provided with a centrifugal separator, the top of the cracking tank 9 is provided with an exhaust port 11, the exhaust port 11 is provided with a cyclone machine 10, and the bottom of the cracking tank 9 is provided with a discharge port 8.
The temperature of the high-pressure tank 7 is adjusted to 5 ℃, the high-pressure tank 7 is sealed and then pressurized, carbon dioxide is introduced, the pressure is increased to 1MPa, the pressure and the temperature are stably kept for 5h, then the temperature is increased to 25 ℃, a bottom discharge pipe is opened, and the fermented materials in the high-pressure tank 7 rapidly enter the cracking tank 9 under the pressure compression. The pressure difference between the cracking tank 9 and the high-pressure tank 7 is kept to be not less than 1MPa by pressurizing the high-pressure tank 7, and the material discharge speed in the high-pressure tank 7 is not lower than 1L/min. The cracking tank 9 is communicated with the outside through the exhaust port 11 and is not sealed, so that pressure is not formed; when the lactobacillus material in the high-pressure tank 7 is pressed by pressure and quickly enters the cracking tank 9 (pressure reducing container), the lactobacillus can be quickly cracked under the condition of quick pressure reduction, and the content of the lactobacillus can be released. The content is filtered and centrifuged to remove solid impurities, and 75.1kg of a mixture of the lactobacillus fermentation liquor and the lactobacillus lysate, which is called lactobacillus fermentation lysate for short, can be obtained.
Culturing lactic acid bacteria: 3kg of xylo-oligosaccharide, 5kg of pearl powder, 6kg of glucose, 0.5kg of lactic acid bacteria (dry weight), 6kg of milk powder and 79.5kg of water. As shown in FIG. 1, the lid 3 is opened, the fermentation material is placed in a fermentation tank 2 capable of containing 200L of water, stirred uniformly by a stirrer 4, kept at a constant temperature of 25 ℃ by a thermostat 1, and kept at the constant temperature for 10 hours to obtain 98.7kg of fermentation product, and then filtered and centrifuged to remove solid impurities, thereby obtaining 73.3kg of lactobacillus fermentation liquor.
Breaking the cell wall of the lactobacillus by adopting a spray drying method:
culturing lactic acid bacteria: 3kg of xylo-oligosaccharide, 5kg of pearl powder, 6kg of glucose, 0.5kg of lactic acid bacteria (dry weight), 6kg of milk powder and 79.5kg of water. As shown in fig. 1, the cover 3 is opened, the fermentation raw materials are put into a fermentation tank 2 which can be filled with 200 liters of water, the materials are uniformly stirred by a stirrer 4, the constant temperature of the constant temperature machine 1 is kept at 25 ℃, the temperature is kept for 10 hours, 98.7kg of the obtained fermented product is taken out, a protective agent is directly added into the lactobacillus fermented product, the material is added into a spray dryer, the lactobacillus cells are crushed by adjusting the operation parameters of the spray dryer, the intracellular lactase is released, and the lactase solid powder is obtained by spraying, wherein the protective agent is a mixture of dextrin, skimmed milk powder and dextrin, and the addition amount of the dextrin is as follows: 10-20 percent of the total weight of the mixture, g/100mL of the mixture, wherein the adding amount of the mixture of the skimmed milk powder and the dextrin is 15-20 percent, and g/100mL of the mixture. The air inlet temperature of the spray dryer is 125-135 ℃. Filtering, centrifuging to remove solid impurities to obtain 81.2kg lactobacillus fermentation liquid.
The lactobacillus fermentation lysate obtained in the embodiments 1 to 4 of the invention, the lactobacillus fermentation liquid obtained in the control group 1 and the control group 2 were subjected to the following efficacy test.
Tyrosinase inhibition assay: tyrosinase activity was measured by dopa-rate oxidation. Taking B16F10 melanoma cells in logarithmic growth phase, adding the prepared solution to be tested with mass fractions of 2.5%, 5% and 7.5%, setting negative control group and blank group, placing at 37 deg.C and 5% CO2The cells were removed after 24h incubation in the incubator. The resuspended cells were collected into 96-well plates, 90. mu.L of 1% Triton X-100 was added to each well, and the plates were placed in a-80 ℃ freezer and thawed at room temperature after 30 min. Preheating in 37 deg.C water bath to thoroughly lyse cells, adding 100 μ L of 10 g/L-DOPA solution into each well, and placing in 37 deg.C water bath for 4 hr. The microplate reader measures each well a490 and the tyrosinase inhibition was calculated according to the formula: tyrosinase inhibition rate ═ 1- (A experimental TYR/A control TYR)]X 100%, wherein: the TYR of the experiment group A is the tyrosinase activity of the culture medium added with the tested solution and the MRS broth of the negative control group, and the TYR of the control group A is the tyrosinase activity of the culture medium added only. The measurement results are shown in table 1.
Polypeptide content determination: the polypeptide content of the fermentation supernatant was measured by the OPA method (o-phthalaldehyde method), and the measurement results are shown in table 1.
Determination of the cleavage efficiency: the results of the biomicroscopy were shown in Table 1, in which the number of disrupted cells was counted per 100 cells after staining with 0.2% toluidine blue.
TABLE 1
As can be seen from the data in Table 1, the cracking rate, the inhibition rate and the polypeptide content of the lactobacillus fermentation lysate obtained by introducing carbon dioxide and applying pressure are all higher than those of the control group 1 and the control group 2. Meanwhile, the invention has differences in cracking effects obtained by applying different pressure differences: in example 4, when the applied pressure is 1MPa, the inhibition rate and the polypeptide content are significantly different from those in examples 1 to 3, so that the cracking effect is better when the pressure difference is 3 to 10 MPa. Although the control group 2 employed the spray high temperature expansion method to swell and demulsify the cells of the acid bacteria, a larger amount of the contents of the cells of the lactic acid bacteria could be obtained. However, high temperatures destroy temperature-sensitive active ingredients, and although this method adds a protective agent, the protective agent also inhibits the activity of the active.
The above disclosure is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the scope of the present invention, therefore, the present invention is not limited by the appended claims.
Claims (9)
1. A preparation method of lactobacillus fermentation lysate is characterized in that: putting the fermented lactobacillus culture solution into a high-pressure resistant container, introducing carbon dioxide and applying pressure to ensure that the carbon dioxide fully permeates into lactobacillus thallus, then conveying the lactobacillus thallus into a pressure reduction container to quickly reduce the pressure, quickly expanding the carbon dioxide in the thallus, and bursting the thallus, thereby releasing the content of the lactobacillus thallus.
2. The method for preparing a lactic acid bacteria fermentation lysate according to claim 1, characterized in that: the pressure applied by introducing carbon dioxide is 0.5-100 MPa.
3. The method for preparing a lactic acid bacteria fermentation lysate according to claim 2, characterized in that: the pressure applied by introducing carbon dioxide is 3-10 MPa.
4. The method for preparing a lactic acid bacteria fermentation lysate according to claim 1, characterized in that: the lactobacillus fermentation raw material contains pearl powder.
5. The method for preparing a lysate obtained by fermenting lactic acid bacteria according to claim 4, comprising: the lactic acid bacteria culture solution is prepared by fermenting xylo-oligosaccharide, pearl powder, glucose, lactic acid bacteria, milk powder and water.
6. The method for producing a lactic acid bacterium fermentation lysate according to claim 1 or 2, characterized in that: the temperature of the high-pressure resistant container is adjusted to 0-5 ℃, the high-pressure resistant container is sealed and then pressurized, carbon dioxide is introduced, the pressure is increased to 0.5-100 MPa, the pressure and the temperature are stably maintained for 5-10 hours, then the temperature is increased to 15-30 ℃, a discharge pipe at the bottom of the high-pressure resistant container is opened, and the fermented product in the high-pressure resistant container rapidly enters the pressure reduction container under pressure.
7. A cracking apparatus for preparing a lactic acid bacteria fermentation lysate according to any one of claims 1 to 6, characterized in that: the fermentation tank is internally provided with a stirrer, one end of the fermentation tank is connected with a constant temperature machine, the bottom of the fermentation tank is connected with the high-pressure resistant container through a discharge pipe, the high-pressure resistant container is provided with a temperature regulating device and a pressurizing device, and the top of the high-pressure resistant container is provided with a pipeline into which gas can be introduced; the bottom of the high pressure resistant container is connected with the pressure reducing container through a discharge pipe, a centrifugal separator is arranged in the pressure reducing container, and a discharge hole is formed in the bottom of the pressure reducing container.
8. The lysis device of claim 7, wherein: the high pressure resistant container is a high-pressure tank, and the pressure reducing container is a cracking tank.
9. The lysis device of claim 8, wherein: the top of the cracking tank is provided with an exhaust port, and the exhaust port is provided with a cyclone.
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| CN (1) | CN112795483A (en) |
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