CN113403247B - High-yield extracellular polysaccharide culture medium for bacillus polymyxa and application method thereof - Google Patents
High-yield extracellular polysaccharide culture medium for bacillus polymyxa and application method thereof Download PDFInfo
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Abstract
The invention relates to the technical field of microorganisms, and particularly discloses a bacillus polymyxa high-yield extracellular polysaccharide culture medium and a fermentation method. The culture medium comprises a seed culture medium and a fermentation culture medium, wherein the fermentation culture medium contains a fermentation regulator, the fermentation regulator is formed by mixing a component A and a component B, the component A is selected from one or a mixture of two of palmitoyl tripeptide-1 and palmitoyl tripeptide-7, the component B is selected from one or a mixture of two of asiaticoside and madecassoside, and the mass ratio of the component A to the component B is 4 to 1-1. The culture medium is applied to bacillus polymyxa fermentation exopolysaccharide, can effectively improve the exopolysaccharide yield, and has short exopolysaccharide production period and high production efficiency.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and particularly relates to a bacillus polymyxa high-yield extracellular polysaccharide culture medium and an application method thereof.
Background
The microbial extracellular polysaccharide is a carbohydrate compound secreted outside a cell wall by microorganisms in the growth and metabolism processes, and some of the carbohydrate compound adhere to the cell wall of the microorganisms to form capsules, which are called capsular polysaccharide; some enter the culture medium to form mucus, called mucopolysaccharide, which is the product of the microbial adaptation to the environment. The structure of the microbial extracellular polysaccharide is more complex than that of the plant polysaccharide, and the microbial extracellular polysaccharide has unique physicochemical property and biological activity, so the microbial extracellular polysaccharide is widely applied to various industries. However, with the social development, the demand for extracellular polysaccharide of microorganism is continuously increased, and the current technology and yield for producing extracellular polysaccharide cannot meet the demand of people for extracellular polysaccharide of microorganism, so that the improvement of the yield of extracellular polysaccharide produced by microbial fermentation has important significance.
Disclosure of Invention
The invention aims to provide a bacillus polymyxa high-yield extracellular polysaccharide culture medium and an application method thereof.
In order to achieve the purpose, the invention provides a culture medium for high yield of exopolysaccharides by bacillus polymyxa, which comprises a seed culture medium and a fermentation culture medium, wherein the seed culture medium comprises the following components in parts by mass: 20-40 g/L of yeast powder, 25-40 g/L of glucose, 10-15 g/L of peptone, 0.1-8 g/L of monopotassium phosphate, 0.2-2 g/L of ammonium nitrate, 0.05-0.1 g/L of sodium chloride, 5-15 g/L of magnesium sulfate heptahydrate and 0.05-0.1 g/L of ferric chloride;
the fermentation medium comprises the following components in percentage by mass: 50-80 g/L of sucrose, 0.5-0.6 g/L of fermentation regulator, 20-45 g/L of peptone, 35-45 g/L of yeast powder, 0.5-10 g/L of monopotassium phosphate, 5-15 g/L of magnesium sulfate heptahydrate, 0.2-2 g/L of ammonium nitrate, 0.05-0.1 g/L of sodium chloride, 0.2-2 g/L of ammonium nitrate and 0.05-0.1 g/L of ferric chloride;
the fermentation regulator is formed by mixing a component A and a component B, wherein the component A is selected from one or a mixture of two of palmitoyl tripeptide-1 and palmitoyl tripeptide-7, the component B is selected from one or a mixture of two of asiaticoside and madecassoside, and the mass ratio of the component A to the component B is 4.
Further, in the above technical scheme, the culture medium includes a seed culture medium and a fermentation culture medium, and the seed culture medium includes, by mass concentration: 30g/L of yeast powder, 25g/L of glucose, 13g/L of peptone, 4g/L of monopotassium phosphate, 1g/L of ammonium nitrate, 0.08g/L of sodium chloride, 15g/L of magnesium sulfate heptahydrate and 0.07g/L of ferric chloride;
the fermentation medium comprises the following components in percentage by mass: 60g/L of sucrose, 0.55g/L of fermentation regulator, 35g/L of peptone, 40g/L of yeast powder, 4g/L of monopotassium phosphate, 10g/L of magnesium sulfate heptahydrate, 1g/L of ammonium nitrate, 0.08g/L of sodium chloride, 1g/L of ammonium nitrate and 0.07g/L of ferric chloride;
the fermentation regulator is formed by mixing a component A and a component B, wherein the component A is palmitoyl tripeptide-1, the component B is asiaticoside, and the mass ratio of the component A to the component B is 3.
The method for preparing extracellular polysaccharide by fermenting the bacillus polymyxa comprises the following steps:
(1) Weighing yeast powder, glucose, peptone, potassium dihydrogen phosphate, ammonium nitrate, sodium chloride, magnesium sulfate heptahydrate and ferric chloride according to the formula proportion of a seed culture medium, adding water for dissolving, sterilizing to obtain the seed culture medium, and inoculating bacillus polymyxa into the seed culture medium for culture to obtain a seed solution;
the seed culture medium comprises the following components in percentage by mass: 20-40 g/L of yeast powder, 25-40 g/L of glucose, 10-15 g/L of peptone, 0.1-8 g/L of monopotassium phosphate, 0.2-2 g/L of ammonium nitrate, 0.05-0.1 g/L of sodium chloride, 5-15 g/L of magnesium sulfate heptahydrate, 0.05-0.1 g/L of ferric chloride and the balance of water;
(2) Weighing sucrose, fermentation regulator, peptone, yeast powder, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, ammonium nitrate, sodium chloride, ammonium nitrate and ferric chloride according to the formula proportion of the fermentation medium, adding water for dissolving, sterilizing to obtain the fermentation medium, inoculating the seed solution into a fermentation tank filled with the fermentation medium for fermentation culture, and obtaining extracellular polysaccharide fermentation liquid after the fermentation is finished;
the fermentation medium comprises the following components in percentage by mass: 50-80 g/L of sucrose, 0.50-0.60 g/L of fermentation regulator, 20-45 g/L of peptone, 35-45 g/L of yeast powder, 0.5-10 g/L of monopotassium phosphate, 5-15 g/L of magnesium sulfate heptahydrate, 0.2-2 g/L of ammonium nitrate, 0.05-0.1 g/L of sodium chloride, 0.2-2 g/L of ammonium nitrate, 0.05-0.1 g/L of ferric chloride and the balance of water;
the fermentation regulator is formed by mixing a component A and a component B, wherein the component A is selected from one or a mixture of two of palmitoyl tripeptide-1 and palmitoyl tripeptide-7, the component B is selected from one or a mixture of two of asiaticoside and madecassoside, and the mass ratio of the component A to the component B is (4).
Further, in the above technical scheme, the sterilization temperature in the step (1) is 120 ℃ to 130 ℃.
Further, in the above technical scheme, the sterilization time in the step (1) is 15-30 min.
Further, in the technical scheme, the inoculation amount of the bacillus polymyxa inoculated into the seed culture medium in the step (1) is 6-12%.
Further, in the technical scheme, the bacillus polymyxa in the step (1) is inoculated into a seed culture medium and cultured for 36-38 h at the temperature of 30-37 ℃ and the rotating speed of 200-240 r/min.
Further, in the above technical scheme, the sterilization temperature in the step (2) is 120-130 ℃ and the time is 15-30 min.
Further, in the above technical scheme, the seed liquid inoculation amount in the step (2) is 6-12%.
Further, in the above technical scheme, the fermentation culture conditions in step (2) include: the charging coefficient is 0.6, the temperature is 30-37 ℃, the pressure is 0.04-0.07 MPa, the ventilation volume is 0.8-1.0 v/v.min, and the fermentation time is 46-48 h.
Compared with the prior art, the invention has the following beneficial effects:
1. the invention provides a bacillus polymyxa high-yield extracellular polysaccharide culture medium which is rich in nutrition, wherein magnesium sulfate heptahydrate and ferric chloride added can maintain the normal metabolic demand of microorganisms, a fermentation regulator is added, the regulator consists of one or two of palmitoyl tripeptide-1 and palmitoyl tripeptide-7 and one or two of asiaticoside and madecassoside, and the propagation of bacillus polymyxa and the secretion of extracellular polysaccharide are promoted by regulating the processes of physiological reaction and the like of bacillus polymyxa, so that the yield of extracellular polysaccharide prepared by fermenting bacillus polymyxa is effectively improved, and the extracellular polysaccharide yield improvement effect of the bacillus polymyxa high-yield extracellular polysaccharide culture medium provided by the invention is obvious.
2. The culture medium of the invention promotes the physiological reaction rate of the bacillus polymyxa, shortens the fermentation period, is beneficial to improving the extracellular polysaccharide production efficiency of the bacillus polymyxa through fermentation, and saves time; secondly, the method for producing the exopolysaccharide by fermenting the bacillus polymyxa is simple and easy to operate, the preparation cost of the culture medium is low, and the fermentation adaptive condition range is wide.
Detailed Description
The following detailed description of specific embodiments of the invention is provided, but it should be understood that the scope of the invention is not limited to the specific embodiments.
Example 1
Medium sample 1: comprises a seed culture medium and a fermentation culture medium;
seed medium (by mass concentration): 30g/L of yeast powder, 25g/L of glucose, 13g/L of peptone, 4g/L of potassium dihydrogen phosphate, 1g/L of ammonium nitrate, 0.08g/L of sodium chloride, 15g/L of magnesium sulfate heptahydrate, 0.07g/L of ferric chloride and the balance of water;
fermentation medium (by mass concentration): 60g/L of sucrose, 0.55g/L of fermentation regulator, 35g/L of peptone, 40g/L of yeast powder, 4g/L of monopotassium phosphate, 15g/L of magnesium sulfate heptahydrate, 1g/L of ammonium nitrate, 0.08g/L of sodium chloride, 1g/L of ammonium nitrate, 0.07g/L of ferric chloride and the balance of water;
the fermentation regulator is formed by mixing a component A and a component B, wherein the component A is palmitoyl tripeptide-1, the component B is asiaticoside, and the mass ratio of the component A to the component B is 3.
Comparative medium 1-1: unlike medium sample 1, which has no fermentation regulator, the other samples are substantially the same as medium sample 1.
Comparative medium 1-2: the same amount of component A as the fermentation regulator was used in the difference from medium sample 1, and the other was substantially the same as medium sample 1.
Comparative media 1-3: the same amount of component B as a fermentation regulator was used as a difference from medium sample 1, and the other was substantially the same as medium sample 1.
Preparing exopolysaccharide by fermentation:
(1) Weighing yeast powder, glucose, peptone, potassium dihydrogen phosphate, ammonium nitrate, sodium chloride, magnesium sulfate heptahydrate and ferric chloride according to the formula proportion of a seed culture medium, adding water for dissolving, sterilizing to obtain the seed culture medium, inoculating bacillus polymyxa into the seed culture medium (the inoculation amount of the bacillus polymyxa into the seed culture medium is 6%) for culture at the sterilization temperature of 120 ℃, and inoculating the bacillus polymyxa into the seed culture medium at the rotation speed of 200r/min for 36h to obtain seed liquid;
(2) Weighing sucrose, fermentation regulators (the fermentation medium in a contrast medium 1-1 does not contain the fermentation regulators), peptone, yeast powder, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, ammonium nitrate, sodium chloride, ammonium nitrate and ferric chloride according to the formula proportion of the fermentation medium, adding water to dissolve, sterilizing (the sterilization temperature is 120 ℃ and the time is 15 min) to obtain the fermentation medium, inoculating the seed solution into a fermentation tank filled with the fermentation medium (the seed solution inoculation amount is 6%) to perform fermentation culture, wherein the charging coefficient is 0.6, the fermentation temperature is 30 ℃, the pressure is 0.05MPa, the ventilation volume is 0.9v/v.min, and fermenting for 46h to obtain the extracellular polysaccharide fermentation liquid.
Yield of extracellular polysaccharide through fermentation culture of different culture media
The yield of extracellular polysaccharide obtained by fermentation culture of different culture media is shown in table 1, wherein the yield of extracellular polysaccharide obtained by adopting culture medium sample 1 is 80.61g/L, the lowest yield of contrast culture medium 1-1 is 13.62g/L, and the yields of contrast culture medium 1-2 and contrast culture medium 1-3 are 41.15g/L and 23.92g/L respectively, which shows that the yield of extracellular polysaccharide obtained by fermentation culture of culture medium sample 1 relative to contrast culture medium is obviously improved obviously.
TABLE 1 fermentation culture of extracellular polysaccharide yields in different media
| Culture medium | Extracellular polysaccharide yield (g/L) |
| Medium sample 1 | 80.61 |
| Comparative Medium 1-1 | 13.62 |
| Comparative Medium 1-2 | 41.15 |
| Comparative Medium 1-3 | 23.92 |
Example 2
Medium sample 2: comprises a seed culture medium and a fermentation culture medium;
seed medium (by mass concentration): 20g/L of yeast powder, 30g/L of glucose, 10g/L of peptone, 0.1g/L of monopotassium phosphate, 0.2g/L of ammonium nitrate, 0.05g/L of sodium chloride, 5g/L of magnesium sulfate heptahydrate, 0.05g/L of ferric chloride and the balance of water;
fermentation medium (by mass concentration): 50g/L of sucrose, 0.5g/L of fermentation regulator, 20g/L of peptone, 35g/L of yeast powder, 0.5g/L of monopotassium phosphate, 5g/L of magnesium sulfate heptahydrate, 0.2g/L of ammonium nitrate, 0.05g/L of sodium chloride, 0.2g/L of ammonium nitrate, 0.05g/L of ferric chloride and the balance of water;
the fermentation regulator is formed by mixing a component A and a component B, wherein the component A is palmitoyl tripeptide-7, the component B is selected from asiaticoside, and the mass ratio of the component A to the component B is 4.
Comparative medium 2-1: unlike medium sample 2, which is essentially the same as medium sample 2, there is no fermentation regulator.
Comparative medium 2-2: the same amount of component A as the fermentation regulator was used as a difference from medium sample 2, and the other was substantially the same as medium sample 2.
Comparative medium 2-3: the same amount of component B as the fermentation regulator was used in contrast to medium sample 2, which was otherwise substantially the same as medium sample 2.
Preparing exopolysaccharide by fermentation:
(1) Weighing yeast powder, glucose, peptone, potassium dihydrogen phosphate, ammonium nitrate, sodium chloride, magnesium sulfate heptahydrate and ferric chloride according to the formula proportion of a seed culture medium, adding water for dissolving, sterilizing to obtain the seed culture medium, inoculating bacillus polymyxa into the seed culture medium (the inoculation amount of the bacillus polymyxa into the seed culture medium is 10%) for culture, and inoculating the bacillus polymyxa into the seed culture medium at the temperature of 35 ℃ and the rotating speed of 220r/min for culture for 36h to obtain a seed solution;
(2) Weighing sucrose according to the formula proportion of a fermentation medium, adding fermentation regulators (the fermentation medium does not contain the fermentation regulators in a comparison medium 2-1), peptone, yeast powder, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, ammonium nitrate, sodium chloride, ammonium nitrate and ferric chloride, dissolving in water, sterilizing (the sterilization temperature is 125 ℃, and the time is 20 min) to obtain the fermentation medium, inoculating a seed solution into a fermentation tank filled with the fermentation medium (the inoculation amount of the seed solution is 10%) for fermentation culture, wherein the charging coefficient is 0.6, the fermentation temperature is 35 ℃, the pressure is 0.05MPa, and the ventilation amount is 0.8v/v.min, and fermenting for 48h to obtain an extracellular polysaccharide fermentation liquid;
yield of extracellular polysaccharide through fermentation culture of different culture media
The yield of extracellular polysaccharide obtained by fermentation culture of different culture media is shown in table 2, wherein the yield of extracellular polysaccharide obtained by adopting culture medium sample 2 is 76.45g/L, the lowest yield of contrast culture medium 2-1 is 13.43g/L, and the yields of contrast culture medium 2-2 and contrast culture medium 2-3 are 33.83g/L and 23.71g/L respectively, which shows that the yield of extracellular polysaccharide obtained by fermentation culture of culture medium sample 2 relative to contrast culture medium is obviously improved obviously.
TABLE 2 fermentation culture of extracellular polysaccharide yields in different media
Example 3
Medium sample 3: comprises a seed culture medium and a fermentation culture medium;
seed medium (by mass concentration): 40g/L of yeast powder, 40g/L of glucose, 15g/L of peptone, 8g/L of potassium dihydrogen phosphate, 2g/L of ammonium nitrate, 0.1g/L of sodium chloride, 10g/L of magnesium sulfate heptahydrate, 0.1g/L of ferric chloride and the balance of water;
fermentation medium (by mass concentration): 80g/L of sucrose, 0.6g/L of fermentation regulator, 45g/L of peptone, 45g/L of yeast powder, 10g/L of monopotassium phosphate, 10g/L of magnesium sulfate heptahydrate, 2g/L of ammonium nitrate, 0.1g/L of sodium chloride, 2g/L of ammonium nitrate, 0.1g/L of ferric chloride and the balance of water;
the fermentation regulator is formed by mixing a component A and a component B, wherein the component A is a mixture of palmitoyl tripeptide-1 and palmitoyl tripeptide-7, the mass ratio of the palmitoyl tripeptide-1 to the palmitoyl tripeptide-7 is 1.
Comparative medium 3-1: unlike medium sample 3, which did not have a fermentation regulator, it was essentially the same as medium sample 3.
Comparative medium 3-2: the same amount of component A as the fermentation regulator was used as a difference from medium sample 3, and the other was substantially the same as medium sample 3.
Comparative medium 3-3: the same amount of component B as a fermentation regulator was used as a difference from medium sample 3, and the other was substantially the same as medium sample 3.
Preparing exopolysaccharide by fermentation:
(1) Weighing yeast powder, glucose, peptone, potassium dihydrogen phosphate, ammonium nitrate, sodium chloride, magnesium sulfate heptahydrate and ferric chloride according to the formula proportion of a seed culture medium, adding water for dissolving, sterilizing to obtain the seed culture medium, inoculating bacillus polymyxa into the seed culture medium (the inoculation amount of the bacillus polymyxa into the seed culture medium is 12%) for culture, inoculating the bacillus polymyxa into the seed culture medium at 37 ℃, rotating speed 240r/min, and culturing for 38h to obtain a seed solution;
(2) Weighing sucrose, fermentation regulators (the fermentation medium in a contrast medium 3-1 does not contain the fermentation regulators), peptone, yeast powder, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, ammonium nitrate, sodium chloride, ammonium nitrate and ferric chloride according to the formula proportion of the fermentation medium, adding water to dissolve the fermentation regulators, sterilizing (the sterilization temperature is 130 ℃ and the time is 30 min) to obtain the fermentation medium, inoculating the seed solution into a fermentation tank filled with the fermentation medium (the seed solution inoculation amount is 12%) for fermentation culture, wherein the charging coefficient is 0.6, the fermentation temperature is 37 ℃, the pressure is 0.07MPa, the ventilation volume is 1.0v/v.min, and fermenting for 48h to obtain extracellular polysaccharide fermentation liquid;
yield of extracellular polysaccharide by fermentation culture of different culture media
The yield of extracellular polysaccharide cultured by different culture medium fermentation is shown in table 3, wherein the extracellular polysaccharide yield of the culture medium sample 3 is 77.19g/L, the lowest yield of the contrast culture medium 3-1 is 13.70g/L, the yields of the contrast culture medium 3-2 and the contrast culture medium 3-3 are 36.92g/L and 23.97g/L respectively, and obviously, the extracellular polysaccharide yield is obviously improved when the culture medium sample 3 is used for producing extracellular polysaccharide by fermentation of bacillus polymyxa compared with the contrast culture medium.
TABLE 3 fermentation culture of extracellular polysaccharide yields in different media
| Culture medium | Extracellular polysaccharide yield (g/L) |
| Medium sample 3 | 77.19 |
| Comparative Medium 3-1 | 13.70 |
| Comparative Medium 3-2 | 36.92 |
| Comparative Medium 3-3 | 23.97 |
Example 4
Medium sample 4: comprises a seed culture medium and a fermentation culture medium;
seed medium (by mass concentration): 35g/L of yeast powder, 30g/L of glucose, 14g/L of peptone, 7g/L of monopotassium phosphate, 1.5g/L of ammonium nitrate, 0.09g/L of sodium chloride, 13g/L of magnesium sulfate heptahydrate, 0.08g/L of ferric chloride and the balance of water;
fermentation medium (by mass concentration): 70g/L of sucrose, 0.58g/L of fermentation regulator, 40g/L of peptone, 40g/L of yeast powder, 8g/L of monopotassium phosphate, 13g/L of magnesium sulfate heptahydrate, 1.5g/L of ammonium nitrate, 0.09g/L of sodium chloride, 1.5g/L of ammonium nitrate, 0.08g/L of ferric chloride and the balance of water;
the fermentation regulator is formed by mixing a component A and a component B, wherein the component A is palmitoyl tripeptide-1, the component B is madecassoside, and the mass ratio of the component A to the component B is 3.
Comparative medium 4-1: unlike medium sample 4, which is essentially the same as medium sample 4, there is no fermentation regulator.
Comparative medium 4-2: the same amount of component A as the fermentation regulator was used as a difference from medium sample 4, and the other was substantially the same as medium sample 4.
Comparative medium 4-3: the same amount of component B as the fermentation regulator was used as a difference from medium sample 4, and the other was substantially the same as medium sample 4.
Preparing extracellular polysaccharide through fermentation:
(1) Weighing yeast powder, glucose, peptone, potassium dihydrogen phosphate, ammonium nitrate, sodium chloride, magnesium sulfate heptahydrate and ferric chloride according to the formula proportion of a seed culture medium, adding water for dissolving, sterilizing to obtain the seed culture medium, inoculating bacillus polymyxa into the seed culture medium (the inoculation amount of the bacillus polymyxa into the seed culture medium is 8%) for culture, inoculating the bacillus polymyxa into the seed culture medium, and culturing at 35 ℃, 220r/min for 36h to obtain a seed solution;
(2) Weighing sucrose, fermentation regulators (the fermentation medium does not contain the fermentation regulators in a contrast medium 4-1), peptone, yeast powder, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, ammonium nitrate, sodium chloride, ammonium nitrate and ferric chloride, adding water for dissolving, sterilizing (the sterilization temperature is 125 ℃, the time is 20 min) to obtain a fermentation medium, inoculating a seed solution into a fermentation tank filled with the fermentation medium (the seed solution inoculation amount is 8%) for fermentation culture, wherein the charging coefficient is 0.6, the fermentation temperature is 37 ℃, the pressure is 0.05MPa, the ventilation volume is 0.9v/v.min, and fermenting for 48h to obtain an extracellular polysaccharide fermentation liquid;
yield of extracellular polysaccharide through fermentation culture of different culture media
The yield of extracellular polysaccharide obtained by fermentation culture of different culture media is shown in table 4, wherein the yield of extracellular polysaccharide of a culture medium sample 4 is 69.28g/L, the lowest yield of a contrast culture medium 4-1 is 13.54g/L, the yields of the contrast culture medium 4-2 and the contrast culture medium 4-3 are 43.82g/L and 20.76g/L respectively, and obviously, the yield of the culture medium sample 4 is obviously improved obviously compared with that of the contrast culture medium for fermentation production of extracellular polysaccharide of bacillus polymyxa.
TABLE 4 fermentation culture of extracellular polysaccharide yields in different media
| Culture medium | Extracellular polysaccharide yield (g/L) |
| Medium sample 4 | 69.28 |
| Comparative Medium 4-1 | 13.54 |
| Comparative Medium 4-2 | 43.82 |
| Comparative Medium 4-3 | 20.76 |
Example 5
Medium sample 5: comprises a seed culture medium and a fermentation culture medium;
seed medium (by mass concentration): 25g/L of yeast powder, 35g/L of glucose, 11g/L of peptone, 3g/L of monopotassium phosphate, 1g/L of ammonium nitrate, 0.07g/L of sodium chloride, 8g/L of magnesium sulfate heptahydrate, 0.08g/L of ferric chloride and the balance of water;
fermentation medium (by mass concentration): 60g/L of sucrose, 0.52g/L of fermentation regulator, 30g/L of peptone, 35g/L of yeast powder, 4g/L of monopotassium phosphate, 6g/L of magnesium sulfate heptahydrate, 1.5g/L of ammonium nitrate, 0.06g/L of sodium chloride, 0.8g/L of ammonium nitrate, 0.07g/L of ferric chloride and the balance of water;
the fermentation regulator is formed by mixing a component A and a component B, wherein the component A is palmitoyl tripeptide-1, the component B is a mixture of asiaticoside and madecassoside, the mass ratio of the asiaticoside to the madecassoside is 1.
Comparative medium 5-1: unlike medium sample 5, which is essentially the same as medium sample 5, there is no fermentation regulator.
Comparative medium 5-2: the same amount of component A as the fermentation regulator was used as it was in the case of medium sample 5, and the other was substantially the same as in medium sample 5.
Comparative medium 5-3: the same amount of component B as the fermentation regulator was used in the difference from medium sample 5, and the other was substantially the same as medium sample 5.
Preparing exopolysaccharide by fermentation:
(1) Weighing yeast powder, glucose, peptone, potassium dihydrogen phosphate, ammonium nitrate, sodium chloride, magnesium sulfate heptahydrate and ferric chloride according to the formula proportion of a seed culture medium, adding water for dissolving, sterilizing to obtain the seed culture medium, inoculating bacillus polymyxa into the seed culture medium (the inoculation amount of the bacillus polymyxa into the seed culture medium is 10%) for culture, and inoculating the bacillus polymyxa into the seed culture medium at 37 ℃ and at the rotating speed of 200r/min for culture for 36h to obtain a seed solution;
(2) Weighing sucrose, fermentation regulators (the fermentation medium in a contrast medium 5-1 does not contain the fermentation regulators), peptone, yeast powder, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, ammonium nitrate, sodium chloride, ammonium nitrate and ferric chloride according to a formula proportion of the fermentation medium, adding water for dissolving, sterilizing (sterilization temperature is 130 ℃ and time is 15 min) to obtain the fermentation medium, inoculating a seed solution into a fermentation tank filled with the fermentation medium (seed solution inoculation amount is 10%) for fermentation culture, wherein the charging coefficient is 0.6, the fermentation temperature is 37 ℃, the pressure is 0.07MPa, the ventilation volume is 0.8v/v.min, and fermenting for 48h to obtain an extracellular polysaccharide fermentation liquid;
yield of extracellular polysaccharide through fermentation culture of different culture media
The yield of exopolysaccharide produced by fermentation of different culture media is shown in table 5, wherein the yield of exopolysaccharide produced by adopting the culture medium sample 3 is 72.37g/L, the lowest yield of adopting the contrast culture medium 5-1 is 13.42g/L, and the yields of adopting the contrast culture medium 5-2 and the contrast culture medium 5-3 are 43.95g/L and 21.84g/L respectively, which shows that the yield of the culture medium sample 5 is obviously improved when being used for producing exopolysaccharide by fermentation of bacillus polymyxa compared with the contrast culture medium.
TABLE 5 fermentation culture of extracellular polysaccharide yields in different media
| Culture medium | Extracellular polysaccharide yield (g/L) |
| Medium sample 5 | 72.37 |
| Comparative Medium 5-1 | 13.42 |
| Comparative Medium 5-2 | 43.95 |
| Comparative Medium 5-3 | 21.84 |
In conclusion, the culture medium provided by the invention is rich in nutrition, good in effect of culturing the bacillus polymyxa, and high in extracellular polysaccharide yield when applied to fermentation preparation of the bacillus polymyxa, and particularly, the mass ratio of palmitoyl tripeptide-1 to asiaticoside is 3. Meanwhile, the bacillus polymyxa fermentation production extracellular polysaccharide has the advantages of short production period, high production efficiency, wide range of fermentation application conditions and stable production.
The foregoing descriptions of specific exemplary embodiments of the present invention have been presented for purposes of illustration and description. It is not intended to limit the invention to the precise form disclosed, and obviously many modifications and variations are possible in light of the above teaching. The exemplary embodiments were chosen and described in order to explain certain principles of the invention and its practical application to enable one skilled in the art to make and use various exemplary embodiments of the invention and various alternatives and modifications as are suited to the particular use contemplated. It is intended that the scope of the invention be defined by the claims and their equivalents.
Claims (10)
1. The culture medium for high yield of exopolysaccharides of bacillus polymyxa is characterized by comprising a seed culture medium and a fermentation culture medium, wherein the seed culture medium comprises the following components in percentage by mass: 20-40g/L yeast powder, 25-40g/L glucose, 10-15g/L peptone, 0.1-8g/L monopotassium phosphate, 0.2-2g/L ammonium nitrate, 0.05-0.1g/L sodium chloride, 5-15g/L magnesium sulfate heptahydrate and 0.05-0.1g/L iron chloride;
the fermentation medium comprises the following components in percentage by mass: 50-80g/L of sucrose, 0.5-0.6 g/L of fermentation regulator, 20-45g/L of peptone, 35-45g/L of yeast powder, 0.5-10g/L of monopotassium phosphate, 0.2-20g/L of magnesium sulfate heptahydrate, 0.2-212g/L of ammonium nitrate, 0.05-0.1g/L of sodium chloride and 0.05-0.1g/L of ferric chloride;
the fermentation regulator is formed by mixing a component A and a component B, wherein the component A is palmitoyl tripeptide-1, the component B is one or a mixture of two of asiaticoside and madecassoside, and the mass ratio of the component A to the component B is 4 to 3.
2. The culture medium for high exopolysaccharide yield of bacillus polymyxa according to the claim 1, wherein the culture medium comprises a seed culture medium and a fermentation culture medium, and the seed culture medium comprises the following components in concentration by mass: 30g/L of yeast powder, 25g/L of glucose, 13g/L of peptone, 4g/L of potassium dihydrogen phosphate, 1g/L of ammonium nitrate, 0.08g/L of sodium chloride, 15g/L of magnesium sulfate heptahydrate and 0.07g/L of ferric chloride;
the fermentation medium comprises the following components in percentage by mass: 60g/L of sucrose, 0.55g/L of fermentation regulator, 35g/L of peptone, 40g/L of yeast powder, 4g/L of monopotassium phosphate, 10g/L of magnesium sulfate heptahydrate, 1g/L of ammonium nitrate, 0.08g/L of sodium chloride and 0.07g/L of ferric chloride;
the fermentation regulator is formed by mixing a component A and a component B, wherein the component A is palmitoyl tripeptide-1, the component B is asiaticoside, and the mass ratio of the component A to the component B is 3.
3. The application of the culture medium for high yield of exopolysaccharides of bacillus polymyxa as claimed in any one of claims 1-2 in preparation of exopolysaccharides through fermentation of bacillus polymyxa.
4. The application of the culture medium for the high-yield exopolysaccharides of bacillus polymyxa in fermentation of bacillus polymyxa for preparing exopolysaccharides is characterized by comprising the following steps of:
(1) Weighing yeast powder, glucose, peptone, potassium dihydrogen phosphate, ammonium nitrate, sodium chloride, magnesium sulfate heptahydrate and ferric chloride according to the formula proportion of a seed culture medium, adding water for dissolving, sterilizing to obtain a seed culture medium, and inoculating bacillus polymyxa into the seed culture medium for culture to obtain a seed solution;
(2) Weighing sucrose, fermentation regulator, peptone, yeast powder, potassium dihydrogen phosphate, magnesium sulfate heptahydrate, ammonium nitrate, sodium chloride and ferric chloride according to the formula proportion of the fermentation medium, adding water for dissolving, sterilizing to obtain the fermentation medium, inoculating the seed liquid into a fermentation tank filled with the fermentation medium for fermentation culture, and obtaining the extracellular polysaccharide fermentation liquid after the fermentation is finished.
5. The application of the culture medium for the high yield of exopolysaccharides of bacillus polymyxa in the fermentation preparation of exopolysaccharides of bacillus polymyxa according to claim 4 is characterized in that the sterilization temperature in the step (1) is 120-130 ℃ and the sterilization time is 15-30min.
6. The application of the culture medium for the high yield of exopolysaccharides of bacillus polymyxa in the fermentation preparation of exopolysaccharides of bacillus polymyxa according to claim 4 is characterized in that the inoculation amount of bacillus polymyxa inoculated into the seed culture medium in the step (1) is 6-12%.
7. The application of the culture medium for the high yield of exopolysaccharides from bacillus polymyxa in the fermentation of bacillus polymyxa for preparing exopolysaccharides is characterized in that in the step (1), bacillus polymyxa is inoculated into a seed culture medium and cultured for 36h to 38h at the temperature of 30 to 37 ℃ and the rotation speed of 200 to 240r/min.
8. The application of the culture medium for the high yield of exopolysaccharides from bacillus polymyxa in fermentation of bacillus polymyxa for preparation of exopolysaccharides is characterized in that the sterilization temperature in the step (2) is 120-130 ℃, and the sterilization time is 15-30min.
9. The application of the culture medium for the high yield of exopolysaccharides in bacillus polymyxa in fermentation of the bacillus polymyxa for preparing exopolysaccharides according to claim 4 is characterized in that the seed liquid inoculation amount in the step (2) is 6-12%.
10. The use of exopolysaccharide highly productive culture medium of bacillus polymyxa in the fermentation preparation of exopolysaccharide by bacillus polymyxa according to claim 4, wherein the fermentation culture conditions in step (2) include: the charging coefficient is 0.6, the temperature is 30 to 37 ℃, the pressure is 0.04 to 0.07MPa, the ventilation volume is 0.8 to 1.0v/v.min, and the fermentation time is 46 to 48h.
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Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5420036A (en) * | 1989-10-04 | 1995-05-30 | Haitai Confectionery Co., Ltd. | Bacillus polymyxa 1 capable of producing polysaccharides and polysaccharides produced by this strain |
| US5547863A (en) * | 1989-08-14 | 1996-08-20 | The United States Of America As Represented By The Secretary Of The Agriculture | Production of fructan (levan) polyfructose polymers using bacillus polymyxa |
| WO2001018226A1 (en) * | 1999-09-07 | 2001-03-15 | Rhodia Chimie | Method for producing exopolysaccharides |
| WO2014147255A1 (en) * | 2013-03-22 | 2014-09-25 | Lipotec S.A. | Exopolysaccharide for the treatment and/or care of the skin, mucous membranes and/or nails |
| EP2868308A1 (en) * | 2013-10-30 | 2015-05-06 | Lipotec, S.A. | Expolysaccharide for the treatment and/or care of skin, culture media and compositions thereof |
| WO2016067218A1 (en) * | 2014-10-28 | 2016-05-06 | Lubrizol Advanced Materials, Inc. | Cosmetic composition containing halomonas ferment extract, and use thereof |
| CN107438428A (en) * | 2015-02-19 | 2017-12-05 | Elc 管理有限责任公司 | Novel skin remodeling strategy |
| WO2020078467A1 (en) * | 2018-10-18 | 2020-04-23 | 浙江大学 | Paenibacillus elgii strain for producing polysaccharide with effect of after-sun repair and application thereof |
| CN111575201A (en) * | 2020-05-09 | 2020-08-25 | 大连理工大学 | High-density mixed fermentation preparation method of composite bacillus agent |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| ES2390033B1 (en) * | 2010-11-30 | 2013-10-31 | Lipotec S.A. | EXOPOLISACÁRIDO FOR THE TREATMENT AND / OR CARE OF SKIN, MUCOSAS, HAIR AND / OR NAILS. |
| KR102448943B1 (en) * | 2014-10-31 | 2022-09-28 | 리포텍 에스.에이. | Cosmetic and/or pharmaceutical composition containing a bacterial extracellular product from pseudoalteromonas antarctica, and use thereof |
| MX2017005597A (en) * | 2014-10-31 | 2017-06-23 | Lubrizol Advanced Mat Inc | Thermoplastic polyurethane film for delivery of active agents to skin surfaces. |
| US20220142910A1 (en) * | 2020-11-06 | 2022-05-12 | Dianne Martin Bastian | Composition including all natural botanical peptide ingredients for topical skin applications |
-
2021
- 2021-05-14 CN CN202110527044.XA patent/CN113403247B/en active Active
Patent Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5547863A (en) * | 1989-08-14 | 1996-08-20 | The United States Of America As Represented By The Secretary Of The Agriculture | Production of fructan (levan) polyfructose polymers using bacillus polymyxa |
| US5420036A (en) * | 1989-10-04 | 1995-05-30 | Haitai Confectionery Co., Ltd. | Bacillus polymyxa 1 capable of producing polysaccharides and polysaccharides produced by this strain |
| WO2001018226A1 (en) * | 1999-09-07 | 2001-03-15 | Rhodia Chimie | Method for producing exopolysaccharides |
| WO2014147255A1 (en) * | 2013-03-22 | 2014-09-25 | Lipotec S.A. | Exopolysaccharide for the treatment and/or care of the skin, mucous membranes and/or nails |
| EP2868308A1 (en) * | 2013-10-30 | 2015-05-06 | Lipotec, S.A. | Expolysaccharide for the treatment and/or care of skin, culture media and compositions thereof |
| WO2016067218A1 (en) * | 2014-10-28 | 2016-05-06 | Lubrizol Advanced Materials, Inc. | Cosmetic composition containing halomonas ferment extract, and use thereof |
| CN107438428A (en) * | 2015-02-19 | 2017-12-05 | Elc 管理有限责任公司 | Novel skin remodeling strategy |
| WO2020078467A1 (en) * | 2018-10-18 | 2020-04-23 | 浙江大学 | Paenibacillus elgii strain for producing polysaccharide with effect of after-sun repair and application thereof |
| CN111575201A (en) * | 2020-05-09 | 2020-08-25 | 大连理工大学 | High-density mixed fermentation preparation method of composite bacillus agent |
Non-Patent Citations (2)
| Title |
|---|
| 一株产胞外多糖植物内生菌EJS-3菌株的分离和鉴定;孙力军等;《食品科学》;20060715;第27卷(第07期);65-68 * |
| 多粘芽孢杆菌发酵条件初步优化;陆旋等;《浙江化工》;20131115;第44卷(第11期);25-28 * |
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