CN113201563B - Nutritive salt for increasing yield of sclerotium rolfsii polysaccharide and application thereof - Google Patents
Nutritive salt for increasing yield of sclerotium rolfsii polysaccharide and application thereof Download PDFInfo
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Abstract
The invention provides a nutritive salt for improving the yield of sclerotium rolfsii polysaccharide and application thereof, belonging to the field of microbial fermentation. The nutrient salt for improving the yield of sclerotium rolfsii polysaccharide comprises the following components: disodium 6-phosphofructose, pyruvic acid, acetic acid, acetaldehyde and glutamic acid. The invention also provides application of the nutrient salt in producing sclerotium rolfsii polysaccharide. The nutritive salt is added at different time periods of fermentation and the pH values at different time periods are adjusted and controlled in a matching way, so that the synthesis of the sclerotium rolfsii polysaccharide is facilitated, and the aims of improving the yield of the sclerotium rolfsii polysaccharide, improving the utilization rate of a substrate and reducing the environmental pollution are fulfilled. The invention opens up a new way for improving the yield of the sclerotium rolfsii polysaccharide by adding the intermediate metabolite, can achieve more ideal polysaccharide yield compared with the prior art, overcomes the defects of complicated fermentation operation and high carbon source demand, and is beneficial to promoting the application and development of the sclerotium rolfsii polysaccharide in industry.
Description
Technical Field
The invention relates to the field of microbial fermentation, in particular to a nutritive salt for improving the yield of sclerotium rolfsii polysaccharide and application thereof.
Background
The microbial exopolysaccharide has the advantages of unique physicochemical properties, rheological characteristics, biological safety and the like, and is widely applied to a plurality of fields of chemical industry, medicine, food, cosmetics, ecological protection and the like. The sclerotium rolfsii polysaccharide, also called scleroglucan, is a microbial extracellular polysaccharide synthesized and secreted by some filamentous fungi of the genus sclerotium, wherein the sclerotium rolfsii polysaccharide is most typically produced by fermentation of sclerotium rolfsii. The sclerotium rolfsii polysaccharide has obvious rheological property and stability in the environment with large variation range of pH value, salinity and temperature, is a neutral nonionic polysaccharide, and has wide application prospect in the fields of petroleum, paint, ceramics, food, cosmetics and the like.
However, the yield and productivity of sclerotium rolfsii polysaccharide produced by fungal fermentation are not high, so that the price of sclerotium rolfsii polysaccharide is always high, which seriously influences the application and development of sclerotium rolfsii polysaccharide in industry. Therefore, how to improve the yield of sclerotium rolfsii polysaccharide is the current research focus.
At present, the means for improving the sclerotium rolfsii polysaccharide in the prior art mainly comprises the following three aspects: screening and mutagenesis of high-yield strains, selection of fermentation culture media and optimization of fermentation conditions. For example, chinese non-patent document "fermentation method for producing Sclerotium rolfsii" discloses a high-yielding strain Sclerotium rolfsii No.1, wherein the yield of polysaccharide reaches 14.14g/L in a 16-liter self-control tank fermentation test; chinese patent document CN108441429A discloses Sclerotinium rolfsii WSH-G01, which is fermented for 56 hours under the condition that the concentration of a glucose carbon source is 75G/L to achieve the effect of 20G/L of scleroglucan yield, and the polysaccharide yields of the two schemes are still to be improved. For example, the Chinese non-patent document "a feeding control strategy for producing scleroglucan by fermenting sclerotium rolfsii" determines a feeding strategy for controlling the stirring speed to 400r/min, starting at 40h of fermentation, and feeding glucose at a constant flow rate to 400g/L, at a flow rate of 6 g/(L.h), and for a feeding maintenance time of 32h, although the method improves the yield of polysaccharide, the method needs to feed for 32h continuously in the fermentation process, the operation is complex, and the carbon source demand is high.
In conclusion, providing a scheme capable of improving the yield of sclerotium rolfsii polysaccharide, being simple and convenient to operate and saving the consumption of carbon sources is an urgent technical problem to be solved in the field.
Disclosure of Invention
Therefore, the invention aims to overcome the defects of low yield of sclerotium rolfsii polysaccharide, complex fermentation operation and high carbon source demand in the prior art, thereby providing the nutritive salt for improving the yield of sclerotium rolfsii polysaccharide and the application thereof.
In a first aspect, the present invention provides a nutritive salt for increasing the yield of sclerotium rolfsii polysaccharide, comprising: disodium 6-phosphofructose, pyruvic acid, acetic acid, acetaldehyde and glutamic acid.
Further, the nutrient salt for improving the yield of the sclerotium rolfsii polysaccharide comprises the following components in parts by weight: 0.01 to 0.2 portion of 6-fructose disodium phosphate, 0.01 to 0.1 portion of pyruvic acid, 0.01 to 0.1 portion of acetic acid, 0.01 to 0.1 portion of acetaldehyde and 0.01 to 0.1 portion of glutamic acid.
Further, the nutrient salt for improving the yield of the sclerotium rolfsii polysaccharide comprises the following components in parts by weight: 0.06-0.15 part of 6-fructose disodium phosphate, 0.01-0.1 part of pyruvic acid, 0.05-0.1 part of acetic acid, 0.02-0.1 part of acetaldehyde and 0.01-0.1 part of glutamic acid.
Further, the nutrient salt for improving the yield of sclerotium rolfsii polysaccharide comprises a component A and a component B, wherein the nutrient salt comprises the components in parts by weight,
the component A comprises: 0.01-0.1 part of 6-fructose disodium phosphate, 0.01-0.1 part of pyruvic acid and 0.01-0.1 part of acetic acid;
the component B comprises: 0.05 to 0.1 portion of 6-fructose disodium phosphate, 0.01 to 0.1 portion of acetaldehyde and 0.01 to 0.1 portion of glutamic acid.
Further, the component A comprises: 0.01 to 0.05 portion of 6-fructose disodium phosphate, 0.01 to 0.1 portion of pyruvic acid and 0.05 to 0.1 portion of acetic acid;
the component B comprises: 0.05 to 0.1 portion of 6-fructose disodium phosphate, 0.02 to 0.1 portion of acetaldehyde and 0.01 to 0.1 portion of glutamic acid.
Further, the component A comprises: 0.01 part of 6-fructose disodium phosphate, 0.01 part of pyruvic acid and 0.05 part of acetic acid;
the component B comprises: 0.05 part of 6-fructose disodium phosphate, 0.02 part of acetaldehyde and 0.01 part of glutamic acid.
Further, the component A comprises: 0.05 part of 6-fructose disodium phosphate, 0.1 part of pyruvic acid and 0.1 part of acetic acid;
the component B comprises: 0.1 part of 6-fructose disodium phosphate, 0.1 part of acetaldehyde and 0.1 part of glutamic acid.
In a second aspect, the invention provides the use of said nutritive salt for increasing the yield of sclerotium rolfsii polysaccharide in the production of sclerotium rolfsii polysaccharide.
Further, the method for producing sclerotium rolfsii polysaccharide comprises the following steps: inoculating sclerotium rolfsii into a fermentation culture medium for fermentation, and adding the nutrient salt into the fermentation culture medium in the fermentation process.
Further, the nutrient salt comprises: 0.01-0.2 g/L of 6-disodium fructose phosphate, 0.01-0.1 g/L of pyruvic acid, 0.01-0.1 g/L of acetic acid, 0.01-0.1 g/L of acetaldehyde and 0.01-0.1 g/L of glutamic acid,
further, the nutrient salt comprises a component A and a component B, based on the volume of the fermentation medium,
the component A comprises: 0.01-0.1 g/L of 6-fructose disodium phosphate, 0.01-0.1 g/L of pyruvic acid and 0.01-0.1 g/L of acetic acid;
the component B comprises: 0.05-0.1 g/L of 6-disodium fructose phosphate, 0.01-0.1 g/L of acetaldehyde and 0.01-0.1 g/L of glutamic acid,
further, the component A comprises: 0.01g/L of 6-fructose disodium phosphate, 0.01g/L of pyruvic acid and 0.05g/L of acetic acid;
the component B comprises: 0.05g/L of 6-disodium fructose phosphate, 0.02g/L of acetaldehyde and 0.01g/L of glutamic acid,
further, the component A comprises: 0.05g/L of 6-disodium fructose phosphate, 0.1g/L of pyruvic acid and 0.1g/L of acetic acid;
the component B comprises: 0.1g/L of 6-disodium fructose phosphate, 0.1g/L of acetaldehyde and 0.1g/L of glutamic acid.
Further, adding the component A into the fermentation medium during fermentation for 0-12 h;
adding the component B into the fermentation medium when fermenting for 12-24 h,
further, adding the component A into the fermentation medium during fermentation for 12 hours;
the component B was added to the fermentation medium at the time of fermentation for 24 h.
Further, controlling the pH value of the fermentation medium to be 4 +/-0.2 from the beginning of fermentation to 48 hours before fermentation;
controlling the pH value of the fermentation medium to be 3 +/-0.2 from 48h to the end of fermentation.
Furthermore, the fermentation temperature is 28 ℃, the rotation speed of a shaking table is 220rpm, and the fermentation time is 72h.
Further, the fermentation medium comprises, based on the volume of the fermentation medium: 75 to 100g/L of sucrose or glucose, 0.5 to 1.5g/L of yeast extract, 2.25 to 3g/L of sodium nitrate, 1 to 2g/L of dipotassium phosphate, 0.4 to 0.5g/L of magnesium sulfate heptahydrate, 0.4 to 0.5g/L of potassium chloride, 0 to 0.05g/L of ferrous sulfate and 0.7 to 1.5g/L of citric acid,
further, the fermentation medium comprises:
100g/L of sucrose, 1g/L of yeast extract, 2.25g/L of sodium nitrate, 2g/L of dipotassium phosphate, 0.5g/L of magnesium sulfate heptahydrate, 0.5g/L of potassium chloride, 0.05g/L of ferrous sulfate and 0.7g/L of citric acid; or alternatively
75g/L glucose, 1g/L yeast extract, 2.25g/L sodium nitrate, 1g/L dipotassium phosphate, 0.5g/L magnesium sulfate heptahydrate, 0.5g/L potassium chloride and 1.5g/L citric acid; or
95g/L glucose, 1g/L yeast extract, 3g/L sodium nitrate, 1g/L dipotassium phosphate, 0.5g/L magnesium sulfate heptahydrate, 0.5g/L potassium chloride and 1.5g/L citric acid.
The technical scheme of the invention has the following advantages:
1. the nutrient salt for improving the yield of the sclerotium rolfsii polysaccharide comprises 6-fructose phosphate disodium, pyruvic acid, acetic acid, acetaldehyde and glutamic acid, and the yield of the sclerotium rolfsii polysaccharide is greatly improved by introducing the nutrient salt into a fermentation culture medium.
2. Based on the staged characteristics of the whole fermentation process, the nutrient salt is further divided into a component A and a component B, and the nutrient salt is supplemented in a targeted manner through different fermentation stages, so that the synthesis rule of the sclerotium rolfsii polysaccharide is better met, and the yield of the sclerotium rolfsii polysaccharide is favorably further improved.
3. The invention also provides the application of the nutritive salt for improving the yield of the sclerotium rolfsii polysaccharide in the production of the sclerotium rolfsii polysaccharide, the nutritive salt is added in different periods of fermentation and the pH values in different periods are regulated and controlled, so that the synthesis of the sclerotium rolfsii polysaccharide is promoted, the yield can reach 34.1g/L, and the aims of improving the yield of the sclerotium rolfsii polysaccharide, improving the utilization rate of a substrate and reducing the environmental pollution are fulfilled.
4. The invention provides a brand-new scheme for improving the yield of the sclerotium rolfsii polysaccharide, develops a new way for improving the yield of the sclerotium rolfsii polysaccharide by adding an intermediate metabolite, can achieve more ideal polysaccharide yield compared with the prior art, overcomes the defects of complex fermentation operation and high carbon source demand, and is beneficial to promoting the industrial application and development of the sclerotium rolfsii polysaccharide.
Detailed Description
The following examples are provided to further understand the present invention, not to limit the scope of the present invention, but to provide the best mode, not to limit the content and the protection scope of the present invention, and any product similar or similar to the present invention, which is obtained by combining the present invention with other prior art features, falls within the protection scope of the present invention.
The examples do not show the specific experimental steps or conditions, and can be performed according to the conventional experimental steps described in the literature in the field. The raw materials or equipment used are all conventional products which can be obtained commercially, including but not limited to the raw materials or equipment used in the examples of the present application.
Sources of raw materials used in the examples:
the Sclerotium rolfsii strain used in the examples is Sclerotium rolfsii (Sclerotium rolfsii) ATCC 15205 which is purchased from China center for the preservation and management of industrial microorganism strains.
The PDA liquid culture medium used in the examples was prepared as follows: peeling 200g potato, cutting into pieces, adding distilled water, boiling for 30min, filtering to obtain filtrate, adding 20g glucose, and replenishing water to 1000mL.
The other raw materials, specifications and sources used in the examples are shown in Table 1.
TABLE 1 raw material specifications and sources
Example 1
The nutritive salt for improving the sclerotium rolfsii polysaccharide and the application thereof in producing the sclerotium rolfsii polysaccharide are as follows:
inoculating mature sclerotium rolfsii liquid into a PDA liquid culture medium (100 mL/250 mL), wherein the inoculation amount is 5%, the temperature is 28 ℃, the rpm is 220, and the culture is carried out for 72 hours, so as to obtain a seed liquid containing a large amount of hyphae;
inoculating the obtained seed liquid into a fermentation culture medium according to the inoculation amount of 5%, culturing at 28 ℃,220rpm for 72h, wherein a component A of nutrient salt is added into the fermentation culture medium when the fermentation is started, a component B of nutrient salt is added into the fermentation culture medium when the fermentation is 12h, the pH value of the fermentation culture medium is controlled to be 4 +/-0.2 before the fermentation is started to 48h, and the pH value of the fermentation culture medium is controlled to be 3 +/-0.2 when the fermentation is 48h to the end of the fermentation.
In this example, the composition of the fermentation medium was: sucrose 100g/L, yeast extract 1g/L, sodium nitrate (NaNO) 3 ) 2.25g/L dipotassium hydrogen phosphate (K) 2 HPO 4 ) 2g/L magnesium sulfate heptahydrate (MgSO) 4 ·7H 2 0.5g/L of O), 0.5g/L of potassium chloride (KCl), and ferrous sulfate (FeSO) 4 ) 0.05g/L and citric acid 0.7g/L.
In this example, the nutrient salt consists of component A and component B,
component A: adding 0.01g of 6-fructose disodium phosphate, 0.01g of pyruvic acid and 0.05g of acetic acid into 1L of fermentation medium;
and (B) component: 0.05g of 6-fructose disodium phosphate, 0.02g of acetaldehyde and 0.01g of glutamic acid are added into 1L of fermentation medium.
Comparative examples 1 to 1
The seed solution obtained in example 1 was inoculated into the fermentation medium of example 1 at an inoculum size of 5%, cultured at 28 ℃ and 220rpm for 72 hours without adding nutritive salts to the fermentation medium or adjusting the pH during the culture.
Comparative examples 1 to 2
The seed solution obtained in example 1 was inoculated into the fermentation medium of example 1 at an inoculum size of 5%, cultured for 72 hours at 28 ℃,220rpm, without adding nutritive salts to the fermentation medium during the culture, and the pH of the fermentation medium was controlled to 4 ± 0.2 from the start of fermentation to 48 hours before the end of fermentation, and to 3 ± 0.2 from the end of fermentation at 48 hours after the start of fermentation.
Example 2
The nutritive salt for improving the sclerotium rolfsii polysaccharide and the application thereof in producing the sclerotium rolfsii polysaccharide are as follows:
the seed solution was obtained in the same manner as in example 1;
inoculating the obtained seed liquid into a fermentation medium according to the inoculation amount of 5%, culturing at 28 ℃,220rpm for 72h, wherein the component A of the nutrient salt is added into the fermentation medium when the fermentation is 12h, the component B of the nutrient salt is added into the fermentation medium when the fermentation is 24h, the pH value of the fermentation medium is controlled to be 4 +/-0.2 before the fermentation is started to 48h, and the pH value of the fermentation medium is controlled to be 3 +/-0.2 when the fermentation is 48h to the end of the fermentation.
In this example, the composition of the fermentation medium was: 75g/L glucose, 1g/L yeast extract, sodium nitrate (NaNO) 3 ) 2.25g/L dipotassium hydrogen phosphate (K) 2 HPO 4 ) 1g/L magnesium sulfate heptahydrate (MgSO) 4 ·7H 2 O) 0.5g/L, potassium chloride (KCl) 0.5g/L, and citric acid 1.5g/L.
In this example, the nutrient salt consists of component A and component B,
and (2) component A: adding 0.025g of 6-fructose phosphate disodium, 0.05g of pyruvic acid and 0.05g of acetic acid into 1L of fermentation medium;
and (B) component: 0.05g of 6-fructose disodium phosphate, 0.025g of acetaldehyde and 0.05g of glutamic acid are added into 1L of fermentation medium.
Comparative example 2-1
The seed solution obtained in example 1 was inoculated into the fermentation medium of example 2 at an inoculum size of 5%, cultured at 28 ℃ and 220rpm for 72 hours, and no nutrient salt was added to the fermentation medium during the culture and no pH adjustment was performed.
Comparative examples 2 to 2
The seed solution obtained in example 1 was inoculated into the fermentation medium of example 2 at an inoculum size of 5%, cultured at 28 ℃ at 220rpm for 72 hours, without adding nutritive salts to the fermentation medium during the culture, and the pH of the fermentation medium was controlled to 4 ± 0.2 from the start of fermentation to 48 hours before the end of fermentation, and to 3 ± 0.2 from the end of fermentation at 48 hours after the start of fermentation.
Example 3
The nutritive salt for improving the sclerotium rolfsii polysaccharide and the application thereof in producing the sclerotium rolfsii polysaccharide are as follows:
the seed liquid was obtained in the same manner as in example 1;
inoculating the obtained seed liquid into a fermentation medium according to the inoculation amount of 5%, culturing at 28 ℃,220rpm for 72h, wherein the component A of the nutrient salt is added into the fermentation medium when the fermentation is 12h, the component B of the nutrient salt is added into the fermentation medium when the fermentation is 24h, the pH value of the fermentation medium is controlled to be 4 +/-0.2 before the fermentation is started to 48h, and the pH value of the fermentation medium is controlled to be 3 +/-0.2 when the fermentation is 48h to the end of the fermentation.
In this example, the composition of the fermentation medium was: 95g/L glucose, 1g/L yeast extract, sodium nitrate (NaNO) 3 ) 3g/L of dipotassium hydrogen phosphate (K) 2 HPO 4 ) 1g/L magnesium sulfate heptahydrate (MgSO) 4 ·7H 2 O) 0.5g/L, potassium chloride (KCl) 0.5g/L, and citric acid 1.5g/L.
In this example, the nutrient salt consists of component A and component B,
and (2) component A: 0.05g of 6-fructose disodium phosphate, 0.1g of pyruvic acid and 0.1g of acetic acid are added into each 1L of fermentation medium;
and (B) component: 0.1g of 6-fructose disodium phosphate, 0.1g of acetaldehyde and 0.1g of glutamic acid are added into 1L of fermentation medium.
Comparative example 3-1
The seed solution obtained in example 1 was inoculated into the fermentation medium of example 3 at an inoculum size of 5%, cultured at 28 ℃ and 220rpm for 72 hours, and no nutrient salt was added to the fermentation medium during the culture and no pH adjustment was performed.
Comparative examples 3 to 2
The seed solution obtained in example 1 was inoculated into the fermentation medium of example 3 at an inoculum size of 5%, and cultured at 28 ℃ at 220rpm for 72 hours, without adding nutritive salts to the fermentation medium during the culture, the fermentation medium pH was controlled to 4 ± 0.2 from the start of fermentation to 48 hours before the end of fermentation, and the fermentation medium pH was controlled to 3 ± 0.2 from the end of fermentation at 48 hours after the start of fermentation.
Example 4
The nutritive salt for improving the sclerotium rolfsii polysaccharide and the application thereof in producing the sclerotium rolfsii polysaccharide are as follows:
the seed solution was obtained in the same manner as in example 1;
inoculating the obtained seed liquid into a fermentation medium according to the inoculation amount of 5%, culturing at 28 ℃,220rpm for 72h, wherein the component A of the nutrient salt is added into the fermentation medium when the fermentation is 12h, the component B of the nutrient salt is added into the fermentation medium when the fermentation is 24h, and the pH value of the fermentation medium is controlled to be 3 +/-0.2 from the beginning to the end of the fermentation.
In this example, the composition of the fermentation medium and the composition of the nutrient salts were the same as in example 3.
Example 5
The nutritive salt for improving the sclerotium rolfsii polysaccharide and the application thereof in producing the sclerotium rolfsii polysaccharide are as follows:
the seed solution was obtained in the same manner as in example 1;
inoculating the obtained seed liquid into a fermentation culture medium according to the inoculation amount of 5%, culturing at 28 ℃,220rpm for 72h, wherein the component A and the component B of nutrient salts are simultaneously added into the fermentation culture medium when the fermentation is 12h, the pH value of the fermentation culture medium is controlled to be 4 +/-0.2 from the beginning of the fermentation to 48h, and the pH value of the fermentation culture medium is controlled to be 3 +/-0.2 from the end of the fermentation to 48 h.
In this example, the composition of the fermentation medium and the composition of the nutrient salts are the same as in example 3.
Examples of the experiments
The fermentation liquid obtained in the example and the comparative example after fermentation for 72 hours was sampled, and sclerotium rolfsii polysaccharide was extracted and the content thereof was measured.
The extraction method of the sclerotium rolfsii polysaccharide comprises the following steps: and (3) filtering the fermentation liquor after fermenting for 72 hours to obtain fermentation filtrate, adding four times of volume of absolute ethyl alcohol, carrying out overnight alcohol precipitation at 4 ℃, centrifuging for 20min at 5000rmp to obtain precipitate, and adding water to the precipitate for redissolving to obtain a sample to be detected.
The content determination method of the sclerotium rolfsii polysaccharide comprises the following steps: phenol-sulfuric acid method, measuring system is 1.00mL sample to be measured, adding 1.0mL 5% phenol solution, then placing in ice bath and quickly adding 5.0mL concentrated sulfuric acid, standing for 30min at room temperature, measuring absorbance at 490nm, calculating the concentration of sclerotium rolfsii polysaccharide according to standard curve, wherein,
standard curve: y =9.0153x-0.0129R 2 ＝0.9963
y-absorbance x-micronucleus polysaccharide concentration
Calculating the weight of the sclerotium rolfsii polysaccharide according to the measured concentration of the sclerotium rolfsii polysaccharide and the volume of a sample to be measured, and calculating the yield of the sclerotium rolfsii polysaccharide according to the following formula:
the yield of sclerotium rolfsii polysaccharide = (weight of sclerotium rolfsii polysaccharide/volume of fermentation broth) × 100%.
The yields of sclerotium rolfsii polysaccharide finally obtained for each example and comparative example are shown in table 2.
TABLE 2 production of sclerotium rolfsii polysaccharides
| Yield of sclerotium rolfsii polysaccharide (g/L) | |
| Example 1 | 32.2 |
| Comparative examples 1 to 1 | 21.7 |
| Comparative examples 1 to 2 | 29.4 |
| Example 2 | 25.4 |
| Comparative example 2-1 | 18.7 |
| Comparative examples 2 to 2 | 22.3 |
| Example 3 | 34.1 |
| Comparative example 3-1 | 21.5 |
| Comparative examples 3 to 2 | 29.6 |
| Example 4 | 23.4 |
| Example 5 | 31.2 |
As can be seen from Table 2, the yield of example 1 is increased by 48.4% compared with the case of not adding nutritive salt and not adjusting pH during fermentation (comparative example 1-1), and the yield is increased by 9.5% compared with the case of not adding nutritive salt and adjusting pH correspondingly during fermentation (comparative example 1-2); example 2 the yield was increased by 35.8% compared to the case of no addition of nutrient salts and no adjustment of pH during fermentation (comparative example 2-1), and 13.9% compared to the case of no addition of nutrient salts and a corresponding adjustment of pH during fermentation (comparative example 2-2); example 3 the yield was increased by 58.6% compared to the case of no addition of nutrient salts and no adjustment of pH during fermentation (comparative example 3-1), and 15.2% compared to the case of no addition of nutrient salts and a corresponding adjustment of pH during fermentation (comparative example 3-2); example 3 compared with the scheme of not regulating the pH value stage by stage (example 4), the yield is improved by 45.7 percent; example 3 the yield was improved by 9.3% compared to the case where component A and component B were added simultaneously (example 5).
Therefore, the nutritive salt provided by the invention can effectively improve the yield of the sclerotium rolfsii polysaccharide, and can further improve the yield by matching with the staged regulation and control of the pH value. In addition, the staged addition of component a and component B also provides a certain increase in yield compared to a single addition of nutrient salt.
It should be understood that the above examples are only for clarity of illustration and are not intended to limit the embodiments. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. And are neither required nor exhaustive of all embodiments. And obvious variations or modifications therefrom are within the scope of the invention.
Claims (6)
1. The application of a nutritive salt for improving the yield of sclerotium rolfsii polysaccharide in the production of sclerotium rolfsii polysaccharide is characterized in that the method for producing sclerotium rolfsii polysaccharide comprises the following steps: inoculating sclerotium rolfsii into a fermentation culture medium for fermentation, adding the nutrient salt into the fermentation culture medium in the fermentation process,
the nutrient salt comprises a component A and a component B, based on the volume of the fermentation medium,
the component A comprises: 0.01-0.1 g/L of 6-disodium fructose phosphate, 0.01-0.1 g/L of pyruvic acid and 0.01-0.1 g/L of acetic acid;
the component B comprises: 0.05-0.1 g/L of 6-disodium fructose phosphate, 0.01-0.1 g/L of acetaldehyde and 0.01-0.1 g/L of glutamic acid,
adding the component A into the fermentation medium when fermenting for 0-12 h;
adding the component B into the fermentation medium when fermenting for 12-24 h,
controlling the pH value of the fermentation medium to be 4 +/-0.2 from the beginning of fermentation to 48 hours before fermentation;
controlling the pH value of the fermentation medium to be 3 +/-0.2 from 48h to the end of fermentation.
2. The use according to claim 1,
the component A comprises: 0.01g/L of 6-disodium fructose phosphate, 0.01g/L of pyruvic acid and 0.05g/L of acetic acid;
the component B comprises: 0.05g/L of 6-disodium fructose phosphate, 0.02g/L of acetaldehyde, 0.01g/L of glutamic acid, or
The component A comprises: 0.05g/L of 6-disodium fructose phosphate, 0.1g/L of pyruvic acid and 0.1g/L of acetic acid;
the component B comprises: 0.1g/L of 6-disodium fructose phosphate, 0.1g/L of acetaldehyde and 0.1g/L of glutamic acid.
3. The use according to claim 1,
adding the component A into the fermentation medium during fermentation for 12 h;
the component B was added to the fermentation medium at the time of fermentation for 24 h.
4. The use according to claim 1, wherein the fermentation temperature is 28 ℃, the shaker speed is 220rpm, and the fermentation time is 72h.
5. Use according to claim 1, wherein the fermentation medium comprises, based on the volume of the fermentation medium: 75-100 g/L of sucrose or glucose, 0.5-1.5 g/L of yeast extract, 2.25-3 g/L of sodium nitrate, 1-2 g/L of dipotassium phosphate, 0.4-0.5 g/L of magnesium sulfate heptahydrate, 0.4-0.5 g/L of potassium chloride, 0-0.05 g/L of ferrous sulfate and 0.7-1.5 g/L of citric acid.
6. Use according to claim 5, wherein the fermentation medium comprises, based on the volume of the fermentation medium:
100g/L of sucrose, 1g/L of yeast extract, 2.25g/L of sodium nitrate, 2g/L of dipotassium phosphate, 0.5g/L of magnesium sulfate heptahydrate, 0.5g/L of potassium chloride, 0.05g/L of ferrous sulfate and 0.7g/L of citric acid; or
75g/L glucose, 1g/L yeast extract, 2.25g/L sodium nitrate, 1g/L dipotassium phosphate, 0.5g/L magnesium sulfate heptahydrate, 0.5g/L potassium chloride and 1.5g/L citric acid; or
95g/L glucose, 1g/L yeast extract, 3g/L sodium nitrate, 1g/L dipotassium phosphate, 0.5g/L magnesium sulfate heptahydrate, 0.5g/L potassium chloride and 1.5g/L citric acid.
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