CN113755407A - Paenibacillus mucilaginosus, exopolysaccharide prepared by same and application of exopolysaccharide in preparation of microbial flocculant - Google Patents

Abstract

The invention belongs to preparation of microbial polysaccharides, and particularly relates to paenibacillus mucilaginosus, prepared exopolysaccharides and application of the exopolysaccharides in preparation of microbial flocculants. The bacillus mucilaginosus with the name of Latin, Paenibaci llus mucilaginosus and the preservation number of CGMCC No.23105 is inoculated into an activation culture medium for activation; inoculating the activated strain into a fermentation medium containing a carbon source, a nitrogen source and nutrient elements, culturing under stirring to prepare a fermentation liquid, and collecting extracellular polysaccharide from the fermentation liquid. The invention solves the technical problems of large using amount, low flocculation capacity and the like of microbial flocculants prepared by the prior art, and has the advantages of good flocculation capacity of the prepared extracellular polysaccharide, easy control of a fermentation process, short period and the like.

Description

Paenibacillus mucilaginosus, exopolysaccharide prepared by same and application of exopolysaccharide in preparation of microbial flocculant

Technical Field

The invention belongs to preparation of microbial polysaccharides, and particularly relates to paenibacillus mucilaginosus, prepared exopolysaccharides and application of the exopolysaccharides in preparation of microbial flocculants.

Background

Flocculation is a solid-liquid separation technology with low cost and simple operation, and is widely applied in the fields of water quality treatment, fermentation and recovery, impurity removal and the like. As modern society develops and population grows, the importance of flocculation is increasingly evident: at present, the discharge amount of various untreated or improperly treated wastewater far exceeds the bearing capacity of the natural environment, for example, the Yangtze river basin with rich total water resource, except Yangtze river dry flow and large reservoirs with strong self-purification capacity of water bodies, most lakes, small reservoirs and plain water networks commonly have the problem of water quality shortage, according to estimation, 58 percent of used water can return to a natural water area in the form of polluted water, so that the water quality is difficult to improve, and the flocculation technology can be used for in-situ protection and restoration of ecological environment and landscape, can also be used for water quality treatment of drinking water plants and sewage plants, and can remove gravel, humus and partial pathogenic bacteria, or treat wastewater from sugarcane industry, animal husbandry and the like; based on the aggregation effect of flocculation on organic matters and microbial cells, industries such as microalgae fermentation and monoclonal antibody production also have the requirement on flocculation, and the method is used for solving a key bottleneck limiting the microbial culture and fermentation industries, namely the high cost of harvesting products from a liquid culture medium; the flocculation technology can also be used for the extraction process of plant small molecules and can remove large molecular impurities such as polysaccharide, protein and the like.

The microbial flocculant is a natural high molecular polymer with flocculation activity produced by a microbial fermentation technology, mainly exists in an extracellular polymer with polysaccharide, protein and nucleic acid as main components, and has the characteristics of safety, biodegradability and the like.

Compared with common inorganic polymeric flocculants such as polyacrylamide and derivatives thereof, the microbial flocculants are biocompatible and do not cause secondary pollution such as carcinogenesis and neurotoxicity. The microbial flocculant taking polysaccharide as a main component has structural characteristics of high molecular weight and various functional groups, can adsorb inorganic matters and organic matters, destabilizes the inorganic matters and the organic matters, aggregates and settles the inorganic matters and the organic matters to realize solid-liquid separation, shows excellent flocculation activity and color removal capability, is generally nontoxic and harmless and has biodegradability, so the microbial flocculant is expected to be a substitute of an artificial polymer flocculant and is a hotspot direction for developing novel green flocculants. But the microbial flocculant has the characteristics of low yield, large dosage and low flocculation efficiency. Research shows that the higher the uronic acid content and the higher the molecular weight of the polysaccharide, the higher the flocculation activity.

The background art retrieved by the applicant includes:

the patent document with publication number CN102952834 discloses a method for producing a microbial polysaccharide fermentation liquor by utilizing Paenibacillus mucilaginosus, wherein the fermentation period is 60h, and the polysaccharide content in the fermentation liquor can reach 7.5g/L by adopting an anthrone colorimetric method. The structure of the polysaccharide was not examined.

The patent document with the publication number CN102628065 discloses a production method and application of a microbial flocculant, wherein a fermentation liquid is obtained by fermenting geotrichum candidum seeds for 3-5 days, the fermentation period is long, the structure of polysaccharide is not detected, 0.1g of the microbial flocculant needs to be added into 50ml of karst water, and the addition amount of the microbial flocculant is large.

The applicant does not find the same or similar literature reports in the prior art as in the present application.

Disclosure of Invention

The invention aims to provide paenibacillus mucilaginosus.

The second purpose of the invention is to provide exopolysaccharides prepared by using Paenibacillus mucilaginosus.

The invention also aims to provide a preparation method of the extracellular polysaccharide.

The fourth purpose of the invention is to provide the application of the exopolysaccharide in the preparation of microbial flocculants.

The overall technical concept of the invention is as follows:

paenibacillus mucilaginosus is named as Paenibacillus mucinosus, and the preservation number is CGMCC No. 23105.

The strain of the invention is submitted to China general microbiological culture Collection center of China general microbiological culture Collection center No. 3 of Xilu No.1 Hospital, Beijing, Chaoyang, on 8.2 days in 2021, wherein the preservation unit is abbreviated as CGMCC and the preservation number is CGMCC No. 23105. The classification named Paenibacillus mucilaginosus strain mucinaginosus is suggested.

The strain is obtained by the applicant through sampling, separating and purifying the wetland in the constant water lake of the constant water city of Hebei province, and the spore is oval and the sporangium is slightly expanded and is close to the center. The colony is round, transparent and convex, and viscous.

The cell morphology and physicochemical characteristics of the strain of the invention are as follows:

the exopolysaccharide prepared by the paenibacillus mucilaginosus has the chemical composition of mannose: glucose: galactose: the molar ratio of glucuronic acid is 2:1:2:1, and the polysaccharide has the following structural formula:

the general structural formula is as follows:

→{3)-α-D-Manp-(1→3)-4-Suc-β-D-Manp-(1→3)-β-D-Glcp-(1→4) -β-D-GlcpA-(1→4)[4，6-Pyr-β-D-Galp-(1→3)]-β-D-Galp-(1→}_n→

wherein n is10⁴-10⁶Suc succinic acid group, Pyr pyruvic acid group.

A method for preparing exopolysaccharide, comprising the steps of:

A. inoculating Paenibacillus mucilaginosus with the name of Latin and the preservation number of CGMCC No.23105 into an activation culture medium for activation;

B. b, inoculating the activated strain in the step A into a fermentation medium containing a carbon source, a nitrogen source and nutrient elements, and culturing under stirring to prepare a fermentation liquid;

C. and C, collecting exopolysaccharides from the fermentation liquor prepared in the step B.

Application of exopolysaccharide in preparation of microbial flocculant.

The concrete and technical conception of the invention are as follows:

the 16S rRNA gene sequence of the strain is shown as SEQ No. 1.

In order to realize industrial production, the preferable technical implementation means is that the step B is to inoculate the activated strain in the step A into a seed culture medium to prepare a seed solution after amplification culture, and inoculate the seed solution into a fermentation culture medium according to the inoculum size of 4-10% of the volume ratio to prepare fermentation liquor.

In order to realize better activation effect, the more preferable technical implementation means is that the activation in the step A is to inoculate the bacillus mucilaginosus with the Latin name of Paenibacillus mucilaginosus and the preservation number of CGMCC No.23105 into an activation culture medium and activate the bacillus mucilaginosus for 24-48 hours at the temperature of 28-40 ℃.

In order to realize the control of the culture end point, the preferable technical means is that the culture of the seed liquid is stopped when the OD value is more than or equal to 1.0 under the condition of 550nm wavelength.

As the viscosity of the fermentation liquor is higher and higher along with the increase of time in the fermentation process, in order to ensure the dissolved oxygen level in the fermentation process and facilitate the growth of thalli and the synthesis of final products, the preferable technical implementation means is that the culture condition in the step B is 28-40 ℃, the stirring speed is 100-300 r/min, and the culture time is 16-50 hours.

In order to realize the control of the fermentation end point in the culture process, the preferable technical implementation means is that the fermentation is stopped when the viscosity of the fermentation liquid is more than or equal to 500cp in the step B.

In order to realize the extraction of the extracellular polysaccharide in the fermentation broth, the preferable technical implementation means is that the ethanol solution with the volume percentage concentration of 80-95% is adopted in the step C to settle the extracellular polysaccharide in the fermentation broth, and the volume ratio of the ethanol solution to the fermentation broth is 2-4.

In order to facilitate the growth of the strain and the synthesis of the final product, the preferred technical implementation is that the culture medium used in the step A, B comprises the following components:

20 g/L to 50 g/L of cane sugar; 1 g/L-2.5 g/L of potassium nitrate; 1 g/L-2 g/L of sodium dihydrogen phosphate; calcium chloride 0.01 g/L-1 g/L; magnesium sulfate heptahydrate 0.1 g/L-1 g/L; ferrous sulfate heptahydrate 0.01 g/L-0.05 g/L; 0.001 g/L-0.01 g/L of manganese sulfate monohydrate; 0.001 g/L-0.01 g/L of zinc chloride; the pH value is 6-8.

The invention achieves the substantive characteristics and obvious technical progress that:

1. the invention provides a paenibacillus jelly, which can synthesize extracellular polysaccharide with flocculation capacity.

2. The preparation method of the extracellular polysaccharide is simple and easy to implement, short in fermentation period and mild in conditions.

3. The extracellular polysaccharide has the advantages of high uronic acid content and large molecular weight, the dosage required for achieving the same flocculation effect (more than 98%) through the test of the applicant is basically consistent with that of polyacrylamide, and the dosage is 1ppm for kaolin suspension and 0.5ppm to 1ppm for coal powder. Has better flocculation activity, and effectively solves the technical problems of low flocculation activity, large flocculant dosage and the like of the existing product.

Drawings

FIG. 1 is a photomicrograph of a plate colony of the strain of the present invention.

FIG. 2 is an infrared chromatogram for determination of the exopolysaccharide component of the invention.

FIG. 3 is a liquid chromatogram for determination of the exopolysaccharide fraction of the present invention.

FIG. 4 shows exopolysaccharides according to the invention¹³C nuclear magnetic resonance spectroscopy.

Fig. 5 is a test presentation of a coal powder suspension at various times.

FIG. 6 is a 16S rRNA sequence listing of the strains of the present invention.

The strain of the invention is submitted to China general microbiological culture Collection center of China general microbiological culture Collection center No. 3 of Xilu No.1 Hospital, Beijing, Chaoyang, on 8.2 days in 2021, wherein the preservation unit is abbreviated as CGMCC and the preservation number is CGMCC No. 23105. The classification named Paenibacillus mucilaginosus strain mucinaginosus is suggested.

In order to verify the structure and flocculation activity of the exopolysaccharide of the present invention, the applicant performed the following experiments:

1. structural analysis of polysaccharide flocculants

(1) Infrared spectroscopy detection

The infrared scanning instrument used was NICOLETIS10 (Seimer Feishell science). And (3) putting the pressed potassium bromide blank sheet on a sample frame of a sample bin of an infrared scanner, confirming to collect a reference background spectrum, and then putting a sample to be detected into a spectrometer to scan the sample. The sample dosage is 2-5 mg each time, and the result is shown in figure 2:

the infrared spectrum result shows that 3000-3700 cm^-1The broad absorption peaks of (a) belong to hydroxyl groups of various environments in polysaccharide molecules; 2931cm^-1The peak of (a) is from C-H stretching and bending vibrations; 1100-1200 cm^-1The peak of (A) belongs to the C-O-C linkage, i.e.the glycosidic bond in the polysaccharide.

(2) High performance liquid chromatography detection

The extracellular polysaccharide florcuronic acid prepared by the strain of the invention is completely hydrolyzed by trifluoroacetic acid, derivatized by PMP, extracted by chloroform and filtered by water phase, and then high-efficiency liquid phase analysis is carried out. FIG. 3 is a liquid chromatogram for the determination of the monosaccharide composition. As can be seen from FIG. 3, the extracellular polysaccharide, fluccuronic acid, prepared by using the strain of the present invention is an acidic heteropolysaccharide composed of mannose, glucose, galactose and glucuronic acid in a molar ratio of 2:1:2: 1.

(3) Nuclear magnetic detection

The extracellular polysaccharide fluccuronic acid prepared by the strain of the invention is partially hydrolyzed by trifluoroacetic acid and is partially hydrolyzed by D₂O was prepared at 80mg/ml, and NMR analysis was performed using TMSP as an internal standard. The one-dimensional nuclear magnetic carbon spectrum of the polysaccharide is shown in FIG. 4. In 13C NMR, there were 6 anomeric carbon signals and a sharp signal between Δ 102-106ppm, indicating that fluccuronic acid has 6 anomeric carbons and 1 pyruvate group. Chemical shifts 179.26ppm in 13C NMR were assigned to the carboxyl carbon atom of glucuronic acid, 178.66ppm to the carboxyl carbon atom of the pyruvic acid group, 183.91ppm and 178.52ppm to the succinyl group, with the succinyl group peaking at another 2 carbon atoms at 34.76ppm and 33.26ppm, respectively.

In combination with the above analysis, it can be confirmed that the structural formula of extracellular polysaccharide flucuronic acid produced by Paenibacillus mucilaginosus (Paenibacillus mucinagino Sus) CGMCC No.23105 is as follows:

wherein n is10⁴-10⁶Suc succinic acid group, Pyr pyruvic acid group.

2. Flocculation activity test (turbidity removal rate) of polysaccharide flocculants

At present, a kaolin suspension or a coal powder suspension is usually used as simulated wastewater in a flocculation capability test, and the most widely used inorganic flocculant polyacrylamide is used as a control group.

100mL of coal ash suspension (the mass concentration is 4g/L) is prepared, a certain amount of flocculant is added for flocculation experiment, and the blank is the coal ash suspension without the flocculant. Magnetic stirring (fast 3 minutes, slow 1 minute, rest 5 minutes), photo record the phenomenon, as in figure 6, and measure the change in absorbance. And (3) taking deionized water as a reference, sucking liquid 2cm below the liquid surface, and measuring blank and absorbance of a flocculation system at 550nm to calculate flocculation activity. The calculation formula of the flocculation activity is as follows:

flocculation activity ═ A₀－A₁)/A₀×100％

Wherein A is₀Absorbance of a blank coal ash suspension, A₁The absorbance of the coal ash suspension was measured. As a result, 0.5ppm, 1ppm and 2ppm of the extracellular polysaccharide floccuronic acid prepared by using the strain of the present invention reached flocculation activities of 89.3%, 93.7% and 91.2%, respectively, as shown in FIG. 5.

3. Amount of flocculant

Through experimental comparison, the dosage of the extracellular polysaccharide prepared by the invention for achieving the same flocculation effect (more than 98%) is basically consistent with that of polyacrylamide, and the dosage is 1ppm for kaolin suspension and 0.5ppm to 1ppm for coal powder.

Detailed Description

The present invention is further described with reference to the following examples, which are not intended to limit the scope of the present invention, and the claims are not to be interpreted as limiting the scope of the present invention.

Example 1

The preparation method of exopolysaccharide in the embodiment comprises the following steps:

A. inoculating Paenibacillus mucilaginosus with the name of Latin and the preservation number of CGMCC No.23105 into an activation culture medium for activation;

B. b, inoculating the activated strain in the step A into a fermentation medium containing a carbon source, a nitrogen source and nutrient elements, and culturing under stirring to prepare a fermentation liquid;

C. and C, collecting exopolysaccharides from the fermentation liquor prepared in the step B.

Application of exopolysaccharide in preparation of microbial flocculant.

The 16S rRNA gene sequence of the strain in the embodiment is shown as SEQ No. 1.

And B, inoculating the activated strain in the step A into a seed culture medium, performing amplification culture to prepare a seed solution, and inoculating the seed solution into a fermentation culture medium according to the inoculum size of 4% by volume to prepare a fermentation liquid.

The activation in the step A is to inoculate the Paenibacillus mucilaginosus with the preservation number of CGMCC No.23105 and activate the Paenibacillus mucilaginosus for 24 hours at the temperature of 28 ℃ under the Latin name.

The preparation of the seed liquid is that the seed liquid culture is stopped when the OD value is more than or equal to 1.0 under the condition of the wavelength of 550 nm.

The culture conditions in step B were a temperature of 28 ℃ and a stirring speed of 100 rpm for 16 hours.

And B, stopping fermentation when the viscosity of the fermentation liquor is more than or equal to 500 cp.

And C, adopting an ethanol solution with the volume percentage concentration of 80% to settle the extracellular polysaccharide in the fermentation liquor, wherein the volume ratio of the ethanol solution to the fermentation liquor is 2.

The culture medium used in step A, B includes the following components:

20 g/L of cane sugar; 1 g/l of potassium nitrate; 1 g/L of sodium dihydrogen phosphate; calcium chloride 0.01 g/l; magnesium sulfate heptahydrate 0.1 g/L; ferrous sulfate heptahydrate 0.01 g/L; 0.001 g/L manganese sulfate monohydrate; zinc chloride 0.001 g/l; the pH value is 6-8.

Example 2

This example differs from example 1 in that:

and B, inoculating the activated strain in the step A into a seed culture medium, performing amplification culture to prepare a seed solution, and inoculating the seed solution into a fermentation culture medium according to the inoculation amount of 10% by volume to prepare a fermentation liquid.

The activation in the step A is to inoculate the Paenibacillus mucilaginosus with the preservation number of CGMCC No.23105 and activate the Paenibacillus mucilaginosus for 48 hours at the temperature of 40 ℃ under the Latin name.

The culture conditions in step B were 40 ℃ temperature, 300 rpm stirring speed and 50 hours culture time.

And C, adopting an ethanol solution with the volume percentage concentration of 95% to settle the extracellular polysaccharide in the fermentation liquor, wherein the volume ratio of the ethanol solution to the fermentation liquor is 4.

The culture medium used in step A, B includes the following components:

50 g/L of cane sugar; 2.5 g/l of potassium nitrate; 2 g/L of sodium dihydrogen phosphate; 1 g/L of calcium chloride; magnesium sulfate heptahydrate 1 g/L; ferrous sulfate heptahydrate 0.05 g/L; 0.01 g/L manganese sulfate monohydrate; zinc chloride 0.01 g/l; the pH value is 6-8.

The rest is the same as in example 1.

Example 3

This example differs from example 1 in that:

and B, inoculating the activated strain in the step A into a seed culture medium, performing amplification culture to prepare a seed solution, and inoculating the seed solution into a fermentation culture medium according to an inoculation amount of 7% by volume to prepare a fermentation liquid.

The activation in the step A is to inoculate the Paenibacillus mucilaginosus with the preservation number of CGMCC No.23105 and activate the Paenibacillus mucilaginosus for 36 hours at the temperature of 34 ℃ under the Latin name.

The culture conditions in step B were 34 ℃ temperature, 200 rpm stirring speed, and 33 hours culture time.

And C, adopting an ethanol solution with the volume percentage concentration of 88% to settle the extracellular polysaccharide in the fermentation liquor, wherein the volume ratio of the ethanol solution to the fermentation liquor is 3.

The culture medium used in step A, B includes the following components:

35 g/L of cane sugar; potassium nitrate 1.7 g/l; 1.5 g/L of sodium dihydrogen phosphate; calcium chloride 0.5 g/l; magnesium sulfate heptahydrate 0.55 g/L; ferrous sulfate heptahydrate 0.03 g/L; manganese sulfate monohydrate 0.005 g/L; zinc chloride 0.005 g/l; the pH value is 6-8.

The rest is the same as in example 1.

Example 4

This example differs from example 1 in that:

and B, inoculating the activated strain in the step A into a seed culture medium, performing amplification culture to prepare a seed solution, and inoculating the seed solution into a fermentation culture medium according to an inoculation amount of 5% by volume to prepare a fermentation liquid.

The activation in the step A is to inoculate the Paenibacillus mucilaginosus with the preservation number of CGMCC No.23105 and activate the Paenibacillus mucilaginosus for 30 hours at the temperature of 31 ℃ in Latin.

The culture conditions in step B were 31 ℃ temperature, 150 rpm stirring speed, and 25 hours culture time.

And C, precipitating the extracellular polysaccharide in the fermentation liquor by adopting an ethanol solution with the volume percentage concentration of 80-95%, wherein the volume ratio of the ethanol solution to the fermentation liquor is 2.5.

The culture medium used in step A, B includes the following components:

25 g/L of sucrose; potassium nitrate 1.3 g/l; 1.3 g/L of sodium dihydrogen phosphate; calcium chloride 0.3 g/l; magnesium sulfate heptahydrate 0.3 g/L; ferrous sulfate heptahydrate 0.02 g/L; 0.003 g/L of manganese sulfate monohydrate; zinc chloride 0.003 g/L; the pH value is 6-8.

The rest is the same as in example 1.

Example 5

This example differs from example 1 in that:

and B, inoculating the activated strain in the step A into a seed culture medium, performing amplification culture to prepare a seed solution, and inoculating the seed solution into a fermentation culture medium according to the inoculation amount of 9% by volume to prepare a fermentation liquid.

The activation in the step A is to inoculate the Paenibacillus mucilaginosus with the preservation number of CGMCC No.23105 and activate the Paenibacillus mucilaginosus for 42 hours at the temperature of 38 ℃ in Latin.

The culture conditions in step B were 38 ℃ temperature, 260 rpm stirring speed and 37 hours culture time.

And C, adopting an ethanol solution with the volume percentage concentration of 92% to settle the extracellular polysaccharide in the fermentation liquor, wherein the volume ratio of the ethanol solution to the fermentation liquor is 3.5.

The culture medium used in step A, B includes the following components:

sucrose 40 g/l; 2.2 g/l of potassium nitrate; 1.8 g/L of sodium dihydrogen phosphate; calcium chloride 0.08 g/l; magnesium sulfate heptahydrate 0.8 g/L; ferrous sulfate heptahydrate 0.04 g/L; 0.008 g/L manganese sulfate monohydrate; zinc chloride 0.008 g/l; the pH value is 6-8.

The rest is the same as in example 1.

Sequence listing

<110> Huchuan Biotech Co., Ltd

<120> Paenibacillus mucilaginosus, exopolysaccharide prepared by same and application of exopolysaccharide in preparation of microbial flocculant

<130> CN102952834

<160> 1

<170> SIPOSequenceListing 1.0

<210> 2

<211> 1405

<212> DNA

<213> Paenibacillus mucilaginosus (2 Ambystoma laterale x Ambystoma jeffersonia)

<400> 2

cggctggctc ccttgcgggt taccccaccg gcttcgggtg ttgtaaactc tcgtggtgtg 60

acgggcggtg tgtacaagac ccgggaacgt attcaccgcg gcatgctgat ccgcgattac 120

tagcaattcc gacttcatgc aggcgagttg cagcctgcaa tccgaactga gaccggcttc 180

taaggattcg ctccatctcg cgacttcgct tcccgttgta ccggccattg tagtacgtgt 240

gtagcccagg tcataagggg catgatgatt tgacgtcatc cccaccttcc tccggtttgt 300

caccggcagt cactctagag tgcccaactc aatgctggca actaaagtca agggttgcgc 360

tcgttgcggg acttaaccca acatctcacg acacgagctg acgacaacca tgcaccacct 420

gtcacctctg tcccgaagga ggaccctatc tctagggctt tcagagggat gtcaagacct 480

ggtaaggttc ttcgcgttgc ttcgaattaa accacatact ccactgcttg tgcgggtccc 540

cgtcaattcc tttgagtttc actcttgcga gcgtactccc caggcggagt gcttattgtg 600

tttacttcgg caccaagggt atcgaaaccc ctaacaccta gcactcatcg tttacggcgt 660

ggactaccag ggtatctaat cctgtttgct ccccacgctt tcgcgcctca gcgtcagtta 720

cagtccagaa agccgccttc gccactggtg ttcctccaca tctctacgca tttcaccgct 780

acacgtggaa ttccgctttc ctctcctgca ctcaagtctt ccagtttccg gtgcgaaccg 840

gggttgagcc ccgggcttaa acaccagact taaaaaaccg cctgcgcgcg ctttacgccc 900

aataattccg gacaacgctt gccccctacg tattaccgcg gctgctggca cgtagttagc 960

cggggctttc ttctcaggta ccgtcattcg cagagcagtt actctccacg acattcttcc 1020

ctggcaacag agctttacga tccgaaaacc ttcatcactc acgcggcgtt gctccgtcag 1080

gcttgcgccc attgcggaag attccctact gctgcctccc gtaggagtct gggccgtgtc 1140

tcagtcccag tgtggccgat caccctctca ggtcggctac gcatcgtcgc cttggtgggc 1200

cgttaccccg ccaactagct aatgcgccgc aggcccatcc gtaagccaca ggttgccccg 1260

tgtttcatga ttccggcatg caccgaaacc agctatccgg tcttagctac cgtttccggt 1320

agttatcccg atcttacagg caggttgcct acgtgttact cacccgtccg ccgctaagca 1380

ccgaagtgct ccgctcgact tgcat 1405

Claims (12)

1. Paenibacillus mucilaginosus is characterized in that the Latin Wen name of the strain is Paenibacillus mucilaginosus, and the preservation number is CGMCC No. 23105.

2. The Paenibacillus mucilaginosus of claim 1, wherein the 16SrRNA gene sequence of the strain is shown in SEQ No. 1.

3. Exopolysaccharides produced by using Paenibacillus mucilaginosus according to claim 1, characterized in that the chemical composition of the polysaccharides is mannose: glucose: galactose: the molar ratio of glucuronic acid is 2:1:2:1, and the polysaccharide has the following structural formula:

the general structural formula is as follows:

→{3)-α-D-Manp-(1→3)-4-Suc-β-D-Manp-(1→3)-β-D-Glcp-(1→4)-β-D-GlcpA-(1→4)[4，6-Pyr-β-D-Galp-(1→3)]-β-D-Galp-(1→}_n→

wherein n is10⁴-10⁶Suc succinic acid group, Pyr pyruvic acid group.

4. A process for the preparation of exopolysaccharides according to claim 3, characterized in that it comprises the following steps:

A. inoculating Paenibacillus mucilaginosus with the name of Latin and the preservation number of CGMCC No.23105 into an activation culture medium for activation;

B. b, inoculating the activated strain in the step A into a fermentation medium containing a carbon source, a nitrogen source and nutrient elements, and culturing under stirring to prepare a fermentation liquid;

C. and C, collecting exopolysaccharides from the fermentation liquor prepared in the step B.

5. The method for preparing extracellular polysaccharide according to claim 4, wherein the step B is to inoculate the activated strain obtained in the step A into a seed culture medium to prepare a seed solution after the seed solution is subjected to amplification culture, and the seed solution is inoculated into a fermentation culture medium according to an inoculation amount of 4-10% by volume to prepare a fermentation solution.

6. The method for preparing extracellular polysaccharide according to claim 4, wherein the activation in the step A is carried out by inoculating Paenibacillus mucilaginosus with a preservation number of CGMCC No.23105, which is named as Latin, into an activation medium, and activating for 24-48 hours at 28-40 ℃.

7. The method for preparing exopolysaccharide of claim 5, wherein the seed solution is prepared by terminating the culture of the seed solution when the OD value is not less than 1.0 at a wavelength of 550 nm.

8. The method for preparing exopolysaccharide according to claim 4, wherein the culture conditions in step B are a temperature of 28 ℃ to 40 ℃, a stirring speed of 100 to 300 rpm, and a culture time of 16 to 50 hours.

9. The method for preparing extracellular polysaccharide according to claim 4, 5 or 8, wherein the fermentation is terminated when the viscosity of the fermentation broth is not less than 500cp in the step B.

10. The method for preparing exopolysaccharide of claim 4, wherein ethanol solution with a concentration of 80-95% by volume is used in step C to settle exopolysaccharide in the fermentation broth, and the volume ratio of the ethanol solution to the fermentation broth is 2-4.

11. The method for preparing extracellular polysaccharide according to any one of claims 4-8, wherein the culture medium used in the step A, B comprises the following components:

20 g/L to 50 g/L of cane sugar; 1 g/L-2.5 g/L of potassium nitrate; 1 g/L-2 g/L of sodium dihydrogen phosphate; calcium chloride 0.01 g/L-1 g/L; magnesium sulfate heptahydrate 0.1 g/L-1 g/L; ferrous sulfate heptahydrate 0.01 g/L-0.05 g/L; 0.001 g/L-0.01 g/L of manganese sulfate monohydrate; 0.001 g/L-0.01 g/L of zinc chloride; the pH value is 6-8.

12. Use of exopolysaccharide according to claim 3 in the preparation of microbial flocculants.

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