Extracellular polymer sphingomonas and application thereof

Abstract

The invention relates to the technical field of oil field biochemistry, in particular to a strain of extracellular polymer sphingomonas sanxanigenens and application of the strain of extracellular polymer sphingomonas sanxanigenens and the application of the strain of extracellular polymer sphingomonas sanxanigenens and the application of the strain of extracellular polymer sphingomonas sanxanigenens. The preservation number of the strain of the extracellular polymer sphingomonas disclosed by the invention is CGMCC (China General Microbiological Culture Collection Center) No.30927. The biological flocculant prepared from the extracellular polymer sphingomonas can flocculate crude oil in crude oil-containing sewage and recover the crude oil from the crude oil-containing sewage, the crude oil recovery efficiency is high, the treated reinjection water is low in solid content and less in stratum pollution, and meanwhile, the biological flocculant has the advantages of high efficiency, simplicity, convenience and low cost.

Description

Technical Field

[0001] This invention relates to the field of oilfield biochemistry technology, specifically to an extracellular polymeric sphingosine monocytogenes strain and its applications. Background Technology

[0002] Sphingosomonas was isolated and identified by scientists in 1990, and it wasn't until 2001 that it was taxonomically divided into four genera based on physical characteristics, phylogenetic differences, and other features: *Sphingosomonas* (…). Sphingomonas ), Sphingosine Bacteria ( Sphingobium ), novel sphingosine bacteria ( Novosphingobium ), Coleoptera ( Sphigopyxis Sphingosomalidosis (Sphingosomalidosis) Sphingomonas Sphingosomalids are Gram-negative, rod-shaped, typically aerobic, chemoheterotrophic bacteria with a unique respiratory metabolic pathway involving coenzyme Q10. Their cell membranes primarily contain glycosphingolipids rather than lipopolysaccharides, and they typically secrete pigments that give their colonies a yellow color. Sphingosomalids are receiving increasing attention in the field of oil and gas microbiology. They also possess biosynthetic capabilities, secreting extracellular polymers such as gellan gum, welan gum, and rhamnose gum, which are widely used in the petroleum industry, primarily relying on the chemical and physical properties of polysaccharides, such as viscosity and gelation.

[0003] For example, CN107058187A discloses a strain of *Sphingomonas pluvialis* and a method for its application, providing a new strain of *Sphingomonas pluvialis*, FJAT-10625 (… Sphingomonas paucimobilis Furthermore, this strain yields a high amount of extracellular polysaccharides, thus increasing the source of strains for extracellular polysaccharide production. CN103421718B discloses a strain of *Sphingomonas sclerosus* and a method for its application. The invented *Sphingomonas sclerosus* strain is an endophytic bacterium isolated and screened from wild microporous grass seeds from the Qinghai-Tibet Plateau, and can be used for the industrial production of gellan gum, providing a new approach for the industrial production of gellan gum by endophytic bacteria. CN115838646A discloses a strain of *Sphingomonas sclerosus* SLDT21 and a method for its application, providing applications of *Sphingomonas sclerosus* SLDT21 in petroleum recovery, microbial enhanced oil recovery, and as a biopolysaccharide flooding agent.

[0004] In addition, the extracellular polysaccharides secreted by *Sphingomonas* can also be used as flocculants. For example, CN113151050B discloses *Sphingomonas* (… Sphingomonas sanxanigenensApplication of HL1 strain in the preparation of microbial flocculants. CN115636515A discloses a microbial oil recovery environmentally friendly wastewater treatment flocculant, which is produced by Sphingomonas sphingosine monophosphate with polysaccharides: arabinose, glucosamine hydrochloride, galactose, glucose, mannan, galacturonic acid and guluronic acid. These novel polysaccharides have good flocculation effects, but require compound polyferric chloride to meet the flocculation effect.

[0005] However, for the treatment of crude oil-containing wastewater that requires reinjection, and the recovery of crude oil from oily wastewater, technologies such as filtration, flotation, and membrane separation are currently used. For example, the Yushulin Oilfield uses a process of aeration + vortex flotation + sand filtration + PVC hollow fiber ultrafiltration membrane; the Changqing Oilfield uses gravity separation sedimentation oil removal + walnut shell coarse filter + fiber ball fine filtration; the Liaohe Oilfield uses a deep treatment system for heavy oil wastewater with ion exchange as the core; the Jinzhou Oilfield uses ozone-biocarbon technology; and the Gudao Oilfield uses flotation flocculation + biological contact oxidation. However, the composition of oily wastewater generated during oil extraction and processing is complex, and different processes also have certain limitations in actual operation, such as substandard reinjection water quality, low crude oil recovery rate which increases the difficulty of downstream treatment, weak shock resistance, frequent filter media replacement, harsh application environment of membrane modules, and high operating costs. Summary of the Invention

[0006] To overcome the problems of low crude oil recovery efficiency, high solids content, and formation pollution in existing crude oil wastewater treatment technologies, this invention provides an extracellular polymeric sphingosine monoclonal bacterium and its application. The bioflocculant prepared by this extracellular polymeric sphingosine monoclonal bacterium can flocculate crude oil in crude oil wastewater, recover crude oil from crude oil wastewater, achieve high crude oil recovery efficiency, and produce reinjection water with low solids content and minimal formation pollution.

[0007] To achieve the above objectives, the first aspect of the present invention provides an extracellular polymeric sphingosine monocytogenes strain (Sphingosine monocytogenes). Sphingomonas sanxanigenens The strain JD24001WB, whose preservation number is CGMCC No. 30927, is an extracellular polymeric sphingosine monocytogenes.

[0008] A second aspect of the present invention provides the application of the extracellular polymeric sphingosine monocytogenes described herein in the preparation of flocculants.

[0009] A third aspect of the present invention provides a method for preparing a bioflocculant, the method comprising: inoculating the extracellular polymeric sphingosine monocytogenes described in the present invention into a liquid culture medium for cultivation.

[0010] The fourth aspect of the present invention provides a bio-flocculator prepared by the preparation method described in the present invention.

[0011] The fifth aspect of this invention provides the application of the extracellular polymeric sphingosine monocytogenes and / or its fermentation broth or the bioflocculant described in this invention in flocculating oily wastewater.

[0012] The sixth aspect of the present invention provides a flocculation method for crude oil-containing wastewater, the method comprising: mixing the fermentation broth of a strain containing the extracellular polymeric sphingosine monocytogenes described in the present invention or the bioflocculant described in the present invention with the oil-containing wastewater.

[0013] Through the above technical solution, the extracellular polymeric sphingosine monocytogenes of the present invention has strong environmental adaptability and simple nutritional requirements. It can secrete a large amount of extracellular products in a high-sugar environment. The fermentation broth of this strain can be used to treat crude oil-containing wastewater, which can flocculate the crude oil in the wastewater and recover crude oil from the wastewater. It has the advantages of high efficiency, simplicity and low cost in recovering crude oil from crude oil-containing wastewater, high crude oil recovery efficiency, low solid content in the treated reinjection water, and less formation pollution.

[0014] Biological Preservation The extracellular polymeric sphingosine monocytogenes of the present invention (Sphingosine monocytogenes) Sphingomonas sanxanigenens Strain JD24001WB was deposited on June 11, 2024, at the China General Microbiological Culture Collection Center (Address: No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, Postcode: 100101) (abbreviation of depositary institution: CGMCC), with accession number CGMCC No. 30927. Attached Figure Description

[0015] Figure 1 This is a colony diagram of strain JD24001WB on the culture medium; Figure 2 These are Gram-stained micrographs of strain JD24001WB; Figure 3 This is a cell morphology diagram of strain JD24001WB; Figure 4 These are flocculation results for each group in Example 3; Figure 5 This is a bar chart of the flocculation rate of each group in Example 3. Detailed Implementation

[0016] The endpoints and any values ​​of the ranges disclosed herein are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of the various ranges, the endpoint values ​​of the various ranges and individual point values, and individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed herein.

[0017] The first aspect of this invention provides an extracellular polymeric sphingosine monocytogenes strain (Sphingosine monocytogenes) Sphingomonas sanxanigenens The strain JD24001WB is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 30927.

[0018] A second aspect of the present invention provides the application of the extracellular polymeric sphingosine monocytogenes described herein in the preparation of flocculants.

[0019] A third aspect of the present invention provides a method for preparing a bioflocculant, the method comprising: inoculating the extracellular polymeric sphingosine monocytogenes described in the present invention into a liquid culture medium for cultivation.

[0020] In this invention, the liquid culture medium can be a conventional culture medium capable of culturing extracellular polymeric sphingosine monocytogenes. Preferably, the nutrient medium contains 30-50 g / L glucose, 0.1-2.0 g / L yeast extract, 0.5-2.0 g / L sodium nitrate, 0.5-1.5 g / L dipotassium hydrogen phosphate, 0.2-1 g / L potassium dihydrogen phosphate, 0.1-0.6 g / L magnesium sulfate, 0.1-0.5 g / L calcium carbonate, 0.1-1 g / L potassium chloride, and a pH of 7.0-8.0. In the embodiments of this invention, a medium containing 50 g / L glucose, 2.0 g / L yeast extract, 2.0 g / L sodium nitrate, 1.5 g / L dipotassium hydrogen phosphate, 0.8 g / L potassium dihydrogen phosphate, 0.4 g / L magnesium sulfate, 0.1 g / L calcium carbonate, 0.5 g / L potassium chloride, and a pH of 7.0 is used as an example, but this does not limit the scope of the invention.

[0021] According to the present invention, the cultivation conditions may include: a temperature of 28-35°C and a time of 24-72 hours. In an embodiment of the present invention, cultivation at 30°C for 72 hours is used to exemplify the advantages of the invention, but this does not limit the scope of the invention.

[0022] According to the present invention, the extracellular polymeric sphingosine monocytogenes with accession number CGMCC No.30927 has the ability to secrete extracellular products and has the ability to flocculate various substances, including activated sludge, mud, soil solid suspension, bottom sediment, fly ash, activated carbon powder, silica gel powder, alumina, kaolin, cellulose powder, and colloidal substances. However, in a preferred embodiment, it has more prominent flocculation activity for oily wastewater. When used to treat oily wastewater, it can flocculate the crude oil in the oily wastewater and recover crude oil from the oily wastewater, thus treating the wastewater.

[0023] The fourth aspect of the present invention provides a bio-flocculator prepared by the preparation method described in the present invention.

[0024] The fifth aspect of this invention provides the application of the extracellular polymeric sphingosine monocytogenes and / or its fermentation broth or the bioflocculant described in this invention in flocculating oily wastewater.

[0025] According to the present invention, the fermentation broth of the strain can be provided by a culture of *Sphingomonas extracellular polymericus*, that is, the fermentation broth of *Sphingomonas extracellular polymericus* can be prepared according to the method described above. Specifically, *Sphingomonas extracellular polymericus* is inoculated into a liquid culture medium for cultivation. According to a preferred embodiment of the present invention, the cultivation conditions are as described above and will not be repeated here.

[0026] According to a preferred embodiment of the present invention, the strain uses glucose or xylose as a carbon source and inorganic nitrogen as a nitrogen source to ferment and produce the strain fermentation broth containing extracellular products.

[0027] The extracellular polymeric sphingomonas strain and / or its fermentation broth, or the bioflocculant described in this invention, can be used to treat crude oil-containing wastewater, thereby flocculating the crude oil in the wastewater and recovering the crude oil from the wastewater, thus treating the wastewater. A sixth aspect of this invention provides a method for flocculating oily wastewater, comprising: mixing the fermentation broth of a strain containing the extracellular polymeric sphingomonas strain described in this invention, or the bioflocculant described in this invention, with the oily wastewater.

[0028] In this invention, the mixing temperature can be selected from a wide range. This is an illustrative example, but it does not limit the scope of the invention. According to a preferred embodiment of the invention, the mixing temperature is 20-60°C.

[0029] In this invention, the pH range of the mixed solution is relatively wide, which is illustrative but does not limit the scope of the invention. According to a preferred embodiment of the invention, the pH of the mixed solution is 3-10.

[0030] According to a preferred embodiment of the present invention, the oily wastewater includes water, crude oil, and optionally solid substances.

[0031] In this invention, the oily wastewater contains light crude oil. According to a preferred embodiment of the invention, the crude oil contains 50-90 wt% alkanes and 10-30 wt% aromatics.

[0032] In this invention, the crude oil content in the oily wastewater can be selected from a wide range. This is an illustrative example, but it does not limit the scope of this invention. According to a preferred embodiment of this invention, the crude oil content in the oily wastewater is 50-200 mg / L, for example, it can be 60 mg / L, 80 mg / L, 100 mg / L, 120 mg / L, 150 mg / L, 180 mg / L, or any two of the above values.

[0033] In this invention, the solid matter in the oily wastewater is generally sand or gravel, and the mass content of the solid matter can be selected from a wide range. This is an illustrative example, but it does not limit the scope of this invention. According to a preferred embodiment of this invention, the mass content of solid matter in the oily wastewater is 0-100 mg / L, for example, it can be 10 mg / L, 20 mg / L, 40 mg / L, 50 mg / L, 60 mg / L, 80 mg / L, or any two of the above values.

[0034] In this invention, the sources of oily wastewater can be selected from a wide range. This is an illustrative example, but it does not limit the scope of the invention. According to a preferred embodiment of the invention, the oily wastewater is selected from the outlet wastewater of the three-phase separator of an oilfield wastewater treatment plant.

[0035] In the present invention specification and context, the following embodiments are included where the crude oil content is detected using ultraviolet light: extraction with petroleum ether is used, and measurement is performed on a UV spectrophotometer. Standard curves are prepared using standard oil solutions of different concentrations obtained by diluting with petroleum ether.

[0036] According to a preferred embodiment of the present invention, the method further includes solid-liquid separation of the flocculated system and extraction of the solid to obtain recovered crude oil.

[0037] According to one embodiment of the present invention, solid-liquid separation can be performed by means of filtration, centrifugation, etc.

[0038] According to one embodiment of the present invention, petroleum ether can be used to extract and recover crude oil from the separated solid; preferably, the boiling point of petroleum ether is 60-90℃.

[0039] According to one embodiment of the present invention, the flocculated sample is filtered and separated to obtain flocculents, which are then extracted with petroleum ether and purged with a gas (e.g., nitrogen) to obtain recovered crude oil.

[0040] Compared with the prior art, the bioflocculant prepared by the extracellular polymeric sphingosine monocytogenes of the present invention can flocculate the crude oil in the crude oil-containing wastewater when used to treat crude oil-containing wastewater. It can recover crude oil from crude oil-containing wastewater without the need to add additives. The additives refer to inorganic flocculants, such as polyferric chloride, polyaluminum chloride, polyaluminum sulfate, polyferric sulfate, etc.

[0041] To further understand the present invention, preferred embodiments of the present invention are described below in conjunction with examples. However, it should be understood that these descriptions are only for further illustrating the features and advantages of the present invention, and not for limiting the scope of the claims of the present invention.

[0042] In the context and embodiments of this invention, the crude oil-containing wastewater is taken from the outlet wastewater of the three-phase separator at the oilfield wastewater treatment plant.

[0043] In the context and embodiments of this invention, the formula for calculating the flocculation rate is as follows:

[0044] Where A represents the absorbance of the blank control oily wastewater at 550 nm, and B represents the absorbance of the flocculated sample system at 550 nm.

[0045] Example 1 This embodiment illustrates the results of morphological identification and molecular genetic classification of the extracellular polymeric sphingosine monoclonal bacteria described in this invention.

[0046] A bacterial strain was isolated from the produced fluid of well Wei5-27 in block Wei5 of Jiangsu Oilfield and deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 30927.

[0047] Extracellular polymeric sphingosine monocytogenes was cultured on solid LB agar plates (10g tryptone, 5g yeast extract, 10g sodium chloride, 15g agar, 1000mL deionized water, pH 7.2). After 2 days of culture, if... Figure 1 As shown, the colonies are yellow, with a smooth, moist, and sticky surface; they are droplet-shaped, round, raised, opaque, and have relatively neat edges; Gram staining is as follows. Figure 2 As shown, Gram staining is negative.

[0048] Colonies of the bacterial strain cultured on LB agar plates for 2 days were selected and sequentially fixed with glutaraldehyde, washed with phosphate buffer (pH 7.2), dehydrated with graded ethanol, treated with isoamyl acetate, and dried at the critical point of carbon dioxide. The samples were then sputter-coated with gold and placed in an observation chamber for scanning electron microscopy (SEM). The morphology was observed under the SEM. Figure 3 As shown, the extracellular polymeric sphingosine monocytogenes is rod-shaped, non-spore-forming, and non-flagellated, with a size of 0.53-0.70 μm × 1.06-1.67 μm.

[0049] Based on morphological observation, the 16S rRNA of the strain was sequenced, and the sequence is shown in SEQ ID NO:1. A similarity search was performed in the NCBI database, and it was identified as *Sphingosophora* genus *Extracellular Polymer Sphingosophora*. Sphingomonas sanxanigenens )microorganism.

[0050] Example 2 This embodiment illustrates the method for culturing the fermentation broth of the extracellular polymeric sphingosine monoclonal strain described in this invention.

[0051] (1) Activation of the strain The extracellular polymeric sphingosine monocytogenes strain JD24001WB was streaked onto an activation medium using an inoculation loop and cultured in a constant temperature incubator for 24-72 h at a temperature of 30±2℃. The activation medium could be YPD medium or LB medium.

[0052] YPD medium (by weight percentage): yeast extract 1%, peptone 2%, glucose 2%, agar powder 2%, prepared with distilled water, pH 7.2.

[0053] LB medium (by weight percentage): 2% glucose, 0.5% yeast extract, 1% peptone, 1% sodium chloride, 2% agar powder, prepared with deionized water, pH 7.2.

[0054] (2) Seed culture Using an inoculation loop, pick up a single well-grown colony from the activation plate and inoculate it into a 250mL Erlenmeyer flask containing 50mL of nutrient medium. Incubate at 30℃ and 200rpm on a shaker for 24 hours.

[0055] Nutrient culture medium: glucose 50 g / L, yeast extract 2.0 g / L, sodium nitrate 2.0 g / L, dipotassium hydrogen phosphate 1.5 g / L, potassium dihydrogen phosphate 0.8 g / L, magnesium sulfate 0.4 g / L, calcium carbonate 0.1 g / L, potassium chloride 0.5 g / L, pH 7.0.

[0056] (3) Preparation of fermentation broth The cultured seed solution was inoculated into a 250 mL Erlenmeyer flask containing 50 mL of nutrient medium at an inoculation rate of 2% (v / v), and then placed in a shaker at 30°C and 200 rpm for 72 h.

[0057] Example 3 This embodiment illustrates the flocculation effect of the extracellular polymeric sphingosine monocytogenes strain and its fermentation broth on oily wastewater.

[0058] The extracellular polymeric sphingosine monocytogenes and its fermentation broth prepared in Example 2 were used to separate the bacterial cells from the culture medium by centrifugation at a rotor speed of 10,000 r / min for 5 min. The supernatant was then collected to determine its flocculation activity. The precipitate was washed with sterile water and then resuspended in sterile water of the same volume as the supernatant to obtain a bacterial cell suspension.

[0059] The flocculation rate of crude oil-containing wastewater solutions was tested according to the blank group, HPAM (anionic polyacrylamide) group, fermentation broth group, supernatant group, and bacterial cell group. The crude oil-containing wastewater was taken from the outlet of the three-phase separator at the oilfield wastewater treatment plant, with a crude oil content of 175.03 mg / L and 77.67 mg / L of sand and gravel particles. The oil type was light oil, containing 85.83 wt% alkanes and 12.79 wt% aromatics. Specific implementation methods: According to the grouping, 1 mL of fermentation broth, 1 mL of supernatant, 1 mL of bacterial cell suspension, 1 mL of anionic polyacrylamide, and 1 mL of sterile water were added to each of the 5 groups of 49 mL crude oil-containing wastewater. The mixture was stirred at 250 rpm for 1 min and then at 100 rpm for 4 min. The temperature of the flocculation system was 30℃ and the pH was 8.0. After standing for 5 min, the absorbance before and after flocculation was measured at 550 nm.

[0060] The distribution of flocculation activity is as follows: Figure 4 As shown, the fermentation broth of the extracellular polymeric sphingosine monocytogenes strain exhibits excellent flocculation efficiency and can effectively emulsify crude oil, as well as effectively flocculate solid matter; the flocculation rate was calculated to obtain... Figure 5 The bar chart shown, combined with Figure 4 , Figure 5 It was found that after removing the bacterial cells, the flocculation rate of the supernatant was 93%, similar to that of the original fermentation broth (98.7%), and significantly better than that of the anionic polyacrylamide flocculant (76%). The flocculation rate of the bacterial cell suspension (20.1%) was significantly lower, indicating that the flocculation effect was achieved by the metabolic products produced and released extracellularly by the microorganisms during cultivation. Furthermore, this flocculation method does not require the addition of other flocculants, is highly efficient and environmentally friendly, and will not cause secondary pollution to the formation after reinjection.

[0061] The sample was filtered and separated after flocculation of the fermentation broth of the extracellular polymeric sphingosine monocytogenes strain. The flocculent was then extracted multiple times with equal amounts of petroleum ether (boiling point 60-90℃). After nitrogen blowing and volatilization, the recovered crude oil was obtained with a recovery rate of 96.3%.

[0062] Example 4 The wastewater containing crude oil had a crude oil content of 68.55 mg / L, a sand and gravel content of 44.32 mg / L, and the oil type was light oil. The alkane content was 63.06 wt% and the aromatic content was 27.64 wt%. 1 mL of the extracellular polymeric sphingosine monocytogenes fermentation broth prepared in Example 2 was added to 49 mL of the wastewater containing crude oil. The mixture was stirred at 250 rpm for 1 min and then at 100 rpm for 4 min. The flocculation system temperature was 30 °C and the pH was 8.0. After standing for 5 min, the absorbance before and after flocculation was measured at 550 nm.

[0063] The results showed that the flocculation rate of the fermentation broth of the extracellular polymeric sphingosine monocytogenes strain was 99.0%, and the crude oil recovery rate was 95.6%.

[0064] Example 5 The crude oil-containing wastewater contained 124.34 mg / L of crude oil and 66.87 mg / L of sand and gravel. The oil type was light oil, with an alkane content of 72.82 wt% and an aromatic content of 19.49 wt%. 1 mL of the extracellular polymeric sphingosine monocytogenes fermentation broth prepared in Example 2 was added to 49 mL of the crude oil-containing wastewater. The mixture was stirred at 250 rpm for 1 min and then at 100 rpm for 4 min. The flocculation system temperature was 30 °C and the pH was 8.0. After standing for 5 min, the absorbance before and after flocculation was measured at 550 nm.

[0065] The results showed that the flocculation rate of the fermentation broth of the extracellular polymeric sphingosine monocytogenes strain was 99.2%, and the crude oil recovery rate was 96.7%.

[0066] The preferred embodiments of the present invention have been described in detail above; however, the present invention is not limited thereto. Within the scope of the inventive concept, various simple modifications can be made to the technical solutions of the present invention, including combinations of various technical features in any other suitable manner. These simple modifications and combinations should also be considered as the content disclosed in the present invention and are all within the protection scope of the present invention.

Claims

1. A strain of extracellular polymeric sphingosine monocytogenes (Sphingosine monocytogenes) Sphingomonas sanxanigenens strain JD24001WB, characterized in that, The extracellular polymeric sphingosine monocytogenes strain has the accession number CGMCC No. 30927.

2. The application of the extracellular polymeric sphingosine monocytogenes according to claim 1 in the preparation of flocculants.

3. A method for preparing a bioflocculant, characterized in that, The method includes: inoculating the extracellular polymeric sphingosine monocytogenes of claim 1 into a liquid culture medium for cultivation.

4. The method according to claim 3, wherein, The culture medium contains: glucose 30-50 g / L, yeast extract 0.1-2.0 g / L, sodium nitrate 0.5-2.0 g / L, dipotassium hydrogen phosphate 0.5-1.5 g / L, potassium dihydrogen phosphate 0.2-1 g / L, magnesium sulfate 0.1-0.6 g / L, calcium carbonate 0.1-0.5 g / L, potassium chloride 0.1-1 g / L, and a pH of 7.0-8.0; and / or The cultivation conditions include: a temperature of 28-35℃ and a time of 24-72h.

5. The bio-flocculator prepared by the preparation method according to claim 3 or 4.

6. The application of the extracellular polymeric sphingomonas strain of claim 1 and / or its fermentation broth, and the bioflocculant of claim 5 in flocculating oily wastewater.

7. A flocculation method for wastewater containing crude oil, characterized in that, The method includes mixing the fermentation broth of a strain containing the extracellular polymeric sphingosine monocytogenes of claim 1 or the bioflocculant of claim 5 with oily wastewater.

8. The method according to claim 7, wherein, The mixing temperature is 20-60℃; and / or The pH of the mixed solution is 3-10.

9. The method according to claim 7 or 8, wherein, The oily wastewater includes: water, crude oil, and solid matter, preferably, the crude oil content is 50-200 mg / L; and / or the solid matter content is 0-100 mg / L; More preferably, the crude oil is light crude oil, wherein alkanes comprise 50-90 wt% and aromatics comprise 10-30 wt%.

10. The method according to any one of claims 7-9, wherein, The method also includes solid-liquid separation of the flocculated system and extraction of the solid to obtain recovered crude oil.

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