CN112195115A

CN112195115A - Brevibacillus borstelensis, preparation, method for producing surfactant and application

Info

Publication number: CN112195115A
Application number: CN202010814071.0A
Authority: CN
Inventors: 佘跃惠; 张凡; 张峙; 姚普勇; 李飞; 董浩; 孙姗姗; 喻高明; 易绍金; 张文达; 胡琳琪; 冯阳阳; 郑安应; 李杨
Original assignee: Yangtze University
Current assignee: Yangtze University
Priority date: 2020-08-13
Filing date: 2020-08-13
Publication date: 2021-01-08
Anticipated expiration: 2040-08-13
Also published as: CN112195115B; NL2028866B1; NL2028866A

Abstract

The invention relates to the technical field of application of bioengineering technology in microbial oil recovery, and discloses a Brevibacillus borstelensis, a preparation, a method for producing a surfactant and application. The Brevibacillus borstelensis and the preparation thereof can effectively improve the recovery ratio of crude oil; the method for producing the surfactant can ensure that the lipopeptide biosurfactant has good physical properties, effectively reduces the surface tension and has good emulsifying property on petroleum and various hydrocarbons and lipids.

Description

Brevibacillus borstelensis, preparation, method for producing surfactant and application

Technical Field

The invention relates to the technical field of application of bioengineering technology in microbial oil recovery, in particular to Brevibacillus borstelensis, a preparation, a method for producing a surfactant and application.

Background

As the difficulty of increasing the yield of the oil field is increased continuously in the middle and later stages of the development, people are seeking various economic, effective and environment-friendly new technologies to improve the crude oil recovery rate continuously. Microbial oil recovery is receiving more and more attention because of the advantages of wide application range, simple process, low cost, environmental protection and the like, and generally, microbial oil recovery refers to injecting proper strains, nutrients or fermentation liquor into an oil reservoir to ensure that the strains propagate and metabolize in the oil reservoir to generate gas or active substances and reduce the oil-water interfacial tension so as to improve the crude oil recovery ratio.

Over the past decades, many patents have reported that many different species of microorganisms use different carbon sources to metabolize to produce surfactants and their use in the petroleum industry. The microorganisms capable of producing surfactants are of various types, and most studied are mainly Pseudomonas, Bacillus, Acinetobacter, Rhodococcus, Candida, and the like. Biosurfactants are also generally divided into two main classes, one class is a biosurfactant with larger molecular weight, and the biosurfactant is usually amphiphilic polysaccharide, protein and the like, and the most researched biosurfactants are Emulsan and Alasan produced by Acinetobacter; a class of biosurfactants are micromolecular biosurfactants which are mainly divided into two classes of glycolipids (mainly producing strains comprise Pseudomonas, Rhodococcus, Candida, Burkholderia and the like) and lipopeptides (mainly producing strains comprise Bacillus), because of the unique environment of an oil reservoir, a large variety of hydrocarbon degrading bacteria and surfactant producing bacteria are contained in the oil reservoir, and usually, a single bacterium separated from the oil reservoir can be more suitable for the extreme environment of the oil reservoir, the microorganism is the key point of microbial oil extraction research, surfactant producing microorganisms are screened and separated from the oil reservoir, or the proportion of the surfactant producing microorganisms in the oil reservoir is regulated by injecting microbial inoculum and nutrition obtained by culture, and the biosurfactant producing microorganisms are one of the main contents of the microbial oil extraction research.

In the process of microbial oil recovery, different oil reservoirs have certain requirements on strains, a large number of strains for producing biosurfactants are reported at present, and a large number of microorganisms for producing the biosurfactants are also reported from the oil reservoirs, but documents and patents are rarely reported for producing the surfactants by Brevibacillus borstelensis (Brevibacillus borstelensis) in the oil reservoirs. The prior art has some relevant patents of the short bacillus borsteinm, and the application of the short bacillus borsteinm in brewing, pharmacy, food, animal husbandry, organic pollutant degradation, food waste treatment and oil exploitation is respectively reported. However, there is no report on the metabolic production of biosurfactants by Brevibacillus borstelensis.

Based on the technical blank in the prior art, the invention provides a Brevibacillus borstelensis, a preparation, a method for generating a surfactant and application.

Disclosure of Invention

The invention provides a Brevibacillus borstelensis, a preparation, a method for producing a surfactant and application.

The invention adopts the following technical scheme:

a Brevibacillus borstelensis (Brevibacillus borstelensis) strain is named as YZ-2, and the preservation number is as follows: CGMCC No. 9981.

In one aspect, the invention provides a borstem brevibacillus preparation, which contains the following preservation numbers: the short bacillus borsteinq with CGMCC No.9981 is a solid or liquid bacterial preparation.

In another aspect, the invention provides a method of preparing a lipopeptide biosurfactant, the method comprising fermentatively culturing a Brevibacillus borstelensis or a Brevibacillus borstelensis preparation in a nutrient medium to produce the lipopeptide biosurfactant.

Further, the nutrient medium is LB medium.

Further, the nutrient medium comprises: 10-50g/L of sucrose, 10-40g/L of corn steep liquor dry powder, 3-20g/L of peptone and MgSO₄0.1-2g/L，KCl 2-18g/L，MnSO₄5-9 g/L，CuSO₄5-10 g/L，ZnSO₄5-12 g/L， KH₂PO₄1-7g/L, and the conditions for producing the lipopeptide biosurfactant are as follows: the pH value of the mixture is 5-9.5, and the fermentation temperature is 20-60 ℃.

Further, the method may further comprise the step of removing the bacterial cells after the fermentation.

Further, the fermentative culture of Brevibacillus borstelensis in a nutrient medium comprises:

step 1.1: the slant preserved strain is activated by inoculating loop on a flat plate and streaked, and is cultured for 20h at the constant temperature of 30 ℃, then tricyclic strain is picked from the flat plate and inoculated into a first-class seed shake flask, and is cultured for 18h at the temperature of 30 ℃ and at the speed of 180 r/min;

step 1.2: 2% of the volume of the first-stage seed shake flask is taken to be inoculated into a second-stage shake flask, and the second-stage seed shake flask is cultured for 18h under the same condition as the step 1.1;

step 1.3: 2% of the volume in the secondary flask was transferred to the fermentation flask and cultured under the same conditions as in step 1.1 until the spore yield reached 100%.

On the other hand, the invention provides the application of the lipopeptide biosurfactant in oil displacement.

In yet another aspect, the invention provides a method for preparing a lipopeptide biosurfactant, wherein the lipopeptide biosurfactant is produced by the metabolism of Brevibacillus borstelensis.

In another aspect, the invention provides the use of Brevibacillus borstelensis for enhanced oil recovery.

In yet another aspect, the invention provides the use of a Brevibacillus borstelensis preparation in the enhanced oil recovery.

Compared with the prior art, the invention has the following advantages:

1. according to the method for preparing the lipopeptide biosurfactant, the lipopeptide biosurfactant has good physical properties, can effectively reduce the surface tension, and has good emulsifying performance on petroleum, various hydrocarbons and lipids;

2. the Brevibacillus borstelensis preparation can effectively improve the recovery ratio of crude oil.

Deposit description

And (4) storage address: the institute of microbiology, China academy of sciences, No. 3, Xilu No. 1, Beijing, Chaoyang, China;

the preservation date is as follows: 11 month and 18 days 2014;

the strain name is as follows: brevibacillus borstelensis;

latin name: brevibacillus borstelensis;

the preservation organization: china general microbiological culture Collection center;

the preservation organization is abbreviated as: CGMCC;

the preservation number is: CGMCC No. 9981.

Drawings

FIG. 1 is a schematic diagram of an oil drain circle of a fermentation broth of strain YZ-2 in example 1 of the present invention;

FIG. 2 is a tree of evolutionary trees of the 16S rRNA-based surfactant-producing strain YZ-2 of example 1 of the present invention;

FIG. 3 is an infrared spectrum of a crude product of a YZ-2 bacteria metabolite in example 2 of the present invention;

FIG. 4-1 is a TIC spectrum of part 1 of a metabolite of the YZ-2 strain in example 2 of the present invention;

FIG. 4-2 is a TIC spectrum of part 2 of the metabolite of the YZ-2 strain in example 2 of the present invention;

FIG. 5-1 is a schematic diagram of mass spectrometry of a metabolite part 1 of strain YZ-2 in example 2 of the present invention;

FIG. 5-2 is a schematic illustration of mass spectrometry analysis of local 2 of metabolites of strain YZ-2 in example 2 of the present invention;

FIG. 6 is a flow chart of a flooding simulation test in embodiment 4 of the present invention;

FIG. 7 is a schematic representation of YZ-2 surfactant vs. enhanced oil recovery efficiency in example 4 of the present invention.

Detailed Description

The present invention will be further described with reference to the following specific examples, which are not intended to limit the present invention, but are merely illustrative of the technical effects of Brevibacillus borstelensis YZ-2, the characteristics and the applications of the Brevibacillus borstelensis of the present invention. The experimental methods used in the following examples are not specifically described, and the materials, reagents and the like used in the following examples are generally commercially available under the usual conditions without specific descriptions.

The invention provides a Brevibacillus borstelensis strain separated from an oil reservoir environment, research data show that the strain has good oil recovery characteristics, and a physical simulation oil displacement experiment shows that a biological preparation of the Brevibacillus borstelensis strain can greatly improve the crude oil recovery rate in a rock core. The invention provides a method for producing lipopeptide biosurfactant by fermenting Brevibacillus borstelensis and a process for extracting a metabolite of the Brevibacillus borstelensis, and provides a preparation of the Brevibacillus borstelensis and the application of the preparation in improving the crude oil recovery ratio.

Example 1 isolation, identification and preservation of surfactant strains

1. Isolation of surfactant-producing strains

According to a conventional strain screening method, 10mL of water sample collected from an oil field is taken and inoculated into a sterilized crude oil culture medium (2% crude oil, v: v) containing 100mL, the constant temperature shaking culture is carried out for 72h at 35 ℃ and 150rpm, an experimental group with high crude oil emulsifying and dispersing degree is selected, 100 mu L of fermentation liquor is taken and coated on an LB agar plate culture medium, and the culture is carried out for 48h at 35 ℃; picking single colonies with different forms, streaking and purifying on an LB agar plate culture medium, and culturing at 35 ℃ for 48 h. Selecting single colony, enriching culture, inoculating into original inorganic salt culture medium with crude oil as carbon source, culturing at 35 deg.C and 150rpm for 72 hr, observing crude oil emulsifying and dispersing degree, measuring surface tension of fermentation liquid, and selecting strain with maximum surface tension reduction.

In this example, it was found that microorganisms capable of emulsifying and dispersing crude oil and reducing the surface tension of the culture solution were present in the sample by the preliminary screening. After the cultivation and domestication, 1 strain producing the biosurfactant is separated and named as YZ-2, as shown in figure 1, the diameter of an oil discharge ring of the YZ-2 is 7.21cm, the surface tension of fermentation liquor is reduced to 30.103mN/m, as shown in table 1, the metabolites of the YZ-2 have excellent surface activity, and the biosurfactant is applied to microorganisms to improve the crude oil recovery ratio.

TABLE 1 identification of surfactant-producing microorganisms

2. Strain identification and preservation

According to the manual of identifying common bacteria systems, the strain YZ-2 separated by the invention is identified, the genus is mainly identified from individual morphological characteristics, colony characteristics, staining reaction, physiological and biochemical reactions and the like of the strain, the contents comprise morphological observation, gram staining, hydrogen peroxide catalase reaction, oxidase experiment, glucose oxidation fermentation experiment and methyl red experiment, and the identification result is shown in Table 2.

TABLE 2 physiological and biochemical characterization of Strain YZ-2

In addition, according to a conventional strain identification method, genome DNA of the strain YZ-2 is respectively extracted, primers are designed for PCR amplification, amplification products are detected by agarose gel electrophoresis, and ideal PCR products are selected and sent to Shanghai worker sequencing for DNA sequencing. PCR amplification product sequencing results for YZ-2 NCBI submitted for BLAST search and homology comparison and the phylogenetic tree was constructed using Clustalx1.83, Mega, etc. software, as shown in FIG. 2, the strain YZ-2 was determined to be Brevibacillus borstelensis (similarity of Brevibacillus borstelensis strain 1CK49 is 99%) by 16S rDNA sequence analysis (corresponding representative species in Genbank see Table 3 below).

Table 3 alignment of 16S rRNA sequences of YZ-2 with Genbank database sequences

The strain YZ-2 of the invention can be preserved by the following method:

(1) short-term preservation: the strain is streaked on a slant culture medium, cultured for 48 hours at 35 ℃, and then preserved at 4 ℃.

(2) And (3) long-term storage: adopting a glycerol cryopreservation method: several loopful of bacteria were scraped from a fresh slant medium and transferred to a glycerin tube containing 1.5mL of 30% sterilized glycerin and stored at-80 ℃ by freezing. Or adopting a skim milk freezing preservation method: scraping 2-ring bacteria on fresh slant culture medium, transferring into glycerol tube filled with sterilized skimmed milk, and freezing at-80 deg.C for storage.

The Brevibacillus borstelensis YZ-2 screened in this example was deposited in the general microbiological culture Collection center of China Committee for culture Collection at 11 months and 18 days 2014 (address: Beijing, Naja-Kogyo No. 1 Hozeh No. 3, institute of microbiology, China academy of sciences), with the following deposition numbers: CGMCC No. 9981.

Example 2 extraction and analysis of YZ-2 metabolites

1. Fermentation of YZ-2

Brevibacillus borstelensis (Brevibacillus borstelensis) YZ-2 provided in this example was cultured in a fermentation medium. Wherein the fermentation medium comprises the following components in percentage by weight (g/L): 10-50g/L of sucrose, 10-40g/L of corn steep liquor dry powder, 3-20g/L of peptone and MgSO₄1-5 g/L，KCl 2-18g/L，MnSO₄5-9 g/L，CuSO₄5-10 g/L，ZnSO₄5-12 g/L，KH₂PO₄1-7g/L, pH 6-8g/L, sterilizing at 115 deg.C for 30 min.

The slant preserved strain is activated by inoculating loop on a flat plate through lineation, the strain is cultured for 20 hours at the constant temperature of 30 ℃, then, a strain with three loops (each inoculating loop strain contains more than 3 single colonies with obvious characteristics) is selected from the flat plate, inoculated into a first-stage seed shake flask (250mL triangular flask with the liquid loading of 50mL), cultured for 18 hours at the temperature of 30 ℃ and 180r/min, 2 percent of the volume is inoculated into a second-stage shake flask (250mL conical flask with the liquid loading of l00 mL), and cultured for 18 hours under the same conditions. The spore content is 2% of the total volume of the culture medium, the culture medium is inoculated to a fermentation shake flask (500mL conical flask, liquid containing 150mL), and the culture medium is cultured under the same condition until the spore content reaches 100%.

2. Extraction and identification of YZ-2 lipopeptide

10L of the fermentation broth were centrifuged at 8000r/min for 10 minutes at room temperature to remove the precipitate, the pH was then adjusted to 2.0 with 6mol/mL hydrochloric acid and the mixture was stored at 4 ℃ for 24 hours. Centrifuging the acidified solution at room temperature at 8000r/min for 10 min, removing supernatant, collecting precipitate, leaching with methanol for more than 3 times, centrifuging the methanol extract under the same condition, rotary evaporating and concentrating the supernatant (45 deg.C), dissolving with distilled water, adjusting pH to 7.0, vacuum filtering, adjusting pH to 2.0, standing at 4 deg.C overnight, centrifuging, collecting precipitate, leaching with methanol equal to the fermentation broth for 3 times, centrifuging, collecting supernatant, concentrating under reduced pressure with rotary evaporator, oven drying at 60 deg.C until constant weight is reached, measuring weight to obtain yellowish brown solid powder, grinding the obtained yellowish brown solid powder and potassium bromide in a mortar, tabletting, performing infrared spectroscopic analysis, as shown in FIG. 3, showing that there is typical cyclic lactam bond in lipopeptide, the nitrogen hydrogen bond of amino acid, deducing that the strain YZ-2 metabolizes to produce lipopeptide biosurfactant.

3. Mass spectrometric analysis of YZ-2 metabolites

Chromatographic conditions are as follows: mobile phase: 0.1% aqueous acetic acid + acetonitrile; flow rate: 0.2 ml/min; column temperature: 35 ℃; sample introduction volume: 5 mu L of the solution; a chromatographic column: WatersAcquity UPLC C181.7um 2.1X 100 mm.

Mass spectrum conditions: an ion source: ESI (+); capillary voltage: 3.0 KV; taper hole voltage: 40V; the temperature of the desolvation: 300 ℃; an ion source: 150 ℃; atomizing: 650L/h; an acquisition mode: ms1 Scan.

Liquid chromatography separation as shown in FIGS. 4-1 and 4-2, analysis of metabolites of strain YZ-2 showed 8 peaks (1.101, 6.173,6.814,7.249,7.602,7.936,8.354min, respectively) in time periods, and as shown in FIG. 5-2, analysis of molecular weight 965.96-1078.48 showed 8 fragment peaks, main peaks 993.95, 994.99, indicating surfactant and eformin, and there were several homologs containing short chain lipopeptide. In addition, there are also 1463.99-1702.25 four small peaks, indicating the presence of the interleukin lipopeptide.

Analysis of the YZ-2 bacteria metabolites showed peaks in 8 time periods (1.101, 6.173,6.814,7.249,7.602,7.936,8.354min, respectively). As shown in FIG. 5-1, the molecular weight of 979.92-1052.22 has 9 fragment peaks, main peaks 1008.06, 1022.06, 1036.15 and 1037.14, which indicate that there are many homologs of mainly surfactant and efonin.

Example 3 optimization of YZ-2 lipopeptide-producing fermentation System

Seed bacterial liquid: inoculating a seed solution into an enrichment medium, wherein the basic composition of the seed solution comprises: yeast 5g/L, NaCl 10g/L and Peptone 10 g/L. The cells were cultured to a concentration of 108 cells/mL and 109cells/mL as seed solutions for experimental studies of this example.

1. Carbon source optimization

Preparing a culture medium: weighing 1000mL of distilled water, and weighing corn steep liquor10-40g/L of dry powder and MgSO₄1-5 g/L， KCl 2-18g/L，MnSO₄5-9 g/L，CuSO₄5-10 g/L，ZnSO₄5-12 g/L，KH₂PO₄1-7g/L, and adjusting the pH value to 6-8. The carbon sources are respectively added into 100mL portions in 8 conical flasks with 250mL portions, and the following eight carbon sources are respectively added into the conical flasks: sucrose (No. 1), glucose (No. 2), corn steep liquor dry powder (No. 3), starch (No. 4), olive oil (No. 5), lactose (No. 6), glycerol (No. 7), liquid paraffin (No. 8) (content: 2%), after sterilization, 5% seed solution was inoculated, cultured at 45 ℃ or below, shaking cultured at 150rpm for 72 hours, the fermentation broth was diluted to various gradients and the surface tension of the fermentation broth (unit: mN/m) was measured, the results were as shown in table 4:

TABLE 4 Effect of different carbon sources on the fermentative production of lipopeptides by the Strain YZ-2

The measured surface tension data was observed, and the surface tension was 38.2 mN/m after adding carbon source medium bacterial liquid No. 1 and diluting 20 times, the dry weight of the cells was 1.39g/L, the surface tension was 37.1mN/m after adding carbon source medium bacterial liquid No. 3 and diluting 20 times, and the dry weight of the cells was 0.89 g/L. And (3) integrating the dry weight of the thalli and the reduction condition of the surface tension of the fermentation liquor, and selecting sucrose as an optimal carbon source for fermentation. And then, optimizing the adding amount of sucrose, collecting a product by adopting an acid precipitation method, drying and weighing, simultaneously collecting thalli by ultracentrifugation, and drying to dry weight, and finding that when the adding amount of the sucrose is 1-4g/L, the output of the surfactant reaches the highest value, so that 1-4% of the sucrose is selected as the optimal carbon source for producing lipopeptide by using the strain YZ-2.

2. Nitrogen source optimization

Preparing a culture medium: weighing 1000mL of distilled water, weighing 10-40g/L of sucrose and MgSO₄1-5 g/L，KCl 2-18 g/L，MnSO₄5-9 g/L，CuSO₄5-10 g/L，ZnSO₄5-12 g/L，KH₂PO₄1-7 g/L. After being stirred uniformly, the mixture is divided into 100mL portions in 6 conical flasks with the volume of 250 mL. 6 different nitrogen sources were prepared according to the following table and added to the medium (content 0.5%): ammonium chloride (No. 1), ammonium nitrate (No. 1)2) The peptone (number 3), the corn steep liquor dry powder (number 4), the sodium glutamate (number 5) and the sodium nitrate (number 6) are inoculated into 5% seed liquid after sterilization, cultured at the temperature below 45 ℃, subjected to shaking culture at 150rpm for 72 hours, and the fermentation liquid is diluted to different gradients and the surface tension of the fermentation liquid is measured, so that the results are shown in table 5:

TABLE 5 Effect of different carbon sources on the fermentative production of lipopeptides by Strain YZ-2

The research result shows that when the inorganic nitrogen source NaNO is added₃Ammonium chloride (number 1) and ammonium nitrate (number 2), the surface tension of the fermentation liquor is not obviously reduced after three days of culture, and the surface tension of the fermentation liquor is reduced to about 30mN/m by adopting organic nitrogen source fermentation.

Example 4 oil displacement simulation test

1. Preparation of oil displacement bacteria liquid

150ml of a surfactant-containing bacterial fermentation medium and 3L of an inorganic salt medium were prepared. Sterilizing with high pressure steam at 121 deg.C for 20 min. Activating a YZ-2 strain, transferring the activated strain to a fermentation culture medium, carrying out shake culture at 40 ℃ and 150r/min, adding 10% of fermentation liquor into 3L of oil displacement culture medium after 24h, carrying out shake culture at 40 ℃ and 150r/min, taking out after two days, and putting into a refrigerator at 4 ℃ for later use.

2. Core displacement of reservoir oil model test

As shown in fig. 6, an oil displacement process (in fig. 6, 1-distilled water container, 2-advection pump, 3-six-way valve, 4-intermediate container, 5-pressure gauge, 6-core holder, 7-core, 8-confining pressure pump, 9-measuring cylinder metering) is connected, the oil displacement process is composed of a micro-magnitude fast pump (gas cylinder for pressure source of constant pressure displacement), a high-pressure container, a core model and an oil-water separation metering tube, the four parts are connected by pipelines and valves, the pressure gauge is installed according to requirements to display pressure value, and the part needing to keep experimental temperature is installed in a thermostat.

Selecting two artificial rock cores D21 and D22 with gas permeability of about 300md, injecting YZ-2 bacterial liquid 2PV, closing the well for 2 days, and respectively measuring the oil displacement effect according to the following steps.

(1) Saturated water of rock core

Weighing dry weight of an artificial core with gas permeability of 300md, putting the artificial core into a wide-mouth bottle, evacuating for three hours by using a vacuum pump, adding simulated formation water to completely immerse the core into water, starting the vacuum pump until the reading of a vacuum meter is zero, and taking out the artificial core and weighing wet weight to obtain the pore volume of the core.

(2) Saturated oil

The rock core is loaded into a rock core holder, the rock core holder is placed into a thermostat at 40 ℃, dehydrated crude oil is added into a steel container, a pipeline is connected, a advection pump is started to press the crude oil into the rock core, when 3-4 mL of crude oil flows out of the outlet end of the rock core holder, saturated oil is finished, and the saturation of the crude oil is obtained according to the volume of the drained water.

(3) Water drive

And (3) filling simulated formation water into a steel container, connecting an oil displacement pipeline, starting a constant flow pump to displace oil, connecting crude oil to the outlet end of the rock core holder by using a 5mL test tube, and counting one time every 10 minutes until the water content reaches 95%.

(4) Microbial flooding

Filling the oil displacement bacterial liquid into a steel container, connecting a pipeline to displace oil, recording the amount of the oil displaced, closing the advection pump after reaching the specified PV number, and closing the well at 40 ℃ for 2 days according to a plan.

(5) Subsequent water drive

And after the specified well closing time, performing subsequent water drive on the rock core, recording the driven crude oil amount, and calculating the value for improving the crude oil recovery rate. Cores used to produce surfactant bacteria YZ-2 were D21 and D22, and the results of physical property measurements on two homogeneous synthetic cores are shown in Table 6. And (3) after the two cores are saturated with oil, injecting a product surfactant bacterial liquid when the water content is 95%, closing the well for 2 days, and then performing subsequent water flooding to obtain the recovery ratio, wherein the results are shown in a table 6.

Table 6 measurement results of physical properties of homogeneous core

From table 6, it is seen that the physical property measurements of D21 and D22 show that the physical parameters of the two cores are approximately the same, the gas permeability is about 300md, the void volume is about 14%, and the oil saturation is about 83%. From the YZ-2 oil displacement experimental data shown in FIG. 7, for two groups of cores D21 and D22, the oil recovery rate is improved by 5.6% and 11.69% after the body is displaced by 120mg/LYZ-2 surfactant solution, and the oil displacement effect is obvious.

The present invention is not limited to the above-described embodiments, which are described in the specification and illustrated only for illustrating the principle of the present invention, but various changes and modifications may be made within the scope of the present invention as claimed without departing from the spirit and scope of the present invention. The scope of the invention is defined by the appended claims.

Claims

1. A Brevibacillus borstelensis (Brevibacillus borstelensis) with a preservation number of: CGMCC No. 9981.

2. A Brevibacillus borstelensis preparation is characterized in that the Brevibacillus borstelensis preparation contains the following preservation numbers: the short bacillus borsteinq with CGMCC No.9981 is a solid or liquid bacterial preparation.

3. A method for preparing a lipopeptide biosurfactant, which comprises fermenting and culturing a Brevibacillus borstelensis or Brevibacillus borstelensis preparation in a nutrient medium to produce the lipopeptide biosurfactant.

4. The method of claim 3, wherein the nutrient medium is LB medium.

5. The method of claim 3, wherein the nutrient medium comprises: 10-50g/L of sucrose, 10-40g/L of corn steep liquor dry powder, 3-20g/L of peptone and MgSO₄ 0.1-2g/L，KCl 2-18g/L，MnSO₄ 5-9g/L，CuSO₄ 5-10g/L，ZnSO₄ 5-12g/L，KH₂PO₄1-7g/L, and the conditions for producing the lipopeptide biosurfactant are as follows: the pH value of the mixture is 5-9.5, and the fermentation temperature is 20-60 ℃.

6. The method for producing a lipopeptide biosurfactant according to claim 3 further comprising the step of removing the biomass after fermentation.

7. The use of the lipopeptide biosurfactant of any one of claims 3-6 in oil displacement.

8. A method for preparing a lipopeptide biosurfactant is characterized in that Brevibacillus borstelensis metabolizes to generate the lipopeptide biosurfactant.

9. Use of Bacillus borstelensis as claimed in claim 1 for enhanced oil recovery.

10. Use of a brevibacillus borstelensis preparation according to claim 2 for enhanced oil recovery.