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Bacillus subtilis for anaerobic production of lipopeptid surfactant and application of bacillus subtilis

A technology of Bacillus subtilis and surfactants, applied in the direction of bacteria, drilling compositions, microbial-based methods, etc., can solve problems such as weakening oil displacement efficiency, increasing production costs, and affecting the metabolic activity of surfactant-producing bacteria , to achieve the effects of saving crude oil production costs, improving crude oil recovery, and strong crude oil emulsification activity

Active Publication Date: 2017-05-31
SHENYANG INST OF APPL ECOLOGY CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the anoxic environment of the oil reservoir affects the metabolic activity of the surfactant-producing bacteria, thus weakening its oil displacement efficiency; air injection in the oil reservoir will increase the production cost

Method used

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  • Bacillus subtilis for anaerobic production of lipopeptid surfactant and application of bacillus subtilis
  • Bacillus subtilis for anaerobic production of lipopeptid surfactant and application of bacillus subtilis
  • Bacillus subtilis for anaerobic production of lipopeptid surfactant and application of bacillus subtilis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Example 1: Screening and identification of bacterial strain AnPL-1

[0035] The isolation and screening process of strain AnPL-1 is as follows:

[0036] Take 10ml of oil-water samples from an oilfield in China, add them to an anaerobic bottle containing 190ml enrichment medium, and carry out anaerobic enrichment culture at 37°C and 80rpm for 15 days.

[0037] Take 1ml of the anaerobic enrichment solution for gradient dilution with sterile distilled water, dilute to 10 4 times, 10 5 times, 10 6 times, 10 7 Double the volume, take 0.1ml samples of different dilution factors and spread them on the LB solid medium plate containing 5% (v / v) sterile sheep blood, and culture the plate upside down at 37°C. Pick the colony with a large hemolytic circle and streak it on a new LB solid medium plate containing 5% sterile sheep blood for separation and purification. Streak the purified single colony on the LB solid medium slant for temporary bacterial colony kind of preservatio...

Embodiment 2

[0043] Embodiment 2: Extraction and identification of the surfactant product of bacterial strain AnPL-1 under anaerobic conditions

[0044] Strain AnPL-1 is a facultative anaerobic bacterium capable of producing biosurfactants under both aerobic and anaerobic conditions. Strain AnPL-1 was cultured in LB medium at 37°C and 160rpm for 24h. Take the LB culture solution and centrifuge, discard the supernatant, then wash with an equal volume of NaCl solution with a mass concentration of 0.9% and resuspend the bacterial pellet to make a seed solution for inoculation; inoculate to 200ml fermenter with a volume ratio of 1:20 In the anaerobic bottle of the culture medium, the anaerobic culture was carried out at 37° C. and 80 rpm for 10 days.

[0045] Take the above-mentioned anaerobic fermentation broth, centrifuge to remove bacteria, adjust the pH value of the supernatant to 2.0 with 6mol / L HCl solution, and place it overnight at 4°C; then use an equal volume of chloroform / methanol ...

Embodiment 3

[0050] Embodiment 3: the environmental condition evaluation of the anaerobic efficient production lipopeptide surfactant of bacterial strain AnPL-1

[0051] The ability of Bacillus subtilis AnPL-1 to anaerobically produce lipopeptide surfactant under different temperature, pH and salinity conditions was investigated using an anaerobic tube equipped with 20ml fermentation medium. (v / v) inoculum amount was inoculated, and anaerobic culture was carried out at 37° C. for 8 days. The ability of strain AnPL-1 to produce lipopeptide anaerobic surfactant is characterized by the surface tension of the anaerobic fermentation broth and the diameter of the oil drainage circle. AnPL-1 is capable of efficiently producing lipopeptide surfactants.

[0052] Temperature experiment treatment group: 4 temperature experiment groups were set at 30°C, 35°C, 40°C and 45°C (initial pH value was 7.0, NaCl mass concentration was 0.1%), and three parallel experiments were set up in each group. In the t...

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Abstract

The invention relates to bacillus subtilis AnPL-1 capable of generating a lipopeptid surfactant under an anaerobic condition, with a preservation number of CGMCC NO. 13461. The strain AnPL-1 is a facultative anaerobe; a temperature range for efficient production of lipopeptid under the anaerobic condition is 30-40 DEG C; a pH (potential of hydrogen) value range is 6.0-7.5; a tolerance range of NaCl is 0-5%. An emulsion coefficient of an anaerobic fermentation solution of the strain for crude oil is equal to or greater than 56%; the diameter of an oil drainage circle is equal to or greater than 15mm; the lipopeptid surfactant produced by the strain AnPL-1 in situ in a physical simulation rock core increases a crude oil recovery factor by 9.9%. The strain has important popularization and application values in the field of microbial oil recovery and environmental pollution remediation.

Description

technical field [0001] The invention belongs to the field of environmental biotechnology and microbial oil recovery, in particular, it relates to an anaerobic lipopeptide biosurfactant Bacillus subtilis (Bacillus subtilis) AnPL-1 and its application in the field of microbial oil recovery and environmental pollution restoration application. Background technique [0002] Biosurfactants are surface-active metabolites synthesized and secreted by microorganisms, mainly including lipopeptides, glycolipids, and phospholipids. Compared with chemically synthesized surfactants, biosurfactants have the advantages of high surface activity, safety, non-toxicity, and biodegradability, so they are widely used in environmental pollution remediation, oil exploration, food, medicine and other fields. Among the biosurfactants studied, the lipopeptide surfactants had a lower critical micelle concentration and higher surface activity. [0003] In the remediation of petroleum hydrocarbon pollut...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P21/00C09K8/582A62D3/02C12R1/125A62D101/20
CPCA62D3/02A62D2101/20C09K8/582C12P21/00C12N1/205C12R2001/125
Inventor 赵峰张颖史荣久韩斯琴
Owner SHENYANG INST OF APPL ECOLOGY CHINESE ACAD OF SCI
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