Biological oil and sulfur removal composite method and crude oil associated gas biological oil and sulfur removal method

Abstract

The invention relates to the technical field of environmental microorganisms, in particular to a biological oil and sulfur removal composite method and a crude oil associated gas biological oil and sulfur removal method, and the method comprises the following steps: (1) contacting liquid containing sulfide and crude oil with a system containing oil removal bacteria; and (2) mixing the product obtained in the step (1) with a system containing desulfurization bacteria. The method has the advantages of being low in cost, easy to operate, good in effect, environmentally friendly and the like, the crude oil and sulfur in the liquid containing the sulfide and the crude oil can be efficiently removed, and the method is an environment-friendly and sustainable environment treatment technology.

Description

Technical Field

[0001] This invention relates to the field of environmental microbiology technology, specifically to a combined biological oil and sulfur removal method and a method for biological oil and sulfur removal from associated gas of crude oil. Background Technology

[0002] Associated gas is natural gas extracted from the ground along with crude oil during oil extraction. This gas is mainly composed of hydrocarbons such as methane and ethane, but may also contain certain amounts of non-hydrocarbon components such as hydrogen sulfide and carbon dioxide. Hydrogen sulfide is a particularly harmful component of associated gas. It is a colorless, highly toxic, acidic gas with a strong, pungent odor. It poses a significant threat to human health; even low concentrations can irritate the eyes and respiratory tract, causing headaches, nausea, and vomiting; high concentrations can rapidly lead to coma and even death. Furthermore, hydrogen sulfide corrodes metal equipment, shortening its lifespan and increasing the cost of oilfield extraction and production. Direct discharge of associated gas without treatment can also cause severe environmental pollution.

[0003] The recovery and utilization of associated gas is a crucial aspect of the petroleum development industry. Early methods involved venting and combustion, which polluted the environment and wasted significant energy. As development technologies matured, associated gas is increasingly being treated through recovery methods, but these methods still face challenges such as high costs, significant difficulties, and the presence of hydrogen sulfide. Currently, some associated gas deoiling devices primarily remove light hydrocarbons from the associated gas through centrifugation, filtration, and cooling. However, these devices suffer from drawbacks such as high energy consumption and the need for regular replacement of consumables. Summary of the Invention

[0004] The inventors discovered through research that the commonly used desulfurization technologies in oil fields are mainly wet desulfurization and dry desulfurization. Biological desulfurization is also gradually being applied to associated gas. However, a small amount of crude oil in associated gas has a significant impact on desulfurization microorganisms. How to remove crude oil from associated gas is a key issue for biological desulfurization systems.

[0005] To overcome the problem that the associated gas may contain crude oil, which has a significant impact on the biological desulfurization system in existing associated gas biological desulfurization methods, this invention provides a combined biological oil removal and desulfurization method and a method for biological oil removal and desulfurization of associated gas. The combined biological oil removal and desulfurization method of this invention can effectively remove crude oil and sulfur from liquids containing sulfides and crude oil.

[0006] To achieve the above objectives, the first aspect of the present invention provides a combined biological oil and sulfur removal method, the method comprising:

[0007] (1) Contact the liquid containing sulfides and crude oil with a system containing oil-removing bacteria;

[0008] (2) Mix the product of step (1) with the system containing desulfurizing bacteria.

[0009] A second aspect of the present invention provides a method for biological oil and sulfur removal from associated gas of crude oil, the method comprising:

[0010] (I) Contacting associated gas from crude oil with an alkaline solution yields an alkaline liquid containing sulfides and crude oil;

[0011] (II) De-oiling and desulfurization of alkaline liquids containing sulfides and crude oil according to the composite method described in this invention.

[0012] Through the above technical solution, the biological oil and sulfur removal composite method of the present invention can effectively remove crude oil and sulfur from liquids containing sulfides and crude oil.

[0013] Especially for associated gas from crude oil, after absorbing crude oil and sulfides from the associated gas with alkaline solution, the composite method described in this invention is used to de-oil the associated gas through a system containing oil-removing bacteria and desulfurize it through a system containing desulfurizing bacteria; this effectively removes crude oil from the liquid, reduces the impact on the desulfurizing bacteria in the system containing desulfurizing bacteria, and achieves effective desulfurization of associated gas from crude oil.

[0014] Biological Preservation

[0015] The *Pseudomonas aeruginosa* strain G028 provided by this invention was deposited on April 11, 2014, 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, 100101, China) (abbreviation of depositary institution: CGMCC), with accession number CGMCC No. 9039.

[0016] The *Geobacillus thermodenitrificans* strain JSHD-2 provided by this invention was deposited on March 5, 2013, 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, 100101, China) (abbreviation of depositary institution: CGMCC), with accession number CGMCCNo.7271.

[0017] The Geobacillus toebii strain JSHD-4 provided by this invention was deposited on March 5, 2013, 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, 100101, China) (abbreviation of depositary institution: CGMCC), with accession number CGMCC No. 7273. Attached Figure Description

[0018] Figure 1 This is a comparison chart showing the co-culture of oil-removing bacteria G028 and desulfurizing bacteria JSHD-4 with the culture of desulfurizing bacteria JSHD-4 alone. Detailed Implementation

[0019] 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.

[0020] The first aspect of this invention provides a combined biological oil and sulfur removal method, the method comprising:

[0021] (1) Contact the liquid containing sulfides and crude oil with a system containing oil-removing bacteria;

[0022] (2) Mix the product of step (1) with the system containing desulfurizing bacteria.

[0023] According to a preferred embodiment of the present invention, the system containing oil-removing bacteria contains Pseudomonas aeruginosa strain G028; by adding the strain of the present invention to the liquid containing sulfides and crude oil, the oil-removing bacteria remove the crude oil from the liquid, reducing the impact on the desulfurizing bacteria, and subsequent treatment with desulfurizing bacteria achieves a highly efficient desulfurization effect.

[0024] The G028 strain described in this invention was isolated and screened from the produced fluid of an oil well in Jiangsu Oilfield. The screening method can be any conventional method used in the art for screening new strains. For example, the screening method can be as follows: collect the produced fluid from the oilfield and inoculate it at a ratio of 5% into a culture medium (MgSO4 0.2g, CaCl2 0.02g, KH2PO4 0.8g, K2HPO4 1.2g, NH4NO3 1.2g, FeCl3 0.05g, H2O 500mL, pH 7.2-7.5, with 0.5% crude oil as the sole carbon source). Incubate in a closed, shaking incubator at 37°C for 3-5 days. Stop the incubation when the culture medium becomes turbid or flocculent. Dilute the enrichment solution sequentially with sterile water, spread the diluted solution onto solid culture plates, and incubate at 37°C for 24-48 hours until single colonies appear. Select a single colony and place it into an EP tube containing 1 mL of liquid culture medium. Incubate in a shaker for 24 hours, and streak the plate again. Repeat the single colony selection process 3-5 times.

[0025] Genomic DNA of the bacterial strain was extracted using a bacterial genomic DNA extraction kit and in accordance with the instructions. After PCR amplification and DNA quality assessment using universal 16S rDNA primers 805R and 8F, the DNA was sent to Nanjing Genscript Biotech Co., Ltd. for 16S rRNA gene sequencing. The gene sequence is shown in SEQ ID NO:1.

[0026] The obtained gene sequence was compared with the NCBI database (https: / / www.ncbi.nlm.nih.gov / ). The results showed that SEQ ID NO:1 was most closely related to Pseudonmonas aeruginosa, with a gene sequence identity of 98%. The strain was identified as Pseudonmonas aeruginosa and named G028.

[0027] The Pseudontomosus aeruginosa strain G028 described in this invention is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 9039 and accession date of April 11, 2014.

[0028] In this invention, the method of adding *Pseudomonas aeruginosa* strains to the oil-removing bacteria system is relatively wide. It can be added in the form of the strain's fermentation broth or as a bacterial agent, which can be a solid or liquid bacterial agent. According to a preferred embodiment of this invention, the *Pseudomonas aeruginosa* strains are added to the oil-removing bacteria system in the form of a liquid bacterial agent.

[0029] According to a preferred embodiment of the present invention, the viable count of *Pseudomonas aeruginosa* G028 in the bacterial agent is 10-1. 8 cfu / mL or higher.

[0030] In this invention, the preparation method of the bacterial agent can be a conventional preparation method in the art; this is an example, but does not limit the scope of the invention. According to a preferred embodiment of the invention, the above-mentioned Pseudomonas aeruginosa is inoculated into a liquid culture medium for cultivation.

[0031] According to the present invention, the liquid culture medium can be a conventional culture medium capable of culturing Pseudomonas aeruginosa. In a preferred embodiment, the liquid culture medium contains 0.2 g of MgSO4, 0.02 g of CaCl2, 0.8 g of KH2PO4, 1.2 g of K2HPO4, 1.2 g of NH4NO3, 0.05 g of FeCl3, 500 mL of H2O, and has a pH of 7.2-7.5, with crude oil as the carbon source.

[0032] In this invention, the crude oil source is crude oil from oil wells in Jiangsu Oilfield.

[0033] According to the present invention, the culture conditions may include: a temperature of 30-37°C and a time of 16-24 hours.

[0034] In this invention, the type of desulfurizing bacteria in the system containing desulfurizing bacteria is not particularly limited, as long as the desulfurizing bacteria and the oil-removing bacteria of this invention do not have an antagonistic effect and can grow together.

[0035] According to a preferred embodiment of the present invention, the system containing desulfurizing bacteria contains one or more of the following strains: thermal denitrifying Bacillus strain and composting Bacillus strain.

[0036] In this invention, there are no particular limitations on the method of adding desulfurizing bacteria to the system containing desulfurizing bacteria, as long as the purpose of this invention can be achieved. Desulfurizing bacteria can be added directly, or a bacterial agent containing desulfurizing bacteria can be added.

[0037] According to a preferred embodiment of the present invention, the thermal denitrifying Bacillus includes thermal denitrifying Bacillus strain JSHD-2.

[0038] The JSHD-2 strain described in this invention was isolated and screened from the produced fluid of an oil well in Jiangsu Oilfield. The screening method can be any conventional method used in the field for screening new strains. For example, the screening method can be as follows: collect the produced fluid from the oil well, add NRB enrichment and isolation medium (0.5g sodium nitrate, 2g sodium acetate, 0.5g KH2PO4, 1.0g MgSO4·7H2O, 0.05g FeCl3·6H2O, 0.1g CaCl2, 0.25g yeast extract, 500ml distilled water; pH 7.0-7.2), and incubate at 35℃ for 5 days to obtain an enriched culture sample. Using the above enriched sample as the initial inoculum, the strain was transferred and cultured three times with NRB enrichment and isolation medium, with a culture period of 2 days. Then, the strain was streaked onto plates for isolation, and a single strain was selected and cultured in the medium for 24 hours. This process of single-strain isolation was repeated three times.

[0039] Genomic DNA was extracted from the strain using a bacterial genomic DNA extraction kit and following the instructions. PCR amplification and DNA quality assessment were performed using universal 16S rDNA primers 805R and 8F. The DNA was then sent to Nanjing GenScript Biotech Co., Ltd. for 16S rRNA gene sequencing. The gene sequence was most closely related to *Geobacillus thermodenitrificans*, with a sequence identity of 97%. The strain was identified as *Geobacillus thermodenitrificans* and named JSHD-2.

[0040] The *Geobacillus thermodenitrificans* strain JSHD-2 described in this invention is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 7271 and deposit date of March 5, 2013.

[0041] According to a preferred embodiment of the present invention, the compost Bacillus includes Bacillus compost strain JSHD-4.

[0042] The Geobacillus toebii strain JSHD-4 described in this invention is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 7273 and deposit date of March 5, 2013.

[0043] According to a preferred embodiment of the present invention, the viable count of *Bacillus thermodenitrile* in the bacterial agent containing *Bacillus thermodenitrile* is 10. 8 cfu / mL or higher.

[0044] According to a preferred embodiment of the present invention, the viable count of *Bacillus compostii* in the inoculant is 10. 8 cfu / mL or higher.

[0045] In this invention, the preparation method of the inoculum containing heat-denitrifying Bacillus can be a conventional preparation method in the art; this is an exemplary description, but does not limit the scope of the invention. According to a preferred embodiment of the invention, the above-mentioned heat-denitrifying Bacillus is inoculated into a liquid culture medium for cultivation.

[0046] According to the present invention, the liquid culture medium can be a conventional culture medium capable of culturing thermo-denitrifying Bacillus. In a preferred embodiment, the liquid culture medium contains 1.0 g / L Na2NO3, 4 g / L sodium acetate, 1.0 g / L MgSO4, 0.1 g / L FeCl3, 0.1 g / L CaCl2, 1.0 g / L KH2PO4, and 0.5 g / L yeast extract.

[0047] In this invention, the preparation method of the inoculant containing Bacillus compostii can be a conventional preparation method in the art; this is an example, but does not limit the scope of the invention. According to a preferred embodiment of the invention, the above-mentioned Bacillus compostii is inoculated into a liquid culture medium for cultivation.

[0048] According to the present invention, the liquid culture medium can be a conventional culture medium capable of culturing Bacillus compostii. In a preferred embodiment, the liquid culture medium contains 1.0 g / L Na2NO3, 4 g / L sodium acetate, 1.0 g / L MgSO4, 0.1 g / L FeCl3, 0.1 g / L CaCl2, 1.0 g / L KH2PO4, and 0.5 g / L yeast extract.

[0049] In this invention, after the liquid containing sulfides and crude oil comes into contact with the system containing oil-removing bacteria, the concentration of oil-removing bacteria in the liquid environment can be selected within a wide range. This is illustrative but does not limit the scope of the invention. According to a preferred embodiment of the invention, the concentration of oil-removing bacteria is 10. 7 -10 8 cfu / mL.

[0050] In this invention, the ratio of the oil-removing bacteria system to the liquid can be selected within a wide range, as long as the concentration of the oil-removing bacteria in the mixed liquid environment is 10. 7 -10 8 cfu / mL, illustrative but not limiting of the scope of the invention, according to a preferred embodiment of the invention, the system containing oil-removing bacteria is 0.1%-5% of the total liquid volume.

[0051] In this invention, after the product of step (1) is mixed with the system containing desulfurizing bacteria, the concentration of desulfurizing bacteria in the liquid environment can be selected within a wide range. This is illustrative but does not limit the scope of the invention. According to a preferred embodiment of the invention, the concentration of desulfurizing bacteria is 10. 7 -10 8 cfu / mL.

[0052] In this invention, the ratio of the desulfurizing bacteria-containing system to the product of step (1) can be selected within a wide range, as long as the concentration of desulfurizing bacteria in the mixed liquid environment is 10. 7 -10 8 cfu / mL, illustrative but not limiting of the scope of the invention, according to a preferred embodiment of the invention, the system containing desulfurizing bacteria is 1%-5% of the total volume of the product of step (1).

[0053] In this invention, the system containing oil-removing bacteria also contains nutrients required for the growth of oil-removing bacteria, such as water, carbon source, nitrogen source, and inorganic salts.

[0054] In this invention, the system containing desulfurizing bacteria also contains nutrients required for the growth of desulfurizing bacteria, such as water, carbon source, nitrogen source, and inorganic salts.

[0055] In this invention, the *Pseudomonas aeruginosa* strain has a wide tolerance range for acids and alkalis, and there are no particular limitations on the contact conditions with liquids containing sulfides and crude oil. Conventional contact conditions in the art are sufficient. According to a preferred embodiment of this invention, the contact conditions include: a temperature of 30-37°C; and / or a pH of 7.5-8.5.

[0056] In this invention, the product obtained in step (1) can achieve the purpose of this invention whether it contains crude oil or not. The crude oil content 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 is not higher than 30 mg / L, preferably 5-20 mg / L.

[0057] According to a preferred embodiment of the present invention, the pH of the product obtained in step (1) is 7.5-8.5.

[0058] In this invention, the mixing conditions in step (2) are not particularly limited, and conventional mixing conditions in the art are acceptable. According to a preferred embodiment of this invention, the mixing conditions include a temperature of 30-37°C.

[0059] In this invention, the range of types of sulfides that can be selected from the liquid containing sulfides and crude oil is relatively wide. This is an illustrative example, but does not limit the scope of the invention. According to a preferred embodiment of the invention, the sulfides are selected from sulfides of Group IA metals and / or hydrosulfides of Group IA metals, such as sodium sulfide and sodium hydrosulfide.

[0060] In this invention, the sulfide content in the liquid containing sulfides and crude oil can be selected within 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 sulfide content, calculated as sulfur element, is 50-200 mg / L.

[0061] In this invention, the crude oil content in the liquid containing sulfides and crude oil can be selected from a wide range. This is an illustrative example, but does not limit the scope of the invention. According to a preferred embodiment of the invention, the crude oil content is 5-80 mg / L, preferably 5-20 mg / L.

[0062] A second aspect of this invention provides a combined method for biological oil and sulfur removal from associated gas of crude oil, the method comprising:

[0063] (I) Contacting associated gas from crude oil with an alkaline solution yields an alkaline liquid containing sulfides and crude oil;

[0064] (II) De-oiling and desulfurization of alkaline liquid containing sulfides and crude oil according to the composite method of the present invention. After absorbing crude oil and sulfides in the associated gas of crude oil with alkaline solution, the composite method of the present invention is used to de-oil the associated gas through a system containing oil-removing bacteria and desulfurize it through a system containing desulfurizing bacteria; effectively removing crude oil from the liquid, reducing the impact on the desulfurizing bacteria in the system containing desulfurizing bacteria, and achieving effective desulfurization of the associated gas of crude oil.

[0065] In this invention, the flow rate of the associated gas from crude oil can be selected within a wide range. This is an illustrative example and does not limit the scope of the invention. According to a preferred embodiment of the invention, the flow rate of the associated gas from crude oil is 40-60 m / s. 3 / h.

[0066] In this invention, the range of alkaline concentrations is relatively wide. This is an illustrative example, but it does not limit the scope of the invention. According to a preferred embodiment of the invention, the alkaline concentration is 30-40 g / L.

[0067] In this invention, the range of types of alkali in the alkaline solution is relatively wide. This is an illustrative example, but it does not limit the scope of the invention. According to a preferred embodiment of the invention, the alkali is selected from one or more of alkali metal hydroxides, carbonates, and bicarbonates, such as sodium hydroxide, sodium carbonate, sodium bicarbonate, etc.

[0068] In this invention, there are no particular limitations on the contact conditions; conventional contact conditions in the art are sufficient. According to a preferred embodiment of the invention, the contact conditions include a temperature of 30-37°C.

[0069] 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.

[0070] The present invention specification and context, including the following embodiments, use spectrophotometry to measure the absorbance of bacterial suspensions at a wavelength of 600 nm by measuring the OD value of the bacterial suspension.

[0071] 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.

[0072] In the present invention specification and context, including the following embodiments, the national standard HJ / T60-2000 is used to determine the sulfide content in the solution, and the iodometric method is used to detect the sulfide content.

[0073] Formula for calculating desulfurization rate:

[0074] X – Desulfurization rate, %;

[0075] V1 – Sulfur content in wastewater before treatment;

[0076] V2 – Sulfur content in the treated wastewater.

[0077] In the following examples, the G028 bacterial agent is the product of Pseudomonas aeruginosa strain G028 after being cultured in liquid medium (MgSO4 0.2g, CaCl2 0.02g, KH2PO4 0.8g, K2HPO4 1.2g, NH4NO3 1.2g, FeCl3 0.05g, H2O 500mL, pH 7.5, 0.5wt% crude oil as carbon source) at 30°C.

[0078] In the following examples, the JSHD-2 bacterial agent is the product of *Bacillus thermodenitrificans* JSHD-2 cultured in liquid medium (1.0 g / L Na2NO3, 4 g / L sodium acetate, 1.0 g / L MgSO4, 0.1 g / L FeCl3, 0.1 g / L CaCl2, 1.0 g / L KH2PO4, 0.5 g / L yeast extract) at 30°C.

[0079] In the following examples, the JSHD-4 inoculant is the product of Bacillus compostii JSHD-4 cultured in liquid culture medium (1.0 g / L Na2NO3, 4 g / L sodium acetate, 1.0 g / L MgSO4, 0.1 g / L FeCl3, 0.1 g / L CaCl2, 1.0 g / L KH2PO4, 0.5 g / L yeast extract) at 30°C.

[0080] Example 1

[0081] 0.1 mL of viable bacteria with a concentration of 103 was inoculated into 100 mL of culture medium (0.4 g / L MgSO4, 0.04 g / L CaCl2, 1.6 g / L KH2PO4, 2.4 g / L K2HPO4, 2.4 g / L NH4NO3, 0.1 g / L FeCl3, 0.5 g / L yeast extract). 8 G028 bacterial agent with cfu / mL and 0.9 mL of viable bacteria concentration of 10 8 JSHD-2 bacterial agent at cfu / mL was inoculated into 1.0 mL of a 10 CFU / mL culture medium. 8 A JSHD-2 bacterial suspension of cfu / mL was used as a control group and cultured at 30℃ to observe the growth and reproduction of the two bacteria under a common nutrient system.

[0082] Using a blank culture medium as a blank control, the spectrophotometer was zeroed. Small amounts of the cultured liquid were collected after 24h, 48h, and 72h of incubation to determine the OD600 value. Figure 1As shown, after 16 hours of cultivation, the OD600 value of the liquid was 0.690, indicating that the bacteria were in the logarithmic growth phase. A small amount of the liquid was centrifuged and Gram-stained for microscopic examination. Most bacteria in the field of view were stained blue-purple, with a few red, indicating significant growth of the JSHD-2 strain. A small amount of G028 also showed growth. There was no significant difference in the OD600 value (0.675) after 16 hours of cultivation compared to the control group inoculated with JSHD-2 alone. After 24 hours of cultivation, the OD600 value of the liquid was 0.846, indicating that the bacterial concentration reached its maximum, while the OD600 value of the control group was 0.819. These results indicate that there is no significant competition for growth between the two bacteria in the mixed bacterial culture, and they can co-grow.

[0083] Example 2

[0084] (1) The flow rate is 40m 3 A 30 g / L sodium hydroxide solution is passed through crude oil associated gas at a flow rate of / h. The solution temperature is 30℃, and a liquid product containing sodium sulfide is obtained, in which the sulfide content is 50 mg / L, the oil content is 15 mg / L, and the pH is 8.2.

[0085] (2) An alkaline liquid containing sulfides and crude oil was added to bioreactor A. Tank A contained 0.4% MgSO4, 0.04% CaCl2, 1.6% KH2PO4, 2.4% K2HPO4, 2.4% NH4NO3, and 0.1% FeCl3, by total liquid volume. The temperature of Tank A was set to 30℃, and the pH of the liquid was adjusted to 7.5. 3% of the total liquid volume of G028 bacterial agent was added to Tank A. The concentration of G028 in the inoculated solution was 10%. 8 cfu / mL, Tank A is filled with rope-shaped packing material, oil-removing bacteria grow on the packing material and remove crude oil from the liquid, and the bottom of Tank A of the bioreactor is equipped with an aeration device to provide oxygen to the oil-removing bacteria.

[0086] (3) After treatment with G028 bacterial agent, the oil content in the liquid of tank A was 10 mg / L. The supernatant from tank A entered bioreactor tank B. Tank B contained 1% Na2NO3, 4% sodium acetate, 2% MgSO4, 0.1% FeCl3, 0.05% CaCl2, 1% KH2PO4, and 0.5% yeast powder, by total liquid volume. The temperature was set to 30℃, the pH of the liquid was adjusted to 7.5, and 3% of the total liquid volume of tank B was added with JSHD-2 bacterial agent. The concentration of JSHD-2 in the inoculated solution was 10 mg / L. 8 cfu / mL; the bottom of bioreactor B is equipped with an aeration device to provide oxygen to the desulfurizing bacteria.

[0087] After adding JSHD-4 bacterial agent to pool B, small amounts of liquid from pool B were taken at 3, 7, and 10 days of cultivation to determine the oil content and desulfurization rate. At 3 days of cultivation, the oil content in pool B was 0.5 mg / L, and the desulfurization rate was 85.5%. At 7 days of cultivation, the oil content in pool B was 0.1 mg / L, and the desulfurization rate was 90.3%. At 10 days of cultivation, the oil content in pool B was 0 mg / L, and the desulfurization rate was 97.5%.

[0088] Example 3

[0089] (1) The flow rate is 50m 3 A crude oil associated gas of / h is passed through a 35g / L sodium carbonate solution at a solution temperature of 32℃ to obtain a liquid product containing sodium sulfide, wherein the sulfide content is 60mg / L, the liquid oil content is 15mg / L, and the pH is 8.3.

[0090] (2) An alkaline liquid containing sulfides and crude oil was added to bioreactor A. Tank A contained 0.4% MgSO4, 0.04% CaCl2, 1.6% KH2PO4, 2.4% K2HPO4, 2.4% NH4NO3, and 0.1% FeCl3, by total liquid volume. The temperature of tank A was set to 30℃, and the pH of the liquid was adjusted to 7.5. 5% of the total liquid volume of G028 bacterial agent was added to tank A. The concentration of G028 in the inoculated solution was 10%. 8 cfu / mL, tank A is filled with rope-shaped packing material, and oil-removing bacteria will grow on the packing material and remove crude oil from the liquid. The bottom of tank A of the bioreactor is equipped with an aeration device to provide oxygen to the oil-removing bacteria.

[0091] (3) After treatment with G028 bacterial agent, the oil content in the liquid of tank A was 8 mg / L. The supernatant from tank A entered bioreactor tank B. Tank B contained 1% Na2NO3, 4% sodium acetate, 2% MgSO4, 0.1% FeCl3, 0.05% CaCl2, 1% KH2PO4, and 0.5% yeast powder, by total liquid volume. The temperature of tank B was set to 30℃, the pH of the liquid was adjusted to 7.5, and 5% of the total liquid volume of tank B was added with JSHD-2 bacterial agent. The concentration of JSHD-2 in the inoculated solution was 10. 8 cfu / mL; the bottom of bioreactor B is equipped with an aeration device to provide oxygen to the desulfurizing bacteria.

[0092] After adding JSHD-4 bacterial agent to pool B, small amounts of liquid from pool B were taken at 3, 7, and 10 days of cultivation to determine the oil content and desulfurization rate. At 3 days of cultivation, the oil content in pool B was 0.6 mg / L, and the desulfurization rate was 84.1%. At 7 days of cultivation, the oil content in pool B was 0.15 mg / L, and the desulfurization rate was 89.7%. At 10 days of cultivation, the oil content in pool B was 0 mg / L, and the desulfurization rate was 98.1%.

[0093] Example 4

[0094] (1) The flow rate is 40m 3 A crude oil associated gas of / h is passed into a 40g / L sodium carbonate solution at a solution temperature of 32℃ to obtain a liquid product containing sodium hydrosulfide, wherein the sulfide content is 65mg / L, the liquid oil content is 20mg / L, and the pH is 8.2.

[0095] (2) An alkaline liquid containing sulfides and crude oil was added to bioreactor A. Tank A contained 0.4% MgSO4, 0.04% CaCl2, 1.6% KH2PO4, 2.4% K2HPO4, 2.4% NH4NO3, and 0.1% FeCl3, by total liquid volume. The temperature of tank A was set to 30℃, and the pH of the liquid was adjusted to 7.5. 5% of the total liquid volume of G028 bacterial agent was added to tank A. The concentration of G028 in the inoculated solution was 10%. 8 cfu / mL, tank A is filled with rope-shaped packing material, and oil-removing bacteria will grow on the packing material and remove crude oil from the liquid. The bottom of tank A of the bioreactor is equipped with an aeration device to provide oxygen to the oil-removing bacteria.

[0096] (3) After treatment with G028 bacterial agent, the oil content in the liquid of tank A is 10 mg / L. The supernatant in tank A will enter bioreactor tank B. In tank B, based on the total liquid volume, 1% Na2NO3, 4% sodium acetate, 2% MgSO4, 0.1% FeCl3, 0.05% CaCl2, 1% KH2PO4, and 0.5% yeast powder are added. The temperature is set to 30℃, and the pH of the liquid is adjusted to 7.5. 5% of the total liquid volume of tank B is added with JSHD-4 bacterial agent. The concentration of JSHD-4 bacteria in the inoculated solution is 10 mg / L. 8 cfu / mL; the bottom of bioreactor B is equipped with an aeration device to provide oxygen to the desulfurizing bacteria.

[0097] After adding JSHD-2 bacterial agent to pool B, small amounts of liquid from pool B were taken at 3, 7, and 10 days of cultivation to determine the oil content and desulfurization rate. At 3 days of cultivation, the oil content in pool B was 0.5 mg / L, and the desulfurization rate was 84.8%. At 7 days of cultivation, the oil content in pool B was 0.12 mg / L, and the desulfurization rate was 90.2%. At 10 days of cultivation, the oil content in pool B was 0 mg / L, and the desulfurization rate was 98.3%.

[0098] Example 5

[0099] (1) The flow rate is 50m 3A crude oil associated gas of / h is passed through a 35g / L sodium hydroxide solution at a solution temperature of 35℃ to obtain a liquid product containing sodium sulfide, wherein the sulfide content is 100mg / L, the oil content is 10mg / L, and the pH is 8.3.

[0100] (2) An alkaline liquid containing sulfides and crude oil was added to bioreactor A. Tank A contained 0.4% MgSO4, 0.04% CaCl2, 1.6% KH2PO4, 2.4% K2HPO4, 2.4% NH4NO3, and 0.1% FeCl3, by total liquid volume. The temperature of tank A was set to 30℃, and the pH of the liquid was adjusted to 7.5. 5% of the total liquid volume of G028 bacterial agent was added to tank A. The concentration of G028 in the inoculated solution was 10%. 8 cfu / mL, tank A is filled with rope-shaped packing material, and oil-removing bacteria will grow on the packing material and remove crude oil from the liquid. The bottom of tank A of the bioreactor is equipped with an aeration device to provide oxygen to the oil-removing bacteria.

[0101] (3) After processing the product from step (2) to a crude oil content of 0 mg / L, it is fed into bioreactor B tank. Tank B contains, by total liquid volume, 1% Na₂NO₃, 4% sodium acetate, 2% MgSO₄, 0.1% FeCl₃, 0.05% CaCl₂, 1% KH₂PO₄, and 0.5% yeast powder. The temperature of tank B is set to 30℃, the pH of the liquid is adjusted to 7.5, and 5% of the total liquid volume of JSHD-2 bacterial agent is added. The concentration of JSHD-2 bacteria in the inoculated solution is 10... 8 cfu / mL; the bottom of bioreactor B is equipped with an aeration device to provide oxygen to the desulfurizing bacteria.

[0102] After adding JSHD-4 bacterial agent to pool B, small amounts of liquid from pool B were taken at 3, 7, and 10 days of cultivation to determine the oil content and desulfurization rate. At 3 days of cultivation, the desulfurization rate in pool B was 84.1%. At 7 days of cultivation, the desulfurization rate was 89.7%. At 10 days of cultivation, the desulfurization rate was 98.2%.

[0103] Example 6

[0104] (1) Crude oil (from an oil well in Jiangsu Oilfield) and sodium sulfide solution were added to bioreactor A until the oil content was 15 mg / L and the sodium sulfide content was 60 mg / L. Tank A contained 0.4% MgSO4, 0.04% CaCl2, 1.6% KH2PO4, 2.4% K2HPO4, 2.4% NH4NO3, and 0.1% FeCl3, by total liquid volume. The temperature of Tank A was set to 30℃, and the pH of the liquid was adjusted to 7.5. 5% of the total liquid volume of G028 bacterial agent was added to Tank A. The concentration of G028 in the inoculated solution was 10%. 8cfu / mL, tank A is filled with rope-shaped packing material, and oil-removing bacteria will grow on the packing material and remove crude oil from the liquid. The bottom of tank A of the bioreactor is equipped with an aeration device to provide oxygen to the oil-removing bacteria.

[0105] (2) After processing the product from step (1) to a crude oil content of 10 mg / L, it was fed into bioreactor B tank. Tank B contained 1% Na₂NO₃, 4% sodium acetate, 2% MgSO₄, 0.1% FeCl₃, 0.05% CaCl₂, 1% KH₂PO₄, and 0.5% yeast powder, by total liquid volume. The temperature of tank B was set to 30℃, the pH of the liquid was adjusted to 7.5, and 5% of the total liquid volume of JSHD-2 bacterial agent was added. The concentration of JSHD-2 bacteria in the inoculated solution was 10 mg / L. 8 cfu / mL; the bottom of bioreactor B is equipped with an aeration device to provide oxygen to the desulfurizing bacteria.

[0106] After adding JSHD-4 bacterial agent to pool B, small amounts of liquid from pool B were taken at 3, 7, and 10 days of cultivation to determine the oil content and desulfurization rate. At 3 days of cultivation, the oil content in pool B was 0.6 mg / L, and the desulfurization rate was 84.4%. At 7 days of cultivation, the oil content in pool B was 0.18 mg / L, and the desulfurization rate was 89.2%. At 10 days of cultivation, the oil content in pool B was 0 mg / L, and the desulfurization rate was 98.0%.

[0107] Comparative Example 1

[0108] (1) The flow rate is 50m 3 A crude oil associated gas of / h is passed through a 35g / L sodium hydroxide solution at a solution temperature of 35℃ to obtain a liquid product containing sodium sulfide, wherein the sulfide content is 100mg / L, the liquid oil content is 10mg / L, and the pH is 8.3.

[0109] (2) An alkaline liquid containing sulfides and crude oil was added to a bioreactor. The bioreactor contained 1% Na₂NO₃, 4% sodium acetate, 2% MgSO₄, 0.1% FeCl₃, 0.05% CaCl₂, 1% KH₂PO₄, and 0.5% yeast powder, by total liquid volume. The temperature was set to 30℃, and the pH of the liquid was adjusted to 7.5. 5% of the total liquid volume of JSHD-2 bacterial agent was added to the bioreactor. The concentration of JSHD-2 in the inoculated solution was 10%. 8 cfu / mL; the bioreactor is equipped with an aeration device at the bottom to provide oxygen to the desulfurizing bacteria.

[0110] Small amounts of the bioreactor effluent were taken at 3, 7, and 10 days of incubation to determine the oil content and desulfurization rate. At 3 days of incubation, the oil content at the bioreactor effluent was 4.8 mg / L, and the desulfurization rate was 50.5%. At 7 days of incubation, the oil content at the bioreactor effluent was 4.7 mg / L, and the desulfurization rate was 55.1%. At 10 days of incubation, the oil content at the bioreactor effluent was 4.7 mg / L, and the desulfurization rate was 60.3%.

[0111] 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 combined biological oil and sulfur removal method, characterized in that, The method includes: (1) Contact the liquid containing sulfides and crude oil with a system containing oil-removing bacteria; (2) Mix the product of step (1) with the system containing desulfurizing bacteria.

2. The composite method according to claim 1, wherein, The system containing oil-removing bacteria contains Pseudontomosus aeruginosa strain G028; the preservation number of Pseudontomosus aeruginosa is CGMCC No. 9039.

3. The composite method according to claim 1 or 2, wherein, The system containing desulfurizing bacteria contains one or more of the strains of *Geobacillus thermodenitrificans* and *Geobacillus toebii*.

4. The composite method according to any one of claims 1-3, wherein, The *Bacillus denitrifyingus* species includes *Bacillus denitrifyingus* strain JSHD-2, with accession number CGMCC No. 7271; and / or The Bacillus subtilis strain mentioned includes Bacillus subtilis strain JSHD-4, with accession number CGMCC No. 7273.

5. The composite method according to any one of claims 1-4, wherein, When a liquid containing sulfides and crude oil comes into contact with a system containing oil-removing bacteria, the concentration of the oil-removing bacteria is 10. 7 -10 8 cfu / mL; and / or The system containing oil-removing bacteria is 0.1%-5% of the total volume of the liquid.

6. The composite method according to any one of claims 1-5, wherein, After the product of step (1) is mixed with the system containing desulfurizing bacteria, the concentration of desulfurizing bacteria is 10. 7 -10 8 cfu / mL; and / or The system containing desulfurizing bacteria is 1%-5% of the total volume of the product from step (1).

7. The composite method according to any one of claims 1-6, wherein, In step (1), the contact conditions include: a temperature of 30-37°C and / or a pH of 7.5-8.5; and / or The crude oil content in the product obtained in step (1) is not higher than 30 mg / L, preferably 5-20 mg / L; and / or In step (2), the mixing conditions include a temperature of 30-37°C and / or a pH of 7.5-8.

5.

8. The composite method according to any one of claims 1-7, wherein, The sulfide is selected from sulfides of Group IA metals and / or hydrosulfides of Group IA metals, preferably one or more of sodium sulfide and sodium hydrosulfide; and / or In liquids containing sulfides and crude oil, the sulfide content, calculated as elemental sulfur, is 50-200 mg / L; and / or The crude oil content is 5-80 mg / L.

9. A method for biological oil and sulfur removal from associated gas of crude oil, characterized in that, The method includes: (I) Contacting associated gas from crude oil with an alkaline solution yields an alkaline liquid containing sulfides and crude oil; (II) The composite method according to any one of claims 1-8 for degreasing and desulfurizing alkaline liquids containing sulfides and crude oil.

10. The method according to claim 9, wherein, The flow rate of associated gas from crude oil is 40-60 m / s. 3 / h; and / or The concentration of the alkaline solution is 30-40 g / L. Preferably, the alkali in the alkaline solution is selected from one or more of alkali metal hydroxides, carbonates, and bicarbonates; and / or Contact conditions include a temperature of 30-37℃.

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