Petroleum degradation solid microbial inoculum, preparation method and application thereof

Abstract

The invention discloses a petroleum degradation solid microbial inoculum and a preparation method and application thereof, and the petroleum degradation solid microbial inoculum comprises a solid culture medium and a composite bacterial liquid; wherein the compound bacteria liquid is obtained by compounding pseudomonas aeruginosa and bacillus clausii, and the inoculum size of the compound bacteria liquid is 1-10% of the mass of the solid culture medium; the solid culture medium comprises the following components in parts by mass: 100-600 parts of vinasse particles, 3-10 parts of a nitrogen source, 5-10 parts of a carbon source, 3-10 parts of inorganic salt, 1-3 parts of monopotassium phosphate, 2-6 parts of dipotassium phosphate, 1-3 parts of magnesium sulfate heptahydrate and 50-100 parts of an alkaline substance. The problems that an existing microbial immobilization technology is limited in application range and difficult to exert effects independently are solved. The petroleum degradation solid microbial inoculum can effectively improve the effective viable count of petroleum degradation bacteria in a pollution system, and realizes field planting and reproduction of strains in soil. The water-retaining capacity of the soil is improved, the content of organic matters and part of trace elements is improved, and the nutrition in the soil is increased.

Description

Technical Field

[0001] The present application relates to the field of microorganisms and environmental biotechnology, and in particular to a petroleum-degrading solid bacterial agent, a preparation method and an application thereof. Background Art

[0002] The impact of petroleum pollutants is expanding day by day. Its main characteristics are deep pollution, wide range and great harm. Specifically, it has harm and impact on ecological environment, soil health, water health, human health, animals and plants. After petroleum pollutants enter the soil, they will change the physical and chemical properties of the soil. After pollution, the total organic carbon concentration and pH of the soil will increase relatively, and the total nitrogen concentration will decrease relatively. In addition, petroleum pollutants are easy to adhere to the soil and have strong hydrophobicity, which will inhibit the air permeability and water permeability of the soil, causing the soil to clump and harden, affecting the physical properties of the soil. In addition, petroleum is rich in reactive groups, which can combine with inorganic nitrogen and phosphorus and limit nitrification and dephosphorylation, thereby reducing the effective nitrogen and phosphorus concentrations in the soil and affecting the absorption of crops. Furthermore, after petroleum substances enter the soil, they will undergo a series of actions in the soil ecosystem, leaving residues in various parts of the soil and crops, affecting the quality of food and reducing the quality of food.

[0003] In recent years, microorganisms have been considered the most promising and green sustainable remediation technology for organic pollutants, which can be removed by metabolizing and transforming them to achieve true harmless treatment. In order to improve the environmental adaptability of microorganisms and enhance the removal effect of pollutants, the immobilization technology of microorganisms has received more and more attention and is widely used in the remediation of soil contaminated by organic pollutants. However, it also faces some challenges in practical applications, including environmental conditions, slow degradation rate and limited scope of application. Therefore, in practical applications, it is often necessary to combine other remediation technologies and make comprehensive use of their advantages to achieve the best pollution remediation effect. Summary of the invention

[0004] The embodiments of the present application provide a petroleum-degrading solid bacterial agent, a preparation method and an application thereof, thereby solving the problem that the existing microbial immobilization technology has a limited scope of application and is difficult to exert its effect alone.

[0005] In a first aspect, an embodiment of the present application provides a petroleum-degrading solid bacterial agent, comprising a solid culture medium and a composite bacterial liquid; the components of the solid culture medium and their mass fractions include: 100-600 parts of wine lees particles, 3-10 parts of a nitrogen source, 5-10 parts of a carbon source, 3-10 parts of an inorganic salt, 1-3 parts of potassium dihydrogen phosphate, 2-6 parts of dipotassium hydrogen phosphate, 1-3 parts of magnesium sulfate heptahydrate and 50-100 parts of an alkaline substance; the composite bacterial liquid is obtained by compounding Pseudomonas aeruginosa and Bacillus clausii, and the inoculation amount of the composite bacterial liquid is 1% to 10% of the mass of the solid culture medium.

[0006] In combination with the first aspect, in a possible implementation manner, the pH value of the petroleum-degrading solid bacterial agent is 7.2.

[0007] In a second aspect, an embodiment of the present application provides a method for preparing a petroleum-degrading solid bacterial agent, comprising: crushing air-dried and sun-dried vinasse, and screening out vinasse particles therefrom; inoculating the bacterial strains into a liquid fermentation medium respectively, and culturing at a constant temperature of 180 rpm and 37°C for 24 hours to 48 hours to obtain a single bacterial liquid; preparing the single bacterial liquid in proportion to obtain a composite bacterial liquid; adding nutrients to the vinasse particles, and adjusting the water content and pH value thereof to obtain a solid culture medium; adding the composite bacterial liquid to the solid culture medium, stirring evenly, and fermenting and culturing at 30°C to 40°C for 48 hours to 72 hours to obtain a petroleum-degrading solid bacterial agent; wherein the inoculation amount of the composite bacterial liquid is 1%-10% of the mass of the solid culture medium.

[0008] In combination with the second aspect, in a possible implementation, the wine grains include alcohol grains and / or white wine grains and / or yellow wine grains and / or beer grains.

[0009] In combination with the second aspect, in a possible implementation, the particle size of the lees particles is 1 mm-3 mm.

[0010] In combination with the second aspect, in a possible implementation, the nutrients include a nitrogen source, a carbon source and an inorganic salt; wherein the nitrogen source is yeast powder, the carbon source is molasses, and the inorganic salt is potassium dihydrogen phosphate, dipotassium hydrogen phosphate, and magnesium sulfate heptahydrate.

[0011] In combination with the second aspect, in a possible implementation, adjusting the moisture content and pH value to obtain a solid culture medium includes: adding water to the lees particles and the nutrients to make their moisture content 20%-50%; adding alkali to the lees particles and the nutrients to make their pH value 5-9.

[0012] In combination with the second aspect, in a possible implementation manner, the bacterial species include Bacillus Clausii and Pseudomonas aeruginosa, and the ratio of Pseudomonas aeruginosa to Bacillus Clausii is 1:5 to 5:1.

[0013] In a third aspect, an embodiment of the present application provides an application of a petroleum-degrading solid bacterial agent for treating petroleum pollutants.

[0014] One or more technical solutions provided in the embodiments of the present application have at least the following technical effects or advantages:

[0015] The embodiment of the present application uses waste vinasse as an immobilized carrier, solid fermentation with petroleum-degrading bacteria, and preparation of a petroleum-degrading solid bacterial agent to achieve the restoration of oil-containing soil. The petroleum-degrading solid bacterial agent of the present application can effectively increase the effective live bacteria of petroleum-degrading bacteria in the pollution system, realize the colonization and reproduction of strains in the soil, and improve the water permeability and air permeability of the soil on the basis of efficient degradation. At the same time, vinasse can significantly improve the water retention capacity of the soil, increase the content of organic matter and some trace elements, and increase the nutrition in the soil. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings required for use in the embodiments of the present application or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0017] Figure 1 An example diagram of a petroleum-degrading solid bacterial agent provided in an embodiment of the present application;

[0018] Figure 2 A flow chart of a method for preparing a petroleum-degrading solid bacterial agent provided in an embodiment of the present application;

[0019] Figure 3 A graph showing the relationship between the oil content and the degradation rate of the petroleum-degrading solid bacterial agent provided in the examples of the present application;

[0020] Figure 4 A graph showing the relationship between the degradation temperature and degradation rate of the petroleum-degrading solid bacterial agent provided in the embodiments of the present application;

[0021] Figure 5 A graph showing the relationship between the inoculation amount and the degradation rate of the petroleum-degrading solid bacterial agent provided in the embodiments of the present application;

[0022] Figure 6 This is a graph showing the relationship between the degradation time and degradation rate of the petroleum-degrading solid bacterial agent provided in the examples of the present application. DETAILED DESCRIPTION

[0023] The technical solutions in the embodiments of the present application will be described clearly and completely below in conjunction with the drawings in the embodiments of the present application. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0024] The following describes some of the techniques involved in the embodiments of the present application to facilitate understanding, and they should be considered as merely exemplary. Therefore, it should be appreciated by those of ordinary skill in the art that various changes and modifications may be made to the embodiments described herein without departing from the scope and spirit of the present application. Similarly, for the sake of clarity and conciseness, some descriptions of well-known functions and structures are omitted in the following description.

[0025] The weight of the relevant components mentioned in the description of the embodiments of the present invention may not only refer to the specific content of each component, but also indicate the weight ratio between the components. Therefore, as long as the content of the relevant components is proportionally enlarged or reduced according to the description of the embodiments of the present invention, it is within the scope disclosed in the description of the embodiments of the present invention. Specifically, the units described in the description of the embodiments of the present invention may be mass units known in the chemical industry such as mm, g / L, h, kg, etc.

[0026] like Figure 1 As shown, an example diagram of a solid culture bacterial agent is provided. It includes a solid culture medium and a composite bacterial solution; the components and their mass fractions of the solid culture medium include: 100-600 parts of lees particles, 3-10 parts of nitrogen source, 5-10 parts of carbon source, 3-10 parts of inorganic salt, 1-3 parts of potassium dihydrogen phosphate, 2-6 parts of dipotassium hydrogen phosphate, 1-3 parts of magnesium sulfate heptahydrate and 50-100 parts of alkaline substances; the composite bacterial solution is obtained by compounding Pseudomonas aeruginosa and Bacillus clausii, and the inoculation amount of the composite bacterial solution is 1% to 10% of the mass of the solid culture medium.

[0027] The pH value of the single activated bacterial solution is 7.2. The preferred inoculation amount of the composite bacterial solution is 5% of the mass of the solid culture medium.

[0028] like Figure 2 As shown, the present application also provides a method for preparing a solid culture agent for treating oily sludge, including steps 201 to 205, as follows.

[0029] Step 201: crush the air-dried wine lees and screen out the wine lees particles. Specifically, there are complex biochemical changes in the fermentation and winemaking process, which leads to the appearance of new components in the final wine lees, such as RNA, purine, pyrimidine, etc. These substances can create a good environment for the growth of microorganisms.

[0030] In the embodiment of the present application, the used lees can be one or more of alcohol lees, white wine lees, yellow wine lees and beer lees. The discarded lees are naturally air-dried and air-dried, and the air-dried lees are crushed and 1mm-3mm lees particles are screened out for later use.

[0031] Step 202: Inoculate the strains into the liquid fermentation medium respectively, and culture them at a constant temperature of 180 rpm and 37°C for 24 to 48 hours to obtain a single bacterial solution. In the embodiment of the present application, the pH of the liquid culture medium is about 7.2, and high temperature sterilization is performed. Take multiple liquid culture media and add strains thereto, and culture them at a constant temperature of oscillation to obtain a single bacterial solution. Among them, the strains used here are Pseudomonas aeruginosa and Bacillus clausii.

[0032] Exemplarily, the rotation speed of the constant temperature shaking culture is 180 rpm and the temperature is 37°C.

[0033] Step 203: Prepare the single bacterial solution in proportion to obtain a composite bacterial solution. In an embodiment of the present application, several of the multiple single activated bacterial solutions obtained by the above-mentioned constant temperature oscillation culture are prepared in proportion to obtain a composite bacterial solution. The ratio of Pseudomonas aeruginosa and Bacillus clausii is 1:5 to 5:1. Exemplarily, the ratio here can be 1:5 of Pseudomonas aeruginosa and Bacillus clausii. The specific configuration ratio and the single activated bacterial solution used can be determined according to actual needs.

[0034] Step 204: Add nutrients to the lees particles, and adjust the water content and pH value to obtain a solid culture medium. In the embodiment of the present application, the nutrients include a nitrogen source, a carbon source, and an inorganic salt; wherein the nitrogen source is ammonium nitrate, potassium nitrate, yeast extract powder, peptone, ammonium sulfate, or ammonium chloride, the carbon source is molasses, glucose, sucrose, fructose, or starch, and the inorganic salt is sodium chloride, calcium chloride, magnesium chloride, potassium chloride, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, potassium sulfate, magnesium sulfate, sodium sulfate, or zinc sulfate.

[0035] Exemplarily, the water content here is 20%-50%, and the pH value here is 5-9.

[0036] After adding nutrients and adjusting the water content and pH, the obtained solid culture medium is packaged and sterilized at high temperature. For example, the high temperature sterilization here is sterilized at 121° C. for 15 min-20 min.

[0037] Step 205: Add the composite bacterial solution to the solid culture medium, stir evenly, and ferment and culture at 30°C to 40°C for 48h to 72h to obtain a petroleum-degrading solid bacterial agent; wherein the inoculation amount of the composite bacterial solution is 1% to 10% of the mass of the solid culture medium. Specifically, add the composite bacterial solution prepared in step 203 to the solid culture medium, and the inoculation amount of the added composite bacterial solution is 1% to 10% of the mass of the solid culture medium. After shaking evenly, culture at a constant temperature for 24h to 72h, and use the solid culture medium made of wine lees as a carrier for the bacteria to attach and grow, to obtain a petroleum-degrading solid bacterial agent.

[0038] Exemplarily, the temperature of the constant temperature culture here is 30°C-40°C.

[0039] The solid culture bacterial agent prepared by the above steps can be used to treat oily sludge and has a good degradation effect on hydrocarbons in petroleum.

[0040] The following examples are carried out according to the above steps 201 to 205 to verify the effect of the microbial agent on the degradation of petroleum pollutants by the petroleum-degrading solid microbial agent obtained in the present application under different petroleum hydrocarbon concentrations, different temperatures, different inoculation amounts and different degradation time conditions.

[0041] Before conducting the experiment, it is first necessary to prepare oil-containing culture media of different concentrations, add different doses of oil-degrading solid bacterial agents into them to obtain microbial degradation of oil, and measure its oil content to study the effect of oil-degrading solid bacterial agents in degrading petroleum pollutants under different influencing factors, as follows.

[0042] Example 1

[0043] Prepare oil-containing culture medium: add 0.5% NaCl, 0.15% K2HPO4, 0.3% KH2PO4, 0.01% MgSO4·7H2O and 0.001% CaCl2·2H2O to water to prepare liquid culture medium, divide 100 mL of liquid culture medium into several conical flasks, add 50 mg of petroleum to make the concentration 500 mg / L, and finally sterilize at 121°C for 20 minutes.

[0044] Microbial degradation of petroleum: 7 g of the solid petroleum-degrading bacterial agent prepared in the present application was added to the above-mentioned oil-containing culture medium and subjected to microbial degradation at 45°C.

[0045] Determination of oil content: The oil-containing culture medium after 12 days of degradation was extracted and filtered, and the oil content was measured using an infrared oil meter. The degradation rate was 57.64%.

[0046] Example 2

[0047] Prepare oil-containing culture medium: add 0.5% NaCl, 0.15% K2HPO4, 0.3% KH2PO4, 0.01% MgSO4·7H2O and 0.001% CaCl2·2H2O to water to prepare liquid culture medium, divide 100 mL of liquid culture medium into several conical flasks, add 100 mg of petroleum to make the concentration 1000 mg / L, and finally sterilize at 121°C for 20 minutes.

[0048] Microbial degradation of petroleum: 7 g of the solid petroleum-degrading bacterial agent prepared in the present application was added to the above-mentioned oil-containing culture medium and subjected to microbial degradation at 45°C.

[0049] Determination of oil content: The oil-containing culture medium after 12 days of degradation was extracted and filtered, and the oil content was measured using an infrared oil meter. The degradation rate was 56.88%.

[0050] Example 3

[0051] Prepare oil-containing culture medium: add 0.5% NaCl, 0.15% K2HPO4, 0.3% KH2PO4, 0.01% MgSO4·7H2O and 0.001% CaCl2·2H2O to water to prepare liquid culture medium, divide 100 mL of liquid culture medium into several conical flasks, add 400 mg of petroleum to make the concentration 4000 mg / L, and finally sterilize at 121°C for 20 minutes.

[0052] Microbial degradation of petroleum: 7 g of the petroleum-degrading solid bacterial agent prepared in the present application was inoculated into the above-mentioned oil-containing culture medium, and microbial degradation was carried out at 45°C.

[0053] Determination of oil content: The oil-containing culture medium after 12 days of degradation was extracted and filtered, and the oil content was measured using an infrared oil meter. The degradation rate was 49.09%.

[0054] Example 4

[0055] Prepare oil-containing culture medium: add 0.5% NaCl, 0.15% K2HPO4, 0.3% KH2PO4, 0.01% MgSO4·7H2O and 0.001% CaCl2·2H2O to water to prepare liquid culture medium, divide 100 mL of liquid culture medium into several conical flasks, add 500 mg of petroleum to make the concentration 5 g / L, and finally sterilize at 121°C for 20 minutes.

[0056] Microbial degradation of petroleum: 7 g of the solid petroleum-degrading bacterial agent prepared in the present application was added to the above-mentioned oil-containing culture medium and subjected to microbial degradation at 40°C.

[0057] Determination of oil content: The oil-containing culture medium after 12 days of degradation was extracted and filtered, and the oil content was measured using an infrared oil meter. The degradation rate was 50.73%.

[0058] Example 5

[0059] Prepare oil-containing culture medium: add 0.5% NaCl, 0.15% K2HPO4, 0.3% KH2PO4, 0.01% MgSO4·7H2O and 0.001% CaCl2·2H2O to water to prepare liquid culture medium, divide 100 mL of liquid culture medium into several conical flasks, add 500 mg of petroleum to make the concentration 5 g / L, and finally sterilize at 121°C for 20 minutes.

[0060] Microbial degradation of petroleum: 7 g of the petroleum-degrading solid bacterial agent prepared in the present application was inoculated into the above-mentioned oil-containing culture medium, and microbial degradation was carried out at 45°C.

[0061] Determination of oil content: The oil-containing culture medium after 12 days of degradation was extracted and filtered, and the oil content was measured using an infrared oil meter. The degradation rate was 65.56%.

[0062] Example 6

[0063] Prepare oil-containing culture medium: add 0.5% NaCl, 0.15% K2HPO4, 0.3% KH2PO4, 0.01% MgSO4·7H2O and 0.001% CaCl2·2H2O to water to prepare liquid culture medium, divide 100 mL of liquid culture medium into several conical flasks, add 500 mg of petroleum to make the concentration 5 g / L, and finally sterilize at 121°C for 20 minutes.

[0064] Microbial degradation of petroleum: 7 g of the solid petroleum-degrading bacterial agent prepared in the present application was added to the above-mentioned oil-containing culture medium and subjected to microbial degradation at 50°C.

[0065] Determination of oil content: The oil-containing culture medium after 12 days of degradation was extracted and filtered, and the oil content was measured using an infrared oil meter. The degradation rate was 49.80%.

[0066] Example 7

[0067] Prepare oil-containing culture medium: add 0.5% NaCl, 0.15% K2HPO4, 0.3% KH2PO4, 0.01% MgSO4·7H2O and 0.001% CaCl2·2H2O to water to prepare liquid culture medium, divide 100 mL of liquid culture medium into several conical flasks, add 500 mg of petroleum to make the concentration 5 g / L, and finally sterilize at 121°C for 20 minutes.

[0068] Microbial degradation of petroleum: 5 g of the solid petroleum-degrading bacterial agent prepared in the present application was inoculated into the above-mentioned oil-containing culture medium and subjected to microbial degradation at 45°C.

[0069] Determination of oil content: The oil-containing culture medium after 12 days of degradation was extracted and filtered, and the oil content was measured using an infrared oil meter. The degradation rate was 51.84%.

[0070] Example 8

[0071] Prepare oil-containing culture medium: add 0.5% NaCl, 0.15% K2HPO4, 0.3% KH2PO4, 0.01% MgSO4·7H2O and 0.001% CaCl2·2H2O to water to prepare liquid culture medium, divide 100 mL of liquid culture medium into several conical flasks, add 500 mg of petroleum to make the concentration 5 g / L, and finally sterilize at 121°C for 20 minutes.

[0072] Microbial degradation of petroleum: 9 g of the solid petroleum-degrading bacterial agent prepared in the present application was inoculated into the above-mentioned oil-containing culture medium and subjected to microbial degradation at 45°C.

[0073] Determination of oil content: The oil-containing culture medium after 12 days of degradation was extracted and filtered, and the oil content was measured using an infrared oil meter. The degradation rate was 57.64%.

[0074] Example 9

[0075] Prepare oil-containing culture medium: add 0.5% NaCl, 0.15% K2HPO4, 0.3% KH2PO4, 0.01% MgSO4·7H2O and 0.001% CaCl2·2H2O to water to prepare liquid culture medium, divide 100 mL of liquid culture medium into several conical flasks, add 500 mg of petroleum to make the concentration 5 g / L, and finally sterilize at 121°C for 20 minutes.

[0076] Microbial degradation of petroleum: 7 g of the solid petroleum-degrading bacterial agent prepared in the present application was added to the above-mentioned oil-containing culture medium and subjected to microbial degradation at 45°C.

[0077] Determination of oil content: The oil-containing culture medium after 6 days of degradation was extracted and filtered, and the oil content was measured using an infrared oil meter. The degradation rate was 44.70%.

[0078] Example 10

[0079] Prepare oil-containing culture medium: add 0.5% NaCl, 0.15% K2HPO4, 0.3% KH2PO4, 0.01% MgSO4·7H2O and 0.001% CaCl2·2H2O to water to prepare liquid culture medium, divide 100 mL of liquid culture medium into several conical flasks, add 500 mg of petroleum to make the concentration 5 g / L, and finally sterilize at 121°C for 20 minutes.

[0080] Microbial degradation of petroleum: 7 g of the solid petroleum-degrading bacterial agent prepared in the present application was added to the above-mentioned oil-containing culture medium and subjected to microbial degradation at 45°C.

[0081] Determination of oil content: The oil-containing culture medium after 9 days of degradation was extracted and filtered, and the oil content was measured using an infrared oil measuring instrument. The degradation rate was 48.18%.

[0082] Examples 1 to 3 studied the degradation effects of the prepared solid bacterial agent for petroleum degradation at different petroleum hydrocarbon concentrations (oil content). Figure 3 As shown in the figure, the solid culture medium prepared from lees particles of different particle sizes has different culture effects on bacteria. Among them, when the petroleum hydrocarbon concentration is 500 mg / L, the degradation rate is the highest, up to 57.64%; when the petroleum hydrocarbon concentration is 4000 mg / L, the degradation rate is the lowest, at 49.09%. It can be seen that the lower the petroleum hydrocarbon concentration, the higher the degradation rate.

[0083] Examples 4 to 6 studied the degradation effects of the prepared petroleum-degrading solid bacterial agent at different degradation temperatures. Figure 4 As shown, under different degradation temperature conditions, the degradation rate of petroleum by microorganisms is different. At about 45°C, the degradation effect is the best, with a degradation rate of 65.56%.

[0084] Examples 5, 7 and 8 studied the degradation effects of the prepared solid petroleum-degrading bacterial agent at different inoculation amounts. Figure 5 As shown, the inoculation amount of the oil-degrading solid bacterial agent is different, and the oil degradation rate is different. From the experimental example, it can be seen that with the increase of the inoculation amount, the degradation rate first increases and then decreases. When the inoculation amount is 7%, the degradation rate is the highest, up to 65.02%, and the degradation effect is the best.

[0085] Examples 5, 9 and 10 studied the degradation effects of the prepared petroleum-degrading solid bacterial agent at different degradation times. Figure 6 As shown in the figure, the degradation rate of oil is different with different degradation time. With the increase of degradation time, the degradation rate first increases and then decreases. When the degradation time is 12 days, the degradation rate is the highest, up to 54.71%, and the degradation effect is the best.

[0086] The petroleum-degrading solid bacterial agent prepared in the present application has efficient biodegradation performance, and the vinasse can be used as a bulking agent to improve the air permeability and water permeability of the soil. At the same time, the petroleum-degrading solid bacterial agent can be added to the soil as a soil conditioner, and the nutrients in the bacterial agent can improve the content of trace elements in the soil. Through the resource utilization of the waste vinasse, not only the degradation of petroleum hydrocarbons in the soil is achieved, but also the physical and chemical properties of the soil itself are improved, and the soil reclamation rate is increased.

[0087] Although the present application provides method operation steps as described in the embodiments or flowcharts, more or fewer operation steps may be included based on conventional or non-creative labor. The order of steps listed in this embodiment is only one way of executing the order of many steps and does not represent the only execution order. When the actual device or client product is executed, it can be executed in sequence or in parallel according to the method shown in this embodiment or the accompanying drawings (for example, in a parallel processor or multi-threaded processing environment).

[0088] The various embodiments in this specification are described in a progressive manner, and the same or similar parts between the various embodiments can be referenced to each other. Each embodiment focuses on the differences from other embodiments.

[0089] The above embodiments are only used to illustrate the technical solutions of the present application, rather than to limit the present application. Although the present application has been described in detail with reference to the aforementioned embodiments, a person of ordinary skill in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some or all of the technical features thereof may be replaced by equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the present application.

Claims

1. A petroleum degradation solid bacterial agent, characterized in that: Including solid culture medium and composite bacterial liquid; The components of the solid culture medium and their weight fractions include: 100-600 parts of distiller's grains particles, 3-10 parts of nitrogen source, 5-10 parts of carbon source, 3-10 parts of inorganic salt, 1-3 parts of potassium dihydrogen phosphate, 2-6 parts of dipotassium hydrogen phosphate, 1-3 parts of magnesium sulfate heptahydrate and 50-100 parts of alkaline substance; The composite bacterial liquid is obtained by compounding Pseudomonas aeruginosa and Bacillus clausii, and the inoculation amount of the composite bacterial liquid is 1% to 10% of the mass of the solid culture medium.

2. The solid culture bacterial agent according to claim 1, characterized in that: The pH value of the petroleum-degrading solid bacterial agent is 7.

2.

3. A method for preparing a petroleum-degrading solid bacterial agent, characterized in that: include: The air-dried and sun-drying distiller's grains are crushed and the distiller's grains particles are screened out; The bacterial strains were inoculated into liquid fermentation medium respectively, and cultured at a constant temperature of 180 rpm and 37°C for 24 to 48 hours to obtain a single bacterial solution; The single bacterial solution is prepared in proportion to obtain a composite bacterial solution; Adding nutrients to the vinasse particles and adjusting the water content and pH value thereof to obtain a solid culture medium; The composite bacterial liquid is added to the solid culture medium, stirred evenly, and fermented and cultured at 30° C. to 40° C. for 48 h to 72 h to obtain a petroleum-degrading solid bacterial agent; wherein the inoculation amount of the composite bacterial liquid is 1% to 10% of the mass of the solid culture medium.

4. The method according to claim 3, characterized in that The lees include alcohol lees and / or white wine lees and / or yellow wine lees and / or beer lees.

5. The method according to claim 3, characterized in that: The particle size of the vinasse particles is 1 mm to 3 mm.

6. The method according to claim 3, characterized in that The nutrients include a nitrogen source, a carbon source and an inorganic salt; wherein the nitrogen source is yeast powder, the carbon source is molasses, and the inorganic salt is potassium dihydrogen phosphate, dipotassium hydrogen phosphate, and magnesium sulfate heptahydrate.

7. The method according to claim 3, characterized in that The step of adjusting the water content and pH value to obtain a solid culture medium comprises: Adding water to the lees particles and the nutrients to make the moisture content of the lees particles be 20%-50%; Add alkali to the lees particles and the nutrients to make the pH value of the lees particles be 5-9.

8. The method according to claim 3, characterized in that The bacterial species include Bacillus clausii and Pseudomonas aeruginosa, and the ratio of Pseudomonas aeruginosa to Bacillus clausii is 1:5-5:

1.

9. An application of a petroleum degradation solid bacterial agent, characterized in that: Used to treat petroleum pollutants.

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