Culture medium for producing special bacteria for petrochemical wastewater and preparation method of culture medium

Abstract

The invention discloses a culture medium for producing special bacteria for petrochemical wastewater and a preparation method of the culture medium. The culture medium comprises the following components in parts by weight: 30-85 parts of glucose, 10-40 parts of cane sugar, 8-20 parts of monopotassium phosphate, 4-10 parts of magnesium sulfate, 6-18 parts of sodium bicarbonate, 20-39 parts of peptone, 13-30 parts of urea, 11-26 parts of yeast extract and 10000-20000 parts of distilled water. Compared with a conventional culture medium, the optimal culture medium and the production process provided by the invention are relatively low in cost, have the effect of promoting the concentration and the growth speed of the special bacteria of the petrochemical wastewater, and are remarkable in effect.

Description

Technical Field

[0001] The invention relates to microbial fermentation engineering technology, in particular to a culture medium for producing petrochemical wastewater-specific bacteria and a preparation method thereof. Background Art

[0002] The composition of petrochemical wastewater is extremely complex. It contains not only a variety of difficult-to-degrade organic pollutants, such as aromatic hydrocarbons and polycyclic aromatic hydrocarbons, but also various inorganic substances and heavy metals. Traditional physical and chemical treatment methods are often difficult to achieve ideal wastewater treatment results due to their extremely high costs and limited treatment effects when dealing with petrochemical wastewater. For this reason, making full use of the biodegradation effect of microorganisms has gradually become one of the important means of treating petrochemical wastewater.

[0003] According to the characteristics of petrochemical wastewater, in the process of developing specialized bacteria, researchers screened microorganisms with specific degradation capabilities from nature, or modified existing strains through genetic engineering technology to give them new degradation functions. Under suitable conditions, these specialized bacteria can rapidly reproduce and degrade pollutants in wastewater, converting them into harmless substances, thereby achieving the purpose of purifying wastewater.

[0004] However, the existing culture medium formulas often fail to fully consider the unique needs of petrochemical wastewater-specific bacteria. This results in the bacteria growing slowly or their metabolic activity being low, which greatly affects the overall wastewater treatment efficiency. In order to ensure that these exclusive bacteria can work effectively in the treatment system, it is necessary to provide them with the most suitable growth conditions. As one of the key factors to ensure the normal growth and reproduction of exclusive bacteria, optimize treatment performance and improve wastewater treatment effects, the selection of appropriate culture medium is of great significance in the field of petrochemical wastewater-specific bacteria cultivation. Summary of the invention

[0005] 1. Technical issues to be resolved

[0006] The purpose of the present invention is to provide a culture medium for producing petrochemical wastewater-specific bacteria and a preparation method thereof, so as to solve the technical problem that the existing culture medium has low culture efficiency for petrochemical wastewater-specific bacteria.

[0007] (II) Technical solution

[0008] In the first aspect, the present invention provides a culture medium for the production of petrochemical wastewater-specific bacteria, which comprises, by weight: 30-85 parts of glucose, 10-40 parts of sucrose, 8-20 parts of potassium dihydrogen phosphate, 4-10 parts of magnesium sulfate, 6-18 parts of sodium bicarbonate, 20-39 parts of peptone, 13-30 parts of urea, 11-26 parts of yeast extract, and 10,000-20,000 parts of distilled water.

[0009] Furthermore, the invention comprises, by weight: 42-70 parts of glucose, 18-30 parts of sucrose, 9-18 parts of potassium dihydrogen phosphate, 6-10 parts of magnesium sulfate, 8-16 parts of sodium bicarbonate, 24-34 parts of peptone, 17-26 parts of urea, 14-22 parts of yeast extract, and 13,000-18,000 parts of distilled water.

[0010] Furthermore, the invention comprises, by weight: 50-62 parts of glucose, 22-26 parts of sucrose, 12-15 parts of potassium dihydrogen phosphate, 6-8 parts of magnesium sulfate, 11-14 parts of sodium bicarbonate, 28-32 parts of peptone, 20-24 parts of urea, 16-20 parts of yeast extract, and 16400-17900 parts of distilled water.

[0011] Preferably, the mixture comprises, by weight: 50 parts of glucose, 25 parts of sucrose, 14 parts of potassium dihydrogen phosphate, 7 parts of magnesium sulfate, 12 parts of sodium bicarbonate, 30 parts of peptone, 22 parts of urea, 16 parts of yeast extract, and 17,000 parts of distilled water.

[0012] In a second aspect, the present invention also provides a method for preparing the above culture medium, comprising the following steps:

[0013] S100: preparing raw materials; weighing each raw material according to weight;

[0014] S200: preparing a culture medium; adding various raw materials into a glass container, heating to 90-100° C., and stirring evenly until completely dissolved;

[0015] S300: adjusting pH: adjusting the pH of the culture medium obtained in step S200;

[0016] S400: high temperature sterilization: sterilize the culture medium obtained in step S300 by high pressure steam sterilization process.

[0017] (III) Beneficial effects

[0018] Construction of a multi-dimensional nutrient supply system: The present invention creatively uses sucrose to partially replace glucose and urea to partially replace peptone to construct a diversified carbon and nitrogen source supply system. This innovation not only enriches the types of nutrients available in the culture medium, but also effectively simulates the real chemical composition of petrochemical wastewater, providing more suitable growth conditions for exclusive strains. Experimental results show that compared with traditional single or limited combination of nutrient sources, this diversified nutrient supply system can better support the rapid proliferation and long-term stable growth of exclusive strains.

[0019] Enhanced microbial vitality and survival rate: Based on the culture medium formula designed by the present invention, the cultured petrochemical wastewater-specific bacteria show biological characteristics that are significantly better than those of the strains cultured in conventional culture medium. Specifically, after the same storage period, the exclusive bacteria in the culture medium of the present invention maintain higher cell vitality and quantity. This advantage is crucial to ensure the effectiveness and durability of the exclusive strains in practical applications, especially when treating complex and challenging petrochemical wastewater.

[0020] It should be understood that within the scope of the present invention, the above-mentioned various technical features of the present invention and the various technical features specifically described in the following embodiments can be combined with each other to form a new or preferred technical solution. Due to space limitations, they will not be described one by one here. DETAILED DESCRIPTION

[0021] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention is clearly and completely described. 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.

[0022] The petrochemical wastewater-specific bacteria referred to in the present invention can be selected from one of the following strains:

[0023] Pseudomonas

[0024] It has strong metabolic diversity and can use a variety of organic compounds as carbon sources and energy sources. It can degrade aromatic hydrocarbons in petrochemical wastewater, such as benzene, toluene, xylene, etc. Pseudomonas can gradually oxidize and decompose these complex aromatic hydrocarbon molecules by producing specific enzymes, such as monooxygenase and dioxygenase.

[0025] Rhodococcus

[0026] It has good degradation ability for long-chain alkanes, polycyclic aromatic hydrocarbons and other difficult-to-degrade organic matter in petrochemical wastewater. It can use enzymes such as cytochrome P450 monooxygenase in cells to oxidize the complex structure of polycyclic aromatic hydrocarbons, and then gradually decompose these long-chain organic matter through subsequent metabolic steps, such as β-oxidation.

[0027] Bacillus

[0028] In petrochemical wastewater treatment, Bacillus can degrade organic pollutants in wastewater, such as oils and fats. It can secrete a variety of extracellular enzymes, which can decompose oils and fats into small molecules such as fatty acids and glycerol, and then further metabolize and decompose these small molecules.

[0029] Acinetobacter

[0030] It can degrade some organic compounds such as short-chain hydrocarbons and alcohols in wastewater. It can convert these organic compounds into harmless substances through metabolic processes such as redox reactions.

[0031] Those skilled in the art may also apply the culture medium disclosed in the present invention to other known bacterial species. Furthermore, those skilled in the art may also tentatively apply the culture medium disclosed in the present invention to the cultivation of any bacterial species.

[0032] The invention discloses a culture medium for producing petrochemical wastewater-specific bacteria. The culture medium comprises, by weight, 30-85 parts of glucose, 10-40 parts of sucrose, 8-20 parts of potassium dihydrogen phosphate, 4-10 parts of magnesium sulfate, 6-18 parts of sodium bicarbonate, 20-39 parts of peptone, 13-30 parts of urea, 11-26 parts of yeast extract and 10000-20000 parts of distilled water.

[0033] Preferably, the mixture comprises, by weight: 42-70 parts of glucose, 18-30 parts of sucrose, 9-18 parts of potassium dihydrogen phosphate, 6-10 parts of magnesium sulfate, 8-16 parts of sodium bicarbonate, 24-34 parts of peptone, 17-26 parts of urea, 14-22 parts of yeast extract, and 13,000-18,000 parts of distilled water.

[0034] More preferably, the composition comprises, by weight: 50-62 parts of glucose, 22-26 parts of sucrose, 12-15 parts of potassium dihydrogen phosphate, 6-8 parts of magnesium sulfate, 11-14 parts of sodium bicarbonate, 28-32 parts of peptone, 20-24 parts of urea, 16-20 parts of yeast extract, and 16400-17900 parts of distilled water.

[0035] In an optional embodiment, the glucose is glucose monohydrate, the sucrose is food grade sucrose, and the peptone is tryptone.

[0036] The present invention also discloses a method for preparing the culture medium, comprising the following steps:

[0037] S100: preparing raw materials; weighing each raw material according to weight;

[0038] S200: preparing a culture medium; adding various raw materials into a glass container, heating to 90-100° C., and stirring evenly until completely dissolved;

[0039] S300: adjusting pH: adjusting the pH of the culture medium obtained in step S200;

[0040] S400: high temperature sterilization: sterilize the culture medium obtained in step S300 by high pressure steam sterilization process.

[0041] Preferably, in step S300, the pH is adjusted to 7.0-7.5.

[0042] According to the above formula and preparation method, the culture medium was prepared, and the results were as follows:

[0043] Embodiment 1:

[0044] A culture medium formula for petrochemical wastewater-specific bacteria comprises the following raw materials in proportion by weight: 50 parts of glucose, 25 parts of sucrose, 14 parts of potassium dihydrogen phosphate, 7 parts of magnesium sulfate, 12 parts of sodium bicarbonate, 30 parts of peptone, 22 parts of urea, 16 parts of yeast extract, and 17,000 parts of distilled water.

[0045] A method for preparing a culture medium formula for petrochemical wastewater-specific bacteria comprises the following steps:

[0046] Step S100: prepare raw materials: weigh each raw material according to its weight;

[0047] Step S200: preparing a culture medium: adding each raw material into a glass container, heating to 90-100° C., and stirring evenly until completely dissolved;

[0048] Step S300: adjusting pH: adjusting the pH of the culture medium obtained in step S200 to 7.0;

[0049] Step S400 high temperature sterilization: sterilize the culture medium obtained in step S300 by high pressure steam sterilization process at a pressure of 0.11-0.13 MPa, 121° C., for 30 minutes.

[0050] The petrochemical wastewater-specific bacteria are inoculated into the prepared culture medium for cultivation, and the culture temperature is controlled at 30-35°C.

[0051] Example 2

[0052] A culture medium formula for petrochemical wastewater-specific bacteria comprises the following raw materials in proportion by weight: 30 parts of glucose, 40 parts of sucrose, 20 parts of potassium dihydrogen phosphate, 4 parts of magnesium sulfate, 18 parts of sodium bicarbonate, 20 parts of peptone, 26 parts of urea, 11 parts of yeast extract, and 10,000 parts of distilled water.

[0053] The preparation method is the same as that in Example 1, and the method for culturing petrochemical wastewater-specific bacteria is the same as that in Example 1.

[0054] Example 3

[0055] A culture medium formula for petrochemical wastewater-specific bacteria comprises the following raw materials in proportion by weight: 85 parts of glucose, 10 parts of sucrose, 8 parts of potassium dihydrogen phosphate, 10 parts of magnesium sulfate, 6 parts of sodium bicarbonate, 39 parts of peptone, 13 parts of urea, 26 parts of yeast extract, and 20,000 parts of distilled water.

[0056] The preparation method is the same as that in Example 1, and the method for culturing petrochemical wastewater-specific bacteria is the same as that in Example 1.

[0057] Example 4

[0058] A culture medium formula for petrochemical wastewater-specific bacteria comprises the following raw materials in proportion by weight: 42 parts of glucose, 30 parts of sucrose, 18 parts of potassium dihydrogen phosphate, 6 parts of magnesium sulfate, 16 parts of sodium bicarbonate, 24 parts of peptone, 26 parts of urea, 14 parts of yeast extract, and 13,000 parts of distilled water.

[0059] The preparation method is the same as that in Example 1, and the method for culturing petrochemical wastewater-specific bacteria is the same as that in Example 1.

[0060] Example 5

[0061] A culture medium formula for petrochemical wastewater-specific bacteria comprises the following raw materials in proportion by weight: 70 parts of glucose, 18 parts of sucrose, 9 parts of potassium dihydrogen phosphate, 10 parts of magnesium sulfate, 8 parts of sodium bicarbonate, 34 parts of peptone, 17 parts of urea, 22 parts of yeast extract, and 18,000 parts of distilled water.

[0062] The preparation method is the same as that in Example 1, and the method for culturing petrochemical wastewater-specific bacteria is the same as that in Example 1.

[0063] Example 6

[0064] A culture medium formula for petrochemical wastewater-specific bacteria comprises the following raw materials in proportion by weight: 50 parts of glucose, 22 parts of sucrose, 15 parts of potassium dihydrogen phosphate, 6 parts of magnesium sulfate, 14 parts of sodium bicarbonate, 28 parts of peptone, 24 parts of urea, 20 parts of yeast extract, and 16,400 parts of distilled water.

[0065] The preparation method is the same as that in Example 1, and the method for culturing petrochemical wastewater-specific bacteria is the same as that in Example 1.

[0066] Example 7

[0067] A culture medium formula for petrochemical wastewater-specific bacteria comprises the following raw materials in proportion by weight: 62 parts of glucose, 26 parts of sucrose, 12 parts of potassium dihydrogen phosphate, 8 parts of magnesium sulfate, 11 parts of sodium bicarbonate, 32 parts of peptone, 20 parts of urea, 14 parts of yeast extract, and 17,900 parts of distilled water.

[0068] The preparation method is the same as that in Example 1, and the method for culturing petrochemical wastewater-specific bacteria is the same as that in Example 1.

[0069] Subjects:

[0070] Exclusive species: Pseudomonas

[0071] Controlled Experimental Design

[0072] Experimental groups: The culture media with different formulations described in Examples 1-7 were used.

[0073] Control group: Use traditional petrochemical wastewater-specific bacterial culture medium, the ingredients include 50 parts of glucose, 10 parts of potassium dihydrogen phosphate, 5 parts of magnesium sulfate, 10 parts of sodium bicarbonate, 25 parts of peptone, 15 parts of yeast extract, and 15,000 parts of distilled water.

[0074] Evaluation indicators

[0075] Bacterial density (OD600): evaluate the growth efficiency of Pseudomonas.

[0076] COD removal efficiency (%): Evaluate the ability of Pseudomonas to degrade organic pollutants in petrochemical wastewater.

[0077] Viability and viable count (CFU / mL): Assess the activity and viability of Pseudomonas.

[0078] Control experiment results

[0079]

[0080]

[0081] Results Analysis

[0082] From the data in the table, it can be seen that compared with the traditional culture medium, all the new formula culture mediums showed higher cell density, better COD removal rate, and higher vitality and survival number. In particular, Example 1 showed the best results in all three evaluation indicators:

[0083] Bacterial density: reached 1.4 OD600, 75% higher than the control group, indicating that the formula can significantly promote the growth of Pseudomonas.

[0084] COD removal rate: reached 88%, 23 percentage points higher than the control group, indicating that the culture medium of Example 1 can greatly improve the degradation ability of Pseudomonas to organic pollutants in petrochemical wastewater.

[0085] Viability and survival number: reached 5.5×10^7CFU / mL, much higher than the 3.0×10^7CFU / mL of the control group, showing the superiority of this culture medium in maintaining and enhancing the activity of Pseudomonas.

[0086] In summary, the culture medium formula provided in Example 1 is the most effective for the cultivation of Pseudomonas and can be used as a preferred solution in the application of petrochemical wastewater treatment. This not only improves the growth efficiency of the exclusive bacteria, but also enhances its ability to degrade pollutants in the wastewater, thereby improving the overall wastewater treatment effect.

[0087] It should be noted that, in this article, relational terms such as first and second, etc. are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Moreover, the terms "include", "comprises" or any other variations thereof are intended to cover non-exclusive inclusion, so that a process, method, article or device that includes a series of elements includes not only those elements, but also other elements that are not explicitly listed, or also includes elements that are inherent to such process, method, article or device.

[0088] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that not every embodiment includes only one independent technical solution, and in the absence of a conflict of solutions, the various technical features mentioned in each embodiment can be combined in any manner to form other implementation methods that can be understood by those skilled in the art.

[0089] In addition, without departing from the scope of the present invention, the technical solutions described in the aforementioned embodiments may be modified, or some of the technical features may be replaced by equivalents, without causing the essence of the corresponding technical solutions to depart from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A culture medium for producing bacteria specific for petrochemical wastewater, comprising by weight: 30-85 parts of glucose, 10-40 parts of sucrose, 8-20 parts of potassium dihydrogen phosphate, 4-10 parts of magnesium sulfate, 6-18 parts of sodium bicarbonate, 20-39 parts of peptone, 13-30 parts of urea, 11-26 parts of yeast extract, and 10,000 parts to 20,000 parts of distilled water.

2. A culture medium for producing bacteria specific for petrochemical wastewater according to claim 1, characterized in that: The invention comprises, by weight: 42-70 parts of glucose, 18-30 parts of sucrose, 9-18 parts of potassium dihydrogen phosphate, 6-10 parts of magnesium sulfate, 8-16 parts of sodium bicarbonate, 24-34 parts of peptone, 17-26 parts of urea, 14-22 parts of yeast extract and 13000-18000 parts of distilled water.

3. A culture medium for producing bacteria specific for petrochemical wastewater according to claim 1, characterized in that: The invention comprises, by weight: 50-62 parts of glucose, 22-26 parts of sucrose, 12-15 parts of potassium dihydrogen phosphate, 6-8 parts of magnesium sulfate, 11-14 parts of sodium bicarbonate, 28-32 parts of peptone, 20-24 parts of urea, 16-20 parts of yeast extract and 16400-17900 parts of distilled water.

4. A culture medium for producing bacteria specific for petrochemical wastewater according to claim 1, characterized in that: The ingredients include, by weight: 50 parts of glucose, 25 parts of sucrose, 14 parts of potassium dihydrogen phosphate, 7 parts of magnesium sulfate, 12 parts of sodium bicarbonate, 30 parts of peptone, 22 parts of urea, 16 parts of yeast extract and 17,000 parts of distilled water.

5. A culture medium for producing bacteria specific for petrochemical wastewater according to any one of claims 1 to 4, characterized in that: The glucose is glucose monohydrate.

6. A culture medium for producing bacteria specific for petrochemical wastewater according to any one of claims 1 to 4, characterized in that: The sucrose is edible grade sucrose.

7. A culture medium for producing bacteria specific for petrochemical wastewater according to any one of claims 1 to 4, characterized in that: The peptone is tryptone.

8. The method for preparing the culture medium according to any one of claims 1 to 4, comprising the steps of: S100: preparing raw materials; weighing each raw material according to weight; S200: preparing a culture medium; adding various raw materials into a glass container, heating to 90-100° C., and stirring evenly until completely dissolved; S300: adjusting pH: adjusting the pH of the culture medium obtained in step S200; S400: high temperature sterilization: sterilize the culture medium obtained in step S300 by high pressure steam sterilization process.

9. The preparation method according to claim 8, characterized in that: In step S300, the pH is adjusted to 7.0-7.5.

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