Method for improving biological manganese oxidation performance of manganese oxidizing bacteria and application

Abstract

The invention provides a method for improving the biological manganese oxidation performance of manganese oxidizing bacteria and application, and belongs to the technical field of sewage treatment. According to the method, the biological manganese oxidation performance of the manganese oxidizing bacteria is improved by adding the biochar, and after the biochar is added into the manganese oxidizing bacteria, the divalent manganese is added, so that the oxidation rate of the divalent manganese can be obviously improved, and the removal efficiency of heavy metals in the wastewater is improved. Wherein the biochar can be used as an electron shuttle, so that the transfer capability of electrons can be improved, and the oxidation rate of divalent manganese can be increased; the biochar also improves the gene abundance and expression ability of the manganese-oxidizing bacteria and the extracellular superoxide concentration, improves the biological activity of the manganese-oxidizing bacteria, and accelerates the oxidation of divalent manganese. Moreover, the biochar is wide in source and low in cost, resource recycling can be achieved, meanwhile, the treatment cost is reduced, and the biochar is easy to prepare and put and convenient to use in practical application.

Description

Technical Field

[0001] The invention relates to the technical field of sewage treatment, and in particular to a method for improving the biological manganese oxidation performance of manganese oxidizing bacteria and its application. Background Art

[0002] Manganese oxidizing bacteria (MnOB) are a class of microorganisms that can rapidly catalyze the oxidation of Mn(II) and simultaneously induce the formation of biological manganese oxides (BMO). Under neutral, room temperature, and pressure conditions, manganese oxidizing bacteria can induce the production of biological manganese oxides through biosynthesis. Compared with chemically synthesized manganese oxides, biosynthesized BMO has smaller particle size, weaker crystallinity, higher Mn valence, more holes in the octahedral structure, larger specific surface area, and stronger reaction activity. Under natural conditions, the biological oxidation rate of divalent manganese by manganese oxidizing bacteria is 10 times that of chemical oxidation. 5 In addition, due to its special crystal structure and surface chemical properties, biological manganese oxide has a strong adsorption capacity for a variety of heavy metal ions, and can also adsorb and degrade some organic pollutants, such as pesticides, endocrine disruptors, antibiotics, polycyclic aromatic hydrocarbons, etc. Therefore, manganese oxidizing bacteria play an important role in the field of polluted environment remediation.

[0003] In the process of manganese oxidizing bacteria inducing biological manganese oxides, biological manganese oxidation performance plays a vital role, which can be reflected by the oxidation rate of divalent manganese and the generation rate of biological manganese oxides. Many environmental factors will affect the biological manganese oxidation performance, such as DO, pH, Eh, temperature, divalent manganese concentration, strain type, etc. In general, the faster the biological manganese oxidation rate, the higher the biological manganese oxide, and the more conducive to the remediation of pollutants.

[0004] At present, there are two main ways to improve the performance of biological manganese oxides. First, add stimulating factors to promote the manganese oxidation reaction, accelerate the oxidation rate of divalent manganese and the generation rate of manganese oxides. Second, add a carrier to increase the contact area and improve the physical structure of the reaction system, thereby improving the manganese oxidation performance. However, the first method of improving the biological manganese oxidation performance by adding stimulating factors is difficult to implement in actual remediation projects, and the improvement is very limited. Most of the second method is used. Therefore, it is urgent to develop a green, low-cost, and secondary pollution-free method to improve the biological manganese oxidation performance. Summary of the invention

[0005] The purpose of the present invention is to provide a method for improving the biological manganese oxidation performance of manganese oxidizing bacteria and its application, which can significantly increase the oxidation rate of divalent manganese and improve the removal efficiency of heavy metals in wastewater.

[0006] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0007] The present invention provides a method for improving the biological manganese oxidation performance of manganese oxidizing bacteria, comprising the following steps:

[0008] (1) After pretreatment, the straw is treated at high temperature in a CO2 atmosphere, and then cooled, ground, and sieved to obtain black powder, and the black powder is treated with acid, washed, and dried to obtain biochar powder;

[0009] (2) inoculating manganese oxidizing bacteria into a manganese oxidizing bacteria culture medium, then adding the biochar from step (1) into the manganese oxidizing bacteria culture medium for cultivation, and adding a divalent manganese solution after the cultivation is completed.

[0010] Preferably, in step (1), the straw pretreatment method is:

[0011] The straw is dried at 60-80°C to reduce the moisture content of the straw to 10-15%, and is crushed to a straw length of 1-5 cm.

[0012] Preferably, in step (1), the temperature of the high temperature treatment is 300-600° C., and the time is 7-12 hours; and the mesh size of the sieving is 60-300 meshes.

[0013] Preferably, in step (1), the step of acid treatment of the black powder is:

[0014] The black powder is added to a 0.2-1.0 mol / L hydrochloric acid solution and stirred for 8-24 hours; the mass volume ratio of the black powder to the hydrochloric acid solution is 1 g:10-20 mL.

[0015] Preferably, in step (2), the manganese oxidizing bacteria is P. putida strain MnB1; the concentration of the manganese oxidizing bacteria used for inoculation is 1 to 3×10 7 CFU / ml, the inoculation amount of the manganese oxidizing bacteria is 2-10%, the pH of the manganese oxidizing bacteria culture medium is 6.0-7.5; the addition amount of the biochar is 1.0-10.0 g / L, the culture time is 1-2 days; and the divalent manganese concentration is 2.5-12.0 mg / L.

[0016] The present invention also provides an application of a method for improving the biological manganese oxidation performance of manganese oxidizing bacteria in purifying sewage, wherein the application method is:

[0017] Manganese oxidizing bacteria were inoculated into sewage, biochar was added for cultivation, and then divalent manganese solution was added.

[0018] Preferably, the pH of the sewage is 6.0-7.5; the inoculation amount of manganese oxidizing bacteria in the sewage is 2-10%, and the concentration of the manganese oxidizing bacteria is 1-3×10 7CFU / ml; the dosage of the biochar is 1.0-10.0 g / L, the culture time is 1-2 days; and the divalent manganese concentration is 5-12.0 mg / L.

[0019] The beneficial effects of the present invention compared with the prior art are:

[0020] (1) The present invention improves the biological manganese oxidation performance of manganese oxidizing bacteria by adding biochar. After adding biochar to manganese oxidizing bacteria, adding divalent manganese can significantly increase the oxidation rate of divalent manganese and improve the removal efficiency of heavy metals in wastewater. Among them, biochar can act as an electron shuttle, which can improve the electron transfer ability and accelerate the oxidation rate of divalent manganese; biochar also increases the gene abundance and expression ability of manganese oxidizing bacteria and the extracellular superoxide concentration, improves the biological activity of manganese oxidizing bacteria, and accelerates the oxidation of divalent manganese.

[0021] (2) As a green and environmentally friendly material, biochar will not cause secondary pollution to the soil, water and air during use. In addition, biochar is widely available and low-cost, which can achieve resource reuse while reducing processing costs. It is also easy to prepare and put into use, making it convenient for use in practical applications.

[0022] (3) Compared with free bacteria, the addition of biochar provides a biological carrier for manganese oxidizing bacteria, which helps the attachment and growth of manganese oxidizing bacteria and creates a good living environment for manganese oxidizing bacteria. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative labor.

[0024] Figure 1 This is a scanning electron microscope image of the biochar in Example 1 of the present invention;

[0025] Figure 2 This is a scanning electron microscope image of the biochar after bacterial colony attachment in Example 1 of the present invention. DETAILED DESCRIPTION

[0026] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0027] It should be understood that the terms described in the present invention are only for describing special embodiments and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. Each smaller range between the intermediate value in any stated value or stated range and any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0028] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0029] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to the skilled artisan. The present invention description and examples are exemplary only.

[0030] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0031] A method for improving the biological manganese oxidation performance of manganese oxidizing bacteria comprises the following steps:

[0032] (1) After pretreatment, the straw is treated at high temperature in a CO2 atmosphere, and then cooled, ground, and sieved to obtain black powder, and the black powder is treated with acid, washed, and dried to obtain biochar powder;

[0033] (2) inoculating manganese oxidizing bacteria into a manganese oxidizing bacteria culture medium, then adding the biochar from step (1) into the manganese oxidizing bacteria culture medium for cultivation, and adding a divalent manganese solution after the cultivation is completed.

[0034] Example 1

[0035] Example 1 of the present invention provides a method for improving the biological manganese oxidation performance of manganese oxidizing bacteria, and the specific steps are as follows:

[0036] (1) Preparation of Pseudomonas putida strain MnB1 bacterial solution:

[0037] Pseudomonas putida strain MnB1 was inoculated into a Pseudomonas putida enrichment medium at a volume ratio of 3%, and then aerobically enriched and cultured at 30°C with horizontal shaking (150 rpm) for 2 days to obtain a Pseudomonas putida strain MnB1 bacterial solution with a bacterial solution concentration of 1×10 7 CFU / ml.

[0038] The culture medium composition is: yeast extract powder, 0.5g / L; acid hydrolyzed casein, 0.5g / L; glucose, 1g / L; calcium chloride dihydrate, 0.29g / L; magnesium sulfate heptahydrate, 0.82g / L; ferric chloride, 0.001g / L; trace elements, 1mL; pH = 7.0. The trace elements are: copper sulfate pentahydrate, 2.49g / L; zinc sulfate heptahydrate, 12.65g / L; cobalt chloride hexahydrate, 4.76g / L; sodium molybdate dihydrate, 3.15g / L.

[0039] The selected Pseudomonas putida strain MnB1 was purchased from the American Type Culture Collection (ATCC) with the serial number 23483.

[0040] (2) Preparation of biochar:

[0041] The straw was dried at 80°C to reduce the moisture content of the straw to 15%, and then pulverized to a straw length of 5 cm.

[0042] 100g of crushed straw was put into a pyrolysis furnace, CO2 protective gas was introduced, the temperature was raised to 600℃, and the temperature was kept constant for 10h, cooled, ground, and passed through a 300-mesh sieve to obtain a black powder; the black powder prepared above was added to a 1.0mol / L hydrochloric acid solution at a mass volume ratio of 1g:10mL, stirred for 24h, filtered, washed with 300ml deionized water to a pH of 8.0, and freeze-dried in a freeze dryer for 24h to obtain biochar powder; its scanning electron microscope image is as shown in Figure 1 shown.

[0043] Figure 1 It shows that the addition of biochar provides a larger specific surface area for manganese oxidizing bacteria, facilitates the attachment of microorganisms, and provides a good environment for the growth of microorganisms.

[0044] (3) Improve the biological manganese oxidation performance of manganese oxidizing bacteria:

[0045] After sterilizing and cooling 250 ml of ATCC279# culture medium system, HEPES buffer was added to adjust the pH to 7.2, and the manganese oxidizing bacteria cultured in step 1) were added at an inoculation ratio of 3%. The amount of biochar prepared in step 2) was 5.0 g / L, and the culture was continued for 2 days (after adding biochar, the colonies attached to the surface of the culture medium were as follows). Figure 2As shown), divalent manganese was added to a final concentration of 12 mg / L, and the mixture was shaken and cultured at 30 degrees and 120 rpm in a vertical full-temperature shaking incubator.

[0046] Example 2

[0047] In Example 2 of the present invention, the method of step (3) of Example 1 was used to detect the manganese oxidation kinetic rate constant and the BMO generation rate under different biochar addition conditions. The difference from Example 1 is that the biochar addition amounts in Example 2 are 0 g / L, 1.0 g / L, 2.5 g / L, 5.0 g / L, and 10.0 g / L, respectively. The culture was shaken in a vertical full-temperature shaking incubator at 30 degrees and 120 rpm for 48 hours. Samples were taken at 4 hours, 8 hours, 12 hours, 24 hours, 36 hours, and 48 hours, respectively, and the divalent manganese and BMO concentrations were determined. The divalent manganese oxidation kinetic rate constant and the BMO generation rate were calculated. The results are shown in Table 1.

[0048] Table 1 Kinetic rate constants of manganese oxidation and BMO generation under different biochar addition conditions

[0049]

[0050] As shown in Table 1, compared with the control group, when biochar was added, the oxidation rate of divalent manganese increased and the production rate of BMO also increased. When the biochar addition amount was 5.0 g / L, the divalent manganese oxidation rate constant was 0.23 mg·L -1 ·h -1 , the BMO production rate is 1.25mg(MnO2)·L -1 ·h -1 , which were increased by 110% and 205% respectively compared with the control group.

[0051] Example 3

[0052] In Example 3 of the present invention, the method of step (3) of Example 1 was used to detect the manganese oxidation kinetic rate and the BMO generation rate under different divalent manganese concentration addition conditions. Different from Example 1, the final divalent manganese concentrations in Example 3 were 2.5 g / L, 5.0 g / L, 10.0 g / L, and 12.0 g / L, respectively. The results are shown in Table 2.

[0053] Table 2 Manganese oxidation kinetic rate and BMO generation rate under different divalent manganese concentration addition conditions

[0054]

[0055] It can be seen from Table 2 that as Mn 2+ As the concentration of Mn increases, the oxidation rate constant of divalent manganese increases, and the production rate of BMO also increases; when Mn 2+When the final concentration was 12.0 mg / L, the divalent manganese oxidation rate constant and BMO generation rate reached the maximum, and the divalent manganese oxidation rate constant was 0.47 mg·L -1 ·h -1 , the BMO generation rate is 1.24 mg (MnO2) L -1 ·h -1 ; and Mn 2+ Compared with the final concentration of 2.5 mg / L, the divalent manganese oxidation rate constant increased by 11 times and the BMO generation rate increased by 5 times.

[0056] Example 4

[0057] In Example 4 of the present invention, the method of step (3) of Example 1 was used to detect the constant rate of manganese oxidation kinetics and the rate of BMO formation under different pH conditions. The difference from Example 1 is that the pH in Example 4 is 6.0, 7.2, and 7.6, respectively. The results are shown in Table 3.

[0058] Table 3 Manganese oxidation kinetic rate and BMO generation rate under different pH conditions

[0059]

[0060]

[0061] It can be seen from Table 3 that within a certain range, as the pH increases, the divalent manganese oxidation rate constant also increases, and the BMO generation rate also increases; when the pH is 7.2, the divalent manganese oxidation rate constant and the BMO generation rate reach the maximum, and the divalent manganese oxidation rate constant is 0.50 mg·L -1 ·h -1 , the BMO generation rate is 0.87 mg (MnO2) L -1 ·h -1 .

[0062] Example 5

[0063] Example 5 of the present invention refers to the method of Example 1 to detect the Cd removal efficiency of Cd-containing wastewater, and the specific method is as follows:

[0064] Take 1L of Cd-containing wastewater with an initial concentration of 1.0 mg / L, adjust the pH of the wastewater to 7.2, add manganese oxidizing bacteria strain P. putida strain MnB1 at an inoculation ratio of 3%, add biochar at 5.0 g / L, and culture for 2 days; then add divalent manganese to a final concentration of 12 mg / L, place it in an incubator, and culture it at 30 degrees and 140 rpm for 48 hours. The Cd concentration is measured regularly at different time periods, and the Cd removal efficiency is calculated. The results are shown in Table 4.

[0065] Table 4 Cd removal rate

[0066]

[0067] It can be seen from Table 4 that with the change of time, the Cd removal rate gradually increases, and the removal effect is better; within the first 12 hours, the removal rate has reached 74.52%, indicating that the removal of Cd has made rapid progress during this period; within 24h-48h, the removal rate continues to increase, but the growth rate slows down; the removal rate reaches a maximum of 96.75% in the 48th hour, an increase of 22.23% compared with the 12th hour.

[0068] Example 6

[0069] Example 6 of the present invention adopts the method of Example 5 to detect the Cd removal efficiency of Cd-containing wastewater. The difference from Example 5 is that the concentration of Cd-containing wastewater in Example 6 is 2.0 mg / L. The results are shown in Table 5.

[0070] Table 5 Cd removal rate

[0071]

[0072] It can be seen from Table 5 that with the change of time, the Cd removal rate gradually increases, and the removal effect is better; within the first 12 hours, the removal rate has reached 70.36%, indicating that the removal of Cd has made rapid progress during this period; within 24h-48h, the removal rate continues to increase, but the growth rate slows down; the removal rate reaches a maximum of 92.58% in the 48th hour, an increase of 22.22% compared with the 12th hour.

[0073] Example 7

[0074] Example 7 of the present invention adopts the method of Example 5 to detect the Cd removal efficiency of Cd-containing wastewater. The difference from Example 5 is that the concentration of Cd-containing wastewater in Example 7 is 4.0 mg / L. The results are shown in Table 6.

[0075] Table 6 Cd removal rate

[0076]

[0077] It can be seen from Table 6 that with the change of time, the Cd removal rate gradually increases, and the removal effect is better; within the first 12 hours, the removal rate has reached 70.39%, indicating that the removal of Cd has made rapid progress during this period; within 24h-48h, the removal rate continues to increase, but the growth rate slows down; the removal rate reaches a maximum of 93.76% in the 48th hour, an increase of 23.37% compared with the 12th hour.

[0078] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for improving the biological manganese oxidation performance of manganese oxidizing bacteria, characterized in that: The steps include: (1) After pretreatment, the straw is treated at high temperature in a CO2 atmosphere, and then cooled, ground, and sieved to obtain black powder, and the black powder is treated with acid, washed, and dried to obtain biochar powder; (2) inoculating manganese oxidizing bacteria into a manganese oxidizing bacteria culture medium, then adding the biochar from step (1) into the manganese oxidizing bacteria culture medium for cultivation, and adding a divalent manganese solution after the cultivation is completed.

2. The method for improving the biological manganese oxidation performance of manganese oxidizing bacteria according to claim 1, characterized in that: In step (1), the straw pretreatment method is: The straw is dried at 60-80°C to reduce the moisture content of the straw to 10-15%, and is crushed to a straw length of 1-5 cm.

3. The method for improving the biological manganese oxidation performance of manganese oxidizing bacteria according to claim 1, characterized in that: In step (1), the temperature of the high temperature treatment is 300-600° C., and the time is 7-12 hours; and the mesh size of the sieving is 60-300 meshes.

4. The method for improving the biological manganese oxidation performance of manganese oxidizing bacteria according to claim 1, characterized in that: In step (1), the step of acid treatment of the black powder is: The black powder is added into a 0.2-1.0 mol / L hydrochloric acid solution and stirred for 8-24 hours; the mass volume ratio of the black powder to the hydrochloric acid solution is 1 g:10-20 mL.

5. The method for improving the biological manganese oxidation performance of manganese oxidizing bacteria according to claim 1, characterized in that: In step (2), the manganese oxidizing bacteria is P. putida strain MnB1; the concentration of the manganese oxidizing bacteria used for inoculation is 1 to 3×10 7 CFU / ml, the inoculation amount of the manganese oxidizing bacteria is 2-10%, the pH of the manganese oxidizing bacteria culture medium is 6.0-7.5; the addition amount of the biochar is 1.0-10.0 g / L, the culture time is 1-2 days; and the divalent manganese concentration is 2.5-12.0 mg / L.

6. Use of the method for improving the biological manganese oxidizing performance of manganese oxidizing bacteria as claimed in any one of claims 1 to 5 in purifying sewage, characterized in that: The method of application is: The manganese oxidizing bacteria of claim 1 are inoculated into sewage, the biochar prepared in step (1) of claim 1 is added for culture, and then a divalent manganese solution is added.

7. Application of the method for improving the biological manganese oxidizing performance of manganese oxidizing bacteria according to claim 6 in purifying sewage, characterized in that: The pH of the sewage is 6.0-7.5; the inoculation amount of manganese oxidizing bacteria in the sewage is 2-10%, and the concentration of the manganese oxidizing bacteria is 1-3×10 7 CFU / ml; the dosage of the biochar is 1.0-10.0 g / L, the culture time is 1-2 days; and the divalent manganese concentration is 5-12.0 mg / L.

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