Method for improving manganese-oxidizing bacteria's manganese-oxidizing performance and application thereof
By adding straw-based biochar to the manganese-oxidizing bacteria culture medium, the problem of improving manganese oxidation performance was solved, achieving efficient and low-cost improvement in manganese oxidation performance and heavy metal removal.
Patent Information
- Application Number
- CN202411614470.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2024-11-13
- Publication Date
- 2026-02-10
- Estimated Expiration
- 2044-11-13
AI Technical Summary
Existing technologies are insufficient to effectively improve the bio-manganese oxidation performance of manganese-oxidizing bacteria, and traditional methods are costly and environmentally unfriendly.
Biochar was prepared by using straw, and manganese oxidizing bacteria were inoculated into a manganese oxidizing bacteria culture medium. Divalent manganese solution was added, and biochar was used as an electron shuttle to improve the biological activity of manganese oxidizing bacteria and the oxidation rate of divalent manganese.
It significantly improves the oxidation rate of divalent manganese and the removal efficiency of heavy metals in wastewater. The use of biochar is green, environmentally friendly, and inexpensive, and provides a good environment for microbial growth.
Smart Images
Figure CN119330513B_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The present application relates to the technical field of sewage treatment, and particularly relates to a method for improving biological manganese oxidation performance of manganese-oxidizing bacteria and application. BACKGROUND
[0002] Manganese-oxidizing bacteria (MnOB) is a kind of microorganism that can quickly catalyze the oxidation of Mn(II) and induce the formation of biological manganese oxide (BMO). Under neutral, normal temperature and pressure, manganese-oxidizing bacteria can induce the formation of biological manganese oxide by biosynthesis. Compared with chemically synthesized manganese oxide, the biosynthetic BMO has smaller particle size, weaker crystallization, higher Mn valence, more holes in the octahedral structure, larger specific surface area and stronger reactivity. Under natural environment, the biological oxidation rate of manganese-oxidizing bacteria to divalent manganese is 10 times higher than that of chemical oxidation. 5 In addition, due to its special crystal structure and surface chemical properties, biological manganese oxide has strong adsorption capacity for a variety of heavy metal ions and can adsorb and degrade some organic pollutants such as pesticides, endocrine disruptors, antibiotics and polycyclic aromatic hydrocarbons. Therefore, manganese-oxidizing bacteria plays an important role in the field of contaminated environment remediation.
[0003] During the process of manganese-oxidizing bacteria inducing biological manganese oxide, the biological manganese oxidation performance plays a crucial role, which can be reflected by the oxidation rate of divalent manganese and the generation rate of biological manganese oxide. Many environmental factors can affect the biological manganese oxidation performance, such as DO, pH, Eh, temperature, divalent manganese concentration and strain type. Overall, the faster the biological manganese oxidation rate, the higher the biological manganese oxide, and the more conducive to the remediation of pollutants.
[0004] Currently, there are mainly two ways to improve the performance of biological manganese oxide. First, increase the stimulating factors to promote the manganese oxidation reaction, accelerate the oxidation rate of divalent manganese and the generation rate of manganese oxide. Second, add carriers 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 engineering, and the improvement is very limited, so the second method is mostly used. Therefore, it is urgent to develop a green, low-cost and non-secondary pollution method for improving the performance of biological manganese oxidation. SUMMARY
[0005] The purpose of the present application is to provide a method for improving the biological manganese oxidation performance of manganese-oxidizing bacteria and application, which can significantly improve the oxidation rate of divalent manganese and improve the removal efficiency of heavy metals in wastewater.
[0006] In order to achieve the above-mentioned purpose of the application, the present application provides the following technical solutions.
[0007] The application provides a method for improving biological manganese oxidation performance of manganese-oxidizing bacteria.
[0008] (1) after the straw is pretreated, high-temperature treatment is conducted under a CO2 atmosphere, and then cooling, grinding and sieving are conducted to obtain black powder, and acid treatment, washing and drying of the black powder are conducted to obtain biochar powder;
[0009] (2) the manganese-oxidizing bacteria are inoculated into a manganese-oxidizing bacteria culture medium, and then the biochar of step (1) is added into the manganese-oxidizing bacteria culture medium for culture, and after the culture is completed, a divalent manganese solution is added.
[0010] Preferably, in step (1), the method for pretreating the straw is as follows:
[0011] The straw is dried at 60-80 DEG C to reduce the water content of the straw to 10-15%, and the straw is crushed to a length of 1-5 cm.
[0012] Preferably, in step (1), the temperature of the high-temperature treatment is 300-600 DEG C, and the time is 7-12 h; and the mesh number of the sieving is 60-300 meshes.
[0013] Preferably, in step (1), the acid treatment of the black powder is as follows:
[0014] The black powder is added into a 0.2-1.0 mol / L hydrochloric acid solution, and stirring is conducted for 8-24 h; 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 are P.putida strain MnB1; the concentration of the manganese-oxidizing bacteria for inoculation is 1-3 x 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, and the culture time is 1-2 d; and the concentration of the divalent manganese is 2.5-12.0 mg / L.
[0016] The application further provides an application of the method for improving the biological manganese oxidation performance of the manganese-oxidizing bacteria to purifying sewage.
[0017] The manganese-oxidizing bacteria are inoculated into sewage, biochar is added for culture, and then a divalent manganese solution is added.
[0018] Preferably, the PH of the sewage is 6.0-7.5; the inoculation amount of the manganese-oxidizing bacteria in the sewage is 2-10%, and the concentration of the manganese-oxidizing bacteria is 1-3 x 10 7CFU / ml; the adding amount of the biochar is 1.0-10.0g / L, the culture time is 1-2d, and the concentration of divalent manganese is 5-12.0mg / L.
[0019] Compared with the prior art, the present application has the following beneficial effects:
[0020] (1) The present application improves the biological manganese oxidation performance of manganese-oxidizing bacteria by adding biochar. After adding biochar to the manganese-oxidizing bacteria and then adding divalent manganese, the oxidation rate of divalent manganese can be significantly improved, and the removal efficiency of heavy metals in wastewater can be improved. The biochar can act as an electron shuttle, improve the electron transfer capacity, and accelerate the oxidation rate of divalent manganese. The biochar also improves the gene abundance and expression capacity 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 soil, water and air during use. Moreover, biochar is widely available and low in cost, can realize resource recycling and reduce treatment cost, and is easy to prepare and put into use, facilitating the use in practical applications.
[0022] (3) Compared with free bacteria, the addition of biochar provides a biological carrier for manganese-oxidizing bacteria, helps the attached growth of manganese-oxidizing bacteria, and creates a good living environment for manganese-oxidizing bacteria. BRIEF DESCRIPTION OF DRAWINGS
[0023] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings needed in the embodiments will be briefly introduced as follows. Obviously, the drawings in the following description only constitute some embodiments of the present application, and those skilled in the art can also obtain other drawings according to these drawings without creative labor.
[0024] Figure 1 It is a scanning electron microscope image of biochar in Example 1 of the present application.
[0025] Figure 2 It is a scanning electron microscope image of biochar after colony attachment in Example 1 of the present application. DETAILED DESCRIPTION
[0026] The various exemplary embodiments of the present application will now be described in detail, which should not be considered as limiting the present application, but should be understood as a more detailed description of certain aspects, characteristics and embodiments of the present application.
[0027] It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the present application. In addition, where particular ranges of values are given, understand that each intervening value, to the upper or lower limit of the ranges is also specifically included. The upper and lower limits of these smaller ranges can independently be included or excluded in the range, and are also encompassed within the application, subject to any specifically excluded limit in the stated
[0028] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. Although preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present application. All documents mentioned herein are incorporated by reference to disclose and describe in detail the methods and / or materials which are related to the present application. In the case of conflict between the content of the specification and that of any document incorporated herein by reference, the content of the specification prevails.
[0029] Many modifications and variations of the present application described in the specification are possible without departing from the scope or spirit of the application. Other embodiments of the application will be apparent to those skilled in the art from consideration of the specification and practice of the application. The specification and examples are illustrative only.
[0030] As used herein, the terms "comprises", "comprising", "includes", "including", "has", "having", and the like are open-ended terms that are intended to mean including, but not limited to.
[0031] A method for improving the biological manganese oxidation performance of manganese-oxidizing bacteria, comprising the following steps:
[0032] (1) After the straw is pretreated, it is treated at high temperature in a CO2 atmosphere, then cooled, ground, and sieved to obtain black powder, and the black powder is acid-treated, washed, and dried to obtain biochar powder;
[0033] (2) The manganese-oxidizing bacteria are inoculated into a manganese-oxidizing bacteria culture medium, then the biochar of step (1) is added to the manganese-oxidizing bacteria culture medium for culture, and a divalent manganese solution is added after the culture is completed.
[0034] Example 1
[0035] The method for improving the biological manganese oxidation performance of manganese-oxidizing bacteria provided in Example 1 of the present application has the following specific steps:
[0036] (1) Preparation of P. putida strain MnB1 bacterial solution:
[0037] Pseudomonas putida strain MnB1 was inoculated into Pseudomonas putida enrichment medium at a transfer rate of 3% (v / v), and then aerobically enriched for 2 days with horizontal shaking (150 rpm) at 30°C to obtain Pseudomonas putida strain MnB1 bacterial suspension with a concentration of 1×10⁻⁶. 7 CFU / ml.
[0038] The culture medium consisted of: yeast extract, 0.5 g / L; acid-hydrolyzed casein, 0.5 g / L; glucose, 1 g / L; calcium chloride dihydrate, 0.29 g / L; magnesium sulfate heptahydrate, 0.82 g / L; ferric chloride, 0.001 g / L; trace elements, 1 mL; pH = 7.0. The trace element composition was: copper sulfate pentahydrate, 2.49 g / L; zinc sulfate heptahydrate, 12.65 g / L; cobalt chloride hexahydrate, 4.76 g / L; sodium molybdate dihydrate, 3.15 g / L.
[0039] The selected *Pseudomonas putida* strain MnB1 was purchased from the American Type Culture Collection (ATCC), catalog number 23483.
[0040] (2) Preparation of biochar:
[0041] The straw is dried at 80℃ to reduce its moisture content to 15%, and then shredded to a length of 5cm.
[0042] 100g of crushed straw was placed in a pyrolysis furnace, a protective CO2 gas was introduced, the temperature was raised to 600℃, and the temperature was maintained for 10 hours. After cooling, the straw was ground and passed through a 300-mesh sieve to obtain a black powder. The black powder obtained above was added to a 1.0 mol / L hydrochloric acid solution at a mass-to-volume ratio of 1g:10mL, stirred for 24 hours, filtered, washed with 300ml of deionized water until the pH reached 8.0, and freeze-dried in a freeze dryer for 24 hours to obtain biochar powder. Its scanning electron microscope image is shown below. Figure 1 As shown.
[0043] Figure 1 The results showed that the addition of biochar provided a larger specific surface area for manganese-oxidizing bacteria, which helped microorganisms attach and provided a favorable environment for their growth.
[0044] (3) Improve the bio-manganese oxidation performance of manganese-oxidizing bacteria:
[0045] After sterilizing and cooling 250 ml of ATCC279# culture medium, add HEPES buffer to adjust the pH to 7.2. Add the manganese-oxidizing bacteria cultured in step 1) at an inoculation ratio of 3%, and the biochar prepared in step 2) at a dosage of 5.0 g / L. Culture for 2 days (after adding biochar, the colonies adhere to its surface as shown). Figure 2As shown in the figure, divalent manganese was added to make a final concentration of 12 mg / L, and the mixture was shaken and cultured in a vertical full-temperature shaking incubator at 30 degrees and 120 rpm.
[0046] Example 2
[0047] In Example 2 of this invention, the method of step (3) in Example 1 was used to detect the constant rate of manganese oxidation kinetics and the rate of BMO formation under different biochar dosages. Unlike Example 1, the biochar dosages in Example 2 were 0 g / L, 1.0 g / L, 2.5 g / L, 5.0 g / L, and 10.0 g / L, respectively. The biochar was cultured at 30 degrees Celsius and 120 rpm in a vertical full-temperature shaking incubator for 48 h. Samples were taken at 4 h, 8 h, 12 h, 24 h, 36 h, and 48 h to determine the concentrations of divalent manganese and BMO. The constant rate of divalent manganese oxidation kinetics and the rate of BMO formation were calculated. The results are shown in Table 1.
[0048] Table 1. Kinetic rate constants of manganese oxidation and BMO formation rate constants under different biochar addition levels.
[0049]
[0050] Table 1 shows that, compared with the control group, the oxidation rate of divalent manganese and the production rate of BMO increased with the addition of biochar. When the biochar dosage was 5.0 g / L, the oxidation rate constant of divalent manganese was 0.23 mg·L⁻¹. -1 ·h -1 The BMO production rate was 1.25 mg (MnO2)·L. -1 ·h -1 Compared with the control group, the increases were 110% and 205%, respectively.
[0051] Example 3
[0052] In Example 3 of this invention, the method of step (3) of Example 1 was used to detect the constant rate of manganese oxidation kinetics and the BMO formation rate under different concentrations of divalent manganese. Unlike Example 1, the final concentrations of divalent manganese 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. Kinetic rates of manganese oxidation and BMO formation rate under different concentrations of divalent manganese.
[0054]
[0055] As shown in Table 2, with Mn 2+ As Mn concentration increases, the oxidation rate constant of divalent manganese increases, and the formation rate of BMO also increases; when Mn 2+At a final concentration of 12.0 mg / L, the oxidation rate constant of divalent manganese and the formation rate of BMO reached their maximum values, with the oxidation rate constant of divalent manganese being 0.47 mg·L⁻¹. -1 ·h -1 The BMO formation rate was 1.24 mg(MnO2)·L. -1 ·h -1 ; and Mn 2+ Compared to a final concentration of 2.5 mg / L, the oxidation rate constant of divalent manganese increased by 11 times, and the BMO formation rate increased by 5 times.
[0056] Example 4
[0057] In Example 4 of this invention, the method of step (3) of Example 1 was used to detect the constant rate of manganese oxidation kinetics and the BMO formation rate under different pH conditions. Unlike Example 1, the pH values in Example 4 were 6.0, 7.2 and 7.6, respectively. The results are shown in Table 3.
[0058] Table 3. Kinetic rates of manganese oxidation and BMO formation rate under different pH conditions.
[0059]
[0060]
[0061] Table 3 shows that, within a certain range, the oxidation rate constant of divalent manganese and the BMO formation rate increase with increasing pH. The oxidation rate constant of divalent manganese and the BMO formation rate reach their maximum at pH 7.2, with the oxidation rate constant of divalent manganese being 0.50 mg·L⁻¹. -1 ·h -1 The BMO formation rate was 0.87 mg (MnO2)·L. -1 ·h -1 .
[0062] Example 5
[0063] In Example 5 of this invention, the Cd removal efficiency of Cd-containing wastewater was tested using the method described in Example 1. The specific method is as follows:
[0064] 1 L of Cd-containing wastewater with an initial concentration of 1.0 mg / L was taken, and the pH of the wastewater was adjusted to 7.2. Manganese-oxidizing bacteria strain P. putida strain MnB1 was added at an inoculation ratio of 3%, and biochar was added at 5.0 g / L. The mixture was cultured for 2 days. Then, divalent manganese was added to a final concentration of 12 mg / L, and the mixture was placed in an incubator and cultured at 30 degrees and 140 rpm for 48 h. The Cd concentration was measured periodically at different time points, and the Cd removal efficiency was calculated. The results are shown in Table 4.
[0065] Table 4 Cd removal rate
[0066]
[0067] As shown in Table 4, the Cd removal rate gradually increased over time, indicating a better removal effect. Within the first 12 hours, the removal rate reached 74.52%, indicating that Cd removal progressed rapidly during this period. From 24 to 48 hours, the removal rate continued to increase, but the growth rate slowed down. The removal rate reached its maximum of 96.75% at the 48th hour, an increase of 22.23% compared to the 12th hour.
[0068] Example 6
[0069] In Example 6 of this invention, the Cd removal efficiency of Cd-containing wastewater was tested using the method of Example 5. The difference from Example 5 is that the concentration of Cd-containing wastewater in Example 6 was 2.0 mg / L. The results are shown in Table 5.
[0070] Table 5 Cd removal rate
[0071]
[0072] As shown in Table 5, the Cd removal rate gradually increased over time, indicating a better removal effect. Within the first 12 hours, the removal rate reached 70.36%, indicating that Cd removal progressed rapidly during this period. From 24 to 48 hours, the removal rate continued to increase, but the rate of increase slowed down. The removal rate reached its maximum of 92.58% at the 48th hour, an increase of 22.22% compared to the 12th hour.
[0073] Example 7
[0074] In Example 7 of this invention, the Cd removal efficiency of Cd-containing wastewater was tested using the method of Example 5. The difference from Example 5 is that the concentration of Cd-containing wastewater in Example 7 was 4.0 mg / L. The results are shown in Table 6.
[0075] Table 6 Cd removal rate
[0076]
[0077] As shown in Table 6, the Cd removal rate gradually increased over time, indicating a better removal effect. Within the first 12 hours, the removal rate reached 70.39%, indicating that Cd removal progressed rapidly during this period. From 24 to 48 hours, the removal rate continued to increase, but the rate of increase slowed down. The removal rate reached its maximum of 93.76% at the 48th hour, an increase of 23.37% compared to the 12th hour.
[0078] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.
Claims
1. A method for improving the bio-manganese oxidation performance of manganese-oxidizing bacteria, characterized in that, Includes the following steps: (1) After straw pretreatment, it is treated at high temperature in CO2 atmosphere, then cooled, ground and sieved to obtain black powder. The black powder is acid treated, washed and dried to obtain biochar powder. (2) Inoculate manganese oxidizing bacteria into manganese oxidizing bacteria culture medium, then add the biochar from step (1) into the manganese oxidizing bacteria culture medium for culture, and add divalent manganese solution after the culture is completed. In step (1), the high-temperature treatment is performed at a temperature of 300–600°C for 7–12 hours; the sieve mesh size is 60–300 mesh. In step (1), the acid treatment of the black powder is as follows: The black powder was added to a 0.2–1.0 mol / L hydrochloric acid solution and stirred for 8–24 h; the mass-to-volume ratio of the black powder to the hydrochloric acid solution was 1 g: 10–20 mL. In step (2), the manganese-oxidizing bacterium is P. putida strain MnB1; the concentration of the manganese-oxidizing bacterium used for inoculation is 1–3 × 10⁻⁶. 7 The concentration of manganese oxidizing bacteria is 2-10% CFU / ml, the pH of the culture medium for manganese oxidizing bacteria is 6.0-7.5, the dosage of biochar is 1.0-10.0 g / L, the culture time is 1-2 days, and the concentration of divalent manganese is 2.5-12.0 mg / L.
2. The method for improving the bio-manganese oxidation performance of manganese-oxidizing bacteria according to claim 1, characterized in that, In step (1), the method for pretreating the straw is as follows: The straw is dried at 60-80℃ to reduce its moisture content to 10-15%, and then shredded to a length of 1-5cm.
3. The application of a method for improving the bio-manganese oxidation performance of manganese-oxidizing bacteria as described in any one of claims 1 to 2 in wastewater purification, characterized in that, The method of application is as follows: The manganese-oxidizing bacteria of claim 1 were inoculated into the sewage, and the biochar prepared in step (1) of claim 1 was added for culture, followed by the addition of divalent manganese solution.
4. The application of the method for improving the bio-manganese oxidation performance of manganese-oxidizing bacteria according to claim 3 in wastewater purification, characterized in that, The wastewater has a pH of 6.0–7.5; the inoculum amount of manganese-oxidizing bacteria in the wastewater is 2–10%, and the concentration of the manganese-oxidizing bacteria is 1–3 × 10⁻⁶. 7 CFU / ml; the amount of biochar added is 1.0-10.0 g / L, the culture time is 1-2 days; the concentration of divalent manganese is 5-12.0 mg / L.