Modified porous composite material as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of soil improvement, and particularly relates to a modified porous composite material as well as a preparation method and application thereof. The composite material comprises modified ceramsite and microorganisms, the modified ceramsite comprises a ceramsite matrix and a calcium alginate gel layer; the ceramsite matrix has a porous structure; the calcium alginate gel layer is loaded on the surface and the interior of the ceramsite matrix, and the surface and the interior of the ceramsite matrix and the interior of the calcium alginate gel layer are coated with microorganisms by the calcium alginate gel layer. The composite material with a specific structure can improve microbial activity, has an obvious promotion effect on ferrous oxidation, and can effectively avoid degradation of organic carbon in soil, so that a stable carbon sequestration effect is achieved; meanwhile, generation of nitrous oxide in the process of iron-oxidizing bacterium ferrous oxidation coupling nitrate reduction can be effectively inhibited, greenhouse gas emission in the process of iron-oxidizing bacterium nitrate reduction coupling ferrous oxidation is reduced, the DOC concentration is reduced, and good carbon sequestration and emission reduction effects are achieved.

Description

Technical Field

[0001] The invention belongs to the technical field of soil improvement, and particularly relates to a modified porous composite material and a preparation method and application thereof. Background Art

[0002] Soil carbon is an important component of the ecosystem, and carbon sequestration helps enhance the stability and anti-disturbance capacity of the ecosystem. The carbon sequestration process can increase the organic matter content in the soil, improve soil fertility and productivity, and thus help increase crop yields. Soil microorganisms actively participate in the decomposition and transformation of organic matter through a variety of metabolic pathways, play a key role in the carbon cycle of the soil system, and help stabilize organic carbon, thereby affecting the storage and turnover of soil carbon.

[0003] At present, the main carbon fixation and emission reduction measures include organic fertilizer application, straw return to the field, conservation tillage, etc., but the application of these measures may have limitations, such as the accumulation of heavy metals in the soil and the increase of the number of antibiotics in the soil, thereby increasing environmental risks. In addition, porous materials such as ceramsite and biochar can also be used as a means of carbon fixation and emission reduction, but there are ecological and health risks in their production and application. For example, pollutants such as heavy metals that may exist in synthetic raw materials will be enriched in the process of biochar preparation. Studies have shown that functional microorganisms such as iron oxidizing bacteria and iron reducing bacteria can actively participate in the decomposition and transformation of organic matter through a variety of metabolic pathways, and play a key role in the carbon cycle of soil systems. However, there are also certain disadvantages in the application of functional microorganisms. For example, there will be competition and antagonism between exogenous microorganisms and microorganisms in the ecosystem of the use area, and the original ecological balance will be broken, which may cause a decrease in the number of both microorganisms and slow growth; it may also inhibit the growth of other types of microorganisms, thereby causing secondary pollution to the environment. In addition, as exogenous strains, there must be competition between artificially added functional microorganisms and indigenous microorganisms for nutrients and living environment, which will make it difficult for exogenously added functional microorganisms to colonize.

[0004] Some studies have used polymer materials such as alginate or polyvinyl alcohol as carriers, combined with other materials to immobilize microorganisms, but there are still disadvantages such as being limited by environmental media and having a single function. Some studies have used attapulgite to cross-link with calcium alginate containing bacterial strains. Although the fixation of microorganisms was achieved, the fixation effect only existed on the surface of the material, the fixation effect was single, and it was mainly used in sewage treatment, and it was difficult to play a role in the soil. Some studies have used biochar as a skeleton to embed bacterial strains in seaweed microspheres to achieve the fixation of heavy metals, but the biochar particles are small, making it difficult for the gel microspheres to maintain their original form in the soil for a long time, and the main purpose is to fix soil heavy metals; at the same time, the large amount of iron in the soil is easy to combine with microbial cells, co-precipitate with microorganisms, and cause the loss of microbial activity. In short, the above methods still have shortcomings in soil carbon sequestration and emission reduction.

[0005] Therefore, it is of great significance to provide a material with good carbon fixation and emission reduction effects, which can improve soil microbial activity and reduce dissolved organic carbon (DOC) concentration. Summary of the invention

[0006] The present invention aims to solve one or more technical problems existing in the above-mentioned prior art and at least provide a beneficial option. Specifically, the present invention provides a composite material, which has good carbon fixation and emission reduction effects, can increase soil microbial activity and reduce DOC concentration.

[0007] The inventive concept of the present invention is as follows: the composite material of the present invention comprises modified ceramsite and microorganisms; the modified ceramsite comprises a ceramsite matrix and a calcium alginate gel layer; the ceramsite matrix has a porous structure; the calcium alginate gel layer is loaded on the surface and inside of the ceramsite matrix, and the calcium alginate gel layer encapsulates the microorganism on the surface, inside of the ceramsite matrix and in the calcium alginate gel layer; the inside refers to the pore surface of the porous structure. The calcium alginate gel layer and the inside of the ceramsite matrix of the present invention are both porous structures, which can provide nucleation sites for the secondary minerals of the microorganisms, thereby reducing the wrapping pressure of the secondary minerals on the microorganisms themselves, thereby indirectly enhancing the biological activity of the microorganisms, thereby improving the ferrous iron oxidation ability of the microorganisms, and the same is true for the nitrate reduction ability; and the porous structure can also provide a stable colonization environment for the microorganisms, avoid the loss of microorganisms, promote the reproduction of microorganisms, and directly improve the ferrous iron oxidation and nitrate reduction abilities of the microorganisms. The iron minerals formed during ferrous oxidation can combine with functional groups (such as carboxyl, phenolic and amine) in organic carbon to form a stable structure, which can effectively avoid the degradation of soil organic carbon and achieve a stable carbon fixation effect; at the same time, the composite material can effectively inhibit the generation of nitrous oxide during the ferrous oxidation coupled with nitrate reduction by iron-oxidizing bacteria, reduce greenhouse gas emissions during the nitrate reduction coupled with ferrous oxidation by iron-oxidizing bacteria, and reduce DOC concentration. In addition, the microorganisms are coated on the surface, inside and in the calcium alginate gel layer of the ceramsite matrix, which protects the functional microorganisms, prevents their loss and provides them with a stable colonization space, which can maintain the good biological activity of the functional microorganisms.

[0008] Accordingly, a first aspect of the present invention provides a modified porous composite material.

[0009] Specifically, the modified porous composite material includes modified ceramsite and microorganisms;

[0010] The modified ceramsite comprises a ceramsite matrix and a calcium alginate gel layer;

[0011] The ceramsite matrix has a porous structure; the calcium alginate gel layer is loaded on the surface and inside of the ceramsite matrix, and the calcium alginate gel layer covers the microorganism on the surface, inside of the ceramsite matrix and in the calcium alginate gel layer.

[0012] Specifically, the interior refers to the pore surface of the porous structure.

[0013] Preferably, the ceramsite matrix is ​​acidified ceramsite.

[0014] Preferably, the particle size of the ceramsite is 3.5-5.5 mm; further preferably, the particle size of the ceramsite is 4-5 mm.

[0015] Preferably, the mass ratio of the ceramsite matrix to the calcium alginate gel layer is (1.8-3.3):1; further preferably, the mass ratio of the ceramsite matrix to the calcium alginate gel layer is (2.0-3.0):1.

[0016] Preferably, the microorganisms include iron oxidizing bacteria.

[0017] The second aspect of the present invention provides a method for preparing the modified porous composite material according to the first aspect of the present invention.

[0018] Specifically, the method for preparing the modified porous composite material comprises the following steps:

[0019] (1) mixing a ceramsite matrix, a calcium salt, and a solvent to obtain a ceramsite matrix loaded with calcium salt; mixing an alginate solution and a microbial culture liquid to obtain a mixture;

[0020] (2) mixing the calcium salt-loaded ceramsite matrix obtained in step (1) with the mixture to obtain the modified porous composite material.

[0021] Preferably, the method for preparing the ceramsite matrix comprises the following steps:

[0022] The ceramsite and acid solution are mixed and acidified to obtain the ceramsite matrix.

[0023] Preferably, the ceramsite is first placed in a rotary agitator for a stirring treatment, and then mixed with the acid solution.

[0024] Preferably, the rotation speed of the stirring treatment is 140-190 rpm, and the stirring treatment time is 1.5-2.5 h; further preferably, the rotation speed of the stirring treatment is 150-180 rpm, and the stirring treatment time is 1.8-2.2 h.

[0025] Specifically, the stirring treatment is magnetic stirring, and the purpose of the stirring treatment is to remove impurities and worn skin on the surface of the ceramsite through preliminary friction, thereby increasing the surface porosity.

[0026] Preferably, the acid solution comprises hydrochloric acid.

[0027] Preferably, the concentration of the acid solution is 0.8-1.2M; further preferably, the concentration of the acid solution is 0.9-1.1M; further preferably, the concentration of the acid solution is 1.0M.

[0028] Preferably, the stirring speed of the acidification treatment is 180-270 rpm; further preferably, the stirring speed of the acidification treatment is 200-250 rpm.

[0029] Preferably, the acidification treatment time is 4.5-6.5 h; further preferably, the acidification treatment time is 5-6 h.

[0030] Preferably, the acidification treatment further includes washing and drying processes.

[0031] Preferably, in step (1), the ceramsite matrix, calcium salt and solvent are mixed by stirring, the stirring speed is 180-270 rpm, and the stirring time is 25-35 min; further preferably, the stirring speed is 200-250 rpm, and the stirring time is 27-33 min.

[0032] Preferably, in step (1), the solvent comprises water; further preferably, the water is deionized water.

[0033] Preferably, the ratio of the solvent to the calcium salt is 1 mL: (1.1-1.6) g; further preferably, the ratio of the solvent to the calcium salt is 1 mL: (1.3-1.4) g.

[0034] Preferably, in step (1), the mass ratio of the ceramsite matrix to the calcium salt is 1:(3.0-4.0); further preferably, the mass ratio of the ceramsite matrix to the calcium salt is 1:(3.25-3.75).

[0035] Preferably, the calcium salt comprises calcium chloride.

[0036] Preferably, in step (1), the calcium salt and the solvent are first mixed and ultrasonically treated to obtain a supersaturated calcium salt, which is then mixed with the ceramsite matrix to obtain a ceramsite matrix loaded with calcium salt.

[0037] Specifically, the calcium salt is loaded on the surface of the ceramsite matrix and the pore surface of the porous structure.

[0038] Preferably, in step (1), the alginate solution comprises a sodium alginate solution.

[0039] Preferably, the mass fraction of the alginate solution is 2.2-3.8%; further preferably, the mass fraction of the alginate solution is 2.5-3.5%.

[0040] Preferably, the temperature of the alginate solution is 9-16°C; further preferably, the temperature of the alginate solution is 10-15°C.

[0041] Preferably, in step (1), the volume ratio of the alginate solution to the microbial culture liquid is 1:(0.25-0.45) mL; further preferably, in step (1), the volume ratio of the alginate solution to the microbial culture liquid is 1:(0.3-0.4) mL.

[0042] Preferably, the cell concentration in the microbial culture liquid is (3.0-5.5)×10 7 cells / mL; further preferably, the cell concentration in the microbial culture liquid is (3.2-5.0)×10 7 Pieces / mL.

[0043] Preferably, the alginate solution and the microbial culture liquid are mixed by stirring, the stirring speed is 90-160 rpm, and the stirring time is 4-6 min; more preferably, the stirring speed is 100-150 rpm, and the stirring time is 4.5-5.5 min.

[0044] Preferably, in step (2), the ratio of the amount of the ceramsite substrate loaded with calcium salt to the alginate solution in the mixture is 7-13 g: 1 mL; further preferably, in step (2), the ratio of the amount of the ceramsite substrate loaded with calcium salt to the alginate solution in the mixture is 8-12 g: 1 mL.

[0045] Preferably, in step (2), the mixing is carried out by stirring, the stirring speed is 180-270 rpm, and the stirring time is 13-18 min; more preferably, the stirring speed is 200-250 rpm, and the stirring time is 14-16 min.

[0046] Preferably, in step (2), the ceramsite matrix loaded with calcium salt is added to the mixture at a rate of 9-16 g / min; further preferably, in step (2), the ceramsite matrix loaded with calcium salt is added to the mixture at a rate of 10-15 g / min.

[0047] Preferably, the microbial liquid comprises an iron oxidizing bacteria liquid.

[0048] Preferably, the iron-oxidizing bacteria species in the iron-oxidizing bacteria liquid is selected from any one of Acidovorax sp.BoFeN1 (BoFeN1), Pseudogulbenkiania sp.strain 2002 (SP 2002) and Klebsiella pneumoniae (L17).

[0049] Specifically, the BoFeN1 and SP 2002 were purchased from the German Culture Collection Center, and L17 was purchased from the China Marine Microbial Culture Collection Center.

[0050] Preferably, the absorbance of the microbial culture at 600 nm is 0.7-1.1; further preferably, the absorbance of the microbial culture at 600 nm (OD 600 ) is 0.8-1.0.

[0051] Preferably, the culture process of the iron oxidizing bacteria liquid comprises the following steps:

[0052] (1) adding an iron-oxidizing bacterial strain to an anaerobic culture medium for culturing to obtain an activated iron-oxidizing bacterial liquid; and then adding the iron-oxidizing bacterial strain to a new anaerobic culture medium for culturing again to obtain a bacterial liquid 1;

[0053] (2) Centrifuging the bacterial solution obtained in step (1), removing the supernatant, mixing the bacterial cells with a buffer solution to obtain bacterial solution 2, and adjusting the absorbance of bacterial solution 2 to obtain an iron-oxidizing bacterial solution.

[0054] Preferably, in step (1), the components of the anaerobic culture medium include CaCl2·2H2O, KH2PO4, MgCl2·6H2O, NaCl, NH4Cl, piperazine-1,4-diethanesulfonic acid (PIPES), CH3COONa and NaNO3.

[0055] Preferably, in step (1), the volume fraction of the iron oxidizing bacteria inoculated is 8-12%; further preferably, the volume fraction of the iron oxidizing bacteria inoculated is 9-11%; further preferably, the volume fraction of the iron oxidizing bacteria inoculated is 10%.

[0056] Preferably, in step (2), after adjusting the absorbance of the bacterial solution 2, the step further includes introducing nitrogen to remove oxygen in the bacterial solution.

[0057] The third aspect of the present invention provides a use of the modified porous composite material described in the first aspect of the present invention in soil improvement.

[0058] Compared with the prior art, the technical solution provided by the present invention has the following beneficial effects:

[0059] (1) The composite material of the specific structure of the present invention has a significant promoting effect on ferrous iron oxidation. The iron minerals formed during ferrous iron oxidation can combine with functional groups (such as carboxyl, phenolic and amine groups) in organic carbon to form a stable structure, which can effectively avoid the degradation of soil organic carbon, thereby achieving a stable carbon fixation effect; at the same time, it can effectively inhibit the generation of nitrous oxide during the ferrous iron oxidation coupled with nitrate reduction by iron oxidizing bacteria, reduce greenhouse gas emissions during the nitrate reduction coupled with ferrous iron oxidation by iron oxidizing bacteria, and reduce DOC concentration. In addition, the microorganisms are coated on the surface, inside and in the calcium alginate gel layer of the ceramsite matrix, which protects the functional microorganisms, prevents their loss and provides a stable colonization space for them, and can maintain the good biological activity of the functional microorganisms.

[0060] (2) The ceramsite matrix of the present invention is a porous material, which has good water and fertilizer retention. After being modified by calcium alginate gel, the gel, as a three-dimensional network structure material with high hydrophilicity, can absorb a large amount of water, which makes the composite material have excellent water retention capacity. Under drought conditions, this water retention performance significantly improves the drought resistance of the soil and reduces the demand for irrigation, which is of great significance for water-saving agriculture. In addition, the porous structure can provide a rich habitat for microorganisms, increase the diversity and number of microorganisms in the soil. This not only helps to improve the biological activity of the soil, but also promotes the reproduction and activity of beneficial bacteria, and enhances its stability and durability in the soil.

[0061] (3) The high specific surface area and pore structure of the ceramsite matrix make it an efficient carrier that can adsorb and fix nutrients in the soil, such as nitrogen, phosphorus, potassium, etc., thereby reducing nutrient loss and improving fertilizer utilization efficiency. This slow-release effect helps to achieve precise fertilization and reduce environmental pollution problems caused by excessive fertilization. At the same time, the pore structure and adsorption properties can be used to fix and degrade organic pollutants and heavy metals in the soil, reduce soil pollution, and restore soil ecological functions. At the same time, in the composite material, the porous structure of the ceramsite matrix can also improve the physical properties of the soil, such as increasing the porosity and air permeability of the soil, and promoting the healthy development of the root system. A good soil structure helps the effective exchange of water and air, providing a more superior environment for plant growth.

[0062] (4) The present invention has a simple production process, low material price, good application value and economic benefits, and is convenient for large-scale production and application. BRIEF DESCRIPTION OF THE DRAWINGS

[0063] Figure 1 This is a morphology diagram of the modified porous composite material of Example 1 of the present invention;

[0064] Figure 2The surface structure scanning electron microscope images of the ceramsite of Example 1 and the modified porous composite material of Example 1 of the present invention are shown;

[0065] Figure 3 It is a diagram showing the structural stability results of the modified porous composite material of Example 1 of the present invention and the iron oxidizing bacteria-calcium alginate gel composite material of the control group;

[0066] Figure 4 The scanning electron microscope images of the iron oxidizing bacteria in the modified porous composite material of Example 1 of the present invention and the control group after microcosmic culture;

[0067] Figure 5 The ferrous oxidation results of the modified porous composite material of Example 2 of the present invention and the control group material are shown in FIG.

[0068] Figure 6 It is a graph showing the nitrate reduction results of the modified porous composite material of Example 2 of the present invention and the control group material;

[0069] Figure 7 This is a graph showing the effect of the modified porous composite material of Example 3 of the present invention and the control group material on the change of nitrous oxide concentration;

[0070] Figure 8 This is a graph showing the influence of the modified porous composite material of Example 3 of the present invention and the control group material on the change of dissolved organic carbon concentration. DETAILED DESCRIPTION

[0071] In order to make the technical scheme of the present invention more clearly understood by those skilled in the art, the following embodiments are listed for illustration. It should be pointed out that the following embodiments do not limit the protection scope of the present invention.

[0072] Unless otherwise specified, the raw materials, reagents or devices used in the following examples can be obtained from conventional commercial sources or by existing known methods.

[0073] Example 1

[0074] A method for preparing a modified porous composite material comprises the following steps:

[0075] (1) 0.25 kg of ceramsite with a diameter of 4-5 mm is placed in a rotary stirrer and mixed at a speed of 175 rpm for 2 hours. After the stirring is completed, the ceramsite is placed on a 10-mesh sieve to screen out impurities left after friction; after the screening is completed, the ceramsite is placed in a 1L beaker, and a 1.0M hydrochloric acid solution and a magnetic rotor and rotor of a magnetic stirrer are added. The acid solution is sufficient to cover the ceramsite by about 5 cm. The magnetic stirrer is stirred at a speed of 220 rpm. After the ceramsite is acidified for 6 hours, the ceramsite is taken out; it is washed with deionized water 5 times, and dried at 60° C. to obtain a ceramsite matrix for standby use;

[0076] (2) Add 500 mL of deionized water and 700 g of calcium chloride to a 1 L beaker, and perform ultrasonic treatment for 30 min to ensure that the calcium chloride solution is supersaturated and solids are precipitated at the bottom of the beaker; take 0.2 kg of the ceramsite matrix and magnet obtained in step (1), put them into the supersaturated calcium chloride solution, stir at a rate of 220 rpm for 30 min, take out the ceramsite matrix, place it in an oven and dry it at 85° C. to obtain a ceramsite matrix loaded with calcium chloride particles;

[0077] (3) The frozen strain BoFeN1 was activated in an anaerobic medium (pH = 7.0). Specifically, the strain BoFeN1 stored in glycerol in a -80°C refrigerator was taken out and after thawing, it was injected into a sterile anaerobic medium with an inoculum volume fraction of 10% using a sterile syringe. The composition of the sterile anaerobic medium was as follows: 0.68 mmol·L -1 CaCl2·2H2O, 1.03 mmol·L -1 KH2PO4, 2.07 mmol·L -1 MgCl2·6H2O, 5.14mmol·L -1 NaCl, 5.60 mmol·L -1 NH4Cl, 30mmol·L -1 PIPES, 5.0 mmol·L -1 CH3COONa, 10.0mmol·L -1 NaNO3; in a constant temperature incubator, culture at 30°C in the dark for 24 h to obtain an activated bacterial solution; transfer the activated bacterial solution to a new anaerobic culture medium in the same way, culture at 30°C in the dark for 20 h, centrifuge the obtained bacterial solution at 4°C for 5 min (8000 g), discard the supernatant, and use sterile PIPES buffer (30 mmol·L -1 , pH = 7.0) and then centrifuged (8000g, 5min, 4°C) for 3 times to obtain bacterial cells; the bacterial cells were resuspended in PIPES buffer and mixed thoroughly to obtain a bacterial suspension, and the absorbance of the bacterial suspension at 600nm (OD 600 ) is 0.8-1.0, adjust the OD 600 The bacterial suspension was passed through nitrogen (filtered through a sterile filter) for 40 minutes to remove oxygen from the bacterial suspension, and the iron-oxidizing bacterial solution was obtained, which was placed in an anaerobic workstation for standby use;

[0078] (4) Add 500 mL of deionized water to a beaker, add a certain amount of sodium alginate to make its mass fraction 33.5%, and fully dissolve it; centrifuge the cultured iron-oxidizing bacterial solution obtained in step (2), resuspend it, and adjust the OD600 =0.4, the cell concentration is about 4.5×10 7 / mL; add 7.5mL of iron oxidizing bacteria solution to every 17.5mL of cooled sodium alginate solution, and stir at a rate of 150rpm for 5min using a magnetic stirrer, then adjust the stirring speed to 250rpm, gradually add to the ceramsite matrix obtained in step (1) at a rate of 15g / min, add a total of 250g, continue stirring for 15min after the addition, and finally filter using a 10-mesh sieve to obtain a modified porous composite material.

[0079] The morphology of the composite material prepared in Example 1 was observed, and the results were as follows: Figure 1 As shown, Figure 1 Figure (a) shows the overall morphology of the composite material. Figure 1 Figure (b) is a local scanning electron microscope image of the composite material at low magnification.

[0080] The surface structures of the ceramsite in Example 1 and the composite material prepared in Example 1 were observed by scanning electron microscopy. Figure 2 As shown. Among them, Figure 2 Figure (a) is a scanning electron microscope image of the ceramsite of Example 1; Figure 2 Figure (b) is a scanning electron microscope image of the composite material of Example 1.

[0081] Depend on Figure 2 It can be seen that compared with the rough surface of the ceramsite before modification, a smooth calcium alginate "protective film" is formed on the surface of the modified ceramsite after modification. The formation of the "protective film" is through the cross-linking reaction between the calcium chloride particles loaded on the ceramsite matrix and the sodium alginate solution, forming a calcium alginate gel layer on the surface of the ceramsite matrix. The calcium alginate gel layer can protect the functional microorganisms loaded on the modified ceramsite, prevent their loss and provide a stable customized environment for them.

[0082] The structural stability of the composite material prepared in Example 1 was tested, and a single system of iron oxidizing bacteria-calcium alginate gel composite material was set as a control group. The composite material of Example 1 and the iron oxidizing bacteria-calcium alginate gel composite material of the control group were placed naturally in a dry environment at 30°C to observe their structural stability.

[0083] Among them, the preparation process of the iron-oxidizing bacteria-calcium alginate gel composite material is as follows: first, prepare a sodium alginate solution, weigh 2.0g of sodium alginate in an anaerobic workstation, stir and dissolve it in 70mL of oxygen-free ultrapure water, and sterilize it in a high-temperature sterilizer (121°C, 20min), then put it back into the anaerobic workstation and cool it to room temperature for use. Prepare a CaCl2 solution with a mass fraction of 2.0wt%, and pass nitrogen for 40min to remove the dissolved oxygen in the solution. Similarly, sterilize the CaCl2 solution at high temperature (121°C, 20min), cool it to room temperature, and place it in the anaerobic workstation for use. In the anaerobic workstation, add 7.5mL, OD 600 =1.33 bacterial suspension was uniformly mixed with 17.5 mL of sodium alginate solution to prepare an iron oxidizing bacteria-sodium alginate mixed solution. The OD of the obtained mixed solution was 600 The theoretical value is about 0.4. Subsequently, the iron oxidizing bacteria-sodium alginate mixture was drawn with a disposable sterile syringe and added dropwise to a 2.0wt% CaCl2 solution (stirred continuously with a magnetic stirrer) to form gel microspheres with a diameter of 0.3-0.5cm. After the gel microspheres were hardened at room temperature for 2h in an anaerobic workstation, they were washed several times with sterile oxygen-free ultrapure water to obtain an iron oxidizing bacteria-calcium alginate gel composite material.

[0084] The structural stability test results of the composite material of Example 1 and the iron oxidizing bacteria-calcium alginate gel composite material of the control group are as follows Figure 3 As shown. Among them, Figure 3 Figure (a) is a structural morphology of the material before natural placement. The left material in Figure (a) is the composite material of Example 1, and the right material in Figure (a) is the iron oxidizing bacteria-calcium alginate gel composite material of the control group; Figure 3 Figure (b) is a structural morphology of the material after natural placement for 72 hours. The material on the left in Figure (b) is the composite material of Example 1, and the material on the right in Figure (b) is the iron-oxidizing bacteria-calcium alginate gel composite material of the control group.

[0085] Depend on Figure 3 It can be clearly seen that the structure of the single system iron oxidizing bacteria-calcium alginate gel composite material will collapse and deform significantly after being placed in a dry room temperature environment, which will severely compress the living space of microorganisms inside the material. However, the composite material of Example 1 has a stable structure and does not shrink or collapse significantly after being placed for 72 hours, indicating that the composite material prepared in Example 1 has the advantage of maintaining a gel structure for a long time.

[0086] The composite material prepared in Example 1 was subjected to a microcosm culture experiment. The specific method was as follows: 50 mL of PIPES buffer, 0.25 mL of ferrous chloride solution, and 0.625 mL of nitrate solution were added to a 100 mL vial to construct a microcosm culture system. Wherein, the concentration of the PIPES buffer was 30 mM, the pH was 7.0, the concentration of the ferrous chloride solution was 6 mM, and the concentration of the nitrate solution was 10 mM. The relevant solutions used in the culture experiment have all been subjected to high temperature sterilization to avoid interfering with the culture process. Wherein, 1.5 g of the composite material of Example 1 was added to the experimental group, wherein the proportion of each component was 1 g of the ceramsite matrix loaded with calcium chloride, 0.5 g of the sodium alginate solution and the bacterial suspension (the volume of the bacterial suspension was 2.5 mL, OD 600 The control group was a pure microcosm culture system with only iron-oxidizing bacteria added, and only 2.5 mL and OD 600 The iron-oxidizing bacteria suspension with a value of 0.4, the corresponding iron-oxidizing bacteria microbial biomass is consistent with that in the experimental group. The microcosm culture experiment was carried out at 30°C in the dark, and the culture time was 72 hours. After the culture was completed, destructive sampling was performed, and the iron-oxidizing bacteria in the composite material of Example 1 and the pure culture system of the control group were scanned and observed. The scanning electron microscope images of the iron-oxidizing bacteria in the composite material of Example 1 and the control group after microcosm culture are as follows Figure 4 As shown. Among them, Figure 4 Figure (a) is a scanning electron micrograph of iron-oxidizing bacteria in the pure microcosm culture system of the control group. Figure 4 Figure (b) is a scanning electron microscope image of iron oxidizing bacteria in the composite material of Example 1.

[0087] Depend on Figure 4 It can be seen that the cell encrustation and agglomeration of iron-oxidizing bacteria in the pure microcosm culture system of the control group are serious, and the secondary minerals and cells form tight aggregates, which are not conducive to the growth and metabolism of microorganisms; although the cell encrustation phenomenon of iron-oxidizing bacteria in the composite material of Example 1 is not completely inhibited, its cell agglomeration phenomenon is weak, the connection between microorganisms and secondary minerals is relatively loose, and the iron-oxidizing bacteria can still maintain a good morphological structure.

[0088] Example 2

[0089] A method for preparing a modified porous composite material comprises the following steps:

[0090] (1) 0.25 kg of ceramsite with a diameter of 4-5 mm is placed in a rotary stirrer and mixed at a rate of 175 rpm for 2 h. After the stirring is completed, the ceramsite is placed on a 10-mesh sieve to screen out impurities left after friction; after the screening is completed, the ceramsite is placed in a 1L beaker, and a 1.0M hydrochloric acid solution and a magnetic rotor and rotor of a magnetic stirrer are added. The acid solution is sufficient to cover the ceramsite by about 5 cm. The magnetic stirrer is stirred at a rate of 220 rpm. After the ceramsite is acidified for 6 h, the ceramsite is taken out; it is washed with deionized water 5 times, and dried at 60° C. to obtain a ceramsite matrix for standby use;

[0091] (2) Add 500 mL of deionized water and 700 g of calcium chloride to a 1 L beaker, and perform ultrasonic treatment for 30 min to ensure that the calcium chloride solution is supersaturated and solids are precipitated at the bottom of the beaker; take 0.2 kg of the ceramsite matrix and magnet obtained in step (1), put them into the supersaturated calcium chloride solution, stir at a rate of 220 rpm for 30 min, take out the ceramsite matrix, place it in an oven and dry it at 80-85° C. to obtain a ceramsite matrix loaded with calcium chloride particles;

[0092] (3) The frozen strain BoFeN1 was activated in an anaerobic medium (pH = 7.0). Specifically, the strain BoFeN1 stored in glycerol in a -80°C refrigerator was taken out and after thawing, it was injected into a sterile anaerobic medium with an inoculum volume fraction of 10% using a sterile syringe. The composition of the sterile anaerobic medium was as follows: 0.68 mmol·L -1 CaCl2·2H2O, 1.03mmol·L -1 KH2PO4, 2.07mmol·L -1 MgCl2·6H2O, 5.14mmol·L -1 NaCl, 5.60mmol·L -1 NH4Cl, 30mmol·L -1 PIPES, 5.0mmol·L -1 CH3COONa, 10.0mmol·L - 1 NaNO3; in a constant temperature incubator, culture at 30°C in the dark for 24 h to obtain an activated bacterial solution; transfer the activated bacterial solution to a new anaerobic culture medium in the same way, culture at 30°C in the dark for 20 h, centrifuge the obtained bacterial solution at 4°C for 5 min (8000 g), discard the supernatant, and use sterile PIPES buffer (30 mmol·L -1, pH = 7.0) and then centrifuged (8000g, 5min, 4°C) for 3 times to obtain bacterial cells; the bacterial cells were resuspended in PIPES buffer and mixed thoroughly to obtain a bacterial suspension, and the absorbance of the bacterial suspension at 600nm (OD 600 ) is 0.8-1.0, adjust the OD 600 The bacterial suspension was passed through nitrogen (filtered through a sterile filter) for 40 minutes to remove oxygen from the bacterial suspension, and the iron-oxidizing bacterial solution was obtained, which was placed in an anaerobic workstation for standby use;

[0093] (4) Add 500 mL of deionized water to a beaker, add a certain amount of sodium alginate to a mass fraction of 3.5%, and fully dissolve it; centrifuge the cultured iron-oxidizing bacterial solution obtained in step (2), resuspend it, and adjust the OD 600 =0.4, the cell concentration is about 4.5×10 7 / mL; add 7.5mL of iron oxidizing bacteria solution to every 17.5mL of cooled sodium alginate solution, and stir at a rate of 150rpm for 5min using a magnetic stirrer, then adjust the stirring speed to 220rpm, gradually add to the ceramsite matrix obtained in step (1) at a rate of 15g / min, add a total of 250g, continue stirring for 15min after the addition, and finally filter using a 10-mesh sieve to obtain a modified porous composite material.

[0094] The ferrous oxidation and nitrate reduction performance of the modified porous composite material of Example 2 was tested. The specific method was to construct a 50 mL neutral, anaerobic microcosm culture system in a 100 mL vial. The microcosm culture system included: ferrous chloride (5 mM), sodium nitrate (7 mM), and PIPES solution (30 mM, pH 7.0). The composite material of Example 2 was used as the experimental group, and control groups CK-1 and CK-2 were set up. Only 2.5 mL, OD 2.5 mL, was added to the control group CK-1. 600 The iron-oxidizing bacteria suspension with an OD of 0.4 had the same microbial biomass as that in the experimental group. In the control group CK-2, 1 g of unmodified ceramsite and 2.5 mL of OD 600 The composite material of Example 2 and the materials of the control groups CK-1 and CK-2 were added to the above microcosm culture system respectively, and the vials were placed in a constant temperature incubator and cultured in the dark at 30°C for 6 days. The concentration changes of ferrous iron and nitrate in the reaction system were measured at intervals of 24 hours.

[0095] The extraction and determination method of ferrous iron is as follows: ferrous iron in the above-mentioned culture system is extracted using aminosulfonic acid (40 mM, pH = 1.8) as an extractant, the vial cultured to a specific time point is placed in a sterile operating table, the sample is shaken, 200 μL of the mixed suspension is taken into a centrifuge tube containing 1800 μL of aminosulfonic acid, the centrifuge tube is placed in a shaker (30°C, 200 rpm), extracted at room temperature for 5 minutes, and then centrifuged at 4°C for 5 minutes (8000g); 100 μL of the supernatant is taken, 400 μL of aminosulfonic acid is added to adjust the pH value of the solution, and 500 μL of phenanthroline is added as a color developer; after the sample is shaken and colored, the absorbance of the sample at 562 nm is determined using an enzyme marker, and the ferrous iron concentration is calculated in combination with the standard curve.

[0096] The method for determining the nitrate concentration is as follows: mix the sample in the culture system in the above-mentioned vial, use a clean 5mL syringe to draw 1mL of the suspension, place it in a centrifuge tube containing 4mL of deionized water, and extract it on an oscillator for 5 minutes. The extraction parameters are light-proof, 30°C, and 200rpm; after extraction, centrifuge the centrifuge tube for 3 minutes, and the centrifugation parameters are 25°C and 6000rpm; after centrifugation, take the supernatant, filter it with a 0.22μm filter head, and then use an ion chromatograph to determine the nitrate in the filtrate, and use the nitrate mark as a benchmark to calculate the actual nitrate concentration in the extracted sample.

[0097] The ferrous oxidation results of the modified porous composite material of Example 2 and the control group material are as follows: Figure 5 As shown, the horizontal axis Time (d) represents time (days), the vertical axis Fe (II) mM represents the concentration of ferrous iron, and TL represents the modified porous composite material of Example 2. The nitrate reduction effect of the modified porous composite material of Example 2 and the control group material is shown as follows Figure 6 As shown, the horizontal axis Time (d) represents time (days), and the vertical axis [NO3 - ]mM represents the concentration of nitrate, and TL represents the modified porous composite material of Example 2.

[0098] Depend on Figure 5It can be seen that as the reaction proceeds, the ferrous iron concentration in the control group CK-1 drops to 2.82mM; in the control group CK-2, the ferrous iron concentration drops to 2.11mM; in the experimental group of the modified porous composite material in Example 2, the ferrous iron concentration drops to 1.42mM at the end of the reaction, and a total of 3.57mM ferrous iron is consumed, indicating that compared with the addition of the iron oxidizing bacteria suspension alone, and the addition of the iron oxidizing bacteria suspension and ceramsite separately, the modified porous composite material in Example 2 has a significant promoting effect on ferrous iron oxidation. The modified porous composite material will form iron minerals during the ferrous iron oxidation process, and iron minerals play a key role in the fixation of soil organic carbon, because iron minerals have a large surface area and highly active surface properties, which can effectively protect organic carbon. At present, up to 80% of the organic carbon in the world is preserved in combination with reactive iron. The iron minerals formed in the modified porous composite material during the ferrous iron oxidation process can combine with the functional groups (such as carboxyl, phenolic and amine groups) in organic carbon to form a stable structure, which can effectively avoid the degradation of organic carbon in the soil, thereby achieving a stable carbon fixation effect. It also shows that the porous structure of the ceramsite matrix and the calcium alginate gel layer structure in the modified porous composite material can effectively improve the biological activity of the iron oxidizing bacteria BoFeN1, promote ferrous iron oxidation, and thus improve the carbon fixation effect.

[0099] Depend on Figure 6 It can be seen that the control groups CK-1 and CK-2 showed similar nitrate reduction effects, the nitrate concentration decreased slowly, and the nitrate reduction ability was weak. In the experimental group of the modified porous composite material of Example 2, the nitrate concentration decreased rapidly, and by the end of the reaction, a total of 3.62 mM nitrate was reduced. This shows that the modified porous composite material of Example 2 can promote the reduction of nitrate.

[0100] Combination Figure 5 and Figure 6 This indicates that the modified porous composite material prepared by the present invention can effectively improve the biological activity of the iron oxidizing bacteria BoFeN1, thereby improving the carbon fixation effect.

[0101] Example 3

[0102] A method for preparing a modified porous composite material comprises the following steps:

[0103] (1) 0.25 kg of ceramsite with a diameter of 4-5 mm is placed in a rotary stirrer and mixed at a rate of 175 rpm for 2 h. After the stirring is completed, the ceramsite is placed on a 10-mesh sieve to screen out impurities left after friction; after the screening is completed, the ceramsite is placed in a 1L beaker, and a 1.0M hydrochloric acid solution and a magnetic rotor and rotor of a magnetic stirrer are added. The acid solution is sufficient to cover the ceramsite by about 5 cm. The magnetic stirrer is stirred at a rate of 220 rpm. After the ceramsite is acidified for 6 h, the ceramsite is taken out; it is cleaned with deionized water 5 times, and dried at 60° C. to obtain a ceramsite matrix for standby use;

[0104] (2) Add 500 mL of deionized water and 700 g of calcium chloride to a 1 L beaker, and perform ultrasonic treatment for 30 min to ensure that the calcium chloride solution is supersaturated and solids are precipitated at the bottom of the beaker; take 0.2 kg of the ceramsite matrix and magnet obtained in step (1), put them into the supersaturated calcium chloride solution, stir at a rate of 220 rpm for 30 min, take out the ceramsite matrix, place it in an oven and dry it at 85° C. to obtain a ceramsite matrix loaded with calcium chloride particles;

[0105] (3) The frozen strain BoFeN1 was activated in an anaerobic medium (pH = 7.0). Specifically, the strain BoFeN1 stored in glycerol in a -80°C refrigerator was taken out and after thawing, it was injected into a sterile anaerobic medium with an inoculum volume fraction of 10% using a sterile syringe. The composition of the sterile anaerobic medium was as follows: 0.68 mmol·L -1 CaCl2·2H2O, 1.03mmol·L -1 KH2PO4, 2.07mmol·L -1 MgCl2·6H2O, 5.14mmol·L -1 NaCl, 5.60mmol·L -1 NH4Cl, 30mmol·L -1 PIPES, 5.0mmol·L -1 CH3COONa, 10.0mmol·L - 1 NaNO3; in a constant temperature incubator, culture at 30°C in the dark for 24 h to obtain an activated bacterial solution; transfer the activated bacterial solution to a new anaerobic culture medium in the same way, culture at 30°C in the dark for 20 h, centrifuge the obtained bacterial solution at 4°C for 5 min (8000 g), discard the supernatant, and use sterile PIPES buffer (30 mmol·L -1, pH = 7.0) and then centrifuged (8000g, 5min, 4°C) for 3 times to obtain bacterial cells; the bacterial cells were resuspended in PIPES buffer and mixed thoroughly to obtain a bacterial suspension, and the absorbance of the bacterial suspension at 600nm (OD 600 ) is 0.8-1.0, adjust the OD 600 The bacterial suspension was passed through nitrogen (filtered through a sterile filter) for 40 minutes to remove oxygen from the bacterial suspension, and the iron-oxidizing bacterial solution was obtained, which was placed in an anaerobic workstation for standby use;

[0106] (4) Add 500 mL of deionized water to a beaker, add a certain amount of sodium alginate to a mass fraction of 3.5%, and fully dissolve it; centrifuge the cultured iron-oxidizing bacterial solution obtained in step (2), resuspend it, and adjust the OD 600 =0.4, the cell concentration is about 4.5×10 7 / mL; add 7.5mL of iron oxidizing bacteria solution to every 17.5mL of cooled sodium alginate solution, and stir at a rate of 150rpm for 5min using a magnetic stirrer, then adjust the stirring speed to 220rpm, gradually add to the ceramsite matrix obtained in step (1) at a rate of 15g / min, add a total of 250g, continue stirring for 15min after the addition, and finally filter using a 10-mesh sieve to obtain a modified porous composite material.

[0107] The effect of the composite material of Example 3 on the changes in nitrous oxide and organic carbon concentrations was tested. The specific method was as follows: 80 mL of soil simulation solution was added to a 150 mL syringe bottle to construct a microcosm culture system; the main component of the soil simulation solution was oxygen-free borax-boric acid buffer, wherein the concentration of the borax-boric acid buffer was 50 mmol / L. The initial dissolved organic carbon concentration of the soil simulation solution was 70 mg / L, which was provided by fulvic acid solution (sterile); in addition, 7 mM nitrate and 5 mM ferrous iron were added to the soil simulation solution. The composite material of Example 3 was used as the experimental group, and a control group CK was set up. Only 2.5 mL, OD 600 The iron oxidizing bacteria suspension with a value of 0.4, the corresponding iron oxidizing bacteria microbial biomass is consistent with that in the experimental group. The composite material of Example 3 and the bacterial solution of the control group CK were added to the vials respectively, and the vials were sealed and shaken, and then the two groups of vials (three each) were placed in a constant temperature incubator and cultured at 28°C, and the concentration changes of nitrous oxide in the reaction system and organic carbon in the solution were measured every 24 hours.

[0108] Among them, the method for determining the nitrous oxide concentration is as follows: the nitrous oxide gas sample is tested before the liquid sample is sampled. Before sampling, the vial is shaken and placed in a dark place for 15 minutes; a clean 1mL syringe is taken, the syringe is cleaned with high-purity nitrogen, and 0.8mL of the headspace gas in the vial is drawn; after sampling, a gas chromatograph is used for measurement, and the final content is determined according to the nitrous oxide mark.

[0109] The method for determining the concentration of dissolved organic carbon is as follows: mix the sample in the vial, use a pipette to draw 10 mL of the suspension into a 15 mL centrifuge tube in an anaerobic chamber, and centrifuge the centrifuge tube for 10 minutes at 25°C and 8000g. After centrifugation, filter the supernatant into a new 15 mL centrifuge tube using a 0.22 μm filter head and add 10 μL HCl (6M) for storage. After sampling, the sample is measured using a TOC analyzer, which is the dissolved organic carbon content.

[0110] Example 3 The effect of the modified porous composite material and the control group material on the change of nitrous oxide concentration is shown in the following results. Figure 7 As shown, the abscissa Time (d) represents time (day), the ordinate N2O (mg / L) represents the concentration of nitrous oxide, and TL represents the modified porous composite material of Example 3.

[0111] Example 3 The effect of the modified porous composite material and the control group material on the concentration change of dissolved organic carbon is shown in the following table. Figure 8 As shown, the abscissa Time (d) represents time (day), the ordinate DOC (mg / L) represents the concentration of dissolved organic carbon, and TL represents the modified porous composite material of Example 3.

[0112] Depend on Figure 7 It can be seen that in the control group where only the iron-oxidizing bacteria suspension was added, the generation rate of nitrous oxide was faster; while in the experimental group of the composite material of Example 3, the generation of nitrous oxide was relatively slow, and the total amount of nitrous oxide generated was low. This experimental result shows that the modified porous composite material can effectively inhibit the generation of nitrous oxide in the process of ferrous iron oxidation coupled with nitrate reduction by iron-oxidizing bacteria, which has a positive effect on reducing greenhouse gas emissions in the process of nitrate reduction coupled with ferrous iron oxidation by iron-oxidizing bacteria.

[0113] Depend on Figure 8It can be seen that for the control group CK to which only the iron-oxidizing bacteria suspension was added, the DOC concentration started from about 65 mg / L on the 0th day, and gradually decreased over time, and dropped to about 25 mg / L on the 4th day. For the experimental group TL, the DOC concentration dropped sharply to about 25 mg / L on the 1st day, and then remained relatively stable in the next few days until it dropped to about 15 mg / L on the 4th day. Throughout the experiment, the DOC concentration of the TL group was always lower than that of the CK group, which shows that the modified porous composite material of the present invention has a significant effect on reducing the DOC concentration, and also shows that the modified porous composite material of the present invention has a positive effect on the fixation of soil organic carbon.

[0114] The ferrous oxidation and nitrate reduction performance of the composite material of Example 1 of the present invention is equivalent to that of Example 2, and the effect on the change of nitrous oxide and organic carbon concentration is equivalent to that of Example 3. The morphology, the property of maintaining the gel structure, and the iron-oxidizing bacteria cell crusting phenomenon of the composite material of Example 2 are equivalent to those of Example 1, and the effect on the change of nitrous oxide and organic carbon concentration is equivalent to that of Example 3. The morphology, the property of maintaining the gel structure, and the iron-oxidizing bacteria cell crusting phenomenon of the composite material of Example 3 are equivalent to those of Example 1, and the ferrous oxidation and nitrate reduction performance are equivalent to those of Example 2.

[0115] In summary, the composite material with a specific structure of the present invention can improve the activity of microorganisms, has a significant promoting effect on ferrous oxidation, and can effectively avoid the degradation of soil organic carbon, thereby achieving a stable carbon fixation effect; at the same time, it can effectively inhibit the generation of nitrous oxide in the process of ferrous oxidation coupled with nitrate reduction by iron-oxidizing bacteria, reduce greenhouse gas emissions in the process of nitrate reduction coupled with ferrous oxidation by iron-oxidizing bacteria, reduce DOC concentration, and achieve good carbon fixation and emission reduction effects.

[0116] The above embodiments are only used to illustrate the technical solution of the present invention rather than to limit the protection scope of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solution of the present invention can be modified or replaced by equivalents without departing from the essence and scope of the technical solution of the present invention.

Claims

1. A composite material, characterized in that: Including modified ceramsite and microorganisms; The modified ceramsite comprises a ceramsite matrix and a calcium alginate gel layer; The ceramsite matrix has a porous structure; the calcium alginate gel layer is loaded on the surface and inside of the ceramsite matrix, and the calcium alginate gel layer covers the microorganism on the surface, inside of the ceramsite matrix and in the calcium alginate gel layer.

2. The composite material according to claim 1, characterized in that The ceramsite matrix is ​​acidified ceramsite.

3. The composite material according to claim 1, characterized in that The mass ratio of the ceramsite matrix to the calcium alginate gel layer is (1.8-3.3):

1.

4. The composite material according to claim 1, characterized in that The microorganisms include iron oxidizing bacteria.

5. The method for preparing the composite material according to any one of claims 1 to 4, characterized in that: The following steps are involved: (1) mixing a ceramsite matrix, a calcium salt, and a solvent to obtain a ceramsite matrix loaded with calcium salt; mixing an alginate solution and a microbial culture liquid to obtain a mixture; (2) mixing the calcium salt-loaded ceramsite matrix obtained in step (1) with the mixture to obtain the composite material.

6. The preparation method according to claim 5, characterized in that: The preparation method of the ceramsite matrix comprises the following steps: The ceramsite and acid solution are mixed and acidified to obtain the ceramsite matrix.

7. The preparation method according to claim 6, characterized in that: The concentration of the acid solution is 0.8-1.2M; and / or the acid solution includes hydrochloric acid; and / or the time of the acidification treatment is 4.5-6.5h.

8. The preparation method according to claim 5, characterized in that: In step (1), the mass ratio of the ceramsite matrix to the calcium salt is 1:(3.0-4.0); and / or the calcium salt includes calcium chloride; and / or the alginate solution includes sodium alginate solution; and / or the volume ratio of the alginate solution to the microbial culture liquid is 1:(0.25-0.45); and / or the cell concentration in the microbial culture liquid is (3.0-5.5)×10 7 Pieces / mL.

9. The preparation method according to claim 5, characterized in that: In step (2), the amount ratio of the calcium salt-loaded ceramsite matrix to the alginate solution in the mixture is 7-13 g: 1 mL; and / or, the mixing is performed by stirring, the stirring speed is 180-270 rpm, and the stirring time is 13-18 min.

10. Use of the composite material according to any one of claims 1 to 4 in soil improvement.

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