阅读说明：本技术 一种土壤微生物原位合成纳米磁铁矿的方法 (Method for in-situ synthesis of nano magnetite from soil microorganisms ) 是由 李田 王鑫 田丽丽 于 2020-11-06 设计创作，主要内容包括：本发明公开一种土壤微生物原位合成纳米磁铁矿的方法,该方法针对土壤传质能力较差等问题,步骤主要包括：土壤微生物的富集、纳米磁铁矿的微生物合成以及纳米磁铁矿的测定。其中土壤微生物的富集主要通过土壤浸提液提取、微生物定向富集等步骤完成,纳米磁铁矿的微生物合成主要是通过前驱体的制备和微生物电子供体的添加等步骤完成,纳米磁铁矿的测定主要是通过盐酸消解测定其中铁离子含量,红外光谱、X射线衍射图谱、透射电镜等步骤完成。该方法操作简单、纳米磁铁矿在土壤中可原位形成且分布均匀,为污染土壤的修复提供了一种新思路。(The invention discloses a method for in-situ synthesis of nano magnetite by soil microorganisms, which aims at the problems of poor mass transfer capacity of soil and the like and mainly comprises the following steps: enrichment of soil microorganisms, microbial synthesis of nano magnetite and determination of nano magnetite. The method mainly comprises the steps of soil leaching liquor extraction, microorganism directional enrichment and the like, the microorganism synthesis of the nano magnetite is mainly completed through the steps of precursor preparation, microorganism electron donor addition and the like, and the determination of the nano magnetite is mainly completed through the steps of hydrochloric acid digestion determination of iron ion content, infrared spectroscopy, X-ray diffraction pattern, transmission electron microscope and the like. The method is simple to operate, and the nano magnetite can be formed in situ in the soil and is uniformly distributed, so that a new idea is provided for the remediation of the polluted soil.)

1. A method for in-situ synthesis of nano magnetite by soil microorganisms is characterized by comprising the following steps:

1) enrichment of soil microorganisms

Weighing 1000-2000g of the organic-inorganic composite material into an acrylic conical leaching solution reactor, wrapping a mesh filter sieve outside the reactor, connecting the lower part of the reactor with an anaerobically sealable leaching solution collector, injecting water flow into the collector, stirring soil deposition liquid, and repeating the whole process for 2-3 times to complete the extraction of the soil leaching solution;

adding sodium acetate and ferric citrate into the extracted soil leaching liquor, performing anaerobic sealing treatment, culturing soil microorganisms and directionally enriching reducible iron microorganisms under the condition of constant temperature, periodically extracting solution, measuring optical density value in the solution, and determining microorganism enrichment effect;

2) microbial synthesis of nano magnetite

Weighing 2000-2500g of farmland soil into the soil reactor, extracting microorganisms from the leaching solution after the soil microorganisms are completely enriched, uniformly mixing the microorganisms with the farmland soil in the soil reactor, adding ferric hydroxide and sodium acetate into the mixed farmland soil, uniformly mixing again, then gradually injecting sterile water until the liquid seal is 3-5cm, and carrying out the in-situ synthesis process of the nano magnetite of the soil microorganisms under the constant temperature condition;

3) determination of Nano Magnetite

In the process of synthesizing the nano magnetite, extracting soil from different positions of a soil reactor by using a sampler every day, adding hydrochloric acid for digestion, and then measuring Fe in the solution²⁺And (3) freeze-drying the extracted soil overnight, then carrying out infrared spectrum test to analyze elements in the soil, and determining that the synthesized substance is the nano magnetite by combining an X-ray spectrum, a transmission electron microscope and the like.

2. The method for in-situ synthesis of nano magnetite from soil microorganisms according to claim 1, wherein the flow rate of the water stream in step 1) is 5-10mL/h, the content of sodium acetate added is 4-5g/L, and the content of ferric citrate added is 1-2 g/L.

3. The method for in-situ synthesis of nano magnetite from soil microorganisms according to claim 1, wherein the extraction microorganisms of step 2) are mainly obtained by passing the leaching solution through a 1 μm filter membrane, the concentration of ferric hydroxide is 100mM, the concentration of sodium acetate is 4-5g/L, and the constant temperature condition is 30 ℃.

4. The method for in situ synthesis of nano-magnetite from soil microorganisms according to claim 1, wherein the hydrochloric acid in step 3) is prepared from 1 volume of water and 3 volumes of concentrated hydrochloric acid, and the freeze drying time is 12 h.

Technical Field

The invention belongs to the field of environmental microorganism geochemistry, and particularly relates to a method for in-situ synthesis of nano magnetite by soil microorganisms.

Background

With the acceleration of the industrialization process, the environmental pollution problem becomes an important factor restricting the economic development, and the serious threat to the survival and development of human beings is generated. Heavy metal and organic matter pollution are two types of pollution common in soil and water environments, microorganisms complete the recovery of heavy metals and the degradation process of organic matters in the two media through an oxidation-reduction process, but compared with the water environment, the mass transfer capacity of the soil environment is poor, the oxidation-reduction process of the microorganisms is greatly hindered, and the removal efficiency of the pollutants is greatly reduced.

Iron (Fe) is an important high-activity and high-abundance redox element in soil, and the biogeochemical cycle of the iron (Fe) is directly related to environmental processes such as soil organic matter mineralization, denitrification, heavy metal fixation and the like. Fe can perform Fe (III) reduction and Fe (II) oxidation processes in the electron transfer process of anaerobic microorganisms, and exists in soil in the form of iron-containing minerals, and the iron-containing minerals can act with extracellular polymers of the microorganisms to participate in the extracellular electron transfer process to accelerate the reduction of nitrate nitrogen, biological electricity generation, interspecies mutualism and the like. Research shows that the electroactive microorganisms can utilize magnetite to complete an interspecific electron transfer process so as to realize mutual nutrient enrichment and microbial metabolism. However, in soil, magnetite and other conductive minerals are often wrapped by organic matters or pollutants and are difficult to act as electronic mediators, and the repair cost is increased by adding magnetite from an external source. Therefore, if nano magnetite can be synthesized in situ in soil by using microorganisms, the removal efficiency of pollutants in soil can be greatly improved.

Disclosure of Invention

The invention aims to provide a method for in-situ synthesis of nano magnetite by soil microorganisms aiming at poor mass transfer in soil and severe influence on the efficiency of repairing soil by the microorganisms.

The invention is realized by the following steps:

1) enrichment of soil microorganisms

Grinding and sieving the collected farmland soil, weighing 1000-2000g of acrylic material conical leaching liquor reactor with the bottom surface diameter of 10cm and the height of 15cm, wrapping a net-shaped filter sieve outside, connecting the lower part of the reactor with an anaerobically sealable leaching liquor collector, injecting water flow with a certain flow rate into the collector, stirring soil deposition liquid by a glass rod every certain time, and repeating the whole process for 2-3 times to complete the extraction of the soil leaching liquor;

adding a certain amount of sodium acetate and ferric citrate into the extracted soil leaching liquor, carrying out anaerobic sealing treatment, carrying out culture of soil microorganisms and microorganism directional enrichment of reducible iron under the constant temperature condition, periodically extracting solution, measuring optical density value therein, and determining microorganism enrichment effect.

2) Microbial synthesis of nano magnetite

Weighing 2000-2500g of farmland soil into the soil reactor, extracting microorganisms from the leaching solution after the soil microorganisms are completely enriched, uniformly mixing the microorganisms with the farmland soil in the soil reactor, adding a certain amount of ferric hydroxide and sodium acetate into the mixed farmland soil, uniformly mixing again, then gradually injecting sterile water until the liquid seal is 3-5cm, and carrying out the in-situ synthesis process of the nano magnetite of the soil microorganisms under the constant temperature condition.

3) Determination of Nano Magnetite

In the process of synthesizing the nano magnetite, 2g of soil is extracted from different positions of a soil reactor by using a sampler every day, 3mL of hydrochloric acid with certain concentration is added for digestion for 8 hours, and then Fe in the solution is measured²⁺And (3) freeze-drying the extracted soil overnight, then carrying out infrared spectrum test to analyze elements in the soil, and determining that the synthesized substance is the nano magnetite by combining an X-ray spectrum, a transmission electron microscope and the like.

The flow rate of the water flow in the step 1) is 5-10mL/h, the content of the added sodium acetate is 4-5g/L, and the content of the added ferric citrate is 1-2 g/L.

The extraction of the microorganism in the step 2) is mainly to pass the leaching liquor through a filter membrane with the diameter of 1 mu m, the concentration of ferric hydroxide is 100mM, the concentration of sodium acetate is 4-5g/L, and the constant temperature condition is 30 ℃.

The hydrochloric acid in the step 3) is prepared from 1 volume of water and 3 volumes of concentrated hydrochloric acid, and the freeze drying time is 12 hours.

The invention has the advantages and beneficial effects that:

the invention realizes the process of in-situ synthesis of the nano magnetite by using indigenous microorganisms in the soil for the first time, greatly improves the mass transfer capacity of the soil environment and provides a new idea for soil remediation.

Drawings

FIG. 1 shows a diagram of a soil leaching solution installation: 1 is a conical leaching liquor reactor, 2 is a mesh filter sieve, 3 is a leaching liquor collector, and 4 is a one-way exhaust valve.

FIG. 2 is a schematic diagram of a microorganism synthesis device for nano magnetite: 1 is a soil reactor, 2 is a liquid seal area, 3 is a soil area, and 4 is a microorganism synthesis schematic diagram.

FIG. 3 is a graph showing the change of the content of the biosynthetic nano magnetite.

FIG. 4 is a transmission electron microscope image of the biosynthetic nano magnetite.

FIG. 5X-ray diffraction pattern of biosynthetic nano magnetite.

FIG. 6 is an infrared spectrum of the biosynthesized nano magnetite.

FIG. 7 is a graph of the synthetic content of nano-magnetite without the microbial enrichment step.

FIG. 8 is a graph showing the change in optical density upon enrichment with microorganisms.

FIG. 9 is a graph showing the synthetic content of nano magnetite in soil without iron hydroxide.

Detailed Description

One, experiment group

1) Enrichment of soil microorganisms

Grinding and sieving the harvested farmland soil, and then weighing 1000g of acrylic conical leaching liquor reactor with the bottom surface diameter of 10cm and the height of 15cm, wrapping a net-shaped filter sieve outside the reactor, connecting the lower part of the reactor with an anaerobically-sealed leaching liquor collector, injecting 5mL/h of water flow into the collector, stirring the soil deposition solution by a glass rod every certain time, and repeating the whole process for 2-3 times to complete the extraction of the soil leaching liquor, wherein the device diagram is shown in figure 1;

adding 4g/L sodium acetate and 1g/L ferric citrate into the extracted soil leaching liquor, carrying out anaerobic sealing treatment, carrying out culture of soil microorganisms and microorganism directional enrichment of reducible iron at the constant temperature of 30 ℃, periodically extracting the solution, measuring the optical density value in the solution, and determining the microorganism enrichment effect.

2) Microbial synthesis of nano magnetite

Weighing 2000g of farmland soil into a soil reactor, extracting microorganisms from the leaching liquor by using a filter membrane of 1 mu m after the enrichment of soil microorganisms is finished, uniformly mixing the microorganisms with the farmland soil in the soil reactor, adding 100mM of ferric hydroxide and 4g/L of sodium acetate into the mixed farmland soil, uniformly mixing again, then gradually injecting sterile water until the liquid seal is 3cm, and carrying out the in-situ synthesis process of the nano magnetite of the soil microorganisms at the constant temperature of 30 ℃, wherein the device and the synthesis principle are shown in figure 2.

3) Determination of Nano Magnetite

In the process of synthesizing the nano magnetite, 2g of soil is extracted from different positions of a soil reactor by using a sampler every day, 3mL of special hydrochloric acid with the mixture ratio of 1 volume of water to 3 volumes of concentrated hydrochloric acid is added for digestion for 8 hours, and then Fe in the solution is measured²⁺The concentration and the content change of the nano magnetite are shown in figure 3, the synthesis of the nano magnetite is completed within 13 days, the extracted soil is frozen and dried for 12 hours, then the infrared spectrum test is carried out to analyze elements in the soil, the synthetic substance is determined to be the nano magnetite by combining an X-ray spectrum, a transmission electron microscope and the like, and as shown in figures 4-6, the comprehensive characterization result shows that the nano magnetite is synthesized in situ by indigenous microorganisms in the soil.

II, control group 1

1) Microbial synthesis of nano magnetite

2200g of farmland soil is weighed into a soil reactor, 100mM ferric hydroxide and 4g/L sodium acetate are added into the mixed farmland soil to be uniformly mixed again, then sterile water is gradually injected until the liquid seal is 3cm, the in-situ synthesis process of the nano magnetite of the soil microorganism is carried out under the condition of constant temperature of 30 ℃, and the device and the synthesis principle are shown in figure 2.

2) Determination of Nano Magnetite

In the process of synthesizing the nano magnetite, 2g of soil is extracted from different positions of a soil reactor by using a sampler every day, 1 volume of water and 3 bodies are added3mL of special hydrochloric acid with concentrated hydrochloric acid ratio is dissolved for 8h, and then Fe in the solution is measured²⁺The concentration and the content change are shown in FIG. 7, and the synthesis of the nano magnetite is completed in about 32 days.

II, control group 2

1) Enrichment of soil microorganisms

Grinding and sieving the harvested farmland soil, weighing 2000g of acrylic conical leaching liquor reactor with the bottom surface diameter of 10cm and the height of 15cm, wrapping a net-shaped filter sieve outside the reactor, connecting the lower part of the reactor with an anaerobically-sealed leaching liquor collector, injecting 10mL/h of water flow into the collector, stirring the soil deposition liquor by a glass rod every certain time, and repeating the whole process for 2-3 times to complete extraction of the soil leaching liquor, wherein the device is shown in figure 1;

adding 5g/L sodium acetate and 2g/L ferric citrate into the extracted soil leaching solution, performing anaerobic sealing treatment, culturing soil microorganisms and directionally enriching reducible iron microorganisms at constant temperature of 30 ℃, periodically extracting the solution, measuring the optical density value of the solution, and determining the microorganism enrichment effect, as shown in figure 8.

2) Microbial synthesis of nano magnetite

And then 2500g of farmland soil is weighed into the soil reactor, after the soil microorganism enrichment is finished, microorganisms are extracted from the leaching liquor by using a filter membrane with the diameter of 1 mu m, the microorganisms are uniformly mixed with the farmland soil in the soil reactor, 5g/L of sodium acetate is added into the mixed farmland soil for uniform mixing again, then sterile water is gradually injected until the liquid seal is 3cm, the in-situ synthesis process of the nano magnetite of the soil microorganism is carried out under the condition of constant temperature of 30 ℃, and the device and the synthesis principle are shown in figure 2.

3) Determination of Nano Magnetite

In the process of synthesizing the nano magnetite, 2g of soil is extracted from different positions of a soil reactor by using a sampler every day, 3mL of special hydrochloric acid with the mixture ratio of 1 volume of water to 3 volumes of concentrated hydrochloric acid is added for digestion for 8 hours, and then Fe in the solution is measured²⁺The concentration, the content change of which is shown in fig. 9, almost no nano magnetite was synthesized.