CN110303040B - In-situ restoration agent for efficiently degrading tetracycline antibiotics in soil and preparation method thereof - Google Patents
In-situ restoration agent for efficiently degrading tetracycline antibiotics in soil and preparation method thereof Download PDFInfo
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- CN110303040B CN110303040B CN201910715643.7A CN201910715643A CN110303040B CN 110303040 B CN110303040 B CN 110303040B CN 201910715643 A CN201910715643 A CN 201910715643A CN 110303040 B CN110303040 B CN 110303040B
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
- B09C—RECLAMATION OF CONTAMINATED SOIL
- B09C1/00—Reclamation of contaminated soil
- B09C1/08—Reclamation of contaminated soil chemically
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
- B09C—RECLAMATION OF CONTAMINATED SOIL
- B09C1/00—Reclamation of contaminated soil
- B09C1/10—Reclamation of contaminated soil microbiologically, biologically or by using enzymes
- B09C1/105—Reclamation of contaminated soil microbiologically, biologically or by using enzymes using fungi or plants
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
- B09C—RECLAMATION OF CONTAMINATED SOIL
- B09C2101/00—In situ
Abstract
The invention discloses an in-situ restoration agent for efficiently degrading tetracycline antibiotics in soil and a preparation method thereof, and is characterized by comprising the following components in parts by weight: Pt/TiO 2230-40 parts of modified porous silkworm excrement, 2-6 parts of graphene, 1-5 parts of cation exchange resin, 4-6 parts of methylobacterium and 4-6 parts of pseudomonas syriacus; the methylobacterium and the pseudomonas syriacus are respectively obtained by continuously carrying out passage domestication on original strains in a basic solid culture medium containing tetracycline; the Pt/TiO2The preparation process of the modified porous silkworm excrement comprises the following steps: swelling original silkworm excrement in 6-12% urea solution, drying, carbonizing at 200 ℃ for 1-2h under the action of protective gas, adding 0.1-1% bromoplatinic acid solution and tetrabutyl titanate diluent for quenching, heating to 450 ℃ and 500 ℃, and roasting for 3-5 h. The method can directly carry out in-situ remediation on the soil polluted by the tetracycline antibiotics, has short period, combines the means of physical adsorption, photocatalytic degradation, biodegradation and the like to carry out in-situ removal, and has high conversion rate.
Description
Technical Field
The invention belongs to the technical field of contaminated soil remediation, and particularly relates to an in-situ remediation agent capable of efficiently degrading tetracycline antibiotics in soil and a preparation method thereof.
Background
The tetracycline antibiotics are a broad-spectrum antibiotics produced by actinomycetes, and include aureomycin, oxytetracycline, tetracycline and semisynthetic derivatives of methacycline, doxycycline, dimethylamino tetracycline, and the like, and the structures of the antibiotics all contain tetracene basic skeleton. The action mechanism of the tetracycline antibiotics is that the tetracycline antibiotics enter cells from an energy-dependent transfer system on an inner membrane through hydrophilic pores on an outer membrane of the cells, are specifically combined with ribosome 30S subunits, and prevent aminoacyl-tRNA from being linked, so that peptide chain elongation and protein synthesis are inhibited, and meanwhile, the permeability of bacterial cell membranes can be changed, so that important components in cells leak out, DNA replication is rapidly inhibited, and the growth of the bacteria is inhibited.
Tetracycline is the most used antibiotic in all countries of the world due to low price, and in 2014, the tetracycline antibiotic accounts for 43% of the antibiotics used in animal husbandry in the United states. Because the tetracycline is relatively stable in property and is difficult to degrade and absorb after entering an organism, more than 80 percent of the tetracycline is discharged into the soil environment in the form of a parent compound and continuously accumulated to induce microorganisms in the soil to gradually generate resistance, so that the enrichment of drug-resistant flora and the mutagenesis and generation of drug-resistant genes are caused to destroy the soil environment, and meanwhile, the tetracycline antibiotics accumulated in the soil are absorbed by crops and enter a human body through a food chain to harm the health of the human body. In the prior art, the method mainly aims at repairing and degrading the water body polluted by tetracycline antibiotics, and for example, a plant repairing method for tetracycline polluted wastewater is disclosed in a patent with the application number of CN 201310011876.1; for example, patent No. CN20-1610299434.5 discloses a method for preparing a hierarchical porous carbon material for adsorbing tetracycline antibiotics; for example, patent with application number CN201710610505.3 discloses a preparation method and application of a tetracycline polluted water body remediation material; for example, patent with application number CN20181-10103963.2 discloses a method for degrading tetracycline in wastewater by photo-response promoted oxidation free radicals; for example, patent with application number CN201810106340.0 discloses a preparation method and application of chitosan magnetic tetracycline molecular imprinting adsorbent; however, research on the remediation of soil polluted by tetracycline antibiotics is less and less intensive, and microbial agents are mainly used for degradation, but a single biological method has a long treatment period, such as a patent with application number CN201410-348058.5, which discloses a mycorrhizal fungi agent and an application thereof in the degradation of residual tetracycline antibiotics; for example, patent with application number of CN201710115130.3 discloses a mixed microbial inoculum for degrading antibiotics in soil and a preparation method thereof; for example, patent with application number of CN2017102000-22.6 discloses a strain of tetracycline degrading bacteria and application thereof in livestock and poultry manure composting treatment.
Based on the statement, the in-situ restoration agent for rapidly and efficiently degrading the tetracycline antibiotics in the soil has extremely wide market prospect and application value.
Disclosure of Invention
Aiming at the defects of the prior art, the invention aims to provide an in-situ restoration agent for efficiently degrading tetracycline antibiotics in soil and a preparation method thereof.
The technical scheme of the invention is summarized as follows:
an in-situ restoration agent for efficiently degrading tetracycline antibiotics in soil comprises the following components in parts by weight: Pt/TiO 2230-40 parts of modified porous silkworm excrement, 2-6 parts of graphene, 1-5 parts of cation exchange resin, 4-6 parts of methylobacterium and 4-6 parts of pseudomonas syriacus; the Pt/TiO2The modified porous silkworm excrement is prepared by swelling and carbonizing original silkworm excrement by urea, quenching the original silkworm excrement by bromoplatinic acid and tetrabutyl titanate, and finally roasting the original silkworm excrement at the temperature of 450-DEG C and 500 ℃; the methylobacterium and the pseudomonas syriacus are respectively obtained by continuously carrying out passage domestication on original strains in a basic solid culture medium containing tetracycline.
Preferably, the Pt/TiO2The preparation process of the modified porous silkworm excrement is as follows: soaking silkworm excrement in 6-12% urea solution for 3-6 hr to swell the silkworm excrement fully, drying at 80 deg.c to dewater, carbonizing at 200 deg.c under the protection of mixed gas of nitrogen and hydrogen for 1-2 hr, adding 0.1-1% bromoplatinic acid solution and tetrabutyl titanate diluent for quenching, heating to 500 deg.c and roasting for 3-5 hr, and naturally cooling to obtain Pt/TiO2Modified porous silkworm excrement.
Preferably, the volume ratio of the nitrogen gas to the hydrogen gas is 1: (0.1-0.2).
Preferably, the tetrabutyl titanate diluent is a mixed solution of tetrabutyl titanate and absolute ethyl alcohol, and the volume ratio of the tetrabutyl titanate to the absolute ethyl alcohol is 1: (3-4).
Preferably, the mass ratio of the silkworm excrement to the urea solution to the bromoplatinic acid solution to the tetrabutyl titanate diluent is 1: 2: 0.3: 0.1.
preferably, the passage acclimation process: inoculating methylobacterium protospecies or pseudolaricis protospecies in a freezing storage tube to a basic solid culture medium containing 150mg/L tetracycline, culturing for 2-4 days at 25-37 ℃, screening out tolerant live bacteria, transferring into a basic solid culture medium containing 300mg/L tetracycline for subculture, repeating for 4 times, and increasing the concentration of the tetracycline by 150mg/L each time, namely the final concentration of the tetracycline is 750mg/L, thereby completing the subculture.
Preferably, the subculture temperature is 25-35 ℃ and the number of days is 1-3 d.
High-efficiency degradationThe preparation method of the in-situ remediation agent of the tetracycline antibiotics in the soil comprises the following steps: mixing Pt/TiO according to the weight portion2Modified porous silkworm excrement, graphene, cation exchange resin, methylobacterium and pseudomonas syriacus to obtain the in-situ repairing agent.
The invention has the beneficial effects that:
(1) the repairing agent produced by the invention can directly carry out in-situ repair on the soil polluted by tetracycline antibiotics, and has the advantages of simple method, convenient operation, short repairing period, high efficiency, strong broad spectrum and easy popularization;
(2) the invention is beneficial to the rapid recovery of the functional diversity and stability of microorganisms in the polluted soil, is environment-friendly and does not cause secondary pollution;
(3) the invention can realize the high-efficient degradation of tetracycline antibiotics and residual pesticides simultaneously, decompose and remove the antibiotics and organic pollutants in the soil by utilizing three means of physical adsorption, photocatalytic degradation, biodegradation and the like, has high conversion rate, and adsorbs and removes heavy metal ions in the soil;
(4) the invention first utilizes Pt/TiO2The microporous structure of the modified porous silkworm excrement adsorbs tetracycline antibiotics, organic pollutants and heavy metals in soil, and Pt supported by the silkworm excrement is utilized4+Nano Pt and nano TiO2The adsorbed harmful substances are photolyzed in cooperation with graphene, and macromolecular compounds are finally converted into CO through free radical peroxide and singlet oxygen generated by excitation2And H2And O, meanwhile, the domesticated methylobacterium and the pseudomonas hibiscus have strong stress resistance and are rich in various degrading enzyme systems, so that the biological degradation of tetracycline antibiotics is realized, the structure and the physicochemical properties of the tetracycline antibiotics are changed, and the soil environment is improved.
Detailed Description
The present invention is further described in detail below with reference to examples so that those skilled in the art can practice the invention with reference to the description.
The invention provides an implemented in-situ restoration agent for efficiently degrading tetracycline antibiotics in soil, which comprises the following components in parts by weight: Pt/TiO 2230-40 parts of modified porous silkworm excrement, 2-6 parts of graphene, 1-5 parts of cation exchange resin, 4-6 parts of methylobacterium and 4-6 parts of pseudomonas syriacus.
The in-situ restoration agent utilizes triple means such as physical adsorption, photocatalytic degradation and biodegradation to efficiently degrade and remove tetracycline antibiotics and residual pesticides in soil, and the specific action mechanism is as follows: first, Pt/TiO is utilized2The microporous structure of the modified porous silkworm excrement adsorbs tetracycline antibiotics, organic pollutants and heavy metals in soil, and Pt supported by the silkworm excrement is utilized4+Nano Pt and nano TiO2The adsorbed harmful substances are photolyzed in cooperation with graphene, and macromolecular compounds are finally converted into CO through free radical peroxide and singlet oxygen generated by excitation2And H2And O, meanwhile, the domesticated methylobacterium and the pseudomonas hibiscus have strong stress resistance and are rich in various degrading enzyme systems, so that the biological degradation of tetracycline antibiotics is realized, the structure and the physicochemical properties of the tetracycline antibiotics are changed, and the soil environment is improved.
The above Pt/TiO2The modified porous silkworm excrement is prepared by swelling and carbonizing original silkworm excrement by urea, quenching the original silkworm excrement by bromoplatinic acid and tetrabutyl titanate, and finally roasting the original silkworm excrement at the temperature of 450-500 ℃, and the specific preparation process is as follows: according to the following steps: 2: 0.3: weighing raw silkworm excrement, urea solution, bromoplatinic acid solution and tetrabutyl titanate diluent according to the mass ratio of 0.1, wherein the tetrabutyl titanate diluent is prepared from tetrabutyl titanate and absolute ethyl alcohol according to the weight ratio of 1: (3-4) mixing the raw silkworm excrement with a mixed solution composed of 6-12% of urea solution according to the volume ratio, soaking for 3-6h to fully swell the silkworm excrement, drying and dehydrating at 80 ℃, and then mixing the raw silkworm excrement with the urea solution according to the volume ratio of 1: (0.1-0.2) under the protection of mixed gas of nitrogen and hydrogen, carbonizing at 180-200 ℃ for 1-2h, adding 0.1-1% bromoplatinic acid solution and tetrabutyl titanate diluent for quenching, heating to 450-500 ℃, roasting for 3-5h, and naturally cooling to obtain Pt/TiO2Modified porous silkworm excrement.
The methylobacterium and the pseudomonas hibiscus are respectively obtained by continuously carrying out passage domestication on original strains in a basic solid culture medium containing tetracycline, and the specific passage domestication process is as follows: inoculating methylobacterium protospecies or pseudolaricis protospecies in a freezing storage tube to a basic solid culture medium containing 150mg/L tetracycline, culturing for 2-4d at 25-37 ℃, screening out tolerant viable bacteria, transferring into a basic solid culture medium containing 300mg/L tetracycline for subculture, repeating for 4 times, and increasing the tetracycline concentration by 150mg/L each time, namely the final tetracycline concentration is 750mg/L, thereby completing the subculture.
The preparation method of the in-situ restoration agent for efficiently degrading the tetracycline antibiotics in the soil comprises the following steps: mixing Pt/TiO according to the weight portion2Modified porous silkworm excrement, graphene, cation exchange resin, methylobacterium and pseudomonas syriacus to obtain the in-situ repairing agent.
EXAMPLE 1 preparation of Pt/TiO2Modified porous silkworm excrement
S1: according to the following steps: 2: 0.3: weighing raw silkworm excrement, urea solution, bromoplatinic acid solution and tetrabutyl titanate diluent according to the mass ratio of 0.1, wherein the tetrabutyl titanate diluent is prepared from tetrabutyl titanate and absolute ethyl alcohol according to the weight ratio of 1: 3.5 of mixed solution;
s2: soaking silkworm excrement in 9% urea solution for 5 hr to swell the silkworm excrement fully, drying at 80 deg.c to dewater, and treating in a volume ratio of 1: carbonizing at 190 ℃ for 1.5h under the protection of 0.15 of mixed gas of nitrogen and hydrogen;
s3: adding 0.5% bromoplatinic acid solution and tetrabutyl titanate diluent into the carbonized silkworm excrement obtained in S2 for quenching, heating to 475 ℃, roasting for 4h, and naturally cooling to obtain Pt/TiO2Modified porous silkworm excrement.
Example 2 domestication of passaged Methylobacterium and Pseudomonas Hibisci
(1) Domesticating methylobacterium: inoculating methylobacterium protospecies in a freezing storage tube to a basic solid culture medium containing 150mg/L tetracycline, culturing for 3d at 32 ℃, screening out tolerant live bacteria, transferring into the basic solid culture medium containing 300mg/L tetracycline for subculture, repeating for 4 times at the subculture temperature of 30 ℃ and the days of 2d, and increasing the concentration of the tetracycline by 150mg/L each time, namely the final concentration of the tetracycline is 750mg/L, thereby completing the subculture.
(2) Domesticated pseudomonas syriacus: inoculating the pseudomonas syriacus original strain in the frozen storage tube to a basic solid culture medium containing 150mg/L tetracycline, culturing for 3 days at 32 ℃, screening out tolerant live bacteria, transferring into the basic solid culture medium containing 300mg/L tetracycline for subculture, repeating for 4 times at the subculture temperature of 30 ℃ and the days of 2 days, and increasing the concentration of the tetracycline by 150mg/L each time, namely the final concentration of the tetracycline is 750mg/L, thereby completing the subculture.
Example 3 preparation of in situ remediation Agents
35 parts of Pt/TiO2And uniformly mixing the modified porous silkworm excrement, 4 parts of graphene, 3 parts of cation exchange resin, 5 parts of methylobacterium and 5 parts of pseudomonas syriacus to obtain the in-situ restoration agent.
According to the following steps of 1: 2000, the in-situ restoration agent obtained in the embodiment 3 is mixed in the soil polluted by aureomycin, terramycin and tetracycline, after 24 hours, the degradation rates of aureomycin, terramycin and tetracycline reach 69.1%, 65.8% and 72.4% respectively, and after 72 hours, the degradation rates of aureomycin, terramycin and tetracycline reach 87.9%, 84.7% and 91.6% respectively, so that the in-situ restoration agent produced by the invention has extremely strong degradation effect on tetracycline antibiotics, and has the advantages of short restoration period, simple operation and easy agricultural popularization and implementation.
While embodiments of the invention have been disclosed above, it is not limited to the applications listed in the description and the embodiments, which are fully applicable in all kinds of fields of application of the invention, and further modifications may readily be effected by those skilled in the art, so that the invention is not limited to the specific details without departing from the general concept defined by the claims and the scope of equivalents.
Claims (8)
1. An in-situ restoration agent for efficiently degrading tetracycline antibiotics in soil is characterized by comprising the following components in parts by weight: Pt/TiO 2230-40 parts of modified porous silkworm excrement, 2-6 parts of graphene, 1-5 parts of cation exchange resin, 4-6 parts of methylobacterium and 4-6 parts of pseudomonas syriacus; the Pt/TiO2The modified porous silkworm excrement is prepared by swelling and carbonizing original silkworm excrement by urea, quenching by bromoplatinic acid and tetrabutyl titanate, and roasting at the temperature of 450-Preparing; the methylobacterium and the pseudomonas syriacus are respectively obtained by continuously carrying out passage domestication on original strains in a basic solid culture medium containing tetracycline.
2. The in-situ remediation agent for efficiently degrading tetracycline antibiotics in soil of claim 1, wherein the Pt/TiO is selected from the group consisting of Pt, TiO, and mixtures thereof2The preparation process of the modified porous silkworm excrement is as follows: soaking silkworm excrement in 6-12% urea solution for 3-6 hr to swell the silkworm excrement fully, drying at 80 deg.c to dewater, carbonizing at 200 deg.c under the protection of mixed gas of nitrogen and hydrogen for 1-2 hr, adding 0.1-1% bromoplatinic acid solution and tetrabutyl titanate diluent for quenching, heating to 500 deg.c and roasting for 3-5 hr, and naturally cooling to obtain Pt/TiO2Modified porous silkworm excrement.
3. The in-situ remediation agent for efficiently degrading tetracycline antibiotics in soil according to claim 2, wherein the volume ratio of the nitrogen to the hydrogen is 1: (0.1-0.2).
4. The in-situ restoration agent for efficiently degrading tetracycline antibiotics in soil according to claim 2, wherein the tetrabutyl titanate diluent is a mixed solution of tetrabutyl titanate and absolute ethyl alcohol, and the volume ratio of the tetrabutyl titanate diluent to the absolute ethyl alcohol is 1: (3-4).
5. The in-situ restoration agent for efficiently degrading tetracycline antibiotics in soil according to claim 2, wherein the mass ratio of the silkworm excrement to the urea solution to the bromoplatinic acid solution to the tetrabutyl titanate diluent is 1: 2: 0.3: 0.1.
6. the in-situ remediation agent for efficiently degrading tetracycline antibiotics in soil according to claim 1, wherein the passage domestication process comprises: inoculating methylobacterium protospecies or pseudolaricis protospecies in a freezing storage tube to a basic solid culture medium containing 150mg/L tetracycline, culturing for 2-4 days at 25-37 ℃, screening out tolerant live bacteria, transferring into a basic solid culture medium containing 300mg/L tetracycline for subculture, repeating for 4 times, and increasing the concentration of the tetracycline by 150mg/L each time, namely the final concentration of the tetracycline is 750mg/L, thereby completing the subculture.
7. The in-situ remediation agent for efficiently degrading tetracycline antibiotics in soil according to claim 6, wherein the subculture temperature is 25-35 ℃ and the number of days is 1-3 d.
8. A preparation method of an in-situ restoration agent for efficiently degrading tetracycline antibiotics in soil is characterized by comprising the following steps: mixing Pt/TiO according to the weight portion2Modified porous silkworm excrement, graphene, cation exchange resin, methylobacterium and pseudomonas syriacus to obtain the in-situ repairing agent.
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