CN115058252A - Microbial soil conditioner for phthalate ester contaminated soil remediation and application thereof - Google Patents
Microbial soil conditioner for phthalate ester contaminated soil remediation and application thereof Download PDFInfo
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Abstract
The application provides a microbial soil conditioner for repairing phthalate ester contaminated soil, which comprises microbial carried biochar, organic matters, a sugar carbon source and nutrients, wherein the microbial carried biochar consists of sphingomonas, bacillus subtilis and biochar; the endophyte of the conditioner can also colonize the interior of plants, promote the degradation of Pollutants (PAEs) in the plants, achieve the effect of simultaneously repairing inside and outside, integrate the repairing and the growth promotion, promote the growth of the plants while repairing and removing the pollution of plasticizers, adjust the micro-ecology of soil, reduce the use of chemical fertilizers, and is favorable for the sustainable development of high-quality farmland construction and agriculture.
Description
Technical Field
The invention relates to the technical field of fertilizers, in particular to a preparation method of a microbial soil conditioner for repairing soil polluted by toxic organic matters and application of the microbial soil conditioner in phthalate ester degradation.
Technical Field
Phthalates, also known as Plasticizers (PAEs), are phthalate-based compounds that are widely used as plasticizers in toys, cosmetics, building materials, pesticide carriers, and the like. Some PAEs are endocrine disrupting substances, 6 plasticizers such as n-butyl phthalate (DBP) and di (2-ethylhexyl) phthalate (DEHP), which are potential carcinogens, teratogens and mutagenics for animals and humans, are listed as pollutants to be controlled by the United states environmental protection agency of America preferentially, and 3 PAEs such as dimethyl phthalate are also considered as pollutants to be controlled by the China department of environmental protection preferentially. Due to the wide use of plasticizers, the plasticizers are generally left in the air, soil, water and other environments. In the agricultural product producing area, the detected concentration and the detected rate of the plasticizer in facilities and the soil and vegetables of the film-covered vegetable producing area are far higher than those of the conventional producing area. Plasticizers in the environment are easily absorbed and enriched by crops, which not only affect the quality of crops, but also cause potential health risks to human bodies.
Plasticizer repair technology has two forms of bioremediation and non-bioremediation, wherein the bioremediation comprises microbial remediation and plant remediation; and the non-bioremediation technique includes hydrolysis technique, physical technique, advanced oxidation technique, and the like. Compared with non-biological repair, biological repair is more suitable for being used in farmlands, but microorganisms have poor stability in the environment, so that the microbial repair technology is difficult to popularize in a large area; while phytoremediation can disrupt the normal farming rhythm. The technology for remedying the farmland plasticizer pollution, which has low cost, high efficiency and high speed and does not introduce new pollutants, is still very lacking. At present, the application of the microbial soil conditioner with plasticizer repair and plant growth promotion functions is not reported.
Disclosure of Invention
In order to solve the problems, the invention provides a microbial soil conditioner with a plasticizer degradation function. The soil conditioner can promote the degradation of plasticizers in a vegetable-soil planting system, has the effect of promoting plant growth, obviously improves the biomass of vegetables, improves the microbial community structure of soil, improves the cultivated land quality and greatly reduces the use of chemical fertilizers.
Specifically, the invention is realized by the following technical scheme:
firstly, the application provides a microbial soil conditioner for repairing phthalate ester polluted soil, wherein the microbial soil conditioner comprises microbial carried biochar, organic matters, a sugar carbon source and nutrients. The microorganism-carried biochar consists of biochar and endophytes, and the content of the endophytes in the microorganism soil conditioner is not less than 1.5 multiplied by 10 8 cfu/g, the endophyte is obtained by mixing sphingomonas and bacillus subtilis.
Preferably, the bacteria content ratio of sphingomonas to bacillus subtilis in the endophyte is 1: 1.
Preferably, the mass percentages of the organic matters, the sugar carbon source and the nutrients in the microbial soil conditioner are 65%, 5% and 5% in sequence, and the balance is organic matters carried by microbes.
Preferably, the nutrient includes at least one of urea, calcium superphosphate, and potassium chloride.
Preferably, the organic matter is one or two of bean pulp and rapeseed pulp; the sugar carbon source is preferably D-cellobiose or other poly-oligosaccharides;
preferably, the biochar is rice hull biochar or straw (preferably corn straw) biochar.
Secondly, the application also provides the application of the microbial soil conditioner in repairing phthalate ester polluted soil. Specifically, the microbial soil conditioner is used as a base fertilizer to be broadcast and applied one week before vegetable sowing, and then the conditioner is uniformly stirred with the surface soil (about 15cm deep) of a field, wherein the using amount is 110 kg/mu.
The principle of the microbial soil conditioner for organic pollution remediation constructed in the application is shown in figure 1, the conditioner is constructed by taking beneficial endophytes as a core, and the stability of the endophytes in the environment is improved by carrying the endophytes by using charcoal; the biochar can also adsorb organic pollutants in the soil environment, block the absorption of plants on the organic pollutants, and provide an environment for degrading the pollutants intensively for endophytes; sugar carbon sources are added in an auxiliary mode, so that the degradation capability of endophytes is improved; organic matters are added to strengthen rhizosphere microbial communities; meanwhile, a small amount of nutrients are added to ensure the requirement of the early growth of crops.
The soil conditioner provided by the application can be used for repairing pollutants in soil, and the endophyte can also be used for colonizing the interior of a plant, so that the degradation of Pollutants (PAEs) in the plant is promoted, and the effect of simultaneously repairing the interior and the exterior is achieved. The microbial regulator integrates restoration and growth promotion, can promote plant growth while restoring and removing plasticizer pollution, regulates soil micro-ecology, reduces the use of chemical fertilizers, and is beneficial to high-quality farmland construction and sustainable development of agriculture.
Drawings
FIG. 1 is a schematic diagram illustrating a construction principle of a microbial soil conditioner for organic pollution remediation according to the present application.
Fig. 2 is a schematic diagram illustrating that the addition of a sugar carbon source significantly improves the ability of endophytes to degrade plasticizers.
FIG. 3 is a graph showing the effect of different treatments on PAEs concentrations in vegetables (a) and soil (b);
different letters in fig. 3 indicate that the two differ significantly at the α -0.05 level, the same below.
FIG. 4 is a graph showing the effect of different treatments on vegetable biomass.
FIG. 5 is a graph showing the effect of different treatments on the enzymatic activities of soil urease (a), sucrase (b) and acid phosphatase (c).
FIG. 6 is a schematic representation of the effect of different treatments on the microbial community at the root of vegetables.
FIG. 7 is a graph showing PAEs concentrations in the stems and leaves (a), roots (b) and soil (c) of vegetables tested in a plot.
Fig. 8 is a graph showing chlorophyll and vitamin C contents of different treated vegetables.
Detailed Description
In the following examples, Sphingomonas and Bacillus subtilis were isolated from plants by an organic pollution reduction and control innovation team in agricultural product producing areas of the academy of agricultural sciences of Jiangsu province, purified, identified and stored. Wherein, the bacillus subtilis has already been published by Chinese patent 201910079682.2 (publication No. 109666612A with the preservation number of CGMCC NO.16233), and the sphingomonas has already been published by Chinese patent ZL201510775953.X (publication No. 105274031B with the preservation number of CGMCC No. 11031).
The starting materials and media mentioned in the examples:
LB liquid medium: 10g/L tryptone, 5g/L yeast extract, 10g/L sodium chloride and pH7.0;
inorganic salt liquid medium MSM: MgSO (MgSO) 4 ·7H 2 O(0.4g)，FeSO 4 ·7H 2 O(0.2g)，K 2 HPO 4 (0.2g)，(NH 4 ) 2 SO 4 (0.2g) and CaSO 4 (0.08g) 1L of deionized water, and the pH value is 7.0-7.2;
seed culture medium: k 2 HPO 4 (4.8g)，KH 2 PO 4 (3.5g)，(NH 4 ) 2 SO 4 (2g)，MgCl 2 (0.16g)，CaCl 2 (0.02g)，NaMoO 4 .2H 2 O(0.0024g)，FeCl 3 (0.0018g)，MnCl 2 .2H 2 O (0.0015g), PH7.0, water to 1L.
Inorganic salt solid medium: agar was added to the inorganic salt liquid medium to a final concentration of 20 g/L.
The high temperature start-up furnace was purchased from Shanghai optical precision mechanics research institute of Chinese academy of sciences, model SG-GL 1100K.
The instruments and materials used in the other examples were purchased commercially unless otherwise indicated.
Example 1 preparation of soil conditioner
Respectively inoculating purified endophytes (Sphingomonas bacteria and Bacillus subtilis) into LB liquid culture medium, and culturing at 32 + -1 deg.C for 24 hr to obtain endophyte fermentation broth with bacterial content of 10 10 Left and right for standby.
The method comprises the steps of taking rice hulls as raw materials, preparing biomass charcoal under the anaerobic condition of 450 ℃ by adopting a high-temperature open-type tubular furnace, crushing, sieving with a 100-mesh sieve, sterilizing in a sterilization pot at 121 ℃ for 1 hour, and taking out to obtain the rice hull biomass charcoal for later use. In the embodiment, the biochar prepared by anaerobic firing at 450 ℃ has relatively high adsorption capacity and electron transfer capacity, and is suitable for soil organic pollution remediation.
Sterilizing bean pulp at 121 ℃ and grinding the bean pulp to about 1mm to obtain organic matters for later use;
sphingomonas and Bacillus subtilis were mixed according to a ratio of cell content 1:1, mixing to obtain an endophyte liquid, and inoculating the endophyte liquid to the charcoal, wherein the mass ratio of the endophyte liquid to the charcoal is 1: 4; culturing for 2 days, air drying to water content<25 percent of the obtained microorganism-carried biochar, wherein the content of endophytes in the biochar is 10 10 About cfu/g.
Uniformly mixing bean pulp, D-cellobiose, nutrients (urea, calcium superphosphate and potassium chloride are 2:3:1 in parts by mass) and microorganism-carried biochar to obtain a microorganism soil conditioner, wherein the organic matter bean pulp, the sugar-carbon source D-cellobiose and the nutrients respectively account for 65% by mass, 5% by mass and 5% by mass of the microorganism soil conditioner, and the balance is the microorganism-carried biochar; the obtained conditioner has endophyte content ≧ 1.5 × 10 8 cfu/g。
In the specific implementation process, one or more of endophytes with corresponding degradation capability and endophytes with growth promoting capability can be selected according to the target pollutant to be repaired, and a proper sugar carbon source is selected to prepare the soil conditioner.
Sugar carbon source alignment experiment: adding various plasticizers (DMP, DEP, DBP, BBP, DEHP, DnP) into LB liquid culture medium respectively, wherein the final concentration of the plasticizer is 40 mg/kg; respectively measuring the bacterial liquid concentration OD 600 1mL of 1.0 endophyte solution was inoculated into 50mL of medium, and then 10mmol of D-cellobiose was added to the medium of test group 1 (bacteria + sugar), and no sugar was added to test group 2 (bacteria), while a control group without inoculation and sugar was set up. The concentration of residual plasticizer in the petri dish after 7d was measured, and the measurement results are shown in fig. 2. As can be seen from fig. 2, the addition of the sugar carbon source significantly increased the amount of plasticizer degradation.
Example 2 bowl test
The microbial soil conditioner obtained in example 1 was mixed with artificially contaminated soil (DBP and DEHP were added to non-contaminated soil at the same time to final concentrations of 20 mg/kg), and after the solvent was evaporated, the contaminated soil was loaded into pots, each of which was loaded with 2 kg.
The soil used in the test was collected from Jiangsu Nanjing and Sichuan Meishan.
The test crop is Chinese cabbage, 4 treatments are set in the test, NF: no fertilizer is applied; CF: applying urea; OF: applying a soil conditioner without added bacteria (except that no endophyte is added, the preparation method is the same as the preparation method of the microbial soil conditioner in the example 1); EOF: applying the microbial soil conditioner obtained in example 1; the CF and OF treatment groups remained the same as the EOF treatment group Nutrients (NPK) except NF; the amount of the soil conditioner used in the EOF treatment group was 6 g/bowl.
The treatment is carried out in a manner that other management measures are consistent except different varieties of applied fertilizers, each treatment is repeated for 3 times, the experiment is carried out in 2020, 12 and 10 days, 2021, 1 and 22 days, vegetables and soil samples are collected during harvesting, the concentration of PAEs in the vegetables and the soil, the biomass of the vegetables, the soil fertility effect and the microbial diversity are analyzed, and the detection results are as follows:
1) effect of different treatments on PAEs concentrations in vegetables and soil
The experimental results are shown in fig. 3, in which (a) is the result of detecting the concentration of PAEs in vegetables, and (b) is the result of detecting the concentration of PAEs in soil. As can be seen from fig. 3, the concentrations of typical plasticizers DBP and DEHP in the roots and stems of brassica campestris were significantly lower in the EOF treatment group applied with the microbial soil conditioner containing bacillus subtilis than in the other treatment groups; compared with the control group, the DBP and DEHP concentrations of the roots, the stem leaves and the leaves of the pakchoi in the EOF treatment group are reduced by about 70 percent; compared with the OF group, the reduction is more remarkable. The result shows that the soil conditioner added with the bacillus subtilis can effectively remove DBP and DEHP in the green vegetables. DBP and DEHP are environmentally more and preferentially controlled plasticizer types and are relatively more toxic and therefore serve as indicators of the efficacy of plasticizer contamination remediation.
In general, the soil conditioner containing two endophytes significantly reduced the concentrations of PAEs in the soil, and compared with the control, the DBP concentration was reduced by about 42%, and the DEHP concentration was reduced by about 48%.
2) Experiment of influence of different treatments on green vegetable biomass
The results of the experiment are shown in FIG. 4. As can be seen from FIG. 4, the soil conditioner prepared in example 1 can significantly increase the green vegetable biomass in the soil polluted by 20mg/kg plasticizer. Compared with the NF, CF and OF treatment groups, the EOF improves the total biomass OF the green vegetables by about 150 percent, 30 percent and 12 percent respectively. The EOF has obvious growth promoting effect on stems, leaves and roots of green vegetables.
3) The results of the measurements of the changes in soil properties for the different treatments are shown in tables 1, 2 and FIG. 5 below
TABLE 1 fertility status of Nanjing soil under different treatments
TABLE 2 fertility of the Szechwan mountains under different treatments
The results in tables 1 and 2 show that compared with NF and CF, EOF has certain improvement effect on soil with different properties, and improves the content of organic matters, available phosphorus and the like in the soil.
In fig. 5, (a), (b), and (c) are the results of the effects of different treatments on the activities of soil urease, sucrase, and acid phosphatase, respectively, and it can be seen that, compared with the control and the fertilizer application group, EOF significantly enhances the activities of soil urease, sucrase, and acid phosphatase, and significantly improves the properties of plasticizer-contaminated soil. 4) Influence of soil conditioner on microbial community of vegetable rhizosphere
Effect results as shown in the results of fig. 6, the application of EOF significantly increased the abundance of beneficial bacteria such as sphingomonas, flavobacterium, lysobacter, and geobacter xanthans, wherein multiple strains of sphingomonas and flavobacterium were reported to have plasticizer-degrading function, lysobacter had resistance to phytopathogen, nitrogen fixation, and degradation of organic matter, and geobacter xanthans had rhizosphere growth promoting function.
Example 3 field test
And (3) field plot experiment: DBP and DEHP are prepared into solution with a certain concentration, and the solution is sprayed into a test cell, turned over (15 cm of surface soil) and uniformly mixed. The soil was contaminated with DBP and DEHP at a concentration of 20mg/kg (in 15cm soil thickness of the top layer). After the soil is artificially polluted, the test cell is aired for 2 days. And (3) fertilizing or treating with a soil conditioner after 2 days, uniformly spreading the fertilizer or the soil conditioner on the surface layer of the soil, and spreading a certain amount of vegetable seeds after uniformly stirring.
The test crop is Chinese cabbage, 4 treatments are set in the test, NF: no fertilization, CF: applying a fertilizer, EOF: applying the soil conditioner obtained in example 1; CF and EOF treatment group Nutrients (NPK) are identical except for NF; the using amount of the soil conditioner in the EOF treatment group is 110 kg/mu.
The treatment is carried out according to the same management measures except different varieties of applied fertilizers, 3 management measures are repeated in each cell, the vegetables are harvested 45 days after being planted, samples of the vegetables and soil are collected during harvesting, and the concentrations of PAEs, chlorophyll and VC in the vegetables and the soil are analyzed.
The test results are as follows:
1) detection of PAEs (DBP and DEHP) concentration in vegetables and soil
The detection results are shown in fig. 7, wherein a and b in fig. 7 are the detection results of the concentrations of the stem leaves of the brassica chinensis, the roots of the brassica chinensis and PAEs (DBP and DEHP) in the cell test respectively. Therefore, the concentrations of PAEs in the stem leaves and the roots of the vegetables are obviously lower than those of the vegetables which are not fertilized and are used by a fertilizer group by using the soil conditioner group added with the bacillus subtilis, and compared with the group which is not fertilized, the concentrations of DBP and DEHP in the stem leaves of the vegetables are respectively reduced by 15 percent and 31 percent; the DBP and DEHP concentration of the vegetable roots are reduced by 46 percent and 60 percent respectively.
In fig. 7, c is the measurement result of PAEs (DBP and DEHP) concentration in soil, and it can be seen that PAEs concentration in soil of the soil conditioner treatment group is significantly lower than that of the non-fertilization and fertilization groups, and DBP and DEHP concentrations in soil of the soil conditioner group are respectively reduced by 56% and 61% compared with those of the non-fertilization group.
3) Detection results of chlorophyll and VC content of different treated vegetables
The detection results are shown in fig. 8, wherein a and b in fig. 8 are the detection results of the contents of chlorophyll and vitamin C in the green vegetables, respectively. As can be seen, the contents of chlorophyll and vitamin C in the green vegetables in the soil treatment agent group are obviously higher than those in the non-fertilization or fertilizer application group, which shows that the quality index of the vegetables is effectively improved by soil conditioning.
The results show that the soil conditioner constructed by the invention has multiple functions of reducing toxicity and promoting growth, can effectively reduce the plasticizer pollution in vegetables and soil, improve the soil microbial community, promote the vegetable growth, greatly reduce the use of chemical fertilizers, improve the production field quality of agricultural products and promote the sustainable development of agriculture.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that there are objectively infinite specific structures due to the limited character expressions, and it will be apparent to those skilled in the art that a plurality of modifications, decorations or changes may be made without departing from the principle of the present invention, and the technical features described above may be combined in a suitable manner; such modifications, variations, combinations, or adaptations of the invention in other instances, which may or may not be practiced, are intended to be within the scope of the present invention.
Claims (7)
1. A microbial soil conditioner for repairing phthalate ester contaminated soil is characterized by comprising microbial carried biochar, organic matters, a sugar carbon source and nutrients;
the nutrient comprises at least one of urea, calcium superphosphate and potassium chloride;
the organic matter is at least one of bean pulp and rapeseed pulp;
the sugar carbon source is poly-oligosaccharide;
the microorganism-carried biochar consists of biochar and endophytes, and the content of the endophytes in the microorganism soil conditioner is not less than 1.5 multiplied by 10 8 cfu/g, wherein the endophyte is obtained by mixing sphingomonas and bacillus subtilis.
2. The microbial soil conditioner for phthalate-contaminated soil remediation according to claim 1 wherein the endophyte comprises sphingomonas and bacillus subtilis in a bacteria content ratio of 1: 1.
3. The microbial soil conditioner for phthalate-contaminated soil remediation according to claim 1 wherein said sugar carbon source is D-cellobiose.
4. The microbial soil conditioner for phthalate-contaminated soil remediation according to claim 1 wherein said biochar is rice hull biochar or straw biochar.
5. The microbial soil conditioner for remediating phthalate-contaminated soil as claimed in any one of claims 1 to 4, wherein the organic matter, the sugar carbon source and the nutrient account for 65%, 5%, 5% and the balance of the microbial soil conditioner in sequence by mass.
6. The use of the microbial soil conditioner for remediation of phthalate-contaminated soil according to any one of claims 1 to 4 for remediation of phthalate-contaminated soil.
7. The use according to claim 6, wherein the microbial soil conditioner is applied as a base fertilizer in an amount of 110 kg/acre for one week before vegetable sowing.
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| CN116640580A (en) * | 2023-06-06 | 2023-08-25 | 山东农大肥业科技股份有限公司 | Soil conditioner and preparation method and application thereof |
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