阅读说明：本技术 一种修复多环芳烃污染土壤的生物制剂 (Biological agent for repairing polycyclic aromatic hydrocarbon polluted soil ) 是由 高慧鹏 张全 关浩 王蒙 彭绍忠 于 2019-10-31 设计创作，主要内容包括：一种修复多环芳烃污染土壤的生物制剂,其制备方法为：以木质纤维素原料预处理后并进行纤维素酶水解后的水解液进行乙醇发酵,发酵液进行乙醇提取,将提取后的残液调节pH为6.0-8.0,灭菌,固液分离,将所得的固体干燥后磨成粉剂Ⅰ；将部分粉剂Ⅰ进行煅烧,粉碎成粉剂Ⅱ；将粉剂Ⅰ和粉剂Ⅱ混合制成所述生物制剂,其中粉剂Ⅱ的质量百分比占5%-50%。本发明的生物制剂来源于秸秆等生物质资源,无需添加化学品,不会造成二次污染。将木质纤维素水解发酵残余废渣用于土壤修复,实现废物利用,可以进一步降低纤维素乙醇的生产和排污成本。(A biological agent for repairing polycyclic aromatic hydrocarbon contaminated soil is prepared by the following steps: pretreating lignocellulose raw material, performing ethanol fermentation on hydrolysate subjected to cellulase hydrolysis, performing ethanol extraction on fermentation liquor, adjusting the pH of the extracted residual liquid to 6.0-8.0, sterilizing, performing solid-liquid separation, drying the obtained solid, and grinding into powder I; calcining part of the powder I, and crushing into powder II; and mixing the powder I and the powder II to prepare the biological preparation, wherein the mass percent of the powder II accounts for 5-50%. The biological agent is derived from straw and other biomass resources, does not need to add chemicals, and does not cause secondary pollution. The residual waste residue of lignocellulose hydrolysis and fermentation is used for soil remediation, so that waste utilization is realized, and the production and pollution discharge cost of cellulosic ethanol can be further reduced.)

1. A preparation method of a biological agent for repairing polycyclic aromatic hydrocarbon contaminated soil comprises the following steps:

(1) pretreating lignocellulose raw material, performing ethanol fermentation on hydrolysate subjected to cellulase hydrolysis, performing ethanol extraction on fermentation liquor, adjusting the pH of the extracted residual liquid to 6.0-8.0, sterilizing, performing solid-liquid separation, drying the obtained solid, and grinding into powder I;

(2) calcining part of the powder I, and crushing into powder II;

(3) and mixing the powder I and the powder II to prepare the biological preparation, wherein the mass percent of the powder II accounts for 5-50%.

2. The preparation method according to claim 1, wherein the mass percentage of the powder II in the biological agent is 10% -30%.

3. The method of claim 1, wherein the lignocellulosic feedstock is a straw, wood chips or energy plant containing cellulose, hemicellulose and lignin.

4. The method according to claim 1, wherein the pretreatment is at least one selected from the group consisting of mechanical pulverization, irradiation, microwave, acid treatment, alkali treatment, steam explosion, and solvent treatment.

5. The preparation method according to claim 1, wherein the solid-liquid separation in step (1) is realized by centrifugation or plate-and-frame filtration.

6. The method according to claim 1, wherein the drying in step (1) is carried out at a temperature of 40-90 ℃ for a time of 8-24 hours.

7. The process according to claim 1, wherein the solid particles in powder I and powder II have a particle size of not more than 5 mm.

8. The method of claim 1, wherein the calcining of step (2) is heating at 400 ℃ to 600 ℃ for 3 to 5 hours.

9. A biological agent produced by the method of any one of claims 1 to 8.

10. Use of the biological agent of claim 9 for remediation of polycyclic aromatic hydrocarbon contaminated soil.

11. The use according to claim 10, wherein the biological agent is administered at a dose of 10-100 g-kg^-1The soil is applied to the polluted soil in the concentration, the water is supplemented in the period of maintaining the water content of the surface soil to be 10% -35%, and the soil restoration period is 20-200 days.

Technical Field

The invention relates to the technical field of soil remediation, and particularly relates to a biological preparation prepared from cellulose hydrolysis fermentation residues and used for remedying polycyclic aromatic hydrocarbon contaminated soil.

Background

With the continuous consumption of fossil energy and the increasing prominence of environmental problems in recent years, the development of biomass energy has attracted extensive attention in all world circles. Since the 70 s in the 20 th century, biological energy sources represented by fuel ethanol and biodiesel are vigorously developed, and the development of the first generation of biomass fuel is based on the large occupation and consumption of agricultural resources, which causes a series of controversy about the problem of 'food safety'. The development of non-grain biofuels (second generation biofuels) is an important issue of worldwide interest and is also an important direction for the development of biofuels.

Lignocellulose is the most widely distributed and abundant renewable carbohydrate resource in nature. The cellulose raw materials such as crop straws and the like contain a large amount of cellulose and hemicellulose polysaccharide substances. These polysaccharides are stable in nature, and even after pretreatment, they are hydrolyzed into monosaccharides by the action of a catalyst, and then they are fermented to prepare cellulosic ethanol. The most commonly used catalysts for hydrolysis are mineral acids and cellulase preparations. The enzymolysis utilizes a microbial enzyme system to degrade natural cellulose and hemicellulose into fermentable monosaccharides, and compared with a chemical hydrolysis method, the method has the advantages of high yield of the fermentable monosaccharides, few byproducts, mild reaction conditions, low energy consumption and environmental friendliness.

However, how to effectively reduce the production cost of the whole process of preparing ethanol from lignocellulose is a key problem whether the technology is widely applied, a large amount of lignin-type waste residues are generated after the cellulose ethanol is hydrolyzed and fermented, the fermented waste residues are fully utilized, the economic value of the waste residues is improved, and the production cost of the cellulose ethanol can be effectively reduced.

Polycyclic Aromatic Hydrocarbons (PAHs) are organic compounds consisting of two or more benzene rings or heterocycles in a linear, bent, or clustered fashion, typical persistent organic pollutants present in the environment. In 1979, the U.S. Environmental Protection Agency (EPA) published 16 PAHs with priority control. PAHs mostly have higher boiling points and hydrophobicity, but lower volatility and water solubility. As the molecular weight of PAHs increases, the boiling point and the hydrophobicity of the PAHs are increased, the volatility and the water solubility are lower, the molecular structure is more stable, and the half-life period in the environment is longer. PAHs are generally biologically toxic and indeed carcinogenic to humans, and the biological toxicity increases with increasing molecular weight. Due to the severity of PAHs pollution in soil, from the 70 th century ago, countries in Europe and the United states began to develop remediation techniques for PAHs polluted soil. The soil remediation technology mainly comprises the following steps: physical repair techniques, chemical repair techniques, biological repair techniques, and joint repair techniques. In recent years, the utilization of organic wastes and the like for soil pollution control to realize resource utilization of wastes has become a hot problem of academic research.

CN101618394A discloses a method for restoring polycyclic aromatic hydrocarbon contaminated soil by using biochar, which specifically uses organic waste livestock manure produced in livestock and poultry breeding industry to prepare biochar material, and then adds the biochar into the contaminated soil to remove PAHs. The repair method of the invention mainly realizes the physical adsorption of the PAHs by the biochar, but can not realize the degradation of the PAHs, and has poor removal capability of the low molecular weight PAHs.

Disclosure of Invention

The invention provides a biological agent for repairing polycyclic aromatic hydrocarbon contaminated soil for the prior art, which takes cellulose hydrolysis fermentation residues as raw materials to prepare the biological repairing agent, solves the problem of recycling of the residues, improves the economy of the cellulose raw materials, and has good repairing performance and good application prospect.

In order to solve the technical problems, the technical scheme adopted by the invention is as follows:

the technical purpose of the first aspect of the invention is to provide a preparation method of a biological agent for repairing polycyclic aromatic hydrocarbon contaminated soil, which comprises the following steps:

(1) pretreating lignocellulose raw material, performing ethanol fermentation on hydrolysate subjected to cellulase hydrolysis, performing ethanol extraction on fermentation liquor, adjusting the pH of the extracted residual liquid to 6.0-8.0, sterilizing, performing solid-liquid separation, drying the obtained solid, and grinding into powder I;

(2) calcining part of the powder I, and crushing into powder II;

(3) and mixing the powder I and the powder II to prepare the biological preparation, wherein the mass percent of the powder II accounts for 5-50%.

Further, the mass percentage of the powder II in the biological preparation is preferably 10% -30%.

Further, the lignocellulose raw material is straw, wood chips or energy plants containing cellulose, hemicellulose and lignin, and the straw is preferred.

Further, the pretreatment is carried out by at least one selected from mechanical pulverization, radiation, microwave, acid treatment, alkali treatment, steam explosion and solvent treatment.

Further, the solid-liquid separation in the step (1) is realized by adopting a centrifugal or plate-frame filtration mode.

Further, the drying temperature in the step (1) is 40-90 ℃, and the time is 8-24 h.

Furthermore, the particle size of solid particles in the powder I is not more than 5 mm.

Further, the calcination in the step (2) is heating for 3-5 hours at 400-600 ℃.

Furthermore, the particle size of solid particles in the powder II is not more than 5 mm.

The technical object of the second aspect of the present invention is to provide a biological agent prepared by the above method.

The technical purpose of the third aspect of the invention is to provide the application of the biological agent in repairing polycyclic aromatic hydrocarbon contaminated soil, wherein the biological agent is 10-100 g.kg^-1The soil is applied to the polluted soil in the concentration, the water is supplemented in the period of maintaining the water content of the surface soil to be 10% -35%, and the soil restoration period is 20-200 days.

Compared with the prior art, the invention has the following beneficial effects:

(1) the biological agent is derived from straw and other biomass resources, does not need to add chemicals, and does not cause secondary pollution.

(2) The residual waste residue of lignocellulose hydrolysis and fermentation is used for soil remediation, so that waste utilization is realized, and the production and pollution discharge cost of cellulosic ethanol can be further reduced.

(3) The biological agent physically adsorbs pollutants by utilizing the porous structure of the biochar, promotes biodegradation of polycyclic aromatic hydrocarbon of the pollutants by biostimulation of fermented residual nutrients, has good soil remediation effect, and is convenient for industrial production and application.

Additional features and advantages of the invention will be set forth in the detailed description which follows.

Detailed Description

The following non-limiting examples are presented to enable those of ordinary skill in the art to more fully understand the present invention and are not intended to limit the invention in any way.

The lignocellulose raw material used in the embodiment of the invention is corn straw, wherein the cellulose accounts for 38.2wt%, the hemicellulose accounts for 22.1wt%, the lignin accounts for 20.2wt%, and the ash accounts for 3.9wt%, and the raw material is crushed to the particle size of 1-5 cm by a crusher.

The method for analyzing PAHs in soil comprises the following specific steps: taking a sample with the surface layer of 0-20cm polluted or restored soil, removing animal and plant residues and root systems, uniformly mixing, naturally drying in the shade for 48h, grinding the soil, sieving by using a 80-mesh sieve to obtain a soil sample to be analyzed, and storing in a brown bottle in a dark place. During analysis, 2g of soil sample to be detected is taken each time, 1g of anhydrous sodium sulfate is added and uniformly mixed, vortex oscillation extraction is carried out for 5min by using 4mL of n-hexane/acetone mixed solution (1: 1, v/v), extraction is carried out for three times, standing is carried out for 30min after each extraction, and extraction liquid obtained by three times of extraction is taken out and combined. After passing the extract through a 0.22 μm organic filter by vacuum filtration, the extract was transferred to a 100mL round-bottomed flask, the extract was concentrated using a rotary evaporator, and when the concentrate was concentrated to nearly 1mL, the concentrate was transferred to a 12mL brown bottle, and the brown bottle was previously rinsed with 2mL of n-hexane. The brown bottle is placed under a nitrogen blow dryer to completely volatilize n-hexane in the bottle, and then the volume of the brown bottle is increased to 2mL by using the n-hexane. 1mL of n-hexane was taken out and transferred to a liquid chromatography sample bottle, and the concentration of PAHs was analyzed by HPLC. The recovery rate of each PAH in the sample is proved to be more than 90% by the extraction method through calculation of the recovery rate.

And (3) quantitatively detecting PAHs in the sample by using Agilent 1200 high performance liquid chromatography. The chromatographic column is PAH analysis special column (ZORBAX Eclipse PAH, 3.0 × 250mm filler particle diameter5 μm, Agilent, USA). The flow rate of the mobile phase was 0.85 ml min^-1The column temperature is 25 ℃, the detector is an ultraviolet variable wavelength detector, the wavelength is 220 nm, and the sample injection amount is 10 mu L. The mobile phase was acetonitrile and water, the initial ratio was 45% acetonitrile and 55% water, the acetonitrile ratio rose to 100% after 17.5min and was held for 20 min. And (3) establishing a standard curve between the peak area and the concentration of each compound by using PAHs standard substances with different concentrations, and quantifying the concentration of the PAHs in the sample by an external standard method.

Example 1

(1) Adjusting the water content of corn straws to 88%, soaking for one hour, putting the corn straws into a feeder of a steam explosion device, extruding the corn straws into a digester through a screw rod, wherein the cooking temperature is 180 ℃, the cooking time is 10min, extruding materials in the digester through a combined screw rod device, putting the extruded materials into a detention device, keeping the temperature at 200 ℃, the pressure at 1.8MPa for 10min, and then instantaneously performing pressure relief explosion to obtain materials pretreated by steam explosion;

preparing the material pretreated by steam explosion and water into a mixed solution with the solid-liquid mass ratio of 12.6%, adding Ctec2 cellulase of Novoxil, and adding 10FPU unit of enzyme activity cellulase solution into each gram of cellulose; performing enzymolysis for 120h at 50 ℃, pH of 5.0 and rotation speed of 130 rpm; adjusting pH of the enzymolysis solution to 6.0, mixing at 30 deg.C, adding temperature-resistant Saccharomyces cerevisiae FE-B provided in CN200910204295.3, and fermenting with ethanol; the preparation process of the fermented seed liquid comprises the following steps: the seed culture medium is YPD culture medium, wherein the yeast extract powder is 10g/L, the peptone is 20g/L, and the glucose is 20 g/L; inoculating zymophyte to a seed culture medium, wherein the culture conditions are as follows: the culture temperature is 40 ℃, the rotating speed of a shaking table is 250rpm, and the culture time is 24 hours; inoculating the seed solution into the enzymolysis solution according to the inoculum size of 10% of the volume ratio, and culturing for 72h at 120 rpm.

Distilling ethanol in the fermentation liquor, adjusting pH of the residual fermentation liquor to 6.5, sterilizing at 121 deg.C for 30min, centrifuging to obtain solid precipitate, drying at 80 deg.C for 12 hr, and grinding into powder I with particle size not greater than 5 mm.

(2) Calcining part of the powder I in a muffle furnace at 500 ℃ for 4 hours, cooling and crushing into powder II with the particle size not larger than 5 mm.

(3) And mixing the powder I and the powder II according to a mass ratio of 5: 1 mixing to obtain the biological preparation.

The biological agent is used for repairing polycyclic aromatic hydrocarbon polluted soil:

the polycyclic aromatic hydrocarbon contaminated soil is surface soil obtained from the vicinity of a chemical plant, and the biological agent is added at a ratio of 80 g/kg^-1Uniformly mixing the concentration of the soil and the polluted soil, and replenishing water to keep the water content of the surface soil to be 20%; after 50 days of remediation, the concentrations of PAHs in the soil were analyzed, and the results are shown in Table 1, wherein various PAHs were reduced to different degrees, the average removal rate of the PAHs was 68.72%, and the total removal amount was 63.07mg/kg soil.

Table 1.

Example 2

The steps (1) and (2) are the same as the step (1), and in the step (3), the powder I and the powder II are mixed according to the mass ratio of 5: 2 mixing to obtain the biological preparation.

The soil was restored in the same manner as in example 1, and after 50 days, the restoration results are shown in Table 2, in which the average removal rate of PAHs was 65.42% and the total removal amount was 54.48mg/kg of soil. Wherein the removal rate of low molecular weight PAHs is reduced, and the removal rate of high molecular weight PAHs is increased.

Table 2.

Comparative example 1

As a blank control group, no preparation was added to the contaminated soil, and the soil was restored by self-cleaning ability, and after 50 days, the results are shown in Table 3, in which the average removal rate of PAHs was 4.22% and the total removal amount was 5.32mg/kg of soil.

Table 3.

Comparative example 2

Only powder I prepared in step (1) of example 1 was used as a soil remediation agent at a dose of 80 g/kg^-1After the soil is added into the soil, the repairing result is shown in table 4, the average removal rate of PAHs is 38.12%, the total removal amount is 49.12mg/kg of soil, and the effect of removing high molecular weight PAHs is not ideal after 50 days.

Table 4.

Comparative example 3

Only the powder II prepared in the step (2) in the example 1 is taken as a soil repairing agent in an amount of 80 g/kg^-1After the soil is added into the soil, the results are shown in table 5, the average removal rate of PAHs is 4.22%, the total removal amount is 30.64mg/kg of soil, and the removal effect of low molecular weight PAHs is not ideal after 50 days.

Table 5.