A treatment method for removing cadmium from contaminated biomass using Aspergillus niger

Abstract

The invention discloses a treatment method for removing cadmium from contaminated biomass by using Aspergillus niger, and the treatment method comprises the following steps: using Aspergillus niger as an exogenous added fungus to extract cadmium from contaminated biomass alone, optimizing the optimal conditions by using the response surface method, and using the pretreatment method based on the optimal conditions of the response surface to work together with Aspergillus niger to complete the removal of cadmium from contaminated biomass. The treatment method of the invention performs shake flask extraction and culture of Aspergillus niger and contaminated biomass in a microbial extraction manner, which can achieve effective treatment of cadmium-contaminated biomass after plant restoration, has the advantages of good treatment effect, low investment, green environmental protection, low technical management requirements, simple operation, etc., and can also achieve the reuse of the treated biomass as fertilizer, which has good environmental benefits, social benefits and economic benefits.

Description

Technical Field

[0001] The invention belongs to the technical field of heavy metal contaminated biomass disposal, relates to a method for treating contaminated biomass, and specifically relates to a method for removing cadmium from contaminated biomass by using Aspergillus niger. Background Art

[0002] The 2014 National Soil Pollution Survey Bulletin shows that the overall soil environmental situation in my country is not optimistic. Soil pollution is serious in some areas, the environmental quality of cultivated land soil is worrying, and the soil environmental problems in industrial and mining wasteland are prominent. Among them, the point-to-point excess rate of cadmium among inorganic pollutants is the highest, reaching 7%. Plant extraction is a method of using some special plants with strong enrichment ability for heavy metals to absorb heavy metals from the soil, transfer and store them to the aboveground part, and remove pollutants in the soil by harvesting the aboveground part of the plant. It is also a common method for repairing heavy metal pollution in farmland at this stage. Compared with traditional remediation methods, plant remediation has the advantages of being green, environmentally friendly, and economical. However, after plant remediation, the plants are removed from the site, which may lead to the accumulation of a large amount of harmful substances in the biomass. Oil crops are often used to repair heavy metal pollution in farmland because of their large biomass, low heavy metal content in oil, and certain economic value. Studies have shown that the distribution of heavy metal Cd in oil sunflower is: leaves> roots> fruit pods> stems> seeds; after oil extraction, most cadmium remains in the cake. The cadmium content in rapeseed cake is twice that of rapeseed oil. Oil can be used as an energy source to solve my country's energy crisis. The treatment of contaminated plant materials is one of the obstacles to the commercial implementation of plant extraction. Therefore, as the last link of phytoremediation, how to safely dispose of heavy metal-enriched plant biomass is considered to be an important factor in measuring whether phytoremediation is truly successful.

[0003] The existing methods for biomass treatment and heavy metal purification are mainly physical and chemical leaching methods. In order to improve the efficiency of phytoremediation, many scholars have studied the close interaction between plants, microorganisms and heavy metals in rhizosphere soil. Bioleaching is the process of leaching valuable metals from ores or other materials into solutions with the help of microorganisms. It is a multidisciplinary cross-field that integrates hydrometallurgy, mineral processing, chemical engineering, environmental engineering and microbiology. It has many advantages such as simple process, short process, simple equipment, small investment, low cost, light pollution and small resource consumption. At present, the research on microbial extraction technology is mainly focused on the leaching of copper, nickel and other metals in ores or contaminated soils and the enrichment or fixation of heavy metals by microorganisms and plants. There is still a gap in the research on the leaching and purification of cadmium in cadmium-containing biomass (leaves, straw and oil cakes, etc.) after plant extraction using microorganisms, as well as the safe utilization of leached biomass. Summary of the invention

[0004] The technical problem to be solved by the present invention is to overcome the deficiencies of the prior art and provide a treatment method for removing cadmium from contaminated biomass by using Aspergillus niger with good treatment effect and economic rationality.

[0005] In order to solve the above technical problems, the technical solution adopted by the present invention is:

[0006] A method for removing cadmium from contaminated biomass using Aspergillus niger, the method comprising the following steps:

[0007] (1) Aspergillus niger was used as an exogenous fungus to extract cadmium from contaminated biomass under different treatments;

[0008] (2) After completing step (1), the optimal conditions for leaching cadmium from contaminated biomass by applying Aspergillus niger alone are optimized using response surface methodology;

[0009] (3) After completing step (2), the pretreatment method is used in combination with Aspergillus niger based on the optimal conditions of the response surface to remove cadmium from the contaminated biomass.

[0010] The above treatment method is further improved in that the polluted biomass is crop straw, which is one of sunflower straw, rape straw and sesame straw.

[0011] The above treatment method is further improved, the Aspergillus niger is purchased from China Industrial Microbiological Culture Collection Management Center, and the culture collection number is CICC 2214; the concentration of the Aspergillus niger spore suspension is 4.25×10 5 ~10 8 pc·mL -1 ; The inoculation amount of Aspergillus niger is 8% to 12%.

[0012] The above treatment method is further improved, wherein the different treatments are performed with a sucrose content of 70 to 120 g·L -1 , the inoculation amount is 8% to 12%, and the pH is at least one of 2 to 5.

[0013] The above processing method is further improved, wherein the response surface is based on a central structure design to form a three-factor five-level experimental condition; the response surface uses Design Expert 10 software to generate 20 experimental operation groups.

[0014] The above treatment method is further improved, wherein the contaminated biomass pretreatment includes at least one of water treatment, hydrothermal treatment, alkali treatment, and enzyme treatment.

[0015] The above-mentioned treatment method is further improved, wherein the hydrothermal treatment is maintained at 160°C for 10 minutes, cooled to 120°C and maintained for 10 minutes; the alkali treatment is 0.5% to 1% by mass of sodium hydroxide, soaked for 4 hours; the enzyme treatment is cellulase produced from Aspergillus niger, hydrolyzed at 50°C for 72 hours; the water treatment is ultrapure water at room temperature for the same time as hydrothermal treatment, soaked in ultrapure water for 4 hours after extrusion filtration, and maintained in ultrapure water at 50°C for 72 hours after extrusion filtration.

[0016] The above treatment method is further improved in that the shaking flask culture extraction system is 100 mL, the temperature is 30° C., the culture time is 11 to 15 days, and the rotation speed is 150 to 220 rpm.

[0017] In the present invention, the crop straw used is the residual biomass harvested after the plant restoration of oil crops. The straw contains rich organic matter, nitrogen, phosphorus, potassium and trace elements, is a multi-purpose and renewable biological resource, and is also an important source of organic fertilizer required for agricultural production. Therefore, how to safely dispose of heavy metal-contaminated biomass is extremely important; and oilseeds can be used to extract oil, and cakes can be used as high-protein feed, which can bring certain economic benefits and has a very broad market potential.

[0018] In the present invention, the Aspergillus niger used is a fungus of the genus Aspergillus of the family Aspergillus, which produces organic acids such as oxalic acid, gluconic acid and citric acid during its growth and metabolism; it can also produce enzyme preparations such as amylase, cellulase, acid protease, glucose oxidase, pectinase, etc.

[0019] Compared with the prior art, the advantages of the present invention are:

[0020] (1) The present invention provides a treatment method for removing cadmium from contaminated biomass using Aspergillus niger, wherein Aspergillus niger is used as an exogenous added fungus to extract cadmium from contaminated biomass alone, and the optimal conditions are optimized using the response surface method. Based on the optimal conditions of the response surface, the pretreatment method is used in combination with Aspergillus niger to complete the removal of cadmium from contaminated biomass. In the present invention, Aspergillus niger can grow normally in a culture medium containing cadmium-contaminated biomass and produce spores. At the same time, Aspergillus niger can secrete a variety of secretions (such as organic acids, including citric acid, oxalic acid, and lactic acid), and the content of the secretions is very high. On the one hand, the Aspergillus niger secretions maintain or reduce the initial pH value of the culture medium, which is conducive to the leaching of cadmium from the contaminated biomass straw; on the other hand, the enzyme preparation (including cellulase) produced by Aspergillus niger decomposes the biomass straw cellulose, loosens the contaminated biomass straw, increases the porosity, and allows the leached nutrients or nutrients added in the culture medium to be adsorbed on the surface of the oil sunflower straw. On this basis, in the present invention, the sucrose content in the culture medium, the Aspergillus niger inoculation amount, and the initial pH value of the system are set as variables. In the present invention, sucrose is added as an exogenous carbon source into the culture medium, and Aspergillus niger can use simple high molecular carbohydrate sucrose to convert into low molecular organic acids such as citric acid, oxalic acid, lactic acid, etc., resulting in a decrease in the pH value of the extraction system, thereby better improving the leaching effect of cadmium in the contaminated biomass straw. In addition, in the process of Aspergillus niger extracting contaminated biomass straw, different single factors are combined to control the cultivation (such as, the initial pH value of the extraction system, the inoculation amount of Aspergillus niger, etc.). On the one hand, acidification is conducive to metal migration, which is the most important mechanism in fungal bioleaching, and is to leach metals from solid biomass and transfer them to the leaching solution through protonation, so as to achieve the effect of reducing the cadmium content in the contaminated biomass straw; and the appropriate initial pH value of the system can promote the growth of Aspergillus niger, which is beneficial to acid and enzyme production. On the other hand, the impact of the inoculation amount of Aspergillus niger on the extraction system is also very important. Using a larger inoculation amount can shorten the time for mycelial reproduction to reach a peak, so that the formation of the product arrives in advance, and the growth opportunity of miscellaneous bacteria can be reduced. However, if the inoculum size is too large, it will cause insufficient dissolved oxygen, and the nutrient organic matter will be consumed too quickly, which will affect the synthesis of the product, and too much metabolic waste will be transferred in; if the inoculum size is too small, the culture time will be prolonged. In the present invention, according to the single factor control experiment, a response surface optimization method based on the central structure design (CCD) is designed. Based on the response value of each factor level, the predicted optimal response value and the experimental conditions of the response can be found to reduce the cadmium content in the contaminated biomass straw as much as possible. In order to better improve the leaching effect of cadmium in the contaminated biomass straw, the contaminated biomass straw is pretreated, and Aspergillus niger extraction is performed based on the optimal conditions of the response surface. Biomass straw is rich in lignocellulose, which is mainly composed of cellulose, hemicellulose and lignin. Due to the protective effect of lignin and hemicellulose on cellulose and the crystal structure of cellulose itself, lignocellulose forms a dense and impermeable high-level structure, making it difficult for enzyme preparations to contact cellulose.In order to make the enzyme react better with cellulose, the present invention uses at least one or a combination of water treatment, hydrothermal treatment, alkali treatment, and enzyme treatment to pretreat biomass straw samples, so that the plant surface lignocellulose is changed, the accessibility of the enzyme preparation is improved, and the cadmium extraction of Aspergillus niger based on the optimal conditions of the response surface is more sufficient. The present invention discloses a treatment method for removing cadmium from contaminated biomass using Aspergillus niger, using Aspergillus niger as an exogenous added fungus to extract the cadmium in the contaminated biomass alone, optimizing the optimal conditions using the response surface method, and using the pretreatment method based on the optimal conditions of the response surface to work together with Aspergillus niger to complete the removal of cadmium from the contaminated biomass. The treatment method of the present invention, in the form of microbial extraction, Aspergillus niger and contaminated biomass are subjected to shake flask extraction and culture, which can achieve effective treatment of cadmium-contaminated biomass after plant repair, with the advantages of good treatment effect, low investment, green environmental protection, low technical management requirements, simple operation, etc., and can also achieve the reuse of the treated biomass as fertilizer, with good environmental benefits, social benefits and economic benefits.

[0021] (2) In the treatment method of the present invention, Aspergillus niger is used as the extraction strain, which can grow normally in the culture medium containing cadmium-contaminated biomass straw, and can not only leach cadmium in the biomass straw, but also use the sucrose in the culture medium to produce low molecular weight organic acids, maintain or even reduce the initial pH value of the system, so as to improve the efficiency of cadmium leaching from biomass straw.

[0022] (3) In the treatment method of the present invention, at least one of the sucrose content, the inoculum size of Aspergillus niger, and the initial pH value of the system is used as a single variable for a control experiment, and a response surface optimization method based on central structure design (CCD) is performed according to the result to optimize a set of optimal sucrose content, Aspergillus niger inoculum size, and the initial pH value of the system for Aspergillus niger to extract cadmium from biomass straw contaminated by Aspergillus niger, so that the Aspergillus niger extraction of cadmium from biomass straw can achieve the maximum efficiency.

[0023] (4) In the treatment method of the present invention, the contaminated biomass straw is pretreated with at least one of water treatment, hydrothermal treatment, alkali treatment, and enzyme treatment, and the biomass straw is pretreated by Aspergillus niger extraction based on the optimal conditions of the response surface, so as to maximize the removal of cadmium in the contaminated biomass straw and ensure the safe utilization of the contaminated biomass straw. DETAILED DESCRIPTION

[0024] The present invention is further described below in conjunction with specific preferred embodiments, but the protection scope of the present invention is not limited thereby.

[0025] The materials and instruments used in the following examples are all commercially available. In the examples of the present invention, unless otherwise specified, the following data are the average values ​​of more than three tests.

[0026] Embodiment 1:

[0027] A method for removing cadmium from contaminated biomass by using Aspergillus niger, specifically using Aspergillus niger to treat contaminated oil sunflower straw, wherein the oil sunflower is planted in Liuyang City, Changsha City, Hunan Province, and the cadmium concentration of the soil in the oil sunflower planting area is 1.01 mg kg -1 The cadmium concentration of sunflower straw was 4.22 mg kg -1 The culture is carried out with at least one of sucrose content, Aspergillus niger inoculation amount, and initial pH value of the system as a single control variable, comprising the following steps:

[0028] (1) The basic culture steps of the single control variable are: the mass of sunflower straw is 5.0000 g; the standard extraction medium includes: NaNO3 1.5 g·L -1 , KH2PO4 0.5g·L -1 , MgSO4.7H2O 0.025g·L -1 , KCl 0.025 g·L -1 , yeast extract 1.6 g·L -1 , sterilized at 121°C for 20 min, the pH value of the culture medium was 5.67; the extraction conditions included: the system volume was 100 mL, the time was 15 d, the temperature was 30°C, and the rotation speed was 150 rpm.

[0029] (2) Taking sucrose content as the single control variable, the steps are as follows: sucrose content control range 40~120g·L -1 , 20g·L -1 The inoculum size of Aspergillus niger was 2% and the pH value was the natural pH value.

[0030] (3) The steps of taking the inoculum size of Aspergillus niger as the single control variable are as follows: the inoculum size of Aspergillus niger is controlled in the range of 2% to 10%, with 2% as a gradient; the sucrose content is 100 g L -1 ; pH value is natural pH value.

[0031] (4) The steps of taking the initial pH value of the system as the single control variable are as follows: the initial pH value of the system is controlled in the range of 3 to 7, increasing in increments of 1; the sucrose content is 100 g·L -1 ; The inoculation amount of Aspergillus niger was 2%; a control group without adding bacteria was set.

[0032] Collection of oil sunflower straw residue after extraction: After one extraction cycle, the oil sunflower straw residue and the extraction supernatant were harvested. The oil sunflower straw residue was fully washed with ultrapure water, dried and recovered at a constant temperature of 55°C; the extraction supernatant was recovered by centrifugation at 4000rpm for 10min; the total cadmium concentration of the oil sunflower straw residue and the total cadmium concentration of the extraction supernatant were determined, and the removal rate, leaching rate and Aspergillus niger adsorption rate of the oil sunflower straw cadmium were calculated. The total cadmium concentration and removal rate of the oil sunflower straw residue with the largest removal amount, the total cadmium concentration and leaching rate of the extraction supernatant and the Aspergillus niger adsorption rate are shown in Table 1.

[0033] Table 1 Total cadmium content and removal rate of sunflower straw residue with the largest removal amount under different control variables, total cadmium content and leaching rate of leaching supernatant and adsorption rate of Aspergillus niger

[0034]

[0035] As can be seen from Table 1, under different control variables, the total cadmium concentration of oil sunflower straw in the optimal pH treatment was lower than that in the sucrose content treatment and the inoculum amount treatment; the cadmium concentration in the supernatant was greater in the optimal pH treatment than in the inoculum amount treatment and the sucrose content treatment. However, the adsorption rate of Aspergillus niger was better in the optimal sucrose content treatment than in the inoculum amount treatment and pH treatment. In summary, the optimal pH treatment acidified the original pH value of the culture medium and increased the H + , which changes the lignocellulose on the surface of the sunflower straw, improves the accessibility of Aspergillus niger to the sunflower straw, and thus increases the leaching of cadmium from the sunflower straw. Sucrose is a simple high-molecular-weight carbon source added as an exogenous source, which can quickly provide energy for the growth and reproduction of Aspergillus niger, allowing Aspergillus niger to reproduce in large quantities. This may be the reason why the adsorption rate of Aspergillus niger under the sucrose content treatment is better than that under the inoculation amount treatment and pH value treatment; at the same time, Aspergillus niger can also use sucrose to produce low-molecular organic acids, maintain or even reduce the pH value of the system, and remove cadmium from the sunflower straw. However, the acidification effect of low-molecular organic acids is weaker than that of H + Therefore, this is the reason why the cadmium concentration in sunflower straw under sucrose content treatment is higher than that under pH treatment. In summary, in order to maximize the reduction of cadmium concentration in sunflower straw, the three treatments of sucrose content, Aspergillus niger inoculation amount and pH value need to work together. Therefore, the subsequent design is based on the response surface optimization method of central structure design (CCD) to obtain the optimal condition combination for removing cadmium from sunflower straw by Aspergillus niger extraction.

[0036] Embodiment 2:

[0037] A method for removing cadmium from contaminated biomass using Aspergillus niger, specifically using high performance liquid chromatography to analyze the concentration of low molecular weight organic acids in a solution, wherein the sample to be measured is the supernatant collected after one leaching cycle in Example 1. The specific steps are as follows:

[0038] (1) Determination of low molecular weight organic acid concentration: The concentration of low molecular weight organic acid in the solution was analyzed by high performance liquid chromatography. The chromatographic column was Agilent 5HC-C18 (150×4.6 mm, 5 μm), the mobile phase was 10 mM KH2PO4 (pH adjusted to 2.6 with H3PO4): chromatographic grade methanol = 93:7, and the flow rate was 0.8 mL min -1; Column temperature was 25℃; Detection wavelength was 220nm. In this experiment, the peak times of oxalic acid, tartaric acid, lactic acid, citric acid and succinic acid were 3.657min, 4.107min, 5.828min, 6.700min and 8.611min respectively.

[0039] (2) Preparation of the sample to be tested: Take 1.5 mL of the extraction supernatant, centrifuge at 13000 rpm for 10 min, take 0.95 mL of the supernatant and add 0.05 mL of 10% (w / w) H3PO4 solution, vortex mix and filter with a 0.22 μm filter membrane to obtain the sample for liquid phase determination. Determine the content of oxalic acid, tartaric acid, lactic acid, citric acid and succinic acid, and calculate the proportions as shown in Table 2.

[0040] Table 2 Proportion of organic acid in the extract under different inoculation conditions

[0041] deal with oxalic acid(%) tartaric acid(%) Lactic acid(%) Citric acid(%) Succinic acid(%) No bacteria added 8.85 8.35 16.48 9.27 57.05 2% inoculation 10.40 10.24 22.86 30.69 25.72 10% inoculation 5.76 8.01 27.69 43.74 15.07

[0042] Note: The sucrose content in different treatments was 100 g·L -1 ; The initial pH value of the system is the natural pH value of 5.67.

[0043] As can be seen from Table 2, the proportion of succinic acid was the largest in the treatment without bacteria; under the 2% inoculation condition, the proportion of succinic acid was greatly reduced, while the proportion of oxalic acid, tartaric acid, lactic acid and citric acid increased, among which the proportion of citric acid increased the most, which was 30.69%; under the 10% inoculation condition, the proportion of succinic acid decreased by 31.98%, the proportion of oxalic acid decreased by 3.09%, and the proportion of tartaric acid, lactic acid and citric acid increased, among which the proportion of citric acid increased the most, which was 43.74%. Compared with the treatment without bacteria, the addition of Aspergillus niger can increase the citric acid in the system, and the 10% inoculation condition treatment is more than the 12% inoculation condition treatment. This is because a large number of Aspergillus niger can use sufficient sucrose to produce more citric acid under acidic conditions; in addition, the oxalic acid under the 2% inoculation condition treatment increased by 1.55% compared with the treatment without bacteria, and the proportion of oxalic acid under the 10% inoculation condition treatment decreased by 3.09% compared with the treatment without bacteria. This is because the increase in the inoculation amount of Aspergillus niger inhibits the formation of oxalic acid that is harmful to Aspergillus niger. In summary, sufficient sucrose and a large amount of Aspergillus niger can be used in subsequent experiments to produce citric acid and inhibit the formation of oxalic acid, thereby achieving the removal of cadmium from sunflower straw.

[0044] Embodiment 3:

[0045] A method for removing cadmium from contaminated biomass using Aspergillus niger, specifically using a pH meter to measure the pH value in a solution, wherein the sample to be measured is the supernatant collected after one extraction cycle in Example 1. The initial pH value of the sucrose content treatment and the inoculation amount treatment is the pH value of the prepared culture medium of 5.67. The pH value in the culture system after extraction under different initial pH value treatments is measured, as shown in Table 3.

[0046] Table 3 pH values ​​in the culture system after extraction under different initial pH values

[0047]

[0048]

[0049] As can be seen from Table 3, the pH value of the extract without inoculation of Aspergillus niger changes, resulting in the pH value in the extract being maintained between 4.98 and 5.73 under different initial pH conditions. It can be seen from Example 1 that when the pH is 3, Aspergillus niger can significantly reduce the concentration of cadmium in oil sunflower straw, and the removal rate of cadmium in oil sunflower straw is the highest. Under the conditions of the initial pH value of the leaching system of the present invention of 3 and 4, Aspergillus niger can maintain the initial pH value of the leaching system and even reduce the pH value of the leaching system; and the cadmium concentration in the leaching solution is higher than that of the control without bacteria, wherein the leaching rate is the highest at 35.67% when the initial pH value is 3. This may be due to the fact that Aspergillus niger can produce more citric acid under the condition of low initial pH value, thereby realizing the leaching of cadmium in oil sunflower straw. In addition, when the initial pH value is above 5, the cadmium concentration in the leaching solution inoculated with Aspergillus niger is lower than that of the leaching solution not inoculated with Aspergillus niger, which is related to the fact that the initial pH value is greater than 5, which is conducive to the germination of Aspergillus niger spores, and Aspergillus niger spores have a certain adsorption effect on cadmium in the leaching solution. In summary, the initial pH value of the extraction system needs to be reduced to below 5 in subsequent extraction tests.

[0050] Embodiment 4:

[0051] A method for removing cadmium from contaminated biomass using Aspergillus niger, specifically using Aspergillus niger to treat contaminated sunflower straw using a response surface optimization method based on central structure design (CCD), wherein the cadmium concentration of the sunflower straw is 4.22 mg kg -1 , the system volume is 100 mL, the time is 15 days, the temperature is 30°C, the rotation speed is 220 rpm, the response factors are sucrose content, Aspergillus niger inoculation amount and the initial pH value of the system, including the following steps:

[0052] (1) The response surface optimization method based on central composite design (CCD) was used to determine the optimal conditions for extracting cadmium from sunflower straw by Aspergillus niger. Three factors and five levels of experimental conditions were formed according to the response factors, and 20 experimental run groups were generated using Design Expert 10 software. The central composite experimental factors and levels are shown in Table 4, and the central composite experimental design is shown in Table 5.

[0053] Table 4 Central composite test factors and levels

[0054]

[0055] Table 5 Central composite experimental design

[0056]

[0057]

[0058] (2) Based on the results of the central composite experimental design, the optimal conditions for extracting cadmium from sunflower straw by Aspergillus niger were optimized using Design Expert 10 software, and the optimal conditions were verified. The conditions after software optimization and the verification results are shown in Table 6.

[0059] Table 6 Response surface optimization and validation results

[0060] <![CDATA[Sucrose content (g·L -1 )]]> Inoculation amount (%) Initial pH value of the system Cadmium removal rate (%) optimization 76.266 10 3 76.20 verify 76.266 10 3 67.67

[0061] Collection of sunflower straw residue after extraction: After one extraction cycle, the sunflower straw residue was harvested. The sunflower straw residue was fully washed with ultrapure water and dried at 55°C for recovery. The variance analysis function in Design Expert 10 software was used to model and analyze the response surface test data of cadmium extraction of sunflower straw by Aspergillus niger, and the results are shown in Table 7.

[0062] Table 7 Variance analysis of binomial model constructed by response surface methodology for Aspergillus niger extraction of sunflower straw

[0063] source sum of squares Degrees of Freedom Mean Square F-number P-value Significance Model 0.0479 9 0.0053 11.4731 0.0004 Very significant A-pH 0.0096 1 0.0096 20.7598 0.0010 B-Sucrose content 0.0059 1 0.0059 12.6719 0.0052 C-Inoculum size 0.0013 1 0.0013 2.8760 0.1208 AB 0.0041 1 0.0041 8.8588 0.0139 AC 0.0034 1 0.0034 7.4059 0.0215 <![CDATA[B 2 ]]> 0.0024 1 0.0024 5.2431 0.0450 <![CDATA[C 2 ]]> 0.0070 1 0.0070 15.0052 0.0031 <![CDATA[A 2 B]]> 0.0055 1 0.0055 11.8981 0.0062 <![CDATA[A 2 B 2 ]]> 0.0111 1 0.0111 23.9567 0.0006 Residual 0.0046 10 0.0005 Lack of Fit 0.0004 5 0.0001 0.0924 0.9897 Not significant Pure Error 0.0043 5 0.0009 All items 0.0526 19

[0064] As can be seen from Table 7, the P value of the model selected for the response surface test of Aspergillus niger extracting cadmium from sunflower straw is 0.0004, which is less than 0.01 and has a very significant effect, indicating that the model is statistically significant. The P value of the lack of fit term is 0.9897, which is greater than 0.05, which is favorable to the model and there is no lack of fit factor. In addition, A-pH, B-sucrose content, AB, AC, B 2 , C 2 , A 2 B. A 2 B 2The effect on the response value is significant (P<0.05). Therefore, the effect of each experimental factor on the response value is not a simple linear relationship. 2 =0.9117, AdjR 2 =0.8322, indicating that there is a variation possibility of 8.83% in the model, and the fitting effect is good; the coefficient of variation is 4.07%, which is a low value, indicating that the test is accurate and reliable, and can be used for theoretical prediction of the optimal process of cadmium removal from sunflower straw by Aspergillus niger extraction. After regression fitting of each factor, the quadratic response surface regression equation is: Cadmium removal rate = 0.54-0.03A+0.03B-0.01C-0.02AB-0.02AC-0.01B 2 +0.02C 2 -0.04A 2 B-0.05A 2 B 2 According to the regression model, the optimal process conditions for cadmium removal from sunflower straw by Aspergillus niger extraction were predicted to be: sucrose content of 76.266 g·L -1 , inoculation amount of 10%, initial pH value of the system is 3, then the removal rate of cadmium in oil sunflower straw can theoretically reach 76.20%. In order to verify whether the predicted results are consistent with the actual situation, a verification test was carried out. The results showed that the actual value of cadmium removal rate based on the optimal process conditions was 67.67%, and the concentration of cadmium in oil sunflower straw was 1.36 mg kg -1 The cadmium concentration in sunflower straw increased from the original concentration of 4.22 mg kg -1 Reduced to 1.36 mg kg -1 , in line with the standards of my country's "Limits of Toxic and Hazardous Substances in Fertilizers (GB 38400-2019)". Among them, the limit of total cadmium content in inorganic fertilizers is ≤10mg·kg -1 , the limit of total cadmium content in other fertilizers is ≤3mg·kg -1 , providing the possibility for the recycling of sunflower straw after Aspergillus niger extraction.

[0065] Embodiment 5:

[0066] A method for removing cadmium from contaminated biomass using Aspergillus niger, specifically using Aspergillus niger to treat contaminated sunflower straw based on time variation, wherein the cadmium concentration of the sunflower straw is 4.22 mg kg -1 , the system volume was 100 mL, the temperature was 30 °C, the rotation speed was 220 rpm, and the response surface optimization conditions (sucrose content was 76.266 g·L -1 , inoculation volume of 10%, initial pH value of the system of 3) were used for the experiment. The specific steps are as follows: the culture time is used as a single control variable, specifically 1d, 3d, 5d, 7d, 9d, 11d, 13d, 15d in sequence.

[0067] Collection of sunflower straw residue after extraction: After one extraction cycle, the sunflower straw residue was harvested. The sunflower straw residue was fully washed with ultrapure water and dried at 55°C for recovery. The concentration of cadmium in the sunflower straw was determined, and the cadmium removal rate was calculated. The results showed that the cadmium concentration in the sunflower straw gradually decreased with the increase of time when Aspergillus niger was optimized based on the response surface. On the 11th day, the cadmium concentration in the sunflower straw reached the lowest level of 1.36 mg kg -1 ; The cadmium concentration in sunflower straw was maintained at 1.36 mg kg on the 13th and 15th days. -1 It can be seen that the present invention uses Aspergillus niger to extract sunflower straw based on the response surface optimization conditions, which reduces 4 days compared with the previous 15-day shake flask culture test, saving time and thus increasing the economic effect.

[0068] Embodiment 6:

[0069] A method for removing cadmium from contaminated biomass by using Aspergillus niger, specifically comprising pre-treating sunflower straw by using at least one of water treatment, hydrothermal treatment, alkali treatment and enzyme treatment, and then extracting the sunflower straw by using Aspergillus niger, wherein the cadmium concentration of the sunflower straw is 4.22 mg kg -1 , the system volume was 100 mL, the temperature was 30 °C, the rotation speed was 220 rpm, and the response surface optimization conditions (sucrose content was 76.266 g·L -1 , inoculation amount of 10%, initial pH value of the system was 3) for the test. The specific steps are as follows:

[0070] (1) Enzyme treatment (E): The crushed sunflower straw was added with cellulase and citric acid buffer solution, pH 5.0, and hydrolyzed in a 50°C incubator for 72 h (the above step is T3). The amount of cellulase used was 10 U g -1 . After washing with ultrapure water, dry it.

[0071] (2) Water treatment (W0): The crushed sunflower straw is added to ultrapure water and treated at room temperature for the same time as the hydrothermal treatment. After squeezing and filtering, it is soaked in ultrapure water for 4 hours. After squeezing and filtering, it is maintained in ultrapure water at 50°C for 72 hours (the above steps are T3). After squeezing and filtering, it is dried.

[0072] (3) Hydrothermal treatment-water treatment-enzyme treatment (WW): Add crushed sunflower straw to ultrapure water, heat to 160°C in a high-pressure reactor, react for 10 minutes, then cool to 120°C, react for 10 minutes, and cool down to terminate the reaction (the above step is T1). After the harvested reactant is cooled, it is squeezed, filtered and washed with water, soaked in ultrapure water for 4 hours, and then quickly squeezed and filtered (the above step is T2). Add the leached residue to a cellulase and citric acid buffer solution with a pH value of 5.0 and hydrolyze in a 50°C incubator for 72 hours (the above step is T3). Then wash the sunflower straw with water until the pH value of the solution after washing is about 7.0, and dry it for storage.

[0073] (4) Hydrothermal treatment - 0.5% alkali treatment - enzyme treatment (W-0.5B): The hydrothermal treatment is T1 as shown in (3). After the harvested reactants are cooled, squeezed, filtered and washed with water, the filter residue is mixed with a 0.5% NaOH solution and soaked for 4 hours, and then quickly squeezed and filtered (the above step is T2). After washing with water, the pH value of the filter residue reaches about 7.0, and cellulase and citric acid buffer solution are added, the pH value is 5.0, and hydrolyzed in a 50°C incubator for 72 hours (the above step is T3). Then the sunflower straw is washed with water until the pH value of the solution after washing is about 7.0, and dried for storage.

[0074] (5) Hydrothermal treatment - 1% alkali treatment - enzyme treatment (W-1B): The hydrothermal treatment is T1 as shown in (3). After the harvested reactants are cooled, squeezed, filtered and washed with water, the filter residue is mixed with a 1% NaOH solution and soaked for 4 hours, and then quickly squeezed and filtered (the above step is T2). After washing with water, the pH value of the filter residue reaches about 7.0, and cellulase and citric acid buffer solution are added, the pH value is 5.0, and hydrolyzed in a 50°C incubator for 72 hours (the above step is T3). Then the sunflower straw is washed with water until the pH value of the solution after washing is about 7.0, and then dried and stored.

[0075] (6) The sunflower straw pretreated with the above five schemes was combined with Aspergillus niger, and a microbial cadmium leaching test was carried out on the sunflower straw based on the response surface optimization conditions.

[0076] Collection of sunflower straw residue after extraction: After one extraction cycle, the sunflower straw residue was harvested and fully washed with ultrapure water, and then dried and recovered at a constant temperature of 55°C. The concentration of cadmium in the pretreated sunflower straw and the sunflower straw after Aspergillus niger extraction was determined, and the removal rate of cadmium in the sunflower straw after Aspergillus niger extraction was calculated. The results are shown in Table 8.

[0077] Table 8 Cadmium concentration in pretreated sunflower straw and cadmium concentration and cadmium removal rate in sunflower straw after Aspergillus niger extraction

[0078]

[0079] As can be seen from Table 8, the cadmium concentration in sunflower straw under treatment E was 2.21 mg kg -1 The removal rate was 47.63%; the cadmium concentration in sunflower straw under W0 treatment was 2.65 mg kg -1 The removal rate was 37.20%. In the present invention, after the hydrothermal alkaline enzyme composite pretreatment, as the NaOH concentration in the second step of alkali treatment increases, the cadmium concentration in the oil sunflower straw decreases more. After different alkali treatments, the cadmium concentration in the oil sunflower straw is 2.24 mg kg -1 (water immersion), 1.98 mg kg -1 (0.5% NaOH) and 1.87 mg kg -1 (1% NaOH); however, the cadmium concentration in oil sunflower straw was basically the same after enzyme treatment. The three-step pretreatment can significantly reduce the cadmium concentration in oil sunflower straw, all below 1 mg kg -1 , respectively 0.9 mg kg -1 (WW), 0.96mg·kg -1 (W-0.5B), 0.9mg·kg -1 (W-1B), the removal rates were 78.67%, 77.25% and 78.67% respectively. After different pretreatments and cadmium leaching with Aspergillus niger in the present invention, the cadmium concentration in the oil sunflower straw decreased, and there was a significant difference in the cadmium concentration of the oil sunflower straw between different pretreatments and cadmium leaching with Aspergillus niger. Compared with single enzyme treatment and water treatment, the continuous three-step pretreatment can help Aspergillus niger extract the cadmium concentration of oil sunflower straw and improve the removal rate of cadmium in oil sunflower straw. The cadmium concentration in the oil sunflower straw obtained was lower than 0.4 mg kg -1 , and the addition of Aspergillus niger after W-0.5B treatment could reduce the cadmium concentration in sunflower straw to 0.33 mg kg -1 , and finally a cadmium removal rate of up to 92.15% can be achieved.

[0080] In summary, the above six embodiments selected a method of hydrothermal treatment-0.5% alkali treatment-enzyme treatment combined with Aspergillus niger extraction, wherein a standard extraction medium (NaNO3 1.5 g·L -1 , KH2PO4 0.5g·L -1 , MgSO4.7H2O 0.025g·L -1 , KCl0.025g·L -1 , yeast extract 1.6 g·L -1 ), the culture volume was 100 mL, the temperature was 30 °C, the rotation speed was 220 rpm, the culture time was 11 days, and the response surface optimization conditions (sucrose content was 76.266 g·L -1, inoculation amount of 10%, and initial pH value of the system of 3) to achieve maximum cadmium removal.

[0081] The above is only a preferred embodiment of the present invention, and the protection scope of the present invention is not limited to the above embodiment. All technical solutions under the concept of the present invention belong to the protection scope of the present invention. It should be pointed out that for ordinary technicians in this technical field, improvements and modifications without departing from the principle of the present invention should also be regarded as the protection scope of the present invention.

Claims

1. A method for removing cadmium from contaminated biomass using Aspergillus niger, characterized in that: The processing method comprises the following steps: (1) Pre-treatment of contaminated biomass; (2) Extraction medium: NaNO3 1.5~3 g·L -1 , KH2PO4 0.1~1 g·L -1 ,MgSO4 . 7H2O 0.025~1g·L -1 , KCl 0.025~0.5 g ·L -1 , yeast extract 1.6~5 g·L -1 ; Sucrose content is 70-120 g·L -1 , initial pH is 2-5; (3) adding the contaminated biomass pretreated in step (1) to the standard extraction medium of step (2) and inoculating Aspergillus niger for extraction; The contaminated biomass pretreatment includes at least one of water treatment, hydrothermal treatment, alkali treatment, and enzyme treatment; The polluted biomass is crop straw, which is one of sunflower straw, rape straw and sesame straw; The Aspergillus niger was purchased from the China Industrial Microbiological Culture Collection Administration Center, with the culture collection number being CICC 2214. The Aspergillus niger was a spore suspension with a concentration of 4.25×10 5 ~10 8 mL -1 ; The inoculation amount of Aspergillus niger is 8% to 12%.

2. The processing method according to claim 1, characterized in that: The hydrothermal treatment is to maintain it at 160°C for 10 min, then cool it to 120°C and maintain it for 10 min; the alkali treatment is 0.5% to 1% sodium hydroxide by mass, soaking for 4 h; the enzyme treatment is cellulase produced from Aspergillus niger, hydrolyzed at 50°C for 72 h; the water treatment is ultrapure water at room temperature for the same time as the hydrothermal treatment, soaking in ultrapure water for 4 h after extrusion filtration, and maintaining ultrapure water at 50°C for 72 h after extrusion filtration.

3. The processing method according to any one of claims 1-2, characterized in that: In step (2), the sucrose content is 76.266 g·L -1 The inoculation amount of Aspergillus niger was 10%, and the initial pH value of the standard extraction medium was 3.

4. The processing method according to claim 1, characterized in that: In step (3), the extraction temperature is 30°C, the incubation time is 11 to 15 days, and the rotation speed is 150 to 220 rpm.

5. The processing method according to claim 4, characterized in that: The culture time is 11 days and the rotation speed is 220 rpm.

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