Iron-source synergistic penicillium oxalicum agent and stepped expanding culture preparation and application thereof for accelerating red mud soil formation

Abstract

The invention belongs to the technical field of microbial culture and red mud storage yard ecological restoration, and particularly discloses an iron-source synergistic penicillium oxalicum agent and stepped expanding culture preparation and application thereof for accelerating red mud soil formation. The preparation method of the penicillium oxalicum agent comprises the following steps: inoculating activated penicillium oxalicum spore suspension into a liquid culture medium for culturing, adding ferric salt, and culturing to obtain a primary seed culture solution; inoculating the first-stage seed culture solution into a liquid culture medium, adding ferric salt, and culturing to obtain a second-stage seed culture solution; and inoculating the secondary seed culture solution into a production culture medium of a production tank, adding ferric salt, introducing air, and culturing to obtain the iron-source synergistic penicillium oxalicum agent which is mature by expanding culture. The viable count of the penicillium oxalicum after propagation and the activity of citric acid and cellulase secreted by the penicillium oxalicum are remarkably improved, the saline-alkaline property of the red mud storage yard can be effectively improved, and a good development habitat is provided for plants. The invention further provides a method for applying the synergistic fungicide to red mud soil.

Description

Technical Field

[0001] This invention relates to the fields of microbial culture and ecological restoration of red mud dumps, and in particular to an iron-enhancing Penicillium oxalate agent and its step-by-step expansion preparation and application to accelerate the soilification of red mud. Background Technology

[0002] Red mud is a solid waste generated during alumina production. Due to its inherent characteristics such as high alkalinity, high salinity, and poor physical structure, its resource utilization rate is currently less than 5%. Red mud soilification is a core environmental remediation approach prioritized by alumina companies and environmental management departments. This involves using physical, chemical, or biological methods to regulate the physicochemical properties and structure of red mud, enabling it to possess basic soil functional characteristics, including water and fertilizer retention capacity and plant growth support. Microbial remediation technology has become a research hotspot in red mud soilification due to its advantages such as low cost, ease of cultivation, environmental friendliness, and strong sustainability.

[0003] Penicillium oxalicum, due to its ability to secrete organic acids such as oxalic acid and citric acid to reduce the alkalinity of red mud, has become a core microbial species in current red mud alkalinity reduction and remediation technologies. However, the secretion of these organic acids and the activity of their extracellular enzymes both require iron as a key coenzyme factor. Iron ions, as an essential cofactor for enzymatic reactions such as citric acid synthase, are a crucial material basis for promoting the efficient operation of microbial metabolic pathways and the synthesis of organic acids. The available iron content in red mud matrix is ​​extremely low (usually <0.05 g / kg), and it mostly exists in the form of insoluble iron oxides such as hematite and goethite, which are difficult for Penicillium oxalicum to effectively absorb and utilize. In red mud remediation projects, Penicillium oxalate, when applied alone, is susceptible to the inhibitory stress of the extreme physicochemical environment of red mud (high alkalinity, low available iron, and low organic matter). Insufficient supply of free available iron leads to limited acid secretion and decreased functional enzyme activity, ultimately restricting the metabolic efficiency of the fungi and the remediation effect. This technical bottleneck is a significant factor hindering the widespread application of Penicillium oxalate in large-scale red mud remediation. Therefore, increasing the iron source during cultivation and expanding the culture to obtain Penicillium oxalate strains with strong metabolic activity and high functional stability can effectively enhance its organic acid secretion efficiency and organic substrate decomposition capacity in the red mud environment, thereby efficiently promoting the soilification of red mud in the remediation site. This provides core technical support for large-scale vegetation reconstruction in red mud. However, significant technological gaps still exist in this field, urgently requiring breakthroughs and filling.

[0004] Therefore, it is necessary to optimize and improve the Penicillium oxalate agent applied to red mud soil treatment to enhance the agent's performance and its effect on improving red mud soil treatment. Summary of the Invention

[0005] This invention aims to at least solve one of the technical problems existing in the prior art. To this end, this invention provides a method for preparing an iron-enhanced Penicillium oxalate inoculant. This invention utilizes a stepwise iron source introduction method for Penicillium oxalate cultivation. By adding different concentrations of iron source at different cultivation stages, Penicillium oxalate is directionally induced to proliferate and synthesize functional products. The viable count and the activities of citric acid and cellulase secreted by the expanded Penicillium oxalate inoculant are significantly increased. Applying this inoculant to red mud soil can effectively improve the salinity of red mud dumps, providing a favorable growing environment for plants.

[0006] The present invention also provides a Penicillium oxalate antifungal agent.

[0007] This invention also provides a method for red mud soilification.

[0008] In a first aspect, the present invention provides a method for preparing an iron-enhancing Penicillium oxalate fungicide, comprising the following steps:

[0009] S1. Inoculate the activated Penicillium oxalate spore suspension into a liquid culture medium and add iron salt to make the final concentration of iron salt in the culture medium 0.005~0.05g / L. After culturing for 1~2 days, the first-level seed culture medium is obtained.

[0010] S2. Inoculate the primary seed culture medium into the liquid culture medium, add iron salt to make the final concentration of iron salt in the culture medium 0.03~0.1g / L, and culture for 2~4 days to obtain the secondary seed culture medium;

[0011] S3. Inoculate the secondary seed culture medium into the production culture medium of the production tank, add iron salt to make the final concentration of iron salt in the culture medium 0.04~0.2g / L, introduce air, and after culturing for 2~4 days, the expanded and mature iron source enhanced Penicillium oxalate agent is obtained.

[0012] The spore concentration of the Penicillium oxalate spore suspension was 5 × 10⁻⁶. 7 ~1×10 8 per mL.

[0013] According to some embodiments of the present invention, the method for preparing the Penicillium oxalate spore suspension in step S1 is as follows: scrape the activated Penicillium oxalate spores from the culture medium plate and mix them with sterile water to prepare a spore suspension, which is then refrigerated and stored at 4°C for inoculation and later use.

[0014] According to some embodiments of the present invention, the iron salt is Fe2(SO4)3 or FeCl3.

[0015] According to some preferred embodiments of the present invention, the iron salt is Fe2(SO4)3.

[0016] According to some embodiments of the present invention, the concentration of iron salt in the culture medium in step S1 is preferably 0.008~0.03 g / L; more preferably 0.008~0.02 g / L.

[0017] According to some embodiments of the present invention, the concentration of iron salt in the culture medium in step S2 is preferably 0.03~0.08 g / L; more preferably 0.04~0.07 g / L.

[0018] According to some embodiments of the present invention, the concentration of iron salt in the culture medium in step S3 is preferably 0.03~0.15 g / L; more preferably 0.04~0.07 g / L.

[0019] According to some embodiments of the present invention, the liquid culture medium is sterile potato glucose liquid culture medium (PDB).

[0020] According to some embodiments of the present invention, the components of the production culture medium include glucose, yeast extract, sodium chloride and straw, wherein the mass ratio of glucose, yeast extract, sodium chloride and straw is 45~55:20~30:4~6:1.

[0021] According to some embodiments of the present invention, the straw is one or more of corn straw, wheat straw or cotton straw.

[0022] According to some embodiments of the present invention, the length of the straw is controlled to be within 1 cm.

[0023] According to some embodiments of the present invention, in step S1, the volume ratio of the Penicillium oxalate spore suspension to the liquid culture medium is 1:80~120; in step S2, the volume ratio of the primary seed culture medium to the liquid culture medium is 1:8~15; in step S3, the volume ratio of the secondary seed culture medium to the production culture medium is 1:20~30.

[0024] This invention further explores the number of strains in the culture system during the stepped culture process, and, in conjunction with the induction effect of different concentrations of iron source at each stage, further promotes the growth and reproduction of strains and the synthesis of metabolites. In the production culture stage, by precisely controlling the strain density, iron source concentration and other nutrient content, Penicillium oxalate is directionally induced to synthesize a large amount of functional products such as oxalic acid and cellulase, while humic acid is used to enhance the stability of the inoculum.

[0025] According to some embodiments of the present invention, the culture conditions for steps S1 to S3 are as follows: the temperature is 25 to 30°C, the rotation speed of the shaker or stirrer is 150 to 250 r / min, and the air flow rate introduced in step S3 is 1 to 3 L / min.

[0026] According to some embodiments of the present invention, if a common production tank that cannot strictly meet the conditions of temperature and vibration is used in step S3, i.e., stirring and shaking are carried out at room temperature, the strain expansion status needs to be checked once every 48 hours. When the corn stalks in the production tank disappear and there are ≥10 mycelial balls per 50 mL of bacterial solution, the third-level expansion is considered mature.

[0027] In a second aspect, the present invention provides a Penicillium oxalate fungicide, wherein the Penicillium oxalate fungicide is prepared by the preparation method described in the first aspect of the present invention.

[0028] According to some embodiments of the present invention, the viable colony count in the Penicillium oxalate fungicide is 4 × 10⁻⁶. 9 ~1×10 10 CFU / L, mycelial ball density is 20~50 / L.

[0029] A third aspect of the present invention provides a method for red mud soilification, comprising the following steps:

[0030] S10. After the surface of the red mud stockpile is crushed and air-dried, it is subjected to pre-alkali treatment to obtain pre-alkali red mud.

[0031] S20. Spray the oxalic acid penicillin agent described in the second aspect of the present invention onto the surface of the pre-dealkalized red mud;

[0032] S30. Keep the surface of the pre-dealkali red mud moist, control the moisture content to 40%~60%, and the soil-like red mud will be obtained after 7~14 days.

[0033] According to some embodiments of the present invention, the method of pre-alkali treatment is as follows: 1% to 4% by weight of calcium-containing industrial solid waste is mixed with red mud, water is added to adjust the moisture content of the red mud to 55% to 75%, and the mixture is naturally cured for 3 to 7 days to obtain pre-alkali-treated red mud.

[0034] According to some embodiments of the present invention, the pH value of the pre-dealkali red mud is 9.0~10.5.

[0035] According to some embodiments of the present invention, the calcium-containing industrial solid waste includes one or more combinations of desulfurized gypsum and phosphogypsum (the main component of which is CaSO4·2H2O).

[0036] According to some embodiments of the present invention, the spraying rate of the Penicillium oxalate fungicide is ≥8 L / m³. 3 .

[0037] According to some embodiments of the present invention, the method of red mud soilification further includes sowing seeds on the soilified red mud; the seeds are one or more of herbaceous or shrub seeds; the herbaceous plants are one or more of pioneer plants represented by bermudagrass, tall fescue, alfalfa, and ryegrass.

[0038] The beneficial effects of this invention are:

[0039] 1) In addition to promoting the synthesis of functional products and increasing the number of effective viable bacteria, the iron-enhancing Penicillium oxalate pretreatment agent of this invention also reduces the amount of calcium-containing modifiers such as gypsum by 50% in the red mud dealkalization pretreatment stage, under the same dealkalization effect (compared to the technical solution of Chinese patent CN109224364A). In the red mud dealkalization pretreatment stage, traditional methods rely on a large amount of calcium-containing industrial solid waste (>4%) to neutralize alkalinity; while the iron ion introduction of this invention enhances the oxalic acid secretion capacity of Penicillium oxalate, continuously neutralizing the alkalinity of red mud through microbial metabolic acid production. At the same time, the iron salt serves as a nutrient, ensuring that the fungi maintain high activity in a high-alkaline environment, significantly reducing material costs and the risk of secondary pollution, while avoiding the red mud compaction problem caused by excessive gypsum.

[0040] 2) Iron-enhanced Penicillium oxalate fungicide can significantly improve the soil-forming efficiency of red mud. In this invention, the step-by-step cultivation of Penicillium oxalate, in conjunction with iron source regulation, directionally activates the secretion of cellulase and ligninase in Penicillium oxalate, accelerating the decomposition of organic materials such as straw, and rapidly replenishing key soil components such as humus for red mud. Simultaneously, iron ions promote the synthesis of extracellular polysaccharides and iron oxide complexes by the fungi, significantly enhancing the stability of red mud aggregates (increasing the proportion of aggregates >0.25mm). This invention shortens the traditional red mud soil-forming engineering process from 60-90 days to 30-45 days, greatly improving ecological restoration efficiency and creating conditions for rapid vegetation establishment.

[0041] 3) The innovation of this invention lies in its pioneering preparation process of Penicillium oxalate-containing fungal agents, which combines iron source enhancement with step-by-step expansion cultivation. By precisely controlling the type, concentration, and timing of iron source addition in stages, a high degree of matching is achieved between fungal growth, metabolite synthesis, and the needs of red mud soilification, distinguishing it from traditional single iron source addition or simple expansion cultivation methods. This reveals a new mechanism by which iron sources regulate Penicillium oxalate metabolism, clarifying that iron ions promote the synergistic secretion of functional products such as oxalic acid and cellulase by influencing siderophore synthesis, enzyme activity, and gene expression, thus providing multi-dimensional improvements for red mud soilification. When applied to red mud dump ecological restoration projects, this technology can reduce the required amount of iron-containing industrial solid waste by 50%, effectively supporting vegetation reconstruction and providing an efficient technical solution for red mud soilification and ecological restoration.

[0042] Other features and advantages of the invention will be set forth in the description which follows, and will be apparent in part from the description, or may be learned by practicing the invention. Attached Figure Description

[0043] The present invention will be further described below with reference to the accompanying drawings and embodiments, wherein:

[0044] Figure 1Figure A shows the activity characteristics of the bacterial agents in each example and comparative example after expansion culture; Figure B shows the concentration of viable bacteria in the bacterial agents prepared in each example and comparative example, and Figure B shows the mycelial ball density of the bacterial agents prepared in each example and comparative example.

[0045] Figure 2 The changes in citric acid production in the remediated red mud after application are shown in the examples and comparative examples.

[0046] Figure 3 The changes in cellulase activity in red mud were repaired in each of the various embodiments and comparative examples after application.

[0047] Figure 4 Figure A shows the salinity and alkalinity characteristics of the red mud after application in each embodiment and comparative example; Figure B shows the pH value of the red mud after treatment with the bacterial agents prepared in each embodiment and comparative example, and Figure B shows the EC value of the red mud after treatment with the bacterial agents prepared in each embodiment and comparative example.

[0048] Figure 5 For bacterial culture preparation, propagation and field application;

[0049] Figure 6 A real-life scene of vegetation reconstruction applied in red mud soil conversion engineering. Detailed Implementation

[0050] The following will describe the concept and technical effects of the present invention clearly and completely with reference to embodiments, so as to fully understand the purpose, features and effects of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are all within the scope of protection of the present invention.

[0051] The *Penicillium oxalicum* strain used in the examples was derived from the *Penicillium oxalicum* strain described in Chinese patent application CN109224364A (the preparation of the *Penicillium oxalicum* agent in the examples can also utilize commercially available *Penicillium oxalicum*). Unless otherwise specified, conditions in the examples were performed under standard conditions or as recommended by the manufacturer. Reagents or instruments whose manufacturers were not specified were all commercially available products.

[0052] Example 1

[0053] This embodiment provides the preparation and application of Penicillium oxalate fungicide regulated by stepwise low concentration Fe2(SO4)3.

[0054] The preparation method of the oxalic acid penicillin agent in this embodiment is as follows:

[0055] 1) Preparation of spore suspension: *Penicillium oxalate* strains were inoculated onto PDA plates (potato extract powder 20 g / L, glucose 20 g / L, agar 15 g / L, sterilized at 121℃ for 20 min). *Penicillium oxalate* colonies activated for 72 h were collected from the plates, and surface spores were scraped off using a sterile inoculation loop and added to a 50 mL centrifuge tube containing 10 mL of sterile water. The tubes were vortexed for 30 s to fully disperse the spores. The spores were filtered through four layers of sterile gauze to remove hyphal fragments. A hemocytometer was used for counting, and the spore concentration was adjusted to 9 × 10⁻⁶ spores with sterile water. 7 Quantity / mL, store at 4℃ for later use;

[0056] 2) Primary seed propagation: Prepare sterilized potato glucose liquid culture medium (PDB) (weigh 200g potato extract powder, add 1000mL distilled water, heat to boiling for 30 min, filter through 4 layers of gauze to collect the filtrate, add 20g glucose to the filtrate and stir until completely dissolved, add distilled water to make up to 1000mL, adjust pH to 5.5-6.0, dispense and sterilize at 121 ℃, 0.1 MPa for 20 min); after the sterilized PDB medium cools to below 50℃, add Fe2(SO4)3 solution sterilized through a 0.22 μm filter membrane to make the final Fe2(SO4)3 concentration in the medium 0.01 g / L; inoculate the prepared spore suspension into the medium at a 1% (v / v) inoculation rate, and incubate in a constant temperature shaker at 28℃, 180 r / min for 2 days. After the incubation, use a hemocytometer to determine the spore concentration of the primary seeds to ensure that it is maintained at 9×10⁻⁶. 7 cells / mL;

[0057] 3) Secondary seed propagation: Prepare and sterilize PDB medium, and after cooling, add sterile Fe2(SO4)3 solution to make the final Fe2(SO4)3 concentration 0.05 g / L; inoculate the primary seed culture solution into the medium at a volume ratio of 1:10, and incubate in a constant temperature shaker at 28℃ and 180 rpm for 72 h. During this period, take samples every 24 h for microscopic examination to observe the mycelial morphology (to ensure that the mycelial branches are abundant and there is no autolysis). After the culture is completed, the secondary seed is obtained.

[0058] 4) Tertiary production expansion: Prepare the following ingredients according to the ratio of edible glucose: yeast extract: sodium chloride: corn stalks = 50:25:5:1 (corn stalks crushed to within 1 cm in length), dissolve and bring to a final volume with sterile water, dispense into production tanks, and sterilize at 121 ℃ and 0.1 MPa for 30 min; after the culture medium cools, add Fe2(SO4)3 solution sterilized through a 0.22 μm filter membrane to make the final Fe2(SO4)3 concentration 0.05 g / L; inoculate the secondary seed culture medium into the production tank culture medium at a volume ratio of 1:25, set the culture parameters as follows: aeration rate >1 L / min, temperature 28℃, stirring speed 200 r / min, and continue to culture for 72 h. During this period, take samples regularly to detect the mycelial growth status. After the culture is completed, the mature iron-containing Penicillium oxalate culture is obtained.

[0059] This embodiment further utilizes the above-obtained, matured iron-enhancing Penicillium oxalate-containing liquid for accelerating the conversion of red mud into soil. The specific steps are as follows:

[0060] 1) Select red mud from the surface of the red mud dump, first crush it to remove large impurities, and then air dry it naturally until the moisture content is 40%;

[0061] 2) Add desulfurized gypsum at a ratio of 2% of the dry weight of red mud, adjust the moisture content of red mud to 60% with an appropriate amount of water, and place it in a natural environment for 5 days. During this period, stir it regularly to ensure that the reaction is complete, and finally obtain pre-dealkali red mud with a pH of 9.2~10.2.

[0062] 3) Take the above-mentioned matured iron-containing Penicillium oxalate culture and add it at a rate of ≥8 L / m³. 3 The dosage is evenly sprayed onto the surface of the red mud stockpile after pre-alkali removal; after spraying, the surface of the stockpile is kept moist by regularly spraying water to control the moisture content of the red mud at 60% and continue to maintain it for 14 days, during which time external debris pollution and excessive trampling are avoided.

[0063] 4) After 14 days of curing, sow Bermuda grass seeds on the surface of the treated red mud dump at a rate of 15 g / m². 2 .

[0064] Example 2

[0065] This embodiment provides the preparation and application of Penicillium oxalate agent regulated by stepwise low concentration Fe2(SO4)3. The preparation method of this embodiment is basically the same as that of Example 1, except that in the first-stage expansion culture in step 2), Fe2(SO4)3 is added to the PDB medium at a final concentration of 0.03 g / L; in the second-stage expansion culture in step 3), Fe2(SO4)3 is added to the PDB medium at a final concentration of 0.075 g / L; and in the third-stage expansion culture in step 4), Fe2(SO4)3 is added to the production tank medium at a final concentration of 0.15 g / L.

[0066] Example 3

[0067] This embodiment provides the preparation and application of a stepwise low-concentration Fe2(SO4)3-regulated Penicillium oxalate agent. The preparation method of this embodiment is basically the same as that of Example 1. The difference is that in the first-stage expansion culture in step 2), Fe2(SO4)3 is added to the PDB medium at a final concentration of 0.05 g / L; in the second-stage expansion culture in step 3), Fe2(SO4)3 is added to the PDB medium at a final concentration of 0.1 g / L; and in the third-stage expansion culture in step 4), Fe2(SO4)3 is added to the production tank medium at a final concentration of 0.2 g / L.

[0068] Comparative Example 1

[0069] This comparative example provides the preparation and application of conventional Penicillium oxalate propagation agents without the participation of iron source. The preparation methods of this comparative example and Example 1 are basically the same. The difference is that in this comparative example, Fe2(SO4)3 solution is not introduced during the entire Penicillium oxalate propagation process. When applied to soil, desulfurized gypsum is added at a ratio of 4% of the dry weight of red mud.

[0070] Comparative Example 2

[0071] This comparative example provides the preparation and application of Penicillium oxalate inoculum with iron source expansion. The preparation methods of this comparative example and Example 1 are basically the same, except that in this comparative example, Fe2(SO4)3 solution with a final concentration of 0.03 g / L is added only in the first-stage expansion (i.e., step 2 of Penicillium oxalate inoculum preparation), and no iron source is introduced in the second-stage and third-stage expansion processes.

[0072] Comparative Example 3

[0073] This comparative example provides the preparation and application of Penicillium oxalate inoculum with iron source expansion. The preparation methods of this comparative example and Example 1 are basically the same, except that in this comparative example, Fe2(SO4)3 solution with a final concentration of 0.075 g / L is added only in the secondary expansion (i.e. step 3 of Penicillium oxalate inoculum preparation). No iron source is introduced in the primary and tertiary expansion processes.

[0074] Comparative Example 4

[0075] This comparative example provides the preparation and application of Penicillium oxalate agaricus preparation with iron source expansion. The preparation methods of this comparative example and Example 1 are basically the same, except that in this comparative example, Fe2(SO4)3 solution with a final concentration of 0.15 g / L is added only in the third-stage expansion (i.e. step 4 of the preparation of Penicillium oxalate agaricus preparation). No iron source is introduced in the first-stage and second-stage expansion processes.

[0076] Performance test comparison:

[0077] 1) The activity characteristics of the Penicillium oxalate fungicides prepared in each example and comparative example were determined, mainly testing the concentration of viable bacteria and the mycelial density in the fungal agents. The results are as follows: Figure 1 As shown;

[0078] Depend on Figure 1 As shown in Figure A, the viable colony counts of Examples 1, 2, and 3 were significantly higher than those of Comparative Examples 1-4, with Example 1 showing a viable colony count of approximately 8.5 × 10⁻⁶. 9 CFU / L, approximately 7.2 × 10⁻⁶ in Example 2. 9 CFU / L, approximately 5.8 × 10⁻⁶ in Example 3. 9 CFU / L, however, the viable colony count in Comparative Example 1 was only about 2.8 × 10⁻⁶. 8 The CFU / L example demonstrates how introducing a stepped iron source to induce expansion culture can more efficiently promote the reproduction of Penicillium oxalate microorganisms and maintain a high number of viable bacteria.

[0079] Depend on Figure 1 As shown in Figure B, the mycelial ball density of Examples 1, 2, and 3 is significantly higher than that of the comparative example. Example 1 shows approximately 38 cells / L, while Comparative Example 1 shows only approximately 8 cells / L. Combined with Figure A, the groups with higher viable cell counts also have higher mycelial ball density, indicating that abundant viable cells provide sufficient "raw materials" for mycelial ball formation. This further demonstrates that the low-concentration, tiered introduction of iron source in Example 1 is more effective in promoting microbial growth and mycelial ball development, exhibiting a clear advantage in microbial culture or related applications.

[0080] 2) The citric acid yield of the Penicillium oxalate fungicides prepared in each example and comparative example was determined, and the results are as follows: Figure 2 As shown:

[0081] Depend on Figure 2It can be seen that the citric acid yield of Examples 1, 2, and 3 is much higher than that of Comparative Examples 1-4, with Example 1 showing the highest citric acid yield at 1050 mg / L; Example 2 reaching 820 mg / L; and Example 3 reaching 800 mg / L. In contrast, Comparative Example 1 has an extremely low citric acid yield of only about 100 mg / L. The stepped iron source introduction technology used in the embodiments of this invention can significantly increase the yield of citric acid and has obvious advantages in citric acid production-related applications, while the production effect of the comparative examples is far inferior to that of the examples. After introducing a low-concentration stepped iron source to cultivate Penicillium oxalate, the acid production of Penicillium oxalate was significantly increased, and the effects of medium and high concentrations of iron sources were similar, slightly lower than those of low concentrations.

[0082] 3) The cellulase activity of the Penicillium oxalate fungicides prepared in each example and comparative example was determined, and the results are as follows: Figure 3 As shown:

[0083] Depend on Figure 3 It can be seen that the cellulase activities of Examples 1, 2, and 3 are significantly higher than those of the comparative examples, with Example 1 exhibiting the highest cellulase activity, reaching 230 IU / g. This indicates that the stepwise iron source introduction and expansion method of the present invention can significantly enhance the cellulase activity of Penicillium oxalate, demonstrating clear advantages.

[0084] 4) The salinity and alkalinity characteristics of the red mud remediated after the application of the Penicillium oxalate fungicide prepared in each example and comparative example to red mud soil were determined. The results are as follows: Figure 4 As shown in Figures A and B:

[0085] Depend on Figure 4 As shown in Figure A, the pH of the red mud in Examples 1, 2, and 3 decreased significantly, to approximately 7.9, 8.3, and 8.2, respectively; while the pH of the comparative examples was higher, and the pH values ​​were ranked as follows: Comparative Example 1 > Comparative Example 4 > Comparative Example 2 > Comparative Example 3. There was a highly significant difference between the examples and the comparative examples. Figure 4 Figure B shows a comparison of electrical conductivity (EC) tests. Examples 1, 2, and 3 all significantly reduced the EC value of red mud to approximately 230 μS / cm; while Comparative Example 1 had an EC value of approximately 950 μS / cm, and Comparative Examples 2, 3, and 4 also had relatively high EC values, around 600-800 μS / cm. There were also highly significant differences between the examples and the comparative examples. Overall, the stepwise iron source expansion method using iron salts introduced in the embodiments of this invention can significantly improve the alkalinity and salt ion concentration within the red mud system.

[0086] 5) such as Figure 5 As shown, the preparation, propagation, and field application of Penicillium oxalate fungicide in various embodiments of the present invention are illustrated. Bermuda grass seeds were sown on the surface of a treated and cured red mud dump, and the results are as follows: Figure 6As shown, the red mud in each embodiment was successfully fertilized and can support normal plant growth. The fertilization effect of the red mud in the comparative embodiment was significantly worse, resulting in poorer plant growth. The results indicate that the Penicillium oxalate fungicide cultivated according to the present invention can be effectively used for the improvement of red mud.

[0087] Furthermore, in the oxalic acid penicillin agent prepared by this invention, Fe 3+ Complexation-hydrolysis-reprecipitation with oxalic acid induces in-situ formation of goethite / Schwertmannite / jaundice, achieving Na + Synergistic fixation and pH reduction, balancing structural improvement and salinity mitigation. In the bioremediation of red mud, Fe... 3+ As a bridging center, it promotes the binding of humic acid / fulvic acid with minerals, enhances stability and specific surface area, and strengthens nutrient slow release and metal (likely) fixation. When Fe2(SO4)3 is used as the iron source, it can combine with the existing SO4 in the red mud system. 2- Pathway compatibility, will not introduce additional Cl - Elements such as these help reduce the risk of salt damage and corrosion, and improve on-site operability.

[0088] The embodiments of the present invention have been described in detail above. However, the present invention is not limited to the above embodiments. Within the scope of knowledge possessed by those skilled in the art, various changes can be made without departing from the spirit of the present invention. Furthermore, the embodiments of the present invention and the features thereof can be combined with each other unless otherwise specified.

Claims

1. A method for preparing an iron-enhancing Penicillium oxalate fungicide, characterized in that, Includes the following steps: S1. Inoculate the activated Penicillium oxalate spore suspension into a liquid culture medium and add iron salt to make the final concentration of iron salt in the culture medium 0.005~0.05g / L. After culturing for 1~2 days, the first-level seed culture medium is obtained. S2. Inoculate the primary seed culture medium into the liquid culture medium, add iron salt to make the final concentration of iron salt in the culture medium 0.03~0.1g / L, and culture for 2~4 days to obtain the secondary seed culture medium; S3. Inoculate the secondary seed culture medium into the production culture medium of the production tank, add iron salt to make the final concentration of iron salt in the culture medium 0.04~0.2g / L, introduce air, and after culturing for 2~4 days, the expanded and mature iron source enhanced Penicillium oxalate agent is obtained. The spore concentration of the Penicillium oxalate spore suspension was 5 × 10⁻⁶. 7 ~1×10 8 per mL.

2. The preparation method according to claim 1, characterized in that, The iron salt is Fe2(SO4)3 or FeCl3.

3. The preparation method according to claim 1, characterized in that, The liquid culture medium is sterile potato glucose liquid culture medium (PDB); the components of the production culture medium include glucose, yeast extract, sodium chloride and straw.

4. The preparation method according to claim 1, characterized in that, In step S1, the volume ratio of the Penicillium oxalate spore suspension to the liquid culture medium is 1:80~120; in step S2, the volume ratio of the primary seed culture medium to the liquid culture medium is 1:8~15; in step S3, the volume ratio of the secondary seed culture medium to the production culture medium is 1:20~30.

5. The preparation method according to claim 1, characterized in that, The cultivation conditions for steps S1 to S3 are as follows: temperature is 25 to 30°C, and the speed of shaking or stirring is 150 to 250 r / min; the air flow rate in step S3 is 1 to 3 L / min.

6. A Penicillium oxalate antifungal agent, characterized in that, The oxalic acid penicillin agent is prepared by the preparation method according to any one of claims 1 to 5.

7. The oxalate penicillin agent according to claim 6, characterized in that, The viable colony count in the Penicillium oxalate fungicide is 4 × 10⁻⁶. 9 ~1×10 10 CFU / L, mycelial ball density is 20~50 / L.

8. A method for red mud soil conversion, characterized in that, Includes the following steps: S10. After the surface of the red mud stockpile is crushed and air-dried, it is subjected to pre-alkali treatment to obtain pre-alkali red mud. S20. Spray the oxalic acid penicillin agent according to claim 6 onto the surface of the pre-dealkalized red mud; S30. Keep the surface of the pre-dealkali red mud moist, control the moisture content to 40%~60%, and the soil-like red mud will be obtained after 7~14 days.

9. The method according to claim 8, characterized in that, The method for pre-alkali treatment is as follows: 1% to 4% of calcium-containing industrial solid waste by weight of red mud is mixed with red mud, water is added to adjust the moisture content of red mud to 55% to 75%, and the mixture is naturally cured for 3 to 7 days to obtain pre-alkali red mud.

10. The method according to claim 8, characterized in that, The spraying rate of the Penicillium oxalate fungicide is ≥8 L / m³. 3 .

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