A method for improving carbon and nitrogen content and aggregate content of newly reclaimed farmland soil by using microalgae

Abstract

The present application relates to a method for improving the carbon and nitrogen content and the aggregate content of newly reclaimed farmland soil by using microalgae, which comprises: applying mixed algal liquid of chlorella and nitrogen-fixing anabaena to the soil. In order to solve the problem of low carbon and nitrogen content and low productivity of newly reclaimed farmland soil, the present application utilizes the ability of chlorella and nitrogen-fixing anabaena to synergistically and efficiently fix carbon dioxide in the air into organic carbon stored in the soil, and at the same time, through the biological nitrogen fixation effect, the nitrogen in the air is converted into nitrogen available to plants, thereby effectively improving the carbon and nitrogen retention capacity of the soil. In addition, the physiological activities of the two microalgae can also synergistically promote the formation of soil aggregates, improve the soil structure, and enhance the water and fertilizer retention capacity of the soil.

Description

Technical Field

[0001] This invention belongs to the field of soil improvement technology and relates to a method for improving the carbon and nitrogen content and aggregate content of newly reclaimed farmland soil using microalgae. Background Technology

[0002] While urban construction and industrial development occupy arable land, it is necessary to reclaim unused hilly areas and tidal flats to create new arable land. The most critical and common limiting factor for newly reclaimed arable land is its low carbon and nitrogen content. This directly leads to poor soil fertility, resulting in land that is often referred to as "raw land" or "silent land." During reclamation, the topsoil (mature layer), which was originally rich in organic matter and nutrients, is often destroyed or removed, exposing the deep, unweathered subsoil or parent material. These soil layers themselves have extremely low organic matter and nitrogen content. Before reclamation, the original natural vegetation (such as grasslands and shrubs) continuously contributes organic matter to the soil through fallen leaves and roots. After reclamation, this natural material cycle is completely disrupted, and a new artificial input system has not yet been established.

[0003] Traditionally, applying organic fertilizer to newly reclaimed farmland is an effective way to improve the soil, but it also has some significant drawbacks. Newly reclaimed farmland has a very low organic matter content, requiring a very large initial application rate to achieve the desired improvement. This leads to high fertilizer costs and significant labor and material costs for transportation and application. The nutrients in organic fertilizer are slow-release, requiring microbial decomposition before release. For crops that urgently need nutrients in the current season, organic fertilizer cannot provide "quick-acting" nutrients, potentially leading to insufficient nutrition during the seedling stage. Furthermore, some commercial organic fertilizers or livestock manure contain heavy metals (such as arsenic, copper, zinc, and cadmium) in their raw materials (feed, bedding, etc.), which may remain after processing. Long-term, large-scale application can lead to the accumulation of heavy metals in the soil.

[0004] In recent years, with the rapid development of biotechnology, microalgae, as an emerging bioremediation resource, have gradually shown great potential in the field of soil improvement due to their unique ecological functions and environmental adaptability. Summary of the Invention

[0005] In view of the shortcomings of the existing technology, the purpose of this invention is to provide a method for improving the carbon and nitrogen content and aggregate content of newly reclaimed arable land soil using microalgae, aiming to solve the problems of low carbon and nitrogen content and low productivity in newly reclaimed arable land soil.

[0006] To achieve this objective, the present invention adopts the following technical solution:

[0007] In a first aspect, the present invention provides a method for increasing the carbon and nitrogen content and aggregate content of newly reclaimed arable land soil using microalgae, the method comprising: applying a mixed algal solution of Chlorella and nitrogen-fixing Anabaena to the soil.

[0008] This invention utilizes the synergistic and efficient fixation of atmospheric carbon dioxide by Chlorella and nitrogen-fixing Anabaena, converting it into organic carbon stored in the soil. Simultaneously, through biological nitrogen fixation, atmospheric nitrogen is converted into nitrogen available to plants, thereby effectively enhancing the soil's carbon and nitrogen retention capacity. Furthermore, the physiological activities of these two microalgae synergistically promote the formation of soil aggregates, improve soil structure, and enhance the soil's water and fertilizer retention capacity.

[0009] Preferably, the concentration of microalgae cells in the mixed algal solution is 1×10⁻⁶. 7 - 3×10 7 cells / mL, for example 1×10 7 cells / mL, 1.5×10 7 cells / mL, 2×10 7 cells / mL, 2.5×10 7 cells / mL, 3×10 7 The number of cells / mL, etc., can be selected from other specific point values ​​not listed in this range, which will not be elaborated here.

[0010] Preferably, the dry weight ratio of Chlorella vulgaris and nitrogen-fixing Anabaena in the mixed algal solution is (3-5):1, such as 3:1, 3.2:1, 3.4:1, 3.5:1, 3.7:1, 3.8:1, 4:1, 4.2:1, 4.4:1, 4.6:1, 4.8:1, 5:1, etc. Other specific values ​​not listed within this range can be selected, and will not be elaborated here.

[0011] This invention has found that when Chlorella and nitrogen-fixing Anabaena meet the above-mentioned specific mass ratio, they are more effective in increasing the carbon and nitrogen content and aggregate content of newly reclaimed farmland soil.

[0012] Preferably, the Chlorella is Chlorella sp. ZJ, with accession number GDMCC No: 63704, deposited at the Guangdong Provincial Center for Microbial Culture Collection on August 1, 2023.

[0013] This invention has found that Chlorella from the specific source described above is more effective than Chlorella from other sources in increasing the carbon and nitrogen content and aggregate content of newly reclaimed farmland soil.

[0014] Preferably, the mixed algal solution is a mixture of Chlorella algal solution and nitrogen-fixing Anabaena algal solution.

[0015] Preferably, the Chlorella algal solution is obtained by culturing Chlorella using a liquid culture medium containing the following components: NH4Cl, Tris-HCl, CH3COONa, K2HPO4, MgSO4·7H2O, CaCl2·2H2O, Na2CO3, ferric citrate, H3BO4, MnCl2·H2O, ZnSO4·7H2O, CuSO4·5H2O, Na2MoO4·2H2O, and Co(NO3)2·6H2O, with deionized water as the solvent.

[0016] Preferably, the nitrogen-fixing algae solution is obtained by culturing nitrogen-fixing algae in a liquid culture medium containing the following components: NaNO3, K2HPO4, MgSO4·7H2O, CaCl2·2H2O, citric acid, ferric ammonium citrate, EDTA, Na2CO3, H3BO4, MnCl2·H2O, ZnSO4·7H2O, CuSO4·5H2O, Na2MoO4·2H2O, and Co(NO3)2·6H2O, with deionized water as the solvent.

[0017] The present invention has found that when Chlorella and Anabaena are cultured using the culture medium with the above-mentioned specific formula, the resulting algal solution is more effective in increasing the carbon and nitrogen content and aggregate content of newly reclaimed farmland soil.

[0018] Preferably, the field application rate of the mixed algae solution is 100-200 kg / mu, such as 100 kg / mu, 120 kg / mu, 140 kg / mu, 150 kg / mu, 160 kg / mu, 180 kg / mu, 200 kg / mu, etc. Other specific values ​​not listed within this range can be selected, and will not be elaborated here.

[0019] More preferably, the field application rate of the mixed algae solution is 130-170 kg / mu, such as 130 kg / mu, 140 kg / mu, 145 kg / mu, 150 kg / mu, 155 kg / mu, 160 kg / mu, 170 kg / mu, etc. Other specific values ​​not listed within this range can be selected, and will not be elaborated here.

[0020] Preferably, the Chlorella and nitrogen-fixing Anabaena in the mixed algal solution are either alive or inactivated, more preferably alive.

[0021] Preferably, the mixed algae solution is applied by spraying it directly onto the soil surface using a fertilizer applicator.

[0022] Compared with the prior art, the present invention has the following beneficial effects:

[0023] To address the issues of low carbon and nitrogen content and low productivity in newly reclaimed farmland, this invention utilizes the synergistic and efficient fixation of atmospheric carbon dioxide by *Chlorella vulgaris* and nitrogen-fixing *Anabaena*, converting it into organic carbon stored in the soil. Simultaneously, through biological nitrogen fixation, atmospheric nitrogen is converted into nitrogen available to plants, effectively enhancing the soil's carbon and nitrogen retention capacity. Furthermore, the physiological activities of these two microalgae synergistically promote the formation of soil aggregates, improve soil structure, and enhance the soil's water and fertilizer retention capacity.

[0024] The Chlorella sp. ZJ involved in this invention is classified as Chlorella sp., deposited on August 1, 2023, with accession number GDMCC No: 63704, deposited at Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou. Detailed Implementation

[0025] To further illustrate the technical means and effects of the present invention, the following describes the technical solution of the present invention in conjunction with preferred embodiments of the present invention. However, the present invention is not limited to the scope of the embodiments.

[0026] The commercially available Chlorella used in the following preparation examples was purchased from the Chlorella strain product of the brand name Nanhua Qianmu; the nitrogen-fixing Anabaena strain was purchased from the nitrogen-fixing Anabaena strain product of the brand name Moon Microalgae.

[0027] Preparation Example 1-1

[0028] Chlorella (GDMCC No: 63704) was cultured using the following liquid medium: NH4Cl 0.375 g L -1 The following are the ingredients: Tris-HCl 2.42 g / L, CH3COONa 2 g / L, K2HPO4 0.04 g / L, MgSO4·7H2O 0.075 g / L, CaCl2·2H2O 0.036 g / L, Na2CO3 0.02 g / L, ferric citrate 0.006 g / L, H3BO4 0.00286 g / L, MnCl2·H2O 0.00181 g / L, ZnSO4·7H2O 0.000222 g / L, CuSO4·5H2O 0.000079 g / L, Na2MoO4·2H2O 0.00039 g / L, Co(NO3)2·6H2O 0.000049 g / L; the solvent is deionized water. The absorbance of the culture medium was measured using a spectrophotometer. The magnitude of the absorbance reflects the microalgal biomass. When the microalgal cell concentration reached 2 × 10⁻⁶, the biomass was determined. 7 When the number of algae is 1 / mL, the microalgae solution is harvested.

[0029] Preparation Examples 1-2

[0030] The preparation of the microalgae solution differed from that in Example 1-1 only in that Chlorella vulgaris (GDMCC No: 63704) was replaced with Chlorella vulgaris (commercially available), while all other operations remained unchanged.

[0031] Preparation Examples 1-3

[0032] The preparation of microalgae solution differed from that in Example 1-1 only in that Co(NO3)2·6H2O was absent in the liquid culture medium, while other conditions remained unchanged.

[0033] Preparation Example 2-1

[0034] Nitrogen-fixing algae (commercially available) were cultured using the following liquid culture medium: NaNO3 1.00 g / L, K2HPO4 0.04 g / L, MgSO4·7H2O 0.075 g / L, CaCl2·2H2O 0.036 g / L, citric acid 0.006 g / L, ferric ammonium citrate 0.006 g / L, EDTA 0.001 g / L, Na2CO3 0.02 g / L, H3BO4 0.00286 g / L, MnCl2·H2O 0.00181 g / L, ZnSO4·7H2O 0.000222 g / L, CuSO4·5H2O 0.000079 g / L, Na2MoO4·2H2O 0.00039 g / L, Co(NO3)2·6H2O 0.000049 g / L; solvent was deionized water. The absorbance of the culture medium was measured using a spectrophotometer. The absorbance value of the culture medium reflects the microalgal biomass. When the microalgal cell concentration reached 2 × 10⁻⁶ g / L... 7 When the number of algae is 1 / mL, the microalgae solution is harvested.

[0035] Preparation Example 2-2

[0036] The preparation of microalgae solution differed from that in Example 2-1 only in that Co(NO3)2·6H2O was absent in the liquid culture medium, while other conditions remained unchanged.

[0037] Example 1

[0038] (1) Soil sampling: 0-15 cm soil samples were collected from newly reclaimed paddy fields in Xinbei Village, Shangping Town, Heyuan City, Guangdong Province. After removing stones and plant roots, the samples were air-dried and passed through a 0.25 mm sieve for later use.

[0039] (2) Mixing microalgae solutions in different ways:

[0040] (2.1) The Chlorella solution obtained in Preparation Example 1-1 and the nitrogen-fixing Anabaena solution obtained in Preparation Example 2-1 were mixed to obtain sample S1;

[0041] (2.2) The Chlorella solution obtained in Preparation Example 1-2 and the nitrogen-fixing Anabaena solution obtained in Preparation Example 2-1 were mixed to obtain sample S2;

[0042] (2.3) The Chlorella solution obtained in Preparation Example 1-3 and the nitrogen-fixing Anabaena solution obtained in Preparation Example 2-1 were mixed to obtain sample S3;

[0043] (2.4) The Chlorella solution obtained in Preparation Example 1-1 and the nitrogen-fixing Anabaena solution obtained in Preparation Example 2-2 were mixed to obtain sample S4;

[0044] (2.5) The Chlorella solution obtained in Example 1-1 alone is sample S5;

[0045] In all groups except S5, the dry weight ratio of Chlorella vulgaris to nitrogen-fixing Anabaena was 4:1, and the microalgae cell concentration of the mixed microalgae solution in each group was 2 × 10⁻⁶. 7 per mL.

[0046] (3) Prepare several culture pots, take 1 kg of soil sample prepared in step (1) in each pot, and follow the treatment settings in (2) above. The amount of microalgae solution added to each group is 1.875 g / kg soil (equivalent to about 150 kg / mu in the field). Dilute the active microalgae solution of each group with water and add it to the soil. Only water is added to the blank control group. The amount of liquid added to each pot is controlled at 50 mL.

[0047] Each treatment was repeated three times, and the soil of each treatment was placed in a culture tank and cultured for 30 days. Throughout the experiment, deionized water was used to control the soil moisture at 70% of field capacity.

[0048] Destructive sampling was conducted 30 days later: the net increase in total organic carbon and total nitrogen in the soil was measured using a TOC-TN analyzer. The content of super-large aggregates (> 2 mm), large aggregates (1~2 mm), and small aggregates (0.25~1 mm) was determined using the wet sieving method. The aggregate content was used to characterize the stability of soil organic matter. The results are shown in Table 1 (different letters after the data in the table represent significant differences between groups).

[0049] Table 1

[0050]

[0051] As shown in Table 1, the mixed microalgal solution of Chlorella and nitrogen-fixing Anabaena in this invention can synergistically increase the carbon and nitrogen content and soil aggregate content in the soil, effectively enhancing the soil's carbon and nitrogen retention capacity and improving soil structure. Furthermore, using the specific culture medium formulations described above to cultivate Chlorella and Anabaena separately, the resulting algal solutions showed superior effects in increasing the carbon and nitrogen content and aggregate content of newly reclaimed farmland. Meanwhile, the source of Chlorella also influences these effects to some extent.

[0052] Example 2

[0053] (1) Soil sampling: 0-15 cm soil samples were collected from newly reclaimed paddy fields in Xinbei Village, Shangping Town, Heyuan City, Guangdong Province. After removing stones and plant roots, the samples were air-dried and passed through a 0.25 mm sieve for later use.

[0054] (2) Obtaining microalgae solution in different ways:

[0055] (2.1) The Chlorella solution obtained in Preparation Example 1-1 and the nitrogen-fixing Anabaena solution obtained in Preparation Example 2-1 were mixed to obtain sample S1;

[0056] (2.2) The Chlorella solution obtained in Preparation Example 1-1 and the nitrogen-fixing Anabaena solution obtained in Preparation Example 2-1 were mixed and inactivated (specifically, the algal solution was placed in an autoclave and sterilized at 120 °C for 1 hour) to obtain sample S2;

[0057] In all the above groups, the dry weight ratio of Chlorella vulgaris to nitrogen-fixing Anabaena was 3:1, and the microalgae cell concentration of the mixed microalgae solution in each group was 2×10⁻⁶. 7 per mL.

[0058] (3) Prepare several culture pots, take 1 kg of soil sample prepared in step (1) in each pot, and follow the treatment settings in (2) above. The amount of microalgae solution added to each group is 1.875 g / kg soil (equivalent to about 150 kg / mu in the field). Dilute the active microalgae solution of each group with water and add it to the soil. Only water is added to the blank control group. The amount of liquid added to each pot is controlled at 50 mL.

[0059] Each treatment was repeated three times, and the soil of each treatment was placed in a culture tank and cultured for 30 days. Throughout the experiment, deionized water was used to control the soil moisture at 70% of field capacity.

[0060] Destructive sampling was conducted 30 days later: the net increase in total organic carbon and total nitrogen in the soil was measured using a TOC-TN analyzer. The content of super-large aggregates (> 2 mm), large aggregates (1~2 mm), and small aggregates (0.25~1 mm) was determined using the wet sieving method. The aggregate content was used to characterize the stability of soil organic matter. The results are shown in Table 2 (different letters after the data in the table represent significant differences between groups).

[0061] Table 2

[0062]

[0063] As shown in Table 2, compared with the inactivated microalgae solution treatment, the activated microalgae solution has a more significant effect on improving the content of total organic carbon, total nitrogen and aggregate components.

[0064] Example 3

[0065] (1) Soil sampling: 0-15 cm soil samples were collected from newly reclaimed farmland in Yongji Village, Tianyuan Town, Heyuan City, Guangdong Province. After removing stones and plant roots, the samples were air-dried and passed through a 0.25 mm sieve for later use.

[0066] (2) Obtain the mixed microalgae solution in the following manner:

[0067] The Chlorella solution obtained in Preparation Example 1-1 and the nitrogen-fixing Anabaena solution obtained in Preparation Example 2-1 were mixed, such that the dry weight ratio of Chlorella to Anabaena was 5:1, and the microalgae cell concentration of the mixed microalgae solution was 2 × 10⁻⁶. 7 Samples were obtained by measuring per mL.

[0068] (3) Prepare several culture pots, and take 1 kg of the soil sample prepared in step (1) into each pot. Treat the soil according to different addition amounts:

[0069] (S1) The amount of microalgae solution added is 1.875 g / kg soil (equivalent to approximately 150 kg / mu in the field). The active microalgae solution is diluted with water and then added to the soil. The amount of liquid added to each pot is controlled at 50 mL.

[0070] (S2) The amount of microalgae solution added is 1.25 g / kg soil (equivalent to approximately 100 kg / mu in the field). The active microalgae solution is diluted with water and then added to the soil. The amount of liquid added to each pot is controlled at 50 mL.

[0071] (S3) The amount of microalgae solution added is 2.5 g / kg soil (equivalent to approximately 200 kg / mu in the field). The active microalgae solution is diluted with water and then added to the soil. The amount of liquid added to each pot is controlled at 50 mL.

[0072] The blank control group was only added with water. Each treatment was repeated 3 times. The soil of each treatment was placed in a culture tank and cultured for 30 days. Throughout the experiment, deionized water was used to control the soil moisture at 70% of field capacity.

[0073] Destructive sampling was conducted 30 days later: the net increase in total organic carbon and total nitrogen in the soil was measured using a TOC-TN analyzer. The content of super-large aggregates (> 2 mm), large aggregates (1~2 mm), and small aggregates (0.25~1 mm) was determined using the wet sieving method. The aggregate content was used to characterize the stability of soil organic matter. The results are shown in Table 3 (different letters after the data in the table represent significant differences between groups).

[0074] Table 3

[0075]

[0076] The results in Table 3 show that the active mixed microalgae solution, applied at three different rates, significantly increased the net increase in total organic carbon and total nitrogen in the soil, and promoted the formation of various components of aggregates. Comparing the results among the three methods, S1 and S3 were found to be better than S2, and S1 was close to S3. Therefore, considering both technical effectiveness and cost factors, an application rate of approximately 150 kg / mu is the optimal application rate in the field.

[0077] The applicant declares that the technical solution of this invention is illustrated by the above embodiments, but this invention is not limited to the above embodiments, that is, it does not mean that this invention must rely on the above embodiments to be implemented. Those skilled in the art should understand that any improvements to this invention, equivalent substitutions of raw materials for the products of this invention, addition of auxiliary components, selection of specific methods, etc., all fall within the protection scope and disclosure scope of this invention.

[0078] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solution of the present invention, and these simple modifications all fall within the protection scope of the present invention.

[0079] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not describe the various possible combinations separately.

Claims

1. A method for increasing the carbon and nitrogen content and aggregate content of newly reclaimed farmland soil using microalgae, characterized in that, The method includes applying a mixed algal solution of Chlorella and nitrogen-fixing Anabaena to the soil.

2. The method according to claim 1, characterized in that, The microalgal cell concentration of the mixed algal solution is 1×10⁻⁶. 7 -3×10 7 per mL.

3. The method according to claim 1 or 2, characterized in that, The dry weight ratio of Chlorella vulgaris and nitrogen-fixing Anabaena in the mixed algal solution is (3-5):

1.

4. The method according to any one of claims 1-3, characterized in that, The Chlorella species in question is Chlorella sp. ZJ, with accession number GDMCC No: 63704, deposited at the Guangdong Provincial Center for Microbial Culture Collection on August 1, 2023.

5. The method according to any one of claims 1-4, characterized in that, The mixed algal solution is a mixture of Chlorella algal solution and nitrogen-fixing Anabaena algal solution.

6. The method according to claim 5, characterized in that, The Chlorella algal solution was obtained by culturing Chlorella in a liquid culture medium containing the following components: NH4Cl, Tris-HCl, CH3COONa, K2HPO4, MgSO4·7H2O, CaCl2·2H2O, Na2CO3, ferric citrate, H3BO4, MnCl2·H2O, ZnSO4·7H2O, CuSO4·5H2O, Na2MoO4·2H2O, and Co(NO3)2·6H2O, with deionized water as the solvent.

7. The method according to claim 5, characterized in that, The nitrogen-fixing anemone algae solution was obtained by culturing the nitrogen-fixing anemone algae in a liquid culture medium containing the following components: NaNO3, K2HPO4, MgSO4·7H2O, CaCl2·2H2O, citric acid, ferric ammonium citrate, EDTA, Na2CO3, H3BO4, MnCl2·H2O, ZnSO4·7H2O, CuSO4·5H2O, Na2MoO4·2H2O, and Co(NO3)2·6H2O, with deionized water as the solvent.

8. The method according to any one of claims 1-7, characterized in that, The field application rate of the mixed algae solution is 100-200 kg / mu, preferably 130-170 kg / mu.

9. The method according to any one of claims 1-8, characterized in that, The Chlorella and nitrogen-fixing Anabaena in the mixed algal solution are either alive or inactivated, preferably alive.

10. The method according to any one of claims 1-9, characterized in that, The mixed algae solution is applied by spraying it directly onto the soil surface using a fertilizer applicator.

Images