Method for screening metabolites beneficial to soil structure improvement on sunny slopes of high and cold mountainous regions based on metabonomics

Abstract

The invention discloses a method for screening metabolites beneficial to soil structure improvement on a sunny slope of an alpine mountain based on metabonomics, and relates to the technical field of soil structure improvement. Eight sunny slope sample plots with small human activity interference and consistent sample plot vegetation communities are selected to collect undisturbed soil shaken off from plant roots, fresh soil and air-dried soil are subpackaged, and metabolites beneficial to soil structure improvement are screened through metabonomics; especially, it is found that the phenol (PHE) metabolite and the formation and stability of the soil large aggregate are in an obvious positive correlation relationship, a new theoretical basis and practical guidance are provided for soil structure improvement, through correlation analysis, the selective enrichment or rejection relationship of the soil metabolite in the aggregates with different particle sizes is revealed, and the soil metabolite in the soil large aggregate has a good application prospect. Particularly, the positive influence of the phenol metabolites on the stability of the soil aggregate provides a new perspective for understanding the ecological function and nutrient circulation of the soil.

Description

Technical Field

[0001] This invention relates to the field of soil structure improvement technology, specifically a method for screening metabolites beneficial to soil structure improvement on sunny slopes of high-altitude and cold mountains based on metabolomics. Background Technology

[0002] Improving soil structure is fundamental to enhancing soil quality. Soil structure, the spatial arrangement of aggregates of different particle sizes, is the basic unit and crucial component of soil structure. It is also a fundamental condition for maintaining soil physical structure and fertility, directly influencing soil porosity, water holding capacity, erosion resistance, soil fertility, and the cycling of microorganisms and organic matter. Aggregates, as an important indicator of soil quality, are key factors in resisting soil fragmentation by external forces. Their formation and stability affect processes such as soil infiltration, crusting, nutrient cycling, and the separation and movement of soil on slopes. The compositional characteristics and stability of soil aggregates play a crucial role in regulating the coordination of multiple factors such as water, fertilizer, air, and heat in the soil, thus determining the soil's sustainable utilization capacity and being of great significance for maintaining soil ecological functions, nutrient conservation, and stable storage.

[0003] Soil metabolites mainly originate from soil microbial metabolic activities, decomposition products of plant and animal remains, substances exuded by plant roots, and the transformation of soil organic matter. Studying the sources and functions of metabolites is crucial for understanding soil fertility, plant health, and ecological processes. Multiple studies have shown that root exudates are the primary source of soil metabolites. These exudates mediate multi-faceted interactions within the rhizosphere, initiating and regulating the dialogue between roots and soil microorganisms. They serve as important mediators for the exchange of matter and energy and the transmission of information within the dynamic system of the plant-soil-microbe interface, playing a vital role in regulating plant adaptation to the microenvironment, mitigating rhizosphere nutrient competition, shaping the structure of the rhizosphere microbial community, and influencing soil element biogeochemical cycling processes.

[0004] Root exudates can enhance the formation and stability of soil aggregates through soil microorganisms and their own processes. On the one hand, soil microorganisms can fully utilize root exudates as a primary carbon and energy source, thereby enhancing microbial activity and indirectly promoting aggregate formation. On the other hand, as plants grow, increased root exudates lead to increased organic matter and microbial content in the soil, causing soil micro-aggregates to transform into larger aggregates through the cementing effect of organic matter and root exudates, as well as the entanglement of roots and hyphae. Furthermore, plant root exudates can effectively increase the proportion of water-stable aggregates in the soil and influence soil nutrient release and fertility by affecting enzyme activity, thus creating a favorable ecological microenvironment for roots.

[0005] Therefore, in-depth research on the relationship between soil metabolites and soil aggregates, and analysis of material exchange and energy flow at the plant-soil interface, is crucial for the formation and stability of aggregates, as well as for soil structure improvement and functional enhancement. In addition, the wet sieving method is required in the research. After sieving soil aggregates, the sieve tray needs to be moved. When moving the sieve tray, the sample on the sieve tray is easy to fall through the sieve holes. Summary of the Invention

[0006] To address the shortcomings of existing technologies, this invention provides a method for screening metabolites beneficial to soil structure improvement on sunny slopes of high-altitude and cold mountains based on metabolomics, thus solving the problems mentioned in the background art.

[0007] To achieve the above objectives, the present invention provides the following technical solution: a method for screening metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains based on metabolomics, comprising the following steps: Step 1: Soil sample collection: Along the vertical zonation of the mountains on the eastern edge of the Qinghai-Tibet Plateau, select 8 sunny slope plots (sunny slope: 90-270°) with less human disturbance and consistent vegetation communities (dominant species similarity >80%). Collect undisturbed soil from the plant roots and separate it into fresh soil and air-dried soil. Step 2: Soil aggregate preparation and determination: Remove plant residues, straw and large stones from the soil sample, gently break it into small pieces along its natural structure, and let it air dry for later use; S1: Dry sieving method: The content of aggregates of various particle sizes in air-dried soil samples was determined by the dry sieving method. After the soil samples were air-dried, a certain amount of air-dried soil sample was weighed and sieved through a sieve with apertures of 5 mm, 2 mm, 1 mm, 0.5 mm, and 0.25 mm. The sieve was vibrated vertically with an amplitude of 3 cm for 5 minutes. The dry weight of aggregates of various particle sizes was weighed, and the proportion of each aggregate size was calculated. S2: Wet sieving method: Weigh a certain amount of air-dried soil sample, quickly moisten it in pure water, soak for 10 minutes, and then wet sieve it. The sieve apertures are 5mm, 2mm, 1mm, 0.5mm, and 0.25mm respectively. Vibrate the sample up and down 10 times at an amplitude of 2cm. Rinse the agglomerates remaining on each sieve into an aluminum box, dry at 105℃, weigh, and calculate the mass percentage of agglomerates of each aperture size. Soil macroaggregate density (R) 0.25 Measurement: This is a commonly used indicator reflecting the size distribution of soil aggregates. The larger the value, the higher the degree of aggregation and the stronger the stability of the aggregates.

[0008] In the formula: R 0.25 - Proportion of large aggregates (%); M r - Weight of aggregates of various particle sizes (g); M T - Total weight of aggregates (g); Step 3: Determination of soil metabolite composition: S31: Soil sample pretreatment. Take 50g of freshly collected soil and place it in a volumetric flask. Add 100mL of distilled water and place in a shaker at 20℃ for 24h at 100r / min. Then place the volumetric flask in a high-speed centrifuge at 21℃ and rotate at 3500r / min for 15min. Extract the supernatant twice with ethyl acetate, filter and combine the filtrates. Centrifuge at 3500r / min for 15min, collect the supernatant, remove water with anhydrous sodium sulfate, filter through a 0.45μm organic filter, and place in a 1.5ml sample vial for GC-MS detection. S32: GC-MS determination was performed using a gas chromatography-triple quadrupole mass spectrometer. 5 μL of sample was injected splitlessly at an injection port temperature of 260 ℃ and a flow rate of 1.0 mL / min. -1 The temperature program is as follows: 50 ℃ (hold for 2 min) → 20 ℃·min -1 Increase to 150 ℃ → at 5 ℃·min -1 Increase to 220 ℃ → at 6 ℃·min -1 The temperature was increased to 250 °C and held for 15 min. Mass spectrometry conditions: electron impact (EI) ionization, ionization energy 70 eV, ion source temperature 200 °C, interface temperature 280 °C, scan range 33–600 m / z. S33: Metabolite composition analysis. The total ion chromatogram was obtained based on the GC-MS test results. The content of substances was represented by the peak intensity. Metabolites were quantitatively analyzed by peak area. The identified metabolites were annotated using the NIST database. The relative content of various substances was calculated using the area normalization method. The analysis showed that PHE (phenolic compounds) can promote the formation and stability of soil macroaggregates, which is helpful for soil structure improvement and function enhancement.

[0009] Preferably, in S2: wet screening method: the equipment used includes a vibrator and a collection frame. A water cylinder is placed below the vibrator, and a fixed frame is placed inside the water cylinder. The vibrating rod of the vibrator is fixedly connected to the fixed frame by bolts. Five screen plates are placed in the middle of the fixed frame. Screen frames are fixedly connected to the outer side of each screen plate. Insert frames are fixedly connected to the top of each screen frame and above the screen plate. Rewinding boxes are fixedly connected to both sides of each screen frame.

[0010] Preferably, each of the winding boxes has a winding roller rotatably connected to its inner cavity. A baffle is wound around the outside of one of the winding rollers. One end of the baffle passes through the screen frame and extends into the interior of another winding box and is fixedly connected to the outside of the winding roller. A through groove is provided at the top of the baffle and below the screen plate. A socket is fixedly connected to the bottom of the screen frame.

[0011] Preferably, a connecting box is fixedly connected to the front of the screen frame, and two worm gears are rotatably connected inside the connecting box. One end of each winding roller extends into the connecting box and is fixedly fitted with a worm wheel that is connected to the worm gears in a transmission manner. The two worm gears are fixedly connected to each other. A connecting rod is rotatably connected to one side of the inner cavity of the connecting box. A first bevel gear that meshes with the outer side of the connecting rod and the outer side of one of the worm gears is fixedly fitted. A gear is fixedly fitted on the outer side of the connecting rod.

[0012] Preferably, a threaded rod is rotatably connected inside the connecting box and below the worm gear. A second rack is threadedly connected to the outer side of the threaded rod. One end of the second rack is inserted into a groove in the inner wall of the connecting box and slidably connected to the groove. The first bevel gear cooperates with the second rack.

[0013] Preferably, one end of the connecting rod extends to the outside of the connecting box and is fixedly connected to a gear two. A first drive box is fixedly connected to one side of the winding box. A fixed seat is fixedly connected to one side of the first drive box. A threaded block is fixedly connected to the front end of each fixed seat. An mounting seat is fixedly connected to the inside of the fixing frame and below the fixed seat and the threaded block. A screw is inserted into the inside of each mounting seat. One end of each screw passes through the mounting seat and the threaded block and is inserted into a limiting hole opened inside the fixed seat. A moving groove is opened inside the fixed seat and behind the limiting hole. A spring one is fixedly connected to one side of the inner cavity of the moving groove.

[0014] Preferably, one end of the spring is fixedly connected to a push post, one end of the push post extends into the limiting hole and is fixedly connected to a slot, one side of the push post is fixedly connected to a first rack, and one side of the inner cavity of the first drive box is rotatably connected to a rotating shaft.

[0015] Preferably, one end of each rotating shaft extends into the moving groove and is fitted with a first driving gear that engages with the first rack via a one-way bearing.

[0016] Preferably, both ends of the threaded rod extend into the interior of the two first drive boxes, and both ends of the threaded rod and the outer side of the rotating shaft are fixedly fitted with sprockets connected by chain drive.

[0017] This invention provides a method for screening metabolomics-based metabolites that are beneficial to soil structure improvement on sunny slopes of high-altitude and cold mountains, which has the following advantages: 1. This study utilizes metabolomics to screen metabolites beneficial to soil structure improvement on sunny slopes of high-altitude, cold mountains. Through metabolomics screening, metabolites beneficial to soil structure improvement were identified, particularly phenolic (PHE) metabolites, which showed a significant positive correlation with the formation and stability of large soil aggregates. This provides new theoretical basis and practical guidance for soil structure improvement. Correlation analysis revealed the selective enrichment or repulsion relationships of soil metabolites in aggregates of different particle sizes, especially the positive impact of phenolic metabolites on the stability of soil aggregates, providing a new perspective for understanding soil ecological functions and nutrient cycling.

[0018] 2. This method, based on metabolomics screening, is used on the sunny slopes of high-altitude, cold mountains to screen metabolites beneficial to soil structure improvement. In screening mode, a baffle belt aligns the sluice gate with the sieve plate, ensuring unobstructed screening channels. In transfer and backwashing mode, pulling out the screw triggers a mechanical linkage: first rack-threaded rod-second rack-connecting rod-worm / worm wheel, driving the baffle belt to move horizontally, using its non-perforated portion to seal the bottom of the sieve plate. This prevents screened material or blockages from accidentally falling off during the movement of the screening mechanism.

[0019] 3. This method, based on metabolomics screening, for metabolites beneficial to soil structure improvement on sunny slopes in high-altitude, cold mountains utilizes a backwashing mode. As the squeezing bottle and nozzle assembly move to a specific position, the baffle belt retracts accordingly, causing the trough to align with the nozzle. This design avoids the risk of sample loss due to water flow directly carrying the sample back to the sieve plate during rinsing caused by trough misalignment. It also ensures that the high-pressure water flow is concentrated on the exposed sieve openings, dislodging any blocked samples. Attached Figure Description

[0020] Figure 1 This is a schematic diagram of the overall structure of the device of the present invention; Figure 2 This is a schematic diagram of the fixing frame structure of the present invention; Figure 3 This is a cross-sectional view of the connecting box of the present invention; Figure 4 This is a schematic diagram of the mounting base structure of the present invention; Figure 5 This is a cross-sectional internal structural diagram of the winding box, connecting box, and first drive box of the present invention; Figure 6 This is a cross-sectional view of the internal structure of the fixing base of the present invention; Figure 7 For the present invention Figure 6 Enlarged view of point A; Figure 8 This is a schematic diagram of the backwashing device of the present invention; Figure 9 This is a partial structural diagram of the present invention; Figure 10 This is a schematic diagram of the internal structure of the second drive box of the present invention; Figure 11 For the present invention Figure 9 A schematic diagram of the structure viewed from below; Figure 12 This is a schematic diagram of the rear view of the second drive box of the present invention; Figure 13 This is a schematic diagram of the positioning point structure of the present invention; Figure 14 This is a schematic diagram of the structure when the backwashing operation of the present invention reaches the first flushing checkpoint; Figure 15 This is a cross-sectional view of the squeezing bottle of the present invention; Figure 16 This is a top view of the backwashing device of the present invention. Figure 17 For the present invention Figure 9 Enlarged view of point B; Figure 18 For the present invention Figure 5 Enlarged view of point C; Figure 19 This is a top-view cross-sectional schematic diagram of the internal structure of the fixing base of the present invention; Figure 20 This is a schematic diagram of the baffle structure of the device of the present invention; Figure 21 This is a schematic diagram of the correlation analysis of the present invention.

[0021] In the diagram: 1. Vibrator; 2. Water tank; 3. Collection rack; 4. Fixing frame; 5. Mounting frame; 6. Mounting base; 7. Threaded block; 8. Screen frame; 9. Insert frame; 10. Screen plate; 11. Rewinding box; 12. First drive box; 13. Fixing base; 14. Screw; 15. Rewinding roller; 16. Baffle belt; 17. Through groove; 18. Connecting box; 19. Moving groove; 20. Slot; 21. Push column one; 22. Spring one; 23. First rack; 24. Rotating shaft one; 25. First drive gear; 26. Threaded rod one; 27. Sprocket one; 28. Second rack; 29. ​​Worm gear; 30. Worm; 31. Connecting rod; 32. First bevel gear; 33. Gear one; 34. Gear II; 35. Fixing component; 36. Drive frame; 37. First magnet block; 38. First guide block; 39. Sound-generating plate; 40. Elastic card; 41. Fourth rack; 42. Threaded shaft; 43. Second sprocket; 44. Connecting rod; 45. Second bevel gear; 46. Second drive box; 47. Second magnet block; 48. Guide groove; 49. Push column II; 50. Second guide block; 51. Spring II; 52. Drive rail; 53. Motor; 54. Threaded rod II; 55. Limiting block; 56. Guide cavity; 57. Fifth rack; 58. Rotating shaft II; 59. Gear III; 60. Bottom groove; 61. Squeezing bottle; 62. Threaded end seat; 63. Nozzle; 64. Socket. Detailed Implementation

[0022] The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings of the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments.

[0023] Example 1 Please see Figure 21 This invention provides a technical solution: a method for screening metabolites beneficial to soil structure improvement on sunny slopes of high-altitude and cold mountains based on metabolomics, comprising the following steps: Step 1: Soil sample collection: Along the vertical zonation of the mountains on the eastern edge of the Qinghai-Tibet Plateau, select 8 sunny slope plots (sunny slope: 90-270°) with less human disturbance and consistent vegetation communities (dominant species similarity >80%). Collect undisturbed soil from the plant roots and separate it into fresh soil and air-dried soil. Step 2: Soil aggregate preparation and determination: Remove plant residues, straw and large stones from the soil sample, gently break it into small pieces along its natural structure, and let it air dry for later use; S1: Dry sieving method: The content of aggregates of various particle sizes in air-dried soil samples was determined using the dry sieving method. After air-drying, a certain amount of the air-dried soil sample was weighed and sieved through a sieve with apertures of 5 mm, 2 mm, 1 mm, 0.5 mm, and 0.25 mm. The sieve was vibrated vertically with an amplitude of 3 cm for 5 minutes. The dry weight of aggregates of various particle sizes was then measured, and the proportion of each aggregate size was calculated. S2: Wet sieving method: Weigh a certain amount of air-dried soil sample, quickly moisten it in pure water, soak it for 10 minutes, and then wet-sieve it. The sieve apertures are 5mm, 2mm, 1mm, 0.5mm, and 0.25mm respectively. Vibrate the sample up and down 10 times at an amplitude of 2cm. Rinse the agglomerates remaining on each sieve into an aluminum box, dry it at 105℃, weigh it, and calculate the mass percentage of agglomerates of each aperture size. (3) Specific gravity of soil macroaggregates (R) 0.25 ) Measurement It is a commonly used indicator reflecting the size distribution of soil aggregates. The larger the value, the higher the degree of aggregation and the stronger the stability of the aggregates.

[0024] In the formula: R 0.25 - Proportion of large aggregates (%); M r - Weight of aggregates of various particle sizes (g); M T - Total weight of aggregates (g); Step 3: Determination of soil metabolite composition: S31: Soil sample pretreatment. Take 50g of freshly collected soil and place it in a volumetric flask. Add 100mL of distilled water and place in a shaker at 20℃ for 24h at 100r / min. Then place the volumetric flask in a high-speed centrifuge at 21℃ and rotate at 3500r / min for 15min. Extract the supernatant twice with ethyl acetate, filter and combine the filtrates. Centrifuge at 3500r / min for 15min, collect the supernatant, remove water with anhydrous sodium sulfate, filter through a 0.45μm organic filter, and place in a 1.5ml sample vial for GC-MS detection. S32: GC-MS determination was performed using a gas chromatography-triple quadrupole mass spectrometer. 5 μL of sample was injected splitlessly at an injection port temperature of 260 ℃ and a flow rate of 1.0 mL / min. -1 The temperature program is as follows: 50 ℃ (hold for 2 min) → 20 ℃·min -1 Increase to 150 ℃ → at 5 ℃·min -1 Increase to 220 ℃ → at 6 ℃·min -1 The temperature was increased to 250 °C and held for 15 min. Mass spectrometry conditions: electron impact (EI) ionization, ionization energy 70 eV, ion source temperature 200 °C, interface temperature 280 °C, scan range 33–600 m / z. S33: Metabolite composition analysis. A total ion chromatogram (TIC) was obtained based on GC-MS results. The horizontal axis represents elution time, and the vertical axis represents elution intensity. Different peaks and elution times are used to distinguish substances, and the content of a substance is indicated by its elution intensity. Metabolites were quantitatively analyzed using peak area. The identified metabolites were annotated using the NIST database, and the relative content of each substance was calculated using area normalization.

[0025] In R4.3.3 software, Spearman's analysis was used to statistically assess the correlation between soil metabolites and soil aggregate stability on sunny slopes of high-altitude and cold mountains.

[0026] S4: Correlation analysis. In the figure, the vertical axis represents the proportion of agglomerates of different particle sizes obtained by wet sieving and dry sieving. On the vertical axis, W0.25mm_2mm and W0.25mm_ represent large agglomerates measured by wet sieving, and D0.25mm_2mm and D0.25mm_ represent large agglomerates measured by dry sieving. All four types of agglomerates belong to R0.25mm_. 0.25 Category of class aggregates.

[0027] The horizontal axis represents the percentage of each major soil metabolite category as determined by GC-MS, and the ratio of the number of specific species within each category to the total number of categories. The major soil metabolites on the horizontal axis are: ALK (alkanes), ALC (alcohols), EST (esters), OA (organic acids), ETH (ethers), ALD (aldehydes), KET (ketones), PHE (phenols), QUI (quinones), ALKEN (olefins), and OTH (others). ALK2, ALC2, EST2, OA2, ETH2, ALD2, KET2, PHE2, QUI2, ALKEN2, and OTH2 represent the percentage of specific species within each of these major metabolite categories to the total number of categories.

[0028] Correlation heatmaps were used to analyze the strength and significance of associations between soil aggregates of different particle sizes (large aggregates, micro aggregates, silt and clay components, etc.) and various soil metabolites (such as amino acids, organic acids, sugars, phenolic compounds, etc.). The heatmaps visually represent the range of correlation coefficients using a color gradient (blue to green), and statistical significance is marked with an asterisk (*) to indicate a significant relationship.

[0029] The analysis results show that the distribution of soil metabolites in aggregates of different particle sizes is not uniform, but exhibits significant selective enrichment or repulsion relationships, among which R 0.25 A significant positive correlation was found with PHE metabolites, suggesting that PHE (phenolic compounds) can promote the formation and stability of large soil aggregates, thus contributing to soil structure improvement and functional enhancement.

[0030] Table 1 Metabolites from the sunny slope

[0031] Table 2 Aggregates on sunny slopes

[0032] Example 2 Please see Figures 1 to 20 This invention provides a technical solution: a method for screening metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains based on metabolomics, including equipment. The equipment includes a vibrator 1 and a collection frame 3. A water cylinder 2 is placed below the vibrator 1, and a fixed frame 4 is placed inside the water cylinder 2. The vibrating rod of the vibrator 1 is fixedly connected to the fixed frame 4 by bolts. Five sieve plates 10 are placed in the middle of the fixed frame 4. A sieve frame 8 is fixedly connected to the outer side of each sieve plate 10. An insert frame 9 is fixedly connected to the top of the sieve frame 8 and above the sieve plate 10. A winding box 11 is fixedly connected to both sides of the sieve frame 8. A winding roller 15 is rotatably connected to the inner cavity of each winding box 11. A baffle 16 is wound around the outer side of one winding roller 15. One end of the baffle 16 passes through the sieve frame 8 and extends into the interior of another winding box 11 and is fixedly connected to the outer side of the winding roller 15. A through groove 17 is opened at the top of the baffle 16 and below the sieve plate 10. A socket 64 is fixedly connected to the bottom of the sieve frame 8.

[0033] The screen frame 8 is fixedly connected to a connecting box 18. Two worm gears 30 are rotatably connected inside the connecting box 18. One end of each winding roller 15 extends into the connecting box 18 and is fixedly fitted with a worm wheel 29 that is connected to the worm gear 30. The two worm gears 30 are fixedly connected. A connecting rod 31 is rotatably connected to one side of the inner cavity of the connecting box 18. The outer side of the connecting rod 31 and the outer side of one of the worm gears 30 are fixedly fitted with a first bevel gear 32 that meshes with each other. A gear 33 is fixedly fitted on the outer side of the connecting rod 31. When the washing machine is working, the first rack 23 is located at the front end of the first drive gear 25. The two do not contact each other to avoid interference.

[0034] Inside the connecting box 18 and below the worm gear 30, there is a threaded rod 26 rotatably connected. The outer side of the threaded rod 26 is threadedly connected to a second rack 28. One end of the second rack 28 is inserted into a groove in the inner wall of the connecting box 18 and is slidably connected to the groove. The first bevel gear 32 cooperates with the second rack 28, so that the second rack 28 can slide along the inner wall of the connecting box 18.

[0035] One end of the connecting rod 31 extends to the outside of the connecting box 18 and is fixedly connected to the gear 34. The first drive box 12 is fixedly connected to one side of the winding box 11. The fixed seat 13 is fixedly connected to one side of the first drive box 12. The front end of the fixed seat 13 is fixedly connected to the threaded block 7. The mounting seat 6 is fixedly connected to the inside of the fixing frame 4 and below the fixed seat 13 and the threaded block 7. The mounting seat 6 is inserted with screws 14. One end of the screws 14 passes through the mounting seat 6 and the threaded block 7 and is inserted into the limiting hole opened inside the fixed seat 13. The fixed seat 13 is provided with a moving groove 19 behind the limiting hole. The moving groove 19 is fixedly connected to one side of the inner cavity of the moving groove 19. The fixed seat 13 is provided with a sealing sleeve between the limiting hole and the moving groove 19. The sealing sleeve is located outside the push column 21 and keeps the moving groove 19 closed.

[0036] One end of the spring 22 is fixedly connected to the push post 21. One end of the push post 21 extends into the limiting hole and is fixedly connected to the slot 20. One side of the push post 21 is fixedly connected to the first rack 23. One side of the inner cavity of the first drive box 12 is rotatably connected to the rotating shaft 24. The connecting rod 31 can be driven to rotate through the rotating shaft 24.

[0037] One end of the rotating shaft 24 extends into the moving groove 19 and is fitted with a first drive gear 25 that cooperates with the first rack 23 via a one-way bearing. Both ends of the threaded rod 26 extend into the two first drive boxes 12 respectively. Both ends of the threaded rod 26 and the outer side of the rotating shaft 24 are fixedly fitted with sprockets 27 connected by chain drive. Only when the screw 14 is pulled out can the first rack 23 drive the threaded rod 26 to rotate.

[0038] The top of the collection frame 3 is fixedly connected to a fixing member 35. The top of the collection frame 3 is equipped with a screening mechanism consisting of two fixing seats 13, a first drive box 12, a winding box 11, a insertion frame 9, a sieve plate 10, and a connecting box 18. The top of the fixing member 35 contacts the bottom of the threaded block 7 and the fixing seat 13. One end of the screw 14 passes through the fixing member 35 and is inserted into the threaded block 7. The top of the collection frame 3 and the front end of the connecting box 18 are fixedly connected to a drive frame 36. The drive frame 36 is rotatably connected to a threaded shaft 42. The outer side of the threaded shaft 42 is threadedly connected to a fourth rack 41. One end of the fourth rack 41 extends to the outer side of the drive frame 36. The fourth rack 41 is located below the gear 24. The connecting box 18 and the connecting rod 44 are rotatably connected to the threaded shaft 42. The outer side of the connecting rod 44 and the threaded shaft 42 are both fixedly fitted with a sprocket 2 43 that is connected by a chain drive. The length of the fourth rack 41 matches the length of the second rack 28.

[0039] The collection rack 3 has a first guide block 38 fixedly connected to both sides, and a nozzle 63 fixedly connected between the two first guide blocks 38. A threaded end seat 62 is fixedly connected to the bottom of the nozzle 63, and a squeezing bottle 61 is threadedly connected inside the threaded end seat 62. Several spray pipes are fixedly connected to the top of the nozzle 63. The bottom of the collection rack 3 and both sides of the sieve plate 10 are provided with bottom grooves 60. The nozzle 63 is located below one bottom groove 60, so that the nozzle 63 can be stored inside the bottom groove 60.

[0040] Six elastic cards 40 are fixedly connected to both sides of the collection rack 3. One end of each elastic card 40 is inserted into the first guide block 38. A sound-emitting plate 39 is fixedly connected to one side of the inner cavity of the first guide block 38. One end of each elastic card 40 is engaged with the surface of the sound-emitting plate 39. When the elastic card 40 strikes the sound-emitting plate 39, a clicking sound is emitted.

[0041] Among them, a second drive box 46 is fixedly connected to one side of the drive frame 36 and the side of the elastic card 40. A second guide block 50 is opened inside the second drive box 46. A spring 51 is fixedly connected to the top of the inner cavity of the second guide block 50. A push post 49 is fixedly connected to one end of the spring 51. The bottom end of the push post 49 extends into the guide groove 48 opened at the bottom of the second drive box 46 and is fixedly connected to a second magnet block 47. A fifth rack 57 is fixedly connected to one side of the push post 49. The fifth rack 57 is located inside the second drive box 46 and inside the guide cavity 56 opened to the right of the second magnet block 47. A rotating shaft 58 is rotatably connected inside the guide cavity 56. A gear 59 that cooperates with the fifth rack 57 is fixedly sleeved on the outside of the rotating shaft 58. One end of the rotating shaft 58 extends into the connecting box 18. A second bevel gear 45 that meshes with the outside of the rotating shaft 58 and the connecting rod 44 is fixedly sleeved on the outside of the connecting rod 44. A drive rail 52 is fixedly connected to the left side of the drive box 46. A threaded rod 54 is rotatably connected inside the drive rail 52. A motor 53 is fixedly connected to the top of the drive rail 52. The output end of the motor 53 is fixedly connected to the top of the threaded rod 54. A limit block 55 is threadedly connected to the outside of the threaded rod 54. One end of the limit block 55 extends into the second guide block 50. A mounting frame 5 is fixedly connected to one end of the first guide block 38. A first magnet 37 is embedded in the top of the mounting frame 5. The first magnet 37 is located below the second magnet 47. The first magnet 37 and the second magnet 47 are magnetically repelled. The output end of the motor 53 can drive the threaded rod 54 to rotate, so that the threaded rod 54 drives the limit block 55 to move. By adjusting the position of the limit block 55, the movable height of each fifth rack 57 can be adjusted, and thus the distance that each second drive box 46 drives the fourth rack 41 to move can be adjusted.

[0042] In summary, this method for screening metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains based on metabolomics involves placing a set of screening mechanisms on the top of the mounting base 6 on the fixed frame 4. Then, one end of the screw 14 is passed through the mounting base 6 and the threaded block 7 and inserted into the limiting hole, so that one end of the screw 14 pushes the slot 20 and the push column 21 to compress the spring 22. At this time, the push column 21 drives the first rack 23 to move without driving the first drive gear 25 to drive the rotating shaft 24 to rotate through the one-way bearing, so that the through groove 17 is located below the sieve plate 10. Then, five screening mechanisms are stacked in sequence, so that the socket 64 of the uppermost screening mechanism is inserted into the outside of the insertion frame 9 of the lowermost screening mechanism. Then, the fixed frame 4 is immersed in the water cylinder 2. Next, the soil sample is placed above the sieve plate 10 in the uppermost screening mechanism. Then, the fixed frame 4 is installed on the telescopic end of the vibrator 1 with bolts. The vibrator 1 drives the fixed frame 4 and the screening mechanism to vibrate for wet screening. After screening is complete, remove the connecting bolts between the fixed frame 4 and the telescopic end of the vibrator 1, remove the water cylinder 2 from under the vibrator 1, then pull the fixed frame 4 out of the water cylinder 2 and drain the water from the screening mechanism. First, tighten screw 14 so that it moves out of the limiting hole, threaded block 7, and mounting base 6. At this time, spring 22 pushes push column 21 and slot 20 to reset, causing push column 21 to drive the first rack 23 to reset. This allows the first rack 23 to drive the rotating shaft 24 via the first drive gear 25 and one-way bearing, which in turn drives the two sprockets 27 and threaded rod 26 to rotate. The second rack 28 is driven to move, causing the second rack 28 to drive the gear 33 to rotate the connecting rod 31. The connecting rod 31 drives the two worm gears 30 to rotate through the two first bevel gears 32. The worm gears 30 drive the take-up roller 15 to rotate, so that the left take-up roller 15 releases the baffle 16, and the right take-up roller 15 takes up the part of the baffle 16 with the through groove 17. When one screw 14 is completely pulled out, the second rack 28 moves to the midpoint of the stroke of the threaded rod 26. Then the other screw 14 is unscrewed, so that the second rack 28 moves to the end point of the stroke of the threaded rod 26, so that the baffle 16 closes the bottom of the screen plate 10. At this point, remove the screening mechanism from the fixed frame 4, clean the screened material on the surface of the screen plate 10 into the aluminum box using a tool, and then place the screening mechanism on top of the collection frame 3. Pass one end of the screw 14 through the fixing piece 35 and screw it into the threaded block 7 to fix the position of the screening mechanism. At this time, gear 2 34 is located on the left side of the drive frame 36. Then, the operator pulls the threaded end seat 62, causing the threaded end seat 62 to drive the nozzle 63 and the first guide block 38 to move along both sides of the collection frame 3, causing the elastic card 40 to deform and the first guide block 38 to detach from the foremost elastic card 40. Then, the first guide block 38 continues to move. When it moves to the next elastic card 40, the elastic card 40, under its own elastic force, will... The front end of the elastic card 40 is inserted into the first guide block 38, and the front end of the elastic card 40 strikes the sound-emitting plate 39, causing the sound-emitting plate 39 to emit a crisp clicking sound, thereby reminding the staff to reach the designated position. At this time, the first magnet block 37 is located below a second drive box 46. At this time, a repulsive force is generated between the first magnet block 37 and the second magnet block 47, causing the second magnet block 47 to push the push column 2 49 to overcome the supporting force of the spring 2 51 and move upward. This causes the push column 2 49 to drive the fifth rack 57 to move until the top of the push column 2 49 contacts the bottom of the limit block 55. At this time, the fifth rack 57 drives the gear 3 59 to drive the rotating shaft 2 58 to rotate through the one-way bearing. This causes the rotating shaft 2 58 to drive the second bevel gear 45 to drive the connecting rod 44 to rotate, thus... The connecting rod 44 drives the threaded shaft 42 to rotate via two sprockets 43, which in turn drives the fourth rack 41 to move. The fourth rack 41 then drives the gear 34 to rotate the connecting rod 31. The connecting rod 31, through two first bevel gears 32, drives two worm gears 30 to rotate. Each worm gear 30, via a worm wheel 29, drives the take-up roller 15 to rotate. The left take-up roller 15 winds up the baffle belt 16, while the right take-up roller 15 releases the baffle belt 16, causing the through groove 17 to gradually move below the screen plate 10. The stroke of one fifth rack 57 can drive one section of the through groove 17 on the baffle belt 16 to move below the screen plate 10. Then, the user squeezes the water bottle 61, squeezing water through the threaded end seat 62 into the nozzle 63. The screen plate 10 is backwashed by the spray pipe, and then the first guide block 38 and the nozzle 63 are pulled to move. When the first magnet block 37 moves to one side of the second drive box 46, the magnetic repulsion between the second magnet block 47 and the first magnet block 37 disappears. At this time, the second spring 51 pushes the second push column 49 and the second magnet block 47 to move down, so that the second push column 49 drives the fifth rack 57 to move down inside the guide cavity 56. At this time, the fifth rack 57 cannot drive the rotating shaft 58 to rotate through the third gear 59 and the one-way bearing. When the first magnet block 37 moves to the next second drive box 46, it continues to work according to the above principle, flushing out the blockage in the screen hole of the screen plate 10, and flushing the blockage out of the outside of the insertion frame 9 by the water flow and falling into the aluminum box placed at the lower end of the insertion frame 9. Until the first magnet block 37 moves out from under the last second drive box 46, the fourth rack 41 moves to its maximum stroke, causing the through groove 17 to move completely under the screen plate 10. Then, one end of the screw 14 is moved out from inside the threaded block 7 and the fixing member 35, removing the screening mechanism from the collection frame 3. Then, the first guide block 38 is pulled to reset and move, causing the first guide block 38 to drive the first magnet block 37 to move sequentially under several second drive boxes 46. At this time, a repulsive force is generated between the first magnet block 37 and the second magnet block 47, causing the second magnet block 47 to push... Push column 2 49 moves upward against the supporting force of spring 2 51, causing push column 2 49 to drive the fifth rack 57 to move until the top of push column 2 49 contacts the bottom of limit block 55. At this time, the fifth rack 57 drives gear 3 59 to drive rotating shaft 2 58 to rotate through one-way bearing. Rotating shaft 2 58 drives second bevel gear 45 to drive connecting rod 44 to rotate. Connecting rod 44 drives threaded shaft 42 to rotate through two sprockets 2 43. Threaded shaft 42 drives fourth rack 41 to move, causing fourth rack 41 to reset and move until fourth rack 41 moves to the initial position.

[0043] The above description is only a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the scope of the technology disclosed in the present invention, based on the technical solution and inventive concept of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A method for screening metabolomics-based metabolomics-based metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains, characterized in that: Includes the following steps: Step 1: Soil sample collection: Along the vertical zonation of the mountains on the eastern edge of the plateau, select 8 sunny slope plots with minimal human disturbance and consistent vegetation communities to collect undisturbed soil from the roots of plants, and separate the fresh soil into fresh soil and air-dried soil. Step 2: Soil aggregate preparation and determination: Remove plant residues, straw and large stones from the soil sample, gently break it into small pieces along its natural structure, and air dry for later use; S1: Dry sieving method: The content of aggregates of various particle sizes in air-dried soil samples was determined by the dry sieving method. After the soil samples were air-dried, a certain amount of air-dried soil sample was weighed and sieved through a sieve with apertures of 5 mm, 2 mm, 1 mm, 0.5 mm, and 0.25 mm. The sieve was vibrated vertically with an amplitude of 3 cm for 5 minutes. The dry weight of aggregates of various particle sizes was weighed, and the proportion of each aggregate size was calculated. S2: Wet sieving method: Weigh a certain amount of air-dried soil sample, quickly moisten it in pure water, soak for 10 minutes, and then wet sieve it. The sieve apertures are 5mm, 2mm, 1mm, 0.5mm, and 0.25mm respectively. Vibrate the sample up and down 10 times at an amplitude of 2cm. Rinse the agglomerates remaining on each sieve into an aluminum box, dry at 105℃, weigh, and calculate the mass percentage of agglomerates of each aperture size. Soil macroaggregate density (R) 0.25 Measurement: This is a commonly used indicator reflecting the size distribution of soil aggregates. The larger the value, the higher the degree of aggregation and the stronger the stability of the aggregates. ； In the formula: R 0.25 - Proportion of large aggregates (%); M r - Weight of aggregates of various particle sizes (g); M T - Total weight of aggregates (g); Step 3: Determination of soil metabolite composition: S31: Soil sample pretreatment. Take 50g of freshly collected soil and place it in a volumetric flask. Add 100mL of distilled water and place in a shaker at 20℃ for 24h at 100r / min. Then place the volumetric flask in a high-speed centrifuge at 21℃ and rotate at 3500r / min for 15min. Extract the supernatant twice with ethyl acetate, filter and combine the filtrates. Centrifuge at 3500r / min for 15min, collect the supernatant, remove water with anhydrous sodium sulfate, filter through a 0.45μm organic filter, and place in a 1.5ml sample vial for GC-MS detection. S32: GC-MS determination was performed using a gas chromatography-triple quadrupole mass spectrometer. 5 μL of sample was injected splitlessly at an injection port temperature of 260 ℃ and a flow rate of 1.0 mL / min. -1 The temperature program is as follows: 50 ℃ for 2 min → 20 ℃·min -1 Increase to 150 ℃ → at 5 ℃·min -1 Increase to 220 ℃ → at 6 ℃·min -1 The temperature was raised to 250 °C and held for 15 min. Mass spectrometry conditions: electron impact (EI) ionization, ionization energy 70 eV, ion source temperature 200 °C, interface temperature 280 °C, scan range 33–600 m / z. S33: Metabolite composition analysis. The total ion chromatogram is obtained based on the GC-MS test results. The content of substances is represented by the peak intensity. Metabolites are quantitatively analyzed by peak area. The identified metabolites are annotated using the NIST database. The relative content of various substances is calculated using the area normalization method. The analysis shows that PHE (phenolic compounds) are conducive to the formation and stability of soil macroaggregates, which helps to improve soil structure and enhance its function.

2. The method for screening metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains based on metabolomics, as described in claim 1, is characterized in that: S2: Wet screening method: The equipment used includes a vibrator (1) and a collection frame (3). A water cylinder (2) is placed below the vibrator (1). A fixed frame (4) is placed inside the water cylinder (2). The vibrating rod of the vibrator (1) is connected to the fixed frame (4) by bolts. Five screen plates (10) are placed in the middle of the fixed frame (4). Screen frames (8) are installed on the outer side of each screen plate (10). A insertion frame (9) is fixedly connected to the top of the screen frame (8) and above the screen plate (10). A winding box (11) is fixedly connected to both sides of the screen frame (8).

3. The method for screening metabolomics-based metabolomics-based metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains according to claim 2, characterized in that: The inner cavity of each winding box (11) is rotatably connected to a winding roller (15). A baffle (16) is wound around the outside of one of the winding rollers (15). One end of the baffle (16) passes through the screen frame (8) and extends into the interior of another winding box (11) and connects to the outside of the winding roller (15). A through groove (17) is provided on the top of the baffle (16) and below the screen plate (10). A socket (64) is fixedly connected to the bottom of the screen frame (8).

4. The method for screening metabolomics-based metabolomics-based metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains according to claim 3, characterized in that: A connecting box (18) is fixedly connected to the front of the screen frame (8). Two worm gears (30) are rotatably connected inside the connecting box (18). One end of the take-up roller (15) extends into the connecting box (18) and is fixedly fitted with a worm wheel (29) that is connected to the worm gear (30) for transmission. The two worm gears (30) are fixedly connected. A connecting rod (31) is rotatably connected to one side of the inner cavity of the connecting box (18). A first bevel gear (32) that meshes with the outer side of the connecting rod (31) and the outer side of one worm gear (30) is fixedly fitted. A gear one (33) is fixedly fitted on the outer side of the connecting rod (31).

5. The method for screening metabolomics-based metabolomics-based metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains according to claim 4, characterized in that: Inside the connecting box (18) and below the worm gear (30), a threaded rod (26) is rotatably connected. The outer side of the threaded rod (26) is threadedly connected to a second rack (28). One end of the second rack (28) is inserted into the groove on the inner wall of the connecting box (18) and slidably connected to the groove. The first bevel gear (32) cooperates with the second rack (28).

6. The method for screening metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains based on metabolomics, as described in claim 5, is characterized in that: One end of the connecting rod (31) extends to the outside of the connecting box (18) and is fixedly connected to the gear two (34). The first drive box (12) is fixedly connected to one side of the winding box (11). The first drive box (12) is fixedly connected to one side of the first drive box (12). The front end of the fixed seat (13) is fixedly connected to the threaded block (7). The mounting seat (6) is fixedly connected to the inside of the fixing frame (4) and below the fixed seat (13) and the threaded block (7). The mounting seat (6) is inserted with screws (14). One end of the screws (14) passes through the mounting seat (6) and the threaded block (7) and is inserted into the limiting hole opened inside the fixed seat (13). The fixed seat (13) is provided with a moving groove (19) inside and behind the limiting hole. The moving groove (19) is fixedly connected to one side of the inner cavity of the moving groove (19).

7. The method for screening metabolomics-based metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains according to claim 6, characterized in that: One end of spring 1 (22) is connected to push post 1 (21), one end of push post 1 (21) extends into the limiting hole and is fixedly connected to slot (20), one side of push post 1 (21) is fixedly connected to first rack (23), and one side of the inner cavity of the first drive box (12) is rotatably connected to rotating shaft 1 (24).

8. The method for screening metabolomics-based metabolomics-based metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains according to claim 7, characterized in that: One end of each of the rotating shafts (24) extends into the moving groove (19) and is fitted with a first drive gear (25) that cooperates with the first rack (23) via a one-way bearing.

9. The method for screening metabolomics-based metabolites beneficial to soil structure improvement on sunny slopes of high-altitude cold mountains according to claim 8, characterized in that: Both ends of the threaded rod (26) extend into the interior of the two first drive boxes (12), and both ends of the threaded rod (26) and the outer side of the rotating shaft (24) are fixedly fitted with sprockets (27) connected by chain drive.

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