Application of sesquiterpene lactone in preparation of soil conditioner

Abstract

The invention discloses application of sesquiterpene lactone in preparation of a soil conditioner. The application comprises a sesquiterpene lactone preparation method, a method for detecting sesquiterpene lactone in soil and a method for detecting physical and chemical properties of the soil. Sesquiterpene lactone is applied to preparation of the soil conditioner, is a natural compound with wide biological activity, serves as a plant defense substance and participates in interaction between plants and the environment, and can influence interaction between the plants and soil microorganisms, so that the soil conditioner can be applied to the soil conditioner. The fertilizer can promote the symbiotic relationship between plant roots and effective microbial communities, help plants to absorb nutrient substances in soil, especially inorganic phosphorus, and influence the generation and nitrogen circulation process of available nitrogen in different forms, so that the enrichment effect is achieved, the soil fertility is increased, the total organic matter content of the soil can be increased, and the pH value condition of the soil is improved.

Description

Technical Field

[0001] The invention relates to the technical field of application of sesquiterpene lactones, and in particular to application of sesquiterpene lactones in the preparation of soil conditioners. Background Art

[0002] Affected by natural and human factors, the ecological environment is deteriorating, and the restoration and reconstruction of the ecosystem is the main problem at present. The purpose of soil improvement can be achieved by improving the soil structure through physical, chemical and biological methods.

[0003] Traditional biological improvement methods such as planting grass to fertilize the field are traditional agricultural technologies that improve soil fertility and soil structure by planting specific herbaceous plants. This method can not only increase the organic matter content in the soil, but also improve the physical properties of the soil through the action of the plant root system, and increase the water retention and air permeability of the soil. Traditional soil improvement methods, such as adding sand to clay, adding loam to sand, etc., the added substances can be called natural soil conditioners. Now, organic extracts, natural minerals or artificial high molecular polymers are mostly used to synthesize soil conditioners. These soil conditioners use soil microorganisms as a medium to affect the nitrogen cycle of the soil environment, thereby increasing soil fertility and providing conditions for the rapid growth of plants.

[0004] The above traditional improvement methods cannot affect the interaction between plants and soil microorganisms, cannot promote the symbiotic relationship between plant roots and beneficial bacteria, cannot help plants absorb nutrients in the soil, and cannot affect the generation of different forms of effective nitrogen and the nitrogen cycle process, and cannot improve the soil environment well. Summary of the invention

[0005] The object of the present invention is to provide an application of sesquiterpene lactone in the preparation of a soil conditioner to solve the problems raised in the above background technology.

[0006] In order to achieve the above-mentioned purpose, the present invention adopts the following technical scheme: including a method for preparing sesquiterpene lactones, a method for detecting sesquiterpene lactones in soil and a method for detecting soil physical and chemical properties. The method for preparing sesquiterpene lactones includes: directly extracting sesquiterpene lactones from the above-ground tissue parts of Mikania micrantha or directly burying the fallen leaves of the above-ground parts of Mikania micrantha in the soil.

[0007] Preferably, the method for detecting sesquiterpene lactones in soil comprises the following steps:

[0008] (1) Soil nutrient extraction;

[0009] (2) UPLC-MS quantitative testing;

[0010] (3) Experimental data analysis.

[0011] Preferably, the soil nutrient extraction specifically comprises the following steps:

[0012] A. Select fresh soil samples and air-dry them naturally, then filter them through a 30-mesh sterile sieve to remove the fallen leaves of Mikania micrantha in the soil;

[0013] B. Take 15g soil powder, add 30ml methanol solvent, and use ultrasonic extraction for 15min, a total of two extractions;

[0014] C. Combine the extracts and centrifuge at 500 rpm for 4 min.

[0015] D. Take the supernatant and concentrate it under reduced pressure, then use 1.5 ml of chromatographic methanol to reconstitute it, and then filter it through a 0.22 μm PETE filter membrane for use.

[0016] Preferably, the UPLC-MS quantitative test specifically comprises the following steps:

[0017] A. The samples were separated and analyzed by ThermoScientific quadrupole-orbitrap high-resolution liquid chromatography-mass spectrometry. The injector temperature was set at 4°C and random injection was performed.

[0018] B. Detection conditions: chromatographic column temperature 25°C, flow rate 0.25 ml / min, injection volume 1 μl, mobile phase A 0.1% formic acid, mobile phase B acetonitrile;

[0019] C. Gradient elution program: total program duration 30 min, 0-2.5 min, 95% B, 2.5-25 min, B phase linearly changes from 95% to 5%; 25-30 min, B phase linearly changes from 5% to 95%;

[0020] D. Set up QC samples during the experiment to monitor the stability of the instrument and experimental data, and use them to calculate the relative content of sesquiterpene lactones;

[0021] E. Heated electrospray ionization (HESI) negative ion mode kit: In negative ion mode, the spray voltage is 2800 V, the capillary temperature is 340°C, the maximum allowable spray current of the ion source is 100 mA, the probe heater temperature is 340°C, and the mass-to-charge ratio scanning range is 80-1000 Da.

[0022] Preferably, the experimental data analysis specifically includes the following steps:

[0023] A. Browse, check and preliminarily analyze the original data from the machine through Xcalibur, including the total ion current (TIC) in full scan mode, selected ion monitoring (SIM) data and secondary mass spectrometry (MS / MS) spectra;

[0024] B. Use the software CompoundDiscover3.3 (Thermo) to identify metabolites and search common online metabolomics databases such as mzCloud, MassList, ChemSpider, and Thermo Fisher's own databases (such as mzVault TM Spectral library or mass list), peak alignment, retention time correction and peak area extraction were performed to clarify the types and relative contents of metabolites. The retention times of five sesquiterpene lactones, 3-epi, Deo, Anh, Scan and Dih, were 9.30min, 9.30min, 9.20min, 9.20min and 8.44min, respectively. The above-mentioned sesquiterpene lactones were detected in each soil sample by comparison with the standard.

[0025] Preferably, the soil physical and chemical property detection method comprises the following steps:

[0026] ①Detection of organic matter;

[0027] ② Determination of soil pH;

[0028] ③ Detection of total nitrogen;

[0029] ④ Detection of total phosphorus;

[0030] ⑤ Detection of total potassium;

[0031] ⑥ Detection of hydrolyzable nitrogen;

[0032] ⑦ Detection of available phosphorus;

[0033] ⑧Detection of fast-acting potassium.

[0034] Compared with the prior art, the present invention has the following beneficial effects:

[0035] The present invention applies sesquiterpene lactones in the preparation of soil conditioners. Sesquiterpene lactones are a class of natural compounds with broad biological activity. As a plant defense substance and participating in the interaction between plants and the environment, sesquiterpene lactones can affect the interaction between plants and soil microorganisms, promote the symbiotic relationship between plant roots and beneficial bacteria, help plants absorb nutrients in the soil, especially inorganic phosphorus, and affect the generation of effective nitrogen in different forms and the nitrogen cycle process, thereby achieving an enrichment effect, increasing soil fertility, increasing the total organic matter content in the soil, and improving soil pH conditions. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 It is a soil nutrient comparison bar chart of the present invention. DETAILED DESCRIPTION

[0037] In order to further explain the technical solution of the present invention, it is described in detail below through specific embodiments.

[0038] The invention provides an application of sesquiterpene lactone in preparing a soil conditioner, including a sesquiterpene lactone preparation method, a sesquiterpene lactone detection method in soil and a soil physical and chemical property detection method. The sesquiterpene lactone preparation method comprises: directly extracting sesquiterpene lactone from the above-ground tissue of Mikania micrantha or directly burying the fallen leaves of the above-ground part of Mikania micrantha in the soil, wherein the sesquiterpene lactone is Sca, Anh, Deo, Dih and 3epi; the soil improvement effect is to increase the content of soil nutrients and adjust the pH value of the soil; the soil nutrients mainly include total organic matter, nitrogen, phosphorus, potassium and other nutrients; the soil nutrients such as nitrogen, phosphorus and potassium include total phosphorus, total nitrogen, total potassium, hydrolyzable nitrogen, available phosphorus and quick-acting potassium.

[0039] The method for detecting sesquiterpene lactones in soil comprises the following steps:

[0040] (1) Soil nutrient extraction, specifically comprising the following steps:

[0041] A. Select fresh soil samples and air-dry them naturally, then filter them through a 30-mesh sterile sieve to remove the fallen leaves of Mikania micrantha in the soil;

[0042] B. Take 15g soil powder, add 30ml methanol solvent, and use ultrasonic extraction for 15min, a total of two extractions;

[0043] C. Combine the extracts and centrifuge at 500 rpm for 4 min.

[0044] D. Take the supernatant and concentrate it under reduced pressure, then use 1.5 ml of chromatographic methanol to re-dissolve it, and then filter it through a 0.22 μm PETE filter membrane for later use;

[0045] (2) UPLC-MS quantitative test, specifically including the following steps:

[0046] A. The samples were separated and analyzed by ThermoScientific quadrupole-orbitrap high-resolution liquid chromatography-mass spectrometry. The injector temperature was set at 4°C and random injection was performed.

[0047] B. Detection conditions: chromatographic column temperature 25°C, flow rate 0.25 ml / min, injection volume 1 μl, mobile phase A 0.1% formic acid, mobile phase B acetonitrile;

[0048] C. Gradient elution program: total program duration 30 min, 0-2.5 min, 95% B, 2.5-25 min, B phase linearly changes from 95% to 5%; 25-30 min, B phase linearly changes from 5% to 95%;

[0049] D. Set up QC samples during the experiment to monitor the stability of the instrument and experimental data, and use them to calculate the relative content of sesquiterpene lactones;

[0050] E. Heated electrospray ionization (HESI) negative ion mode kit: In negative ion mode, the spray voltage is 2800 V, the capillary temperature is 340 °C, the maximum allowable spray current of the ion source is 100 mA, the probe heater temperature is 340 °C, and the mass-to-charge ratio scanning range is 80-1000 Da;

[0051] (3) Experimental data analysis, specifically including the following steps:

[0052] A. Browse, check and preliminarily analyze the original data from the machine through Xcalibur, including the total ion current (TIC) in full scan mode, selected ion monitoring (SIM) data and secondary mass spectrometry (MS / MS) spectra;

[0053] B. Use the software CompoundDiscover3.3 (Thermo) to identify metabolites and search common online metabolomics databases such as mzCloud, MassList, ChemSpider, and Thermo Fisher's own databases (such as mzVault TM Spectral library or mass list), peak alignment, retention time correction and peak area extraction were performed to clarify the types and relative contents of metabolites. The retention times of five sesquiterpene lactones, 3-epi, Deo, Anh, Scan and Dih, were 9.30min, 9.30min, 9.20min, 9.20min and 8.44min, respectively. The above-mentioned sesquiterpene lactones were detected in each soil sample by comparison with the standard.

[0054] The soil physical and chemical property detection method comprises the following steps:

[0055] ① Organic matter detection: Adopting the NY / T 1121.6-2006 standard "Soil Testing Part 6: Determination of Soil Organic Matter", this method usually involves mixing the soil sample with an oxidant to oxidize the organic carbon, and then determining the remaining inorganic carbon by titration to calculate the organic matter content;

[0056] ② Determination of soil pH: According to the NY / T 1121.2-2006 standard "Soil Testing Part 2: Determination of Soil pH", soil pH is usually determined by mixing the soil sample with water and then using a pH meter to measure the pH of the mixture;

[0057] ③Total nitrogen detection: Use the NY / T1121.24-2012 standard "Soil Testing Part 24: Determination of Total Nitrogen in Soil - Automatic Nitrogen Determination Method". This method involves using a Kjeldahl nitrogen analyzer to convert nitrogen in the soil sample into ammonia, and then determine the ammonia content by acid titration to calculate the total nitrogen content;

[0058] ④ Total phosphorus detection: According to the NY / T 88-1988 standard "Determination of total phosphorus in soil", this method usually involves mixing the soil sample with acid to dissolve the phosphorus in the soil, and then using a UV-visible spectrophotometer to measure the phosphorus concentration in the solution;

[0059] ⑤Total potassium detection: Using the NY / T 87-1988 standard "Determination of total potassium in soil", the determination of total potassium usually involves mixing the soil sample with acid to dissolve the potassium in the soil, and then using an atomic absorption spectrophotometer to measure the potassium concentration in the solution;

[0060] ⑥ Detection of hydrolyzable nitrogen: According to the standard LY / T 1228-2015 "Determination of forest soil nitrogen", this method involves mixing the soil sample with acid to release hydrolyzable nitrogen, and then titrating it with an acid burette to determine the content of hydrolyzable nitrogen;

[0061] ⑦ Detection of available phosphorus: According to the NY / T 1121.7-2014 standard "Soil Testing Part 7: Determination of Available Phosphorus in Soil", the determination of available phosphorus usually involves mixing the soil sample with a specific extractant and then using a UV-visible spectrophotometer to measure the phosphorus concentration in the extract;

[0062] ⑧ Detection of available potassium: Adopt the standard of "Determination of available potassium and slow-release potassium content in soil" NY / T 889-2004. Determination of available potassium usually involves mixing soil samples with specific extractants and then measuring the potassium concentration in the extract using a flame photometer.

[0063] Soil physical and chemical properties testing methods, specifically:

[0064] ①Detection of organic matter:

[0065] Measurement steps: accurately weigh 0.05g~0.5g (accurate to 0.0001g, the sample weight depends on the organic matter content range) of the air-dried sample that has passed through a 0.25mm aperture sieve, and put it into a hard test tube; accurately add 10.00mL0.4mol / L potassium dichromate-sulfuric acid solution from the automatic zeroing burette and shake well; insert a glass funnel into the mouth of each test tube, insert the test tubes into the wire cage one by one, and then sink the wire cage into the oil bath pot that has been heated to 185℃~190℃ on the electric furnace, so that the liquid level in the tube is lower than the oil level; control temperature: the oil bath temperature is required to drop to 170℃~180℃ after being placed in the test tube, and start timing when the solution in the test tube boils; at this moment, the temperature of the electric furnace must be controlled to prevent the solution from boiling violently. Shake evenly: during this period, the wire cage can be gently lifted and shaken in the oil bath pot several times to make the liquid temperature uniform and maintain it at 17 0℃~180℃; after 5min±0.5min, take out the wire cage from the oil bath, cool it for a while, and wipe off the oil (wax) liquid outside the test tube; transfer the digestion liquid and soil residue in the test tube into a 250mL conical flask without damage, rinse the test tube and small funnel with water, and add the washing liquid into the conical flask to control the total volume of the solution in the conical flask to 50mL~60mL; add indicator: add 3 drops of o-phenanthroline indicator; titrate the remaining K2Cr2O7 with ferrous sulfate standard solution, the color change process of the solution is orange-yellow→blue-green→brown-red; if the milliliters of ferrous sulfate solution used for titration are less than 1 / 3 of the milliliters of ferrous sulfate solution consumed in the blank test, the amount of soil sample should be reduced and re-measured; for each batch of analysis, 2 blank tests must be done at the same time, that is, take about 0.2g of calcined pumice powder or soil instead of soil sample, and the other steps are the same as soil sample determination.

[0066] Result calculation

[0067]

[0068] Where:

[0069] OM – mass fraction of soil organic matter, in grams per kilogram (g / kg);

[0070] V0——The volume of ferrous sulfate standard solution consumed in the blank test, in milliliters (mL);

[0071] V——The volume of ferrous sulfate standard solution consumed in the sample determination, in milliliters (mL);

[0072] c——Concentration of ferrous sulfate standard solution, in moles per liter (mol / L);

[0073] 0.003——The millimolar mass of 1 / 4 carbon atom, in grams (g);

[0074] 1.724 – coefficient for converting organic carbon to organic matter;

[0075] 1.10 – Oxidation correction factor;

[0076] m——Weigh the mass of the dried sample, in grams (g);

[0077] 1000——Converted into content per kilogram.

[0078] ② Determination of soil pH

[0079] Instrument calibration: adjust the instrument temperature compensator to the same temperature value as the test solution and the standard buffer solution; insert the electrode into the pH 4.01 standard buffer solution, adjust the instrument so that the pH value of the standard solution is consistent with the instrument indication; remove the electrode, rinse with water, dry with filter paper, and insert into the pH 6.87 standard buffer solution; check the instrument readings, and the absolute difference between the two standard solutions is allowed to be 0.1 pH unit; repeat several times until the instrument is stable; if it exceeds the specified allowable difference, check whether there is a problem with the instrument electrode or the standard solution; when the instrument is calibrated correctly, it can be used for sample determination;

[0080] Determination of soil water pH: weigh 10g (accurate to 0.01g) of air-dried sample that has passed through a 2mm aperture sieve into a 50mL tall beaker, add 25mL of water to remove CO2 (soil-liquid ratio is 1:2.5), stir with a stirrer for 1min to fully disperse the soil particles, and measure after leaving for 30min; insert the electrode into the sample suspension (note that the lower part of the glass electrode bulb is located at the soil-liquid interface, and the calomel electrode is inserted into the upper clear liquid), gently rotate the beaker to remove the water film of the electrode to promote rapid equilibrium, let it stand for a while, press the reading switch, and record the pH value when the reading is stable; release the reading switch, take out the electrode, wash it with water, and use filter paper to absorb the water before measuring the second sample; after measuring 5 to 6 samples, the standard solution should be used to check the positioning;

[0081] Expression of analysis results: When measuring pH with an acidity meter, the pH can be read directly without calculation.

[0082] ③ Detection of total nitrogen

[0083] Weigh about 1g of air-dried soil sample that has passed through a 0.25mm sieve (accurate to 0.0001g, containing about 1mg of nitrogen); place the sample at the bottom of a dry digestion tube, add 1mL of potassium permanganate solution, and shake the digestion tube; slowly add 2mL of sulfuric acid solution, keep rotating the digestion tube, and then leave it for 5 minutes; add 1 drop of octanol; use a long-necked funnel to place 0.5g (±0.01g) of reduced iron powder at the bottom of the digestion tube, add a reflux device at the mouth of the tube or place a small curved-necked glass funnel; rotate the digestion tube to allow the iron powder to contact the acid, and when the violent reaction stops (about 5 minutes), remove the digestion tube. Place the tube on a digestion furnace and heat slowly for 45 minutes; stop heating, wait for the digestion tube to cool, add 2g accelerator and 5mL sulfuric acid through a long-necked funnel, and shake well; digest until the soil liquid turns yellow-green, and continue to digest for 1 hour; after digestion is completed, cool and wait for distillation; refer to the instrument manual and use sulfuric acid or hydrochloric acid standard titration solution; set to add 10mL~30mL of water, 25mL of sodium hydroxide solution and 20mL~30mL of boric acid absorption solution; place the digestion tube on an automatic nitrogen analyzer for distillation and titration; use a blank solution, and the other steps are the same as the determination of the sample solution.

[0084] The total nitrogen (N) content in the soil sample is expressed as mass fraction ω and the value is expressed as percentage (%) and is calculated according to the following formula:

[0085] Where:

[0086]

[0087] C H ——Concentration of acid standard titration solution, in moles per liter (mol / L);

[0088] V——The volume of ferrous sulfate standard solution consumed in the sample determination, in milliliters (mL);

[0089] V0——The volume of ferrous sulfate standard solution consumed in the blank test, in milliliters (mL);

[0090] m——Weigh the mass of the dried sample, in grams (g);

[0091] f——Oxidation correction factor.

[0092] This formula is used to calculate the total nitrogen content in the soil sample, taking into account factors such as volume change during titration, concentration of the standard solution, and mass of the sample. Through this formula, the measured nitrogen content can be converted to a percentage based on the oven-dried soil standard.

[0093] ④Total phosphorus detection

[0094] Soil sample preparation: Take the air-dried soil sample that has passed through a 1 mm pore size sieve and spread it on kraft paper into a thin layer, and divide it into many small squares; use a small spoon to extract an equal amount of soil sample in each square (total amount is not less than 20 g) and further grind it in an agate mortar so that it can all pass through a 0.149 mm pore size sieve; mix well and put it into a ground-mouth bottle for later use;

[0095] Melting sample: accurately weigh 0.25g of air-dried sample, accurate to 0.0001g, and carefully put it into the bottom of nickel (or silver) crucible, and do not stick to the wall; add 3-4 drops of anhydrous ethanol to wet the sample, and spread 2g of sodium hydroxide on the sample; put the crucible into a high-temperature electric furnace and heat it up; when the temperature rises to about 400℃, cut off the power supply and pause for 15min; then continue to heat it to 720℃ and keep it for 15min, take it out and cool it; add 10mL of water at about 80℃, and after the molten block is dissolved, transfer the solution into a 100mL volumetric flask without loss, and wash the crucible with 10mL of 3mol / L sulfuric acid solution and water several times, and the washing liquid is also transferred to the volumetric flask; cool and adjust the volume; filter with phosphorus-free qualitative filter paper or centrifuge to clarify; and do a blank test at the same time;

[0096] Draw a calibration curve: pipette 0, 2, 4, 6, 8, 10 mL of 5 mg / L phosphorus standard solution into a 50 mL volumetric flask, add a blank solution and 2-3 drops of dinitrophenol indicator of the same volume as the sample solution used for colorimetric determination; adjust the solution to just turn slightly yellow with 10% sodium carbonate solution or 5% sulfuric acid solution; accurately add 5 mL of molybdenum antimony anti-color developer, shake well, add water to make up to volume, and obtain a series of standard solutions with phosphorus contents of 0.0, 0.2, 0.4, and 0.8 mg / L, respectively; shake well, place at a temperature above 15°C for 30 minutes, and then measure its absorbance at a wavelength of 700 nm; draw a calibration curve on a grid paper with absorbance as the ordinate and phosphorus concentration (mg / L) as the abscissa;

[0097] Quantification of phosphorus in sample solution: Pipette 2-10mL of the sample solution to be tested (containing 0.04-1.0μg phosphorus) into a 50mL volumetric flask, and dilute with water to about 3 / 5 of the total volume; add 2-3 drops of dinitrophenol indicator, and adjust the solution with 10% sodium carbonate solution or 5% sulfuric acid solution until it is just slightly yellow; accurately add 5mL of molybdenum antimony anti-color developer, shake well, and add water to make up to volume; leave it at room temperature above 15°C for 30 minutes; the colored sample solution is placed on a spectrophotometer with a 700nm, 1cm optical path cuvette, and the instrument is zeroed with a blank test as the reference solution, and a colorimetric determination is performed to read the absorbance; the corresponding phosphorus content is obtained from the calibration curve.

[0098] Analysis results: The percentage of total phosphorus in the soil (calculated on the basis of oven-dried soil) is given by the following formula:

[0099]

[0100] Where:

[0101] c——The phosphorus content in the sample solution to be tested is obtained from the calibration curve, in milligrams per liter (mg / L);

[0102] m——sample weight, in grams (g);

[0103] V1——the constant volume of the sample after melting, in milliliters (mL);

[0104] V2——The volume of the solution when developing color, in milliliters (mL);

[0105] V3——the volume taken from the melt after constant volume, in milliliters (mL);

[0106] 10 -4 - Conversion factor for converting mg / L concentration unit into percentage content;

[0107] H is the percentage of moisture content in air-dried soil.

[0108] ⑤Total potassium detection:

[0109] Sample digestion: weigh 0.1g of air-dried soil that has passed a 0.149mm pore size sieve, accurate to 0.0001g, and place it in a platinum crucible or a polytetrafluoroethylene crucible; add 3mL of nitric acid and 0.5mL of perchloric acid, place it in an electric sand bath or a hot plate covered with asbestos cloth, and heat it in a fume hood until the nitric acid is completely driven out and part of the perchloric acid decomposes to produce a large amount of white smoke. When the sample becomes a paste, remove it and cool it; use a plastic pipette to add 5mL of hydrofluoric acid and 0.5mL of perchloric acid, place it in a sand bath at 200-225℃ and heat it to decompose minerals such as silicates, and continue to heat it until the remaining hydrofluoric acid and perchloric acid are completely driven out; when the white smoke stops, remove it and cool it; add 10mL of 3mol / L hydrochloric acid solution and continue to heat it until the residue is dissolved; remove it and cool it, add 2mL of 2% boric acid solution; quantitatively transfer it into a 100mL volumetric flask with deionized water, make up to volume, and mix it well; this is the soil digestion solution.

[0110] Note: If the residue is not completely dissolved, evaporate the solution to dryness, then add 3-5 mL of hydrofluoric acid and 0.5 mL of perchloric acid to continue digestion.

[0111] At the same time, prepare the reagent blank solution as above.

[0112] Calibration curve drawing: accurately pipette 10mL of 1000mg / L potassium standard solution into a 100mL volumetric flask, dilute to volume with deionized water, and mix well; this is the 100mg / L potassium standard solution; according to the linear detection range of potassium of the instrument used, dilute the 100mg / L potassium standard solution with deionized water into a series of standard solutions of no less than five concentrations; before making up to volume, add appropriate amounts of sodium chloride solution and reagent blank solution so that the sodium ion concentration of the series standards is 1000mg / L, and the reagent blank solution and the soil digestion solution are equal; then measure according to the instrument instruction manual, and adjust the instrument zero point with the solution with zero potassium concentration in the series standard solutions; draw the calibration curve with grid coordinate paper, or calculate the linear regression equation.

[0113] Quantitative determination of potassium: Absorb a certain amount of soil digestion solution and dilute it with deionized water until the potassium ion concentration is equivalent to the concentration range of the potassium series standard solution. This is the soil test solution; before making up to volume, add an appropriate amount of sodium chloride solution to make the sodium ion concentration 1000 mg / L; then perform the determination according to the instrument instruction manual, and adjust the instrument zero point with a solution with zero potassium concentration in the series of standard solutions; find out the potassium concentration in the test solution from the calibration curve or calculate it from the linear regression equation.

[0114] Percentage of total potassium in soil (calculated based on dry soil)

[0115]

[0116] in:

[0117] C is the potassium concentration obtained from the calibration curve or regression equation, in milligrams per liter (mg / L);

[0118] V1——digestion solution volume, mL;

[0119] V2——digestion solution absorption, mL;

[0120] V3——volume of the liquid to be tested, mL;

[0121] m——sample weight, g;

[0122] 10 －4 ——The coefficient for converting mg / L to percentage;

[0123] H——moisture content in air-dried soil

[0124] ⑥ Detection of hydrolyzable nitrogen

[0125] Digestion: Weigh about 1g of air-dried soil sample that has passed through a 0.25mm sieve (accurate to 0.0001g, containing about 1mg of nitrogen); place the sample into the bottom of a dry digestion tube, drip a small amount of deionized water (about 0.5mL~1mL) to wet the sample; add 2g of accelerator and 5mL of sulfuric acid, and shake gently; add a reflux device or place a small curved-necked glass funnel at the mouth of the tube, and place it in a digestion furnace for low-temperature heating; when the reaction in the tube is moderate (about 10~15 minutes), raise the furnace temperature to 360℃~380℃; it is best to condense and reflux H2SO4 vapor at the upper 1 / 3 of the bottleneck; after the digestion liquid and soil particles have all turned grayish white with a slight green color, continue digestion for another hour; after digestion is completed, cool it and wait for distillation;

[0126] Distillation and titration of ammonia: refer to the instrument manual and use sulfuric acid or hydrochloric acid standard titration solution; set to add 10mL~30mL of water, 25mL of sodium hydroxide solution and 20mL~30mL of boric acid absorption solution; place the digestion tube on the automatic nitrogen analyzer for distillation and titration; use a blank solution, and the other steps are the same as the determination of the sample solution.

[0127] Result calculation

[0128] The total nitrogen (N) content in the soil sample is expressed as mass fraction ω and the value is expressed as percentage (%) and is calculated according to the following formula:

[0129]

[0130] Where:

[0131] C H ——Concentration of acid standard titration solution, in moles per liter (mol / L);

[0132] V——The volume of ferrous sulfate standard solution consumed in the sample determination, in milliliters (mL);

[0133] V0——The volume of ferrous sulfate standard solution consumed in the blank test, in milliliters (mL);

[0134] m——Weigh the mass of the dried sample, in grams (g);

[0135] f——Oxidation correction factor.

[0136] ⑦ Detection of available phosphorus

[0137] Extraction of available phosphorus: weigh 2.50 g of air-dried sample that has passed through a 2 mm sieve and place it in a 200 mL plastic bottle; add 50.00 mL of sodium bicarbonate extractant at (25 ± 1) °C;

[0138] Drawing of standard curve: Pipette 0.00mL, 0.50mL, 1.00mL, 2.00mL, 3.00mL, 4.00mL, 5.00mL of phosphorus standard solution into 25mL volumetric flask respectively; add 10.00mL of sodium bicarbonate extractant and 5.00mL of molybdenum antimony anti-color developer, shake slowly, expel CO2 and add water to make up the volume; obtain phosphorus standard series solutions containing 0.00mg / L, 0.10mg / L, 0.20mg / L, 0.40mg / L, 0.60mg / L, 0.80mg / L, 1.00mg / L; after standing for 30 minutes at room temperature above 20℃, use 1cm optical path cuvette at wavelength 880nm, adjust to the zero point of standard solution, and perform colorimetric determination to draw standard curve;

[0139] Determination: Pipette 10.00mL of sample solution into a 50mL volumetric flask or conical flask, slowly add 5.00mL of molybdenum antimony anti-color developer, shake slowly to expel CO2; then add 10.00mL of water, shake well to expel CO2; let stand for 30 minutes at room temperature above 20℃, and then use a 1cm light path cuvette to adjust the zero point of the standard solution at a wavelength of 880nm for colorimetric determination; if the measured phosphorus mass concentration exceeds the range of the standard curve, dilute the sample solution with an extractant and re-determine the colorimetric determination; at the same time, determine the blank solution.

[0140] Calculation results:

[0141] The available phosphorus (P) content in soil samples is expressed as mass fraction ω and the value is expressed in milligrams per kilogram (mg / kg) and is calculated according to the following formula:

[0142]

[0143] Where:

[0144] p——The phosphorus concentration in the colorimetric solution obtained from the standard curve, in milligrams per liter (mg / L);

[0145] ρ0——Phosphorus concentration in blank sample obtained from standard curve, in milligrams per liter (mg / L);

[0146] V——the volume of the color developing solution, in milliliters (mL);

[0147] D——fractionation multiple, the ratio of sample extract volume to fraction volume;

[0148] m——sample mass, in grams (g);

[0149] 1000 – coefficient for converting mL to L and g to kg;

[0150] ⑧Detection of fast-acting potassium

[0151] Weigh 5 g (accurate to 0.01 g) of air-dried soil sample that has passed through a 1 mm pore size sieve into a 200 mL plastic bottle (or 100 mL conical flask); add 50.0 mL of ammonium acetate solution (soil-liquid ratio is 1:10) and cover the bottle tightly; oscillate at 150 to 180 rpm for 30 minutes at 20 to 25 °C; after dry filtration, measure the filtrate directly on a flame photometer; and perform a blank test at the same time;

[0152] Drawing of standard curve: Pipette the following volumes (mL) of potassium standard solution: 0.00, 3.00, 6.00, 9.00, 12.00, 15.00 into a 50mL volumetric flask; make up to volume with ammonium acetate solution to obtain a series of potassium standard solutions with concentrations (μg / mL) of 0, 6, 12, 18, 24, 30; adjust the instrument zero point with a solution with a potassium concentration of 0, measure with a flame photometer, and draw a standard curve or find the regression equation.

[0153] Result calculation: The available potassium content in soil is expressed as the mass fraction of potassium (K) ω1, and the value is expressed in milligrams per kilogram (mg / kg), calculated according to the following formula:

[0154]

[0155] Where:

[0156] c1——The concentration of potassium in the test solution is obtained by checking the standard curve or finding the regression equation, in micrograms per milliliter.

[0157] (μg / mL);

[0158] V1——the value of the volume of the extractant, in milliliters (mL);

[0159] m1 – the numerical value of the mass of the sample, in grams (g).

[0160] After the sesquiterpene lactone of the present invention is used in the preparation of a soil conditioner, in comparison with a soil experiment without sesquiterpene lactone, the soil nutrients are shown in Table 1 below:

[0161] Table 1 Soil nutrients

[0162]

[0163] Note: GD: Guangdong, HN: Hainan, YN: Yunnan, GX: Guangxi; MM refers to the soil where Mikania micrantha is planted (containing sesquiterpene lactones)

[0164] According to the experimental results and Table 1, compared with the soil without sesquiterpene lactones, the contents of organic matter (OM), total nitrogen (AN), hydrolyzable nitrogen (HN), total phosphorus (AP), available phosphorus (HP) and available potassium (HK) in the soil with sesquiterpene lactones increased significantly. The nutrient comparison bar chart is shown in Figure 1. Figure 1 shown.

[0165] The present invention provides the use of sesquiterpene lactone in preparing a soil conditioner. Sesquiterpene lactone is a class of natural compounds with extensive biological activity. As a plant defense substance and participating in the interaction between plants and the environment, sesquiterpene lactone can affect the interaction between plants and soil microorganisms, promote the symbiotic relationship between plant roots and fungi, help plants absorb nutrients in the soil, especially inorganic phosphorus, and affect the generation of effective nitrogen in different forms and the nitrogen cycle process, thereby achieving an enrichment effect, increasing soil fertility, increasing the total organic matter content in the soil, and improving soil pH conditions.

[0166] The above description is only a preferred example of the present invention and is not intended to limit the present invention. Although the present invention is described in detail with reference to the aforementioned embodiments, those skilled in the art can still modify the technical solutions described in the aforementioned embodiments or replace some of the technical features therein by equivalents. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. The use of sesquiterpene lactone in the preparation of soil conditioner is characterized by: The invention comprises a method for preparing sesquiterpene lactone, a method for detecting sesquiterpene lactone in soil and a method for detecting soil physical and chemical properties. The method for preparing sesquiterpene lactone comprises: directly extracting sesquiterpene lactone from the above-ground tissue of Mikania micrantha or directly burying the fallen leaves of the above-ground part of Mikania micrantha in the soil.

2. The use of sesquiterpene lactone in the preparation of a soil conditioner according to claim 1, characterized in that: The method for detecting sesquiterpene lactones in soil comprises the following steps: (1) Soil nutrient extraction; (2) UPLC-MS quantitative testing; (3) Experimental data analysis.

3. The use of sesquiterpene lactone in the preparation of a soil conditioner according to claim 2, characterized in that: The soil nutrient extraction specifically comprises the following steps: A. Select fresh soil samples, air-dry them, and filter them through a 30-mesh sterile sieve to remove the fallen leaves of Mikania micrantha in the soil; B. Take 15g soil powder, add 30ml methanol solvent, and use ultrasonic extraction for 15min, a total of two extractions; C. Combine the extracts and centrifuge at 500 rpm for 4 min. D. Take the supernatant and concentrate it under reduced pressure, then use 1.5 ml of chromatographic methanol to reconstitute it, and then filter it through a 0.22 μm PETE filter membrane for use.

4. The use of sesquiterpene lactone in the preparation of a soil conditioner according to claim 2, characterized in that: The UPLC-MS quantitative test specifically comprises the following steps: A. The samples were separated and analyzed by ThermoScientific quadrupole-orbitrap high-resolution liquid chromatography-mass spectrometry. The injector temperature was set at 4°C and random injection was performed. B. Detection conditions: chromatographic column temperature 25°C, flow rate 0.25 ml / min, injection volume 1 μl, mobile phase A 0.1% formic acid, mobile phase B acetonitrile; C. Gradient elution program: total program duration 30 min, 0-2.5 min, 95% B, 2.5-25 min, B phase linearly changes from 95% to 5%; 25-30 min, B phase linearly changes from 5% to 95%; D. Set up QC samples during the experiment to monitor the stability of the instrument and experimental data, and use them to calculate the relative content of sesquiterpene lactones; E. Heated electrospray ionization (HESI) negative ion mode kit: In negative ion mode, the spray voltage is 2800 V, the capillary temperature is 340°C, the maximum allowable spray current of the ion source is 100 mA, the probe heater temperature is 340°C, and the mass-to-charge ratio scanning range is 80-1000 Da.

5. The use of sesquiterpene lactone in the preparation of a soil conditioner according to claim 2, characterized in that: The experimental data analysis specifically includes the following steps: A. Browse, check and preliminarily analyze the original data from the machine through Xcalibur, including the total ion current (TIC) in full scan mode, selected ion monitoring (SIM) data and secondary mass spectrometry (MS / MS) spectra; B. Use the software CompoundDiscover3.3 (Thermo) to identify metabolites and search common online metabolomics databases such as mzCloud, MassList, ChemSpider, and Thermo Fisher's own databases (such as mzVault TM Spectral library or mass list), peak alignment, retention time correction and peak area extraction were performed to clarify the types and relative contents of metabolites. The retention times of five sesquiterpene lactones, 3-epi, Deo, Anh, Scan and Dih, were 9.30min, 9.30min, 9.20min, 9.20min and 8.44min, respectively. The above-mentioned sesquiterpene lactones were detected in each soil sample by comparison with the standard.

6. The use of sesquiterpene lactone in the preparation of a soil conditioner according to claim 1, characterized in that: The soil physical and chemical property detection method comprises the following steps: ①Detection of organic matter; ② Determination of soil pH; ③ Detection of total nitrogen; ④Total phosphorus detection; ⑤ Detection of total potassium; ⑥ Detection of hydrolyzable nitrogen; ⑦ Detection of available phosphorus; ⑧Detection of fast-acting potassium.

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