Method for detecting mineralization characteristics of nitrogen in organic material and application of method

Abstract

The invention provides a method for detecting organic material nitrogen mineralization characteristics and application thereof, and belongs to the technical field of organic fertilizer mineralization detection. The method for detecting the mineralization characteristic of the nitrogen in the organic material comprises the following steps: carrying out 15N labeling on the organic material, mixing the organic material subjected to 15N labeling with soil, collecting a mixed sample, determining the content of nitrate nitrogen, the content of ammonium nitrogen, the abundance of 15N in nitrate nitrogen and the abundance of 15N in ammonium nitrogen, and then calculating the net mineralization rate of the nitrogen in the organic material. The method for detecting the mineralization characteristics of the nitrogen in the organic material can detect the mineralization process of the organic material after the organic material is added into soil, and has the advantages of high detection sensitivity and high accuracy. By measuring the nitrogen mineralization rate of the organic materials (such as corn straw and chicken manure organic fertilizer), the nitrogen release dynamic state of the organic materials in the soil can be quantified, and the optimal fertilization time and dosage are matched for the growth cycle of crops.

Description

Technical Field

[0001] This invention belongs to the field of organic fertilizer mineralization detection technology, specifically relating to a method for detecting the nitrogen mineralization characteristics of organic materials and its application. Background Technology

[0002] Soil nitrogen mainly exists in the form of organic nitrogen. In the application of organic and inorganic fertilizers, the nitrogen in the organic fertilizer is also primarily organic nitrogen, which typically needs to be mineralized and decomposed by microorganisms into inorganic nitrogen before being absorbed and utilized by crops. Therefore, analyzing the mineralization process of organic nitrogen is of great significance for gaining a deeper understanding of soil nitrogen supply potential and the nitrogen cycle, and for fully realizing the yield-increasing benefits of organic fertilizers.

[0003] Mineralization, as a crucial pathway for organic nitrogen to participate in the nitrogen cycle, has long been a focus of attention. The mineralization rate of organic nitrogen directly relates to the soil nitrogen turnover rate and productivity level. Studies have shown that the organic nitrogen mineralization process is not only closely related to soil properties such as soil temperature, moisture, pH, and SOC content, but also influenced by various factors such as fertilizer type and application rate. Fertilization can alter soil physicochemical properties, microbial activity, and aggregate structure, thereby affecting the nitrogen mineralization process and consequently influencing soil nitrogen supply and crop yield, especially when inorganic and organic fertilizers are applied in combination. A reasonable combination of organic and inorganic fertilizers can improve the soil organic nitrogen mineralization rate, enhance soil nitrogen supply, and promote crop growth and development. Previous nitrogen mineralization experiments have mostly focused on the mineralization of soil nitrogen after fertilization; research on the mineralization of organic fertilizers themselves in the soil is limited, and their nitrogen supply characteristics, supply potential, and mineralization mechanisms remain unclear, requiring further investigation.

[0004] Currently, various methods have been developed for detecting the nitrogen mineralization characteristics of organic materials, mainly divided into indoor culture methods and field in-situ culture methods. Among indoor culture methods, non-leaching aeration culture and undisturbed soil column culture can maintain the integrity of soil structure, but the accumulation of mineral nitrogen in the system can interfere with the subsequent organic nitrogen mineralization process, leading to an underestimation of the organic nitrogen mineralization potential. Intermittent leaching aeration culture, by simulating the absorption and removal of mineral nitrogen by plants, is suitable for rapid determination of large numbers of samples and can eliminate the influence of mineralized nitrogen accumulation, making it widely used. However, this method damages soil structure and changes aeration conditions, which can easily lead to an overestimation of the organic nitrogen mineralization potential. Among in-situ field culture methods, the polyethylene bag culture method is the most widely used, but it has the problem of being impermeable to water, which cannot truly reflect the field moisture conditions. Moreover, sampling damages the soil structure, and soil animals and plant roots may damage the culture bag during the culture process, leading to the loss of mineralized nitrogen. Although the top-cover buried tube culture method overcomes the defects of impermeability and easy damage of polyethylene bags, it still cannot avoid the loss of mineralized nitrogen. The ion exchange resin core method can be used to cultivate without damaging the original soil condition, and it is sensitive to soil temperature, humidity and aeration conditions. It can eliminate the impact of mineralized nitrogen accumulation. However, the operation process is time-consuming and labor-intensive, and ion competition and resin adsorption saturation point must be considered in advance.

[0005] Current research faces numerous challenges in determining the nitrogen mineralization characteristics of organic materials. On the one hand, different detection methods have limitations, making it difficult to accurately reflect the actual mineralization situation and limiting their guidance for the rational application of organic fertilizers in agricultural production. On the other hand, many factors influence the nitrogen mineralization of organic materials, such as organic fertilizer characteristics (C / N ratio, carbon and nitrogen compound composition, degree of maturity, etc.), temperature, moisture, and soil texture. These factors are intertwined, making it difficult for existing research to comprehensively consider them and construct accurate models for predicting the nitrogen mineralization characteristics of organic materials, thus failing to meet the needs of agricultural production in different regions. Therefore, developing a more accurate and efficient method for detecting the nitrogen mineralization characteristics of organic materials that can comprehensively consider multiple factors and overcome the shortcomings of existing methods has become a key issue that urgently needs to be addressed in this field. Summary of the Invention

[0006] In view of this, the purpose of the present invention is to provide a method for detecting the nitrogen mineralization characteristics of organic materials, which can detect the mineralization of the organic materials themselves and has the advantages of high detection sensitivity and high accuracy.

[0007] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0008] This invention provides a method for detecting the nitrogen mineralization characteristics of organic materials, comprising the following steps: processing the organic materials... 15 The N flag will 15 N-labeled organic matter was mixed with soil, and mixed samples were collected to determine the nitrate nitrogen content, ammonium nitrogen content, and nitrate nitrogen concentration. 15N abundance and ammonium nitrogen 15 N abundance is calculated using the following formula to determine the net mineralization rate of nitrogen in organic materials: Organic materials 15 N - Net mineralization rate = Mixed sample 15 N - Net nitrification rate + Mixed sample 15 N-net ammoniation rate, in which mixed samples 15 Net nitrification rate (mg·kg) -1 ·d 1 )=[c(NO3 --15 N) i+1 -c(NO3) --15 N) i ] / (t i+1 -t i ), mixed samples 15 Net ammoniation rate of N- (mg·kg) -1 ·d 1 )=[c(NH4 +-15 N) i+1 -c(NH4) +-15 N) i ] / (t i+1 -t i In the formula, i is the number of samples, t is the sampling time, and t is the sampling time. i+1 -t i The interval between the two samplings is 10 days, and c represents the measurement result; the organic material includes corn stalks or chicken manure organic fertilizer.

[0009] Preferably, when the organic material is corn stalks, the organic material is processed... 15 The nitrogen labeling method was as follows: double-labeled ammonium nitrate was applied at a rate of 80.77 mg N / kg before corn sowing and at the jointing stage. 15 NH4- 15 NO3.

[0010] Preferably, when the organic material is chicken manure organic fertilizer, the organic material is processed... 15 The method for N-labeling is as follows: Double-labeled ammonium nitrate... 15 NH4- 15 NO3 was mixed with water and wheat bran, fed to chickens, and the chicken manure was collected and decomposed in a sealed container. 15 N-labeled chicken manure organic fertilizer.

[0011] Preferred, dual-labeled ammonium nitrate 15 NH4- 15 The weight ratio of NO3, water, and wheat bran is 2:500:750.

[0012] Preferably, the room temperature for fermentation is 27-30°C, the fermentation time is 30-35 days, and the mixture is stirred once every 9-11 days during the fermentation process.

[0013] Preferably, a 2 mol / L KCl solution is used, and the mixture is prepared by mixing water and sample in a ratio of 5:1, shaking at 180 rpm for 1 hour, filtering, and collecting the filtrate to determine the nitrate nitrogen content and ammonium nitrogen content.

[0014] Preferably, nitrate nitrogen 15 The method for determining nitrogen abundance is as follows: denitrifying bacteria are used to convert nitrate nitrogen in the filtrate into N2O, and TraceGas combined with isotope mass spectrometry is used to determine the nitrogen isotope of N2O. USGS32, USGS34 and IAEAN3 are used as standards, and the measured gas is corrected using a two-point correction method.

[0015] Preferably, ammonium nitrogen 15 The method for determining nitrogen abundance is as follows: the filtrate is concentrated by diffusion absorption, and then measured using an elemental analyzer combined with an isotope mass spectrometer.

[0016] This invention also provides applications of the above-described method in agricultural production, ecological research, or environmental management.

[0017] The beneficial effects of this invention are:

[0018] The method for detecting the nitrogen mineralization characteristics of organic materials provided by this invention can detect the mineralization process of organic materials themselves after they are added to soil, and has the advantages of high detection sensitivity and high accuracy. By measuring the nitrogen mineralization rate of organic materials (such as corn stalks and chicken manure organic fertilizer), this method can quantify the nitrogen release dynamics in the soil, allowing for optimal fertilization time and dosage matching for crop growth cycles. Attached Figure Description

[0019] Figure 1 The content of nitrate nitrogen, ammonium nitrogen, and nitrate nitrogen in soil of different groups were measured. 15 N content and ammonium nitrogen 15 The nitrogen content detection results are shown in (a) for nitrate nitrogen, (b) for ammonium nitrogen, and (c) for nitrate nitrogen. 15 The nitrogen content detection results (d) show the ammonium nitrogen content. 15 N content test results;

[0020] Figure 2 For different groups of soil 15 N-Net nitrification rate, 15 N-net ammoniation rate and 15 The N-net mineralization rate results, where (a) to (c) represent different groups. 15 The net nitrification rate results (d) to (f) are for different groups. 15The net N-ammoniation rate results, (g)~(i) represent different groups. 15 N-Net Mineralization Rate Results. Detailed Implementation

[0021] This invention provides a method for detecting the nitrogen mineralization characteristics of organic materials, comprising the following steps: processing the organic materials... 15 The N-mark will 15 N-labeled organic matter was mixed with soil, and mixed samples were collected to determine the nitrate nitrogen content, ammonium nitrogen content, and nitrate nitrogen concentration. 15 N abundance and ammonium nitrogen 15 N abundance is calculated using the following formula to determine the net mineralization rate of nitrogen in organic materials: Organic materials 15 N - Net mineralization rate = Mixed sample 15 N - Net nitrification rate + Mixed sample 15 N-net ammoniation rate, in which mixed samples 15 Net nitrification rate (mg·kg) -1 ·d 1 )=[c(NO3 --15 N) i+1 -c(NO3) --15 N) i ] / (t i+1 -t i ), mixed samples 15 Net ammoniation rate of N- (mg·kg) -1 ·d 1 )=[c(NH4 +-15 N) i+1 -c(NH4) +-15 N) i ] / (t i+1 -t i In the formula, i is the number of samples, t is the sampling time, and t is the sampling time. i+1 -t i The interval between the two samplings is 10 days, and c represents the measurement result; the organic material includes corn stalks or chicken manure organic fertilizer.

[0022] Stable isotope tracing technology, as an effective technique for studying material cycling and element migration and transformation, can accurately distinguish the source and distribution of nutrients. Although different fertilization methods may lead to different results... 15 Numerous reports have documented how nitrogen-labeled controlled-release urea or labeled ordinary urea can increase yield and improve nitrogen fertilizer utilization. However, most studies focus on single-labeled urea, and applications are less well-documented. 15 N-labeled straw or 15 The quantitative study of N-labeled organic fertilizer in combination with ordinary urea on the characteristics of organic nitrogen mineralization in straw and organic fertilizer is still insufficient.

[0023] In this invention, when the organic material is corn stalks, the organic material is processed... 15 The preferred method for N labeling is to apply double-labeled ammonium nitrate at a rate of 80.77 mg N / kg before sowing and at the jointing stage of maize. 15 NH4- 15 NO3. This invention relates to dual-labeled ammonium nitrate. 15 NH4- 15 There are no specific limitations on the specific source of NO3; any commercially available product conventional in this field is acceptable. In this invention, when the organic material is chicken manure organic fertilizer, the organic material is processed... 15 The preferred method for N-labeling is: to double-label ammonium nitrate. 15 NH4- 15 NO3 was mixed with water and wheat bran, fed to chickens, and the chicken manure was collected and decomposed in a sealed container. 15 N-labeled chicken manure organic fertilizer. This invention does not specifically limit the source of wheat bran; conventional commercially available products in the art are acceptable. In this invention, dual-labeled ammonium nitrate... 15 NH4- 15 The preferred weight ratio of NO3 water to wheat bran is 2:500:750. The preferred room temperature for fermentation is 27-30°C, more preferably 28-29°C, and the preferred fermentation time is 30-35 days, more preferably 32-34 days. During the fermentation process, it is preferred to stir once every 9-11 days.

[0024] In this invention, 15 When nitrogen-labeled organic materials are mixed with soil, especially when the organic material is corn stalks... 15 The preferred weight ratio of nitrogen-labeled corn stalks to dry soil is 0.3:100, especially when the organic material is chicken manure organic fertilizer. 15 The preferred amount of nitrogen-labeled chicken manure organic fertilizer added to the soil, based on nitrogen content, is 80.77 mg / kg. -1 This is approximately equivalent to a nitrogen fertilizer application rate of 210 kg N·hm. -2 The bulk density is 1.3 g·cm³. -3 calculate.

[0025] In this invention, when collecting mixed samples for measurement, the time interval between the two collections is preferably 5 to 14 days. Before measurement, it is preferable to extract nitrate nitrogen and ammonium nitrogen from the mixed sample. A preferred extraction method is to use a 2 mol / L KCl solution, mix the sample with water at a ratio of 5:1, shake at 180 rpm for 1 hour, filter, collect the filtrate, and then determine the nitrate nitrogen and ammonium nitrogen content. This invention does not specifically limit the method for determining the nitrate nitrogen and ammonium nitrogen content; conventional methods for detecting nitrate nitrogen and ammonium nitrogen content in the art can be used.

[0026] In this invention, nitrate nitrogen 15 The preferred method for determining nitrogen abundance is as follows: Denitrifying bacteria convert nitrate nitrogen in the filtrate into N₂O, and TraceGas combined with isotope mass spectrometry is used to determine the nitrogen isotopes of N₂O. USGS32, USGS34, and IAEA N₃ are used as standards, and a two-point calibration method is employed to correct the measured gas. (Ammonium nitrogen...) 15 The preferred method for determining nitrogen abundance is as follows: the filtrate is concentrated using diffusion absorption, followed by determination using an elemental analyzer combined with isotope mass spectrometry. This invention is applicable to the determination of nitrogen abundance in nitrate nitrogen. 15 N abundance and ammonium nitrogen 15 The specific operational steps in the method for determining nitrogen abundance are not particularly limited; conventional operational steps in the field can be used. In a specific embodiment of the present invention, the above-mentioned determination process was conducted by the Institute of Agricultural Environment and Sustainable Development, Chinese Academy of Agricultural Sciences, and the specific determination method is as follows:

[0027] In nitrate nitrogen 15 Method for determining nitrogen abundance: Instrumentation: DeltaV-Precon (Thermo Fisher Scientific, Germany), instrument accuracy: 15 N≤0.5‰, 18 O ≤ 1‰. The δ¹⁴ of NO₃⁻N was determined using a bacterial denitrification method. 15 N- is first converted to N2O by denitrifying bacteria (Pseudomonas aeruginosa, ATCC 13985, USA) lacking N2O reductase activity, through a process called δ-. 15 The nitrogen (N) determination method is the same as that in the invention patent with patent number ZL201210228975.0, entitled "A Method for Cultivating Denitrifying Bacteria and Determining the Nitrate Nitrogen Isotope Composition in Water". The generated N₂O is analyzed using a micro-gas concentration preparation device (Precon, Finnigan, Germany) and an isotope ratio mass spectrometer (IRMS) (DeltaVplus, Finnigan, Germany) with a delta-grain ratio. 15 N₂O analysis. N₂O samples were headspace-sampled using an autosampler, and H₂O / CO₂ was removed using two chemical traps (magnesium perchlorate trap and sodium hydroxide trap, Merck KGaA). After cryogenic trapping and concentration, N₂O was transferred at 35°C to a capillary column (PoraPlot Q, 25m, 0.32mm id, 10mm df, Agilent Technologies, USA) and subsequently analyzed by IRMS. The N₂O sample was analyzed using a USGS32 (δ¹²) microarray. 15 N is 180.0±1.0‰), USGS34 (δ 15 N is 1.8±0.2‰), IAEAN3 (δ 15(N = 4.7 ± 0.2‰) Calibration of δ 15 N.

[0028] ammonium nitrogen 15 Method for determining nitrogen abundance: Measure 30 mL of the filtrate and place it in a 60 mL HDPE bottle. Add heavy magnesium oxide to the bottle. Using a diffusion enrichment ammonium ion separation device, wrap a filter membrane containing potassium bisulfate droplets with a hydrophobic membrane and float it in the middle of a quartz tube. Place the tube in the HDPE bottle, seal the bottle, and incubate at 40℃ for 7–10 days. Ammonium ions in the water are converted into ammonia gas. The ammonia gas reacts with the NaHSO4 solution in the filter membrane to form sulfate, which is fixed by the filter membrane. Afterward, remove the filter membrane and place it in a desiccator containing desiccant and concentrated sulfuric acid. After 2 days, remove the membrane, wrap it in a foil cup, and insert the wrapped sample into an elemental analyzer (Vario PYRO cube, Elementar, Germany) via an autosampler. The sample is converted into pure CO2 and N2 gases through combustion and reduction (combustion furnace temperature 1020℃, reduction furnace temperature 650℃, carrier gas He flow rate 230 mL·min). -1 The CO2 was then diluted using a diluent and finally analyzed using a stable isotope mass spectrometer (IsoPrime 100, Isoprime, UK). One laboratory standard sample (δ¹²) was placed between every 12 samples. 15 N air =7.1‰), used for quality control of measurement results. Results are corrected using a two-point calibration. That is, using USGS40 (δ) 15 N air =-4.52‰) and USGS41a(δ 15 N air =+47.55‰) to correct the nitrogen determination results.

[0029] This invention also provides applications of the above-mentioned method in agricultural production, ecological research, or environmental management. The method provided by this invention can dynamically assess the soil's nitrogen supply capacity after applying organic fertilizer. By measuring the nitrogen mineralization rate of organic materials, the dynamics of nitrogen release in the soil can be quantified, allowing for optimal fertilization timing and dosage to be matched to the crop growth cycle. By accurately detecting the nitrogen mineralization characteristics in organic materials using the method of this invention, the nitrogen mineralization rate of organic materials can be precisely matched with crop requirements, reducing the amount of both organic and chemical fertilizers used.

[0030] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0031] Unless otherwise specified, the following embodiments are all conventional methods.

[0032] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0033] Example 1

[0034] A method for detecting nitrogen mineralization characteristics in corn stalks, comprising the following steps:

[0035] A pot experiment was conducted, and double-labeled ammonium nitrate was applied to maize before sowing and at the jointing stage at a concentration of 80.77 mg N / kg soil. 15 NH4- 15 NO3 affects corn stalks 15 N-marked, collect mature corn stalks, obtain 15 N-labeled corn stalks, after testing, 15 The carbon and nitrogen content of nitrogen-labeled corn stalks was 482.53 g·kg⁻¹. -1 and 5.9 g·kg -1 , 15 The nitrogen abundance was 4.08%.

[0036] Will 15 N-labeled corn stalks were mixed with the soil at a rate of 0.3% of the dry soil weight (equivalent to 7500 kg·hm²). -2 The mixed sample (the amount of nitrogen returned to the field) was placed in a 500mL culture bottle with a sealed rubber stopper (the rubber stopper had a hole punched in the middle, through which a hollow glass tube was passed, with its lower end placed in the middle of the upper space inside the bottle, and the upper end connected to a PVC hose with a three-way valve; the device was checked for sealing before use). The culture bottle was incubated in a constant temperature incubator at 25℃ in the dark. Every two days during the incubation process, the soil moisture content was controlled to be 60% of field capacity using a weighing method. Mixed samples were collected on days 5, 14, 28, and 42. Three samples were randomly selected from each treatment for destructive sampling to determine the nitrate nitrogen content, ammonium nitrogen content, and nitrate nitrogen concentration. 15 N abundance and ammonium nitrogen 15 N abundance was determined by the following method:

[0037] First, use a 2 mol / L KCl solution to mix the sample with water at a ratio of 5:1, shake at 180 rpm for 1 hour, filter, collect the filtrate, and determine the nitrate nitrogen content and ammonium nitrogen content.

[0038] In nitrate nitrogen 15 Nitrogen abundance determination: Denitrifying bacteria converted nitrate nitrogen in the filtrate into N₂O. The nitrogen isotopes of N₂O were determined using TraceGas combined with isotope mass spectrometry (Isoprame-100, UK). USGS32, USGS34, and IAEAN3 were used as standards, and a two-point calibration method was employed to correct the measured gas. Specific detection methods: Instrument: DeltaV-Precon (Thermo Fisher Scientific, Germany); Instrument accuracy:15 N≤0.5‰, 18 O ≤ 1‰. The δ¹⁴ of NO₃⁻N was determined using a bacterial denitrification method. 15 N- is first converted to N2O by denitrifying bacteria (Pseudomonas aeruginosa, ATCC 13985, USA) lacking N2O reductase activity, through a process called δ-. 15 The nitrogen (N) determination method is the same as that in the invention patent with patent number ZL201210228975.0, entitled "A Method for Cultivating Denitrifying Bacteria and Determining the Nitrate Nitrogen Isotope Composition in Water". The generated N₂O is analyzed using a micro-gas concentration preparation device (Precon, Finnigan, Germany) and an isotope ratio mass spectrometer (IRMS) (DeltaVplus, Finnigan, Germany) with a delta-grain ratio. 15 N₂O analysis. N₂O samples were headspace-sampled using an autosampler, and H₂O / CO₂ was removed using two chemical traps (magnesium perchlorate trap and sodium hydroxide trap, Merck KGaA). After cryogenic trapping and concentration, N₂O was transferred at 35°C to a capillary column (PoraPlot Q, 25m, 0.32mm id, 10mm df, Agilent Technologies, USA) and subsequently analyzed by IRMS. The N₂O sample was analyzed using a USGS32 (δ¹²) microarray. 15 N is 180.0±1.0‰), USGS34 (δ 15 N is 1.8±0.2‰), IAEAN3 (δ 15 (N = 4.7 ± 0.2‰) Calibration of δ 15 N.

[0039] ammonium nitrogen 15 Determination of nitrogen abundance: The filtrate was concentrated by diffusion absorption and then measured using an elemental analyzer combined with an isotope mass spectrometer (Isoprame-100, UK). The specific detection method is as follows: Measure 30 mL of the filtrate and place it in a 60 mL HDPE bottle. Add heavy magnesium oxide to the bottle. Using a diffusion enrichment ammonium ion separation device, wrap a filter membrane containing potassium bisulfate droplets with a hydrophobic membrane and float it in the middle of a quartz tube. Place the tube in the HDPE bottle, seal the bottle, and incubate at 40℃ for 7–10 days. Ammonium ions in the water are converted into ammonia gas. The ammonia gas reacts with the NaHSO4 solution in the filter membrane to form sulfate, which is fixed by the filter membrane. Afterward, remove the filter membrane and place it in a desiccator containing desiccant and concentrated sulfuric acid. After 2 days, remove the membrane, wrap it in a foil cup, and insert the wrapped sample into an elemental analyzer (Vario PYRO cube, Elementar GmbH, Germany) via an autosampler. The sample is converted into pure CO2 and N2 gases through combustion and reduction (combustion furnace temperature 1020℃, reduction furnace temperature 650℃, carrier gas He flow rate 230 mL·min).-1 The CO2 was then diluted using a diluent and finally analyzed using a stable isotope mass spectrometer (IsoPrime 100, Isoprime, UK). One laboratory standard sample (δ¹²) was placed between every 12 samples. 15 N air =7.1‰), used for quality control of measurement results. Results are corrected using a two-point calibration. That is, using USGS40 (δ) 15 N air =-4.52‰) and USGS41a(δ 15 N air =+47.55‰) to correct the nitrogen determination results.

[0040] The above measurement process was conducted by the Institute of Agricultural Environment and Sustainable Development, Chinese Academy of Agricultural Sciences. The net mineralization rate of nitrogen in organic materials was then calculated using the following formula: Organic materials 15 N - Net mineralization rate = Mixed sample 15 N - Net nitrification rate + Mixed sample 15 N-net ammoniation rate, in which mixed samples 15 Net nitrification rate (mg·kg) -1 ·d 1 )=[c(NO3 --15 N) i+1 -c(NO3) --15 N) i ] / (t i+1 -t i ), mixed samples 15 Net ammoniation rate of N- (mg·kg) -1 ·d 1 )=[c(NH4 +-15 N) i+1 -c(NH4) +-15 N) i ] / (t i+1 -t i In the formula, i is the number of samples, and t is the sampling time; t i+1 -t i This represents the number of days between two consecutive samplings.

[0041] Example 2

[0042] A method for detecting the nitrogen mineralization characteristics of chicken manure organic fertilizer, comprising the following steps:

[0043] Four adult Zhengzhou Red chickens were selected. Three days prior to the selection, they were fed unlabeled wheat bran, then starved for 15 hours to empty their intestines and remove any remaining feces. Afterward, double-labeled ammonium nitrate was used. 15 NH4- 15NO3 (Shanghai Chemical Research Institute, abundance 10.11%) was added to water and wheat bran, and then mixed with double-labeled ammonium nitrate. 15 NH4- 15 The weight ratio of NO3, water, and wheat bran was 2:500:750. Chickens were fed twice daily, morning and evening. Before each feeding, chicken manure was collected, and the mixed wheat bran was removed before being stored in a storage box. This feeding period lasted for 5 days. After the feeding period, all collected chicken manure was brought back to the laboratory for composting at room temperature (27-30°C) in a sealed environment. The composting process began on October 20, 2023, with chicken manure being divided into small storage boxes and composted, stirred every 9 days. The composting process ended on November 25, 2023. The composted chicken manure was then mixed, ground, and sieved to obtain… 15 N-labeled chicken manure organic fertilizer. Tests showed that... 15 The carbon and nitrogen content of nitrogen-labeled chicken manure organic fertilizer was 336.35 g·kg⁻¹. -1 and 67.8 g·kg -1 , 15 The nitrogen abundance was 1.27%.

[0044] Will 15 N-labeled chicken manure organic fertilizer is mixed with the soil and added... 15 The amount of nitrogen-labeled chicken manure organic fertilizer is based on an N content of 80.77 mg / kg soil (approximately equivalent to a nitrogen fertilizer application rate of 210 kg N·hm). -2 The bulk density is 1.3 g·cm³. -3 (Calculation). The mixed sample was placed in a 500mL culture bottle with a sealed rubber stopper (a hole was punched in the center of the rubber stopper, through which a hollow glass tube was passed, with its lower end placed in the middle of the upper space inside the bottle, and the upper end connected to a PVC hose with a three-way valve; the device was checked for sealing before use). The culture bottle was incubated in a constant temperature incubator at 25℃ in the dark. Every two days during the incubation process, the soil moisture content was controlled to be 60% of field capacity using a weighing method. Mixed samples were collected on days 5, 14, 28, and 42. Three samples were randomly selected from each treatment for destructive sampling to determine the nitrate nitrogen content, ammonium nitrogen content, and nitrate nitrogen concentration. 15 N abundance and ammonium nitrogen 15 N abundance was determined by the following method:

[0045] First, a 2 mol / L KCl solution was used to mix the sample with water at a ratio of 5:1. The mixture was shaken at 180 rpm for 1 hour and then filtered. The filtrate was collected, and the contents of nitrate nitrogen and ammonium nitrogen were determined. (The remaining text appears to be incomplete and requires further context.) 15Nitrogen abundance determination: Denitrifying bacteria converted nitrate nitrogen in the filtrate into N₂O. The nitrogen isotopes of N₂O were determined using TraceGas combined with isotope mass spectrometry (Isoprame-100, UK). USGS32, USGS34, and IAEAN3 were used as standards, and a two-point calibration method was employed to correct the measured gas. Specific methods: Instrumentation: DeltaV-Precon (Thermo Fisher Scientific, Germany); Instrument accuracy: 15 N≤0.5‰, 18 O ≤ 1‰. The δ¹⁴ of NO₃⁻N was determined using a bacterial denitrification method. 15 N- is first converted to N2O by denitrifying bacteria (Pseudomonas aeruginosa, ATCC 13985, USA) lacking N2O reductase activity, through a process called δ-. 15 The nitrogen (N) determination method is the same as that in the invention patent with patent number ZL201210228975.0, entitled "A Method for Cultivating Denitrifying Bacteria and Determining the Nitrate Nitrogen Isotope Composition in Water". The generated N₂O is analyzed using a micro-gas concentration preparation device (Precon, Finnigan, Germany) and an isotope ratio mass spectrometer (IRMS) (DeltaVplus, Finnigan, Germany) with a delta-grain ratio. 15 N₂O analysis. N₂O samples were headspace-sampled using an autosampler, and H₂O / CO₂ was removed using two chemical traps (magnesium perchlorate trap and sodium hydroxide trap, Merck KGaA). After cryogenic trapping and concentration, N₂O was transferred at 35°C to a capillary column (PoraPlot Q, 25m, 0.32mm id, 10mm df, Agilent Technologies, USA) and subsequently analyzed by IRMS. The N₂O sample was analyzed using a USGS32 (δ¹²) microarray. 15 N is 180.0±1.0‰), USGS34 (δ 15 N is 1.8±0.2‰), IAEAN3 (δ 15 (N = 4.7 ± 0.2‰) Calibration of δ 15 N.

[0046] ammonium nitrogen 15Determination of nitrogen abundance: The filtrate was concentrated by diffusion absorption and then measured using an elemental analyzer combined with an isotope mass spectrometer (Isoprame-100, UK). The specific detection method is as follows: Measure 30 mL of the filtrate and place it in a 60 mL HDPE bottle. Add heavy magnesium oxide to the bottle. Using a diffusion enrichment ammonium ion separation device, wrap a filter membrane containing potassium bisulfate droplets with a hydrophobic membrane and float it in the middle of a quartz tube. Place the tube in the HDPE bottle, seal the bottle, and incubate at 40℃ for 7–10 days. Ammonium ions in the water are converted into ammonia gas. The ammonia gas reacts with the NaHSO4 solution in the filter membrane to form sulfate, which is fixed by the filter membrane. Afterward, remove the filter membrane and place it in a desiccator containing desiccant and concentrated sulfuric acid. After 2 days, remove the membrane, wrap it in a foil cup, and insert the wrapped sample into an elemental analyzer (Vario PYRO cube, Elementar GmbH, Germany) via an autosampler. The sample is converted into pure CO2 and N2 gases through combustion and reduction (combustion furnace temperature 1020℃, reduction furnace temperature 650℃, carrier gas He flow rate 230 mL·min). -1 The CO2 was then diluted using a diluent and finally analyzed using a stable isotope mass spectrometer (IsoPrime 100, Isoprime, UK). One laboratory standard sample (δ¹²) was placed between every 12 samples. 15 N air =7.1‰), used for quality control of measurement results. Results are corrected using a two-point calibration. That is, using USGS40 (δ) 15 N air =-4.52‰) and USGS41a(δ 15 N air =+47.55‰) to correct the nitrogen determination results.

[0047] The above measurement process was conducted by the Institute of Agricultural Environment and Sustainable Development, Chinese Academy of Agricultural Sciences. The net mineralization rate of nitrogen in organic materials was then calculated using the following formula: Organic materials 15 N - Net mineralization rate = Mixed sample 15 N - Net nitrification rate + Mixed sample 15 N-net ammoniation rate, in which mixed samples 15 Net nitrification rate (mg·kg) -1 ·d 1 )=[c(NO3 --15 N) i+1 -c(NO3) --15 N) i ] / (t i+1 -t i ), mixed samples 15 Net ammoniation rate of N- (mg·kg) -1 ·d 1 )=[c(NH4+-15 N) i+1 -c(NH4) +-15 N) i ] / (t i+1 -t i In the formula, i is the number of samples, and t is the sampling time; t i+1 -t i This represents the number of days between two consecutive samplings.

[0048] Example 3

[0049] The culture experiment was set up with 8 treatments: (1) no fertilizer (NF); (2) labeled urea ( 15 F); (3) Marking corn stalks ( 15 S)(same as Example 1); (4) Marking chicken manure organic fertilizer ( 15 M)(same as Example 2); (5) Mark corn stalks + urea ( 15 S+F); (6) Mark chicken manure organic fertilizer + urea ( 15 M+F); (7) Corn stalks + labeled urea (S+ 15 F); (8) Chicken manure organic fertilizer + labeled urea (M+ 15 F). Each treatment was replicated 12 times, and samples were taken at 4 time points. Before the experiment, the soil moisture of the treated soil was adjusted to 60% of field capacity. The soil was then mixed with the above groups and incubated in the dark at 25°C for 7 days to activate microorganisms and eliminate the wet-dry effect.

[0050] Mixed samples were collected on days 5, 14, 28, and 42. Three samples were randomly selected from each treatment. Destructive soil sampling was used to determine the content of nitrate nitrogen and ammonium nitrogen, as well as the concentration of nitrate nitrogen in... 15 N abundance and ammonium nitrogen 15 N abundance, the specific detection method is the same as in Example 1.

[0051] The results are as follows Figure 1 and Figure 2 As shown.

[0052] like Figure 1 As shown, compared to 15 Treatment F showed a decrease in soil nitrate nitrogen content with the addition of organic materials (corn stalks and chicken manure organic fertilizer), and except for 15 Except for treatment S, there were no significant differences among the fertilization treatments. 15 S treatment significantly reduced soil nitrate nitrogen content; treatment with organic fertilizer combined with chemical fertilizer ( 15 M+F and M+ 15 F) and 15 During the cultivation period, the ammonium nitrogen content in treatment F showed a trend of first increasing and then decreasing, and 15 M+F, M+15 F and 15 The ammonium nitrogen content in treatment F was higher than that in the early stage of cultivation. 15 M processing; 15 S+F and S+ 15 The ammonium nitrogen content in treatment F showed a trend of first decreasing, then increasing, and then decreasing again. 15 The ammonium nitrogen content in the S treatment showed an increasing trend during the incubation period. 15 M+F, M+ 15 F, 15 S and 15 S+F processing compared to 15 F significantly reduced nitrate nitrogen - 15 N content, of which M+ 15 F treatment of nitrate nitrogen - 15 N content is higher than 15 M+F processing, while 15 S and 15 S+F treatment of nitrate nitrogen - 15 The lowest N content; 15 M+F, 15 M and 15 S processing compared to 15 F significantly increased ammonium nitrogen- 15 N content, of which 15 M+F treatment of ammonium nitrogen 15 The nitrogen content showed a decreasing trend during cultivation, while 15 S and 15 Treatment M showed an increasing trend. Overall, during the incubation period, the contents of nitrate nitrogen and ammonium nitrogen in the soil fluctuated among the treatments. 15 M+F and 15 S treatment reduced nitrate nitrogen - 15 N content, but can increase ammonium nitrogen - 15 N content, treatment with organic materials alone ( 15 S and 15 M) Ammonium nitrogen- 15 The nitrogen content showed an increasing trend.

[0053] like Figure 2 As shown, soil 15 N- net nitrification rate is higher than 15 N-net ammoniation rate, with different organic materials added 15 N-Net nitrification rate and 15 The trends in the net N-ammonization rate varied among the treatments, but the differences between the treatments were significant. 15 N-net mineralization rate and 15 The net nitrification rate of N- shows a similar trend. Compared to 15 F-treatment, combined with organic material treatment (S+) 15 F and M+ 15F) both reduced the early stage (day 5) and late stage (day 42) of culture. 15 N- Net nitrification and mineralization rate, M+ 15 Treatment F improved the results on days 14 and 28. 15 N-Net nitrification, mineralization rate, and day 5 15 N-net ammoniation rate; 15 S and 15 S+F treatment soil 15 There was no significant difference in net nitrification and mineralization rates, except on day 5. 15 S processing 15 The net ammonification rate of N- was higher than that of all other processes. 15 S+F processing; compared to 15 M processing, 15 M+F processing 15 The net nitrification and mineralization rates decreased on day 5 and increased on days 14 and 28, while... 15 The net ammonification rate of N- is opposite. Generally speaking, the combined application of organic fertilizer and chemical fertilizer (M+) 15 F) processing compared to 15 F treatment improved the condition on days 14 and 28. 15 N-Net nitrification and mineralization rate, single application of straw ( 15 S) or apply organic fertilizer alone ( 15 M) treatment compared to the combined application of organic and inorganic fertilizers ( 15 S+F, 15 M+F) improved the 14th, 28th and 42nd days. 15 N-net ammoniation rate.

[0054] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A method for detecting the nitrogen mineralization characteristics of organic materials, characterized in that, The process includes the following steps: processing organic materials. 15 The N flag will 15 N-labeled organic matter was mixed with soil, and mixed samples were collected to determine the nitrate nitrogen content, ammonium nitrogen content, and nitrate nitrogen concentration. 15 N abundance and ammonium nitrogen 15 N abundance is calculated using the following formula to determine the net mineralization rate of nitrogen in organic materials: Organic materials 15 N - Net mineralization rate = Mixed sample 15 N - Net nitrification rate + Mixed sample 15 N-net ammoniation rate, in which mixed samples 15 Net nitrification rate (mg·kg) -1 ·d 1 )=[c(NO3 --15 N) i+1 -c(NO3) --15 N) i ] / (t i+1 -t i ), mixed samples 15 Net ammoniation rate of N- (mg·kg) -1 ·d 1 )=[c(NH4 +-15 N) i+1 -c(NH4) +-15 N) i ] / (t i+1 -t i In the formula, i is the number of samples, t is the sampling time, and t is the sampling period. i+1 -t i The interval between the two samplings is 10 days, and c represents the measurement result; the organic material includes corn stalks or chicken manure organic fertilizer.

2. The method according to claim 1, characterized in that, When the organic material is corn stalks, the organic material is processed... 15 The method for N labeling is as follows: double-labeled ammonium nitrate is applied at a rate of 80.77 mg N / kg before corn sowing and at the jointing stage. 15 NH4- 15 NO3.

3. The method according to claim 1, characterized in that, When the organic material is chicken manure organic fertilizer, the organic material is processed... 15 The method for N-labeling is as follows: Double-labeled ammonium nitrate... 15 NH4- 15 NO3 was mixed with water and wheat bran, fed to chickens, and the chicken manure was collected and decomposed in a sealed container. 15 N-labeled chicken manure organic fertilizer.

4. The method according to claim 3, characterized in that, Double-labeled ammonium nitrate 15 NH4- 15 The weight ratio of NO3, water, and wheat bran is 2:500:

750.

5. The method according to claim 3, characterized in that, The fermentation process takes place at a room temperature of 27–30°C for 30–35 days, and is stirred every 9–11 days during the fermentation process.

6. The method according to claim 1, characterized in that, Mix the sample with 2 mol / L KCl solution at a ratio of 5:1 (water:sample), shake at 180 rpm for 1 hour, filter, and collect the filtrate to determine the nitrate nitrogen content and ammonium nitrogen content.

7. The method according to claim 6, characterized in that, In nitrate nitrogen 15 The method for determining nitrogen abundance is as follows: denitrifying bacteria are used to convert nitrate nitrogen in the filtrate into N2O, and TraceGas combined with isotope mass spectrometry is used to determine the nitrogen isotope of N2O. USGS32, USGS34 and IAEAN3 are used as standards, and the measured gas is corrected using a two-point correction method.

8. The method according to claim 6, characterized in that, ammonium nitrogen 15 The method for determining nitrogen abundance is as follows: the filtrate is concentrated by diffusion absorption, and then measured using an elemental analyzer combined with an isotope mass spectrometer.

9. The application of the method according to any one of claims 1 to 8 in agricultural production, ecological research or environmental management.

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