A multi-component hierarchical detection method and detection system for phosphorus in soil
The integrated reaction vessel and two-step digestion process solve the problems of sample loss and safety in soil phosphorus classification and testing, and achieve efficient and accurate determination of soil phosphorus components.
Patent Information
- Application Number
- CN202610304374.5
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2026-03-13
- Publication Date
- 2026-05-29
AI Technical Summary
Existing methods for classifying and detecting phosphorus in soil suffer from problems such as physical loss of samples during multi-stage extraction, time-consuming digestion steps, and insufficient operational safety, which affect the accuracy and efficiency of the test results.
Multi-stage extraction is performed using an integrated reaction vessel, and a quantitative non-destructive transfer kit and a two-step digestion procedure are used, combined with potassium persulfate solution to digest organophosphorus compounds, ensuring sample integrity and safety.
It significantly improves the accuracy and safety of soil phosphorus component determination, reduces physical losses, shortens processing time, and enhances analytical efficiency.
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Figure CN122108721A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of soil chemical analysis technology, specifically to a multi-component classification detection method and system for phosphorus in soil. Background Technology
[0002] Soil phosphorus is a crucial nutrient element essential for plant growth and one of the main factors limiting agricultural productivity. Simultaneously, the migration and loss of soil phosphorus is a significant environmental problem leading to eutrophication of water bodies. Therefore, accurate classification and detection of different forms and availability of phosphorus components in soil are of vital theoretical and practical significance for scientifically guiding fertilization, evaluating soil phosphorus supply potential, and conducting environmental risk assessments.
[0003] Currently, chemical sequential extraction is widely used internationally for phosphorus fractionation in soil, with sequential extraction procedures being the most prevalent technique. This method typically involves sequential, multi-stage extraction and separation of the same soil sample using a series of extractants with different properties, following a decreasing order of chemical stability and bioavailability of phosphorus components. Finally, the extracted soil residue is digested with strong acid. In this way, phosphorus in the soil can be classified into multiple components, including water-extractable phosphorus, weakly adsorbed phosphorus, iron-aluminum bound phosphorus, calcium-bound phosphorus, and residual phosphorus, thus obtaining detailed information on the distribution of phosphorus speciation in the soil.
[0004] However, in the actual operation of existing technical solutions, researchers face a series of technical bottlenecks, which directly affect the accuracy of the test results and the efficiency and safety of the entire analytical process. The core problem stems from the physical treatment of soil samples during multi-stage extraction. After each stage of extraction and centrifugation, the supernatant needs to be separated from the soil residue, and the residue is then used for the next stage of extraction. Traditional operations often involve transferring the moist soil residue from the centrifuge tube to a new reaction vessel, or pouring the supernatant into the same container before proceeding. Due to the strong adhesiveness of moist soil particles, some sample particles inevitably adhere to and remain on the walls of the original container during pouring or transfer, resulting in physical sample loss. Although this loss may seem small in a single step, after four to five consecutive extraction stages, the accumulated sample loss becomes considerable. This leads to a significantly smaller sample volume entering the residual phosphorus digestion step compared to the initial sample volume, resulting in a systematically lower residual phosphorus determination and affecting the balance calculation of total phosphorus in the soil.
[0005] Furthermore, when processing the final soil residue to determine the most inert residual phosphorus, digestion with a strong oxidizing acid at high temperatures is usually required. While perchloric acid has extremely strong oxidizing power, direct contact with residual organic matter in the sample at high temperatures poses a safety hazard, potentially causing a violent reaction or even an explosion. To mitigate this risk, operators must carefully control the heating rate, which not only demands high operational skills but also makes the digestion process lengthy. Conversely, using less vigorous digestion conditions for safety may result in incomplete digestion of phosphorus encapsulated within stable mineral lattices, thus affecting the accuracy of the determination.
[0006] Another issue that cannot be ignored is that when determining total phosphorus in partial extracts, the dissolved organic phosphorus needs to be oxidized and digested. Existing digestion methods also suffer from cumbersome procedures and long processing times, which significantly reduces overall analytical efficiency when analyzing large batches of samples, constituting a time bottleneck in the entire detection process. Therefore, current technologies still have significant shortcomings in ensuring a balance between accuracy, operational safety, and analytical efficiency in graded detection. Summary of the Invention
[0007] To address the shortcomings of existing technologies, this invention provides a multi-component classification detection method and system for phosphorus in soil. This solves the problems of existing multi-component classification detection methods for phosphorus in soil, which involve multiple sample transfers during continuous extraction, leading to soil particle loss and affecting the accuracy of subsequent component determinations, especially residual phosphorus. At the same time, when determining organic phosphorus and residual phosphorus, there are problems such as long digestion steps and high operational safety risks.
[0008] To achieve the above objectives, the present invention provides the following technical solution: a multi-component classification detection method and system for phosphorus in soil, comprising: The first aspect of this invention provides a multi-component classification detection method for phosphorus in soil. This method utilizes an integrated reaction vessel to complete multi-step extraction and optimizes the procedures and reagents in the key digestion steps, aiming to improve the system accuracy and operational safety of the entire classification detection process.
[0009] In this embodiment, the method includes the following steps: S1. Place the soil sample in a centrifuge tube; S2. The soil sample is extracted in the centrifuge tube with at least two stages of different extractants. After each stage of extraction, solid-liquid separation is performed. The supernatant is collected and the centrifuge tube containing soil residue is retained for the next stage of extraction. S3. After completing all extraction steps, transfer the soil residue in the centrifuge tube to the digestion container; S4. Digest the soil residue in the digestion container to obtain residual phosphorus digestion solution; S5. Determine the phosphorus concentration in the supernatant collected from each extraction stage and the residual phosphorus digestion solution, respectively.
[0010] The advantage of this invention lies in the fact that by completing all extraction steps in a single centrifuge tube, it fundamentally avoids the physical loss of soil solid samples caused by multiple container transfer operations such as pouring, rinsing, and scraping in traditional methods. This ensures that the amount of soil involved in the reaction remains constant from the first step to the last, thereby significantly improving the extraction of phosphorus components at each stage, especially the final residual phosphorus. The accuracy and reliability of the measurement results.
[0011] Furthermore, to ensure the integrity of sample transfer from the extraction endpoint to the digestion starting point, in step S3, a quantitative non-destructive transfer kit is preferably used to completely transfer the soil residue from the centrifuge tube to the digestion container. This kit ensures that the initial sample amount for residual phosphorus analysis is the actual remaining amount after all extraction steps are completed.
[0012] Furthermore, to enhance the safety and thoroughness of residual phosphorus determination, a two-step digestion procedure is preferably adopted in step S4. This procedure utilizes the reaction characteristics of different acids at different temperatures: firstly, at a lower temperature, concentrated sulfuric acid is used for dehydration and carbonization to gently pretreat the more reactive organic matter in the residue, reducing the intensity of subsequent high-temperature reactions; then, the temperature is raised to a higher temperature, utilizing the strong oxidizing properties of perchloric acid to completely decompose the remaining recalcitrant organic matter and minerals after pretreatment. This programmed operation effectively avoids the risk of boiling or explosion that may be caused by direct high-temperature contact between organic matter and perchloric acid, while ensuring the complete release of the most inert phosphorus pool.
[0013] Furthermore, to address the time-consuming digestion of organophosphorus compounds in the supernatant, this invention also includes a step of digesting the supernatant containing organophosphorus compounds. This step involves adding a potassium persulfate solution and heating the solution at a specific temperature for digestion. Potassium persulfate, as a highly efficient oxidant, can rapidly convert complex organophosphorus compounds in the extract into measurable orthophosphates under these conditions, thereby shortening the sample pretreatment time.
[0014] In a preferred embodiment, the sequential extraction in step S2 includes: Extraction was performed using deionized water to obtain the water-extractable phosphorus. The supernatant; Extraction with sodium bicarbonate solution yielded a solution for determining weakly adsorbed inorganic phosphorus. and weakly adsorbed total phosphorus The supernatant; Extraction with sodium hydroxide solution yielded inorganic phosphorus for determining iron-aluminum bound states. Total phosphorus in iron-aluminum bound state The supernatant; Extraction was performed using hydrochloric acid solution to obtain phosphorus for the determination of calcium-bound phosphorus. The supernatant.
[0015] After the above extraction was completed, the remaining soil residue was digested, and the residual phosphorus content was determined. .
[0016] Among them, weakly adsorbed organic phosphorus Organophosphorus compounds bound to iron and aluminum The content is calculated by the difference method: ; ; Soil total phosphorus This is the sum of all measured components: ; A second aspect of the present invention provides a multi-component classification detection system for phosphorus in soil, which serves as the physical carrier for implementing the aforementioned method, characterized in that it comprises: An extraction module is used to contain soil samples and sequentially accept multiple extraction agents to separate the supernatant and soil residue; the extraction module includes centrifuge tubes; the design aims to complete multi-step extraction in a single container, ensuring the integrity of the sample in continuous processing through hardware.
[0017] An organophosphorus treatment module, connected to the extraction module, is used to perform organophosphorus digestion treatment on at least one received supernatant; the organophosphorus treatment module is configured to perform a potassium persulfate solution digestion procedure; this module aims to achieve efficient and rapid digestion of a specific supernatant.
[0018] The residual phosphorus treatment module, connected to the extraction module, is used to digest the received soil residue. The residual phosphorus treatment module includes a quantitative non-destructive transfer kit and is configured to perform a two-step digestion procedure. This module integrates two major functions: non-destructive transfer and safe digestion, and is used to accurately and safely determine the most difficult-to-decompose phosphorus components.
[0019] The detection module is connected to the extraction module, the organophosphorus treatment module and the residual phosphorus treatment module respectively, and is used to determine the phosphorus concentration in each liquid after treatment.
[0020] This system provides a systematic solution by solidifying optimized chemical methods and procedures into a functional hardware suite through the collaborative work of its various modules.
[0021] This invention provides a method and system for the multi-component classification and detection of phosphorus in soil. It has the following beneficial effects: 1. This invention utilizes centrifuge tubes to continuously complete multi-stage extraction steps of phosphorus from water-extractable to calcium-bound states in a single container. This fundamentally avoids the physical loss of solid particles caused by repeated pouring, rinsing, and scraping of samples between different containers in traditional methods. This ensures a constant amount of soil participating in each stage of the reaction, guarantees material conservation throughout the process, and thus significantly improves the system accuracy of phosphorus component content calculation and the reliability of the final results.
[0022] 2. This invention solves the technical problem of the difficulty in completely transferring moist residue due to its strong adhesion by setting up a quantitative non-destructive transfer kit for transferring soil residue from centrifuge tubes to digestion containers after all extraction steps are completed. The kit ensures the integrity of the sample to be digested through the synergistic effect of mechanical scraping and micro-rinsing, so that the final measured residual phosphorus content can truly reflect the amount of phosphorus remaining after all previous components are extracted, providing a key guarantee for the accurate closure of the entire phosphorus balance calculation.
[0023] 3. This invention employs a two-step digestion process for residual phosphorus: first, pre-digestion with concentrated sulfuric acid at a lower temperature to carbonize the organic matter; then, heating to a higher temperature for complete oxidation with perchloric acid. This achieves programmed and safe control of the strong acid digestion process. This staged treatment method effectively avoids the risk of boiling or explosion that may be caused by direct contact between residual organic matter in the sample and high-temperature strong oxidants. While ensuring the complete decomposition of the most inert phosphorus pool, it significantly improves the safety of experimental operations. Attached Figure Description
[0024] Figure 1 This is a system architecture diagram of the present invention; Figure 2 This is a flowchart of the method of the present invention. Detailed Implementation
[0025] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.
[0026] Example: Please see the appendix Figure 2 This invention provides a method for multi-component classification and detection of phosphorus in soil, comprising the following steps: S1. Place the soil sample in a centrifuge tube; In this embodiment, in order to achieve accurate separation and determination of water-extractable phosphorus components in soil samples, and to lay an accurate material basis for all subsequent continuous extraction steps, the specific operation method of step S1, the key devices used, and its underlying technical principles will be described in more detail below.
[0027] The initial step of this process involves a series of standardized pretreatment procedures on the collected raw soil samples. Preferably, the soil samples collected in the field are placed in a clean, well-ventilated environment for natural air drying. This effectively terminates the life activities of microorganisms in the soil, preventing them from continuing to transform phosphorus forms during sample storage and processing, thus freezing the chemical state of the sample at the time of collection. The air-dried samples are then ground and passed through a sieve with a specific aperture. This pretreatment process aims to break the soil into uniform, fine particles. This not only greatly improves the homogeneity of the sample, ensuring that subsequent small-scale samples highly represent the average chemical composition of the entire batch, but also significantly increases the specific surface area of the soil particles. This facilitates sufficient contact between the solvent and sample particles during subsequent extraction, thereby improving extraction efficiency.
[0028] After pretreatment, a certain mass of homogeneous soil sample was accurately weighed using a 0.01 g analytical balance and carefully placed into a centrifuge tube, which serves as the core carrier of the method of this invention. This centrifuge tube is not a conventional centrifuge tube used in this field, but is specifically designed to fundamentally address the core problem in existing technologies: physical loss and cross-contamination caused by multiple transfers of samples between different containers.
[0029] Specifically, the centrifuge tube possesses the following key technical features that have undergone deep optimization: its inner wall surface is treated with a special chemical inerting process, forming a dense, smooth, and chemically stable low-phosphorus adsorption coating. Phosphate ions exhibit a strong surface adsorption tendency in aqueous solutions, especially at low concentrations. Conventional glass or polypropylene containers have numerous polar groups or microscopic defects on their surfaces, which can adsorb some of the analyte phosphate ions through electrostatic attraction, hydrogen bonding, or coordination bonding mechanisms. This wall adhesion phenomenon is a significant source of error in analytical chemistry. The low-phosphorus adsorption coating used in this embodiment works by effectively sealing or shielding the active adsorption sites of the tube wall material through surface modification technology, significantly reducing its surface energy. This makes it difficult for phosphate ions to remain on the tube wall, ensuring that the analytes in the solution can be completely separated and measured.
[0030] Furthermore, the material of the centrifuge tube itself has undergone rigorous screening and optimization to ensure excellent chemical resistance throughout the entire process. Considering that this reaction tube is the only physical carrier throughout all extraction steps from S1 to S2, it must be able to stably withstand prolonged chemical erosion from the initial neutral deionized water, to the strongly alkaline sodium hydroxide solution in subsequent steps, and then to the strongly acidic hydrochloric acid solution, without swelling, cracking, material degradation, or precipitation of any chemicals that might interfere with subsequent colorimetric determinations. Preferably, the tube body can be made of high-performance polymers such as modified polypropylene, polytetrafluoroethylene, or perfluoroalkoxyalkanes. This consistent chemical stability is the physical basis for realizing the integrated extraction concept. Simultaneously, the preferred geometry of the tube is a conical bottom, which facilitates the efficient and compact collection of soil particles into small precipitate clusters during centrifugation, making it easier to absorb the supernatant.
[0031] After the sample is placed in the tube, a precise volume of phosphorus-free deionized water is added as the extraction solvent. The tube is then tightly sealed with the matching cap to prevent changes in the volume of the extract due to solvent evaporation during the several-hour shaking process, which would affect the final concentration calculation. The sealed reaction tube is then placed on a temperature-controlled shaking device and subjected to prolonged, thorough shaking at a standard temperature simulating the soil environment, for example, continuously shaking for 16 hours. The principle behind this step is to utilize water, the gentlest extraction solvent that does not disrupt the soil aggregate structure, to slowly desorb and dissolve the pre-existing free phosphate ions in the soil solution, as well as those phosphate ions adsorbed only on the surface of soil particles through the weakest physical adsorption or van der Waals forces, into the aqueous phase. The extracted water-extractable phosphorus (H2O-P) represents the most active, mobile, and readily absorbed component of the soil phosphorus pool by plant roots. Its content directly affects the phosphorus nutrient status of the current crop and is also a key indicator for assessing the environmental risks posed by soil phosphorus entering water bodies through surface runoff or leaching. The 16-hour oscillation time aims to ensure that this slow desorption process approaches equilibrium as closely as possible, guaranteeing sufficient extraction and reproducibility of results.
[0032] After the oscillation process is complete, the key advantages of the method of this invention are fully demonstrated: the operator does not need to perform any pouring or transfer operations, but simply removes the centrifuge tube containing the soil suspension directly from the shaker and places it into a high-speed centrifuge for solid-liquid separation. Sufficient centrifugal force ensures that most solid particles, including fine clay particles, are tightly compacted at the bottom cone of the tube, forming a clear and stable solid-liquid interface.
[0033] After centrifugation, carefully use a pipette to aspirate the clear supernatant and filter it through a microporous membrane, preferably with a pore size of 0.45 μm. This filtration step is crucial for ensuring the purity of the test liquid, aiming to thoroughly remove any extremely fine soil particles or colloidal substances that may still be suspended in the supernatant and have not been completely settled by centrifugation. If these particles are not removed, they may interfere with accurate absorbance readings in subsequent colorimetric analysis by causing light scattering, and may also continue to slowly release phosphorus during storage, causing the measurement results to become inaccurate over time. The clear filtrate obtained after filtration is collected in a clean storage tube as the final test solution for the final determination of water-extractable phosphorus content.
[0034] It is worth emphasizing again that after the core operation in step S1 is completed, the same centrifuge tube containing all the soil residue is directly and completely preserved, seamlessly entering the subsequent step S2 for the extraction of weakly adsorbed phosphorus. This closed-loop operation mode, which completes the feeding-extraction-separation-residue retention within a single sealed container, constitutes the essence of the integrated concept of this invention. It physically eliminates sample transfer loss during the switching between different steps, ensuring the quality of the soil sample entering the next extraction step. The sample quality is completely consistent with the initial sample quality in both theory and practice, thus providing the most fundamental guarantee for the system accuracy of the entire multi-level grading detection method.
[0035] Finally, the concentration of water-extractable phosphorus in the test solution was determined by colorimetric analysis in subsequent steps. The final content of water-extractable phosphorus in the soil sample was calculated using the following formula. : ; In the formula: The content of water-extractable phosphorus in soil samples is usually reported in milligrams per kilogram; The concentration of phosphorus in the filtrate was determined by spectrophotometry, in milligrams per liter. This refers to the precise volume of the extractant added in this step, in liters. The mass of the initially weighed, pretreated, dried soil sample is expressed in kilograms.
[0036] S2. The soil sample is extracted in centrifuge tubes with at least two different extractants. After each extraction, solid-liquid separation is performed. The supernatant is collected and the centrifuge tube containing soil residue is retained for the next extraction. In this embodiment, after the most active water-extractable phosphorus is precisely separated in step S1, step S2 takes the soil residue left from the previous step and, in the same physical carrier, uses a series of extractants with progressively stronger chemical properties and mechanisms of action to continuously and hierarchically separate multiple phosphorus components in the soil with progressively increasing chemical stability. This step is the core of the entire method, and its design essence lies in integrating the complex process that originally required multiple sample transfers into a single centrifuge tube, thereby achieving streamlined operation and precise results.
[0037] This step begins with the centrifuge tube containing all the soil residue obtained in step S1. Importantly, after the supernatant from the previous step is aspirated and separated, the tube and the moist soil residue inside it do not require any washing or drying and proceed directly to the first stage of extraction in this step, ensuring absolute continuity of material flow.
[0038] First, the extraction of weakly adsorbed phosphorus is performed. A specific volume of sodium bicarbonate solution is precisely added to the tube. In a preferred embodiment, the concentration of the solution used is 0.5 mol / L, and its pH value is precisely adjusted to 8.5 beforehand using sodium hydroxide or dilute acid. The design principle of this step is extremely ingenious, utilizing the synergistic effect of several chemical equilibria: firstly, a higher concentration of bicarbonate ions can competitively exchange and adsorb phosphate ions adsorbed on the surface of soil colloids in the form of electrostatic attraction or weak chemical bonding. According to Le Chatelier's principle, a high concentration of HCO3-... - Firstly, the adsorption equilibrium is driven towards desorption, thereby displacing this portion of phosphorus into the solution phase. Secondly, the weakly alkaline environment of pH 8.5 is maintained by a carbonate buffer system, the equilibrium of which is determined by the second-order dissociation constant of carbonic acid, providing a stable pH environment to ensure the reproducibility of extraction conditions. This pH condition also effectively inhibits the activity of calcium ions in the soil, preventing secondary precipitation of calcium phosphate during extraction, while avoiding significant dissolution of iron and aluminum oxides, thus clearly defining the chemical boundary between weakly adsorbed phosphorus and subsequent iron-aluminum bound phosphorus.
[0039] After adding sodium bicarbonate extractant, the standardized mechanical operation in step S1 is repeated again, i.e., the centrifuge tube is tightly capped and shaken for a long time on a constant-temperature shaker to reach adsorption-desorption chemical equilibrium. Subsequently, the tube is placed directly in a centrifuge for high-speed centrifugation to complete solid-liquid separation. The supernatant obtained after separation, due to its alkaline extraction, usually contains a large amount of dissolved organic matter and is dark in color. Therefore, after filtration through a 0.45 μm microporous membrane, the filtrate needs to be fractionated: one fraction is used directly in subsequent steps to determine the inorganic phosphorus content of this fraction. The other fraction is reserved for subsequent organophosphorus digestion in step S3, to determine the total phosphorus content of that fraction. After the supernatant was collected, the centrifuge tube containing all soil residue was retained for the next stage of more intense extraction.
[0040] Next, iron-aluminum bound phosphorus was extracted from the soil residue in the same tube where sodium bicarbonate extraction had already been performed. A specific volume of sodium hydroxide solution was added to the tube, preferably a 0.1 mol / L solution. The principle behind this step is that higher concentrations of hydroxide ions are stronger ligands than phosphate ions, and can effectively replace phosphate ions that are specifically adsorbed on the hydroxyl groups of amorphous or hydrated iron and aluminum oxides in the soil through a ligand exchange mechanism. Simultaneously, the strongly alkaline environment at this concentration can selectively dissolve a portion of the chemically active and poorly crystallized iron and aluminum oxide colloids, thereby releasing the phosphorus co-precipitated or physically encapsulated therein. This component represents a phosphorus pool with significantly enhanced chemical stability in the soil. Similarly, the shaking and centrifugation operations were repeated, and the resulting supernatant of the darker-colored sodium hydroxide extract was subjected to the same sample processing: one portion was directly used for the determination of iron-aluminum bound inorganic phosphorus. Another sample is reserved for digestion in subsequent step S3 to determine the total phosphorus bound to iron and aluminum. .
[0041] After two stages of alkaline extraction, a final stage of solution extraction, namely the extraction of calcium-bound phosphorus, is performed. A specific volume of hydrochloric acid solution, preferably a 1.0 mol / L solution, is added to the tube. The chemical principle is that a strongly acidic environment provides a large number of hydrogen ions, thereby effectively dissolving phosphorus in the soil in the form of various calcium phosphate minerals. These minerals, such as different forms of apatite, are extremely stable under neutral or alkaline conditions, but their solubility increases sharply as the pH decreases. The strong acid, through an acid-base reaction with phosphate ions, greatly reduces the activity of phosphate ions in the solution, thereby shifting the dissolution equilibrium of calcium phosphate minerals to the right. This component is the main inorganic phosphorus pool in neutral and calcareous soils. After shaking and centrifugation, the resulting acidic supernatant is generally considered to contain mainly inorganic phosphorus. Because organic matter has low solubility in strong acid, this extract usually does not require digestion of organic phosphorus and can be directly used for subsequent determination to obtain calcium-bound phosphorus. The content of.
[0042] At this point, all the sequential extraction sub-steps based on chemical reagents in step S2 have been completed sequentially in a single centrifuge tube. The remaining soil residue in this tube is the final sample prepared for non-destructive transfer in step S4 and ultimately digested in step S5.
[0043] Through the above series of rigorous and continuous extraction operations, data for calculating the content of different forms of phosphorus can be obtained. The calculation formula and necessary limitations are as follows: The calculation of phosphorus content in each fraction follows the basic principle of multiplying the concentration of phosphorus in the extract measured by colorimetry by the volume of solvent used in that fraction extraction, and then dividing by the mass of the initial soil sample.
[0044] In the calculation of weakly adsorbed phosphorus content, its organic phosphorus content... It is not measured directly, but obtained through the difference method, and the necessary calculation relationship is as follows: ; in, and Calculated using the following formula: ; ; Similarly, in calculating the content of iron-aluminum bound phosphorus, its organic phosphorus content... It must also be obtained through the difference method, and the calculation relationship is limited as follows: ; in, and Calculated using the following formula: ; ; Calcium-bound phosphorus content The results are then calculated directly from the extract: ; In all the above formulas, the symbols are defined and restricted as follows: These represent the final content of each corresponding form of phosphorus in the soil sample, in milligrams per kilogram; These are the mass concentrations of phosphorus in the extract to be tested, read from the standard curve method during the determination of each phosphorus form, in milligrams per liter. This refers to the precise volume of extractant added for each stage of extraction in this step, in liters. The mass of the pretreated, dried soil sample weighed at the beginning of step S1 is expressed in kilograms.
[0045] S3. After completing all extraction steps, transfer the soil residue in the centrifuge tube to the digestion container; In this embodiment, step S3 is a chemical transformation process specifically targeting a particular extract component. Its core technical objective is to completely convert the phosphorus component extracted along with inorganic phosphorus in the preceding steps—which exists stably in an organic form and cannot be directly responded to by the conventional molybdenum blue colorimetric method—into an accurately quantifiable inorganic orthophosphate form through an efficient and controllable oxidation-digestion process. This step is a crucial step in achieving accurate quantification of the soil organic phosphorus pool.
[0046] Specifically, this step does not process the entire extract, but selectively targets the two supernatants obtained in step S2 using alkaline extractants, namely sodium bicarbonate solution and sodium hydroxide solution. The rationale for this selection is that an alkaline environment, particularly the strongly alkaline environment created by sodium hydroxide, not only desorbs and dissolves some inorganic phosphorus, but also efficiently dissolves and extracts large amounts of humic substances and other organic matter from the soil. During this process, various organophosphorus compounds, such as phytic acid phosphorus, nucleic acid phosphorus, phospholipids, and other phosphate esters, which are tightly bound to these organic macromolecules or are themselves organic molecules, are also transferred in large quantities to the liquid extract phase. These organophosphorus compounds are important, potentially bioavailable components of the soil phosphorus pool, and their accurate quantification is crucial for comprehensively evaluating the long-term phosphorus supply potential of the soil and for a deeper understanding of the biogeochemical cycling mechanism of phosphorus.
[0047] To this end, embodiments of the present invention provide a specific and optimized wet digestion scheme for organophosphorus compounds. This scheme abandons the complex operation of some traditional methods that require the use of multiple strong acids mixed for heating and are difficult to control. Instead, it uses potassium persulfate solution as a single core oxidant and carries out the reaction under specific, easily standardized high-temperature and high-pressure hydrothermal conditions to achieve efficient and complete conversion of trace organophosphorus compounds in complex matrices.
[0048] In a preferred embodiment, the detailed operation procedure of this step is as follows: First, a certain volume of liquid is precisely transferred from the sodium bicarbonate or sodium hydroxide extract to be tested, derived from step S2, using a calibrated pipette, and placed in a clean, dedicated digestion tube capable of withstanding subsequent high-temperature and high-pressure conditions. Preferably, the digestion tube is made of thick-walled borosilicate glass or polytetrafluoroethylene, etc. Subsequently, a potassium persulfate solution of a specific concentration is added to the digestion tube. The amount of oxidant added is experimentally optimized to ensure sufficient oxidant to completely destroy any organic matter in the sample that may be present in a wide range of concentrations.
[0049] The digestion tubes containing the added oxidant are thoroughly shaken to ensure homogeneous mixing of their contents, and then sealed tightly with the matching screw caps to ensure a closed reaction system during subsequent heating and pressurization. This batch of digestion tubes is then placed in an autoclave. The equipment is started, and the reaction conditions are set at a specific temperature and the corresponding pressure determined by the saturated steam at that temperature. Preferably, the target temperature is 121°C. At this temperature, the pressure inside the chamber will automatically reach a gauge pressure of approximately 103 kPa. This condition not only provides a uniform, stable, and efficient heat input for the chemical reaction but also effectively raises the boiling point of water molecules, ensuring the reaction proceeds stably in the liquid phase. Under these conditions, the heating and digestion are maintained for a programmed time, for example, 30 minutes.
[0050] The core chemical principle employed in this step lies in the fact that potassium persulfate is a strong oxidizing agent, and its molecular structure contains peroxy bonds. Under high-temperature and high-pressure hydrothermal conditions of 121°C, these peroxy bonds undergo homolytic cleavage, decomposing to produce sulfate free radicals with extremely high oxidizing activity. These sulfate free radicals are even stronger and more active oxidizing agents than the persulfate ion itself, possessing extremely high standard redox potentials. Once generated, these free radicals indiscriminately and rapidly attack the diverse and complex organophosphate molecules in the soil extract. Whether it's nucleic acids and phospholipids linked by phosphoester bonds, or phytate phosphorus present through phosphate ester bonds in inositol hexaphosphate, their normally stable chemical bonds are effectively broken under the intense attack of sulfate free radicals. Through a series of complex chain-like free radical reactions, the carbon elements in the organic molecular skeleton are ultimately oxidized to carbon dioxide, the hydrogen elements to water, and the phosphorus elements bound in the organic structure are completely released and converted into the most stable and simplest inorganic orthophosphate form. This process is mineralization.
[0051] Compared to other methods, the specific combination disclosed in this embodiment, which uses potassium persulfate solution and digestion under high pressure at 121°C, provides an efficient, safe and reproducible solution for treating soil alkaline extract matrix with complex composition and severe background interference.
[0052] After the digestion process is complete, wait for the autoclave cycle to finish and the pressure and temperature inside the chamber to automatically drop to a safe range before opening the door, removing the digestion tube, and allowing it to cool naturally at room temperature. The solution inside the tube at this point is the converted test solution. The significance of this test solution lies in the fact that the phosphorus it contains is the sum of the initial inorganic phosphorus and initial organic phosphorus converted from the original extract sample. Therefore, this solution can be directly used for colorimetric determination in subsequent steps to obtain the total phosphorus concentration of this extraction fraction. or .
[0053] Step S3 itself does not directly produce a calculation formula, but it provides crucial experimental data for subsequent calculations of organophosphorus content. It enables the following conceptual formula, expressed in words, to hold true: the organophosphorus concentration of a certain extraction fraction is equal to the total phosphorus concentration measured after treatment in step S3 minus the inorganic phosphorus concentration directly measured without treatment in this step. These concentration values achieved in this step are essential for subsequent calculations of weakly adsorbed organophosphorus content. content of organophosphorus compounds bound to iron and aluminum An indispensable data foundation provides the necessary technical support for a comprehensive and accurate analysis of the intrinsic composition of soil phosphorus speciation.
[0054] S4. Digest the soil residue in the digestion container to obtain residual phosphorus digestion solution; In this embodiment, step S4 plays a crucial physical transition role in the entire multi-component fractionation detection method. It is both the endpoint of all preceding chemical extraction steps and the starting point of the subsequent final destructive digestion step. The core technical objective of this step is to achieve a quantitative, non-destructive, and clean transfer of soil residue from the centrifuge tube, which serves as the extraction carrier, to a specialized digestion container. The rigor and effectiveness of this step directly determine the accuracy of the final measured residual phosphorus data and have a decisive impact on the quality balance closure of the entire phosphorus fractionation system.
[0055] Specifically, after completing the entire continuous extraction process of water, sodium bicarbonate, sodium hydroxide, and hydrochloric acid covered in step S2, the initial soil sample matrix has undergone multiple rounds of chemical action and physical separation. At this point, the moist solid remaining at the bottom of the centrifuge tube is the soil residue. This residue is the most chemically stable component of the soil, mainly composed of acid- and alkali-resistant primary minerals, highly crystalline secondary silicate minerals, and organic matter tightly cemented to the minerals with a highly resilient structure. Phosphorus, deeply embedded or encased within these substances, or existing in a lattice substitution form, is the residual phosphorus that the method of this invention aims to ultimately quantify.
[0056] Since the determination of residual phosphorus requires the extreme chemical conditions employed in step S5—namely, a high-temperature and highly corrosive mixed acid system—and the centrifuge tubes used as extraction containers, despite their excellent chemical stability against acid and alkali extractants, are not designed in terms of materials and structure to withstand such extreme high-temperature and strong oxidative digestion conditions, it is logically and practically necessary to completely transfer this portion of soil residue from the centrifuge tubes and place it into a specially designed container capable of safely withstanding high-temperature and strong acid digestion, such as an Erlenmeyer flask or Kjeldahl flask made of borosilicate glass.
[0057] However, this transfer process itself presents significant technical challenges. Traditional transfer methods, such as simple pouring, rinsing, or scraping with a common spatula, have inherent drawbacks that are difficult to overcome. Moist soil residue, especially when it contains a large amount of clay minerals, is highly adhesive and forms a thin film that adheres tightly to the inner wall and conical bottom of the centrifuge tube. Simply pouring and tapping is far from sufficient to completely remove it. If large amounts of liquid are used for repeated rinsing in pursuit of thorough transfer, it will have a series of negative impacts on the subsequent step S5: excessive water will greatly dilute the concentrated acid, reducing the initial digestion efficiency; in the early stages of heating, the evaporation of a large amount of water will significantly prolong the entire digestion process and may cause acid splashing due to violent boiling, posing a serious safety hazard. Most importantly, any residue particles that are not successfully transferred represent a direct loss of the analyte, which will directly lead to a systematic and irreversible underestimation of the final measured residual phosphorus content, thus introducing a systematic error that is difficult to assess and correct.
[0058] To fundamentally overcome the aforementioned technical challenges, this invention provides and employs a specially designed quantitative non-destructive transfer kit. This kit is the core physical tool for achieving the technical objective of this step; its ingenious design lies in achieving a near-perfect transfer effect through the synergistic effect of mechanics and micro-fluids. In a preferred embodiment, the kit includes two key functional components: Firstly, there is a specialized scraper precisely fitted to the internal geometry of the centrifuge tubes used. Preferably, the scraper head is made of polytetrafluoroethylene (PTFE), a chemically extremely stable material with very low surface energy. PTFE is known for its extremely low coefficient of friction and excellent anti-adhesion properties, ensuring that soil residue does not adhere to the scraper during the scraping process. The scraper head is meticulously ergonomically and hydrodynamically designed to fit snugly against the cylindrical inner wall and conical bottom of the centrifuge tube. With a slight mechanical force applied by the operator, it effectively scrapes and peels off almost all soil residue particles adhering to the tube wall, collecting them at the bottom of the tube.
[0059] Secondly, there is a specially designed miniature anti-splash funnel used to assist in micro-rinsing. The lower end of the funnel is designed to be long and thin, so that it can easily reach into the neck of the receiving container, thus preventing the rinsing liquid from wetting the inner wall of the bottle mouth; while the upper end of the funnel mouth is designed to be wide enough to completely cover and seal the mouth of the centrifuge tube, forming a semi-closed rinsing system.
[0060] The specific operation procedure of step S4 is standardized as follows: First, the operator holds the special scraper and inserts it into the centrifuge tube, rotating and moving it up and down along the inner wall of the tube, focusing on scraping the conical bottom carefully to remove all visible moist soil residue from the tube wall and collect it at the bottom. Next, a miniature splash-proof funnel is placed at the mouth of a pre-prepared, clean, and dry conical flask, and the scraped centrifuge tube is inverted on top of the funnel. A gentle tap on the outside of the tube allows most of the scraped, clump-like residue to fall directly into the conical flask due to gravity. Finally, a very small amount of deionized water is drawn using a pipette and used to quickly rotate and rinse the inner wall of the centrifuge tube 1 to 2 times through the funnel. This rinsing solution, containing the last trace residue, is collected completely into the conical flask. The miniature funnel serves a triple purpose in this process: guiding the liquid flow, preventing splashing during rinsing, and flushing away any particles that may be adhering to the tube opening.
[0061] This step itself does not involve chemical reactions, therefore there are no stoichiometric calculation formulas. However, its ultimate purpose is to ensure that the important mass conservation relationship holds physically, which can be expressed in words as follows: the dry weight of the soil residue successfully transferred to the digestion container should theoretically be exactly equal to the initial soil sample dry weight minus the total dry weight of substances dissolved and removed in all previous extraction steps. Through the combination of mechanical scraping as the main method and micro-rinsing as a supplement, this step can ensure that this mass conservation relationship is realized in actual operation to the greatest extent, ensuring that the amount of sample to be digested is the true final residual amount after all extraction steps. This provides a real, complete, and reliable material premise for the accurate quantification of residual phosphorus in step S5, perfectly completing the precise transition from the integrated extraction stage to the independent digestion stage.
[0062] S5. Determine the phosphorus concentration in the supernatant and residual phosphorus digest collected at each extraction stage; In this embodiment, step S5 is the final chemical processing step in the entire multi-component grading and detection method, holding a strategic position of endpoint and conclusion in the entire analytical chain. The core technology of this step lies in applying a highly optimized and programmed strong acid digestion strategy to the final residue, which represents the most inert component of the soil and has been accurately quantified and transferred in step S4, to ensure that the phosphorus component, i.e., the residual phosphorus, which exists in the most stable form, can be completely and thoroughly destroyed and converted into an inorganic orthophosphate form that can be detected by modern analytical instruments.
[0063] Specifically, this step involves handling all soil residues and a small amount of wetting solution used for transfer, stored in a specialized digestion container. The chemical composition of this residue is extremely stubborn, and traditional digestion methods often face a dilemma: if the conditions are mild, it is difficult to completely destroy the crystal structure of phosphorus-containing primary minerals such as apatite and zircon, or the phosphorus tightly complexed with highly humified organic matter, leading to systematically low test results; if the conditions are too aggressive, especially when processing samples containing organic matter, strong oxidizing acids can easily trigger violent and uncontrollable exothermic reactions when in direct contact with organic matter at high temperatures, even leading to explosions and posing serious safety hazards.
[0064] To address the core conflict between complete digestion and operational safety, this invention proposes and employs a specially designed two-step digestion procedure. This procedure is not merely a simple combination of reagents and temperature, but a systematic solution incorporating chemical principles, programmed control, and safety engineering considerations.
[0065] In a preferred embodiment, the detailed implementation of the two-step digestion procedure is as follows: First, two strong acids are carefully added to the digestion container containing the soil residue in a specific order and volume. Preferably, 5 mL of concentrated sulfuric acid is slowly added to the container first, and the container is gently shaken to initially wet the residue with the acid. Then, 1 mL of perchloric acid is added. This order of addition is not arbitrary but an optimization based on the different physicochemical properties of the two acids. Concentrated sulfuric acid has a high boiling point, providing a high-temperature reaction medium for the entire digestion system; while the oxidizing power of perchloric acid is strongly temperature-dependent and relatively mild at room temperature. To prevent violent boiling and aerosol splashing that may occur during heating, and to provide some condensation and reflux to reduce acid volatilization loss during prolonged heating and maintain a relatively constant acid concentration within the reaction system, a small glass funnel can be placed at the mouth of the digestion container.
[0066] Subsequently, the pre-treated digestion vessel assembly is placed on a heating device that can be precisely programmed, such as a CNC graphite digester or a CNC hot plate, and the pre-programmed two-step digestion program is started.
[0067] The first stage of the process is defined as the low-temperature pre-digestion and organic matter carbonization stage. In this stage, the program controls the heating device to set the temperature at a relatively low and constant level, preferably 150°C, and maintains this temperature precisely for a period of time. The chemical principle of this stage lies in utilizing the dual properties of concentrated sulfuric acid at this temperature as both an effective dehydrating agent and a moderately strong oxidizing agent. It can gently act on the remaining, relatively complex organic macromolecules in the residue, causing them to undergo dehydration and incomplete oxidation, ultimately carbonizing them into simpler, less chemically reactive elemental carbon particles. This process is relatively gentle, mainly releasing gases such as water vapor and carbon dioxide, effectively avoiding the dangerous, instantaneous, and massive gas-generating reaction that could occur if highly reactive organic matter directly contacts the subsequent strong oxidizing agent at high temperatures. This pretreatment step completely eliminates the main safety hazards for the subsequent high-temperature, strong oxidation steps and is the cornerstone of the program's safety design.
[0068] The second stage of the process is defined as the high-temperature complete oxidation and digestion stage. After the low-temperature carbonization process in the first stage, the program-controlled heating device automatically and slowly raises the temperature linearly from 150°C to a high temperature sufficient to activate the powerful oxidizing ability of perchloric acid, preferably 350°C. This slow and steady temperature increase is another important safety measure, ensuring that any trace amounts of potentially remaining active organic matter in the system have a gradual reaction process, rather than being suddenly exposed to extreme temperatures. At this high temperature of 350°C, the oxidizing property of perchloric acid is fully activated, becoming an extremely powerful oxidant. The principle is that perchloric acid at high temperatures can thoroughly and indiscriminately attack and destroy the extremely stubborn organic residues remaining after the first stage of carbonization, while also effectively decomposing phosphorus-containing primary or secondary minerals that are still difficult to dissolve under acidic conditions. The color of the entire system will undergo characteristic changes as the organic matter and carbon particles are continuously oxidized, serving as a process indicator. It typically changes from an initial dark brown to a clear and transparent color or a slightly yellowish color due to residual trace metal ions. To ensure the absolute completeness of the digestion process, the procedure is set to maintain this high temperature and continue boiling for another 20 minutes after the system solution has clarified. This digestion step is designed to ensure that even phosphorus encased in the core of the most inert mineral particles has sufficient time to be penetrated and dissolved by the acid.
[0069] Through the sophisticated programmed control of the above-mentioned low-temperature carbonization and passivation followed by high-temperature strong oxidation, this step ensures that the most inert phosphorus pool in the soil is completely converted into soluble orthophosphate, while minimizing the operational risks of the entire strong acid digestion process, achieving a high degree of balance between safety and effectiveness.
[0070] After the digestion process is complete, the digestion container and its high-concentration acidic solution are completely cooled to room temperature in a fume hood before subsequent quantitative transfer and dilution. Carefully and repeatedly rinse the inner walls of the container and the small funnel covering the bottle opening with a small amount of deionized water, ensuring that all digestion solution adhering to the container walls is washed off. Transfer all the rinsing solution together and without omission to a clean volumetric flask of a specific volume, and finally dilute to the mark with deionized water. Stopper the flask, invert and shake repeatedly to mix, and let it stand until ready for use. This solution contains all residual phosphorus, has a clear background, and can be directly used for the final colorimetric determination.
[0071] The final result of this step is to provide a reliable sample for calculating the content of residual phosphorus. The calculation must be limited according to the following formula: ; In the formula: The final content of residual phosphorus in the soil sample is reported in milligrams per kilogram (mg / kg). The mass concentration of phosphorus in the test solution is measured by colorimetry in subsequent steps and by referring to a standard curve, in milligrams per liter. This refers to the total fixed volume of the residue digestate after final quantitative transfer in this step, in liters. The initial mass, expressed in kilograms, is the precise weight of the pretreated, dried soil sample taken at the beginning of step S1. The initial mass is used here. The rigor of its logic is based on the premise that step S4 has ensured the non-destructive transfer of the residue.
[0072] Please see the appendix Figure 1 Another embodiment of the present invention discloses a multi-component graded detection system for phosphorus in soil, comprising: The extraction module is used to hold soil samples and sequentially accept multiple extraction reagents to separate the supernatant and soil residue; the extraction module includes centrifuge tubes; An organophosphorus treatment module, connected to the extraction module, is used to perform organophosphorus digestion treatment on at least one received supernatant; the organophosphorus treatment module is configured to execute a potassium persulfate solution digestion program; The residual phosphorus treatment module, connected to the extraction module, is used to digest the received soil residue; the residual phosphorus treatment module includes a quantitative non-destructive transfer kit and is configured to perform a two-step digestion procedure; The detection module is connected to the extraction module, the organophosphorus treatment module, and the residual phosphorus treatment module, respectively, and is used to determine the phosphorus concentration in each of the treated liquids.
Claims
1. A multi-component graded detection method for phosphorus in soil, characterized in that, Includes the following steps: S1. Place the soil sample in a centrifuge tube; S2. The soil sample is extracted in the centrifuge tube with at least two stages of different extractants. After each stage of extraction, solid-liquid separation is performed. The supernatant is collected and the centrifuge tube containing soil residue is retained for the next stage of extraction. S3. After completing all extraction steps, transfer the soil residue in the centrifuge tube to the digestion container; S4. Digest the soil residue in the digestion container to obtain residual phosphorus digestion solution; S5. Determine the phosphorus concentration in the supernatant collected from each extraction stage and the residual phosphorus digestion solution, respectively.
2. The method for multi-component graded detection of phosphorus in soil according to claim 1, characterized in that, Step S3 specifically involves using a quantitative non-destructive transfer kit to transfer the soil residue from the centrifuge tube to the digestion container.
3. The method for multi-component graded detection of phosphorus in soil according to claim 1, characterized in that, The digestion process in step S4 is a two-step digestion procedure, which includes: first, pre-digesting the soil residue at a lower temperature, and then raising the temperature to a higher temperature for complete digestion.
4. The method for multi-component graded detection of phosphorus in soil according to claim 1, characterized in that, It also includes the step of digesting at least one of the supernatants with organophosphorus compounds, the organophosphorus digestion step comprising: adding potassium persulfate solution to the supernatant and heating for digestion at a specific temperature.
5. The method for multi-component graded detection of phosphorus in soil according to claim 4, characterized in that, The step of digesting at least one of the supernatants with organophosphorus compounds is applied to the supernatants obtained by extraction with sodium bicarbonate solution and sodium hydroxide solution, so as to determine the weakly adsorbed total phosphorus and the iron-aluminum bound total phosphorus, respectively.
6. The method for multi-component graded detection of phosphorus in soil according to claim 1, characterized in that, The sequential extraction in step S2 includes: sequentially extracting with deionized water, sodium bicarbonate solution, sodium hydroxide solution, and hydrochloric acid solution.
7. The method for multi-component graded detection of phosphorus in soil according to claim 3, characterized in that, The two-step digestion process is carried out by adding concentrated sulfuric acid and perchloric acid to the digestion container.
8. The method for multi-component graded detection of phosphorus in soil according to claim 7, characterized in that, The lower temperature is 150°C, and the higher temperature is 350°C.
9. The method for multi-component graded detection of phosphorus in soil according to claim 4, characterized in that, In the organophosphorus digestion step, the specific temperature is 121°C.
10. A multi-component classification and detection system for phosphorus in soil, comprising a method for multi-component classification and detection of phosphorus in soil according to any one of claims 1-9, characterized in that, include: The extraction module is used to hold soil samples and sequentially receive multiple extractants to separate the supernatant and soil residue; The extraction module includes centrifuge tubes; An organophosphorus treatment module, connected to the extraction module, is used to perform organophosphorus digestion treatment on at least one received supernatant; the organophosphorus treatment module is configured to execute a potassium persulfate solution digestion program; A residual phosphorus treatment module, connected to the extraction module, is used to digest the received soil residue; the residual phosphorus treatment module includes a quantitative non-destructive transfer kit and is configured to perform a two-step digestion procedure; The detection module is connected to the extraction module, the organophosphorus treatment module and the residual phosphorus treatment module respectively, and is used to determine the phosphorus concentration in each liquid after treatment.