Soil respiration multi-channel real-time analysis method based on carbon dioxide sensor

Abstract

A soil respiration multi-channel real-time analysis method based on a carbon dioxide sensor belongs to the technical field of soil respiration detection, and comprises the following steps: (a) placing a soil sample subjected to standardization treatment in a transparent container, the standardization treatment comprising: controlling the soil water content to be 40%-60% of the maximum soil water-holding capacity; (b) sequentially covering a silica gel soft cover and a sealing film on the top of the container to form a double-sealing structure; (c) starting a micro fan in the container to forcibly drive air circulation; (d) collecting gas data in real time through a non-contact infrared CO2 sensor extending into the container; and (e) transmitting the data of the plurality of detection units to an industrial personal computer through wires to realize multi-channel parallel monitoring and data storage.

Description

Technical Field

[0001] This invention belongs to the field of soil respiration detection technology, specifically relating to a multi-channel real-time analysis method for soil respiration based on a carbon dioxide sensor. Background Technology

[0002] Soil respiration, a biological process by which soil microorganisms, plant roots, and animal metabolism release carbon dioxide (CO2), constitutes a core pathway for soil carbon sink output. It comprises two main components: microbial heterotrophic respiration and root autotrophic respiration. It is not only a key indicator for assessing soil biological activity and carbon conversion efficiency but also a crucial link connecting the underground carbon cycle with the atmospheric system. Global-scale studies show that annual carbon emissions from soil respiration reach 98 ± 12 Pg C, equivalent to 11 times the emissions from fossil fuels, ranking as the second largest carbon flux between terrestrial ecosystems and the atmosphere. Such a massive flux means that even small measurement deviations can significantly affect the accuracy of atmospheric CO2 concentration predictions, thereby interfering with the reliability of global climate models. Therefore, developing high-precision soil respiration detection technology is of great significance for accurately estimating soil carbon flux and carbon sink effects, accurately assessing soil health, and accurately predicting the impacts of climate change on terrestrial ecosystems.

[0003] Soil respiration (rate) is defined as the amount of CO2 released per unit of soil per unit time. Current mainstream analytical methods for soil respiration include alkaline absorption, static chamber methods, and dynamic chamber methods. However, these methods have significant limitations for high-frequency, multi-channel detection of soil respiration, especially for large sample sizes in laboratories. The alkaline absorption method, a classic approach, indirectly calculates emissions by titrating the remaining alkali after CO2 absorption with alkali solution. While simple to operate, it suffers from drawbacks such as long processing time (typically 24-48 hours), low detection frequency, and difficulty in real-time and automated monitoring. The static chamber method uses a closed gas chamber to capture gas, combined with gas chromatography analysis to achieve high-precision CO2 concentration determination. However, CO2 detection relies on expensive equipment, requiring dedicated gas path (safety) devices and personnel for maintenance. Furthermore, each measurement requires manual intervention for instrument and data analysis, resulting in poor timeliness and high costs. While dynamic chamber methods allow for continuous monitoring, achieving multi-channel monitoring requires complex valves for gas path switching and ensuring a strictly sealed gas path system. Therefore, they still suffer from high system complexity and maintenance difficulties. Dynamic chambers are typically used for single-channel gas measurements in the field. Pressure methods are less common due to environmental interference, low accuracy, and difficulty in ensuring data stability. These methods share three common problems: first, insufficient timeliness, with single measurement cycles lasting hours to days, making it impossible to capture instantaneous changes in respiration rate; second, low data density, relying on manual sampling at discrete time points, making it difficult to establish high-resolution time series; and third, limited throughput, limited by CO2 detection equipment, supporting only single-sample serial monitoring, making it difficult to meet the needs of multi-sample parallel monitoring. A more detailed comparison of the advantages and disadvantages of these methods is shown in Table 1.

[0004] As research on soil microbial activity deepens, especially with the increasing demand for quantifying the dynamic processes of basal respiration and substrate-induced respiration (SIR), traditional methods are no longer sufficient to support the acquisition of high spatiotemporal resolution data. For example, when assessing the impact of fertilization on soil respiration, it is necessary to simultaneously monitor the real-time response curves of multiple soil samples; when investigating the microbial metabolic burst after glucose addition, it is necessary to capture the CO2 release inflection point at a frequency of minutes. The current technological system lacks a solution that simultaneously meets the requirements of multi-channel parallel acquisition, high-frequency acquisition, and automated operation, becoming a bottleneck restricting soil carbon cycle research. Summary of the Invention

[0005] Technical problem solved: This invention provides a real-time multi-channel analysis method for soil respiration based on a carbon dioxide sensor, which solves the problems of poor timeliness, low data volume, high cost and excessive reliance on manual operation in traditional soil respiration detection methods.

[0006] Technical Solution: A real-time multi-channel analysis method for soil respiration based on a carbon dioxide sensor, comprising the following steps: (a) placing a standardized soil sample in a transparent container, wherein the standardization process includes: adapting the soil mass to the container volume and controlling the moisture content to 40%-60% of the soil's maximum water holding capacity; (b) sequentially covering the top of the container with a silicone soft cap, forming a double-sealed structure with a sealing film; (c) activating a micro fan inside the container to force air circulation; (d) acquiring gas data in real time using a non-contact infrared CO2 sensor inserted into the container; (e) transmitting data from multiple detection units to an industrial control computer via wires to achieve multi-channel parallel monitoring and data storage.

[0007] In the above-mentioned double sealing structure: the silicone soft cap has a Shore hardness of 40-50 HA and a thickness of 4±0.5 mm; the sealing film is Parafilm PM996 and is fixed by elastic cord sealing.

[0008] The aforementioned miniature fan has a voltage of 5 V, a diameter of 8±0.1 cm, and a speed of 3000±100 rpm.

[0009] The aforementioned transparent container is a lead-free glass bottle with a volume of 2±0.1 L, a bottom diameter of 14±0.2 cm, a height of 21.5±0.3 cm, and a mouth diameter of 8.5±0.1 cm.

[0010] The soil mass mentioned in step (a) is 10.0 ± 0.2 g dry weight.

[0011] Before step (a), pretreatment is also included: pre-culturing the soil sample for 24-48 h; opening the container and starting the fan for 3-5 min to make the CO2 concentration in the container consistent with the environment.

[0012] The range of the infrared CO2 sensor described in step (d) is 0-10000 ppm.

[0013] The industrial computer in step (e) automatically collects and stores data at a frequency of 2 times / min.

[0014] The above method operates at least two detection units simultaneously, and each unit is independently connected to the same industrial control computer.

[0015] The above method is used to determine any of the following types of respiration: basic soil respiration, i.e., soil microbial respiration without the addition of nutrients; and soil induced respiration, i.e., soil microbial respiration after the addition of glucose solution.

[0016] Beneficial Effects: This invention, utilizing a precise CO2 sensor, significantly improves the timeliness and data reliability of soil respiration detection through the synergistic effect of standardized sample processing, a double-sealing structure, and an active gas circulation mechanism. Specifically: multiple detection units are directly connected to an industrial control computer via independent sensors, supporting simultaneous monitoring of at least 10 samples (see stability test in Example 1), overcoming the low throughput of traditional single-channel serial detection. Measured coefficients of variation (CV) are all below 0.01, ensuring the comparability of multi-sample data; a non-contact infrared sensor combined with a data acquisition frequency of 2 times / min (see Examples 3-4) enables minute-level continuous monitoring, accurately capturing the dynamic process of soil respiration (such as the glucose response inflection point in the CMF group in Example 4); double sealing (silicone soft cap + sealing film, oxygen permeability <50 cm³). 3 / m 2 The system minimizes the risk of gas leakage by using a 24-hour atm method; a micro-fan forces circulation (3000±100 rpm) to eliminate the CO2 concentration gradient within the container (compared to the extremely low CV value in the empty bottle test of Example 1); standardized treatment (soil quality adapted to volume + water holding capacity 40%-60%) maintains stable microbial activity, and these three factors work together to ensure data consistency (the respiration curves of parallel samples in the same group in Example 2 are highly consistent); the industrial control computer automatically collects and stores data, replacing manual sampling and chromatographic analysis steps, making short-cycle detection of 30 minutes possible (see Example 3), which is more efficient than the static box method. This synergistic system ultimately achieves high-throughput, low-error, and continuous quantitative analysis of soil respiration, which is crucial for the dynamic correlation study of basal respiration and induced respiration (e.g., Figure 3-4 (As shown) provides technical support.

[0017] Table 1. Comparison of different methods for measuring soil respiration

[0018] Attached Figure Description

[0019] Figure 1 This is a schematic diagram of the detection unit in a multi-channel real-time detection method. 1-Lead-free glass bottle, 2-Silicone soft cap, 3-Rubber band, 4-Sealing film, 5-Soil sample, 6-Fan, 7-Infrared CO2 sensor, 8-Wire.

[0020] Figure 2 This is a complete schematic diagram of a multi-channel real-time monitoring method. 9 represents the industrial computer, and 10 represents the power supply.

[0021] Figure 3 This is a test diagram of airtightness using different sealing methods for multi-channel real-time detection.

[0022] Figure 4 To measure the basic respiration curve of soil using a multi-channel real-time detection method.

[0023] Figure 5 To measure the soil induced respiration curve using a multi-channel real-time detection method. Detailed Implementation

[0024] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described below with reference to specific embodiments. Unless otherwise described in detail, the technical means used in the following embodiments are all conventional means well known to those skilled in the art.

[0025] Example 1:

[0026] A method for detecting soil respiration (hereinafter referred to as the detection method), the schematic diagram of the detection unit is shown below. Figure 1 As shown. A lead-free glass bottle 1 was selected as the measuring chamber, with the following geometric parameters: volume 2±0.1 L, bottom diameter 14±0.2 cm, height 21.5±0.3 cm, and mouth diameter 8.5±0.1 cm. A silicone soft cap 2 was installed at the mouth of the bottle as the main sealing layer (Shore hardness 40-50 HA, thickness 4±0.5 mm), and an outer sealing film 4 (thickness 0.1 mm, oxygen permeability <50 cm³ / m²·24h·atm) was covered by an elastic rope 3. 10.0±0.2 g (dry weight) of soil sample 5 (moisture content of 40%-60% of maximum water holding capacity) was put into the bottle. At the same time, a small embedded fan 6 (voltage 5V, diameter 8±0.1 cm, speed 3000±100 rpm) was started and continuously operated to eliminate the CO2 concentration gradient. An infrared CO2 sensor (range 0-10000ppm) 7 was used to insert the probe into the bottle to collect gas data in real time. Each detection unit is connected to a multi-channel data acquisition unit via wire 8, and the data is ultimately transmitted to an industrial control computer 9 for dynamic display and storage (the industrial control computer model is Hangzhou Maichong Technology F10G20). This enables parallel real-time monitoring of multiple detection units. A small fan is connected to a power supply 10. A complete schematic diagram is shown below. Figure 2 As shown.

[0027] This application uses a 2±0.1 L lead-free glass bottle as the standard testing container and 10 g of soil as the standard sample volume. This choice has many advantages:

[0028] 1. Operation and Usage: This volume provides the necessary minimum space for the built-in sensor probe and its essential mechanical fixing structure, while ensuring sufficient volume for sample placement. Under the premise of meeting the aforementioned minimum functional requirements, the volume is minimized to 2 L. The 2 L container is lighter and smaller, reducing the workload of operators and making sample addition, handling, and replacement processes more convenient and safer. At the same time, this physical size is more conducive to adaptation and layout with multi-channel, highly integrated automated detection equipment, providing a foundation for building a compact, high-throughput real-time detection system. A 10 g soil sample is sufficient to meet statistical representativeness requirements, accurately reflecting the overall characteristics of the tested object, while meeting practical needs and avoiding resource waste caused by excessive samples. The soil sample mass can be adjusted according to the soil respiration level during actual testing.

[0029] 2. Measurement Accuracy: The small-volume chamber design brings a synergistic effect of "signal amplification" and "error reduction": on the one hand, it increases the absolute value of concentration changes, enhancing the sensor's response; on the other hand, it suppresses the relative impact of environmental fluctuations and system noise on the measurement results, thus comprehensively improving the sensitivity, accuracy, and reliability of the measurement. Furthermore, the small volume significantly reduces the diffusion path and time required for CO2 gas, ensuring that the CO2 concentration in the sensing element's detection area and the average concentration throughout the container quickly reach thermodynamic equilibrium within the measurement cycle. This effectively eliminates measurement deviations caused by uneven concentration gradient distribution, thus laying the foundation for high-precision measurement. The 2 L container volume is optimally matched to the gas volume produced by a 10 g sample. This ensures that a sufficiently significant and easily measurable gas concentration change value can be generated per unit time, thereby significantly reducing the relative measurement error and improving the method's sensitivity and accuracy.

[0030] The typical usage method of multi-channel detection provided in this application is as follows:

[0031] 1. Soil pre-culture: Weigh 10 g of soil sample, adjust the soil moisture content, and pre-culture for 24-48 h.

[0032] 2. Ventilation: Open the soft cap of the measuring bottle and start the embedded fan to ventilate for 3-5 minutes to make the CO2 concentration inside the bottle the same as that in the outside air.

[0033] 3. Sample loading: Transfer the pre-cultured soil samples into lead-free glass bottles for further culture.

[0034] 4. Device sealing: Cover the bottle mouth with a soft cap and seal the bottle mouth with a sealing film to prevent air leakage.

[0035] 5. Multi-channel measurement: If multiple samples need to be measured simultaneously, repeat the above steps in other detection units.

[0036] 6. Start Measurement: Start the industrial control computer's software, and the CO2 sensor will begin collecting data.

[0037] 7. Data Acquisition: After the required time for incubation and testing in the bottle, data is acquired through industrial control computer software.

[0038] 8. Calculation of soil respiration intensity: Calculate the soil respiration intensity based on the obtained data.

[0039] Stability Test of a Multi-channel Real-time Detection Method for Soil Respiration

[0040] To measure the stability of this detection method, 10 detection units were used. Each unit was placed in an empty beaker, covered with a silicone soft cap, and the CO2 probe was ensured to be centered in the bottle. After confirming this, the bottle opening was sealed a second time with a rubber band and sealing film. The fan was turned on, and after stabilizing for 10 minutes, the industrial control computer measurement software was opened to start counting. The empty bottle was left to stand for 30 minutes, and the automatic data collection interval was set to 2 minutes. The coefficient of variation (CV) within each group was calculated and retained to 4 decimal places. The results are shown in Table 2 (unit: ppm). The CV values ​​of all 10 groups were less than 0.01, indicating that the detection method has good stability.

[0041] Table 2. Stability test results of the multi-channel detection method

[0042] Accuracy Test of Multi-channel Real-time Detection Method for Soil Respiration

[0043] Weigh 1.06 g of sodium carbonate into a beaker, place the beaker into the detection unit, cover with a silicone soft cap, ensuring the CO2 probe is centered in the bottle, turn on the fan, and start the measurement using the industrial control computer software, setting the automatic data collection interval to 30 seconds. Open the silicone soft cap, quickly add excess dilute hydrochloric acid to the beaker, then immediately cover with the silicone soft cap again, and seal the bottle opening a second time with a rubber band and sealing film. Record the data, repeating this process three times. Calculate the actual CO2 emissions using the acquired data, and simultaneously calculate the theoretical CO2 emissions that 1.06 g of sodium carbonate can produce. Comparing the two sets of data, the actual CO2 emissions measured by the three channels were 0.4856, 0.4747, and 0.4604 g, respectively, while the theoretical emission was 0.44 g. The relative deviation was less than 10%, indicating that this method can accurately detect the amount of CO2 produced. The slightly higher results may be due to water vapor inside the bottle potentially affecting the results.

[0044] Table 3. Accuracy test results of multi-channel real-time detection method

[0045]

[0046] Comparison Example 1: Fan on vs. Fan off

[0047] To verify the effectiveness of the small fan, six identical wet soil samples, each weighing 10 g (dry weight), were weighed. These samples were pre-cultured in a constant temperature incubator (27℃±3℃) for 24 hours. The six samples were then placed into beakers, and the beakers were placed into the detection units, with silicone soft caps on, ensuring the CO2 probe was centered. After confirming everything was correct, the bottle openings were sealed a second time with rubber bands and sealing film. The fans were turned on in three detection units, and off in the remaining three. After stabilizing for 10 minutes, the industrial control computer measurement software was opened to start counting. The empty bottles were left to stand for 20 minutes, with an automatic data collection interval of 2 minutes. Linear fitting was performed on each group to calculate the soil respiration intensity. The results are shown in Table 4 (unit: ppm). The calculated soil respiration intensity of the group without fans was lower than that of the group with fans, indicating that turning on the fan is effective in this detection method and can ensure a uniform distribution of CO2 within the measurement system. Without the fan, CO2 deposition is likely, causing the CO2 concentration in the detection area of ​​the glass bottle to be lower than that in the probe detection area, resulting in lower detection results.

[0048] Table 4. Results of small fan performance verification

[0049]

[0050] Comparative Example 2: Comparison of Double-Layer Seals and Single-Layer Seals

[0051] To verify the effectiveness of the double-sealed structure, four detection units were used, each containing an empty beaker. Carbon dioxide was filled into four of the measuring bottles to maintain a high concentration of carbon dioxide inside, and then silicone soft caps were placed on top, ensuring the CO2 probe was centered within the bottle. After confirming correct installation, two of the detection units were resealed using rubber bands to secure the sealing film; the remaining two units were not resealed as a control. The fan was turned on, and after stabilizing for 10 minutes, the industrial control computer measurement software was started to collect data. All measuring bottles were continuously monitored for 24 hours in a static state, with the automatic data collection interval set to 0.5 minutes. The experimental results are as follows: Figure 3 As shown, the rate of decrease in carbon dioxide concentration in the double-sealed measuring bottle is significantly slower, indicating that the double-sealed structure effectively improves the airtightness of the detection unit, which helps to reduce measurement errors caused by gas leakage, thereby making the measurement results more reliable and rigorous.

[0052] Determination of Soil Basal Respiration Based on Multichannel Detection Method

[0053] Soil basal respiration refers to the respiration rate of soil without added nutrients. Two soil samples were collected: conventionally fertilized alluvial soil (NPK) and unfertilized alluvial soil (CK), each weighing 10g (dry weight), with two replicates for each soil type. The samples were pre-incubated in a constant temperature incubator (27℃±3℃) for 24 hours. After pre-incubation, the soil samples were placed in 100 mL beakers. The soft cap of the glass bottle was opened, the beaker was placed inside, and the silicone soft cap was replaced, ensuring the CO2 probe was centered in the bottle. After confirming this, the bottle opening was sealed a second time with a rubber band and sealing film. The fan was turned on, and after stabilizing for 10 minutes, the industrial control computer measurement software was started to begin counting. The samples were incubated and measured in the measurement unit for 30 minutes, with an automatic data collection interval set to 2 minutes. The results were calculated as follows: Figure 3 As shown, CK1 and CK2 represent unfertilized wet soil, NPK1 and NPK2 represent fertilized wet soil, and the blank represents the control group. A total of 5 detection units were used. It can be seen that the conventionally fertilized wet soil emitted more CO2 in the same time period, and the results were parallel and stable within the same group, indicating that the detection method has good reproducibility.

[0054] Determination of soil-induced respiration based on multi-channel real-time detection method

[0055] Soil-induced basal respiration refers to the respiration rate of soil after the addition of nutrients such as glucose as a substrate. 10 g of each of the three treatments of wet soil were weighed: soil samples treated with conventional fertilizer (NPK), mushroom residue fertilizer (MRF), and chicken manure fertilizer (CMF), with two replicates for each treatment. After pre-incubation, the soil samples were placed in 100 mL beakers, and 1.5 mL of 40 g / L glucose solution was added. The mixture was shaken for 3 min to ensure thorough mixing. After shaking, the soft cap of the bottle was opened, and the beaker was placed inside the test glass bottle. The silicone soft cap was then replaced, ensuring the CO2 probe was centered in the bottle. After confirming this, the bottle opening was sealed a second time with a rubber band and sealing film. The fan was turned on, and after stabilizing for 10 min, the industrial control computer measurement software was started to begin counting. The samples were incubated and measured in the measurement unit for 36 h, with an automatic data collection interval set to 0.5 h. The results were calculated as follows: Figure 4 As shown, NPK1 and NPK2 were fertilized wet soils, MRF1 and MRF2 were wet soils fertilized with mushroom residue fertilizer, CMF1 and CMF2 were wet soils fertilized with chicken manure fertilizer, and the blank was the control group. A total of 7 detection units were used. It can be seen that the soil samples with the same treatment have good parallelism and stable measurement data. The induced respiration curves of different treatments have significantly different characteristics. Among them, the CMF group has the shortest soil respiration lag period, the fastest response to glucose, and the highest CO2 release rate during the logarithmic growth phase.

[0056] The above examples are merely illustrative of the technical concept and features of the present invention, intended to enable those skilled in the art to understand the content of the invention and implement it accordingly, and should not be construed as limiting the scope of protection of the present invention. All equivalent transformations or modifications made in accordance with the spirit and essence of the present invention should be covered within the scope of protection of the present invention.

Claims

1. A method for real-time multi-channel analysis of soil respiration based on a carbon dioxide sensor, characterized in that, Includes the following steps: (a) The standardized soil sample is placed in a transparent container, the standardization process including controlling the soil moisture content to 40%-60% of the soil's maximum water holding capacity; (b) Cover the top of the container with a silicone soft cap in sequence, and the sealing film forms a double sealing structure; (c) Start the micro fan inside the container to force air circulation; (d) Collect gas data in real time through a non-contact infrared CO2 sensor that extends into the container; (e) Transmit the data from multiple detection units to the industrial control computer through wires to realize multi-channel parallel monitoring and data storage.

2. The method according to claim 1, characterized in that, In the double sealing structure: the silicone soft cap has a Shore hardness of 40-50 HA and a thickness of 4±0.5 mm; the sealing film is Parafilm PM996 and is fixed by elastic cord sealing.

3. The method according to claim 1, characterized in that, The miniature fan has a voltage of 5 V, a diameter of 8±0.1 cm, and a speed of 3000±100 rpm.

4. The method according to claim 1, characterized in that, The transparent container is a lead-free glass bottle with a volume of 2±0.1 L, a bottom diameter of 14±0.2 cm, a height of 21.5±0.3 cm, and a mouth diameter of 8.5±0.1 cm.

5. The method according to claim 1, characterized in that, The mass of the soil sample in step (a) is 10.0 ± 0.2 g dry weight.

6. The method according to claim 1, characterized in that, Before step (a), pretreatment is also included: pre-culturing the soil sample for 24-48 h; opening the container and starting the fan for 3-5 min to make the CO2 concentration in the container consistent with the environment.

7. The method according to claim 1, characterized in that, The range of the infrared CO2 sensor described in step (d) is 0-10000 ppm.

8. The method according to claim 1, characterized in that, The industrial computer in step (e) automatically collects and stores data at a frequency of 2 times / min.

9. The method according to any one of claims 1-8, characterized in that, The method operates at least two detection units simultaneously, and each unit is independently connected to the same industrial control computer.

10. The method according to claim 9, characterized in that, The method is used to determine any of the following types of respiration: basic soil respiration, i.e., soil microbial respiration without the addition of nutrients; and soil induced respiration, i.e., soil microbial respiration after the addition of glucose solution.

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