Method for graded extraction and stress determination of micro-plastics in activated sludge

Abstract

The invention discloses a method for graded extraction and stress determination of micro-plastics in activated sludge, and belongs to the technical field of sewage treatment. In order to solve the problems of insufficient extraction of micro-plastics in a sewage treatment plant and insufficient determination of the stress of the micro-plastics on activated sludge at present, the invention can accurately extract the micro-plastics in sewage through an efficient separation and detection technology, and determine the stress of the micro-plastics on the activated sludge based on the occurrence state of the micro-plastics. Therefore, a technical support and a solution are provided for removal of micro-plastic pollution and performance protection of activated sludge in a sewage treatment plant. The innovation fills the blank of micro-plastic removal mechanism research, and provides a new thought and technical means for sewage treatment process optimization and ecological protection.

Description

Technical Field

[0001] The invention belongs to the technical field of sewage treatment, and specifically relates to a method for graded extraction of microplastics in activated sludge and a method for determining their stress. Background Art

[0002] Sewage treatment plants are an important part of urban sewage treatment, and they are also key places for the collection and release of microplastics. With the extensive use and disposal of plastic products, microplastics, as a new type of pollutant, have attracted widespread attention from the world. Sewage treatment plants not only undertake the important task of sewage purification, but also play an important role in the removal and re-release of microplastics. Existing studies have shown that microplastics in sewage treatment plants can be partially removed through treatment processes, but their removal mechanism is still in the initial exploration stage. Although there have been many research results at home and abroad on the identification of microplastics, the effect of process removal and the seasonal changes of microplastics, there is still a lack of in-depth research on the occurrence state of microplastics, the removal mechanism and its impact on activated sludge in sewage treatment plants.

[0003] First, the removal mechanism of microplastics is still unclear, resulting in obvious limitations in the removal efficiency of existing processes. The biological treatment processes commonly used in sewage treatment plants mainly rely on the metabolic function of activated sludge to remove pollutants such as organic matter, nitrogen, and phosphorus, but there is a lack of systematic research and verification on how microplastics are captured or discharged in these processes. Therefore, the current problem of microplastic removal remains in the "black box mode", and process optimization is difficult, making it impossible to achieve low-cost and efficient microplastic removal.

[0004] Secondly, the existence state of microplastics in sewage treatment plants is complex and diverse. They may exist in different treatment units in three forms: free, loosely adsorbed, and tightly attached. Microplastics in different existence states have different effects on the biological functions of activated sludge. Activated sludge is a core component in sewage treatment, and its performance is crucial to the effect of sewage treatment. Microorganisms in activated sludge complete the degradation of organic pollutants and the removal of nitrogen and phosphorus through metabolism. However, it is not clear whether microplastics affect the metabolic function of activated sludge, especially its influence mechanism on biological denitrification, phosphorus removal, and carbon removal processes. Existing studies have no clear quantitative indicators for the stress of microplastics on activated sludge, and lack in-depth assessment of its potential impact.

[0005] In addition, the detection and extraction technology of microplastics in sewage treatment plants is still imperfect. Existing microplastic detection methods are mostly concentrated in water bodies and sediments, and there are few studies on the quantitative extraction of microplastics in sewage treatment processes and the classification of their occurrence states. How to effectively extract microplastics from sewage in a graded manner and accurately evaluate the impact of their different occurrence states on activated sludge has become a key issue in the optimization of sewage treatment processes.

[0006] Therefore, it is particularly important to propose an innovative method that can efficiently and accurately extract microplastics from sewage in a graded manner and measure their stress to activated sludge. This can not only help sewage treatment plants understand the removal rules of microplastics in the treatment process and optimize the process flow, but also provide a scientific basis for microplastic pollution control and ecological risk assessment in sewage treatment. The graded extraction of microplastics in their state of existence and the determination of their stress to activated sludge can help researchers better understand the potential impact of microplastics on the biological treatment process in sewage treatment plants, and thus provide technical support for sewage treatment plants to formulate effective microplastic removal strategies. Summary of the invention

[0007] In view of the current shortcomings in the extraction of microplastics in sewage treatment plants and the determination of their stress on activated sludge, the present invention provides a method for the graded extraction of microplastics in different states of existence in activated sludge and the determination of their stress. It is possible to effectively separate microplastics from various treatment links in sewage treatment plants and evaluate their impact on activated sludge. The method specifically performs graded extraction of free, loosely adsorbed and tightly attached microplastics to ensure the efficiency and accuracy of the extraction process.

[0008] In order to achieve the above object, the present invention adopts the following technical solutions:

[0009] A method for fractionating and extracting microplastics from activated sludge and determining their stress, comprising the following steps:

[0010] Step 1, centrifuge the activated sludge to be tested, collect solid sample A and supernatant A; use a filter membrane to filter the supernatant A; place the filter membrane in a glass culture dish, add alkaline potassium persulfate solution to digest it, and dry the filter membrane; place the filter membrane back in a glass culture dish, add Fenton reagent solution to digest it, and dry the filter membrane, which is the filter membrane enriched with free microplastics; place the filter membrane enriched with free microplastics under an electron microscope, take pictures and count them;

[0011] Step 2, drying the solid sample A in step 1, adding a flotation liquid to repeatedly flotate for multiple times, shaking and standing, mixing the supernatants after multiple flotations to obtain a supernatant B and a solid sample B;

[0012] Filter the supernatant B using a filter membrane; place the filter membrane in a glass culture dish, add an alkaline potassium persulfate solution to digest it, and dry the filter membrane; place the filter membrane back in a glass culture dish, add a Fenton reagent solution to digest it, and dry the filter membrane, which is the filter membrane that enriches loosely adsorbed microplastics; place the treated filter membrane under an electron microscope, take pictures, and count;

[0013] Step 3, drying the solid sample B in step 2, adding Fenton's reagent to digest; centrifuging the digested sample and collecting the supernatant C; using a filter membrane to filter the supernatant; placing the filter membrane in a glass culture dish, adding alkaline potassium persulfate solution to digest, and drying the filter membrane; placing the filter membrane back in a glass culture dish, adding Fenton's reagent solution to digest, and drying the filter membrane, which is a filter membrane enriched with tightly attached microplastics; placing the treated filter membrane under an electron microscope, taking pictures and counting;

[0014] Step 4, the activated sludge to be tested is activated sludge containing free microplastics, loosely adsorbed microplastics and enriched tightly attached microplastics; solid sample A is activated sludge containing loosely adsorbed microplastics and enriched tightly attached microplastics; solid sample B is activated sludge containing tightly attached microplastics;

[0015] The activated sludge to be tested, solid sample A and solid sample B were fixed to the initial volume, nutrients were added, and the nitrification activity, denitrification activity, carbon metabolism activity and polyphosphate activity of the sludge were measured respectively; by comparing the activated sludge containing microplastics in different existence states, the stress of microplastics in different existence states on the activated sludge was calculated.

[0016] Furthermore, the filter membrane is a 0.22-0.45 μm water filter membrane.

[0017] Furthermore, during the digestion process of the alkaline potassium persulfate solution, the digestion solution is 1-2 mol / L alkaline potassium persulfate.

[0018] Furthermore, in the digestion of the Fenton reagent solution, the concentration of the Fenton reagent is 5-10% H2O2 and 1.25-2.5mmol / L FeSO4.

[0019] Furthermore, the centrifugal speed in step 1, step 2 and step 3 is 5000-7000 r / min, and the centrifugal time is 5-15 min.

[0020] Furthermore, in the flotation, the flotation liquid is deionized water.

[0021] Further, the flotation liquid is added and repeatedly floated for multiple times, and the shaking and standing are specifically: placing in a constant temperature shaker at 20-30° C. and 150-180 r / min for 12-24 hours; placing the shaken sample in a constant temperature incubator and standing for 6-12 hours, transferring the supernatant after standing to a beaker for standby use, and repeating the flotation of the sample 2-5 times.

[0022] Furthermore, the drying temperature is 60-80°C.

[0023] Furthermore, the calculation formula for the stress of microplastics in different states on activated sludge is:

[0024] MPS FS,i =SA LB+TB,i -SA FS+LB+TB,i

[0025] MPS LB,i =SA TB,i -SA LB+TB,i

[0026] MPS TB,i =SA L，i -SA TB,i

[0027] Among them, MPS FS,i MPS is the stress of free microplastics on the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge per unit mass, mg / (mg·min); LB,i MPS is the stress of loosely adsorbed microplastics on the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of unit mass of activated sludge, mg / (mg·min); TB，i SA is the stress of tightly attached microplastics on the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of unit mass of activated sludge, mg / (mg·min); LB+TB，i is the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge containing loosely adsorbed microplastics and tightly attached microplastics, mg / (L·min); SA FS+LB+TB，i is the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge containing free microplastics, loosely adsorbed microplastics and tightly attached microplastics, mg / (L·min); SA LB,i is the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge containing tightly attached microplastics, mg / (L·min); SA L,i are the theoretical nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge, mg / (L·min); FS is free microplastics; LB is loosely adsorbed microplastics; TB is tightly attached microplastics; i1, i2, i3 and i4 are used to represent the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge, respectively.

[0028] Compared with the prior art, the present invention has the following advantages:

[0029] 1. Through a precisely designed three-step separation process (centrifugation, flotation, digestion), free, loosely adsorbed, and tightly attached microplastics in the sewage treatment plant were successfully separated. Compared with other technologies, this graded extraction method is more comprehensive in that it only extracts a single microplastic form, and can identify and distinguish the different states of microplastics in activated sludge, improving the accuracy of separation and analysis.

[0030] 2. Multiple flotation processes are used to effectively release loosely adsorbed microplastics, and the optimized digestion steps ensure the complete separation of tightly attached microplastics. Compared with other methods that only perform a single flotation or simplify the digestion steps, the present invention can more comprehensively extract all microplastic forms, ensuring the comprehensiveness and reliability of the measurement results.

[0031] 3. Collect microplastics through filter membrane filtration, and introduce alkaline potassium persulfate and Fenton reagent to digest and remove non-microplastic organic matter on the filter membrane, ensuring that only microplastic particles remain on the filter membrane, and then accurately count them with an electron microscope. This digestion and counting method significantly improves the accuracy of quantitative analysis of microplastics, and is more efficient and accurate than traditional methods.

[0032] 4. The present invention provides a comprehensive biological activity assessment system by measuring the comprehensive stress of microplastics in different states on biological functions such as nitrification, denitrification, phosphorus removal and carbon metabolism of activated sludge. Different from other measurement methods that only focus on a single biological function, the present invention can comprehensively evaluate the multi-faceted stress effects of microplastics on activated sludge, providing more abundant parameter support for the optimization of sewage treatment processes.

[0033] 5. The reagents (such as alkaline potassium persulfate, Fenton's reagent, etc.) used in the present invention are simple and easy to obtain, and the equipment requirements are relatively low. Unlike the complex reagents and expensive equipment used in other similar patents, the present invention is more operational and cost-effective, and is convenient for large-scale promotion and application.

[0034] 6. Through graded extraction and stress determination, the present invention can help sewage treatment plants identify and reduce the negative impact of microplastics on the activated sludge treatment process, especially the interference effects in the denitrification, phosphorus removal and organic matter degradation processes, providing a clear technical path for sewage treatment plants to improve process design and reduce microplastic emissions. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 Experiment on the inhibition of ammonia oxidation activity of activated sludge by free microplastics;

[0036] Figure 2 Experiment on the inhibition of denitrification activity of activated sludge by loosely adsorbed microplastics;

[0037] Figure 3Experiment on the inhibition of the readily degradable carbon oxidation activity of activated sludge by tightly attached microplastics;

[0038] Figure 4 Photo of free microplastics on the filter membrane under a microscope;

[0039] Figure 5 Photo of loosely adsorbed microplastics on the filter membrane under a microscope;

[0040] Figure 6 Photo of microplastics tightly attached to the filter membrane under a microscope. DETAILED DESCRIPTION

[0041] In order to gain a deeper understanding of the present invention, we will provide a comprehensive and detailed description of the present invention. However, the present invention has multiple implementations and is not limited to the specific examples listed herein. The presentation of these examples is intended to deepen the comprehensive understanding of the disclosure of the present invention.

[0042] Embodiment 1:

[0043] A 2 Experiment on separation and stress determination of free microplastics in aerobic pools

[0044] (1) Separation

[0045] First, take 500mL of the activated sludge sample to be tested, put it into a centrifuge, and centrifuge it at 6000r / min for 10 minutes to separate the supernatant and solid sludge sample. The supernatant mainly contains free microplastic particles. Then, use a 0.22μm filter membrane to filter the supernatant, and the substances captured on the filter membrane are free microplastics.

[0046] After the filter membrane is filtered, it is placed in a glass culture dish, 10mL of 1mol / L alkaline potassium persulfate solution is added, and it is placed in a 60°C oven for digestion for 2 hours to remove the organic matter on the filter membrane. After digestion, the filter membrane is dried, placed in a culture dish again, 10mL of Fenton's reagent (5% H2O2 and 1.25mmol / LFeSO4) is added, and digestion is continued for 2 hours, and then the filter membrane is dried again. The treated filter membrane is placed under an electron microscope, photographed and counted to obtain the number and morphological data of free microplastics.

[0047] (2) Stress measurement

[0048] In order to determine the effect of free microplastics on the nitrification activity of activated sludge, an experimental group and a parallel experimental group were set up, and conventional activated sludge was used as a control group. The solid sludge samples separated in the separation step (1) were taken and 500 mL of simulated wastewater was added respectively. The composition of the simulated wastewater included: COD 400 mg / L, ammonium chloride 30 mg / L, potassium dihydrogen phosphate 8 mg / L, magnesium sulfate 20 mg / L, and calcium chloride 20 mg / L.

[0049] The experiment was conducted under aerobic conditions, and each group was set up with parallel experiments. Samples were taken every 10 minutes for 60 minutes, and the ammonia nitrogen content of the samples was determined by Nessler's reagent colorimetry. The conventional activated sludge group was compared with the experimental group containing loosely adsorbed microplastics and tightly attached microplastics to calculate the stress of free microplastics on the nitrification activity of activated sludge. The data of the experimental group and the parallel experimental group were compared to ensure the stability and accuracy of the experimental results.

[0050] According to the test results, the number of free microplastics in the aerobic tank was 839.41±1.702, and the inhibitory intensity of free microplastics on the ammonia oxidation activity of activated sludge was 8.86±0.5610. -5 mg / (L·min·particles).

[0051] Example 2

[0052] A 2 Separation and stress determination of loosely adsorbed microplastics in anaerobic O tanks

[0053] (1) Separation

[0054] Take 500mL of the activated sludge sample to be tested, put it into a centrifuge, centrifuge it at 6000r / min for 10 minutes, and separate the supernatant and solid sludge samples. Take the solid sludge sample and dry it. The drying temperature is set to 70℃ and the sample is dried to constant weight. Then add 400mL of deionized water as a flotation liquid to the dried sludge, put it into a constant temperature shaker, and shake it for 12 hours at 25℃ and 150r / min. After shaking, let it stand for 6 hours to release the loosely adsorbed microplastics from the sludge.

[0055] After standing, collect the supernatant after flotation and filter with a 0.22μm filter membrane. The particles collected on the filter membrane are loosely adsorbed microplastics. According to the digestion steps of Implementation Case 1, place the filter membrane in a glass culture dish and use 1mol / L alkaline potassium persulfate solution and Fenton reagent (5% H2O2 and 1.25mmol / LFeSO4) to digest the organic matter on the surface of the filter membrane. Each digestion time is 2 hours, and the filter membrane is dried at 70°C after digestion. The filter membrane is photographed and counted using an electron microscope to record the number of loosely adsorbed microplastics.

[0056] (2) Stress measurement

[0057] In order to determine the effect of loosely adsorbed microplastics on the denitrification activity of activated sludge, an experimental group and a parallel experimental group were set up, and the activated sludge containing loosely adsorbed microplastics and tightly attached microplastics was used as the control group. The solid sludge sample that released loose microplastics in the separation step (1) was taken and 500 mL of simulated wastewater was added. The composition of the simulated wastewater included: 30 mg / L sodium nitrate, 8 mg / L potassium dihydrogen phosphate, 20 mg / L magnesium sulfate, and 20 mg / L calcium chloride.

[0058] The experiment was conducted under anaerobic conditions, and the experimental group and the parallel experimental group were operated independently. Sampling was performed every 10 minutes for 60 minutes, and the changes in the content of nitrate nitrogen were determined by ultraviolet spectrophotometry. By comparing the control group containing loosely adsorbed microplastics and tightly attached microplastics with the experimental group containing tightly attached microplastics, the stress of loosely adsorbed microplastics on the denitrification activity of activated sludge was calculated. The experimental results were verified by parallel groups to ensure the reliability of the data.

[0059] According to the test results, the number of loosely adsorbed microplastics in the anaerobic tank was 461.42±15.32, and the inhibitory intensity of loosely adsorbed microplastics on the denitrification activity of activated sludge was 29.64±0.2810 -5 mg / (L·min·particles).

[0060] Example 3

[0061] A 2 Isolation and stress determination of tightly attached microplastics in anaerobic O tanks

[0062] (1) Separation

[0063] Take 500mL of the activated sludge sample to be tested, put it into a centrifuge, centrifuge it at 6000r / min for 10 minutes, and separate the supernatant and solid sludge samples. Take the solid sludge sample and dry it, set the drying temperature to 70℃, and dry it until the sample is constant weight. Then add 400mL of deionized water as a flotation liquid to the dried sludge, put it into a constant temperature shaker, and shake it for 12 hours at 25℃ and 150r / min. After shaking, let it stand for 6 hours to release the loosely adsorbed microplastics from the sludge. After the loosely adsorbed microplastics are separated, the remaining solid sludge samples are dried at a drying temperature of 75℃ until the sample is constant weight. Then add 200mL of Fenton's reagent (6% H2O2 and 2mmol / LFeSO4) to the sample, digest it at room temperature, and the digestion time is 8 hours to release the tightly attached microplastics. After digestion, the sample is centrifuged at 7000r / min for 15 minutes, and the supernatant is collected.

[0064] The supernatant was filtered through a 0.45 μm filter membrane, and the substances enriched on the filter membrane were tightly attached microplastics. According to the method in Implementation Case 1, the filter membrane was digested with alkaline potassium persulfate solution and Fenton's reagent, and after drying, it was observed and counted under an electron microscope to determine the number and morphological characteristics of tightly attached microplastics.

[0065] (2) Stress measurement

[0066] Take the solid sludge sample that releases loose microplastics in the separation step (1), add 500mL of simulated wastewater, and set up a parallel experimental group. The composition of the simulated wastewater is: COD 400mg / L, glucose 375mg / L, potassium dihydrogen phosphate 8mg / L, magnesium sulfate 20mg / L, calcium chloride 20mg / L. The oscillation reaction is carried out under aerobic conditions, and samples are taken every 10 minutes for 60 minutes. The changes in chemical oxygen demand (COD) are determined using a rapid closed catalytic digestion method. By comparing the theoretical carbon metabolic activity of activated sludge with the measured values ​​of the experimental group containing tightly attached microplastics, the stress of tightly attached microplastics on the carbon metabolism of activated sludge is calculated. Parallel experiments ensure the stability and accuracy of the experimental results.

[0067] According to the test results, the number of tightly attached microplastics in the anaerobic tank was 405.23±23.84, and the inhibitory intensity of tightly attached microplastics on the oxidative activity of easily degradable carbon in activated sludge was 10.87±0.3510 -3 mg / (L·min·particles).

[0068] In summary, the present invention provides a method for the graded extraction of microplastics in sludge / sewage and the determination of its active stress on sludge. Microplastics in different existence states (free state, loosely adsorbed state, and tightly attached state) are effectively separated by three-step centrifugation through the use of microplastic graded extraction technology; efficient extraction of microplastics is achieved by suction filtration of the supernatant, and the extraction process operation steps are simple and fast; alkaline potassium persulfate is used to remove organic matter other than microplastics and particulate carbon black, and the filter membrane is digested by Fenton's reagent to achieve the removal of other carbon-containing particles except black carbon. The Nessler reagent colorimetric method and N-1 (naphthyl)-ethylenediamine spectrophotometry were used to determine the contents of ammonia nitrogen and nitrite nitrogen, the ultraviolet spectrophotometry was used to determine the contents of nitrate nitrogen and nitrite nitrogen, the molybdenum antimony anti-spectrophotometry was used to determine the content of phosphate, and the rapid closed catalytic digestion method was used to determine the chemical oxygen demand, so as to realize the determination of the stress of microplastics on sludge. The measurement results are accurate, efficient and stable, which clarifies the source and fate of microplastics and the mechanism by which microplastics affect the activity of activated sludge, laying the foundation for subsequent research on removing and reducing microplastic emissions and improving treatment processes in sewage treatment plants.

[0069] The contents not described in detail in the specification of the present invention belong to the prior art known to the professional and technical personnel in the field. Although the illustrative specific embodiments of the present invention are described above to facilitate the understanding of the present invention by the technical personnel in the field, it should be clear that the present invention is not limited to the scope of the specific embodiments. For the ordinary technical personnel in the field, as long as various changes are within the spirit and scope of the present invention defined and determined by the attached claims, these changes are obvious, and all inventions and creations using the concept of the present invention are protected.

Claims

1. A method for fractionating and extracting microplastics from activated sludge and determining their stress, characterized in that: The following steps are involved: Step 1, centrifuging the activated sludge to be tested, and collecting a solid sample A and a supernatant A; Filter the supernatant A using a filter membrane; The filter membrane is placed in a glass culture dish, an alkaline potassium persulfate solution is added to digest it, and the filter membrane is dried; the filter membrane is placed back in a glass culture dish, a Fenton reagent solution is added to digest it, and the filter membrane is dried. The filter membrane is a filter membrane that enriches free microplastics; Step 2, drying the solid sample A in step 1, adding a flotation liquid to repeatedly flotate for multiple times, shaking and standing, mixing the supernatants after multiple flotations to obtain a supernatant B and a solid sample B; Filter the supernatant B using a filter membrane; The filter membrane is placed in a glass culture dish, an alkaline potassium persulfate solution is added to digest it, and the filter membrane is dried; The filter membrane is placed back in a glass culture dish, Fenton's reagent solution is added to digest it, and the filter membrane is dried. The filter membrane is the filter membrane that enriches loosely adsorbed microplastics; Step 3, drying the solid sample B in step 2, adding Fenton's reagent for digestion; centrifuging the digested sample to collect the supernatant C; and filtering the supernatant using a filter membrane; The filter membrane is placed in a glass culture dish, an alkaline potassium persulfate solution is added to digest it, and the filter membrane is dried; the filter membrane is placed back in a glass culture dish, a Fenton reagent solution is added to digest it, and the filter membrane is dried. The filter membrane is a filter membrane that enriches tightly attached microplastics; Step 4, the activated sludge to be tested is activated sludge containing free microplastics, loosely adsorbed microplastics and enriched tightly attached microplastics; solid sample A is activated sludge containing loosely adsorbed microplastics and enriched tightly attached microplastics; solid sample B is activated sludge containing tightly attached microplastics; The activated sludge to be tested, solid sample A and solid sample B were fixed to the initial volume, nutrients were added, and the nitrification activity, denitrification activity, carbon metabolism activity and polyphosphate activity of the sludge were measured respectively; by comparing the activated sludge containing microplastics in different existence states, the stress of microplastics in different existence states on the activated sludge was calculated.

2. The method for fractionating and extracting microplastics from activated sludge and determining their stress according to claim 1, characterized in that: The filter membrane is a 0.22-0.45 μm water filter membrane.

3. The method for fractionating and extracting microplastics from activated sludge and determining their stress according to claim 1, characterized in that: During the digestion process of the alkaline potassium persulfate solution, the digestion solution is 1-2 mol / L alkaline potassium persulfate.

4. The method for fractionating and extracting microplastics from activated sludge and determining their stress according to claim 1, characterized in that: In the digestion of the Fenton reagent solution, the concentration of the Fenton reagent is 5-10% H2O2 and 1.25-2.5mmol / LFeSO4.

5. The method for fractionating and extracting microplastics from activated sludge and determining their stress according to claim 1, characterized in that: The centrifugal speed in step 1, step 2 and step 3 is 5000-7000 r / min, and the centrifugal time is 5-15 min.

6. The method for fractionating and extracting microplastics from activated sludge and determining their stress according to claim 1, characterized in that: In the flotation, the flotation liquid is deionized water.

7. The method for fractionating and extracting microplastics from activated sludge and determining their stress according to claim 1, characterized in that: The adding of flotation liquid for repeated flotation, shaking and standing are specifically: placing in a constant temperature shaker at 20-30°C and 150-180r / min for 12-24h; placing the shaken sample in a constant temperature incubator for 6-12h, transferring the supernatant after standing for standby, and repeating the flotation of the sample 2-5 times.

8. The method for fractionating and extracting microplastics from activated sludge and determining their stress according to claim 1, characterized in that: The drying temperature is 60-80°C.

9. The method for fractionating and extracting microplastics from activated sludge and determining their stress according to claim 1, characterized in that: The calculation formula for the stress of microplastics in different states on activated sludge is: MPS FS,i =IN LB+TB,i -IN FS+LB+TB,i MPS LB,i =IN TB,i -IN LB+TB,i MPS TB,i =IN L,i -IN TB,i Among them, MPS FS,i MPS is the stress of free microplastics on the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge per unit mass, mg / (mg·min); LB,i MPS is the stress of loosely adsorbed microplastics on the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of unit mass of activated sludge, mg / (mg·min); TB,i SA is the stress of tightly attached microplastics on the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of unit mass of activated sludge, mg / (mg·min); LB+TB,i is the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge containing loosely adsorbed microplastics and tightly attached microplastics, mg / (L·min); SA FS+LB+TB,i is the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge containing free microplastics, loosely adsorbed microplastics and tightly attached microplastics, mg / (L·min); SA LB,i is the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge containing tightly attached microplastics, mg / (L·min); SA L,i are the theoretical nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge, mg / (L·min); FS is free microplastics; LB is loosely adsorbed microplastics; TB is tightly attached microplastics; i1, i2, i3 and i4 are used to represent the nitrification activity, denitrification activity, carbon metabolism activity and phosphorus accumulation activity of activated sludge, respectively.

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