A method for measuring the sedimentation rate of plastics

Abstract

The present invention belongs to the technical field of water environment detection, and particularly relates to a method for measuring the sedimentation rate of plastics. In the measurement method of the present invention, the requirements for the experimental environment are not high, and accurate measurement can be carried out under most temperatures or light intensities. The sample to be measured only needs to reach a uniform dispersion state for detection, avoiding the cumbersome sample treatment before analysis. Moreover, in the measurement method of the present invention, the calculation of the sedimentation rate is quite simple, scientific, and general. Only by measuring the density of plastic particles before and after sedimentation can the sedimentation rate of plastics be calculated.

Description

Technical Field

[0001] The present invention belongs to the technical field of water environment detection. More specifically, it relates to a method for determining the sedimentation rate of plastics. Background Art

[0002] Due to the widespread use of plastic products and the poor management of related waste, plastic debris pollution in the aquatic environment has become a new global problem. Plastic debris with a size less than 5 mm is generally recognized as microplastics, which are commonly present in the marine environment and freshwater habitats (such as lakes and rivers). In the aquatic environment, microplastics can be ingested by a variety of aquatic organisms, including zooplankton, fish, invertebrates, and marine mammals, causing adverse effects on the health of these aquatic organisms (such as abrasions and ulcers, digestive tract obstruction, reduced energy reserves, reproductive disruption, and increased mortality). Moreover, microplastics can be transferred upward along the food chain like other pollutants such as heavy metals, accompanied by a biomagnification effect. The study of microplastic sedimentation and its trends helps to effectively remove the sedimented microplastics, thereby reducing the harm of microplastics to aquatic organisms. Therefore, in recent years, the determination of the density of plastic particle groups, plastic sedimentation, and its trends in the water environment has attracted the attention of more and more research teams.

[0003] At present, there are many limitations in the determination of the density of plastic particle groups. Generally, the density of plastic particle groups is used to monitor and detect the sedimentation rate of plastics. Therefore, there are also many limitations in the current determination of the sedimentation rate of plastics, especially for the determination of the sedimentation rate of microplastics. For example, scholars have used a series of methods such as gas chromatography-mass spectrometry, liquid chromatography, stereomicroscopy, scanning electron microscopy (SEM), scanning electron microscopy-energy dispersive X-ray spectroscopy (SEM-EDS), and environmental scanning microscopy-energy dispersive X-ray spectroscopy (ESEM-EDS) to qualitatively and quantitatively analyze and identify plastics. However, most of these methods are time-consuming and laborious, and may cause accidental contamination of samples, which will affect the data quality and prevent the comparison of different research results. In addition, the quantification of plastics requires counting under a microscope in many cases, and the data needs to be interpreted by the naked eye, resulting in a large error and inaccurate calculation of the plastic sedimentation rate. For example, a Chinese patent application discloses an analysis method for microplastic pollutants in drinking water, which counts microplastics through a fluorescence microscope, resulting in a large error and inaccurate calculation of the sedimentation rate. Summary of the Invention

[0004] The technical problem to be solved by the present invention is to overcome the deficiencies in the existing determination of the density of plastic particle groups, where there is a lack of a set of scientific and general calculation formulas, and a microscope is required to count the particles by the naked eye, resulting in large errors and inaccurate results of the calculated sedimentation rate. The present invention provides a method for quickly and accurately determining the plastic sedimentation rate.

[0005] The above object of the present invention is achieved by the following technical solutions:

[0006] The present invention protects a method for determining the plastic sedimentation rate, including the following steps:

[0007] S1. Ultrasonically disperse the sample to be measured evenly to obtain sample y1, add sample y1 to the sedimentation column, and let it stand for sedimentation;

[0008] S2. Take the bottom 5-10% of the volume of the sedimentation column as the lower layer, and the rest as the upper layer, collect the lower layer liquid, and oscillate it evenly to obtain sample y2;

[0009] S3. Detect the densities of samples y1 and y2 respectively with a flow cytometer, and calculate the sedimentation rate using the following formula:

[0010] u = f s l / t

[0011] Where u is the average sedimentation rate of the plastic particle group, l is the height of the upper layer of the sedimentation column, t is the sedimentation time, and f s is the net proportion of the number of plastic particles with negative buoyancy in the total number of particles: f s = B s / B t ;

[0012] B s is the net number of particles that have sunk after standing, B s = V s b s - V s b1;

[0013] B t is the total amount of particles in the sedimentation column, B t = b1V t ;

[0014] b1 is the initial density of the plastic particle group, that is, the density of sample y1, b s is the plastic particle concentration of the lower layer liquid volume V s after standing, that is, the density of sample y2, both of which are measured by a flow cytometer; V s is the lower layer liquid volume, V t is the total volume of the liquid in the sedimentation column.

[0015] Preferably, the sample y1 is divided into a sedimentation column group and a retained sample group. The sedimentation column group and the retained sample group are allowed to settle statically under the same conditions. The b1 is wherein, b 0,0 is the result of density measurement by flow cytometry for the sedimentation column group to be evenly dispersed before sedimentation, and b 0,t is the result of density measurement by flow cytometry for the retained sample group to be evenly dispersed after the sedimentation experiment (the purpose of this measurement is to take into account any change in the amount of particles caused by processes other than sedimentation during the sedimentation experiment).

[0016] Preferably, the derivation process of the formula u = f s l / t is as follows: In a vertically fixed sedimentation column filled with a uniformly distributed plastic sample, the plastic particles are initially located at a distance x above the lower liquid. To sink from the upper layer to the lower layer before time t, these particles must have a sinking rate v ≥ x / t. Therefore, the proportion f x of the number of particles initially at a distance x above the lower liquid and that will sink to the lower layer before time t is:

[0017]

[0018] Taking the average of f x for all x values in the column, i.e., the entire length range, gives the proportion of plastic particles vertically distributed throughout the column and that will sink to the lower layer before time t, which is given by the following equation:

[0019]

[0020] Reversing the integration order of the dummy variables x and v, it becomes:

[0021]

[0022] u is the average sedimentation rate of the plastic particle population. It can be clearly seen from the equation that f s should initially be linear with time (because no particle has an infinite sedimentation rate), and will remain linear until the integral term basically becomes non - zero. Since the average sinking rate can be determined from the slope of the number of particles versus time, the equation can be rearranged to write u as a function that only includes the height l of the upper layer of the sedimentation column, the sedimentation time t, and the proportion f s of the number of plastic particles reaching the lower layer during sedimentation, i.e., u = f s l / t.

[0023] This method is a homogenization method. This method involves using a sedimentation column with known geometric parameters (height, volume, etc.). Before the start of the sedimentation experiment, the plastic particle sample contained in the column must be in a uniformly dispersed state (to ensure the accuracy of the experimental results), and the average sedimentation rate of the whole plastic is calculated based on the change in the vertical distribution of the particulate matter after a given time. The core of this method is to obtain the average sinking rate from the net change in the microplastic distribution in the column after a limited time t (such as two hours). This focus does not rely on the time derivative analysis of the change in the density of the microplastic particle population, thus greatly increasing the simplicity of the whole process and avoiding the errors that may occur in the derivative method.

[0024] Preferably, in step S1, the plastic contained in the sample to be tested is microplastic or nanoplastic.

[0025] More preferably, the size x of the microplastic is: 1 μm < x ≤ 5 mm.

[0026] More preferably, the size of the nanoplastic is ≤ 1 μm.

[0027] Preferably, in step S1, the main component of the plastic is polystyrene, polyvinyl chloride or polyethylene terephthalate.

[0028] More preferably, in step S1, the main component of the plastic is polystyrene or polyvinyl chloride.

[0029] Preferably, in step S1, the time of ultrasonic treatment is 5 - 30 minutes.

[0030] More preferably, in step S1, the time of ultrasonic treatment is 10 - 20 minutes.

[0031] Preferably, in step S1, the time of standing is 2 - 12 hours.

[0032] More preferably, in step S1, the time of standing is 2 - 4 hours.

[0033] Preferably, in step S3, 400 - 600 μL of the samples y1 and y2 are respectively taken for detection by a flow cytometer.

[0034] Specifically, in step S3, 500 μL of the samples y1 and y2 are respectively taken for detection by a flow cytometer.

[0035] Preferably, in step S3, the parameters of the flow cytometer are: the time for a single test is 20 - 40 seconds, and the sample flow rate is 20 - 40 μL / min.

[0036] Specifically, in step S3, the time for a single test of each sample by the flow cytometer is 30 seconds, and the sample flow rate is 30 μL / min.

[0037] The present invention has the following beneficial effects:

[0038] In the measurement method of the present invention, the requirements for the experimental environment are not high, and accurate measurement can be carried out under most temperatures or light intensities. The sample to be measured only needs to reach a uniform dispersion state for detection, avoiding the cumbersome sample treatment before analysis. Moreover, in the measurement method of the present invention, the calculation of the sedimentation rate is quite simple, and only the density of the plastic particles before and after sedimentation needs to be measured to calculate the sedimentation rate of the plastic. Description of the Drawings

[0039] Figure 1 It is the spectrum and statistical result graph presented by the flow cytometer for analyzing the microplastic particle sample.

[0040] Figure 2 It is a physical diagram of the sedimentation column during the specific use process in the experiment.

[0041] Figure 3 It is a schematic diagram of the sedimentation column, where 1-3 are all valves, 4 is the upper layer of the sedimentation column, and 5 is the lower layer of the sedimentation column. Detailed Embodiments

[0042] The present invention will be further described below in conjunction with the drawings of the specification and specific embodiments, but the embodiments do not impose any form of limitation on the present invention. Unless otherwise specified, the reagents, methods, and equipment used in the present invention are conventional reagents, methods, and equipment in the technical field.

[0043] Unless otherwise specified, the reagents and materials used in the following examples are all commercially available.

[0044] Model specification of the flow cytometer: cytoflex.

[0045] Manufacturer of the flow cytometer: Beckman Coulter, Inc., USA.

[0046] Example 1 Analysis of the Feasibility and Accuracy of Measuring the Sedimentation Rate of Plastic Particles

[0047] Method: The core of this sedimentation rate calculation is to calculate the sedimentation rate based on the change in the vertical distribution of the plastic particle group before and after sedimentation. Although the process of calculating this sedimentation rate is simple, it is based on a solid theoretical foundation. For the two samplings (before and after sedimentation) of the microplastic or nanoplastic sample particle group, they must be carried out after the sample reaches a uniform dispersion state to ensure the accuracy of the average sedimentation rate result. The feasibility of detecting the density of the microplastic or nanoplastic particle group with a flow cytometer is crucial. To test the feasibility and accuracy, 20 mg of microplastic sample with a particle size of 10 μm (mainly composed of polystyrene) was added to a conical flask, made up to 200 mL with natural seawater or lake water, oscillated and ultrasonicated for 10 minutes until the microplastic particles were uniformly dispersed, thus obtaining a uniform suspension. 500 μL was pipetted for flow cytometer detection. The model of the flow cytometer used in this example and the following examples is cytoflex, produced by Beckman Coulter, USA. For each sample, the single test time is 30 seconds, the sample flow rate is 30 μL / min, each sample is tested three times repeatedly, and the results are averaged.

[0048] Results: As shown in the flow cytogram ( Figure 1 ), the vast majority of plastic particles are concentrated in distribution, and only a few deviate slightly on the cytogram, which is caused by relatively large differences in particle size or material of a few particles. Within the allowable range, it does not affect the statistical results of the instrument.

[0049] Example 2 Determination of the Sedimentation Rate of Microplastics in Seawater

[0050] S1. After washing the sedimentation column, fix it vertically on the iron stand and let it dry naturally. The iron stand should be placed on a horizontal tabletop (as Figure 2 shown).

[0051] S2. Add 20 mg of microplastic sample with a particle size of 10 μm (mainly composed of polystyrene) to a conical flask, make up to 200 mL with natural seawater, oscillate and ultrasonicate for 10 minutes until the microplastic particles are uniformly dispersed, thus obtaining a uniform suspension (sample y1). Pipette 500 μL for flow cytometer detection, and then add all of sample y1 to the sedimentation column (before pouring the sample into the sedimentation column, valves 1 - 3 of the sedimentation column need to be turned to the closed state first). Let it stand and sediment for 2 hours. During this process, the sedimentation column has been standing still, avoiding shaking, and the surrounding temperature remains constant.

[0052] S3. After 2 hours, carefully and slowly rotate the side valve 2, trying to avoid shaking, so that the upper layer 4 of sample y1 flows out, leaving only the lower layer 5 of seawater and the microplastics in it.

[0053] S4. Slowly and carefully rotate valve 3 to allow the seawater in the lower layer and the microplastics to flow out, collect them using a centrifuge tube or a beaker, and shake to evenly distribute the sedimented microplastic particles (sample y2). Take 500 μL of the sample using a pipette for flow cytometry detection.

[0054] Use flow cytometry to detect the density of 500 μL of samples y1 and y2, and through simple calculations, obtain the density b of the microplastic particle population before and after sedimentation. 0,0 And b 0,t .

[0055] Substitute into the sedimentation rate calculation formula u = f s l / t, (f s = B s / B t ), Calculate the sedimentation rate to be 0.070 m / h.

[0056] Example 3: Determine the sedimentation rate of microplastics in lake water

[0057] S1. After washing the sedimentation column, vertically fix it on an iron stand and let it dry naturally. The iron stand should be placed on a horizontal tabletop (as Figure 2 shown).

[0058] S2. Add a 20 mg nano-plastic sample with a particle size of 500 nm (mainly composed of polystyrene) to a conical flask, make up the volume to 200 mL with natural lake water, shake and sonicate for 20 minutes until the microplastic particles are evenly dispersed, thus obtaining a uniform suspension (sample y1). Take 500 μL of the sample using a pipette for flow cytometry detection, and then add all of sample y1 to the sedimentation column (before pouring the sample into the sedimentation column, all three valves of the sedimentation column need to be first rotated to the closed state. The schematic diagram is as Figure 3 shown), and let it stand and sediment for 2 hours. During this process, the sedimentation column should be kept stationary to avoid shaking, and the surrounding temperature should be kept constant.

[0059] S3. After 2 hours, carefully and slowly rotate the side valve 2, try to avoid shaking, and let the upper layer of sample y1 flow out, leaving only the lake water in the lower layer and the microplastics in it.

[0060] S4. Slowly and carefully rotate valve 3 to allow the lake water in the lower layer and the microplastics to flow out, collect them using a centrifuge tube or a beaker, and shake to evenly distribute the sedimented microplastic particles (sample y2). Take 500 μL of the sample using a pipette for flow cytometry detection.

[0061] Use flow cytometry to detect the density of 500 μL of samples y1 and y2, and through simple calculations, obtain the density b of the microplastic particle population before and after sedimentation. 0,0 And b 0,t .

[0062] Substitute into the sedimentation rate calculation formula u = f s l / t, (f s = B s / B t ), The sedimentation rate is calculated to be 0.058 m / h.

[0063] Example 4: Measuring the sedimentation rate of microplastics in seawater

[0064] S1. After washing the sedimentation column, fix it vertically on the iron stand and let it dry naturally. The iron stand should be placed on a horizontal tabletop (as Figure 2 shown).

[0065] S2. Add a 20 mg microplastic sample with a particle size of 10 μm (mainly composed of polyvinyl chloride) into a conical flask, make up the volume to 200 mL with natural seawater, oscillate and ultrasonicate for 10 minutes until the microplastic particles are evenly dispersed, thus obtaining a uniform suspension (sample y1). Use a pipette to aspirate 500 μL for flow cytometry detection, and then add all of sample y1 into the sedimentation column (before pouring the sample into the sedimentation column, all three valves of the sedimentation column need to be turned to the closed state first), and let it stand and settle for 2 hours. During this process, the sedimentation column has been standing still to avoid shaking, and the surrounding temperature remains constant.

[0066] S3. After 2 hours, carefully and slowly rotate the side valve 2, trying to avoid shaking, so that the upper layer of sample y1 flows out, leaving only the lower layer of seawater and the microplastics in it.

[0067] S4. Carefully and slowly rotate valve 3 to let the lower layer of seawater and microplastics flow out, collect them with a centrifuge tube or a beaker, oscillate to make the sedimented microplastic particles evenly distributed (sample y2), and use a pipette to take a 500 μL sample for flow cytometry detection.

[0068] Use flow cytometry to detect the density of 500 μL of samples y1 and y2. After simple calculation, the density b 0,0 of the microplastic particle population before and after sedimentation is obtained 0,t .

[0069] Substitute into the sedimentation rate calculation formula u = f s l / t, (f s = B s / B t ), The sedimentation rate is calculated to be 0.089 m / h.

[0070] Example 5: Study on whether experimental defects in measuring the sedimentation rate of plastics will affect the analysis results

[0071] The calibration of the results obtained from this sedimentation rate measurement method is also a key point. To determine whether experimental defects (such as particles adhering to the column wall) would affect the analysis results, corresponding sedimentation rate analyses (tested according to the above method) were carried out in a series of sedimentation columns with different surface area to volume ratios (the difference in surface area to volume ratio was up to 5 times). The results showed that the calculated sedimentation rates were similar in all cases (P < 0.01) and were independent of the surface area to volume ratio, highlighting the universality of this sedimentation rate measurement method. This proves that in sedimentation experiments, the size of the surface area does not affect the measurement of the passive sinking rate of the contained samples. In addition, the experiments carried out with this method had quite good repeatability, and the coefficient of variation of repeated experiments was less than 10%.

[0072] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited by the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications made without departing from the spirit and principle of the present invention shall be equivalent replacement methods and are all included in the protection scope of the present invention.

Claims

1. A method for measuring the sedimentation rate of plastics, characterized in that, It includes the following steps: S1. Ultrasonically disperse the sample to be tested evenly to obtain sample y1, add sample y1 into the sedimentation column, and let it stand for sedimentation; S2. Take the bottom 5-10% of the volume of the sedimentation column as the lower layer, and the rest as the upper layer, collect the lower-layer liquid, and oscillate it evenly to obtain sample y2; S3. Detect the density of samples y1 and y2 respectively with a flow cytometer, and calculate the sedimentation rate using the following formula: u = f s l / t where u is the average sedimentation rate of the plastic particle swarm, l is the height of the upper layer of the sedimentation column, t is the sedimentation time, and f s is the net proportion of the number of plastic particles with negative buoyancy in the total number of particles: f s = B s / B t ; B s is the number of net particles that have sunk after standing, B s = V s b s - V s b1; B t is the total amount of particles in the sedimentation column, B t = b1V t ; b1 is the initial density of the plastic particle group, that is, the density of sample y1, b s is the volume V of the lower-layer liquid after standing s of the plastic particle concentration, that is, the density of sample y2, all determined by a flow cytometer; V s is the volume of the lower-layer liquid, V t is the total volume of the liquid in the sedimentation column; In step S1, the plastic contained in the sample to be tested is microplastic or nanoplastic; The size x of the microplastic is: 1 μm < x ≤ 5 mm.

2. The method according to claim 1, characterized in that, The size of the nanoplastic is < 1 μm.

3. The method according to claim 1, wherein The main component of the plastic is polystyrene, polyvinyl chloride or polyethylene terephthalate.

4. The method according to claim 3, wherein The main component of the plastic is polystyrene or polyvinyl chloride.

5. The method according to claim 1, wherein In step S1, the time of the ultrasonic treatment is 5-30 min.

6. The method according to claim 1, wherein In step S1, the standing time is 2-12 hours.

7. The method according to claim 1, wherein In step S3, 400-600 μL of samples y1 and y2 are respectively taken for density detection with a flow cytometer.

8. The method according to claim 1, wherein In step S3, the single-test time of the flow cytometer is 20-40 seconds, and the sample flow rate is 20-40 μL / min.

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