A method for analyzing a pollution toxicity source of coastal high-salinity underground water

Abstract

The application discloses a kind of coastal high-salinity groundwater pollution toxicity source analytical methods, comprising the following steps: S1, determine target pollutant and target form of target pollutant;S2, preparation simulation water sample: S3, determine the toxicity of each target pollutant;S4, determine the main toxicity source.The application uses Vibrio fischeri luminescence method to analyze the toxicity source of coastal high-salinity groundwater pollution, which can quickly indicate the comprehensive ecological toxicity effect of contaminated water body, define the contribution degree of main pollution components in water body to toxicity, effectively identify the pollution components that should be given priority to control in groundwater environmental pollution from the perspective of aquatic ecological toxicity and risk control, and provide scientific support for coastal groundwater environmental multi-component pollution control.

Description

Technical Field

[0001] This invention relates to the field of groundwater detection technology, specifically to a method for analyzing the toxicity sources of pollution in coastal high-salinity groundwater. Background Technology

[0002] Coastal areas have abundant surface runoff, making it easy for land-based pollutants to flow into the ocean via hydraulic action. Simultaneously, the region boasts excellent water accessibility and convenient transportation, attracting numerous chemical enterprises and industrial parks. Influenced by both industrial activities and hydrological conditions, groundwater in coastal areas exhibits high salinity, complex pollutant components, and diverse pollutant forms, posing significant risks to the groundwater ecosystem and necessitating targeted research on pollution identification and control.

[0003] Vibrio fischeri possesses sensitive bioluminescent properties, enabling rapid response to water pollution toxicity, making it suitable for water quality monitoring or pollutant toxicity assessment. Because it originates from marine environments, it is particularly suitable for pollution identification and control in highly saline environments.

[0004] However, existing toxicity detection methods based on Vibrio fischeri usually only assess the overall toxicity effect of a single pollutant or only compare the toxicity between different pollutants. They fail to establish a correlation between the toxicity of pollutants and their actual occurrence forms in complex high-salinity groundwater environments, such as soluble and insoluble forms, different valence states, or different coordination forms. This makes it difficult to accurately identify the specific toxic source forms, resulting in a lack of precise targeting in pollution source control.

[0005] For pollutant dispersion systems where soluble and sparingly soluble forms coexist, or multiple soluble forms coexist, the entire suspension is usually tested, or only the toxicity of each individual form is measured separately. There is a lack of a comprehensive analytical method that can systematically separate the toxicity contribution of each form in the mixed system and identify the preferred control form accordingly. Therefore, in coastal high-salinity groundwater environments with complex pollutant forms, it is difficult to scientifically determine the priority of remediation, which is detrimental to the efficient allocation of limited environmental resources.

[0006] For example, patent publication number CN201010580821 discloses a method for quantitatively detecting water toxicity, comprising: culturing luminescent bacteria in LB solid medium, then inoculating the cultured bacteria into liquid LB medium for further culture, centrifuging the liquid medium containing the luminescent bacteria, suspending it in sterile physiological saline to obtain a bacterial suspension with an OD value of 0.4-0.9; and reacting the bacterial suspension with different concentrations of Cr... 6+ The solution was subjected to a luminescent bacteria toxicity test, and Cr was obtained. 6+ A standard curve for the luminescence inhibition rate was obtained; the water sample to be tested was mixed with the bacterial suspension for a luminescent toxicity test to obtain the luminescent inhibition rate. The luminescent inhibition rate of the water sample to be tested was then substituted into the standard curve, with Cr... 6+Concentration indicates the toxicity of the water sample. The patent claims to have selected Cr... 6+ As a standard toxic substance, the results are consistent and reproducible. Furthermore, by treating the luminescent bacteria, both organic and inorganic toxic substances can inhibit the luminescence intensity of the bacteria, making the assessment of the overall toxicity of water more accurate. However, this patented method does not address high-salinity groundwater background or pollutant speciation; therefore, there may be biases in the analysis of pollution toxicity sources. Summary of the Invention

[0007] To address the aforementioned problems, this invention provides a method for analyzing the toxicity sources of high-salinity groundwater pollution in coastal areas.

[0008] The technical solution of this invention is: A method for apportioning the toxicity sources of high-salinity groundwater pollution in coastal areas includes the following steps: S1. Determine the target pollutants and their target forms: Take coastal high-salinity groundwater as the original water sample, detect the pollutant categories and their concentrations in the original water sample, and select the pollutants with the highest mass concentration as the target pollutants based on the detection results. When a target pollutant exists in multiple soluble and multiple insoluble forms, the highest concentration of the soluble form and the highest concentration of the insoluble form are selected as the two target forms of the pollutant, denoted as soluble form M and insoluble form N, respectively. When the target pollutant has multiple soluble states, the two soluble states with the highest mass concentration are selected as the two target forms of the target pollutant, and are respectively denoted as the soluble state with relatively high mass concentration S and the soluble state with relatively low mass concentration T. S2. Preparation of simulated water samples: For each of the aforementioned target pollutants: ① Prepare a simulated water sample with the same mass concentration of the target pollutant as the original water sample, and designate it as the first simulated water sample; ② Prepare a simulated water sample with the same mass concentration as the two target forms of the target pollutant in the original water sample, and denoted as the second simulated water sample; ③ Prepare a simulated water sample with the same mass concentration as each target soluble form M of the target pollutant in the original water sample, and record it as the third simulated water sample; prepare a simulated water sample with the same mass concentration as each target soluble form S of the target pollutant in the original water sample, and record it as the fourth simulated water sample; S3. Determine the toxicity of each target pollutant: Conduct a toxicity test on the first simulated water sample using Vibrio fischeri bacterial solution to obtain the luminescence intensity of the first simulated water sample. D 、 And the luminescence intensity of the 3% NaCl bacterial solution blank control. D0 The luminescence inhibition rate of each target pollutant in the first simulated water sample on Vibrio fischeri was calculated according to formula (1). E(x) : (1) In the formula: E(x) The value represents the inhibition rate of bioluminescence by the target pollutant on Vibrio fischeri. A positive value indicates inhibition of bioluminescence, i.e., toxicity, while a negative value indicates promotion of bioluminescence, i.e., the hormesis effect. D x This is the luminescence intensity reading for the first simulated water sample. D 0 The reading is the luminescence intensity of the 3% NaCl bacterial solution; According to each target pollutant E(x), Determine the toxicity of each target pollutant, when the luminescence inhibition rate... E(x) When the value is positive, the larger the value, the stronger the toxicity; when the value is negative, the smaller the absolute value, the stronger the toxicity. The toxicity of each target pollutant is compared. S4. Identify the main sources of toxicity: Measure the luminescence intensity of the second simulated water sample. D (mn) , D (st) The luminescence intensity of the third simulated water sample D (m) The luminescence intensity of the fourth simulated water sample D (s) ; Then, based on the above luminescence intensity, determine the luminescence inhibition rate of Vibrio fischeri for each target morphology in each simulated water sample. The toxic contribution of insoluble N or soluble T is calculated by back-calculation according to formula (2). The luminescence inhibition rate of the two target forms of the same pollutant is compared. The target form with the larger inhibition rate is determined as the preferred control form, i.e. the main source of toxicity. E(xy)=1 - [1 - E(x)]·[1 - E(y)] (2) In the formula: E(xy) The combined luminescence inhibition rate of Vibrio fischeri for two target forms of the same pollutant. E(x) The emission suppression rate is the emission rate when the target morphology X exists alone, where X is either soluble M or soluble S. E(y) The emission suppression rate is given by the target morphology Y when it exists alone, where Y is either the sparingly soluble N or the soluble T, calculated by reverse deduction. E(y) .

[0009] Furthermore, the preparation method of the Vibrio fischeri bacterial solution is as follows: after thawing and reviving the lyophilized Vibrio fischeri powder, it is inoculated onto a solid culture medium and cultured at 20±2℃ in the dark for 24±4h. Then, it is transferred to a liquid culture medium and cultured at 20±2℃ and 160~200rpm in the dark for 12~16h.

[0010] Note: By optimizing and controlling the culture temperature, time, rotation speed, and light-protection conditions, Vibrio fischeri is ensured to achieve stable luminescence performance under optimal growth conditions, providing highly active and reproducible bacterial solutions for subsequent toxicity testing, thereby improving the reliability and stability of toxicity test results.

[0011] Furthermore, the components and concentrations of the liquid culture medium are as follows: NaCl 28~32g / L, MgSO4·7H2O 0.1~0.3g / L, NaH2PO4·2H2O 6.5~7g / L, K2HPO4·3H2O 2.5~3g / L, (NH4)2HPO4 0.4~0.6g / L, peptone 4~6g / L, yeast extract 0.4~0.6g / L, glycerol 0.2~0.4mL / L, with the balance being water, and the pH is 7±0.2. The solid culture medium is based on the liquid culture medium with the addition of 10~15g / L agar.

[0012] Note: To address the marine origin of Vibrio fischeri, the salinity and nutrient composition of the culture medium were optimized to more closely resemble the natural marine growth environment of Vibrio fischeri, promoting rapid cell proliferation and maintaining stable bioluminescent metabolic activity. Simultaneously, the formulation correspondence between solid and liquid culture media was clarified to facilitate the integration of strain preservation and large-scale cultivation.

[0013] Furthermore, in S1, the target pollutants are 3 to 4 types.

[0014] Note: Limiting the number of target pollutants to 3-4 types not only covers high-risk pollutants but also controls the experimental scale and workload, enabling efficient completion of toxicity source tracing analysis under limited resource conditions and improving the practicality and operability of the method.

[0015] Furthermore, in S2, when the target pollutant is a heavy metal, the chloride of the heavy metal is selected to prepare the corresponding simulated water sample.

[0016] Note: Chlorides are used for heavy metal pollutants to improve their solubility and stability in the compound system.

[0017] Further, in S4, determining the luminescence inhibition rate of Vibrio fischeri corresponding to each target morphology in each simulated water sample based on the above-mentioned luminescence intensity includes: The luminescence inhibition rates of the two target forms of each target pollutant in the second simulated water sample against Vibrio fischeri were calculated according to formula (1-1).E(mn) : (1-1) In the formula: E(mn) The combined luminescence inhibition rate of two target forms of the target pollutant against Vibrio fischeri. D mn The luminescence intensity reading is for the second simulated water sample; The luminescence inhibition rate of the target speciation soluble M of each target pollutant in the third simulated water sample against Vibrio fischeri was calculated according to formula (1-2). E(m) : (1-2) In the formula: E(m) The luminescence inhibition rate of the target pollutant, soluble form M, against Vibrio fischeri. D m The luminescence intensity reading is for the third simulated water sample; The luminescence inhibition rates of the two target forms of each target pollutant in the second simulated water sample against Vibrio fischeri were calculated according to formula (1-3). E(st) : (1-3) In the formula: E(st) The combined luminescence inhibition rate of two target forms of the target pollutant against Vibrio fischeri. D st The luminescence intensity reading is for the second simulated water sample; The luminescence inhibition rate of the target speciation soluble S of each target pollutant in the fourth simulated water sample against Vibrio fischeri was calculated according to formula (1-4). E(s) : (1-4) In the formula: E(s) The luminescence inhibition rate of the target pollutant, soluble S, against Vibrio fischeri. D m This is the luminescence intensity reading for the fourth simulated water sample.

[0018] Note: The luminescence inhibition rate of Vibrio fischeri under the condition of coexistence of two target forms of the target pollutant can be calculated by various modified formulas of formula (1). E(mn) or E(st) 1. Spectrophotometric inhibition rate of soluble M against Vibrio fischeri E (m) And the luminescence inhibition rate of soluble S on Vibrio fischeri. E(s) This allows for subsequent calculations to obtain the remaining parameters.

[0019] Furthermore, in S4, the luminescence inhibition rate of sparingly soluble N on Vibrio fischeri is calculated according to formula (2-1): (2-1) In the formula: E(n) The luminescence inhibition rate of insoluble N on Vibrio fischeri; Compare E(m) and E(n) Size: If E(m) > E(n) If the target form, soluble M, is the preferred controlled form, i.e., the main source of toxicity; if E(n) > E(m) In this case, sparingly soluble nitrogen is the preferred controlled form, i.e., the main source of toxicity; The luminescence inhibition rate of soluble T against Vibrio fischeri was calculated according to formula (2-2): (2-2) In the formula: E(t) The luminescence inhibition rate of soluble T against Vibrio fischeri; Compare E(s) and E(t) Size: If E(s) > E(t) If soluble sulfur is the preferred controlled form, i.e., the main source of toxicity; if E(t) > E(s) In this case, soluble form T is the preferred controlled form, i.e., the main source of toxicity.

[0020] Note: Since the luminescence inhibition rate of sparingly soluble N cannot be calculated from the normal luminescence inhibition rate, it is calculated by mixing it with soluble M to obtain a more accurate luminescence inhibition rate of sparingly soluble N. Since soluble T may have the risk of inaccurate detection results due to low concentration, it is calculated by mixing it with soluble S to obtain a more accurate luminescence inhibition rate of soluble T, thus enabling a more precise comparison.

[0021] The beneficial effects of this invention are: This invention utilizes Vibrio fischeri luminescence to analyze the sources of toxicity in coastal high-salinity groundwater pollution. It can rapidly indicate the comprehensive ecotoxicological effects of polluted water bodies and define the degree of contribution of major pollutants to toxicity. From the perspective of aquatic ecotoxicity and risk control, it can effectively identify pollutants that should be prioritized for control in groundwater pollution, providing scientific support for the multi-component pollution control of coastal groundwater environments. Attached Figure Description

[0022] Figure 1 This is a schematic diagram of sample loading in a 96-well plate according to Embodiment 1 of the present invention. Detailed Implementation

[0023] Example 1: A method for apportioning the toxicity sources of high-salinity groundwater pollution in coastal areas, comprising the following steps: S1. Determine the target pollutants and their target forms: Take high-salinity coastal groundwater as the original water sample, and test the concentration of pollutants and their occurrence forms in the original water sample. Based on the test results, select the three pollutants with the highest mass concentration as target pollutants; simultaneously test the pH value, the concentration of conventional cations and conventional anions, and the occurrence forms, i.e., all soluble and all poorly soluble forms. The conventional cation is K+. + Ca 2+ Na + Mg 2+ Al 3+ The conventional anion is Cl. - NO3 - SO4 2- CO3 2- HCO3 - ; When a target pollutant exists in multiple soluble and multiple insoluble forms, the highest concentration of the soluble form and the highest concentration of the insoluble form are selected as the two target forms of the pollutant, denoted as soluble form M and insoluble form N, respectively. When the target pollutant has multiple soluble states, the two soluble states with the highest mass concentration are selected as the two target forms of the target pollutant, and are respectively denoted as the soluble state with relatively high mass concentration S and the soluble state with relatively low mass concentration T. S2. Preparation of simulated water samples: For each of the several target pollutants: ① Prepare a simulated water sample with the same mass concentration of the target pollutant as the original water sample, and record it as the first simulated water sample. When the target pollutant is a heavy metal, select the chloride of the heavy metal to prepare the corresponding simulated water sample. ② Prepare a simulated water sample with the same mass concentration as the two target forms of the target pollutant in the original water sample, and denoted as the second simulated water sample; ③ Prepare a simulated water sample with the same mass concentration as each target soluble form M of the target pollutant in the original water sample, and record it as the third simulated water sample; prepare a simulated water sample with the same mass concentration as each target soluble form S of the target pollutant in the original water sample, and record it as the fourth simulated water sample; S3. Determine the toxicity of each target pollutant: Conduct a toxicity test on the first simulated water sample using Vibrio fischeri bacterial solution to obtain the luminescence intensity of the first simulated water sample. D 、 And the luminescence intensity of the 3% NaCl bacterial solution blank control. D 0 The luminescence inhibition rate of each target pollutant on Vibrio fischeri in the first simulated water sample was calculated according to formula (1). E(x) : (1) In the formula: E(x) The value represents the inhibition rate of bioluminescence by the target pollutant on Vibrio fischeri. A positive value indicates inhibition of bioluminescence, i.e., toxicity, while a negative value indicates promotion of bioluminescence, i.e., the hormesis effect. D x This is the luminescence intensity reading for the first simulated water sample. D 0 The reading is the luminescence intensity of the 3% NaCl bacterial solution; According to each target pollutant E(x), Determine the toxicity of each target pollutant, when the luminescence inhibition rate... E(x) When the value is positive, the larger the value, the stronger the toxicity; when the value is negative, the smaller the absolute value, the stronger the toxicity. The toxicity of each target pollutant is compared. Figure 1 As shown, the luminescence intensity test was performed using a 96-well plate, with three replicates for each test. After standing in the dark for 40 minutes, the luminescence intensity was measured using a multi-functional microplate reader, and the average value of each replicate was taken as the luminescence intensity reading of that sample. The preparation method of Vibrio fischeri bacterial suspension is as follows: after thawing and reviving the lyophilized Vibrio fischeri powder, it is inoculated onto a solid culture medium and cultured at 20°C in the dark for 24 hours. Then it is transferred to a liquid culture medium and cultured at 20°C and 180 rpm in the dark for 14 hours. The components and concentrations of the liquid culture medium are as follows: NaCl 30 g / L, MgSO4·7H2O 0.2 g / L, NaH2PO4·2H2O 6.9 g / L, K2HPO4·3H2O 2.75 g / L, (NH4)2HPO4 0.5 g / L, peptone 5 g / L, yeast extract 0.5 g / L, glycerol 0.3 mL / L, with the remainder being water, and the pH is 7. The solid culture medium is based on the liquid culture medium with the addition of 12 g / L agar. S4. Identify the main sources of toxicity: Determine the luminescence intensity of the second simulated water sample. D (mn) , D (st) The luminescence intensity of the third simulated water sample D (m) The luminescence intensity of the fourth simulated water sample D (s) ; Then, based on the above luminescence intensity, determine the luminescence inhibition rate of Vibrio fischeri for each target morphology in each simulated water sample. The toxic contribution of insoluble N or soluble T is calculated by back-calculation according to formula (2). The luminescence inhibition rate of the two target forms of the same pollutant is compared. The target form with the larger inhibition rate is determined as the preferred control form, i.e. the main source of toxicity. E(xy)=1 - [1 - E(x)]·[1 - E(y)] (2) In the formula: E(xy) The combined luminescence inhibition rate of Vibrio fischeri for two target forms of the same pollutant. E(x) The emission suppression rate is the emission rate when the target morphology X exists alone, where X is either soluble M or soluble S. E(y) The emission suppression rate is given by the target morphology Y when it exists alone, where Y is either the sparingly soluble N or the soluble T, calculated by reverse deduction. E(y) ; Based on the above luminescence intensity, the inhibition rate of Vibrio fischeri luminescence by pollutants corresponding to each target morphology in each simulated water sample was determined, including: The luminescence inhibition rates of the two target forms of each target pollutant in the second simulated water sample against Vibrio fischeri were calculated according to formula (1-1). E(mn) : (1-1) In the formula: E(mn) The combined luminescence inhibition rate of two target forms of the target pollutant against Vibrio fischeri. D mn The luminescence intensity reading is for the second simulated water sample; The luminescence inhibition rate of the target speciation soluble M of each target pollutant in the third simulated water sample against Vibrio fischeri was calculated according to formula (1-2). E(m) : (1-2) In the formula: E(m) The luminescence inhibition rate of the target pollutant, soluble form M, against Vibrio fischeri. D m The luminescence intensity reading is for the third simulated water sample; The luminescence inhibition rates of the two target forms of each target pollutant in the second simulated water sample against Vibrio fischeri were calculated according to formula (1-3). E(st) : (1-3) In the formula: E(st) The combined luminescence inhibition rate of two target forms of the target pollutant against Vibrio fischeri. D st The luminescence intensity reading is for the second simulated water sample; The luminescence inhibition rate of the target speciation soluble S of each target pollutant in the fourth simulated water sample against Vibrio fischeri was calculated according to formula (1-4). E(s) : (1-4) In the formula: E(s) The luminescence inhibition rate of the target pollutant, soluble S, against Vibrio fischeri. Dm This is the luminescence intensity reading for the fourth simulated water sample; The luminescence inhibition rate of insoluble nitrogen against Vibrio fischeri was calculated according to formula (2-1): (2-1) In the formula: E(n) The luminescence inhibition rate of insoluble N on Vibrio fischeri; Compare and E(m) Size: If E(n) > E(m) If the target form, soluble M, is the preferred controlled form, i.e., the main source of toxicity; if E(n) In this case, sparingly soluble nitrogen is the preferred controlled form, i.e., the main source of toxicity; The luminescence inhibition rate of soluble T against Vibrio fischeri was calculated according to formula (2-2): (2-2) In the formula: E(n) > E(m) The luminescence inhibition rate of soluble T against Vibrio fischeri; Compare E(t) and E(s) Size: If E(t) If soluble sulfur is the preferred controlled form, i.e., the main source of toxicity; if E(s) > E(t) E(t) > E(s) In this case, soluble form T is the preferred controlled form, i.e., the main source of toxicity.

[0024] Example 2: The difference between this example and Example 1 is that four pollutants were screened out.

[0025] Example 3: The difference between this example and Example 1 is that after the freeze-dried Vibrio fischeri powder is thawed and revived, it is inoculated onto a solid culture medium and cultured at 18°C ​​in the dark for 20 hours. Then it is transferred to a liquid culture medium and cultured at 18°C ​​and 160 rpm in the dark for 16 hours.

[0026] Example 4: The difference between this example and Example 1 is that after the lyophilized Vibrio fischeri powder is thawed and revived, it is inoculated onto a solid culture medium and cultured at 22°C in the dark for 28 hours. Then it is transferred to a liquid culture medium and cultured at 22°C and 200 rpm in the dark for 12 hours.

[0027] Example 5: This example differs from Example 1 in that the components and concentrations of the liquid culture medium are as follows: NaCl 28g / L, MgSO4·7H2O 0.1g / L, NaH2PO4·2H2O 6.5g / L, K2HPO4·3H2O 2.5g / L, (NH4)2HPO4 0.4g / L, peptone 4g / L, yeast extract 0.4g / L, glycerol 0.2mL / L, with the remainder being water, and the pH is 6.8. The solid culture medium is based on the liquid culture medium with the addition of 10g / L agar.

[0028] Example 6: This example differs from Example 1 in that the components and concentrations of the liquid culture medium are as follows: NaCl 32g / L, MgSO4·7H2O 0.3g / L, NaH2PO4·2H2O 7g / L, K2HPO4·3H2O 3g / L, (NH4)2HPO4 0.6g / L, peptone 6g / L, yeast extract 0.6g / L, glycerol 0.4mL / L, with the remainder being water, and the pH is 7.2. The solid culture medium is based on the liquid culture medium with the addition of 15g / L agar.

[0029] Example 7: The difference between this example and Example 1 is that in S3, Vibrio fischeri bacterial solution was used to test the toxicity of each simulated water sample, and luminescence was tested using a 96-well plate. Each test was performed in triplicate. After standing in the dark for 30 minutes, the luminescence intensity was measured using a multi-functional microplate reader.

[0030] Experimental Example: Using CuO NPs and ZnO NPs as research objects, and groundwater from two different coastal areas (A and B) as actual exposure environments, this study simulated groundwater pollution caused by 100 mg / L CuO NPs and ZnO NPs during a 72-hour emergency response period following a leak. The study employed the 30-minute acute toxicity test using Vibrio fischeri from Example 1, combined with analysis of nanoparticle leachates (Cu... 2+ Zn 2 + The concentration of the two nanoparticles was determined to analyze their aquatic ecotoxicity effects and main sources of toxicity within 72 hours after leakage.

[0031] In S1, Vibrio fischeri are cultured in advance and can be used for acute toxicity testing by stable passage.

[0032] In S2, corresponding to S2-1, the original water sample was tested: high-salinity groundwater was collected from two coastal areas, and the pH value and common cations (K) were measured. + Ca 2+ Na + Mg 2+ Al 3+ ) and anions (Cl - NO3 -SO4 2- CO3 2- HCO3 - ) concentration, Cu 2+ and Zn 2+ The concentration.

[0033] Table 1 Physicochemical properties of groundwater samples A and B

[0034] S2-2. Determine the target pollutant and its target form: To identify the preferred controlled form of the pollutant, in a dispersion system where soluble and insoluble CuO NPs coexist, the soluble form with the higher concentration (CuO NPs) is selected. 2+ The target morphology is the insoluble state (CuO nanoparticles). In a dispersion system where soluble and insoluble ZnO NPs coexist, the soluble state (ZnO nanoparticles) with a higher concentration is selected. 2+ The target morphology is the insoluble state (ZnO nanoparticle state). S2-3. Preparation of simulated water samples: Based on the physicochemical properties of the original groundwater samples A and B, appropriate inorganic salts are calculated and weighed, dissolved and dispersed in deionized water, and then diluted to 1L to ensure that the pH value, conventional cation and anion concentrations are the same as those of the original water samples, thus obtaining two simulated water samples.

[0035] Preparation of compound dispersion systems: The concentration of pollutants (CuO NPs and ZnO NPs) was set at 100 mg / L, and the concentration of the nanoparticle suspension was also set at 100 mg / L. This was mainly because the nanoparticles might reach this concentration level in a pollution leak event, which has practical significance for environmental risk assessment. Appropriate amounts of nanoparticles were weighed and added to groundwater simulation systems A and B, respectively. The mixtures were ultrasonically vibrated (150 W, 38 kHz) for 20 min to ensure uniform particle dispersion. The suspensions were prepared and used immediately. The four compound dispersion systems (A CuO NPs, A ZnO NPs, B CuO NPs, and B ZnO NPs) were continuously subjected to constant-temperature magnetic stirring (20℃, 200 r / min) to simulate the hydraulic conditions of groundwater in coastal areas.

[0036] Determination of soluble concentration: 15 mL samples were taken at 24, 48, and 72 h, respectively. After high-speed centrifugation (20000 g, 20℃, 40 min), the supernatant was collected and filtered sequentially through a 0.22 μm aqueous filter and a 0.02 μm cellulose acetate membrane. The filtrate was then determined using inductively coupled plasma atomic emission spectrometry (ICP-AES). 2+ Zn 2+ Concentration, with 3 parallel samples at each time point.

[0037] Table 2 Cu in coastal groundwater suspension dispersion system2+ and Zn 2+ Dissolution concentration (unit: mg / L)

[0038] S3 bacterial suspension toxicity test The luminescence intensity was measured using a 96-well plate. Suspension and filtrate samples were collected at 24, 48, and 72 h, with three replicates for each test. The samples were added to the bacterial culture to be tested, and after standing in the dark for 30 min, the luminescence intensity was measured using a multi-mode microplate reader. The luminescence intensity was read and the average value was taken to obtain the luminescence intensity readings D(total), D(ion), and D0 of the 3% NaCl bacterial culture.

[0039] The luminescence inhibition rate of Vibrio fischeri in the suspension and filtrate of the water sample is calculated according to formula (1). E (total) represents the total toxicity of the suspension (including soluble and insoluble forms) and E (ion) represents the ion toxicity of the soluble form.

[0040] To identify the form (soluble and insoluble) of the pollutants that are to be prioritized for control, the particulate toxicity E (particle) of the insoluble form is calculated using formula (2) in combination with the concentration of the pollutant form, and the pollutants that are the main sources of toxicity are analyzed.

[0041] Table 3. Effects of 100 mg / L CuO NPs and ZnO NPs on the luminescence intensity of Vibrio fischeri in coastal groundwater (positive values ​​indicate inhibition, negative values ​​indicate promotion).

[0042] Data analysis shows that for CuO NPs, the Cu in groundwater of A and B... 2+ The maximum leaching amounts were 0.8 mg / L (72 h) and 0.6 mg / L (72 h), respectively. The leaching of CuO NPs in the two groundwater simulation systems... 2+ At low concentrations (<1 mg / L), acute toxicity primarily originates from the nanoparticles themselves. Their aggregation and sedimentation behavior may influence the contact efficiency between the particles and Vibrio fischeri, thereby dominating the toxic effect.

[0043] For ZnO NPs, A Zn in groundwater 2+ The leaching concentration of 7 mg / L (72 h) was significantly higher than that in groundwater B (2.5 mg / L (72 h)); there were significant differences in the toxic effects of ZnO NPs between groundwater B and groundwater A: in groundwater B, the ZnO NPs suspension (E(total)) and the leached Zn 2+The values ​​of E(ion) were all negative, indicating that they promoted the luminescence of Vibrio fischeri, while the toxicity of the ZnO NPs particles themselves (E(particle)) contributed very little. In groundwater group A, the toxicity of ZnO NPs increased significantly over time, reaching 0.99 (nearly lethal) at 48 h and 1.0 (lethal) at 72 h, corresponding to the leached Zn 2+ The E(ion) also increased synchronously, reaching 0.97 at 48 h and 1.0 at 72 h, while the E(particle) remained below 0.07, indicating that Zn 2+ It is the dominant factor in toxicity.

[0044] In summary, the studies indicate that the sources of toxicity for CuO NPs and ZnO NPs differ significantly between groundwater environments A and B. The acute toxicity of CuO NPs primarily originates from the nanoparticles themselves, while the acute toxicity of ZnO NPs mainly stems from the leached Zn. 2+ This is closely related to its own physicochemical properties and aquatic environmental conditions, providing experimental basis for the aquatic ecological risk assessment and precise pollution control of nano-metal oxides.

Claims

1. A method for analyzing the toxicity sources of high-salinity coastal groundwater pollution, characterized in that, Includes the following steps: S1. Determine the target pollutants and their target forms: Take coastal high-salinity groundwater as the original water sample, detect the pollutant categories and their concentrations in the original water sample, and select the pollutants with the highest mass concentration as the target pollutants based on the detection results. When a target pollutant exists in multiple soluble and multiple insoluble forms, the highest concentration of the soluble form and the highest concentration of the insoluble form are selected as the two target forms of the pollutant, denoted as soluble form M and insoluble form N, respectively. When the target pollutant has multiple soluble states, the two soluble states with the highest mass concentration are selected as the two target forms of the target pollutant, and are respectively denoted as the soluble state with relatively high mass concentration S and the soluble state with relatively low mass concentration T. S2. Preparation of simulated water samples: For each of the aforementioned target pollutants: ① Prepare a simulated water sample with the same mass concentration of the target pollutant as the original water sample, and designate it as the first simulated water sample; ② Prepare a simulated water sample with the same mass concentration as the two target forms of the target pollutant in the original water sample, and denoted as the second simulated water sample; ③ Prepare a simulated water sample with the same mass concentration as each target soluble form M of the target pollutant in the original water sample, and record it as the third simulated water sample; prepare a simulated water sample with the same mass concentration as each target soluble form S of the target pollutant in the original water sample, and record it as the fourth simulated water sample; S3. Determine the toxicity of each target pollutant: Conduct a toxicity test on the first simulated water sample using Vibrio fischeri bacterial solution to obtain the luminescence intensity of the first simulated water sample. D 、 And the luminescence intensity of the 3% NaCl bacterial solution blank control. D 0 The luminescence inhibition rate of each target pollutant on Vibrio fischeri in the first simulated water sample was calculated according to formula (1). E(x) : （1） In the formula: E(x) The value represents the inhibition rate of bioluminescence by the target pollutant on Vibrio fischeri. A positive value indicates inhibition of bioluminescence, i.e., toxicity, while a negative value indicates promotion of bioluminescence, i.e., the hormesis effect. D x This is the luminescence intensity reading for the first simulated water sample. D 0 The reading is the luminescence intensity of the 3% NaCl bacterial solution; According to each target pollutant E(x), Determine the toxicity of each target pollutant, when the luminescence inhibition rate... E(x) When the value is positive, the larger the value, the stronger the toxicity; when the value is negative, the smaller the absolute value, the stronger the toxicity. The toxicity of each target pollutant is compared. S4. Identify the main sources of toxicity: Measure the luminescence intensity of the second simulated water sample. D (mn) , D (st) The luminescence intensity of the third simulated water sample D (m) The luminescence intensity of the fourth simulated water sample D (s) ; Then, based on the above luminescence intensity, determine the luminescence inhibition rate of Vibrio fischeri corresponding to each target morphology in each simulated water sample on Vibrio fischeri. The toxic contribution of insoluble N or soluble T is calculated by back-calculation according to formula (2). The luminescence inhibition rate of the two target forms of the same pollutant is compared. The target form with the larger inhibition rate is determined as the preferred control form, i.e. the main source of toxicity. E(xy) = 1 - [1 - E(x)]·[1 - E(y)] （2） In the formula: E(xy) The combined luminescence inhibition rate of Vibrio fischeri for two target forms of the same pollutant. E(x) The emission suppression rate is the emission rate of target morphology X when it exists alone, where X is either soluble M or soluble S. E(y) The emission suppression rate is given by the target morphology Y when it exists alone, where Y is either the sparingly soluble N or the soluble T, calculated by reverse deduction. E(y) .

2. The method for apportioning the toxicity sources of high-salinity coastal groundwater pollution according to claim 1, characterized in that, The preparation method of Vibrio fischeri bacterial solution is as follows: after thawing and reviving the freeze-dried Vibrio fischeri powder, it is inoculated onto a solid culture medium and cultured at 20±2℃ in the dark for 24±4h. Then, it is transferred to a liquid culture medium and cultured at 20±2℃ and 160~200rpm in the dark for 12~16h.

3. The method for analyzing the toxicity sources of high-salinity coastal groundwater pollution according to claim 2, characterized in that, The liquid culture medium comprises the following components and concentrations: NaCl 28~32 g / L, MgSO4·7H2O 0.1~0.3 g / L, NaH2PO4·2H2O 6.5~7 g / L, K2HPO4·3H2O 2.5~3 g / L, (NH4)2HPO4 0.4~0.6 g / L, peptone 4~6 g / L, yeast extract 0.4~0.6 g / L, glycerol 0.2~0.4 mL / L, with the balance being water, and a pH of 7±0.

2. The solid culture medium is prepared by adding 10~15 g / L of agar to the liquid culture medium.

4. The method for apportioning the toxicity sources of high-salinity coastal groundwater pollution according to claim 1, characterized in that, In S1, the target pollutants are 3 to 4 types.

5. The method for apportioning the toxicity sources of high-salinity coastal groundwater pollution according to claim 1, characterized in that, In S2, when the target pollutant is a heavy metal, the chloride of the heavy metal is selected to prepare the corresponding simulated water sample.

6. The method for apportioning the toxicity sources of high-salinity coastal groundwater pollution according to claim 1, characterized in that, In S4, determining the luminescence inhibition rate of Vibrio fischeri for each target morphology in each simulated water sample based on the above-mentioned luminescence intensity includes: The luminescence inhibition rates of the two target forms of each target pollutant in the second simulated water sample against Vibrio fischeri were calculated according to formula (1-1). E(mn) : （1-1） In the formula: E(mn) The combined luminescence inhibition rate of two target forms of the target pollutant against Vibrio fischeri. D mn The luminescence intensity reading is for the second simulated water sample; The luminescence inhibition rate of the target speciation soluble M of each target pollutant in the third simulated water sample against Vibrio fischeri was calculated according to formula (1-2). E(m) : （1-2） In the formula: E(m) The luminescence inhibition rate of the target pollutant, soluble form M, against Vibrio fischeri. D m The luminescence intensity reading is for the third simulated water sample; The luminescence inhibition rates of the two target forms of each target pollutant in the second simulated water sample against Vibrio fischeri were calculated according to formula (1-3). E(st) : （1-3） In the formula: E(st) The combined luminescence inhibition rate of two target forms of the target pollutant against Vibrio fischeri. D st The luminescence intensity reading is for the second simulated water sample; The luminescence inhibition rate of the target speciation soluble S of each target pollutant in the fourth simulated water sample against Vibrio fischeri was calculated according to formula (1-4). E(s) : （1-4） In the formula: E(s) The luminescence inhibition rate of the target pollutant, soluble S, against Vibrio fischeri. D m This is the luminescence intensity reading for the fourth simulated water sample.

7. The method for apportioning the toxicity sources of high-salinity coastal groundwater pollution according to claim 6, characterized in that, In S4, the luminescence inhibition rate of insoluble N on Vibrio fischeri is calculated according to formula (2-1): （2-1） In the formula: E(n) The luminescence inhibition rate of insoluble N on Vibrio fischeri; Compare E(m) and E(n) Size: If E(m) > E(n) If the target form, soluble M, is the preferred controlled form, i.e., the main source of toxicity; if E(n) > E(m) In this case, sparingly soluble nitrogen is the preferred controlled form, i.e., the main source of toxicity; The luminescence inhibition rate of soluble T against Vibrio fischeri was calculated according to formula (2-2): （2-2） In the formula: E(t) The luminescence inhibition rate of soluble T against Vibrio fischeri; Compare E(s) and E(t) Size: If E(s)>E(t) If soluble sulfur is the preferred controlled form, i.e., the main source of toxicity; if E(t)>E(s) In this case, soluble form T is the preferred controlled form, i.e., the main source of toxicity.

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