CN112198150B - Method for monitoring polluted water body by bioluminescence method - Google Patents
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Abstract
The invention belongs to the technical field of biological environment monitoring, and discloses a method for monitoring a polluted water body by a bioluminescence method, which comprises the following steps: step 1) preparing a rejuvenation culture solution, step 2) rejuvenating luminescent bacteria, and step 3) monitoring sewage. The bioluminescence method disclosed by the invention is rapid and sensitive in monitoring the polluted water body and has a wide market application prospect.
Description
Technical Field
The invention belongs to the technical field of biological environment monitoring, and particularly relates to a method for monitoring a polluted water body by a bioluminescence method.
Background
In recent years, with the improvement of the industrialization process and the great demand for chemical products, the number of various chemical substances is increased dramatically, which has great influence on the living environment of human beings, and the acute toxicity identification of the environmental pollutants which are increased day by day is urgently needed. Environmental monitoring methods can be divided into two categories, physical and chemical analysis techniques and biological monitoring. The physical and chemical analysis technology is often used for testing single physical and chemical indexes, but cannot reflect the toxicity of pollutants. In the traditional biological monitoring, daphnia, algae or fish and the like are taken as subjects, and although the direct influence of poisons on organisms can be reflected, the methods have the biggest defects of long experimental period and tedious experiment. Aiming at the defects of the traditional biotoxicity detection method, the luminous bacteria method is being widely applied as a novel biotoxicity monitoring technology, and attracts the attention of the vast environmental-friendly scientific researchers.
The luminous bacteria method is to utilize a sensitive photoelectric measurement system to determine the influence of poison on the luminous intensity of luminous bacteria. Luminescent bacteria contain luminescent elements such as fluorescein and luciferase, and produce weak fluorescence by intracellular biochemical reactions under aerobic conditions. The use of luminescent bacteria to monitor water pollutants has been increasingly applied to practical applications.
Document CN109946433A discloses a sewage detection method comprising: taking a sewage sample at a sewage discharge position, extracting part of the sewage sample by using dichloromethane, dissolving the part of the sewage sample by using DMSO, placing the part of the sewage sample into a cold box for low-temperature storage, and detecting the part of the sewage sample by using a luminous bacterial toxicity test; if the toxicity of the sewage is not detected, the sewage can be directly recycled; if the toxicity of the sewage is detected, uniformly spraying a sewage treatment agent at the sewage port, and sampling again for detection; if the sewage is detected to have toxicity, the sewage sample put into the refrigerator is subjected to deep detection, and if the sewage at the undetected position has toxicity, the sewage can be recycled.
Document CN108507999a discloses a biotoxicity detection method applied to biotechnology, which comprises the following steps: sampling a target water body; preparing a microbial liquid; and (4) detecting biological toxicity. The target water body is detected through the luminous bacteria, and compared with a daphnia biotoxicity test, an algae toxicity test and a fish toxicity test, the luminous bacteria biotoxicity test has the advantages of short period, good accuracy, wide monitoring range and any detection place.
In actual operation, the luminescent bacteria are generally stored in a freeze-dried powder form, resuscitation and rejuvenation are carried out when the luminescent bacteria are used, the number and the vitality of strains are recovered according to the process, the strains die due to improper resuscitation process, the brightness is not enough, toxicity test is difficult to carry out, and components in resuscitation culture solution may cause certain influence on the luminescence of the strains. The commonly used bacterial strains in the prior art comprise vibrio qinghai, vibrio fischeri, photobacterium leiognathi and the like, but different photobacterium have larger difference in recovery and culture process due to different taxonomy and physicochemical properties.
The prior art has some records on the recovery and rejuvenation culture of the luminescent bacteria.
Document CN102465097a, developed through the development of specialized bacteria resuscitation solutions, include inorganic components (such as sodium chloride, potassium dihydrogen phosphate, disodium hydrogen phosphate, potassium chloride, etc.) and organic components (including tryptone, yeast extract, amino acids, glycerol, vitamins, glucose, etc.). However, the resuscitation solution has no specific component proportion relationship, and cannot be practically applied.
The document CN102604873A discloses a rejuvenation method of marine luminous bacteria, and the main components of a rejuvenation culture medium comprise 2-8g/L of casein hydrolase, 20-25g/L of sodium glycerophosphate and 2.5g/L of yeast extract. The rejuvenation culture medium mainly aims at Photobacterium brightens and Vibrio fischeri, and is not suitable for Photobacterium leiognathi.
Document CN101874105a discloses a media composition comprising: 1.0-3.0% NaCl/KCl, 0.5-7.0% inositol/lactose/trehalose/dextran, 0.0-300mM Mg/Ca, 0.01-0.05% yeast extract/acid hydrolyzed casein, 0.02-1.0% bovine serum albumin/ovalbumin and 0.1-3.5% ethanol/methanol/propanol; the luminogenic bacteria suspension was stored for 2 weeks under conditions to maintain the luminogenic bacteria viable and prevent expansion of the bacteria. The preservation solution mainly aims at maintaining the activity of the bacterial strain, but has little practical significance in application, freeze-dried powder generally needs to be revived and rejuvenated in practical operation, and the luminous activity of the bacteria is quickly restored and the death of the bacteria is reduced in the reviving and rejuvenating process, so that the luminous bacteria can be applied to monitoring pollutants in the environment in the shortest time.
Disclosure of Invention
In order to overcome the defects in the prior art, the invention provides a method for monitoring a polluted water body by a bioluminescence method, and the method can quickly and efficiently monitor pollutants in the water body.
The invention is realized by the following technical scheme.
A method for monitoring a polluted water body by a bioluminescence method comprises the following steps: step 1) preparing a rejuvenation culture solution, step 2) rejuvenating luminescent bacteria, and step 3) monitoring sewage.
Further, the method comprises the steps of:
step 1) preparing a rejuvenation culture solution: sequentially adding sodium chloride, magnesium chloride, adenosine triphosphate, methionine, calcium acetate and sterile defibrinated sheep blood into distilled water, mixing, and sterilizing;
step 2) rejuvenation of luminescent bacteria: placing photobacterium leiognathus freeze-dried powder in a water bath at 37 ℃ for rapid thawing, then placing the rejuvenation culture solution obtained in the step 1) in an incubator at 32 ℃ for shake culture at 200rpm for 20-40min to obtain a bacterial solution;
step 3), sewage monitoring: placing a water sample to be tested in a testing container, adding sodium chloride, controlling the concentration of the sodium chloride to be 30g/L, and then accessing the bacterial liquid obtained in the step 2); placing 30g/L sodium chloride aqueous solution in a control container, and then inoculating a bacterial solution; controlling the density of the thallus in the test container and the density of the thallus in the comparison container to be the same; placing in biochemical luminescence tester for 15min, and measuring luminescence intensity.
Further, the rejuvenation culture solution comprises the following components:
10-30g/L of sodium chloride,
4-7g/L of magnesium chloride,
3-5g/L of adenosine triphosphate,
1-2g/L of methionine,
0.5-1g/L of calcium acetate,
30-50ml/L of aseptic defibered sheep blood.
Preferably, the rejuvenation culture solution has the following components:
26g/L of sodium chloride is added into the solution,
the magnesium chloride is 6g/L,
the concentration of the adenosine triphosphate is 4g/L,
1g/L of methionine is added into the mixture,
0.8g/L of calcium acetate,
35ml/L of sterile defibered sheep blood.
Preferably, the rejuvenation culture solution has the following components:
the concentration of the sodium chloride is 18g/L,
the magnesium chloride is 5g/L,
3g/L of adenosine triphosphate is added,
1g/L of methionine is added into the mixture,
0.5g/L of calcium acetate,
30ml/L of sterile defibered sheep blood.
Preferably, the rejuvenation culture solution has the following components:
the concentration of the sodium chloride is 20g/L,
the magnesium chloride is 6g/L,
adenosine triphosphate of 5g/L is added into the solution,
2g/L of methionine is added, and the content of methionine is 2g/L,
1g/L of calcium acetate is added,
50ml/L of sterile defibered sheep blood.
Preferably, the rejuvenation culture solution has the following components:
22g/L of sodium chloride is added,
the magnesium chloride is 6g/L,
the concentration of adenosine triphosphate is 4g/L,
1.5g/L of methionine,
0.7g/L of calcium acetate,
38ml/L of sterile defibered sheep blood.
More preferably, the temperature conditions for measuring the luminescence intensity are: 20-25 ℃.
Preferably, the polluted water body is a heavy metal polluted water body.
More preferably, the heavy metal is mercury.
The beneficial effects achieved by the invention mainly comprise the following aspects:
both the external conditions and the composition of the medium influence the luminescence of the bacteria. The recovery condition is unfavorable, the survival of the strain is influenced, and the luminous efficiency of the strain is reduced.
The luminescence process of the luminous bacteria is related to the light respiration, the luminescence of the luminous bacteria is a light respiration process, and luciferin is activated after receiving energy provided by ATP and emits fluorescence under the catalysis of luciferase. When the cells are frozen, the cells are in a resting state, most enzymes are not activated, and the metabolic activity and the enzyme activity of the strains can be recovered for a long time after recovery.
Aiming at the invention, the rapid recovery of the luciferase activity is positively promoted by adding magnesium ions; the adenosine triphosphate, upon absorption, provides energy for the activation of fluorescein. The aseptic defibered sheep blood provides hormone and low molecular nutrient substances for maintaining cell viability, provides binding protein, can bind metal ions and methionine, regulates the material viability of bound factors, and is beneficial to absorption and utilization of strains; and the osmotic pressure can be buffered, so that cells are protected from being damaged. Acetate participates in the synthesis of acetyl coenzyme A in cells and further participates in tricarboxylic acid circulation, so that the metabolic strength of the cells is increased; and calcium ions can also have positive influence on the growth of thalli and the synthesis of products.
According to the invention, by optimizing and improving the photobacterium leiognathi rejuvenation culture solution, resting cells are quickly activated within 20-30min, the survival rate of thallus cells and the enzyme activity are obviously improved compared with the prior art, the luminous efficiency is completely recovered, and the photobacterium leiognathi rejuvenation culture solution is beneficial to quickly and accurately monitoring environmental pollutants.
The bioluminescence method disclosed by the invention is rapid and sensitive in monitoring the polluted water body and has a wide market application prospect.
Drawings
FIG. 1: influence of rejuvenation culture solution on the survival rate of somatic cells;
FIG. 2: the influence of rejuvenation culture medium on luminous efficiency;
FIG. 3: comparing the luminous efficiency of each group;
FIG. 4: mercury chloride concentration-relative luminosity regression equation.
Detailed Description
Those skilled in the art can modify the process parameters appropriately to achieve the desired results with reference to the disclosure herein. It is expressly intended that all such similar substitutes and modifications which would be obvious to one skilled in the art are deemed to be included in the invention. While the products and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations and modifications, or appropriate alterations and combinations, of the products and methods described herein may be made and utilized without departing from the spirit, scope, and spirit of the invention. For a further understanding of the present invention, reference will now be made in detail to the following examples.
The reagent consumables used in the present invention can be commercially purchased except for the specific instructions. The specific test strain of photobacterium leiognathi is photobacterium leiognathi ATCC33979.
Example 1
A rejuvenation culture solution of photobacterium leiognathi for monitoring environmental pollutants comprises the following components:
26g/L of sodium chloride is added,
the magnesium chloride is 6g/L,
the concentration of the adenosine triphosphate is 4g/L,
1g/L of methionine is added into the mixture,
0.8g/L of calcium acetate,
35ml/L of sterile defibered sheep blood.
The preparation method comprises the following steps: sequentially adding the raw materials into distilled water, mixing, and sterilizing.
Example 2
A rejuvenation culture solution of photobacterium leiognathi for monitoring environmental pollutants comprises the following components:
the concentration of the sodium chloride is 18g/L,
the magnesium chloride is 5g/L,
3g/L of adenosine triphosphate is added,
1g/L of methionine is added into the mixture,
0.5g/L of calcium acetate,
30ml/L of sterile defibered sheep blood.
Example 3
The rejuvenation culture solution of photobacterium leiognathi for monitoring environmental pollutants comprises the following components:
the concentration of the sodium chloride is 20g/L,
the magnesium chloride is 6g/L,
adenosine triphosphate of 5g/L is added into the solution,
2g/L of methionine is added, and the content of methionine is 2g/L,
1g/L of calcium acetate is added,
50ml/L of sterile defibered sheep blood.
Example 4
The rejuvenation culture solution of photobacterium leiognathi for monitoring environmental pollutants comprises the following components:
22g/L of sodium chloride is added,
the magnesium chloride is 6g/L,
the concentration of the adenosine triphosphate is 4g/L,
1.5g/L of methionine,
0.7g/L of calcium acetate,
38ml/L of sterile defibered sheep blood.
Example 5
The method for rejuvenating photobacterium leiognathi by using the rejuvenation culture solution comprises the following steps:
placing the photobacterium leiognathi freeze-dried powder in a water bath at 37 ℃ for rapid thawing, adding a rejuvenation culture solution with the weight 2 times that of the photobacterium leiognathi freeze-dried powder, and placing the photobacterium leiognathi freeze-dried powder in an incubator at 32 ℃ for shaking culture at 200rpm for 20min to obtain the photobacterium leiognathi freeze-dried powder.
The freeze-dried powder is prepared by a conventional method, or the following method:
the frozen stock solution comprises the following components in parts by weight: 8 parts of skimmed milk powder, 5 parts of cane sugar and 93 parts of distilled water, uniformly mixing, sterilizing at 121 ℃ for 15min, and naturally cooling to obtain the milk powder.
Culturing Photobacterium liquid of Haliotis diversicolor leiognathi (in logarithmic phase, with thallus density of 1 × 10) 8 Individual/ml) and frozen stock according to 2:1, placing at-70 ℃ for freezing for 30min, then carrying out vacuum freeze-drying to obtain freeze-dried powder, and storing in a refrigerator at-20 ℃.
Comparative example 1
The rejuvenation culture solution comprises the following components:
26g/L of sodium chloride is added,
methionine 1g/L.
Comparative example 2
The rejuvenation culture solution comprises the following components:
26g/L of sodium chloride is added,
1g/L of methionine is added into the mixture,
calcium acetate 0.8g/L.
Comparative example 3
The rejuvenation culture solution comprises the following components:
26g/L of sodium chloride is added into the solution,
the magnesium chloride is 6g/L,
1g/L of methionine is added into the mixture,
calcium acetate 0.8g/L.
Comparative example 4
The rejuvenation culture solution comprises the following components:
26g/L of sodium chloride is added,
the magnesium chloride is 6g/L,
the concentration of the adenosine triphosphate is 4g/L,
1g/L of methionine is added into the mixture,
calcium acetate 0.8g/L.
Example 6
1. The effect of rejuvenation medium on the survival of bacterial cells.
Conventional viable cell counting methods were used.
The number of viable bacteria in each tube of lyophilized powder is 4 × 10 8 The bacterial suspension is prepared from 4ml of bacterial liquid and 2ml of frozen stock solution.
The freeze-dried powder for each group test has the same preparation method and conditions and is comparable. The freeze-dried powder used in each group of experiments is stored in a refrigerator at the temperature of 20 ℃ below zero for 30 days and 180 days.
Rejuvenation method reference example 5; the rejuvenation culture medium was selected from examples 1 to 4 and comparative examples 1 to 4.
Strain survival rate = (viable count after rejuvenation/viable count before freeze-drying) × 100%
As shown in FIG. 1, in the test groups frozen and stored for 30d, the survival rates of the groups of examples 1-4 were maintained above 98%, the survival rates of the comparative examples 1-4 were maintained between 60-90%, and the survival rate of the lowest group was 65.8% of that of the comparative example 1. In the test groups frozen and stored for 180d, the survival rates of the groups of examples 1-4 are all maintained to be more than 90%, the survival rates of the comparative examples 1-4 are maintained to be between 50% and 80%, the lowest survival rate is that of the comparative example 1 which is only 51.3%, and then that of the comparative example 2 which is 59.6%.
2. Effect of rejuvenation Medium on luminous efficiency
Diluting the bacterial liquid before freezing and the rejuvenated bacterial liquid to 10 4 The luminescence intensity of each/ml dilution was measured by using an SHG-1 type biochemical luminescence measuring apparatus. Is prepared to contain 2ml of diluentAnd (4) a test tube for releasing liquid. And (3) under the constant temperature condition, putting the sample added with the bacterial liquid into a luminescence tester, measuring the luminescence intensity, obtaining a reading, comparing the luminescence intensity of the rejuvenated bacterial liquid with that of the bacterial liquid before freezing storage, and calculating the relative luminescence intensity. Each set of experiments was repeated three times and the average was calculated. As shown in fig. 2, the relative luminescence intensities of examples 1 to 4 respectively reach 99.1%, 101.3%, 98.9% and 100.6%, and are substantially consistent with the luminescence intensity of the bacteria solution before cryopreservation, which indicates that the photobacterium leiognathi can rapidly recover the luminescence efficiency after the rejuvenation culture solution is revived, and is helpful for rapidly and accurately monitoring environmental pollutants; comparative example 1 had the worst luminous efficiency, and the relative luminous intensity reached only 47.8%, and comparative examples 2 to 4 were 62.5%, 70.6%, and 82.3%, respectively.
3. The applicant tries to add four substances of glucose, yeast extract, albumin and trehalose respectively on the basis of the rejuvenation culture solution in the example 1, the adding amount is 5g/L as a test example, and the proportioning mode is shown in Table 1:
TABLE 1
| Glucose | Yeast extract | Albumin | Trehalose | |
| Group 1 | + | - | - | - |
| Group 2 | - | + | - | - |
| Group 3 | - | - | + | - |
| Group 4 | - | - | - | + |
Through tests, compared with example 1, the survival rates of the bacterial cells of groups 1-4 are not obviously changed, the bacterial cells are maintained at higher survival rates, and the luminous efficiency is changed, as shown in fig. 3, the luminous efficiency of group 2 is obviously reduced, about 20% of that of example 1, the luminous efficiency of groups 1 and 4 is also reduced, and the group 3 is basically the same as the example 1. Probably, the yeast extract interferes with a luminescence mechanism of photobacterium leiognathi in the resuscitation of photobacterium leiognathi, and glucose and trehalose also have certain influence on the luminescence mechanism.
4. The photobacterium leiognathi is tested by using a rejuvenation culture medium aiming at photobacterium leiognathi in the prior art.
CN102604873A (ammonium chloride 0.3, calcium carbonate 1, casein hydrolase 3, magnesium sulfate 0.3, monopotassium phosphate 3, sodium chloride 25, sodium glycerophosphate 24, yeast extract 2.5, unit g/L); the rejuvenation method was repeated three times as in example 5, and the average value was taken, and it was determined that the survival rate of the cells was 68.9%, the relative luminous intensity was only 43.7%, and the luminous efficiency was reduced by more than half. Due to the difference of physicochemical properties of the strains and the difference of components of the rejuvenation culture solution, the luminous efficiency of the strains is greatly influenced.
Example 7
A method for monitoring a polluted water body by a bioluminescence method comprises the following steps:
placing a water sample to be tested into a testing container, adding sodium chloride, controlling the concentration of the sodium chloride to be 30g/L, then inoculating a bacterial liquid, and controlling the density of thalli to be 1 multiplied by 10 4 Per ml; placing 30g/L sodium chloride aqueous solution in a control container, inoculating bacterial liquid, and controlling the bacterial density to 1 × 10 4 Per ml; controlling the equal volume of the solutions of the test container and the control container, placing the containers in an SHG-1 type biochemical luminescence tester for 15min, and then measuring the luminescence intensity.
The calculation method comprises the following steps:
relative luminosity (Y) = test/control × 100%.
Luminescence inhibition ratio (I) = (control-test)/control × 100%.
The measurement conditions were as follows: room temperature: 20-25 ℃, and the temperature fluctuation of the same batch of samples is required not to exceed +/-1 ℃ in the determination process.
If the measured sample has a luminescence inhibition rate of more than 99%, the sample needs to be diluted by 30g/L sodium chloride aqueous solution to five concentrations: 10%, 1%, 0.1%, 0.01% four concentration gradients.
The toxicity standards are referenced in table 2 with mercuric chloride as the reference contaminant.
TABLE 2
| Concentration of mercuric chloride solution mg/L | Toxicity |
| ＜0.07 | Low toxicity |
| ≥0.07，＜0.09 | Poisoning by |
| ≥0.09，＜0.12 | Heavy toxicity |
| ≥0.12，＜0.16 | High toxicity |
| ≥0.16 | High toxicity |
The relative luminosity was measured as described above using 30g/L sodium chloride as the solvent, and mercury chloride concentration 0.04,0.08,0.12,0.16,0.20,0.24 in mg/L was set. Solving a (intercept), b (slope, regression coefficient) and R (correlation coefficient) of a unary linear regression equation: an equation between the relative luminosity Y and the mercury chloride concentration X was established, Y = a + bX mercury chloride, the significance level of the correlation coefficient (P value) was looked up, and the significance level of the R value found was examined, as shown in fig. 4. If P is less than or equal to 0.01 and EC50 mercuric chloride =0.l0mg/L +/-0.02 mg/L, the required correlation equation is established; conversely, this does not hold true, and the light emission of the series of mercury chloride concentrations must be retested. The test results establish an equation and a significant level of correlation coefficient P < 0.1, and EC50 mercuric chloride =0.l18mg/L.
Taking sewage discharged by a certain paper mill and a certain metallurgical plant as test water samples, repeating the three times to obtain an average value, and the specific test results are shown in a table 3:
TABLE 3
| Water sample source | Relative luminosity% | Equivalent mercury ion concentration mg/L | Luminescence inhibition ratio% | Toxicity |
| A certain paper mill | 61.9 | 0.087 | 38.1 | Poisoning by |
| A certain metallurgical plant | 7.5 | 0.231 | 92.5 | High toxicity |
The foregoing lists only preferred embodiments of the invention. It is obvious that the invention is not limited to the above embodiments, but that many variations are possible. All modifications which can be derived or suggested by a person skilled in the art from the disclosure of the present invention are to be considered within the scope of the invention.
Claims (7)
1. A method of monitoring a contaminated water body by bioluminescence, the method comprising the steps of:
step 1) preparing a rejuvenation culture solution: sequentially adding sodium chloride, magnesium chloride, adenosine triphosphate, methionine, calcium acetate and sterile defibrinated sheep blood into distilled water, mixing, and sterilizing;
step 2) rejuvenation of luminescent bacteria: unfreezing photobacterium leiognathi freeze-dried powder in a water bath at 37 ℃, adding the rejuvenation culture solution obtained in the step 1), and placing the mixture in an incubator at 32 ℃ for shake culture at 200rpm for 20-40min to obtain a bacterial solution;
step 3), sewage monitoring: placing a water sample to be tested in a testing container, adding sodium chloride, controlling the concentration of the sodium chloride to be 30g/L, and then accessing the bacterial liquid obtained in the step 2); placing 30g/L sodium chloride aqueous solution in a reference container, and then inoculating the bacterial liquid obtained in the step 2); controlling the density of the thallus in the test container and the density of the thallus in the comparison container to be the same; placing in biochemical luminescence determinator for 15min, and finally determining luminescence intensity;
the rejuvenation culture solution consists of the following components:
10-30g/L of sodium chloride,
4-7g/L of magnesium chloride,
3-5g/L of adenosine triphosphate,
1-2g/L of methionine,
0.5-1g/L of calcium acetate,
30-50ml/L of aseptic defibered sheep blood.
2. The method of claim 1, wherein the rejuvenation broth comprises the following components:
26g/L of sodium chloride is added,
the magnesium chloride is 6g/L,
the concentration of the adenosine triphosphate is 4g/L,
1g/L of methionine is added into the mixture,
0.8g/L of calcium acetate,
35ml/L of sterile defibered sheep blood.
3. The method as claimed in claim 1, wherein the rejuvenation broth has the following composition:
the concentration of the sodium chloride is 18g/L,
the magnesium chloride is 5g/L,
3g/L of Adenosine Triphosphate (ATP),
1g/L of methionine is added into the mixture,
0.5g/L of calcium acetate,
30ml/L of sterile defibered sheep blood.
4. The method as claimed in claim 1, wherein the rejuvenation broth has the following composition:
the concentration of the sodium chloride is 20g/L,
the magnesium chloride is 6g/L,
adenosine triphosphate of 5g/L is added into the solution,
2g/L of methionine is added, and the content of methionine is 2g/L,
1g/L of calcium acetate is added,
50ml/L of sterile defibered sheep blood.
5. The method of claim 1, wherein the rejuvenation broth comprises the following components:
22g/L of sodium chloride is added,
the magnesium chloride is 6g/L,
the concentration of the adenosine triphosphate is 4g/L,
1.5g/L of methionine,
0.7g/L of calcium acetate,
38ml/L of sterile defibered sheep blood.
6. The method of claim 1, wherein the contaminated water is a heavy metal contaminated water.
7. The method of claim 6, wherein the heavy metal is mercury.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
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| CN102465167A (en) * | 2010-11-10 | 2012-05-23 | 中国科学院生态环境研究中心 | Rapid and high-flux acute toxicity test method for luminous bacteria |
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| AU4220297A (en) * | 1996-09-13 | 1998-04-02 | Kyowa Hakko Kogyo Co. Ltd. | Processes for producing sugar nucleotides and complex carbohydrates |
| GB0818453D0 (en) * | 2008-10-08 | 2008-11-12 | Novartis Ag | Fermentation processes for cultivating streptococci and purification processes for obtaining cps therefrom |
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| GB2005018A (en) * | 1977-09-28 | 1979-04-11 | Beckman Instruments Inc | Method for detecting toxic substances in liquids |
| CN102465167A (en) * | 2010-11-10 | 2012-05-23 | 中国科学院生态环境研究中心 | Rapid and high-flux acute toxicity test method for luminous bacteria |
| CN105713952A (en) * | 2014-12-05 | 2016-06-29 | 中国石油化工股份有限公司 | Biotoxicity assessment method of oilfield chemicals |
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