CN105842399B - The method that model organism algae measures agriculture chemicals fungicide toxicity - Google Patents
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- 241000195493 Cryptophyta Species 0.000 title claims abstract description 69
- 238000000034 method Methods 0.000 title claims abstract description 36
- 239000000126 substance Substances 0.000 title claims abstract description 27
- 230000000855 fungicidal effect Effects 0.000 title claims abstract description 22
- 239000000417 fungicide Substances 0.000 title claims abstract description 22
- 231100000419 toxicity Toxicity 0.000 title claims abstract description 19
- 230000001988 toxicity Effects 0.000 title claims abstract description 19
- 238000010790 dilution Methods 0.000 claims abstract description 38
- 239000012895 dilution Substances 0.000 claims abstract description 37
- 239000007788 liquid Substances 0.000 claims abstract description 20
- 238000002474 experimental method Methods 0.000 claims abstract description 18
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 15
- 239000008223 sterile water Substances 0.000 claims abstract description 5
- 239000001963 growth medium Substances 0.000 claims description 24
- 239000000243 solution Substances 0.000 claims description 18
- 241000883966 Astrophytum capricorne Species 0.000 claims description 11
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims description 11
- 239000002574 poison Substances 0.000 claims description 11
- 231100000614 poison Toxicity 0.000 claims description 11
- 230000002401 inhibitory effect Effects 0.000 claims description 10
- 238000012360 testing method Methods 0.000 claims description 10
- 239000000575 pesticide Substances 0.000 claims description 9
- 230000012010 growth Effects 0.000 claims description 8
- 210000004027 cell Anatomy 0.000 claims description 7
- 238000011109 contamination Methods 0.000 claims description 7
- 238000005286 illumination Methods 0.000 claims description 7
- 238000002360 preparation method Methods 0.000 claims description 7
- PXMNMQRDXWABCY-UHFFFAOYSA-N 1-(4-chlorophenyl)-4,4-dimethyl-3-(1H-1,2,4-triazol-1-ylmethyl)pentan-3-ol Chemical compound C1=NC=NN1CC(O)(C(C)(C)C)CCC1=CC=C(Cl)C=C1 PXMNMQRDXWABCY-UHFFFAOYSA-N 0.000 claims description 6
- 239000005839 Tebuconazole Substances 0.000 claims description 6
- 239000011248 coating agent Substances 0.000 claims description 6
- 239000000725 suspension Substances 0.000 claims description 6
- 230000001419 dependent effect Effects 0.000 claims description 4
- 239000012153 distilled water Substances 0.000 claims description 4
- 239000003814 drug Substances 0.000 claims description 4
- 229940079593 drug Drugs 0.000 claims description 4
- 231100000566 intoxication Toxicity 0.000 claims description 4
- 231100000053 low toxicity Toxicity 0.000 claims description 4
- 231100000572 poisoning Toxicity 0.000 claims description 4
- 230000000607 poisoning effect Effects 0.000 claims description 4
- 210000000601 blood cell Anatomy 0.000 claims description 3
- 238000012937 correction Methods 0.000 claims description 3
- 238000011156 evaluation Methods 0.000 claims description 3
- 239000011888 foil Substances 0.000 claims description 3
- 230000009036 growth inhibition Effects 0.000 claims description 3
- 230000000873 masking effect Effects 0.000 claims description 3
- 238000012545 processing Methods 0.000 claims description 3
- 230000001018 virulence Effects 0.000 claims description 3
- 238000004581 coalescence Methods 0.000 claims description 2
- 230000000694 effects Effects 0.000 claims description 2
- 230000001954 sterilising effect Effects 0.000 claims description 2
- 238000004659 sterilization and disinfection Methods 0.000 claims description 2
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 claims 2
- XLYOFNOQVPJJNP-ZSJDYOACSA-N heavy water Substances [2H]O[2H] XLYOFNOQVPJJNP-ZSJDYOACSA-N 0.000 claims 2
- 229910021380 Manganese Chloride Inorganic materials 0.000 claims 1
- GLFNIEUTAYBVOC-UHFFFAOYSA-L Manganese chloride Chemical compound Cl[Mn]Cl GLFNIEUTAYBVOC-UHFFFAOYSA-L 0.000 claims 1
- 229910015667 MoO4 Inorganic materials 0.000 claims 1
- KGBXLFKZBHKPEV-UHFFFAOYSA-N boric acid Chemical compound OB(O)O KGBXLFKZBHKPEV-UHFFFAOYSA-N 0.000 claims 1
- 229910052564 epsomite Inorganic materials 0.000 claims 1
- 239000011565 manganese chloride Substances 0.000 claims 1
- 229910000029 sodium carbonate Inorganic materials 0.000 claims 1
- 230000004071 biological effect Effects 0.000 abstract description 4
- 239000005760 Difenoconazole Substances 0.000 description 5
- BQYJATMQXGBDHF-UHFFFAOYSA-N difenoconazole Chemical compound O1C(C)COC1(C=1C(=CC(OC=2C=CC(Cl)=CC=2)=CC=1)Cl)CN1N=CN=C1 BQYJATMQXGBDHF-UHFFFAOYSA-N 0.000 description 5
- 239000000375 suspending agent Substances 0.000 description 5
- 230000005791 algae growth Effects 0.000 description 4
- 230000005764 inhibitory process Effects 0.000 description 4
- 238000011160 research Methods 0.000 description 4
- 238000012986 modification Methods 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 239000000463 material Substances 0.000 description 2
- 231100000331 toxic Toxicity 0.000 description 2
- 230000002588 toxic effect Effects 0.000 description 2
- 231100000820 toxicity test Toxicity 0.000 description 2
- 231100000041 toxicology testing Toxicity 0.000 description 2
- 231100000215 acute (single dose) toxicity testing Toxicity 0.000 description 1
- 238000011047 acute toxicity test Methods 0.000 description 1
- 238000004166 bioassay Methods 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 239000012897 dilution medium Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 230000035800 maturation Effects 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 239000012452 mother liquor Substances 0.000 description 1
- 244000038651 primary producers Species 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000006641 stabilisation Effects 0.000 description 1
- 238000011105 stabilization Methods 0.000 description 1
- 239000003206 sterilizing agent Substances 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 230000002110 toxicologic effect Effects 0.000 description 1
- 231100000027 toxicology Toxicity 0.000 description 1
Classifications
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
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- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
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- Agricultural Chemicals And Associated Chemicals (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention discloses the methods that one mode biology algae measures agriculture chemicals fungicide toxicity, include the following steps：Agriculture chemicals fungicide is configured to the experiment liquid of gradient concentration；Algae dilution is configured to by biological algae stoste sterile water is dilute；Experiment liquid is mixed with algae dilution, 72h is cultivated under air-proof condition, frustule is counted every for 24 hours, and record the biological effect of frustule and the physical and chemical parameter of experimental liquid；Half effective concentration EC of the agriculture chemicals fungicide to frustule is calculated according to the biological effect parameter of frustule50, judge the toxicity of agriculture chemicals fungicide.The method of the present invention has the advantages such as easy to operate, highly practical, the period is short, and assessment endpoint easily determines.
Description
Technical field
The present invention relates to the sides that cultural technique field more particularly to one mode biology algae measure agriculture chemicals fungicide toxicity
Method.
Background technology
Primary producer of the algae as aquatic ecosystem, balance and stabilization to water body play extremely important work
With.Algal bioassay in biological test is essential method in aquatic ecological toxicologic study, and pesticide is after the use of field
It can migrate into water body, the growth of algae can be had an impact, to influence the entire ecosystem.Algae is commonly used for new chemistry
Algal grown in substance registration inhibits toxicity test research and agriculture chemical registration algal grown to inhibit toxicity test research.
Currently, it is less about the research of agriculture chemicals fungicide toxicity screening report both at home and abroad, even if having part report and correlation
The model of research, but it is a lack of suitability and reliability that relevant data assesses the model, and as maturation and recognize
Can toxicity screening model.Therefore it provides a kind of method of algae screening agriculture chemicals fungicide toxicity easy to operate, highly practical is outstanding
Its is important.
Invention content
The technical problem to be solved by the present invention is to overcome the deficiencies in the prior art, provide a kind of easy to operate, highly practical
The method that model organism algae measures agriculture chemicals fungicide toxicity.
In order to solve the above technical problems, the method that one mode biology algae measures agriculture chemicals fungicide toxicity is provided, including
Following steps：
S1, the experiment liquid that agriculture chemicals fungicide is configured to gradient concentration；
S2, algae dilution is configured to by biological algae stoste sterile water is dilute；
S3, the experiment liquid is mixed with the algae dilution, 72h is cultivated under air-proof condition, every for 24 hours to frustule
It counts, and records the biological effect of frustule and the physical and chemical parameter of experimental liquid；
S4, half effective concentration EC of the agriculture chemicals fungicide to frustule is calculated according to the biological effect parameter of frustule50, sentence
The toxicity of disconnected agriculture chemicals fungicide.
Above-mentioned method, it is preferred that the S1 steps are specially：Agriculture chemicals fungicide and BG11 culture medium dilutions are mixed
It closes, is configured to the experiment liquid of gradient concentration.
Above-mentioned method, it is preferred that the culture medium dilution is that BG11 culture mediums are diluted to the solution after 10 times.
Above-mentioned method, it is preferred that biological algae stoste is the purebred algae in exponential phase described in the step S2.
Above-mentioned method, it is preferred that it is described, in algae dilution frustule a concentration of 1.0 × 104~1.0 × 105A/
mL。
Above-mentioned method, it is preferred that described in the step S3, experiment liquid and the volume ratio of the algae dilution are 1:
1。
Above-mentioned method, it is preferred that the condition cultivated described in the step S3 is：Temperature is 21 DEG C~24 DEG C, illumination
Intensity is 4440Lux~8880Lux, light application ratio 16h/8h.
Above-mentioned method, it is preferred that during culture, be no less than 5 times every the number for shaking solution in triangular flask for 24 hours.
Above-mentioned method, it is preferred that the biological respinse effect of frustule described in the step S3 includes the suppression of frustule
The intoxication conditions of rate processed and frustule；The intoxication conditions of the frustule include：Frustule is lopsided, algae solution bleaches, frustule body
Product becomes smaller, frustule adhesion or coalescence.
Above-mentioned method, it is preferred that the physical and chemical parameter of experimental liquid described in the step S3 includes：The water temperature of experimental liquid,
PH value and intensity of illumination.
Above-mentioned method, it is preferred that statistic software SPSS is used in the step S4, with the logarithm of pesticide sterilizing agent concentration
Value is independent variable x, with the probit value (the concentration changing value for inhibiting probit value, that is, frustule) of inhibiting rate for dependent variable y, establishes poison
Power regression equation calculates the half effective concentration EC that tested material inhibits algal grown50。
Above-mentioned method, it is preferred that pesticide poison grade criterion described in the step S4 is EC50≤0.3mg a.i./
L, high poison；0.300 < EC50≤ 3.00mg a.i./L, poisoning；EC50> 3.0mg a.i./L, low toxicity.
Above-mentioned method, it is preferred that the method that the model organism algae measures agriculture chemicals fungicide toxicity is additionally provided with blank pair
According to group, the blank control group is with the mixture culture biology algae of algae dilution and culture medium.
Compared with the prior art, the advantages of the present invention are as follows：
(1) the present invention provides a kind of model organism algae easy to operate, highly practical to screen agriculture chemicals fungicide toxicity
Method；This method can tentatively judge the toxicity of agriculture chemicals fungicide, and speed is fast, and accuracy is high.
(2) the present invention provides the method that one mode biology algae measures agriculture chemicals fungicide toxicity, acute toxicity tests
Period is short, and assessment endpoint easily determines, has simple, easy-operating advantage.
Specific implementation mode
Below in conjunction with specific preferred embodiment, the invention will be further described, but not thereby limiting the invention
Protection domain.
Embodiment
Material and instrument employed in following embodiment are commercially available.
Embodiment 1：
A kind of method that the model organism algae of the present invention measures agriculture chemicals fungicide toxicity, includes the following steps：
S1, the preparation for testing liquid：
Prepare BG11 culture medium dilutions：BG11 culture medium prescriptions are prepared according to the formula of table 1, preparation method is：It will
Ingredient in table 1 is configured to corresponding mother liquid concentration, and corresponding mother liquor dosage is sequentially transferred to 1000mL appearances successively according to indicating
In measuring bottle, distilled water is used in combination to be settled to 1000mL, the high pressure sterilization 15min at 121 DEG C seals and post label, and 4 DEG C of refrigerators are protected
It deposits, the term of validity 2 months.It is BG11 culture medium dilutions after BG11 culture mediums are diluted 10 times with distilled water.
50% difenoconazole suspending agent 0.1000g is weighed, dissolved with BG11 culture medium dilutions and is settled to 100mL appearances
Measuring bottle obtains the storing solution of a concentration of 500mg a.i./L.Pipette successively above-mentioned storing solution 0.100,0.200,0.400,
0.800,1.600,3.200mL is settled to 250mL in 250mL volumetric flasks with BG11 culture medium dilutions, obtains concentration difference
For the experiment liquid of 0.200,0.400,0.800,1.60,3.20,6.40mg a.i./L.
Table 1：BG11 culture medium prescriptions
The preparation of S2, algae dilution：The cell concentration that algae stoste (goat's horn crescent moon algae) is measured with counting method of blood cell is 4.00
×106A/mL is used in combination sterile water to dilute 100 times and obtains algae dilution, frustule a concentration of 4.00 × 10 in algae dilution4A/
mL。
S3, contamination：
Experimental group：The experiment liquid 50mL for the gradient concentration prepared in step S1 is taken to be separately added into the algae of 50mL in step S2
Dilution obtains the biological algae culture medium of gradient concentration.Algae initial concentration is 2.00 × 10 in experiment4A/mL.
Blank control group：It takes 50mL algaes dilution to be uniformly mixed so as to obtain biological algae with 50mL BG11 culture medium dilutions to cultivate
Base.
The biological algae culture medium of experimental group and blank control group is fitted into triangular flask respectively, is sealed with masking foil, and
It is placed in intelligent growth cabinet and cultivates, 16h is placed in control under illumination condition, and 8h is placed under dark surrounds；Daily daytime is every
It shook every 2 hours 1 time, shakes 5 times altogether.
S4, test data record：
The water temperature of each concentration group is 22.2 DEG C~23.5 DEG C during measuring experiment, and pH value is 6.88~7.26, in incubator
Intensity of illumination be 5200Lux~5650Lux.The suppressed algae solution cell volume of 48h, 72h becomes smaller after contamination, observes and records dye
0h after poison, for 24 hours, 48h, 72h frustule concentration (specific data are shown in Table 2).
Table 2：50% difenoconazole suspending agent is to goat's horn crescent moon algae growth inhibition test data record sheet
Note：Indicate frustule mean concentration (a/mL)
S5, data processing：
Using statistic software SPSS 12.0, using the logarithm of drug concentration as independent variable x, with the probability of (correction) inhibiting rate
Value is dependent variable y, establishes virulence regression equation.Calculate the half effective concentration EC inhibited to algal grown for examination object50With
95% confidence limit, result of calculation are listed in Table 3 below.
Table 3：50% difenoconazole suspending agent is to goat's horn crescent moon algae growth inhibition test result table
50% difenoconazole suspending agent is calculated to the growth inhibiting EC of goat's horn crescent moon algae by the above method50
(72h) is 0.759mg a.i./L, and 95% confidence is limited to 0.603~0.973mg a.i./L.
S6, evaluation of result：
According to EC50Value judges pesticide poison grade.Pesticide presses EC to the growth inhibition toxicity of algae50The size of (72h) is divided into
Three grades：High poison (EC50≤0.300mg a.i./L)；Be poisoned (0.300mg a.i./L < EC50≤3.00mg a.i./L)；
Low toxicity (EC50> 3.00mg a.i./L).
Under this experimental condition, 50% difenoconazole suspending agent is to the growth inhibiting EC of goat's horn crescent moon algae50(72h) is
0.759mg a.i./L, 0.300mg a.i./L < EC50≤ 3.00mg a.i./L, toxic grade are " poisoning ".
Embodiment 2
A kind of method that the model organism algae of the present invention measures agriculture chemicals fungicide toxicity, includes the following steps：
1, drug solution preparing is tested：
Pipette 80 grams per liter Tebuconazole suspension seed-coating agent 0.625mL, with BG11 culture mediums dilution (BG11 dilutions at
Divide same as Example 1) it dissolves and is settled to 100mL volumetric flasks, obtain the storing solution of a concentration of 500mg a.i./L.It moves successively
Above-mentioned storing solution 0.300,0.600,1.200,2.400,4.800mL are taken, is settled to 250mL with BG11 culture medium dilutions respectively
In volumetric flask, it is configured to 2 times of experiment liquids of a concentration of 0.600,1.20,2.40,4.80,9.60mg a.i./L.
The preparation of S2, algae dilution：The cell concentration that algae stoste (goat's horn crescent moon algae) is measured with counting method of blood cell is 4.00
×106, it is used in combination sterile water to dilute 100 times and obtains algae dilution, frustule a concentration of 4.00 × 10 in algae dilution4A/mL.
S3, contamination：
Experimental group：The experiment liquid 50mL for the gradient concentration prepared in step S1 is taken to be separately added into the algae of 50mL in step S2
Dilution obtains the biological algae culture medium of gradient concentration.Algae initial concentration is 2.00 × 10 in experiment4A/mL.
Blank control group：Take 50mL algaes dilution and 50mL BG11 culture medium dilution mixings.
The biological algae culture medium of experimental group and blank control group is fitted into triangular flask respectively, is sealed with masking foil, and
It is placed in intelligent growth cabinet and cultivates, 16h is placed in control under illumination condition, and 8h is placed under dark surrounds；Daily daytime is every
It shook every 2 hours 1 time, shakes 5 times altogether.
S4, test data record：
The water temperature of each concentration group is 22.0 DEG C~23.1 DEG C during measuring experiment, and pH value is 6.74~7.16, in incubator
Intensity of illumination be 5200Lux~5700Lux.The suppressed algae solution cell volume of 48h, 72h becomes smaller after contamination, observes and records dye
0h after poison, for 24 hours, 48h, 72h frustule concentration (specific data are shown in Table 4).
Table 4：80 grams per liter Tebuconazole suspension seed-coating agents are to goat's horn crescent moon algae growth inhibition test data record sheet
Note：Indicate frustule mean concentration (a/mL)
S5, data processing：
Using statistic software SPSS 12.0, using the logarithm of drug concentration as independent variable x, with the probability of (correction) inhibiting rate
Value is dependent variable y, establishes virulence regression equation.Calculate the half effective concentration EC inhibited to algal grown for examination object50With
95% confidence limit, result of calculation are listed in Table 5 below.
Table 5：80 grams per liter Tebuconazole suspension seed-coating agents are to goat's horn crescent moon algae growth inhibition test result table
80 grams per liter Tebuconazole suspension seed-coating agents are calculated to the growth inhibiting EC of goat's horn crescent moon algae by the above method50
(72h) is 1.37mg a.i./L, and 95% confidence is limited to 1.15~1.66mg a.i./L.
S6, evaluation of result：
According to EC50Value judges pesticide poison grade.Pesticide presses EC to the growth inhibition toxicity of algae50The size of (72h) is divided into
Three grades：High poison (EC50≤0.300mg a.i./L)；Be poisoned (0.300mg a.i./L < EC50≤3.00mg a.i./L)；
Low toxicity (EC50> 3.00mg a.i./L).
Under this experimental condition, 80 grams per liter Tebuconazole suspension seed-coating agents are to the growth inhibiting EC of goat's horn crescent moon algae50(72h)
For 1.37mg a.i./L, 0.300mg a.i./L < EC50≤ 3.00mg a.i./L, toxic grade are " poisoning ".
The above described is only a preferred embodiment of the present invention, being not intended to limit the present invention in any form.Though
So the present invention has been disclosed with preferred embodiment as above, and however, it is not intended to limit the invention.It is any to be familiar with those skilled in the art
Member, in the case where not departing from the Spirit Essence and technical solution of the present invention, all using in the methods and techniques of the disclosure above
Appearance makes many possible changes and modifications to technical solution of the present invention, or is revised as the equivalent embodiment of equivalent variations.Therefore,
Every content without departing from technical solution of the present invention is made to the above embodiment any simple according to the technical essence of the invention
Modification, equivalent replacement, equivalence changes and modification, still fall within technical solution of the present invention protection in the range of.
Claims (1)
1. the method that one mode biology algae measures agriculture chemicals fungicide toxicity, which is characterized in that include the following steps：
S1, the preparation for testing liquid：
Prepare BG11 culture medium dilutions：Preparation method is：The NaNO of a concentration of 15.0g/100mL of 10mL is taken successively3、10mL
The K of a concentration of 2.0g/500mL2HPO4, a concentration of 3.75g/500mL of 10mL MgSO4·7H2O, a concentration of 1.8g/ of 10mL
The CaCl of 500mL2·2H2O, the C of a concentration of 0.3g/500mL of 10mL6H8O7, a concentration of 0.3g/500mL of 10mL FeC6H5O7·
NH4The EDTANa of a concentration of 0.05g/500mL of OH, 10mL2, a concentration of 1.0g/500mL of 10mL Na2CO3, 1mL it is a concentration of
2.86g/L H3BO3, a concentration of 1.86g/L of 1mL MnCl2·4H2O, the ZnSO of a concentration of 0.22g/L of 1mL4·7H2O、1mL
The Na of a concentration of 0.39g/L2MoO4·2H2O, the CuSO of a concentration of 0.08g/L of 1mL4·5H2O, the Co of a concentration of 0.05g/L of 1mL
(NO3)2·6H2O is transferred in 1000mL volumetric flasks, distilled water is used in combination to be settled to 1000mL, the high pressure sterilization at 121 DEG C
15min seals and posts label, and 4 DEG C of refrigerators preserve, the term of validity 2 months；Above-mentioned BG11 culture mediums distilled water is diluted 10 times
It is BG11 culture medium dilutions afterwards；
80 grams per liter Tebuconazole suspension seed-coating agent 0.625mL are pipetted, dissolved with BG11 culture medium dilutions and are settled to 100mL appearances
Measuring bottle obtains the storing solution of a concentration of 500mg a.i./L；Pipette successively above-mentioned storing solution 0.300,0.600,1.200,
2.400,4.800mL is settled to 250mL, obtaining concentration is respectively in 250mL volumetric flasks with BG11 culture medium dilutions
0.600,2 times of experiment liquids of 1.20,2.40,4.80,9.60mg a.i./L；
The preparation of S2, algae dilution：The cell concentration that the algae stoste of goat's horn crescent moon algae is measured with counting method of blood cell is 4.00 × 106
A/mL is used in combination sterile water to dilute 100 times and obtains algae dilution, frustule a concentration of 4.00 × 10 in algae dilution4A/mL；
S3, contamination：
Experimental group：The experiment liquid 50mL for the gradient concentration prepared in step S1 is taken to be separately added into the algae dilution of 50mL in step S2
Liquid obtains the biological algae culture medium of gradient concentration；Algae initial concentration is 2.00 × 10 in experiment4A/mL；
Blank control group：50mL algaes dilution is taken to be uniformly mixed so as to obtain biological algae culture medium with 50mL BG11 culture medium dilutions；
The biological algae culture medium of experimental group and blank control group is fitted into triangular flask respectively, is sealed, is placed in masking foil
It is cultivated in intelligent growth cabinet, 16h is placed in control under illumination condition, and 8h is placed under dark surrounds；Daily daytime is small every 2
When shake 1 time, altogether shake 5 times；
S4, test data record：
The water temperature of each concentration group is 22.0 DEG C~23.1 DEG C during measuring experiment, and pH value is 6.74~7.16, the light in incubator
It is 5200Lux~5700Lux according to intensity；The suppressed algae solution cell volume of 48h, 72h becomes smaller after contamination, after observing and recording contamination
0h, for 24 hours, 48h, 72h frustule biological respinse effect, include the intoxication conditions of the inhibiting rate of frustule and frustule；The algae
The intoxication conditions of cell include：Frustule deformity, algae solution bleaches, frustule volume becomes smaller, frustule adhesion or coalescence；
S5, data processing：
Using statistic software SPSS 12.0, using the logarithm of drug concentration as independent variable x, with the probit value of the inhibiting rate after correction
For dependent variable y, virulence regression equation is established；Calculate the half effective concentration EC inhibited to algal grown for examination object50With 95%
Confidence limit；
S6, evaluation of result：
According to EC50Value judges pesticide poison grade；Pesticide is to the growth inhibition toxicity of algae by EC after 72h50Size be divided into three
Grade：High poison：EC50≤0.300mg a.i./L；Poisoning：0.300mg a.i./L < EC50≤3.00mg a.i./L；Low toxicity：
EC50> 3.00mg a.i./L.
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| CN102031280B (en) * | 2009-09-30 | 2014-12-17 | 中国海洋大学 | Method for assessing the acute toxicity of drilling fluid rapidly by utilizing marine microalgae |
| CN102597747A (en) * | 2009-11-09 | 2012-07-18 | 浜松光子学株式会社 | Method for evaluating toxicity of chemical using alga |
| JP5731776B2 (en) * | 2010-09-08 | 2015-06-10 | 浜松ホトニクス株式会社 | Algae cell preparation method and chemical toxicity evaluation kit |
| CN103196884A (en) * | 2013-04-19 | 2013-07-10 | 河北科技大学 | Method for determining biotoxicity of atrazine by utilizing microcystis aeruginosa |
| CN103728284B (en) * | 2013-09-11 | 2016-08-17 | 中国科学院安徽光学精密机械研究所 | A kind of comprehensive water-body toxicity method for quick based on algae chlorophyll fluorescence |
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