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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

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Administrative data

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
8th - 11th July 1997
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study to GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1997

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): L-2-Chloropropionic acid
- Physical state: Liquid
- Analytical purity: 97.2% w/w
- Purity test date: 23rd May 1997
- Lot/batch No.: Bx 19
- Expiration date of the lot/batch: Not specified
- Stability under test conditions: Not specified
- Storage condition of test material: Refrigeration in original container

Sampling and analysis

Details on sampling:
Samples were taken of each test solution and analysed for concentration of the test substance.
At the end of the test each blank solution was sampled and analysed.
Algal cell densities were recorded on days 1, 2 and 3.

Test solutions

Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 120 mg/L solution of the test substance was prepared by the direct addition of test substance to sterile culture. The pH was adjusted to pH 7.65 by the addition of 1M NaOH. Lower test concentrations were prepared by the addition of aliquots of the neutralised nominal 120mg/L solution to 1000ml of sterile medium.

- Controls: The control consisted of culture medium only.
-Blanks: Blank vessels (without algal inoculum) for each control and treatment were incubated concurrently.


Test organisms

Test organisms (species):
Navicula pelliculosa
Details on test organisms:
TEST ORGANISM
- Common name:Navicula pelliculosa
- Strain: UTEX 667
- Source (laboratory, culture collection): Laboratory cultures
- Age of inoculum (at test initiation): Algae in the 'exponential growth stage' were used.
- Method of cultivation: axenic0 conditions

ACCLIMATION
- Acclimation period: Not specified
- Culturing media and conditions (same as test or not): As test

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h

Test conditions

Hardness:
Not specified
Test temperature:
Daily temperature measurements in the incubator ranged from 24.0 - 24.1 oC.
The hourly temperature measurements, recorded automatically, ranged from 23.7 - 23.9 oC.
pH:
The pH of the test solutions at the start of the test were in the range 7.7 - 8.4, at the end of the test the range was 8.0 - 9.6. During the course of the test, the control solutions altered by <0.5 pH units, however increases of up to 1.9 pH units were observed in the test solutions, but were considered not to have affected the validity of the test.
Dissolved oxygen:
Not specified
Salinity:
Not specified
Nominal and measured concentrations:
Nominal Concn(mg/L)/ Measured Initial Concn (mg/L)
0 <0.25
7.5 8.1
15 17.0
30 32
60 60
120 130
Details on test conditions:
TEST SYSTEM
- Test vessel: borosilicate glass conical flasks, 250ml capacity
- Type (delete if not applicable): closed with polyurethane foam bungs
- Material, size, headspace, fill volume: 100mls fill volume
- Aeration: Not specified
- Initial cells density: 1.04 x 10 4 cells/ml
- Control end cells density: 170 x 10 4 cells/ml
- No. of organisms per vessel: nominal cell density of 1.00 x 10 4 cells/ml
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6
- No. of vessels per vehicle control (replicates): Not applicable

GROWTH MEDIUM
- Standard medium used: yes – as described by Miller W E et al. Selenastrum capricornutum Printz. Algal Assay Bottle Test: Experimental Design, Applications and Data Interpretation Protocol EPA-600/9-78-018, Corvallis, OR


OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: yes prior to addition of inoculum.
- Photoperiod: Continuous illumination
- Light intensity and quality: 8980 lux
EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter]: Electronic particle counting using a Coulter counter mode ZB

TEST CONCENTRATIONS
- Test concentrations: 0, 7.5, 15.0, 30, 60 and 120mg/L

Results and discussion

Effect concentrationsopen allclose all
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
120 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
120 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
biomass
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 120 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 120 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
biomass

Any other information on results incl. tables

The algal cell densities are summarised in table 1.

Table 1.

Nominal Conc of L-2-Chloropropionic acid (mg/L)

Replicate

 

 

Algal Cell densty (x104cells/ml)

24 hours

48 hours

72 hours

Culture Medium Control

A

3.10

33.6

147

B

3.11

40.9

164

C

2.46

51.9

234

D

3.73

35.3

158

E

2.69

45.2

171

F

2.86

36.1

144

Mean

2.99

40.5

170

7.5

A

3.10

48.4

174

B

3.13

51.0

176

C

3.27

39.1

178

Mean

3.17

46.2

176

15.0

A

3.56

45.7

179

B

3.50

47.3

175

C

3.42

47.5

176

Mean

3.49

46.8

177

30

A

3.55

44.8

192

B

3.33

52.3

185

C

3.25

41.7

193

Mean

3.38

46.3

190

60

A

4.15

50.3

188

B

4.31

60.7

140

C

4.23

54.4

180

Mean

4.23

55.1

169

120

A

3.78

48.3

193

B

4.28

45.6

182

C

3.84

44.9

182

Mean

3.97

46.3

186

 

The mean cell densities were plotted as growth curves and the area under the curve calculated using the formula cited in OECD guideline 201. The mean area under the growth curve is shown in Table 2.

 

Table 2.

Nominal Conc of L-2-Chloropropionic acid (mg/L)

Mean area under growth curve

 (0 – 72 hour)

% of control

Control

125.7

-

7.5

134.7

107

15.0

136.1

108

30

142.0

113

60

141.4

112

120

140.5

112

 

 

There were no significant differences from the control.

  

The mean growth rates over the test period are shown in Table 3.

Table 3.

Nominal Conc of L-2-Chloropropionic acid (mg/L)

Mean Growth Rate  

(0 – 72 hour)

% of control

Control

1.698

-

7.5

1.710

101

15.0

1.712

101

30

1.736

102

60

1.698

100

120

1.728

102

There were no significant differences from the control.

Applicant's summary and conclusion

Conclusions:
The No Observed Effect Level (NOEC) for L-2-Chloropropionic acid based on the growth curve or the growth rate was 120 mg/L.
The EC50 (biomass) and the EC50 (growth rate ) were both >120mg/L.
Executive summary:

The effect on algae of L-2-Chloropropionic acid was studied on the freshwater diatom Navicula pelliculosa using a growth inhibition test according to OECD 201 over a period of 72 hours.

 

The test was performed at concentrations of 0, 7.5, 15.0, 30, 60 and 120 mg /l using a final volume of 100 ml in triplicate, each inoculated with algae at a nominal cell concentration of 1.0 x104cells/ml. The culture flasks were placed in an incubator at 24 +/-2oC at continuous illumination and kept in suspension by constant orbital shaking. The cell densities were measured by a Coulter counter.

The mean cell densities were plotted as a growth curve. The area under which was calculated in order to determine the NOEC and EbC50 values.

NOEC = 120 mg/L

EbC50 = >120 mg/L

 

The average (days 0 -3) growth rate was calculated. The NOEC and ErC50 based on growth rate were:

NOEC = 120 mg/L

ErC50 = >120mg/L