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Toxicity to aquatic algae and cyanobacteria

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Reference
Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
This study is used for read-across and therefore has been assigned a reliability of 2 (reliable with restrictions). The study, if used in support of terephthalic acid, has a reliability of 1 (reliable without restriction).
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
not reported
Vehicle:
no
Details on test solutions:
The stock solution was prepared once before the beginning of the test. The test material used was dissolved in distilled water to give a nominal concentration of 111.662 mg/L for the range finding study and 1112.82 mg/L for the definitive study. The pH of the solution was adjusted using sodium hydroxide to give a pH of approximately 8.0. The stock solution was then diluted further to give a range of concentrations for the test solutions.
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
Supplier; Institut fur Pflanzenphysiologie, University Gottingen, D-3400 Gottingen. Solid and liquid stocks of S.subspicatus have been cultured at Battelle Europe since August 1987.

Liquid cultures were prepared by adding 10(+4) of a 7 days old liquid culture to 100 mL algal medium. The liquid cultures were incubated in 300 mL Erlenmeyer flasks under a uniform quantum flux density of 120 ±10 µE.sec-1.m-2 provided by the light source OSRAM L36W/25 in a growth cabinet 20 ±1°C; the daily photoperiod was 16 h.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Hardness:
not reported
Test temperature:
22.4/22.6°C min/max.

pH:
approximately 8.0
Dissolved oxygen:
not reported
Salinity:
freshwater
Nominal and measured concentrations:
Nominal: 62.5, 125, 250, 500 and 1000 mg TS/L
Details on test conditions:
For the range finding study the following conditions were maintained;
- Test vessel:
- Erlenmeyer flasks containing 100 mL test solution covered with a cotton plug.:
- No. of cells per mL test solution at beginning of test; 10(+4)
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
-Light: light/dark: (24/0 h) OSRAM L36W/25
-Temperature: 23 ±1°C
-Age of the pre-culture: 4 d
-Number of cells per mL in the pre-culture before inoculating the test solutions: 2.25*10(+6)
-Volume of pre-culture: 0.44 mL

For the definitive test the following conditions were maintained:
- Test vessel: Erlenmeyer flasks containing 300 mL test solution covered with a cotton plug.
-age of the preculture: 4 d
-Volume of pre-culture containing 1*10(+6): 0.150 mL.
-Number of cells at beginning of test 10(+4)
- No. of vessels per concentration (replicates):4
-Number of replicates in the control: 4
-Light: light/dark (24/0 h) OSRAM L36W/25
-Temperature 23 ±1°C

The growth inhibition was the parameter determined. This was expressed as the effect of the test substance concentration that reduced the cell number compared to the control was a certain percentage. Of particular interest were the test substance concentrations that reduced the cell number by 90% (EC90), 50% (EC50) and 0% (EC10).
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
> 668 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks:
ErC50
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
> 668 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
cell number
Remarks:
EbC50
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
668 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
other: growth rate and cell number
Details on results:
Before conducting the test a range-finding study was performed to estimate the range of the test substance which effects the growth of the algae. The highest concentration tested causing no obvious inhibition of growth within the period of the range-finding test relative to the control was 100 mg/L. In none of the concentration levels tested within the period of the range-finding test an effect relative to the control could be determined.

Selection of the test substance concentrations for the definitive assay were based upon the results of the range finding test. The nominal concentrations to which the test organisms were exposed were 62.5, 125.0, 250.0, 500.0 and 1000.0 mg/L.

The analytically determined actual concentrations of the test material; at the beginning of the study were found to be within the range of 70% and 104% with an average of 92.2%. After 96 hours of exposure, analysed concentrations of the test material were considerably lower than at 0 hours. They were found to be <10 mg/L for the nominal concentrations 62.5 and 250 mg/L. The recovery for 1000.0 mg/L was 40.5% on average. It is assumed that large amounts of the test material was absorbed by the test organisms.

Based on the nominal concentrations the NOEC obtained was >1000 mg/L terephthalic acid and terephthalic sodium salt. Based on the measured average concentration (40.9%) of the highest concentration level tested (nominally 1000 mg/L) the NOEC was >409 mg/L terephthalic acid and terephthalic sodium salt.

In the report, the endpoint is given as >409 mg/L, apparently the mean measured value at the highest concentration, but based on the 96-h data only, i.e. (344.48 + 473.15)/2. The mean should include the 0-h data as well, i.e. (948.58 + 905.52 + 344.48 + 473.15)/4 = 668 mg/L.
Results with reference substance (positive control):
The study with potassium dichromate was performed in April 1992. EC10 (72 h) values for potassium dichromate obtained from different laboratories range between 0.9 mg/L and 1.9 mg/L.
Reported statistics and error estimates:
not reported

The tables below show the cell number N*10(+4) per mL for the replicates a, b, c and d after approximately 24, 48, 72 and 96 h.

 

 

Cell density measurements: Control

 

Time (h)

Replicate

a

b

c

d

24

2.0

7.0

6.0

7.0

48

13.0

16.0

9.0

10.0

72

98.0

109.0

78.0

88.0

96

280.0

290.0

370.0

380.0

 

Cell density measurements: 62.5 mg TPA/L

 

Time (h)

Replicate

a

b

c

d

24

5.0

3.0

3.0

6.0

48

7.0

12.0

15.0

9.0

72

105.0

93.0

89.0

96.0

96

310.0

450.0

370.0

350.0

 

Cell density measurements: 125.0 mg TPA/L

 

Time (h)

Replicates

a

b

c

d

24

6.0

9.0

11.0

8.0

48

18.0

17.0

13.0

8.0

72

121.0

103.0

98.0

91.0

96

500.0

240.0

390.0

370.0

 

Cell density measurements: 250.0 mg TPA/L

 

Time (h)

Replicates

a

b

c

d

24

6.0

6.0

4.0

7.0

48

9.0

11.0

11.0

14.0

72

92.0

90.0

83.0

88.0

96

260.0

440.0

330.0

320.0

 

Cell density measurements: 500.0 mg TPA/L

 

Time (h)

Replicates

a

b

c

d

24

6.0

3.0

8.0

5.0

48

17.0

12.0

13.0

10.0

72

84.0

78.0

95.0

86.0

96

350.0

240.0

270.0

280.0

 

Cell density measurements: 1000 mg TPA/L

 

Time (h)

Replicates

a

b

c

d

24

4.0

3.0

8.0

10.0

48

7.0

6.0

9.0

8.0

72

84.0

79.0

76.0

90.0

96

230.0

330.0

290.0

330.0

 

Measured concentrations of terephthalic acid during a 96 hour, static toxicity test with

Scenedesmus subspicatus.

Exposure Period (h)

Terephthalic Acid Nominal mg/L

Measured Terephthalic Acid mg/L

Percentage of nominal %

0

62.5

43.55

58.35

70

93

0

250

250.31

259.74

100

104

0

1000

948.58

905.52

95

91

96

62.5

< 10.0

< 10.0

< 16

< 16

96

250

< 10.0

< 10.0

< 4.0

< 4.0

96

1000

344.48

473.15

34

47

In the report, the endpoint is given as >409 mg/L, apparently the mean measured value at the highest concentration, but based on the 96-h data only, i.e. (344.48 + 473.15)/2. This is incorrect; the mean should include the 0-h data as well, i.e. (948.58 + 905.52 + 344.48 + 473.15)/4 = 668 mg/L.

 

 

 

 

Validity criteria fulfilled:
yes
Conclusions:
The static 96-hour EbC50 and ErC50 of the test substance (converted to its sodium salt prior to exposure) to Scenedesmus subspicatus) were both >668 mg/L, based on the mean measured concentrations at the maximum treatment applied. The NOEC was 668 mg/L (mean measured).
Executive summary:

Desmodesmus subspicatus (formerly known as Scenedesmus subspicatus) was exposed in a static test system over a period of 96 hours to the test substance (TS) after conversion to its sodium salt. The nominal exposure concentrations were 62.5, 125, 250, 500 and 1000 mg TS/L. There were 4 replicate test vessels per treatment, each initially inoculated with 10,000 algal cells/mL. Exposure concentrations were verified analytically at the start and end of the test in three of the treatments. The analytically determined initial concentrations of the test substance were between 70% and 104% of nominals. Residual concentrations at test-end were reduced to below 16% and 4% of nominals in two of the three treatments that were analysed and to between 34% and 47% of nominal at the maximum concentration which defined the endpoint of the study.

The EbC50(96 h) and ErC50(96 h) endpoints both exceeded 668 mg/L test substance (as sodium terephthalate), based on the mean measured exposure concentration at the highest treatment. Based on the absence of statistically significant effects on algal growth at all of the applied treatments, the NOEC was 668 mg/L (mean measured).

Description of key information

Key value for chemical safety assessment

EC50 for freshwater algae:
668 mg/L
EC10 or NOEC for freshwater algae:
668 mg/L

Additional information

The study was conducted using terephthalic acid, the primary degradate of terephthaloyl dichloride, In an OECD 201 Guideline study under GLP, Desmodesmus subspicatus (formerly known as Scenedesmus subspicatus) were exposed in a static test system over a period of 96 hours to the test substance after conversion to its sodium salt. The nominal exposure concentrations were 62.5, 125, 250, 500 and 1000 mg/L with each vessel initially inoculated with 10000 algal cells/mL. Exposure concentrations were verified analytically at the start and end of the test in three of the treatments. Initial concentrations were between 70% and 104% of the nominal values. Residual concentrations at test-end were reduced to below 16% and 4% of the target nominal concentration in two of the three treatments that were analysed and to between 34% and 47% of nominal at the maximum concentration which defined the endpoint of the study.

The EbC50 (96 h) and ErC50 (96 h) endpoints both exceeded 668 mg/L (converted to its sodium salt prior to exposure), based on the mean measured exposure concentration at the highest treatment. Based on the absence of statistically significant effects on algal growth at all of the applied treatments, the NOEC was 668 mg/L (mean measured).