1,3-dihydro-1,3-dioxo-2H-isoindole-2-hexanoic acid

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10-01-94 - 14-01-94

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP study according to internationally accepted guideline. Measured concentrations in test solution deviate > 20% from nominal concentrations.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

1,3-dihydro-1,3-dioxo-2H-isoindole-2-hexanoic acid

EC Number:

224-675-8

EC Name:

1,3-dihydro-1,3-dioxo-2H-isoindole-2-hexanoic acid

Cas Number:

4443-26-9

Molecular formula:

C14H15NO4

IUPAC Name:

6-(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)hexanoic acid

Details on test material:

Name of test material (as cited in study report): PAC
Lot/Batch number: R719
Purity: 95%
Water solubility: 200 mg/l

Sampling and analysis

Analytical monitoring:

yes

Test solutions

Vehicle:

no

Details on test solutions:

0.53 g of test material was dispersed in 1 litre of ISO medium to give a stock solution of 500 mg/l nominal concentration. High energy sonication was used to assure the homogeneity of the dispersion. NaOH 1 M was used to adjust the pH to 6.5. A range of test solutions with nominal concentration 32-320 mg/l were prepared by dilution of the stock solution.

Test organisms

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Strain: not reported
- Laboratory culture: Yes
- Source: not reported
- Age of inoculum (at test initiation): no data
- Method of cultivation: not reported
- Pre-treatment: 3 days before the experiment an inoculum was prepared and incubated under the test conditions
- Initial cell concentration: Average = 1.28 * 10E+04 cells/ml

Study design

Test type:

not specified

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test conditions

Test temperature:

19-24 °C

pH:

7.0-7.6 at start of test
6.1-7.4 at end of test

Nominal and measured concentrations:

Exposure concentrations vary more 20% from nominal values.

- Test concentrations - nominal: 0, 32, 56, 100, 180 and 320 mg/L
- Test concentrations - measured at 0h: 2.6, 30.2, 49.1, 86.5, 164.0, 286.6 mg/L
- Test concentrations - measured at 96h: 0, 0.2, 0, 39.1, 90.2, 225.0 mg/L

Details on test conditions:

TEST SYSTEM
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: no data
- Aeration: no data
- Initial cells density:Average = 1.28 * 10E+04 cells/ml
- Control end cells density: Cell concentration in control cultures increased at least by a factor of 16 within 3 days
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- ISO medium

OTHER TEST CONDITIONS
- Photoperiod: no data
- Light intensity: not measured

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: microscope with counting chamber

TEST CONCENTRATIONS
- Test concentrations - nominal: 0, 32, 56, 100, 180 and 320 mg/L
- Test concentrations - measured at 0h: 2.6, 30.2, 49.1, 86.5, 164.0, 286.6 mg/L
- Test concentrations - measured at 96h: 0, 0.2, 0, 39.1, 90.2, 225.0 mg/L

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

140 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 110-180 95% CL

Effect concentrations 2

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 320 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 3

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

130 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 110-150 95% CL

Effect concentrations 4

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

> 320 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 5

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

79 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 65-97 95% CL

Effect concentrations 6

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

> 256 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 7

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

56 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Effect concentrations 8

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

25 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Details on results:

Nominal concentrations: 96 h NOEC = 56 mg/l. The tested concentrations were 32, 56, 100, 180, 320 mg/l. 

Since the concentrations decreased substantially during the 96 hour exposure period, effect concentrations based on nominal concentrations are probably an understimate of the toxicity of the test substance. A more relevant assessment would be to base the effects on mean measured concentrations. Although in this test this assessment is limited because the two lowest concentrations had fallen to zero, it is considered acceptable since inhibition of growth was probably only significant at 100 mg/l and above. Based on mean measured concentrations the 96 h EbC50 is 79 mg/L and ErC50 >256 mg/l. The 96 h NOEC was 25 mg/L also using mean measured values.

Reported statistics and error estimates:

EC50 calculated by means of the modified probit method.

Any other information on results incl. tables

Cell concentration data:

Test­-Substance Concentration (nominal) [mg/l]	Cell concentrations (mean values) [10^4cells/ml]
	measured     0 h         24 h         48 h         72 h      96 h
0	1.25	6.26	34.9	174	516
32	1.18	5.61	33.0	157	547
56	1.24	6.26	35.8	187	597
100	1.30	4.85	25.0	127	267
180	1.39	4.30	18.5	59.6	101
320	1.29	4.28	3.52	8.11	15.0

Effect data: cell multiplication inhibition:

Based on nominal concentrations

                         Biomass                       Growth rate

                         E_bC50 (95% CL)            E_rC50 (95% CL)

                           [mg/l]                           [mg/l]

24 h                    180(130-270)                280 (200-480)                               

48 h                    150 (120-210)               290 (200-540)

72 h                    140 (110-180)               >320 (n/a)

96 h                    130 (110 -150)              >320 (n/a)

 

Based on measured concentrations:

Biomass                        Growth rate

                         E_bC50 (95% CL)            E_rC50 (95% CL)

                           [mg/l]                           [mg/l]

96 h                    79(65 -97)                      >256 (n/a)

Applicant's summary and conclusion

Executive summary:

The potential of PAC to inhibit algae growth was studied in the unicellular green alga S. capricornutum. Algae were exposed to nominal concentrations ranging from 32 to 320 mg/l for 96 consecutive hours. 3 samples/concentration and 6 samples for controls were used. The cellular growth was evaluated by measuring the cell density. The mean area under the curve and the mean specific growth rate were calculated.

In this study, the 96h NOEC was found to be 25 mg/L (measured concentration), and the EC50 based on the biomass is 79 mg/L (measured concentration). The average growth rate of controls was > 400 times after 72 hours, fulfilling the validity criteria.