Tris(2-ethylhexyl) phosphate

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1997-12-24 to 1998-07-31

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

The study was conducted in accordance with international guidelines and in accordance with GLP. All relevant validity criteria were met.

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

Concentrations: Control, aids control ( 50 mg/L HCO-40), 1.0, 3.2 and 10.0 mg/L. For all concentrations (1 test vessel per concentration), 0.75 mL of test solution was collected four times during the exposure period before and after water renewal, and analysed for test substance concntration.

Vehicle:

yes

Details on test solutions:

100 mg of test substance was dissolved in 250 mg of DMF, 250 mg HCO-40 was added and the solution was mixed. The solution was diluted in 100 mL od of pure water to make a stock of 1000 mg/L. A stock of solvent controls was also made at 5000 mg/L (DMF:HCO-40, 50:50) without test substance. The appropriate amount of stock was added to a 1 L volumetric flask with 1 L Elendt M4 medium to acheive each test concntration. 80 mL of the corresponding solution was used per replicate test vessel (n =10) at each test concentration or control.

Test organisms (species):

Daphnia magna

Details on test organisms:

- Source:National Institute for Environmental Studies, Japan (18 July 1995) and continuously reared at Yokohama Laboratory since obatiainment in Elendt M4 medium.
- Stocks were created with healthy females, the juvenilles from these females were sperated and further cultured until maturation. The neonates (not first brood) from the final subculture were used in the test.
- Test Daphnia were < 24 hours old
- Food: Chlorella vulgaris, 0.15 mg C (organic carbon)/ day
- Exposure method: semi-static (3 times per week, replace the total amount of the test solution)

Test type:

semi-static

Water media type:

freshwater

Total exposure duration:

21 d

Post exposure observation period:

Dissolved oxygen (DO), pH, water temperature and water hardness was measured before and after water renewals.

Parental dphnid mortality and mobility was checked daily. Number of juviniles, dead juveniles, aborted and dormant eggs were counted daily, and removed after counts. Days to first brood was also recorded.

Hardness:

235 - 305 mg CaCO3/L

Test temperature:

20.0 - 20.9 °C

pH:

pH 7.2 - 8.0

Dissolved oxygen:

6.2 -8.8 mg O2/L

Nominal and measured concentrations:

Nominal concentrations: control (0), vehicle control (50 mg/L of vehicle only), 1.00, 3.20, 10.0 mg/L. Measured concentrations were 82-104 %, within 80-120 % as recommended by the guideline. Nominal concentrations were used to estimate and determine the endpoints within the report.

Details on test conditions:

Daphnia were seeded at a density of 1 daphnid/test vessel(80mL) with 10 replicate dpahnids per concnetration. Daphnids were fed Chlorella vulgaris at 0.15 mg C/daphnid/day. Elendt M4 medium was used as the holding water and test medium. The temperature was 20 ± 1 ℃, with a photoperiod of 16 hours light (1200 lux) / 8 hours dark, pH was between 6-9 and dissolved oxygen was 6-9 mg O2/L. The medium of each test vessel/ control was renewed 3 times per week .

Reference substance (positive control):

yes

Remarks:

potassium dichormate

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Remarks on result:

other: Nominal values were adopted for calculation of effect concentration, because test substance concentrations were within ± 20 % of nominal values when they were measured at the time of preparation and before renewal of test solution during exposure period.

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

3.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Remarks on result:

other: Nominal values were adopted for calculation of effect concentration, because test substance concentrations were within ± 20 % of nominal values when they were measured at the time of preparation and before renewal of test solution during exposure period.

Duration:

21 d

Dose descriptor:

EC50

Effect conc.:

> 10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Remarks on result:

other: Nominal values were adopted for calculation of effect concentration, because test substance concentrations were within ± 20 % of nominal values when they were measured at the time of preparation and before renewal of test solution during exposure period.

Details on results:

measured concntrations were 83 - 104 % nominal. Validity criteria were met.
- > 60 live offspring produced per parent female in the control groups
- < 20 % mortality during the test.
Medium parameters remained optimal for Daphnia magna health.

Results with reference substance (positive control):

48 hour EC50 = 0.66 mg/L

Table 1: Measured concentration as % nominal concntration.

			Measured concentration
Nominal Concentration		DAY	0	2	7	9	9	12	19	21
Control			<LOQ	<LOQ	<LOQ	<LOQ	<LOQ	<LOQ	<LOQ	<LOQ
Solvent control			<LOQ	<LOQ	<LOQ	<LOQ	<LOQ	<LOQ	<LOQ	<LOQ
1.00			83	83	88	87	96	89	96	85
3.20			85	82	95	87	100	83	91	86
10.0			96	84	94	92	102	95	104	104

Cumulative Numbers of Juveniles Produced per Adult Alive for 21 Days in Each Test Vessels and Results of Statistical Comparison of the Mean Values (Dunnett’s Multicomparison Test)

Nominal concentration mg/L
Vessel no.	Control	Solvent control	1.00	3.20	10.0
1	130	D	167	134	D
2	142	166	172	135	98
3	142	160	149	138	D
4	125	168	144	121	101
5	148	163	126	148	84
6	137	132	142	133	125
7	145	155	159	154	115
8	140	138	166	138	137
9	160	160	148	117	102
10	150	140	163	140	108
MeanS.D.	141.910.0	153.613.4	153.614.3	135.811.0	108.816.6
Inhibition rate %Significant difference			0.0-	11.6*	29.2**

D: Were not included for calculatioa because the parentalDaphniawas dead during a 21 -days testing period

-: Indicates no si gnificant difference

*: Indicates a significant difference (α = 0.05) from the solvent control

**: Indicates a significant difference (α = 0.01) from the solvent control

No Observed Effect Concentration (NOEC): 1.00 mg/L

Lowest Observed Effect Concentration (LOEC): 3.20 mg/L

Validity criteria fulfilled:

yes

Conclusions:

The 21 day LC50 was estimated to be > 10 mg/L. The NOEC and LOEC for reproduction were 1 mg/L and 3.2 ,g/L, repsectively.

Executive summary:

The toxicity of tris(2 -ethylhexyl)phosphate to Daphnia magna was studied during a Daphnia magna 21 day reproduction test (according to OECD guideline 211; Daphnia manga Reproduction Test). Test solutions were made in Elendt M4 medium, concntrations were 0, 0 (with solvent control), 1, 3.2 and 10 mg/L. A dispersant was used to enhance the solubility of the test item in the test medium. The NOEC and LOEC were determined using the Dunnett's test.

The LC50 was estimated to be > 10 mg/L; and the reproductive NOEC and LOEC were determined to be 1 mg/L and 3.20 mg/L, respectively.

Description of key information

NOEC, LOEC and EC50 (21 d Daphnia magna reproduction test); 1 mg/L, 3.2 mg/L  and >10 mg/L; OECD 211, Anon (1998)

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

1 mg/L

Additional information

The toxicity of the substance to Daphnia magna during a 21 d reproduction was assessed in line with OECD 211 guidelines (Anon, 1998). The NOEC was determined to be 1 mg/L, derived from a significant reduction in fecundity at 21 days. A dispersant was used to enhance the solubility of the test item in test medium and the resulting NOEC is orders of magnitude higher than the limit of water solubility.

Due to the substance not causing reproductive effects at, or above, the limit of water solubility the substance was concluded as having no long-term toxicity to aquatic invertebrates and no hazard has been identified for this endpoint in accordance with REACH Annex I (section 3).