Trimethyl orthoformate

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2002-02-07 to 2002-04-18

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

1984

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Two runs were performed. In the first run test concentrations of 6.5, 11, 19, 35, 60, 100 and 171 mg/L were used. In the second run 265, 473 and 833 mg/L
- Sampling method: Analytical samples were taken at time 0 and after 48 hours. Observations of the Daphnia were made after 24 and 48 hours

Vehicle:

no

Details on test solutions:

One test solution was prepared for each of the two runs. For the first run 0.515 mL of test item were dissolved in 500 mL of synthetic fresh water without NaHCO3. For the second run 0.309 mL were dissolved in 300 mL. The preparation of the water for the test solutions differed from the preparation of the water in the test with organisms, so that negative effects due to inorganic carbon could be avoided during the analytical determinations.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
Daphnia magna Strauss, clone 5

- Source: Originally, received in 1991 from Dr. N. Caspers (Bayer AG, Leverkusen, Germany); subsequently bred in house
- Age at study initiation (mean and range, SD): Less than 24 hours

- Method of breeding: females maintained in M4 medium in 1 L beakers. They are fed Scenedesmus subspicatus, in amounts that it is consumed by the animal
- Feeding during test: no

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

14°dH = 250 mg CaCO3/L

Test temperature:

20±1°C

pH:

Run 1:
pH at the beginning of the test 7.6-7.9
pH at the end of the test 6.5-7.8

Run 2:
pH at the beginning of the test 7.4-7.9
pH at the end of the test 5.5-7.5

Dissolved oxygen:

Run 1:
at the beginning of the test 7.7-7.9
at the end of the test 7.3-7.8

Run 2:
at the beginning of the test 8.6-8.8
at the end of the test 6.9-7.9

Nominal and measured concentrations:

Concentrations: Two runs were performed. In the first run test concentrations of 6.5, 11, 19, 35, 60, 100 and 171 mg/L were used. In the second run 265, 473 and 833 mg/L. Values at the start and end did not deviate by more than 20 % and recoveries were >80%. Therefore nominal values were used.

Details on test conditions:

Test vessels: 22 ml head-space test tubes with 20 ml volume to minimize the loss of the volatile test item
The vessels were closed tightly (gas tight septum).
run 1: 7 concentrations + 1 control
run 2: 3 concentrations + 1 control
20 organisms in 4 parallel sets of 5 each
Density of occupation per animal: ca. 4 ml per test organism
Test in the dark
No aeration

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

690 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CL: 473-833

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 833 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

EC0

Effect conc.:

473 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

EC50 values were calculated graphically.

Results with reference substance (positive control):

Potassium dichromate is tested as reference item quarterly. The results from the last test demonstrated that the sensitivity lies in the normal range.

Cumulative number of immobilised individuals during the tests, as absolute and %

	Concentration (mg/L)	24h			48h
		Individuals used	Individuals immobilised	% immobilised	Individuals used	Individuals immobilised	% immobilised
Run 1	Control	20	0	0	20	0	0
	6.5	20	0	0	20	0	0
	11	20	0	0	20	0	0
	19	20	0	0	20	0	0
	35	20	0	0	20	0	0
	60	20	0	0	20	0	0
	100	20	0	0	20	0	0
	171	20	0	0	20	0	0
Run 2	Control	20	0	0	20	0	0
	265	20	0	0	20	0	0
	473	20	0	0	20	0	0
	833	20	0	0	20	15	75

Validity criteria fulfilled:

yes

Remarks:

In the control less than 10 % of the Daphnia had been immobilised or trapped at the watersurface. The dissolved oxygen concentration at the end of the test was > 60 % (5.45 mg/I) of the air saturation value. The concentration of the test item had been sat

Conclusions:

Trimethylorthoformate was found not acute toxic to daphinds.

Executive summary:

Acute toxicity, 48h, of Trimethylorthoformate (CAS 149-73-5, 99.5% purity) to daphnids (Daphnia magna) was tested under GLP according to guideline EU Method C.2 (Acute Toxicity for Daphnia, 1992) or OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test, 1984). The study is considered valid and conclusive.

Two runs of the test were performed since no effects were observed in the concentration range tested in the first run. The final concentration ranges were 6.5, 11, 19, 35, 60, 100 and 171 mg/L in run 1 and 265, 473 and 833 mg/L in run 2. Analytical samples were taken after 0 and 48h. Recoveries showed that nominal values could be used.

Observations of the immobility of the daphnia were performed after 24 and 48h. The acute toxicity EC50 to Daphnia magna after 48 hours was 690 mg/L, with a range of 473-833 mg/L. The 24 hour EC50 was > 833 mg/L. After 48 hours the highest concentration at which no immobilisation of the Daphnia was observed was 473 mg/L.

In conclusion the test item was not found to be acutely toxic to daphnids.

Based upon the classification criteria according to the Regulation (EC) No 1272/2008 of the European Parliament and of the Council of 16 December 2008, the test item is considered not classified on the basis of this study.

Description of key information

24h EC50 > 833mg/L
48h EC50 = 690mg/L

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

690 mg/L

Additional information

Acute toxicity, 48h, of Trimethylorthoformate (CAS 149-73-5, 99.5% purity) to daphnids (Daphnia magna) was tested under GLP (Enste and Diefenbach 2002b)according to guideline EU Method C.2 (Acute Toxicity for Daphnia, 1992) or OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test, 1984). The study is considered valid and conclusive.

Two runs of the test were performed since no effects were observed in the concentration range tested in the first run. The final concentration ranges were 6.5, 11, 19, 35, 60, 100 and 171 mg/L in run 1 and 265, 473 and 833 mg/L in run 2. Analytical samples were taken after 0 and 48h. Recoveries, based on dissolved organic carbon, showed that nominal values could be used.

Observations of the immobility of the daphnia were performed after 24 and 48h. The acute toxicity EC50 to Daphnia magna after 48 hours was 690 mg/L, with a range of 473-833 mg/L. The 24 hour EC50 was > 833 mg/L. After 48 hours the highest concentration at which no immobilization of the Daphnia was observed was 473 mg/L.

Enste and Diefenbach (2002b) describe an acute daphnia toxicity study conducted at a pH that varied between 5.5-7.9 over the period of the study (48h). From the hydrolysis as a function of pH study reported by Fischer (2004), included in section 5.12 of this dossier, it is known that the stability of Trimethyl orthoformate within this pH range is very poor. Indeed one can calculate that the half-life in this pH range would be <12h. The analytical method employed by Enste and Diefenbach was based on measuring the dissolved organic carbon (DOC) and they reported that >80% of the original DOC remained after 48h in the test medium. As such the acute daphia toxicity study of Enste and Diefenbach can be viewed more in terms of a study on trimethyl orthoformate and its metabolites formed from the hydrolysis in water.

The known metabolites of the aqueous hydrolysis of trimethyl orthoformate in the pH range 5.5-7.9 are methanol and formic acid. It is clear from the IUCLID dataset (IUCLID dataset reported for council regulation EEC No. 793/93) on methanol that it has very low toxicity to invertebrates with several studies reporting the 48h EC50 in excess of 10000mg/l. However, formic acid does display some toxicity to daphnia. The IUCLID dataset for formic acid reports the 48h EC50 to be in the range of 120-150mg/l and the 24h EC50 to be in the range of 410mg/l. 

Given that 1 mole of trimethyl orthoformate should liberate 1 mole of formic acid through hydrolysis with water and that the highest test concentration used was 833mg/l one can speculate that the daphnia toxicity observed in the Enste and Diefenbach study is due to released formic acid through hydrolysis of the test substance. Further evidence for such decomposition is the acidification of the test medium over the 48h period.

In conclusion the test item was not found to be acutely toxic to daphnids.

Based upon the classification criteria according to the Regulation (EC) No 1272/2008 of the European Parliament and of the Council of 16 December 2008, the test item is considered not classified on the basis of this study.