Tall oil, sodium salt

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2005-07-18 to 2005-10-24

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Control, 12.5 mg/L, 50 mg/L and 100 mg/L test loading rates WAFs.

- Sampling method: The vessels used for Exposure Concentration Analysis, including the Control and WAFs prepared at the various loading rates, were kept under the same conditions as the test vessels but without fish present. These were sampled for TOC analysis (TOC = Total Organic Carbon) and quantitative GCMS analysis at the start and end of test periods 0 - 24 h and 72 - 96 h to assess the stability of exposure concentrations.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION

- Method: The test substance was comprised of many chemicals, some of which have low water solubility. The exposures were therefore carried out using WAFs (Water Accommodated Fractions). These were obtained by stirring the test article with de-chlorinated tap water for 24 h and then allowed to stand for 1 h in order for phase separation to take place. To ensure that emulsification and dispersion of undissolved parts of the test materials did not occur, the surface vortex developed by stirring was set at ~ 10% of the total depth of the water. The exposure solution WAFs were prepared in 20 L glass fish tanks, supplied with glass tubes to allow the separation of the water phase by siphoning. Solutions were replenished after each 24 h exposure period. The duration of the stirring and phase separation phases of WAF preparation were investigated in a preliminary study In a preliminary with a test article of a comparably low water solubility (Sylvablend TM PF 40, STZ No. 01-05-001; study sponsor Arizona Chemical).

- Controls: dilution water used to dilute WAFs, that is de-chlorinated tap water.

- Evidence of undissolved material: The test substance was not fully soluble; to avoid adverse physical effects of undissolved or emulsified test material on the animals, the test was carried out with aqueous extracts (WAFs).

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM

- Common name: Zebra Fish

- Source: Max-Planck-Institut für Entwicklungsbiologie, Tübingen

- Length at study initiation: 3.0-3.5 cm

- Feeding during test: No

ACCLIMATION

- Acclimation period: >12 days, the animals were kept in a climate-controlled room

- Acclimation conditions: same as test

- Health during acclimation: mortality < 1 %

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Post exposure observation period:

3 days

Hardness:

Total hardness: 8.97°dH

Test temperature:

23 °C ± 2°C

pH:

8.1-8.5

Dissolved oxygen:

75-95%

Salinity:

Not applicable.

Nominal and measured concentrations:

Nominal concentrations: Control, 6.25 mg/L, 12.5 mg/L, 25 mg/L, 50 mg/L and 100 mg/L WAFs.

Loading rates: Exposure concentrations of dissolved CTO components, determined by GCMS analysis decreased (< 20 % of initial values) by the end of an exposure period (after 24 h), however they increased with concentration. TOC analysis confirmed that concentrations increased with increasing concentration and time, with the exception of 12.5 mg/L where concentrations decreased in testing period 72-96 h (see Tables 4 and 5).

The results of the test were reported and interpreted with reference to nominal loading rates.

Details on test conditions:

TEST SYSTEM

- Test vessel: fish tank

- Material, size, headspace, fill volume: solid glass, 20 L filled with 7 L of test substance.

- Aeration: None

- Renewal rate of test solution: every 24 h

- No. of organisms per vessel: 7 animals per vessel/concentration

- No. of vessels per concentration: 1

- No. of vessels per control: 1


TEST MEDIUM / WATER PARAMETERS

- Total organic carbon: See Table 4.

- Metals: Hg/Cd/Pb/Cu/Ni: < 0.001 mg/L

- Intervals of water quality measurement: start and end of each 24 h testing period, up to 96 h.


OTHER TEST CONDITIONS

- Photoperiod: 14 h light, 10 h dark using Gro-Lux fluorescent tubes

EFFECT PARAMETERS MEASURED: lethality and any sublethal effects (shoaling behaviour, activity, balance, surfacing, breathing frequency, apathy)


TEST CONCENTRATIONS

- Range finding study

- Test concentrations: the range finding study for this test was conducted with CTO sample 1 by Steinbeis-Transfer-zentrum Angewandte und Umwelt-Chemie in 2005 (STZ-No. 11-05-003, study sponsor Arizona Chemical), in the same testing period, with test concentrations: control, 10 mg/L, 100 mg/L, 1000 mg/L.

- Results used to determine the conditions for the definitive study: 0% lethality at 10 mg/L and 100 mg/L, 100 % lethality after 24 h at 1000 mg/L.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LL50

Effect conc.:

20 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

water accommodated fractions (WAFs) loading rate

Basis for effect:

mortality (fish)

Key result

Duration:

96 h

Dose descriptor:

NOELR

Effect conc.:

6.25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

water accommodated fractions (WAFs) loading rate

Basis for effect:

mortality (fish)

Details on results:

- Behavioural abnormalities:
- 6.25 mg/L: shoaling behaviour and activity were normal at 3 h after beginning of the test. At the end of observation period 0–24 h swimming movements were slightly slower compared to the control. Shoaling behaviour and activity were comparable to the control at the end of observation period 24–48 h. Swimming movements again were slightly slower compared to the control at the end of observation periods 48–72 h and 72-96 h.
- 12.5 mg/L: shoaling behaviour and activity were slightly reduced at 3 h after beginning of the test and the fish swam in the lower part of the vessel. At the end of observation periods 0-24 h and 24–48 h activity was slightly reduced, breathing frequency was heightened and the fish used the lower part of the vessel for swimming action. At the end of observation periods 48-72 h and 72–96 h activity was reduced, breathing frequency was heightened and the fish used the lower part of the vessel for swimming action.
- 25 mg/L: at the end of the first time period (0-24 h) the fish showed symptoms such as slower movements, using the lower part of the tank and increased breathing frequency, while one fish showed balance disorder. At the end of observation period 48-72 five fish were agitated, breathing frequency was heightened, one fish was strongly affected, one showed surfacing and one had died. At the end of the whole testing period 4 more fish died, one of the surviving fish was agitated, the others stayed at the bottom of the vessels and breathing frequency was heightened.
- 50 mg/L: at 3 h after beginning of the test the fish showed agitated swimming movements, surfacing and balance disorder could be observed, breathing frequency was heightened. At the end of observation period 0 - 24 h six fish had died, the surviving fish was strongly affected and showed balance disorder, breathing frequency was heightened and the fish stayed mainly at the bottom of the test vessel. At the end of observation period 24-48 h the remaining fish had died.
- 100 mg/L: after 3 hours symptoms were similar to the 50 mg/L test loading rate at the same time; during the first observation period (0 - 24 h) all fish died.


- Other biological observations (feeding after exposure):

- Control: all fish fed immediately
- 6.25 mg/L: all fish fed immediately
- 12.5 mg/L: the six surviving fish fed with retardation (ca. 2 min) compared to the control
- 25 mg/L: one of the two surviving fish fed, the other fish did not feed


- Observations during recovery period: During this period fish derived from the control and test loading rate 6.25 mg/L showed normal behaviour, fish derived from test loading rates 12.5 and 25 mg/L did not fully recover and still fed with retardation compared to the control after 3 days in clean water.


- Mortality of control: None.

- Effect concentrations exceeding solubility of substance in test medium: the loading rate at which the water-accommodated fraction was prepared exceeded the solubility of the test substance.

Reported statistics and error estimates:

The 96 h LL50 value (LL = Lethal Loading) was determined graphically on probability paper for the test with 5 loading rates. NOELR (No Observed Effect Loading Rate) was derived from observations at test loading rate of 6.25 mg/L.

Sublethal observations / clinical signs:

Table 3. Cumulative mortality (%) at various loading rates (mg/L WAFs) during the 96 h exposure period.

Time (h)	Control	6.25 mg/L	12.5 mg/L	25 mg/L	50 mg/L	100 mg/L
24	0	0	0	0	86	100
48	0	0	0	0	100	100
72	0	0	0	14	100	100
96	0	0	14	71	100	100

Cumulative lethality after 96 h was 14 % at a loading rate of 12.5 mg/L, 71% at a loading rate of 25 mg/L and 100 % at loading rates of 50 mg/L and 100 mg/L.

A definite test was also performed as a limit test. Lethality in the definite test with 5 test loading rates (100 % at loading rate of 100 mg/L) was comparable to lethality in the definite limit test (100 % at loading rate of 100 mg/L).

Table 4. TOC concentrations in the control, and at loading rates 12.5 mg/L, 50 mg/L and 100 mg/L at the beginning and the end of periods 0 -24 h and 72 -96 h.  

Period (h)	Control	12.5 mg/L	50 mg/L	100 mg/L
0-24 h start	2.3	5.7	9.1	12
0-24 h end	3.8	5.6	9.7	12
72-96 h start	3.1	4.6	-	-
72-96 h end	2.8	3.7	-	-

TOC values increased with loading rate and confirmed the low water solubility of the test article. TOC values at test loading rate 12.5 mg/L, 50 mg/L and 100 g/L remained stable (within +/- 6%) during observation period 0 -24 h, while the control value increased (+ 40 %) during the same period. TOC concentration at test loading rate 12.5 mg/L decreased by ca. 20 % during observation period 72 -96 h.

At the 0 -24 h observation period TOC value of the 100 mg/L loading rate in the definite test with 5 loading rates was 10 -30 % higher compared to the TOC values at the same loading rate in the definite test as a limit test.

Table 4. Results of GCMS analysis of water accommodated fractions.

Nominal loading rate (mg/L)	Period (h)		9-Octadenoic acid	9,12 -Octadenoic acid	Abietic Acid	3-Carene	Terpinole	Total
Control (0)	0-24 h start		< 0.0040	< 0.016	0.011	< 0.000015	< 0.00027	< 0.031
Control (0)	0-24 h end		< 0.0040	< 0.016	0.0041	< 0.000015	< 0.00027	< 0.024
12.5	0-24 h start		< 0.0040	< 0.016	0.17	< 0.000015	0.0013	0.19
12.5	0-24 h end		< 0.0040	< 0.016	0.15	< 0.000015	0.0012	0.17
50	0-24 h start		< 0.0040	< 0.016	0.32	< 0.000015	0.0051	0.35
50	0-24 h end		< 0.0040	< 0.016	0.33	< 0.000015	0.0047	0.36
100	0-24 h start		< 0.0040	< 0.016	0.50	< 0.000015	0.0096	0.63
100	0-24 h end		< 0.0040	< 0.016	0.52	< 0.000015	0.0098	0.55
Control (0)	72-96 h start		0.0062	< 0.016	0.0095	< 0.000015	< 0.00027	< 0.032
Control (0)	72-96 h end		< 0.0040	< 0.016	0.0028	< 0.000015	< 0.00027	< 0.023
12.5	72-96 h start		0.0096	0.030	0.23	< 0.000015	0.0017	0.27
12.5	72-96 h end		0.0048	< 0.016	0.21	< 0.000015	0.0016	0.23
50	72-96 h start		-	-	-	-	-	-
50	72-96 h end		-	-	-	-	-	-
100	72-96 start		-	-	-	-	-	-
100	72-96 h end		-	-	-	-	-	-
Detection Limit (mg/L)			0.0040	0.016	0.00062	0.000015	0.00027

GCMS analysis of the control and various loading rates reveals that the total concentration of components remained relatively stable over the period 72 -96 h at all loading rates and control, the total amount decreased at the end of each 24 h observation period probably due to evaporation of the test substance.

GCMS concentrations were definitely lower compared to TOC concentrations. This is caused by the fact that only a fingerprint of the sample was detected by GCMS analysis whilst TOC analysis detected Total Organic Carbon as a sum parameter.

Validity criteria fulfilled:

yes

Conclusions:

A 96 h LL50 of 20 mg/L and a NOELR of 6.25 mg/L have been determined for the effects of the test substance on mortality of Danio rerio.

Description of key information

There are no data for short-term toxicity for the registered substance to fish. However reliable results are available for the closely-related substance, Crude Tall Oil (EC number 931-433-1). A 96-h LL₅₀ of 20 mg/L and a NOELR of 6.25 mg/L have been determined for the effects of this substance on mortality of Danio rerio (tested as Water Accommodated Fractions, WAFs).

Read-across justification:

The use of ecotoxicity data for the read-across substance Crude Tall Oil (CTO; list number 931-433-1) is justified on the following basis.

1. HYPOTHESIS FOR THE ANALOGUE APPROACH

A read across from Crude Tall Oil to TOS can be justified on the following basis.

Crude Tall Oil (CTO), list number 931-433-1, is the term applied to the processed mixture of naturally occurring compounds extracted from tree species like pine, spruce, birch and aspen. CTO is formed by the acidification of TOS, therefore, CTO and TOS are similar substances. They differ only in the water content and that CTO does not contain sodium salts of the fatty/rosin acids.

The main constituents of TOS are sodium salts of saturated and unsaturated C14 -20 fatty acids (5-45% w/w), rosin acid sodium salts (10-40% w/w), water (25-45% w/w), sterols (1-10% w/w), and non-volatile lignin/cellulose fibre/oligomeric acids (3-20% w/w). In addition, the following minor constituent blocks are present: terpenes, sesquiterpenes, abietenes and labdanes, C30 branched polyakenes, 3,5 -dimethoxystilbene, rosin alcohol and aldehyde isomers and saturated/unsaturated alcohols and terpene alcohols (each <3% w/w).

The same constituents are present in CTO, with only the following differences:

-The fatty acids and rosin acids in TOS are present as their sodium salts whereas for CTO they are present as acids. In aqueous media, the sodium salts are dissociated into sodium and the parent acid. The sodium, being a ubiquitous and essential element in nature, is not expected to contribute to the toxicity properties of the substance.

-TOS contains 25 -45% water compared to trace amounts in CTO; other constituent groups are therefore present at lower levels in TOS but in similar proportions to CTO.

The hypothesis for the read-across approach is that in the environment and under standard laboratory test conditions organisms would be exposed to essentially the same constituents (and in similar proportions) whether they originate from TOS or CTO and so the ecotoxicity of TOS would be substantially the same as that for CTO.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)

Both CTO and TOS are UVCB substances that contain fatty acids, rosin acids, neutrals such alcohols, sterols, aldehydes etc. For UVCB substances, impurities are not considered relevant. The fatty and rosin acids present in TOS are in the form of the sodium salt. In the environment, the sodium salts of fatty and rosin acids will dissociate and speciate in a similar way to each other. The sodium salts in TOS will ionise into sodium ions and a mixture of the acid anion/parent acid (depending on the pH) and CTO will dissociate into the same mixture of the acid anion/parent acid (again depending on the pH). Sodium is a ubiquitous, essential element in nature and will not contribute to the toxicity of the acid and as such it is considered relevant to read-across the available data for CTO to TOS.

Therefore, it can be concluded that in the environment and under standard laboratory test conditions, the organisms would be exposed to essentially the same constituents and (and in similar proportions) whether they originate from TOS or CTO and so the read-across of ecotoxicity data from CTO to TOS is justified.

Additional information

Calculation of PNECs for the aquatic compartment will be based on data for the blocks of constituents rather than on data for the whole substance.