SRC TR

- 09

- 057

ORPHAN CHEMICAL DATA ADEQUACY REVIEW AND ROBUST SUMMARY ADEQU ACY

REQUIREMENTS



### Fatty acids, tall

- oil, reaction products with diethylenetriamine, acetates

Prepared by:

Emily Cohen, Christopher Janson and Joan D. Garey

Environ mental Science Center

Syracuse Research Corporation

7502 Round Pond Road

North Syracuse, NY 13212

Contract No. EP

- W

- 04

- 019

Task FA411.4.409.004

Submitted to:

OPPT/RAD/HPVCB (7403M)

U.S. Environmental Protection Agency

1200 Pennsylvania Avenue

Washingt on, D.C. 20460

Kathleen C. Kennedy, Project Officer

David Brooks, Work Assignment Manager

January 16, 2009

### Orphan Chemical Data Adequacy Review and Robust Summary Adequacy Requirements

Chemical Name:

Fatty acids, tall

- oil, reaction products with diethylenetriamine, acetates, CAS No. 68153

- 60

- 6

January 2009

Submitted to:

High Production Volume Chemicals Branch

Risk Assessment Division

Office of Pollution Prevention and Toxics

Environmental Protection Agency

WAM: David Brooks

1200 Pennsylvania Avenue, NW

Washington, DC 20460

- 0001

Prepared By:

SRC

Environmental Science Center

2451 Crystal Drive, Suite 804

Arlington , VA 22202ACUTE TOXICITY TO AQUATIC VERTEBRATES

Summary of endpoint: Adequate

Title: Acute toxicity to fish

The study is: Adequate Not Adequate

TEST SUBSTANCE

Identity: (CAS No. 68153

- 60

- 6)

Remarks: Project 02

- 088

- 2

METHOD

Method/guideline followed: P ARCOM. “Protocols on Methods for Testing of Chemicals Used in

Offshore Oil Industry. Part B Protocol for a Fish Acute Toxicity Test”. 1995

Test type: S tatic acute with a 48 hour renewal

GLP (Y/N): Y es

Year study performed: 2003

Species/Strain: Sheepsh ead minnow, Cyprinodon variegatu s

Analytical monitoring: N one, nominal concentrations

Exposure time: 96 hours

Number of animals/sex/dose: 10/dose

Statistical Methods: Trimmed Spea r man

- Karber method.

Conc entration: 0 (control), 0.5

, 1.0

, 2.0

, 4.0

, 8.0 , 16.0 mg/L

Remarks: Deionized water based test substance stocks introduced into glass, rectangular 21

- L aquaria

containing 10 liter of synthetic seawater. Synthetic seawater was prepared using deionized lab oratory

water and Hawaiian Marinemix artificial salts. An identical vessel containing only seawater was

designated the control. Ten sheepshead minnows were randomly placed into each vessel for a total of ten

fish per concentration. Fish mean total length a nd weight were 1.1 cm and 0.017 g, respectively. Fishwere not fed for 24 hours prior to exposure. Six nominal test concentrations (0.5, 1.0, 2.0, 4.0, 8.0 and

16.0 mg/L) were evaluated. Temperature was maintained at nominal 20 +/

- 2 C and photoperiod at 1 6

hours light: 8 hours dark.

Mortality counts and behavioral observations were made every 24 hours during the 96 hour study.

Dissolved oxygen, pH, salinity and temperature were recorded initially and every 24 hours thereafter for

all surviving concentrati ons. Test vessels were not aerated. Test solutions were renewed at 48 hours.

A 2.3 mg/L 3,5

- dichlorophenol reference study was conducted concurrently with this study.

RESULTS

Remarks: The 96 hour LC50 was determined to be 5.3 (4.6

- 6.1) mg/L using the Tr immed Spea r man

Karber method. The no observed effect concentration (NOEC) at 96 hours was 4.0 mg/L. The minimum

concentration causing 100% mortality was 8.0 mg/L. No mortalities were noted in the control. Normal

behavior was noted at all times and in all c oncentrations up to 4.0 mg/L. The one mortality noted in the

1.0 mg/L at 96 hours was not deemed to be toxicant related. Sheepshead minnow exposure to 2.3 mg/L

3,5

- dichlorophenol demonstrated 50% survival at 96 hours; the acceptable range is 20 to 80 perce nt.

The test substance was completely soluble in synthetic seawater. The chemical and physical parameters

monitored during the study were all within normal range: pH 7.8

– 8.0, dissolved oxygen 6.9

– 7.6 ppm,

temperature 18.4

– 20.3 C and salinity 32

– 34 ppt

Cumulative 96 hour mortality

Control = 0

0.5 mg/L = 0

1.0 mg/L = 1

2.0 mg/L = 0

4.0 mg/L = 0

8.0 mg/L = 10

16.0 mg/L = 10

CONCLUSION

The 96 hour, static acute renewal toxicity of the test substance to the sheepshead minnow was 5.3 mg/L.

The corres p onding NOEC value is 4.0 mg/L

RELIABILITY CODE

1

- The test is considered valid since the 2.3 mg/l 3,5

- dichlorophenol reference toxicity study was within

acceptable parameters. Temperature, dissolved oxygen and pH were also within test guidelines.REF ERENCES

Static acute renewal toxicity to the Sheepshead Minnow ( Cyprinodon variegates ) (2006)

Disk 3: 8(d)86070000012 (OECD) (page 3)ACUTE TOXICITY TO AQUATIC INVERTEBRATES

Summary of endpoint: Inadequate

Title: Acute toxicity to invertebrates

The study is: Adequate Not Adequate

This study is not adequate for the acute toxicity to invertebrates endpoint as freshwater species are the

preferred test subjects.

TEST SUBSTANCE

Identity: CAS No. 68153

- 60

- 6

Remarks : Project O

- 70 02F

METHOD

Method/guideline followed: P ARCOM. Ring T est Protocol “Acute Toxicity to the Marine Copepod

Acartia tonsa” & ISO/CD 14669: “Determination of Acute Lethal Toxicity to Marine Copepods”.

Test type: S tatic acute

GLP (Y/N): Y es

Ana lytical monitoring: N one, nominal concentrations in seawater

Year study performed: 2002

Species/strain: Acartia tonsa

Exposure time: 48 hours

Concentrations: 0 (control), 0.1, 0.5, 2.5, 10.0 mg/L

Remarks: Acartia tonsa eggs used for this study were o btained from the University of Copenhagen,

Laboratory of Marine Biology, Helsingar, Denmark on April 12, 2002 and stored in the refrigerator. Test

animals were cultured in small batches and the age of the animals used in this study was

28 days. The test su bstance was dissolved in filtered and aerated seawater. The concentration of this stock

corresponded with the highest concent ration of product tested (10 mg/ l). The stock was mixed in a 1 liter

Erlenmeyer flask on a magnetic stirrer (~100 rpm) for 2

– 4 ho urs. The flask was wrapped in aluminum

foil to avoid exposure to light. Five Acartia tonsa were randomly placed into each replicate (4) for a total

of twenty copepods per concentration. Four nominal test concentrations (0.1, 0.5, 2.5, and 10.0 mg/l) anda control were evaluated. Temperature was maintained at a nominal 20 +/

- 2.4 C with a photoperiod of

16 hours light: 8 hours dark.

Mortality counts were made every 24 hours during the 48 hour study. Water chemistry was also

performed at this time. Test ve ssels were not aerated. A 1.0 mg/l 3,5

- dichlorophenol reference study was

conducted concurrently with this study.

RESULTS

The 48

- hour LC5 0 was determined to be 2.8 mg/l; confidence limits are not available, using the Logistic

Regression Model (JMP Progr am, version 3, 199 8). The 48 hour NOEC was 0.5 mg/ l while the LC10 and

LC90 were respectively 2.3 and 3.4 mg/l. The minimum concentration causing 100% mortality was 10.0

mg/l. No mortalities were noted in the control. Acartia tonsa exposure to 1.0 mg/l 3,5

- dichlorophenol

demonstrated 60% survival at 48 hours; the acceptable range is 20 to 80 percent.

The test substance was completely soluble in seawater. The chemical and physical parameters monitored

during the study were all within normal range: 48 hour c ontrol dissolved oxygen was greater than 4 mg/ l

(i.e., 7.3 mg/ l) and the temperature range during the study was 19.4

- 20.4 C within the test guideline of

20 ± 2.4 o C.

Cumulative 48 Hour Mortality

Control = 0

0.1 mg/ l = 0

0.5 mg/ l = 0

2.5 mg/ l = 5

10.0 m g/l = 20

CONCLUSIONS

The 48

- hour static acute LC5 0 of the test substance to Acartia tonsa is 2.8 mg/l. The corresponding

NOEC value is 0.50 mg/l. These test results are based upon nominal concentrations of the test substance

in seawater.

RELIABILITY COD E

3

– not valid

REFERENCES

AnalyCen Ecotox AS, “Toxicity Test Results with Acartia tonsa for the Test substance (2002)

Disk 3: 8(d)86070000012 (OECD) (page 10)

TOXICITY TO AQUATIC PLANTSSummary of endpoint: Adequate

Title: Toxicity to marin e algae

The study is: Adequate Not Adequate

TEST SUBSTANCE

Identity: CAS No. 68153

- 60

- 6

Remarks: Project 02

- 088

- 1

METHOD

Method/guideline followed: ISO Draft International Standard 1994. ISO/DIS 10253.2, ISO/TC 14 7/SC

5 , 1994

- 05

- 17: Water Quality

– Marine algal growth inhibition test with Skeletonema costatum and

Phae o dactylum tricornutum. ISO Central Secretariat.

Test Type: Static acute growth inhibition

GLP (Y/N): Yes

Analytical monitoring: Electronic particl e counting (Coulter Counter Multisizer 3 with computer

interface/related software)

Year study performed: 2003

Species/Strain: Skeletonema costatum , UTex #LB2308

Element basis: 10,000 cells/mL inoculums, initial cell density 1,501, 390 cells/mL, 6.660 mL inoculum /L

of nutrient seawater media

Exposure period: 72 hours

Statistical methods: L inear regression analysis

Concentration: 0.04, 0.07, 0.13, 0.24, 0.75, 1.3, 2.4, 4.3 mg/L

Remarks: The inoculum used in this study was extracted from four day

- old s tock culture. The

Skeletonema costatum (UTEX #LB 2308) culture used in this study was obtained from stock maintained

in the natural seawater algal assay medium obtained from the original culture from the Department of

Biology’s Culture Collection of Algae at the University of Texas at Austin, Austin, TX 78712. This strainwas used by Petrolite during a PARCOM Ring Test and its growth and response was found to be

comparable to other strains used. ESG incubated the stock cultures under the same environmental

conditions as those used in the definitive test.

Method of cultivation: Stock subcultures were transferred into 250 mL Erlenmeyer flasks containing fresh

medium 1

- 2 times per week to provide 3± 1 day old cultures for assay inoculations.

Controls: Six con trol vessels were prepared w ith culture medium and inoculum but no test substance. A

single concentration series of blanks (media and test substance without inoculum) including a blank

control (medium only) were also employed.

An Eberbach Shaker power uni t (#6140), located in a Forma walk

- in environmental chamber (set at 20

C), was employed and continuously illuminated by 6000

– 10,000 lux of light provided by overhead cool

white fluorescent lights. Flasks, 250 mL in size, were randomly placed on the power unit and rotated

continuously at 100 +/

- 5 oscillations per minute. Random placement of the Erlenmeyers was determined

using an Excel software program. The flasks were repositioned randomly each day to minimize spatial

differences in the chamber. The temp erature was measured and recorded at zero, 24, 48, and 72 hours in

each of three strategically located temperature control flasks

Test temper ature range: Protocol stipulates 20 ±2 degrees Cels ius (19.8

- 21.2 degrees C. measured)

Dilution water source: The natural seawater algal assay medium was prepared with seawater obtained

from Marine Biological Laboratory (MBL), Aquatic Resource Division, Woods Hole, MA. MBL filtered

the water to 0.8 μm prior to shipping. Upon receipt, the seawater was placed in an envi ronmental chamber

set at 20 C or lower. When needed for media preparation, the water was removed from storage and

filtered to 0.2 μm. The salinity was determined and recorded. The salinity of the water used in this test

was found to be 31 ppt.

pH algal me dium: Nutrients obtained from Aquaculture Supply, Dade City, FL and a commercially

prepared trace sodium silicate nutrient concentrate were used to prepare the algal medium f/2 plus silica

formulation recommended by Guillard (1975). Following the addition of nutrients, the pH of the medium

was checked and adjusted if necessary. The pH of the me dia used in this test was 8.0. T he medium was

stored at 20 C or lower until needed for testing.

Exposure vessel type: Test vessels were 250 mL Erlenmeyer flasks fitt ed with 23

- 45 mm air permeable

Jaece foam plugs. Four replicates per test concentration, no aeration.

Stock solutions: Stock concentration of test material (50 mg/mL) prepared in deionized water and added

in the appropriate amounts to 50 mL algal media a chieve each nominal test concentration.

Water chemistry pH: 0 hour: Control (8.01), 0.04 (8.04), 0.07 (8.04), 0.13 (8. 06), 0.24 (8.08), 0.42 (8.07),

0.75 (8.08), 1.3 (8.09), 2.4 (8.10), 4.03 (8.11)72 hour: Control (8.45), 0.04 (8.80), 0.07 (8.56), 0.13 (8.72), 0.24 (8.43), 0.42 (8.77), 0.75 (8.05), 1.3

(8.02), 2.4 (8.00), 4.03 (8.00)

Light levels and quality: Continuous illumination by 6,000

– 10,000 lux of light provided by overhead

cool

- white fluorescent lighting. Readings of 7051, 8954, and 8008 lux were measured at three spatially

random positions on shaker table.

RESULTS

Remarks:

Element value:

Biomass (mg/L)

24 hr EC50 = 0.49

48 hr EC50 = 0.56

72 hr EC50 = 0.58

72 hr EC90 = 0.75

72 hr NOEC = 0.04

Growth rate (mg/L)

24 hr EC50 = 0.49

48 hr EC5 0 = 0.69

72 hr EC50 = 0.74

72 hr EC90 = 0.87

72 hr EC50 = 0.42

Control response: satisfactory

Statistical results: satisfactory, R

- squared 95+(24, 48, 72 hr data sets)

Percent biomass inhibition (%) per concentration (mg/L) observations:

24 hour: 0.04(0) , 0.07(44.9), 0.13(0), 0.24(0), 0.42(6.9), 0.75(87.4), 1.3(114), 2.4(118), 4.03(176)

48 hour: 0.04(0), 0.07(4.4), 0.13(0), 0.24(0), 0.42(0), 0.75(94.0), 1.3(102), 2.4(105), 4.03(110)

72 hour: 0.04(0), 0.07(12.2), 0.13(0), 0.24(0), 0.42(1.7), 0.75(90. 6), 1. 3(101), 2.4(101), 4.03(101 )

Percent growth rate inhibition (%) per concentration observations (mg/L) :

24 hour: 0.04(0), 0.07(44.9), 0.13(0), 0.24(0), 0.42(6.9), 0.75(87), 1.3(114), 2.4(118), 4.03(176)

48 hour: 0.04(0), 0.07(22.8), 0.13(0), 0.24(0.2), 0.42 (0), 0.75(94.9), 1.3(101), 2.4(103), 4.03(100)

72 hour: 0.04(0), 0.07(0.81), 0.13(0), 0.24(0), 0.42(0), 0.75(56.8), 1.3(104), 2.4(114), 4.03(95.6)

CONCLUSIONS

The test substance is biologically active toward Skeletonema; 72 hr EC50 < 1.0 mg/LRELIABIL ITY CODE

1

- The test was valid since the control cell density increased by a factor of more than 16 in 72 hr. This

increase corresponds to a growth rate of 0.04 h

- 1. The pH did not vary by more than 1.0 unit during the

test.

REFERENCES

“Acute Toxicity of _ to the Saltwater Diatom, Skeletonema costatum (72 hr Exposure)”. (2006)

Disk 3: 8(d)86070000012(OECD) (page 5)Summary of Endpoints

Endpoint Study Information

Provided Acceptable Data Gap

Aquatic Effects

Acute Toxicity to Fish Yes Yes No

Acut e Toxicity to Daphnia Yes No Yes

Toxicity to Aquatic Plants Yes Yes No

Health Effects

Acute Toxicity No No Yes

Repeated

- Dose Toxicity No No Yes

Genetic Mutation No No Yes

Chromosomal Aberrations No No Yes

Toxicity to Reproduction No No Yes

Developm ental Toxicity No No Yes