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Short-term toxicity to aquatic invertebrates

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Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
other: experimental study on hydrolysis product Boric acid
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
other: OECD Guideline 203
Qualifier:
according to guideline
Guideline:
other: ASTM E729-96
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
The concentration of boron in test solutions was measured in samples collected at 0 and 96 hours of the definitive test. Control and boric acid-fortified samples were also prepared for analysis at each sample period. Ten-milliliter samples were collected from the control and test substance treatment and diluted with reagent water to provide final sample concentrations within the analytical standard concentrations range. QC samples were prepared by fortifying laboratory saltwater with boric acid at concentrations of 10.1 and 232 mg B/L. All samples were directly analyzed using an ICP-MS system.
Details on test solutions:
A 1.14 mg boric acid/mL primary stock solution (equivalent to 200 mg B/L) was prepared at test initiation by diluting 20.5200 g of boric acid into an 18,000 mL volume of dilution water.
Test organisms (species):
other: Litopenaeus vannemei
Details on test organisms:
Marine shrimp (Litopenaeus vannemei) were received from Shrimp Improvement Systems (Plantation Key, Florida) on June 4, 2010 and maintained at ABC Laboratories, Inc. The initial supplier provided the species identification. The shrimp were at growth stage PL-8 (post larval) upon receipt. The shrimp were maintained in artificial saltwater at a temperature of approximately 23 to 28°C. The shrimp were gradually acclimated from an initial approximate 35‰ saltwater salinity to a target saltwater salinity of 20 ± 2‰ over a period of 11 days. The animals were fed shrimp food and brine shrimp nauplii (Artemia sp. <48 hours old) during the holding period. Periodic lots of feed were screened for contaminants and records of feed suppliers and the results of the contaminant analyses are on file at ABC Laboratories, Inc. A record of shrimp observations and during the culture period was maintained. No mortalities were recorded during the holding period.
Shrimp used for the definitive test were collected on July 9, 2010. The shrimp were at PL-44 (post larval) stage. Shrimp were fed ad libitum newly hatched brine shrimp (Artemia sp.; approximately 24-48 hours old) nauplii with at least two feedings daily. All treatments and controls received approximately the same ration (i.e., volume) of food. The control shrimp were measured at termination and ranged from 14 to 19 mm in length (mean and standard deviation = 17 ± 1.5 mm) and from 0.0357 to 0.1013 g in blotted wet weight (mean and standard deviation = 0.0767 ± 0.0194g).

During the test shrimp were offered brine shrimp ad libitum daily to reduce predation.
Test type:
static
Water media type:
saltwater
Limit test:
no
Total exposure duration:
96 h
Test temperature:
22.8 to 25.1°C
pH:
7.4 to 8.0
Dissolved oxygen:
5.7 to 7.4 mg/L (76 to 99% saturation)
Salinity:
19.0 to 19.9‰
Nominal and measured concentrations:
Nominal Concentrations: 0 (control), 13, 25, 50, 100, and 200 mg B/L
Mean Measured Concentrations as added Boron: 0 (dilution-water control), 14.0, 27.2, 55.8, 103, and 215 mg B/L
Mean Measured Concentrations as Total Boron: 2.23 (dilution-water control), 16.2, 29.5, 58.0, 105, and 217 mg B/L
Details on test conditions:
The dilution water was a laboratory saltwater prepared by adding a commercial sea salt mix (Crystal Sea Marinemix; Marine Enterprises International, Inc. Baltimore, Maryland) to laboratory freshwater at a target salinity 20 ± 2‰. The laboratory freshwater consists of well water blended with well water that was demineralized by reverse osmosis to yield water with a total hardness ranging from 130 to 160 mg CaCO3/L. The dilution water was heated and aerated prior to its delivery into the diluter system. As the dilution water entered the diluter system, it passed through a particulate filter and an ultraviolet sterilizer.

The test chambers utilized during the range-finding test were 1-L glass beakers containing approximately 800 mL of control or test solution. In the definitive test, the test chambers were 10-L glass jars containing approximately 8 L of control or test solution. Animal size during the definitive test required larger test chambers to avoid biological loading problems. The test chambers in both tests were covered with plastic Petri dishes. The test chambers were maintained at 21 to 25°C and within a range of 2°C in a temperature-controlled water bath. Fluorescent lighting was maintained on a 16-hour light and 8-hour dark photoperiod with 30-minute simulated dawn and dusk periods. The measured light intensity at the termination of the definitive test ranged from 658 to 710 lux as measured with a LI-COR Model LI-189 light meter equipped with a photometric sensor.

Ten shrimp were added to test chambers for the dilution water control and each test substance treatment. Shrimp were impartially added to a set of labeled containers with each container representing one treatment. One individual was added to each labeled container starting with the control then proceeding from the low to high test substance treatments. The individuals within each container were then released from each container into the corresponding test chamber. Observations were made daily (± 1 hour of test initiation) for mortality and sublethal effects. The initial loading rate was 0.0959 grams of shrimp per liter of test solution which meets the loading rate criterion.
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
130 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
element
Remarks:
boron
Basis for effect:
mortality
Remarks on result:
other: added boron (110 - 154) (Salinity: 19.0 to 19.9‰)
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
103 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
element
Remarks:
boron
Basis for effect:
mortality
Remarks on result:
other: added boron (Salinity: 19.0 to 19.9‰)
Details on results:
After 96 hours of exposure, mortality was 0, 0, 0, 0, 20, and 100% in the 0 (dilution-water control), 13, 25, 50, 100, and 200 mg B/L nominal treatments. Based on mean measured total boron concentrations, the 24- and 48-hour LC50 was estimated to be >217 mg B/L, the highest concentration tested, as determined by the trimmed Spearman-Karber method. The 72-hour LC10 and LC50 values, based on mean measured total boron concentrations, were estimated to be 106 mg B/L (95% confidence limits of 50.5 and 136 mg B/L) and 163 mg B/L (95% confidence limits of 124 and 221 mg B/L), respectively, as determined by the probit method. The 96-hour LC50, based on mean measured total boron concentrations, was estimated to be 132 mg B/L with 95% confidence limits of 112 and 156 mg B/L as determined by the untrimmed Spearman-Karber method. The slope of the 96-hour concentration-response could not be calculated. There were no sublethal effects noted in the control or test substance treatments during the definitive test. The 96-hour NOEC and LOEC values were 105 and 217 mg B/L, respectively, based on total measured boron concentrations.
Reported statistics and error estimates:
All statistical analyses were performed with SAS software. Estimates of LC50 values and their 95% confidence limits were calculated using the probit method and Spearman-Karber method (trimmed or untrimmed). When the P value for Goodness of Fit was >0.05 and there was no other evidence of questionable convergence, the probit method was selected for reporting. When this criterion was not achieved, the Spearman-Karber method was selected for reporting. The NOEC was determined by using a Fisher’s exact test. A Hochberg adjustment was used to control the experiment wise error rate for the Fisher’s test at the same alpha level.
Validity criteria fulfilled:
yes
Conclusions:
Based on mean measured total boron concentrations, the 24- and 48-hour LC50 was estimated to be >217 mg B/L, the highest concentration tested. The 72-hour LC10 and LC50 values, based on mean measured total boron concentrations, were estimated to be 106 mg B/L (95% confidence limits of 50.5 and 136mgB/L) and 163 mg B/L (95% confidence limits of 124 and 221 mg B/L), respectively. The 96-hour LC50, based on mean measured total boron concentrations, was estimated to be 132 mg B/L with 95% confidence limits of 112 and 156 mg B/L. The slope of the 96-hour concentration-response could not be calculated. There were no sublethal effects noted in the control or test substance treatments during the definitive test. The 96-hour NOEC and LOEC values were 105 and 217 mg B/L, respectively, based on nominal concentrations. The 96-hour NOEC and LOEC values were 103 and 215 mg B/L, respectively, based on added concentrations.
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
other: experimental study on hydrolysis product Boric acid
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Guideline study
Reason / purpose for cross-reference:
reference to other study
Qualifier:
according to guideline
Guideline:
other: ASTM E729-95 Standard Guide for Conducting Acute Toxicity Tests on Test Materials with Fishes, Macroinvertebrates, and Amphibians
Qualifier:
equivalent or similar to guideline
Guideline:
EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
GLP compliance:
no
Remarks:
No data is available on the GLP compliance
Analytical monitoring:
yes
Details on sampling:
Each treatment was sampled at beginning and end of test
Vehicle:
no
Test organisms (species):
Ceriodaphnia dubia
Details on test organisms:
TEST ORGANISM
- Common name: Ceriodaphnia, water flea
- Source: In-house culture
- Age at study initiation (mean and range, SD): <24 h
- Feeding during test: none
- Food type:
- Acclimation: cultured in dilution water
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
90 mg/L
Test temperature:
24.0 °C
pH:
mean = 7.4. control = 7.8, highest boron concentration = 6.8
Dissolved oxygen:
> 6.0 mg/L
Nominal and measured concentrations:
Nominal: Control, 31.3, 62.5, 125, 250, 500 mg-B/L
Measured: 0.64, 31, 62, 125, 245, 490 mg-B/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 50 ml beaker
- Type: open
- Material, size, headspace, fill volume: glass, 40 ml
- Aeration: none
- Renewal rate of test solution: none
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: MHRW (moderately hard reconstituted water, per US EPA 2002)
- Total organic carbon:
- Particulate matter:
- Metals:
- Pesticides:
- Chlorine: 0 nominal
- Ca/mg ratio: 5.40
- Conductivity:
- Salinity:
- Culture medium different from test medium: Cultured in test medium
- Intervals of water quality measurement: boron, pH, conductivity, alkalinity, hardness at beginning and end of test, temperature and D.O. daily

OTHER TEST CONDITIONS
- Adjustment of pH: none
- Photoperiod: 16 L: 8 D

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Mortality (daily)
Duration:
48 h
Dose descriptor:
LC50
Effect conc.:
102 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
element
Basis for effect:
mortality
Remarks on result:
other: (95% CI 82-125 mg-B/L)
Reported statistics and error estimates:
LC50 calculated using trimmed Spearman-Karber method

Data

Concentration (mg B/L) # dead (out of 20) Percent
0.64 (control) 2 10.0%
31 2 10.0%
62 9 45.0%
125 9 45.0%
246 20 100.0%
490 20 100.0%
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
justification for analogue approach: see IUCLID section 13 "Read-across justification"
Reason / purpose for cross-reference:
read-across source
Key result
Duration:
48 h
Dose descriptor:
LC50
Effect conc.:
1 328 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% CL 1123 - 1925 mg/L
Key result
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
415 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
Daphnids of the negative control, 138, 226, and 415 mg/L n-butyl alcohol treatment groups appeared healthy and normal throughout the test. Although no mortality was observed in the 675 mg/L treatment group, seven of the 20 daphnids appeared lethargic at 48-hours. After 48-hours of exposure, mortality in the 1123 and 1925 mg/L treatment groups was 20 and 100%, respectively.anal
Reported statistics and error estimates:
The 24-hour and 48-hour EC50 values and the 95% confidence intervals were calculated when possible by probit analysis, the moving average method or binomial probability with non-linear interpolation using the computer software of C.E. Stephan. The no mortality/immobility concentration and NOEC were determined by visual inspection of the mortality, immobility and clinical observation data.

Measurement of Test Concentrations

Nominal concentrations selected for use in this study were 156, 259,432, 720,1200 and 2000 mg n-Butyl Alcohol/L. Samples collected at the beginning of the test had measured concentrations that ranged from 94 to 102% of nominal. Samples collected at test termination had recoveries that ranged from 76 to 95% of nominal. When the values obtained for all analyses were averaged, the mean measured test concentrations were 138, 226,415, 675, 1123 and 1925 mg n-Butyl Alcohol/L.

Observations and Measurements

Daily observations of mortality, immobility and other clinical signs observed during the test are attached (see 'Attached background material', Table).

Daphnids in the negative control, 138, 226 and 415 mg n-Butyl Alcohol/L treatment groups appeared healthy and normal throughout the test. Although no mortality was observed in the 675 mg n-Butyl Alcohol/L treatment group, seven of the 20 daphnids appeared lethargic at 48-hours. After 48-hours of exposure, mortality in the 1123 and 1925 mg n-Butyl Alcohol/L treatment groups was 20 and 100%, respectively.

EC50 values and 95% confidence limits at 24 and 48 hours were estimated or calculated from the mortality/immobility data.

The 48-hour concentration-response curve is attached (see 'Attached background material', Figure).

Validity criteria fulfilled:
yes
Conclusions:
- 48-hour EC50 = 1328 mg/L (95% CL: 1123 - 1925 mg/L)
- 48-hour NOEC = 415 mg/L
Executive summary:

The objective of this study was to evaluate the acute toxicity of n-Butyl Alcohol to Daphnia magna, during a 48-hour exposure period under static test conditions. The study was conducted according to OECD Guideline for Testing of Chemicals, 202 and TSCA Title 40 of the Code of Federal Regulations, Part 797, Section 1300.

Daphnids were exposed to a geometric series of six test concentrations and a negative (dilution water) control. Two replicate test chambers were maintained in each treatment and control group, with 10 daphnids in each test chamber for a total of 20 daphnids per test concentration. Nominal test concentrations were selected in consultation with the sponsor, and were based upon the results of an exploratory range finding toxicity test. Nominal test concentrations selected were 156. 259. 432, 720, 1200 and 2000 mg/L. Mean measured test concentrations were determined from samples of lest water collected from each treatment and control group at the beginning of the test and at test termination.

Daphnids were indiscriminately assigned to exposure chambers at test initiation. Observations of mortality/immobility and other clinical signs of toxicity were made at approximately 21, 24 and 48 hours after test initiation. Cumulative percent mortality and immobility observed in the treatment groups were used to estimate or calculate EC50 values at 24 and 48 hours. The no mortality/immobility concentration and the no-observed-effect-concentration (NOEC) were determined by visual interpretation of the mortality, immobility and clinical observation data.

The 48-hour EC50 value for Daphnia magna exposed to n-Butyl Alcohol was 1328 mg/L. The 95% confidence limits were 1123 and 1925 mg/L. The no mortality/immobility concentration was 675 mg/L. Based on the lethargy observed in the 675 mg/L treatment group, the NOEC was 415 mg/L.

Description of key information

Tributyl borate is rapidly hydrolyzed to Boric acid and n-Butanol in the presence of water (18.3 sec at 21°C).

Reliable data of the hydrolysis products n-Butanol and Boric acid are used to address the endpoint, which is wholly adequate to draw conclusions on the toxicity of Tributyl borate to aquatic invertebrates:

1) n-Butanol - Toxicity to freshwater invertebrate Daphnia magna

- (48h) EC50: 1328 mg/L (95% CL: 1123 - 1925 mg/L) [EPA OTS 797.1300 and OECD 202; GLP];

- (48h) NOEC: 415 mg/L [EPA OTS 797.1300 and OECD 202; GLP].

2) Boric acid

a) Toxicity to freshwater invertebrate Ceriodaphnia dubia (based on total boron measured values):

- (48h) LC50: 102 mg B/L (95% CL: 82 - 125 mg B/L) [based on: mortality; ASTM E729-95 and EPA OPPTS 850.1010 and OECD 202; GLP]; LC50 considering the boron content (4.6968%) of Tributyl borate: 2172 mg/L.

b) Toxicity to marine invertebrate Litopenaeus vannemei (based on total boron measured values):

- (96h) LC50: 130 mg B/L (95% CL: 110 - 154 mg B/L) [based on: mortality; ASTM E729-96 and OECD 203; GLP]; LC50 considering the boron content (4.6968%) of Tributyl borate: 2768 mg/L;

- (96h) NOEC: 103 mg B/L [ASTM E729-96 and OECD 203; GLP]; NOEC considering the boron content (4.6968%) of Tributyl borate: 2193 mg/L.

Key value for chemical safety assessment

Additional information