Tricyclo[3.3.1.13,7]decane-1-acetic acid, .alpha.-[[(1,1-dimethylethoxy)carbonyl]amino]-3-hydroxy-, (.alpha.S)-

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 February 2005 to 9 January 2006.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was conducted in 2005 according to and OECD Method # 202 and in accordance with GLP. The study material is well characterized.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2006

Report date:

2006

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

other: American Society for Testing and Materials (ASTM) Standards, Water and Environmental Technology, Section 11, Standard Guide for Conducting Acute Toxicity Tests with Fishes, Macroinvertebrates and Amphibians, E 729

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

(2S)-2-{[(tert-butoxy)carbonyl]amino}-2-(3-hydroxyadamantan-1-yl)acetic acid

EC Number:

700-361-0

Cas Number:

361442-00-4

Molecular formula:

C17H27NO5

IUPAC Name:

(2S)-2-{[(tert-butoxy)carbonyl]amino}-2-(3-hydroxyadamantan-1-yl)acetic acid

Constituent 2

Reference substance name:

1-Hydroxyadamantanyl-3-(S)-Boc-glycine

IUPAC Name:

1-Hydroxyadamantanyl-3-(S)-Boc-glycine

Details on test material:

white powder; stored at room temperature; received at testing laboratory on 06-10-2004.

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
No data in report

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Samples (20 mL) were taken from the freshly prepared (new) test media at 0 hours and from pooled replicate vessels at 48 hours (old), for the chemical analysis of BMS 528233-01.

Test solutions

Vehicle:

no

Details on test solutions:

The test medium at the 100 mg/L nominal BMS 528233-01 concentration was prepared by direct addition of 0.1001 g of BMS 528233-01 to a 1-litre volumetric flask. The flask was made up to volume with ASTM water, treated with ultrasound for ca. 30 minutes, stirred for 30 minutes in the dark, treated with ultrasound for a further 10 minutes and finally, stirred for 30 minutes in the dark. The remaining test media at 10, 1 and 0.1 mg/L were prepared by serial dilution of the 100 mg/L test medium. This was achieved by adding 100 mL aliquots of the 100, 10 and 1.0 mg/L test media as each were prepared to 1-litre volumetric flasks and making up to volume with ASTM water.

The dilution water used for the culturing and testing of Daphnia magna is a standard hard water prepared with analytical grade reagents and reverse-osmosis (RO) water and met American Society for Testing and Materials, standard hard water (ASTM).

The dilution water has the following constituents and final concentration:

Compound Formula Final concentration in dilution water (mg/L)
Sodium hydrogen carbonate NaHCO3 192 mg/L
Calcium sulphate dihydrate CaSO4. 2H2O 120 mg/L
Magnesium sulphate heptahydrate MgSO4. 7H2O 246 mg/L
Potassium chloride KCl 8 mg/L

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: daphnia magna
- Source: in-house laboratory cultures
- Age at study initiation (mean and range, SD): less than 24 hours old
- Feeding during test - Food type: concentrated suspension of Chlorella vulgaris
- Frequency: daily

The D. magna were cultured in 600 mL glass beakers containing 500 mL of standard hard water. Each culture vessel and its contents were referred to as a 'culture'. New cultures were started with juvenile D. magna (less than 24-hours old), at a density of approximately 20 animals per litre.The water in each culture was renewed weekly. Juveniles were removed when present in cultures at least three times per week using a sieve. Cultures were maintained up to a maximum of 28 days. Juveniles for use in acute toxicity tests were collected from the second brood onwards. Approximately 24 hours before a test was set up, juveniles present in the cultures were removed and discarded. Over the next 24 hours, juveniles for use in the test were then removed from the culture and transferred to fresh culture medium, using a wide bored pipette. The juveniles were then left for at least 1 hour before selecting actively swimming individuals for use

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Post exposure observation period:

None reported

Test conditions

Hardness:

approximate theoretical total hardness of 165-174 mg/L as CaC03

Test temperature:

Ambient temperature (ºC) 20.6 -21.4
Culture temperature (ºC) 21.2 -21.7

pH:

pH 7.6 - 7.7

Dissolved oxygen:

Dissolved oxygen (% air saturation) 93 - 99

Salinity:

Alkalinity as CaCO3 (mg/L) 79

Nominal and measured concentrations:

The mean measured concentrations (mean of the measured concentrations at the start and end of the range-finding/limit test) of BMS 528233-01 in samples of test media at nominal concentrations of 1.0, 10 and 100 mg/L, were 0.965, 10.3 and 102 mg/L, corresponding to 96.5, 103 and 102% of the nominal BMS 528233-01 exposure concentrations respectively. At 0.1 mg/L, no BMS 528233-01 was detected above the limit of detection (0.2 mg/L).
The measured concentrations of BMS 528233-01 were consistently maintained between 80 and 120 % of the nominal concentrations at the 1, 10 and 100 mg/L concentrations. However, results at the 0.1 mg/L treatment were below the limit of detection of the analytical procedure. Given the high recoveries (96.5 to 103%) at the 1, 10 and 100 mg/L nominal concentrations and the nature of the test media preparation, it is considered that, if better sensitivity of the analytical was achieved, the % recoveries at the 0.1 mg/L nominal concentration would have been of a similar magnitude.
As no immobility occurred in the test and the mean recoveries above 0.1 mg/L nominal BMS528233-01 were approximately 100% of the nominal concentrations, the toxicity values have been expressed in terms of the nominal BMS 528233-01 concentrations.

Details on test conditions:

The dilution water used was American Society for Testing and Materials, standard hard water (ASTM). Four replicate test vessels were prepared for the control treatment (using ASTM only) and the 100 mg/L concentration. Two replicate vessels were prepared at the remaining test concentrations.. Five juvenile D. magna were added (released under the surface of the test media) to each test vessel using a wide bore glass pipette. During the test, the behaviour of the test substance in the test system was observed and recorded. This included whether the test substance appeared to dissolve, form a film on the surface, whether it was more/less dense or viscous than the ASTM and whether it adhered to the wall of the test vessel.

Reference substance (positive control):

not specified

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CL

Effect concentrations 2

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CL

Effect concentrations 3

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CL

Details on results:

All control and BMS 528233-01 test media appeared as clear, colourless solutions for the duration of the test.The number of immobilised. No immobility was observed in any of the control or BMS 528233-01 test media for the duration of the test.

Results with reference substance (positive control):

No data

Reported statistics and error estimates:

Results are expressed in terms of the concentrations of test substance in ASTM, water which immobilised 50% of the D. magna after 24 and 48 hours exposure. No immobile D. magna were recorded during the range-finding limit test. Statistical analysis of the data was therefore not required.
The toxicity values are quoted as the 24 and 48-hour EC50 values and, where appropriate, include the 95% confidence limits. The highest test concentration causing no immobilisation and the lowest concentration causing 100% immobilisation have been reported.

Numerical data may have been rounded for presentation. Therefore, manual recalculation may give slightly different values to those shown.

Chemical Analysis: The linearity of detector response was determined over the concentration range 1.0 to 100 µg/mL. The coefficient of determination (forced through zero) was better than 0.99 for BMS 528233-01.
-The accuracy at 1.0, 10 and 100 µg/m was 100-102 % and repeatability (error in precision) at 1.0, 10 and 100 µg/m was 2.73%, 1.61% and 0.311% respectively.

Any other information on results incl. tables

Table 3
Immobilisation of Daphnia magna after 24 hours Exposure to
BMS 528233-01 during the Range-finding/limit Test

Nominal BMS 528233-01concentration (mg/L)	Number of Daphnia magna exposed	Mobile Daphnia Bottom Surface	Immobile Daphnia Bottom Surface
Control	20	20 -	- -
0.1	10	9 1**	- -
1.0	10	10 -	- -
10	10	10 -	- -
100	20	20 -	- -

Values not appropriate^$ 

**Surface held D. magna resubmerged by single drop of test media applied to animal by Pasteur pipette

Table 4
Immobilisation of Daphnia magna after 48 hours Exposure to
BMS 528233-01 during the Range-finding/Limit Test

NominalBMS 528233-01concentration (mg/L)	Number ofDaphnia magnaexposed	MobileDaphniaBottom Surface	ImmobileDaphniaBottom Surface
Control	20	20 -	--
0.1	10	10 -	--
1.0	10	10 -	--
10	10	10 -	--
100	20	20 -	--

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

The validity criteria of a maximum of 10% immobility in the control treatment and achieving dissolved oxygen concentrations of ≥ 3 mg/L in the control and test vessels at the end of the test were both satisfied.

Conclusions:

There were no immobile D. magna recorded during the test, therefore the 24 and 48-hour EC50 toxicity values are considered to be >100 mg/L, the highest concentration tested. The corresponding no observed effect concentration (NOEC) was considered to be ≥ 100 mg/L. The lowest concentrations causing 100% immobility was considered to be >100 mg/L.

Executive summary:

The acute toxicity of BMS 528233-01 to Daphnia magna was determined at approximately 20°C in a 48-hour static, range-finding/limit test. The test was conducted at nominal BMS 528233-01 concentrations of 0.1, 1.0, 10 and 100 mg/L.

The test medium at the 100 mg/L nominal BMS 528233-01concentration was prepared by direct addition of 0.1001 g of BMS 528233-01 to a 1-litre volumetric flask. The flask was made up to volume with ASTM water, treated with a combination of ultrasound and stirring pre-treatment until the test substance had dissolved. The remaining test media were prepared by serial dilution of the 100 mg/L test medium. Control vessels were prepared by the addition of ASTM only. Four replicate vessels were prepared at the control and 100 mg/L treatment, with two replicates at the remaining concentrations. Five juvenile D. magna(< 24 hours old) were added to each test vessel. The mean measured concentrations of BMS 528233-01 in unfiltered samples of test media at the nominal concentrations of 1.0, 10 and 100 mg/L were 0.965, 10.3 and 102 mg/L respectively, corresponding to 96.5, 103 and 102% of the nominal BMS 528233-01concentrations. No BMS 528233-01was detected in the control or the 0.1 mg/L treatments.

As the mean measured concentrations were >80 and <120% of the nominal concentrations (with the exception of the 0.1 mg/L treatment), the toxicity values are expressed in terms of nominal concentrations.

 

There were no immobile D.magna ecorded during the test, therefore the 24 and 48-hour EC50 toxicity values are considered to be >100 mg/L, the highest concentration tested. The corresponding no observed effect concentration (NOEC) was considered to be≥100 mg/L. The lowest concentrations causing 100% immobility was considered to be >100 mg/L.

The validity criteria of a maximum of 10% immobility in the control treatment and achieving dissolved oxygen concentrations of≥3 mg/L in the control and test vessels at the end of the test were both satisfied.