3-acetoxymethylen-7-amino-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid

Short-term toxicity to fish

Administrative data

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

December 16, 1996 to December 20, 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study (OECD 203) with acceptable restrictions.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1997

Report date:

1997

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

3-acetoxymethylen-7-amino-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid

EC Number:

213-485-0

EC Name:

3-acetoxymethylen-7-amino-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid

Cas Number:

957-68-6

Molecular formula:

C10H12N2O5S

IUPAC Name:

(6R,7R)-3-[(acetyloxy)methyl]-7-amino-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material (as cited in study report): G-7-ACS Mono-Na-Salz
- Physical state: yellowish powder
- Analytical purity: 97.2 %
- Batch No.: Op.1/96
- Expiration date of the batch: stable under conditions of storage
- Storage condition of test material: in a refrigerator in the dark

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Aquaeous solutions Volume: 20mL
Nominal test substance concentrations : Control; 10000 mg/L

Test solutions

Vehicle:

yes

Details on test solutions:

The test substance was weighed into a beaker. Water for dilution was added and the mixture homogenized using an Ultra-Turrax and poured into the chamber under stirring with a giass rod.

After adding the substance the pH fell to 3.7. it was then adjusted to the pH of the test water with 10 molar NaOH.

In order to determine the substance concentration, water samples were taken from the middle of the test chamber by pipetting approx. 20 mL into a screw-neck glass bottle at study start, and after 48 and 96 hours.

Test organisms

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

Species: Zebra fish Brachydanio rerio (HAMILTON-BUCHANAN)
Origin: Corporate Toxicology, Hoechst AG - Hoechst Marion Roussei Preclinical Development - Drug Safety
Date of hatching: July 12, 1996
Delivery date: August 09, 1996
The fish were kept for 12 days before the start of the study in water for dilution (see section 6.3) under the following conditions:
Temperature: 22 ± 1 °C
Oxygen content: > 80 % of the saturation value
Duration of light period: 12 hours daily
Population density: < 1 g fish / L water
Feeding: twice daily ad libitum
Food: Tetra Min, Tetra Werke, Melie (Germany)

Study design

Test type:

static

Water media type:

freshwater

Total exposure duration:

96 h

Test conditions

Hardness:

2.1 - 2.3 mmol Ca2+ + Mg2+ / L

Test temperature:

21.2-22.1°C

pH:

6.0 -8.2

Dissolved oxygen:

3.5-9.9mg/L

Nominal and measured concentrations:

nominal 10000 mg/L and 6093 mg/L after 0h, 4967 mg/L after 48h and 4481 mg/L after 96h

Details on test conditions:

Test conditions:
The study was conducted in a static system. The test chambers, which were calibrated to 4 liters, were made of glass (length 30 cm, width 22 cm, height 24 cm) and stood in a water bath made of Hostalit Z® with a Plexiglas viewing panei. The temperature of the water bath was regulated by a thermostat to 22 + 1 °C. The chambers were illumi¬nated from above from 06.00 a.m to 06,00 p.m. The light intensity directly over the chambers was approximately 700 lux.

The test chamber of the 10000 mg/L group was aerated from 48 hours after study start on, because the maintenance of an oxygen content of at least 60% of the saturation value was not ensured. Aeration was carried out using a glass capillary with a bubble-frequency of 1 - 3 per second.

Preparation of the test concentrations:
The test substance was weighed into a beaker. Water for dilution was added and the mixture homogenized using an Ultra-Turrax and poured into the chamber under stirring with a giass rod.

After adding the substance the pH fell to 3.7. it was then adjusted to the pH of the test water with 10 molar NaOH.

In order to determine the substance concentration, water samples were taken from the middle of the test chamber by pipetting approx. 20 mL into a screw-neck glass bottle at study start, and after 48 and 96 hours.

Study groups:
The study consisted of 1 part. There were 7 fish in each group. The test concentrati¬ons, which were selected on the basis of range-finding tests, were as follows :

Part of study Start Test chamber No. Concentration (mg/L)
1. Dec-16-96 1 /3 0/10000

Test procedure:
After the test concentrations had been prepared and water parameters recorded, 7 fish were assigned to each test and control chamber. The fish received no feed for the en¬tire study period, inspection of the fish took place after 3, 6, 24, 48, 72 and 96 hours and involved recording the lethality and visible changes in appearance and behavior. Dead fish were removed from the chambers. Fish were considered dead when there was a lack of opercular movement and no response to slight mechanical stimulus. The water parameters were measured and recorded before study start and after 0, 24, 48, 72 and 96 hours.

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 10 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

Water parameters:
The following ranges for the concentration groups and the control groups were ob¬tained for the whole study period:

Parameter Concentration groups Controls
pH (1) 6.0 - 8.2* 7.7-8.2
Oxygen content (mg/L) 3.5 - 9.9** 7.2-10.1
Temperature (°C) 21.2-22.1 21.2-22.1

* = After adding the substance the pH fell to 3.7. It was then adjusted to the pH of the
test water with 10 molar NaOH. ** = from 48 hours after study start on the test chamber was aerated

Test concentrations:
The tested concentration was present at the start of the test as yellow-brownish solution, which changed to a light yellow-brownish turbidity 72 to 96 hours after study start. The substance concentration was determined in the 10000 mg/L group.
The values determined were in a range of 44.8 - 60.9 % of the theoretical value. The concentration values given in this report are based on the nominal concentration. The mean of the analytically determined concentrations is given in brackets.

Observations:
The fish showed no changes in appearance and behavior, swimming behavior and respiration rate in the tested concentration group during the entire exposure time.

Lethality:
In this 96-hour acute toxicity study of G-7-ACS Mono-Na-Salz in zebra fish (Brachydanio rerio) no lethality occurred in the 10000 mg/L-group (nominal concentration) and in the control group. A mean concentration value of 5180.3 mg/L was determined analytically.

Applicant's summary and conclusion

Conclusions:

Under the conditions of this test the LC50 of G-7-ACS Mono-Na-Salz after 96 hours was >10000 mg/L(nominal concentration) and >5180.3 mg/L based on the mean of the analytically determined concentrations.

Executive summary:

G-7-ACSMono-Na-Salzwas tested in zebra fish (Brachydanio rerio) over 96 hours in a static system.

The concentrations tested were 10000 mg/L and a negative control (0 mg/L).

After adding the substance the pH fell to a values of 3.7. It was then adjusted to the pH of the test water with 10 molar NaOH.

The tested concentration was present at the start of the test as yellow-brownish solution, which changed to a light yellow-brownish turbidity 72 to 96 hours after study start.

The substance concentration was determined in both test batches. The values determined were in in a range of 44.8 - 60.9 % of the theoretical value with a tendency of decrease. The concentration values given in this report are based on the nominal concentration. The mean of the analytically determined concentrations is given in brackets.

In this 96 -hour acute toxicity study ofG-7-ACSMono-Na-Salzthe following lethality values were determined:

 

	after 24	after 48	after 72	after 96
	hours	hours	hours	hours
LC0 (mg/L)	10000	10000	10000	10000
	(5180.3*)	(5180.3*)	(5180.3*)	(5180.3*)
LC50 (mg/L)	>10000	>10000	>10000	>10000
	(5180.3*)	(5180.3*)	(5180.3*)	(5180.3*)
LC100 (mg/L)	not determined	not determined	not determined	not determined

 

* = mean measured concentrations

In the control group no lethality was observed. The 10000 mg/L group showed no effect in comparison with the negative control.