(7R)-7-(4-carboxybutanamido)cephalosporanate amidohydrolase (enzyme substance)

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 20 March 2001 to 26 April 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was performed according to OECD guideline and GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Deviations:

yes

Remarks:

Temporary deviations from the maximum level for temperature (with a maximum of 1°C) occurred. Based on laboratory historical data these deviations were considered not to have affected the study integrity.

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

deoxyribose-phosphate aldolase EC 4.1.2.4.

Cas Number:

9026-97-5

Molecular formula:

Not applicable, please see remarks

IUPAC Name:

deoxyribose-phosphate aldolase EC 4.1.2.4.

Constituent 2

Reference substance name:

Protein as a constituent of enzyme deriving from the fermentation or extraction process

Molecular formula:

Not available. See remarks.

IUPAC Name:

Protein as a constituent of enzyme deriving from the fermentation or extraction process

Constituent 3

Reference substance name:

Inorganic salts as a constituent of enzyme deriving from the fermentation or extraction process

Molecular formula:

Not available. See remarks.

IUPAC Name:

Inorganic salts as a constituent of enzyme deriving from the fermentation or extraction process

Constituent 4

Reference substance name:

Carbohydrates constituent of enzyme deriving from the fermentation or extraction process.

Molecular formula:

Not available. See remarks.

IUPAC Name:

Carbohydrates constituent of enzyme deriving from the fermentation or extraction process.

Constituent 5

Reference substance name:

Lipids as a constituent of enzyme deriving from the fermentation or extraction process

Molecular formula:

Not available. See remarks.

IUPAC Name:

Lipids as a constituent of enzyme deriving from the fermentation or extraction process

Test material form:

liquid

Remarks:

Tests based on dissolved material in the concentrate

Details on test material:

- Name of test material (as cited in study report): Aldolase (IUB 4.1.2.4)
- Substance type: enzyme
- Physical state: liquid
- Stability under test conditions: data not available
- Storage condition of test material: In freezer in the dark

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany.
- Age at study initiation: young adult animals (approx. 10 weeks old) were selected.
- Weight at study initiation: mean body weight on day 1 was 335 g in males and 229 g in females. Body weight variation did not exceed +/- 20% of the sex mean.
- Fasting period before study: food was withheld overnight (for a maximum of 20 hours) prior to dosing until approximately 3-4 hours after administration of the test substance.
- Housing: group housing of 3 animals per sex per cage in labelled Macrolon cages (type IV; height 15 cm.) containing purified sawdust as bedding material (SAWI, Jelu Werk, Rosenberg, Germany).
- Diet: free access to standard pelleted laboratory animal diet (from Altromin (code VRF I), Lage, Germany)
- Water: free access to tap-water.
- Acclimation period: at least 5 days before start of treatment under laboratory conditions.


ENVIRONMENTAL CONDITIONS
- Temperature: 21+/-3°C
- Humidity: 30-70%
- Air changes: approximately 15 air changes per hour
- Photoperiod: 12 hours artificial fluorescent light and 12 hours dark per day


IN-LIFE DATES: date of treatment 6 March 2001 (no other information available)

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

MAXIMUM DOSE VOLUME APPLIED: 21 ml/kg body weight which corresponds with a dose level of 2000 mg protein/kg (slight exceeding the maximum dose volume as stated in the guideline, but not expected to affect the outcome of the study).

- Rationale for the selection of the starting dose: data not available

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

3 males and 3 females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing:
> Mortality/Viability: twice daily.
> Body weights: days 1 (pre-administration), 8 and 15.
> Clinical signs: at periodic intervals on the day of dosing (day 1) and once daily thereafter, until day 15.
- Necropsy of survivors performed: yes

Statistics:

No statistical analysis was performed (The method used is not intended to allow the calculation of a precise LD50 value).

Results and discussion

Effect levels

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Remarks on result:

other: no mortality observed

Mortality:

No mortality occurred.

Clinical signs:

other: Lethargy, hunched posture, piloerection, ptosis and/or chromodacryorrhoea were noted among the animals between days 1 and 4.

Gross pathology:

No abnormalities were found at. macroscopic post mortem examination of the animals.

Any other information on results incl. tables

Table 1: Results

Dose (mg/kg bw)	Mortality (# dead/total)			Time range of deaths (hours)	Number with evident toxicity(#/total)
	Male	Female	Combined		Male	Female	Combined
2000	0 / 3	0 / 3	0 / 3	N.A.	3 / 3	3 / 3	6 / 6

Applicant's summary and conclusion

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The oral LD50 value of ALDOLASE (IUB 4.1.2.4) in Wistar rats was established to exceed 2000 mg protein /kg body weight.

Executive summary:

The study was carried out based on the guidelines described in: EC Commission Directive 96/54/EC, Part B.l tris "Acute Toxicity-Oral, Acute Toxic Class Method" and OECD No.423, "Acute Oral Toxicity - Acute Toxic Class Method".

ALDOLASE (IUB 4.1.2.4) was administered by oral gavage to three Wistar rats of each sex at 2000 mglkg body weight. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was performed after terminal sacrifice (day 15).

No mortality occurred. Lethargy, hunched posture, piloerection, ptosis and/or chromodacryorrhoea were noted among the animals between days 1 and 4. The mean body weight gain shown by the animals over the study period was considered to be normal. No abnormalities were found at macroscopic post mortem examination of the animals.

The oral LDS0 value of ALDOLASE (IUB 4.1.2.4) in Wistar rats was established to exceed 2000 mg protein /kg body weight.

Based on these results and according to the EC criteria for classification and labelling requirements for dangerous substances and preparations (Guidelines in Commission Directive 93/21/EEC), ALDOLASE (IUB 4.1.2.4) does not have to be classified and has no obligatory labelling requirement for oral toxicity.