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Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2006
Report date:
2006

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)
Deviations:
no
GLP compliance:
yes
Test type:
fixed dose procedure
Limit test:
yes

Test material

Constituent 1
Reference substance name:
Active enzyme protein of Aspergillopepsin I (EC no. 232-796-2, CAS no. 9025-49-4, EC name: Proteinase, Aspergillus acid, Enzyme Class no. 3.4.23.18)
Molecular formula:
Not applicable, see remarks
IUPAC Name:
Active enzyme protein of Aspergillopepsin I (EC no. 232-796-2, CAS no. 9025-49-4, EC name: Proteinase, Aspergillus acid, Enzyme Class no. 3.4.23.18)
Constituent 2
Reference substance name:
Protein as a constituent of enzyme deriving from the fermentation or extraction process
Molecular formula:
Not available
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
Details on test material:
- Lot/batch No.: AFP501K1

Test animals

Species:
rat
Strain:
other: HanTac:WH
Remarks:
SPF (Specific Pathogen Free)
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Taconic M&B A/S, Dk-8680 Ry, Denmark
- Females (if applicable) nulliparous and non-pregnant: not reported
- Age at study initiation: not reported
- Weight at study initiation: 140-151 g
- Fasting period before study: Overnight; after treatment, feed was withheld for a further 3 hours
- Housing: Transparent polycarbonate cages (macrolone type III, floor area 810 cm2). The cages were cleaned and the bedding changed at least twice a week.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21±3°C
- Humidity (%): 55±15%
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12 hrs dark / 12 hrs light

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: distilled water
Details on oral exposure:
VEHICLE: Distilled water

MAXIMUM DOSE VOLUME APPLIED: 16 mL/kg body weight

DOSAGE PREPARATION (if unusual): Prior to use, the test substance was thawed in the refrigerator overnight. The test substance dose formulations were prepared by diluting the test substance with vehicle. The dose formulations were prepared on the respective days of treatment and stored at room temperature until used.
Doses:
2000 mg/kg
No. of animals per sex per dose:
5
Control animals:
yes
Remarks:
For the animal welfare of the sighting study animal, a female companion animal was housed with the sighting study animal. The companion animal was observed for clinical behaviour and subjected to gross necropsy (results not included in report).
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were weighed prior to treatment on Day 1, and on Days 2, 3, 8, and 15; each rat was observed 15 minutes and 1, 3, and 6 hours after administration and thereafter daily for a period of 14 consecutive days.
- Necropsy of survivors performed: yes
Statistics:
None

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 other: mg total protein/kg bw
Based on:
test mat.
Mortality:
No mortality was observed during the study.
Clinical signs:
Sighting Study: At the observations 1 and 3 hours after treatment, the sighting animal appeared less active. Furthermore, the animal had piloerection 3 hours after treatment. However, the animal appeared normal at all other observations.

Main Study: At the observations 1, 3, and 6 hours after treatment, all animals had piloerection. At all other observations all 4 animals appeared normal.
Body weight:
Sighting Study: The sighting animal had a normal body weight gain from treatment until termination of the study.

Main Study: All 4 animals had a normal body weight gain from treatment until termination of the study.
Gross pathology:
No macroscopic abnormality of tissues or organs was noted at necropsy for the sighting study or the main study.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
LD50: >2000 mg total protein/kg bw
Executive summary:

The acute oral toxicity of the test substance was studied in rats after administration of a single oral dose followed by an observation period of 14 days. A preliminary sighting study, using one female animal, was included in order to estimate the dose effect for toxicity and to provide information on dose selection for the main study. The study was conducted in accordance with OECD Guideline 420. The study was initiated with a sighting study in one female rat, in which 2000 mg total protein/kg body weight was examined in order to provide information on dose selection for the main study. On the basis of the results of the sighting study, the main study was performed in 4 additional female rats given a dose of 2000 mg total protein/kg body weight.

All animals survived the treatment. Slight signs of toxicity (piloerection and less activity) were observed among the animals on the day of treatment. However, no signs of toxicity were observed on any of the following days. All four animals had a normal body weight gain from treatment until termination of the study. The post-mortem inspection revealed no abnormalities. Under the experimental conditions, it was found that the minimal lethal dose of the test substance was above 2000 mg total protein/kg body weight.