Chymotrypsin

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25-06-2015 to 27-08-2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

Adopted 17 December 2001.

Deviations:

no

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Active enzyme protein of Chymotrypsin (EC no. 232-671-2, CAS no.9004-07-3, EC name Chymotrypsin, Enzyme class no 3.4.21.1 )

Molecular formula:

n.a.

IUPAC Name:

Active enzyme protein of Chymotrypsin (EC no. 232-671-2, CAS no.9004-07-3, EC name Chymotrypsin, Enzyme class no 3.4.21.1 )

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.: PPA26797
- Expiration date of the lot/batch: 12 June 2019
- Storage condition of test material: Frozen in the dark

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: BIONEEDS INDIA PRIVATE LIMITED (in-house bred)
- Females nulliparous and non-pregnant: Yes
- Age at study initiation: 10 weeks
- Weight at study initiation: 160.07 g to 170.45 g
- Fasting period before study: No
- Housing: Three animals were housed in a standard Polysulfone cage (size: L 430 x B 285 x H 200 mm) with stainless steel mesh top grill having facilities for holding pelleted food and drinking water in water bottle fitted with stainless steel sipper tube. Clean sterilized paddy husk was provided as bedding material. Sterilized paper shreds were provided as nesting material for enrichment.
- Diet: Ad libitum. Nutrilab rodent feed (Manufactured by Provimi Animal Nutrition India Pvt Ltd)
- Water: Ad libitum. Deep bore-well water passed through activated charcoal filter and exposed to ultraviolet rays in Aquaguard water filter purifier was provided in plastic water bottles with stainless steel sipper tubes.
- Acclimation period: Healthy and young adult animals used for step I, step II and step III were acclimatized for seven, nine and eleven days, respectively.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.6 to 22.4°C
- Humidity (%): 50-61%
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 02 July 2015 To: 20 July 2015

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

Undiluted test material for the top dose.

Doses:

The acute toxicity test was conducted with starting dose 1060 mg TOS/kg body weight, followed by 1591 mg TOS/kg body weight and the highest dose level of 2111 mg TOS/kg body weight. Three animals were used per step (total nine animals).
Fixed dose volume of 5.3 mL/kg body weight was used with increased frequency of administration to gain the final dose volume.

No. of animals per sex per dose:

3 (female only)

Control animals:

no

Details on study design:

The total quantity of test item to be administered per day was administered in divided doses of two (Step I), three (Step II) and four (Step III) times with approximately 4 hour interval between each administration. Test item administration was sequential and allowing at least 24 hours before dosing the next set of animals. The time interval between dosing at each level was determined by the onset, duration and severity of toxic signs.
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: All the animals were observed for clinical signs of toxicity and mortality at 30 to 40 min, 1 hr (±10 min), 2 hrs (±10 min), 3 hrs (±10 min) and 4 hrs (±10 min) after each administration on Day 1 and thereafter, once daily for clinical signs of toxicity and twice daily for mortality during the 14 days observation period.
- Necropsy of survivors performed: At the end of observation period, all the animals were sacrificed by exsanguination in deep Isoflurane anaesthesia. A complete gross pathological examination was carried out for terminally sacrificed animals.

Statistics:

No

Results and discussion

Effect levels

Key result

Sex:

female

Dose descriptor:

LD0

Effect level:

>= 2 111 mg/kg bw

Based on:

other: Total Organic Solids (TOS)

Mortality:

No mortality during 14 days of observation period.

Clinical signs:

other: The animals did not reveal any clinical signs of toxicity and mortality during 14 days of observation period.

Gross pathology:

There were no gross pathological changes observed in the animals dosed at 1060, 1591 and 2111 mg TOS/kg body weight for step I, step II and step III, respectively during necropsy.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The animals at all the tested doses (1060, 1591 and 2111 mg TOS/kg body weight) did not reveal any clinical signs of toxicity throughout the observation period and no mortality occurred. The body weight and percent change in body weight of animals at all the steps were increased with respect to Day 1. A complete gross pathological examination was carried out for the animals and there were no gross pathological changes observed in any of the animals.
Based on the results of the experiment and under experimental conditions employed, it was concluded that the test item serine endopeptidase, batch PPA26797 could not be classified based on the GHS criteria.

Executive summary:

The test item serine endopeptidase, batch PPA26797 was evaluated for Acute Oral Toxicity in Sprague Dawley Rats. The acute toxicity test was conducted with starting dose 1060 mg TOS/kg body weight, followed by 1591 mg TOS/kg body weight and the highest dose level of 2111 mg TOS/kg body weight. Three animals were used per step (total nine animals). Fixed dose volume of 5.3 mL/kg body weight was used with increased frequency of administration to gain the final dose volume.

All the animals were observed for clinical signs of toxicity at 30 to 40 minutes, 1 hr (±10 min), 2 hr (±10 min), 3 hr (±10 min) and 4 hr (±10 min) after each administration on Day 1 and once daily thereafter for clinical signs and twice daily for mortality.

The animals at all the tested doses (1060, 1591 and 2111 mg TOS/kg body weight) did not reveal any clinical signs of toxicity throughout the observation period and no mortality occurred.

Individual animal body weight was recorded at receipt, Day 1 before test item administration, Day 7 and Day 14 during the observation period for step I, step II and step Ill animals. The body weight and percent change in body weight of animals at all the steps were increased with respect to Day 1.

All the animals (step I, step II and step Ill) were observed for 14 days, sacrificed by exsanguination in deep Isoflurane anaesthesia on Day 15 and subjected to necropsy. A complete gross pathological examination was carried out for the animals and there were no gross pathological changes observed in any of the animals.

Based on the results of the experiment and under experimental conditions employed, it was concluded that the test item serine endopeptidase, batch PP A26797 could not be classified based on the GHS criteria.