Mannanase, endo-1,4-β-

Acute Toxicity: inhalation

Administrative data

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From August 16, 1999 to November 30, 1999

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1999

Report date:

1999

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: EEC, OECD and US EPA (Health Effects Test Guidelines, OPPTS 870, 1300, Acute Inhalation Toxicity, 5 August 1998) and JMAFF test guidelines for acute inhalation studies

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

standard acute method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Mannanase, endo-1,4-β-

EC Number:

253-446-5

EC Name:

Mannanase, endo-1,4-β-

Cas Number:

37288-54-3

Molecular formula:

Not available. See Remarks.

IUPAC Name:

mannan endo-1,4-beta-mannosidase IUBMB 3.2.1.78

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:

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 4

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 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:

other: Liquid

Details on test material:

Lot/batch No.: PPE 6432
Expiration date of the lot/batch: July 2009

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Source: Charles River UK Ltd, Manston Road, Margate, Kent UK.
- Housing: Five animals per cage
- Housing: In holding cages (size 35 cm x 53 cm x 25 cm height) in animal room with control of temperature and humidity
- Diet: SDS rat and mouse diet (RM1), ad libitum, except during the 4 hr exposure
- Water: Tap water ad libitum, except during the 4 hr exposure
- Acclimation period: 8 days
- Temperature (°C): 20-26°C
- Humidity: 44-66 %

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

air

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: ADG Developments Ltd., Hitchin, Hertfordshire, England
- Exposure chamber volume: 30 L
- Method of holding animals in test chamber: Snout only
- Source and rate of air: A supply of clean dried air was connected to the aerosol generator and the supply pressure was adjusted to give a flow rate of 15 litres/minute measured at the outlet of the generator. An in-line flow meter was used to monitor airflow throughout the exposure.
- Method of conditioning air: Filtered, oil-free compressed air for the production of the test atmosphere was supplied by compressors.
- System of generating particulates/aerosols: The test substance was supplied to the generator via the feed line from a syringe driven at a constant rate by a syringe pump (Precidor® type 5003).
- Method of particle size determination: Two air samples were taken during the exposure at a sampling rate of 2 L/minute using a Marple cascade impactor (Model 296, Graseby Andersen Inc., Atlanta, USA) to determine particle size distribution. The volume of air sampled was measured using a wet-type gas meter (Model DM3B, G. H. Seal Ltd., London, England).
- Temperature, humidity, pressure in air chamber: Mean temp: 19.9°C (control group) and 19.6°C (test group), relative humidity: 49% (control group) and 95% (test group).

TEST ATMOSPHERE
- Brief description of analytical method used: Seven air samples were taken during exposure. Chamber air was drawn at a measured rate of 2 L/minute, through a pre- weighed glass fibre filter (Whatman GF/A) mounted in an open face filter holder. Filters were dried for 15 minutes in an oven at 30-35°C, and then re-weighed for gravimetric analysis of the test aerosol.

TEST ATMOSPHERE
- Particle size distribution: 87% respirable (< 7 µm in aerodynamic diameter)

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

>= 4 h

Concentrations:

0.45 mg enzyme concentrate dry matter/L

No. of animals per sex per dose:

5

Control animals:

yes

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Observations for clinical signs of effect: At the end of the chamber equilibration period, at 0.25, 0.5 and 1.0 hours then at hourly intervals during the exposure, immediately following completion of the exposure and then at 1.0 and 2.0 hours post-exposure. Subsequent, daily observations. Body weight: Just before exposure and weekly during the observation period.
- Necropsy of survivors performed: yes
- Other examinations performed: Organ weights: The lungs, liver and kidneys of each animal were weighed.

Statistics:

Not performed.

Results and discussion

Effect levels

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 0.45 mg/L air (analytical)

Based on:

other: Enzyme concentrate dry matter

Exp. duration:

4 h

Mortality:

No mortality.

Clinical signs:

other: No clinical signs. Fur/skin soiled with excreta was observed in all test and control rats immediately after exposure, but was not present 1 hour later. Brown staining on the head was seen in 2 male and female control rats and 2 male test rats up to 1 hour

Body weight:

The mean bodyweight gain for male test rats was 69 g compared with 47 g for control rats during the 14-day observation period; however, all body weights and body weight gains were within the expected range throughout the study.

Gross pathology:

No abnormalities.

Other findings:

The mean liver weight of test males was higher (+14%) than the control values. This was considered to be associated with the slightly heavier bodyweights of the test rats.

Applicant's summary and conclusion

Interpretation of results:

study cannot be used for classification

Conclusions:

There were no unscheduled deaths or evidence of a toxic response following exposure of rats for 4 hours to a droplet aerosol generated from Mannanase, PPE 6432 at a chamber concentration of 0.45 mg enzyme concentrate dry matter/L.

Executive summary:

The present study was performed in rats, in accordance with GLP and in compliance with EEC, OECD and US EPA (Health Effects Test Guidelines, OPPTS 870, 1300, Acute Inhalation Toxicity, 5 August 1998) and JMAFF test guidelines for acute inhalation studies. One control group and one test group each consisting of 5 females and 5 males were included.

The animals in the test group were exposed by snout-only exposure for 4 hours to air containing a liquid droplet aerosol generated from the test substance, Mannanase, PPE 6432, at a concentration of 0.45 mg enzyme concentrate dry matter/L. Mass median aerodynamic diameter and percentage of particles < 7 µm of Mannanase, PPE 6432, were 2.9 µm and 87%, respectively.

 

The animals were observed during exposure, for two hours after the exposure and daily during the 14-day observation period. After the observation period, the animals were sacrificed and examined pathologically.

 

Fur/skin soiled with excreta was observed in all test and control rats immediately after exposure, but was not present 1 hour later. Brown staining on the head was seen in 2 male and female control rats and 2 male test rats up to 1 hour after exposure. These were temporary signs and were considered to be associated with the tube restraint for inhalation exposure.

 

The mean bodyweight gain for male test rats was 69 g compared with 47 g for control rats during the 14-day observation period; however, all body weights and body weight gains were within the expected range throughout the study period. Moreover, the mean liver weight of test males was higher (+14%) than the control values. This was considered to be associated with the slightly heavier bodyweights of the test rats.

 

In conclusion, as there were no unscheduled deaths or evidence of a toxic response following exposure of rats for 4 hours to a droplet aerosol generated from Mannanase, PPE 6432 at a chamber concentration of 0.45 mg enzyme concentrate dry matter/L in air, and the LC₅₀ for mannanase is thus in excess of 0.45 mg enzyme concentrate dry matter/L.