3,3'-methylenebis[5-methyloxazolidine]

Endpoint summary

Description of key information

The acute toxicity after oral, inhalation and dermal exposure has been investigated in valid studies on rats.

The oral LD50 ranged from 630 (Leuschner 2002) to 900 mg/kg bw (Leuschner 1979). Clinical signs observed in rats after oral application were sedation, ataxia and dyspnea 5-10 minutes after application followed by coma and death. Pathology revealed no treatment related effects. Similar results were reported in two further oral studies (surprisingly no local effects detected).
The LC50 after inhalation exposure for 4h was >1000 mg/m3 (Fraunhofer ITEM 2010) and the cut-off value of 2000 mg/m3 will be used according to OECD Guideline and GHS.

The dermal LD50 in rats ranged from 760 to 1400 mg/kg bw (Leuschner 2002, Stephen 2000). Lethargy, local erythema, abdominal breathing, nostril discharge and piloerection on day 1 and 2 were reported after acute dermal exposure and at higher dose levels additionally tremor and gasping. The local skin effects (necrosis) were not reversible within 14 days (Stephen, 2000). Similar results were presented by Leuschner 2002. In both studies no treatment-related findings were detected at necropsy except local effects (scab formation).
Clinical signs after application and the dose-effect-level suggested similar absorption pattern of the test substance after oral and dermal exposure (presuming that effects are not exclusively secondary to local necrosis after dermal application).

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Reference 1

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

October 30, 2001 to November 12, 2001 (experimental period)

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

According to the guideline the 2nd dose level should be 300 mg/kg bw (authors used 200 mg/kg bw; minor restriction).

Qualifier:

according to guideline

Guideline:

EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)

Principles of method if other than guideline:

The study was originally intended as Limit test (2000 mg/kg bw). As mortalities occured, the lower dose of 200 mg/kg bw was selected to establish the required information for hazard assessment and hazard classification according to Directive 67/548 EEC.

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland GmbH, D 97633 Sulzfeld
- Age at study initiation: males 36 days, females 48 days
- Weight at study initiation: males 183-197 g and females 173-187 g at dosing
- Fasting period before study: 16 hours
- Housing: In groups of 3 in Macrolon Type 3 cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +-3°C
- Humidity (%): 55 +-15%
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From Sept 24, 2001 to Nov 12, 2001

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

VEHICLE
- Concentration in vehicle: Low dose: 10.7%; high dose undiluted
- Amount of vehicle (if gavage): 1.87 mL/kg bw
- Justification for choice of vehicle: To form a homogenous suspension
- Lot/batch no. (if required): 89 H 0149 (Sigma Aldrich)


MAXIMUM DOSE VOLUME APPLIED: 1.87 mL/kg bw

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: Study was intended tp be performed at Limit test at 2000 mg/kg bw. As mortalities occured, a lower does of 200 mg/kg bw was employed

Doses:

Single oral dose of 2000 and 200 mg/kg bw

No. of animals per sex per dose:

3 males and 3 females per dose group

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Clinical observations 5, 15, 30, 60 min, and 3, 6, 24 h after application and then at least once daily.
Body weight measured day 0, 7, and 14.

- Necropsy of survivors performed: yes
- Necropsy of animals which died during post exposure

Statistics:

Calculation by means of regression analysis (Probit)

Sex:

male/female

Dose descriptor:

LD50

Effect level:

ca. 630 mg/kg bw

Based on:

test mat.

Remarks on result:

other: No CL determined

Mortality:

2000 mg/kg bw: lethal for all rats within 6 h after application (except one male which died within 24 h)

200 mg/kg bw: no mortality.

Clinical signs:

other: Reduced motility, ataxia, reduced muscle tone and dyspnoea prior to death

Gross pathology:

No treatment related effects.

Interpretation of results:

harmful

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The acute oral toxicity (LD50) was calculated to be at 630 mg/kg bw. for male and female rats

Executive summary:

Acute toxic class method according to OECD 423. Three males and 3 females per dose received 2000 or 200 mg/kg bw (10.7% in vehicle; high dose undiluted; volume of 1.87 ml/kg bw). All animals died at 2000 mg/kg bw. No mortality occured at 200 mg/kg bw. Afer 2 weeks of the post-exposure observation period, a necropsy was performed.

The acute oral toxicity was at 630 mg/kg bw. Classification: Harmful according to EC Commission Directive 67/548/EEC and subsequent Amendments.

Reference 2

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

October to November 1976

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Remarks:

No guideline study, no details about the test substance, partly limited documentation, but comparable to OECD 401. Test substance is not the substance, but a condensation product of monoethanolamine, 2-hydroxypropylamine and paraformaldehyde; thus it is similar to a certain extent to the substance as specified in Section 1. The results of this study can be used as supporting data.

Qualifier:

no guideline available

Principles of method if other than guideline:

Comparable to OECD 401 with some restrictions. 10 males and 10 females per dose used. The post-observation period was 4 weeks. No information about test item. Limited documentation in report.

GLP compliance:

no

Remarks:

At the time the study was performed, GLP was not compulsory

Test type:

standard acute method

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: S. Ivanovas GmbH & Co, D 7964 Kißlegg
- Age at study initiation: Males: 38 days, Females: 42 days
- Weight at study initiation: 100-105 g
- Fasting period before study: 15-16 hours before application no feed but water ad libitum
- Housing: Single housing in Macrolon Type II cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: No data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24 +- 0.5
- Humidity (%): 60 +- 3
- Air changes (per hr): No data
- Photoperiod (hrs dark / hrs light): No data

IN-LIFE DATES: From October 1976 To November 1976

Route of administration:

oral: gavage

Vehicle:

physiological saline

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 5, 6.4, 7.9, 10, and 12.6%
- Amount of vehicle (if gavage): constant volume of 10mL/kg bw


MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

Doses:

0.5, 0.64, 0.79, 1.00, 1.26 mL/kg bw (no further details, presumably in mL of the neat test substance which is applied at a constant volume of 10 ml vehicle);

No. of animals per sex per dose:

10 males/10 females per group

Control animals:

no

Details on study design:

- Duration of observation period following administration: 28 days
- Frequency of observations and weighing: Clinical observations; food consumption and body weight measured on days 1, 2, and 7.

- Necropsy of survivors performed: yes, necropsy of all surviving rats 4 weeks after application; necropsy of animals which died: during post exposure observation period

Statistics:

LD50 calculated according to Litchfield & Wilcoxon

Sex:

male/female

Dose descriptor:

LD50

Effect level:

ca. 750 mg/kg bw

Based on:

other: Calculation of LD50 in mL and CL's see under remarks

Remarks on result:

other: In mL: For males LD50 = 0.71 mL/kg bw (5% confidence limits: 0.65-0.78 ml/kg bw; For females LD50 = 0.72 mL/kg bw (5% confidence limits: 0.66-0.79 ml/kg bw . At a relative density of 1.05 at 20°C, the LD50 could be calculated as 750 mg/kg bw.

Mortality:

see Table for acute toxicity

Clinical signs:

other: No effects at 0.50 ml/kg bw; at >=0.64 mL/kg bw sedation, ataxia and dyspnea 5-10 minutes after application; in animals which died due to exposure sedation followed by coma and death (apnoea). In all surviving rats clinical effects reversible within 24 h

Gross pathology:

No treatment related effects

Other findings:

No data

Table for Acute Oral Toxicity Dose [ml/kg bw] Number of dead / number of investigated Time of death (range) Observations 0.50 males 0/10, females 0/10 - see text above 0.64 males 2/10, females 2/10 within 24 hours   0.79 males 6/10, females 5/10 within 24 hours   1.00 males 8/10, females 8/10 within 24 hours   1.26 males 10/10, females 10/10 within 24 hours

Interpretation of results:

harmful

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Expressed in mL, the acute oral toxicity for males was at 0.71 mL/kg bw and for females at 0.72 mL/kg bw.
The mortality rate was dose dependent.
Assuming that the test substance as used in this study has a relative density of 1.05 at 20°C, the following value (in mg/kg bw) could be calculated:
LD50 ca. 750 mg/kg bw for males and females.

Executive summary:

An acute oral toxicity study similar to OECD 401 (with some restrictons) was performed in rats.

Test substance is not the substance, but a condensation product of monoethanolamine, 2-hydroxypropylamine and paraformaldehyde and is thus it is similar to a certain extent to the substance as specified in Section 1. The results of this study can be used as supporting data. The acute oral toxicity for male and females rats was at 750 mg/kg bw and classified as "harmful".

Reference 3

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

August to September, 1976

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

No Guideline study, but widely comparable to OECD 401 with some restrictions. No details about the test substance, partly limited documentation. 5 males and 5 females per dose, termination 4 weeks after exposure. Study meets good scientific standards.

Qualifier:

no guideline followed

Principles of method if other than guideline:

6 dose levels with each 5 male and 5 female animals were used. The posttreatment observation period was 4 weeks.

GLP compliance:

no

Remarks:

At the time the study was performed, GLP was not compulsory

Test type:

standard acute method

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: S. Ivanovas GmbH & Co, D 7964 Kißlegg
- Age at study initiation: Males: 52 days, females: 70 days
- Weight at study initiation: 100-130 g
- Fasting period before study: 16 hours
- Housing: Single housing in Macrolon Type III cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: No data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +- 0.5
- Humidity (%): 55 +- 5

IN-LIFE DATES: From August to September 1976

Route of administration:

oral: gavage

Vehicle:

physiological saline

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 2.7, 5.3, 6.7, 8.5, 10.6, 13.4%

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

Doses:

270, 530, 670, 850, 1060 and 1340 mg/kg bw

No. of animals per sex per dose:

5 males and 5 females per dose

Control animals:

no

Details on study design:

- Duration of observation period following administration: 28 days
- Frequency of observations and weighing: Food consumption and body weight measured day 1, 2, and 7
- Necropsy of survivors performed 4 weeks after application
- Necropsy of animals which died during post exposure observation period

Statistics:

LD50 calculated according to Litchfield & Wilcoxon.

Sex:

male

Dose descriptor:

LD50

Effect level:

ca. 900 mg/kg bw

Based on:

test mat.

95% CL:

>= 830 - <= 970

Sex:

female

Dose descriptor:

LD50

Effect level:

ca. 920 mg/kg bw

Based on:

test mat.

95% CL:

>= 830 - <= 1 020

Mortality:

No animal died at 270 or 530 mg/kg bw.
From 670 mg/kg bw onwards, the mortality rate was dose-related. At 1340 mg/kg bw, all animals died.
All rats died within 24 hours after treatment (see Table below); no further increase in mortality rate at day 14; no further data about mortality; confusing documentation about the time of death in one rat of the 850 mg/kg group.

Clinical signs:

other: No effects at 270 and 530 mg/kg bw; at >= 670 mL/kg bw sedation, ataxia and dyspnea ca. 1 hour after application; in animals which died in side position no further symptoms reported. In all surviving rats clinical effects reversible within 24 hours.

Gross pathology:

No treatment-related effects

Other findings:

No data

Table for Acute Oral Toxicity

Dose [mg/kg bw]	Number of dead / number of investigated	Time of death (range)	Observations
270	males 0/5, females 0/5	-	see text above
530	males 0/5, females 0/5	within 24 hours
670	males 2/5, females 0/5	within 24 hours
850	males 1/5, females 2/5	within 24 hours
1060	males 4/5, females 4/5	within 24 hours
1340	males 5/5, females 5/5	within 24 hours

Interpretation of results:

harmful

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The acute oral toxicity was at 900 to 920 mg/kg bw (males or females).

Executive summary:

No guideline study but acceptable for information. 5 males and 5 females per dose received 270, 530, 670, 850, 1060, 1340 mg/kg bw (2.7, 5.3, 6.7, 8.5, 10.6, 13.4% in vehicle); termination 4 weeks after exposure. Mortality was observed from 670 to 1340 mg/kg bw. in a dose-related manner. The acute oral toxicity was at 900 to 920 mg/kg bw (males or females).

 

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

630 mg/kg bw

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)

GLP compliance:

yes (incl. QA statement)

Remarks:

Fraunhofer Institute for Toxicology and Experimental Medicine ITEM, Hannover, Germany

Test type:

acute toxic class method

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

approx. 6-7 weeks of age at delivery, were purchased from Charles River Deutschland, Sulzfeld, Germany.

Route of administration:

inhalation: mixture of vapour and aerosol / mist

Type of inhalation exposure:

nose only

Vehicle:

clean air

Analytical verification of test atmosphere concentrations:

yes

Concentrations:

500 and 1000 mg/m3

No. of animals per sex per dose:

3m + 3f

Control animals:

no

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 1 000 mg/m³ air (nominal)

Based on:

test mat.

Exp. duration:

4 h

Remarks on result:

not determinable due to absence of adverse toxic effects

Mortality:

1 male died at 1000 mg/m3

Clinical signs:

other: All animals of group 2 (500 mg/m3) showed slight clinical signs like coat soiled, salivation, red stains around mouth directly after the end of exposure. During the next 4 hours only a soiled coat was observed. All animals of this group turned back to nor

Body weight:

Body weight was slightly decreased until day 3 in the male group 2 (500 mg/m3). In females of group 3 (1000 mg/m3) the body weight was markedly decreased until day 7 and returned to normal afterwards.

Gross pathology:

No findings were observed in males and females of the mid dose group (group 2).
In group 3 (1000 mg/m3) all animals showed findings in the lung (e.g. red areas, spongy consistency, partly dark red, and glassy areas). In one male of group 3 (1000 mg/m3), red areas in the thymus were observed.

Interpretation of results:

GHS criteria not met

Conclusions:

In an acute inhalation toxicity of MBO following nose only exposure according to the OECD Guideline 436 the LD was >1000 mg/m3 in Wistar rats.
Based on the present study and according to the OECD Guideline 436, the test item will be classified under Category 4 of the Globally Harmonized System of Classification and Labelling of Chemicals (2003). The LC50 cut-off therefore is 2000 mg/m3 (2 mg/L).

Executive summary:

Six male and six female Wistar (WU) rats, approximately 9 weeks old at the day of exposure, purchased from Charles River Deutschland, Sulzfeld, Germany, were exposed for 4 hours to MBO at concentrations of 531 (group 2) or 1173 mg/m 3 (group 3). The temperature was 21.3° C and the relative humidity was between 61.2 (group 2) and 63.1 % (group3) during the exposure period.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LC50

Value:

2 000 mg/m³ air

Acute toxicity: via dermal route

Link to relevant study records

Reference

Reference 1

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

September 24, 2001 to March 07, 2002 (date of Study Plan to Final Report date)

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

No data on the purity of the test substance

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.3 (Acute Toxicity (Dermal))

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

standard acute method

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland GmbH, D 97633 Sulzfeld
- Age at study initiation: Males 33-43 d, females 44-56 d
- Weight at study initiation: Males 215-267 g and females 198-230 g at dosing
- Fasting period before study: 16 hours
- Housing: Single housing in Macrolon type III cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +- 3
- Humidity (%): 55 +- 15
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From November 8, 2001 to December 10, 2001 (day of dosing to termination of in-life phase)

Type of coverage:

semiocclusive

Vehicle:

corn oil

Details on dermal exposure:

TEST SITE
- Area of exposure: dorsal
- % coverage: 10
- Type of wrap if used: Gauze secured with adhesive plaster on the application site

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Residues of test item (if any) were removed
- Time after start of exposure: 24 hours after administration

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 1.88 mL/kg bw
- Concentration (if solution): 13.3%, 40% in corn oil and undiluted
- Constant volume or concentration used: yes

Duration of exposure:

24 hours

Doses:

250, 750, and 2000 mg/kg bw

No. of animals per sex per dose:

5

Control animals:

not required

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: observations 5, 15, 30, 60 min, and 3, 6, 24 h after application and then at least once daily; weighing once weekly
- Necropsy of survivors performed: yes
- Other examinations: skin was observed for erythema and oedema and rated

Statistics:

LD50 calculated by means of regression analysis. The mortality rates at 24 hours and 14 days were used.

Sex:

male/female

Dose descriptor:

LD50

Effect level:

>= 790 mg/kg bw

Based on:

test mat.

Remarks on result:

other: LD50 after 14 days. No CL determined.

Sex:

male

Dose descriptor:

LD50

Effect level:

>= 1 200 mg/kg bw

Based on:

test mat.

Remarks on result:

other: LD50 after 14 days. No CL determined

Sex:

female

Dose descriptor:

LD50

Effect level:

>= 760 mg/kg bw

Based on:

test mat.

Remarks on result:

other: LD50 after 14 days. No CL determined

Mortality:

see Table for Acute Dermal Toxicity

Clinical signs:

other: 250 mg/kg bw: no local or systemic effects 750 mg/kg bw: no local effects but reduced motility, ataxia, dyspnoea in all animals, 1/5 males with slightly reduced muscle tone. 2000 mg/kg bw, systemic effects: like mid dose group with some augmentation; sl

Gross pathology:

No treatment-related findings at necropsy except local effects in the high dose group: severe necrosis in 2/5 females

Table for Acute Dermal Toxicity

Dose [mg/kg bw]	Number of dead / number of investigated	Time of death (range)	Observations
250	males 0/5, females 0/5		See under Clinical signs
750	males 0/5, females 1/5	day 1
2000	males 5/5, females 5/5	day 1-7

Interpretation of results:

harmful

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The LD50 in Sprague-Dawley rats was calculated to be 1200 mg/kg bw for males and 760 mg/kg bw for females.
The dose dependency of observed effects was demonstrated. The test substance showed local and systemic effects. The test substance hydrolysis in contact with water to formaldehyde and 2-hydroxypropylamine. Effects may be mostly related to the hydrolysis product formaldehyde.

Executive summary:

Study according to OECD guideline 402. Five males and 5 females per dose were exposed to 250, 750, and 2000 mg/kg bw (13.3 and 40% in corn oil and undiluted, volume 1.88 mL/kg bw; semi-occlusive). Termination 14 days after dermal exposure.

LD50 after 14 days: 1200 mg/kg bw for males and 760 mg/kg bw for females. Classification: harmful

Reference 2

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

October to November 1976

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study without detailed documentation

Remarks:

No details about the test substance, partly limited documentation; test substance not applied directly to the skin

Qualifier:

according to guideline

Guideline:

other: HAZARDOUS SUBSTANCES, Part 191, Section 11, FDA Washington 1965; comparable to OECD 402 with some restrictions

Principles of method if other than guideline:

5 males and 5 females per dose received semi-occlusive dermal applications of 2.52, 3.18, 4.00, 5.04, 6.35 ml/kg bw (presumably in mL of the neat test substance); post exposure observation period 14 days. The test item was applied to both, intact and scarified skin.

GLP compliance:

no

Remarks:

At the time the study was performed, GLP was not compulsory.

Test type:

other: At the time the study was performed, no Guideline was compulsory. However, the study widely complies to OECD 402. The test item was applied to the intact and scarified skin.

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: S. Ivanovas GmbH & Co, D 7964 Kißlegg
- Age at study initiation: No data
- Weight at study initiation: 100-105 g
- Housing: Single in Macrolon Type III cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: No data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 +-2
- Humidity (%): 60 +-3
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From Octber 1976 to November 1976

Type of coverage:

semiocclusive

Vehicle:

unchanged (no vehicle)

Details on dermal exposure:

TEST SITE
- Area of exposure: dorsal area
- % coverage: 10
- Type of wrap if used: 4-fold gauze, wrapped with aluminium foil and secured by adhesive tape

REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes
- Time after start of exposure: 24 hours

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): Neat substance applied. Volume depended on dose (ca. 0.2-0.6 mL)
- Concentration (if solution): 2.52, 3.18, 4.00, 5.04, 6.35 mL/kg bw (no further details, presumably in ml of the neat test substance)
- Constant volume or concentration used: no

Duration of exposure:

24 hours

Doses:

2.52, 3.18, 4.00, 5.04, 6.35 mL/kg bw (no further details, presumably in ml of the neat test substance)

No. of animals per sex per dose:

5 males 5 females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Clinical observations including irritation. Food consumption and body weight measured at day 1, 2, and 7
- Necropsy of survivors performed: yes
- Necropsy of animals which died during post exposure: yes

Statistics:

LD50 calculated according to Litchfield & Wilcoxon

Sex:

male/female

Dose descriptor:

LD50

Effect level:

>= 5.7 mL/kg bw

Based on:

test mat.

95% CL:

>= 5.18 - <= 6.27

Remarks on result:

other: Values refer to intact skin

Sex:

male/female

Dose descriptor:

LD50

Effect level:

4.7 mL/kg bw

Based on:

test mat.

95% CL:

>= 4.27 - <= 5.17

Remarks on result:

other: Values refer to scarificed skin

Mortality:

see Table below for Acute Dermal Toxicity

Clinical signs:

other: No effects at 2.52 mL/kg bw. At 3.18 mL/kg bw sedation 1-3 after application for 24 hours; no irritation; reduced food consumption on day 1 and 2; no relevant effects on body weight gain. At 4.00 mL/kg bw additional dyspnea 2-3 after application (no furt

Gross pathology:

No treatment-related findings

Table for Acute Dermal Toxicity

Dose [mL/kg bw]	Number of dead / number of investigated	Time of death (range)	Observations
Intact skin
2.52	males 0/5, females 0/5	-	see text above
3.18	males 0/5, females 0/5	-
4.00	males 0/5, females 0/5	-
5.04	males 0/5, females 0/5	-
6.35	males 5/5, females 5/5	12-48 hours
Scarified skin
2.00	males 0/5, females 0/5	-	no effects
2.52	males 0/5, females 0/5	-	LOEL for systemic effects, see above
3.18	males 0/5, females 0/5	-
4.00	males 1/5, females 1/5	12-24 hours
5.04	males 2/5, females 2/5	12-24 hours
6.35	males 5/5, females 5/5	12-48 hours

Interpretation of results:

practically nontoxic

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Intact skin:
For males and females LD50 = 5.7 mL/kg bw (5% confidence limits: 5.18-6.27 mL/kg bw)
Assuming that Grotamar71 is the test substance used here with a relative density of 1.05 at 20°C, the following value could be calculated:
LD50 ca. 6000 mg/kg bw for males and females

Executive summary:

Test substance is not the substance, but a condensation product of monoethanolamine, 2-hydroxypropylamine and paraformaldehyde; thus it is similar to a certain extent to the substance specified in Section 1. The results of this study can be used as supporting data. No guideline study but acceptable for information (comparable to guideline study with acceptable restriction). Five males and 5 females per dose received semi-occlusive dermal applications of 2.52, 3.18, 4.00, 5.04 and 6.35 mL/kg bw; post exposure observation period was 14 days. The acute dermal toxicity for the intact skin was at 5.7 mL/kg bw.

The LD50 value is higher than in the other studies reported on acute dermal toxicity, which might by related to the purity of the active component.

 

Reference 3

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

April 18, 2000 to October 06, 2000

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Guideline study, well performed and reported according to good scientific standards

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Deviations:

no

GLP compliance:

yes

Test type:

standard acute method

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Jai Research Foundation, India
- Age at study initiation: No data
- Weight at study initiation: 210-298 g
- Fasting period before study: no
- Housing: Rats held singly in polypropylene cages after treatment
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 to 24
- Humidity (%): 67 (mean)
- Air changes (per hr): 20 to 25
- Photoperiod 12 hrs dark /12 hrs light

IN-LIFE DATES: From May 27, 2000 to July 03, 2000

Type of coverage:

semiocclusive

Vehicle:

unchanged (no vehicle)

Details on dermal exposure:

TEST SITE
- Area of exposure: dorsal
- % coverage: about 10% of body surface
- Type of wrap if used: porous gauze fastened with hypo-allergical surgical tape

REMOVAL OF TEST SUBSTANCE
- Washing (if done): with cotton moistened with water
- Time after start of exposure: 24 hours after adminstration

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0, 1000, 1350 and 1823 mg/kg bw
- Concentration (if solution): applied undilutedly
- Constant volume or concentration used: no, volume depended on dose

Duration of exposure:

24 hours

Doses:

0, 1000, 1350 and 1823 mg/kg bw

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: animals were observed for mortality and clinical signs at 30 min, 1, 3 and 6 hours post dosing and twice daily until day 14. Body weights were recorded prior to dosing and on days 7 and 14
- Necropsy of survivors performed: yes
- Other examinations performed: histopathology of treated skin

Statistics:

LD50 calculated according to Probit analyis (Finney, 1971).

Preliminary study:

Dose range finding was performed with 2 males and 2 females per group at dose levels of 500, 1000, 2000 and 2500 mg/kg bw

Sex:

male/female

Dose descriptor:

LD50

Effect level:

>= 1 400 mg/kg bw

Based on:

test mat.

95% CL:

> 1 207 - < 1 620

Mortality:

Mortalty rate was 0, 10, 50 and 80% at the doses of 0, 1000, 1350 and 1823 mg/kg bw

(see also Table for acute dermal toxicity)

Clinical signs:

other: No effects in the control group. 1000 mg/kg bw: no effects 6 h after application but lethargy, local erythema, abdominal breathing, nostril discharge and piloerection on day 1 and 2; all animals appeared normal on day 4 except one female which died on

Gross pathology:

Animals found dead in various treatment groups revealed vascular changes in viscera like lungs, liver, spleen and hemorrhagic exudates in the small intestine. In survivors insignificant pneumonic changes and atrophy or enlargement of the spleen, congestion of the kidneys and hydrometra of the uterus were detected.
At >= 1000 mg/kg bw: Local epidermal thickening / erythema in rats found dead and mild to moderate local scab formation in survivors treated with the test substance.

Other findings:

- Histopathology: Skin histopathology: at >= 1000 mg/kg bw survivors showed at termination local effects: cellular debris/necrotic tissue (scab), leucocytic infiltration, epidermal cyst

Table for Acute Dermal Toxicity

Dose [mg/kg bw]	Number of dead / number of investigated	Time of death (range)	Observations
0	males 0/5, females 0/5	-	see text above
1000	males 0/5, females 1/5	day 8
1350	males 3/5, females 2/5	day 1-5
1823	males 3/5, females 5/5	day 1-2

Interpretation of results:

harmful

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

For males and females combined LD50 = 1400 mg/kg bw (95% confidence limits 1207-1620 mg/kg bw).

Executive summary:

Study according to OECD guideline 402. Five males and 5 females per dose received 0, 1000, 1350, 1823 mg/kg bw of the undiluted test substance (semi-occlusive); post exposure observation period 14 days.

The LD₅₀in Wistar rats (males and females combined) was calculated to be 1400 mg/kg bw (95% confidence limits 1207-1620 mg/kg bw).
The dose dependency of observed effects was demonstrated. The test substance induced local effects, which were not reversible within 14 days. GrotaMar71 hydrolysis in contact with water to formaldehyde and 2-hydroxypropylamine. Effects may be mostly related to the hydrolysis product formaldehyde.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

760 mg/kg bw

Additional information

Acute oral and dermal toxicity was investigated in studies according to current guidelines. The oral LD50 of the substance in rats ranged from 630 to 900 mg/k bw. Clinical signs observed in rats after oral application were sedation, ataxia and dyspnea followed by coma and death. Pathology revealed no treatment related effects; surprisingly no local effects were detected.

The LC50 after inhalation exposure is >1000 mg/m3 (Fraunhofer ITEM 2010) and the cut-off of 2000 mg/m3 will be used according OECD Guideline 436 and GHS.

The dermal LD50 in rats ranged from 790 to 1400 mg/kg bw. Lethargy, local erythema, abdominal breathing, nostril discharge and piloerection on day 1 and 2 were reported after acute dermal exposure and at higher dose levels additionally tremor and gasping. The local skin effects (necrosis) were not reversible within 14 days (no further treatment-related findings were detected at necropsy).

Justification for classification or non-classification

Proposal for classification and labelling of the substance: Harmful in contact with skin and if swallowed, R21/22.

According to GHS: dermal Category 3; oral Category 4

In a study conducted according to OECD guideline 404 in rabbits (4 h dermal exposure) to 0.5 mL undiluted substance resulted in destruction of skin tissue. This corrosive properties were also shown for the products of hydrolysis, e.g. formaldehyde.

Threshold concentration for skin irritation of the substance was 5% in Alembicol D, no effects were detected at 1%).