Benzenepropanal, 2-methyl-4-(2-methylpropyl)-

Endpoint summary

Description of key information

Acute Oral Toxicity

The acute oral toxicity with NYMPHEAL was assessed in the rat (Fixed Dose Method) according to the OECD No.420 (2001) " Acute toxicity-oral, Fixed Dose Procedure".

LD50 oral > 2000mg/kg bw.

Acute Dermal Toxicity:

The acute dermal toxicity with NYMPHEAL in the rat was assessed according the OECD No.402 (1987) "Acute Dermal Toxicity".

LD50 dermal > 2000mg/kg bw.

Acute Inhalation Toxicity:

The acute inhalation toxicity with NYMPHEAL in the rat was assessed according the OECD No.403 (2009) "Acute Inhalation Toxicity".

LC50, 4h in the range of 1 -5 mg/L, based only on animal sacrificed on D2 due to clinical observations and where at necropsy only 1/5 Male and 1/5 Female at dose 5mg/L were found with actual macroscopic findings in the lung (several red foci). Moreover, all surviving animals in the lower doses saw their clinical signs recover by D3.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

February 2015 to April 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1100 (Acute Oral Toxicity)

GLP compliance:

yes (incl. QA statement)

Test type:

fixed dose procedure

Limit test:

yes

Specific details on test material used for the study:

Name (as stated in the report): GR-88-0778
Lot No: Batch 6
Aspect: Colourless liquid
Expiration date: March 29, 2017

Species:

rat

Strain:

Wistar

Remarks:

Crl:WI (outbred, SPF-Quality)

Sex:

female

Details on test animals or test system and environmental conditions:

- Source: Charles River Deutschland, Sulzfeld, Germany
- Number of animals: Pilot Study: 1 female, Main Study: 4 females (females were nulliparous and non-pregnant).
- Age and body weight: Young adult animals (approx. 8-9 weeks old) were selected. Body weight variation did not exceed +/- 20% of the sex mean.
- Identification: Earmark
- Health inspection: 4At least prior to dosing. It is ensured that the animals were healthy and without any abnormality that might affect the study integrity

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

- Method: Oral gavage, using plastic feeding tubes. The test substance was stirred on a magnetic stirrer during dosing.

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

a group of 4 females (females were nulliparous and non-pregnant) / 1 dose per group

Control animals:

no

Details on study design:

Initially, GR-88-0778 was administered by oral gavage to one female Wistar rat at 2000 mg/kg body weight. As no mortality occurred and no signs of significant toxicity were observed, the main study was conducted with a fixed dose of 2000 mg/kg body weight administered to four female rats. The animals were subjected to daily observations. Body weights were determined on Days 1, 8 and 15.
Macroscopic examination was performed after terminal sacrifice.

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality occurred.

Clinical signs:

other: Hunched posture, uncoordinated movements and/or piloerection were noted for the animals on Day 1.

Gross pathology:

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

Interpretation of results:

not classified

Conclusions:

The minimum oral lethal dose of GR-88-0778 in rats was established to exceed 2000 mg/kg body weight.

Executive summary:

The acute oral toxicity with GR-88-0778 was assessed in the rat (Fixed Dose Method) according to the OECD No.420 (2001) " Acute toxicity-oral, Fixed Dose Procedure".

Initially, GR-88-0778 was administered by oral gavage to one female Wistar rat at 2000 mg/kg body weight. As no mortality occurred and no signs of significant toxicity were observed, the main study was conducted with a fixed dose of 2000 mg/kg body weight administered to four female rats. The animals were subjected to daily observations. Body weights were determined on Days 1, 8 and 15. Macroscopic examination was performed after terminal sacrifice.

No mortality occurred.

Hunched posture, uncoordinated movements and/or piloerection were noted for the animals on Day 1.

The 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 minimum oral lethal dose of GR-88-0778 in rats was established to exceed 2000 mg/kg body weight.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Quality of whole database:

Reliability 1, according to guideline and GLP study.

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

Study period:

From September 27, 2016 to October 26, 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.2 (Acute Toxicity (Inhalation))

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1300 (Acute inhalation toxicity)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

no

Specific details on test material used for the study:

Appearance: Colourless to pale yellow liquid
Batch: SC00018060
Purity/Composition: 98,3% (sum of isomers)
Test item storage: At room temperature protected from light
Stable under storage conditions until: 28 April 2017 (expiry date)

Species:

rat

Strain:

Wistar

Remarks:

Crl:WI(Han) (outbred, SPF-Quality)

Sex:

male/female

Details on test animals or test system and environmental conditions:

Number of animals: 5 males and 5 females (females were nulliparous and nonpregnant) per exposure level. Two exposure levels
Age and body weight: Young adult animals were selected (approximately 10-11 weeks old). Animals used within the study were of approximately the same age and body weight variation did not exceed +/- 20% of the sex mean.
Identification: Individual unique number by tattoo on hind leg.
Conditions: Environmental controls for the animal room were set to maintain 18 to 24°C, a relative humidity of 40 to 70%, at least 10 air changes/hour, and a 12-hour light/12-hour dark cycle. Any variations to these conditions were maintained in the raw data and had no effect on the outcome of the study. Health inspection: At least prior to exposure. It was ensured that the animals were healthy and without any abnormality that might affect the study integrity.
Accommodation: Group housing of five animals per sex per cage in labelled Makrolon cages (type IV; height 18 cm) containing sterilized sawdust as bedding material (Lignocel S 8-15, JRS -J.Rettenmaier & Söhne GmbH + CO. KG, Rosenberg, Germany) and paper as cageenrichment (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom). Acclimatization period was at least 5 days before start of treatment under laboratory conditions.
Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany) except during exposure to the test item.
Water: Free access to tap water except during exposure to the test item.
Animal husbandry on the day of exposure.
The animals were moved to the inhalation area to in order to perform the exposure. During the exposure, there was no access to food and water. After exposure, the animals were returned to their cages which were placed in a fume cupboard for a short time period to allow test item remnants to evaporate. A sheet of filter paper was used to cover the bedding material to prevent suffocation in case of bad health condition and in order to recover and to aid the clinical observations. The sheet was removed and before the end of the exposure day, the surviving animals were returned to the animal room. Diet, water, bedding and cage enrichment evaluation for contaminants and/or nutrients was performed according to facility standard procedures. There were no findings that could interfere with the study.

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

air

Remarks:

pressurized air

Mass median aerodynamic diameter (MMAD):

>= 2.6 - <= 3 µm

Geometric standard deviation (GSD):

>= 1.9 - <= 2.1

Remark on MMAD/GSD:

The Mass Median Aerodynamic Diameter (MMAD) and geometric standard deviation (gsd) were determined twice during the exposure period. At 5 mg/L, the MMAD was 3.0 μm (gsd 1.9) at both measurements.
At 1 mg/L, the MMAD was 2.6 μm (gsd 2.0) and 2.9 μm (gsd 2.1).

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

4 h

Concentrations:

Tested concentrations: 5 mg/L and 1 mg/L.
For the 5 mg/L exposure group, the time-weighted mean actual concentration was 5.3 ± 0.2 mg/L. The nominal concentration (amount of test item used divided by the volume of pressurized air used) was 6.2 mg/L. The generation efficiency (ratio of actual and nominal concentration) was 85%. For the 1 mg/L exposure group, the time-weighted mean actual concentration was 1.0 ± 0.03 mg/L. The nominal concentration was 1.5 mg/L and the generation efficiency was 70%. The concentration measurements equally distributed over time showed that the item was sufficiently stable.

No. of animals per sex per dose:

Five animals of each sex were exposed for 4 hours to a target concentration of the test item of 5 mg/L.
Based on the results, five animals of each sex were exposed to a target concentration of 1 mg/L.

Control animals:

no

Details on study design:

Mortality and clinical signs were observed daily during the observation period and body weights were determined on Days 1, 2, 4, 8 and 15. Macroscopic examination was performed on the day of death or after terminal sacrifice (Day 15).

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

>= 1 - <= 5 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Mortality:

All animals exposed to 5 mg/L were sacrificed for ethical reasons on Day 2 and one male exposed to 1 mg/L was sacrificed on Day 10. No further mortality occurred.

Clinical signs:

other: At 5 mg/L, slow breathing was seen during exposure (not presented in the table). After exposure, lethargy, hunched posture, laboured respiration, gasping, piloerection, chromodacryorrhoea on the nose, ptosis and hypothermia were noted for the animals. At

Body weight:

Overall body weight gain in surviving males and females was within the range expected for rats of this strain and age used in this type of study and were therefore considered not indicative of toxicity.

Gross pathology:

Macroscopic post mortem examination of the animals that were sacrificed for ethical reasons during the study revealed abnormalities of the lungs (several reddish foci in the lungs of one male and one female exposed to 5 mg/L and foamy contents for the male exposed to 1 mg/L) and the spleen (reduced in size for one male exposed to 5 mg/L).

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

The inhalation LC50, 4h value of NYMPHEAL in Wistar rats was established to be within the range of 1 – 5 mg/L, based on animals sacrificed for ethical reasons.
It is to be noted that during macroscopic examination no abnormalities were found in the sacrificed animals except for one male and one female at 5mg/L and one male at 1mg/L.

Executive summary:

Assessment of acute inhalation toxicity with NYMPHEAL in the rat (nose-only).

The study was carried out based on the guidelines described in:

- OECD Guidelines, Section 4, Health Effects. No.403, "Acute Inhalation Toxicity", Sep 2009.

- Commission Regulation (EC) No 440/2008, B.2. Acute Toxicity (inhalation), L142, May 2008.

- EPA OPPTS 870.1300, Acute inhalation Toxicity. EPA 712-C-98-193, August 1998.

- JMAFF Guidelines (2000), including the most recent revisions.

NYMPHEAL was administered as an aerosol by nose-only inhalation for 4 hours to two groups of five male and five female Wistar rats. Mortality and clinical signs were observed daily during the observation period and body weights were determined on Days 1, 2, 4, 8 and 15. Macroscopic examination was performed on the day of death or after terminal sacrifice (Day 15).

For the 5 mg/L exposure group, the time-weighted mean actual concentration was 5.3 ± 0.2 mg/L. The nominal concentration (amount of test item used divided by the volume of pressurized air used) was 6.2 mg/L. The generation efficiency (ratio of actual and nominal concentration) was 85%. For the 1 mg/L exposure group, the time-weighted mean actual concentration was 1.0 ± 0.03 mg/L. The nominal concentration was 1.5 mg/L and the generation efficiency was 70%. The concentration measurements equally distributed over time showed that the item was sufficiently stable.

The Mass Median Aerodynamic Diameter (MMAD) and geometric standard deviation (gsd) were determined twice during the exposure period. At 5 mg/L, the MMAD was 3.0 μm (gsd 1.9) at both measurements. At 1 mg/L, the MMAD was 2.6 μm (gsd 2.0) and 2.9 μm (gsd 2.1).

All animals exposed to 5 mg/L were sacrificed for ethical reasons on Day 2 and one male exposed to 1 mg/L was sacrificed on Day 10. No further mortality occurred.

At 5 mg/L, slow breathing was seen during exposure. After exposure, lethargy, hunched posture, labored respiration, gasping, piloerection, chromodacryorrhoea on the nose, ptosis and hypothermia were noted for the animals. At 1 mg/L, slow breathing was seen during exposure in one animal. After exposure, lethargy, hunched posture, labored respiration and rales were seen for the animals. The surviving animals had recovered from the clinical signs by Day 3.

Overall body weight gain in surviving males and females was within the range expected for rats of this strain and age used in this type of study and were therefore considered not indicative of toxicity.

Macroscopic post mortem examination of the animals that were sacrificed for ethical reasons during the study revealed abnormalities of the lungs (several reddish foci in the lungs of one male and one female exposed to 5 mg/L and foamy contents for the male exposed to 1 mg/L) and the spleen (reduced in size for one male exposed to 5 mg/L).

The inhalation LC50, 4h value of NYMPHEAL in Wistar rats was established to be within the range of 1 – 5 mg/L.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LC50

Value:

1 000 mg/m³ air

Quality of whole database:

Reliability 1, according to guideline and GLP study.

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

08 September 2016 to 06 October 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Test type:

fixed dose procedure

Limit test:

yes

Specific details on test material used for the study:

Name (as stated in the report): GR-88-0778
Lot No: SC00018060
Aspect: Colourless liquid
Expiration date: 28 April 2017 (expiry date)

Species:

rat

Strain:

Wistar

Remarks:

Crl:WI (Han) (outbred, SPF-Quality).

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Source: Charles River Deutschland, Sulzfeld, Germany.
- Number of animals: 5 males and 5 females (females were nulliparous and nonpregnant).
- Age and body weight: Young adult animals (approx. 10-11 weeks old) were selected. Body weight variation did not exceed +/- 20% of the sex mean.
- Identification: Tail mark with indelible ink.
- Health inspection: At least prior to dosing. It was ensured that the animals were healthy and that the skin to be treated was intact and free from any abnormality.

Type of coverage:

semiocclusive

Vehicle:

unchanged (no vehicle)

Remarks:

The test item was dosed undiluted as delivered by the Sponsor

Details on dermal exposure:

The test item was applied on an area of approx. 10% of the total body surface, i.e. approx. 25 cm² for males and 18 cm² for females. The test item was held in contact with the skin with a dressing, consisting of a surgical gauze patch (Surgy 1D), successively covered with aluminum foil and Coban elastic bandage. A piece of Micropore tape was additionally used for fixation of the bandages in females only.

Duration of exposure:

- Frequence: Single dosage, on Day 1.
- Application period: 24 hours, after which dressings were removed and the skin cleaned of residual test item using tap water.

Doses:

- Dose level (volume): 2000 mg/kg (2.10 mL/kg) body weight. Dose volume calculated as dose level (g/kg) / specific gravity (0.9516).

No. of animals per sex per dose:

1 group of 5 males and 5 females / 1 dose per group

Control animals:

no

Details on study design:

Initially, NYMPHEAL was administered to three female Wistar rats by a single dermal application at 2000 mg/kg body weight for 24 hours. Based on results, an additional group of two females and five males were dosed at 2000 mg/kg body weight. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was scheduled to be performed after terminal sacrifice (Day 15).

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality occurred.

Clinical signs:

other: Flat posture, piloerection, lethargy, quick breathing, shallow respiration, pstosis and/or chromodacryorrhoea on the snout were noted for all animals between Days 1 and 5.

Gross pathology:

No abnormalities were found at macroscopic post mortem examination.

Interpretation of results:

GHS criteria not met

Conclusions:

The dermal LD50 value of NYMPHEAL in Wistar rats was established to exceed 2000 mg/kg body weight.

Executive summary:

The acute dermal toxicity with NYMPHEAL in the rat was assessed according the OECD No.402 (1987) "Acute Dermal Toxicity".

Initially, NYMPHEAL was administered to three female Wistar rats by a single dermal application at 2000 mg/kg body weight for 24 hours. Based on results, an additional group of two females and five males were dosed at 2000 mg/kg body weight. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was scheduled to be performed after terminal sacrifice (Day 15).

No mortality occurred.

Flat posture, piloerection, lethargy, quick breathing, shallow respiration, pstosis and/or chromodacryorrhoea on the snout were noted for all animals between Days 1 and 5. The mean body weight gain during the observation period was within the range expected for rats used in this type of study. No abnormalities were found at macroscopic post mortem examination.

The dermal LD50 value of NYMPHEAL in Wistar rats was established to exceed 2000 mg/kg body weight.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Quality of whole database:

Reliability 1, according to guideline and GLP study.

Justification for classification or non-classification

Based on the data available and key results described in this summary, the substance has shown some acute inhalation toxicity potential mainly via local intolerance in the lungs. Although the final LC50 is based only on sacrificed animals that may not have been given sufficient time to recover and as such may have been oversetimated, as a conservative approach we still recommend the substance to be classified as Acute Toxic (Inhalation) Category 4 according to the (EC) No 1272/2008 Regulation (CLP).

Nympheal has not shown any systemic toxicity by oral or dermal route and as such no further classification is necessary according to the (EC) No 1272/2008 Regulation (CLP).