3,4-dihydro-2H-1,4-benzoxazin-6-ol

Endpoint summary

Description of key information

Hydroxybenzomorpholine has a low order of acute oral toxicity. Based on the available key study ((Lheritier, OECD TG 401, GLP, 1989, Klimisch 1), the acute oral LD50 value of the test item Hydroxybenzomorpholine was found to be between 1000 and 2000 mg/kg bw in male and female rats.

No experimental inhalation or dermal acute toxicity studies are available for Hydroxybenzomorpholine.

Acute inhalation toxicity study was waived because exposure of humans via inhalation is unlikely taking into account the relatively high melting temperature (113.2 °C), the low vapour pressure (1.66 x 10 -3 Pa at 20°C) and with the relatively high average median particle size (143μm).

Acute dermal toxicity could be extrapolated from acute oral toxicity study considernig the in vitro dermal absorption with the low dermal absorption of 2.2%. The LD50 demal for Hydroxybenzomorpholine could be calculated to be between 22 727 mg/kg bw/d and 45 440 mg/kg bw/d. Hence, hydroxybenzomorpholine is not expected to be acutely hazardous via the dermal route.

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:

From 7 june 1989 to 24 november 1989

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Version / remarks:

24 February 1987

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

standard acute method

Limit test:

no

Species:

rat

Strain:

other: OFA.SD. (lOPS Caw)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Rationale for alternative/additional species to rat (if applicable) : not applicable
- Source: Iffa-Credo (69210 L'Arbresle- France)
- Females (if applicable) nulliparous and non-pregnant: yes
- Rationale for use of males (if applicable) : not applicable
- Age at study initiation: young adults between 5 and 7 weeks old
- Weight at study initiation: from 127 g to 172 g (the individual weights for each sex varied by no more than 20 % of the mean weights of the animals).
- Fasting period before study: test article was administered after about 17 to 18 hours of fasting (water regimen only). Animals received food again 4 hours after the administration.
- Housing: housed by sex and in groups of 5 in type MI polycarbonate cages (interior dimensions 365 x 225 x 180 mm);bedding changed once a week ; cages changed for each study
- Historical data: not specifed in the study report
- Diet (e.g. ad libitum): complete pelleted rat-mouse maintenance diet ad libitum (U.A.R., formule A.04 CR- U.A.R., Villemoisson- 91360 Epinay-S/Orge- France)
- Water (e.g. ad libitum): softened and filtered (15 m) mains drinking water ad libitum. Bacteriological and chemical analyses twice a year.
No known contaminant was present in the food or the at a concentration which could have prejudiced the study. Certificates and water analysis results are kept in the testing facility.
- Acclimation period: at least 8 days before the start of treatment
- Microbiological status when known : not specified in the study report
- Method of randomisation in assigning animals to test and control groups: they were taken from the animal house stock after the acclimatisation period and after weighing were allocated to groups according to weight in order to obtain homogeneous groups of each sex. They were then taken as they came to hand from these weight classified groups in order to obtain the required number of groups for the study.
Clinical examinations of animals were performed at reception then before the start of treatment to ensure that only healthy animals are included in the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3°
- Humidity (%): 30 to 70 R.H
- Air changes (per hr): at least 10 per hour
- Photoperiod (hrs dark / hrs light): artificial, 12 hours out of 24 (photoperiod= 7h30- 19h30)

IN-LIFE DATES: From: To: not specified in the study report


.

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Remarks:

as a 0.5 % (W/V) dispersion in distilled water (house preparation)

Details on oral exposure:

VEHICLE
- Concentration in vehicle: test article in a 2.5 - 5.0 - 10.0 and 25.0 % (W/V) suspension in the vehicle.
- Amount of vehicle (if gavage): not specified
- Justification for choice of vehicle: vehicle requested by the sponsor
- Lot/batch no. (if required): 85133
- Purity: not specified

MAXIMUM DOSE VOLUME APPLIED: 20 ml/kg of test article

DOSAGE PREPARATION (if unusual): not applicable

CLASS METHOD (not applicable)

Doses:

5000, 2000, 1000 and 500 mg/kg bw.

No. of animals per sex per dose:

20 males and 20 non-pregnant females.

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: any deaths and abnormal clinical signs were noted 15 minutes after administration of the test article, then at 1, 2 and 4 hours and then daily for the 14 day study period detailing the nature and duration of any clinical signs observed. Animals were weighed the day before treatment (Day -1), immediately before administration of the test article (Day 1), at Days 8 and 15 as well as at time of death from Day 2 onwards.
- Necropsy of survivors performed: yes, performed in all animals that died during the study. After the 14 days study period and the final observation (Day 15), all surviving rats were killed by means of overdosing with carbon dioxide and then necropsied. The abdominal and thoracic cavities were opened and particular attention was paid to the·following organs: liver, heart, kidneys and lungs.
- Clinical signs including body weight: the daily observations performed, amongst others, included changes in the skin and fur, the eyes, mucous membranes, respiratory system, circulator system, autonomic and central nervous systems, as well as somata-motor activity and behaviour. Shivering, convulsions, salivation, diarrhoea, lethargy, sleeping and coma were noted with particular attention.
- Other examinations performed: none.
- Data analyisis:
-> The data were recorded indicating for each group employed the number of animals of each sex at the start of the study, the cumulative mortality for each observation period and for each sex, the number of animals presenting with signs of toxicity, a description of the toxic effects and the individual results of the necropsy examinations.
-> The body weights of the animals were evaluated for each group and each sex, calculating the mean of the values obtained, the standard deviation, the variation coefficient which indicates the homogeneity of the data.
The evaluation:- of these data included the relationship, if any, between exposure of the animals to the test article and the incidence as well as the severity of all abnormalities including changes in behaviour and clinical abnormalities, macroscopic lesions, changes in body weight, mortality and any other toxic effects.
-> LD50 calculation: taking into account the choice of the different dose levels, the dose level which is lethal for 50 out of 100 animals (LD 50) could only be estimated. The results of the LD 50 were expressed in mg of test article per kg of animal for both males and females combined.

Statistics:

None.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 1 000 - < 2 000 mg/kg bw

Based on:

test mat.

Mortality:

Cumulative mortality rates were as follows:

Dose of 5000 mg/kg bw: 100%
Dose of 2000 mg/kg bw: 100%
Dose of 1000 mg/kg bw: 20%
Dose of 500 mg/kg bw: 0%

Details on mortality are presented in Table 1 in section "Any other information on results incl. tables".

Clinical signs:

other: The following significant changes were noted: . at 5000 mg/kg: a comatose state associated with clonic convulsions was observed in all animals within 1/4 hour after treatment and in the 6 survivors at 1 hour, then lethargy was noted in the last survivor a

Gross pathology:

Areas of congestion were observed in the lungs of animals that died during the study. No abnormality was noted in the rats sacrificed at the end of the test (Day 15).

Other findings:

None.

Table 1. Cumulative mortality results

Dose level (mg/kg bw)	Sex	Day																		Total mortality (%)
		1				2	3	4	5	6	7	8	9	10	11	12	13	14	15
		¼ H	1H	2H	4H
500	M	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	F	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
1000	M	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	20
	F	0	0	0	0	2	2	2	2	2	2	2	2	2	2	2	2	2	2
2000	M	0	0	0	1	5	5	5	5	5	5	5	5	5	5	5	5	5	5	100
	F	0	1	3	5	5	5	5	5	5	5	5	5	5	5	5	5	5	5
5000	M	0	1	4	5	5	5	5	5	5	5	5	5	5	5	5	5	5	5	100
	F	0	3	5	5	5	5	5	5	5	5	5	5	5	5	5	5	5	5

 

Table 2. Evolution of the body weights

		Day -1	Day 1	Day 8	Day 15	Day 15 - D -1	Dead
Group 4 (500 mg/kg bw)
Males		175	159	215	271	96
		176	160	202	274	98
		187	164	234	288	101
		181	160	203	254	73
		174	153	204	259	85
	Mean	178.60	159.20	211.6	269.20	90.60
	S.D	5.41	3.96	13.58	13.37	11.55
	C.V. (%)	3.03	2.49	6.42	4.97	12.74
	t
Females		156	142	190	222	66
		162	142	205	225	63
		151	137	183	218	67
		139	127	167	188	49
		153	138	179	195	42
	Mean	152.20	137.20	184.80	209.60	57.40
	S.D	8.47	6.14	14.04	16.89	11.24
	C.V. (%)	5.56	4.48	7.60	8.06	19.58
	t
Group 2 (1000 mg/kg bw)
Males		178	155	161	220	42
		180	158	152	189	9
		190	166	169	226	36
		183	159	152	215	32
		175	156	145	190	15
	Mean	181.20	158.80	155.80	208.00	26.80
	S.D	5.72				14.13
	C.V. (%)	3.16	2.72	5.98	8.33	52.73
	t
Females		162	143	136	198	36
		162	145	156	190	28
		168	148	***	***	***	146
		152	139	162	197	45
		158	141	***	***	***	140
	Mean	160.40	143.20	151.33	195.00	36.33
	S.D	5.90	3.49	13.61	4.36	8.50
	C.V. (%)	3.68	2.44	9.00	2.24	23.41
	t
Group 3 (2000 mg/kg bw)
Males		191	171	***	***	***	169
		177	159	***	***	***	158
		170	152	***	***	***
		185	163	***	***	***	162
		179	156	***	***	***	154
	Mean	180.40	160.20
	S.D	7.99	7.26
	C.V. (%)	4.43	4.53
	t
Females		150	132	***	***	***
		162	146	***	***	***
		155	138	***	***	***
		163	142	***	***	***
		157	142	***	***	***
	Mean	157.40	140.00
	S.D	5.32	5.29
	C.V. (%)	3.38	3.78
	t
Group 3 (5000 mg/kg bw)
Males		191	171	***	***	***	169
		177	159	***	***	***	158
		170	152	***	***	***
		185	163	***	***	***	162
		179	156	***	***	***	154
	Mean	180.40	160.20
	S.D	7.99	7.26
	C.V. (%)	4.43	4.53
	t
Females		150	132	***	***	***
		162	146	***	***	***
		155	138	***	***	***
		163	142	***	***	***
		157	142	***	***	***
	Mean	157.40	140.00
	S.D	5.32	5.29
	C.V. (%)	3.38	3.78
	t

***: dead animal.

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

Under the conditions of this study, the maximal non-lethal dose of hydroxybenzomorpholine was 500 mg/kg, the minimal lethal dose was 1000 mg/kg, and the median lethal dose (LD50) was between 1000 and 2000 mg/kg.

Executive summary:

A single oral (gavage) 5000 mg/kg bw dose of the test substance to 5 week old rats (5 per sex) after overnight fasting. Due to the mortalities, 5 days later, further groups were dosed at 2000 and 1000 mg/kg bw as 10 and 5% (w/v) suspension, followed by a final group 14 days later, at 500 mg/kg bw dose as 2.5% (w/v) suspension.

Animals were observed at 15 min, 1, 2 and 4h and then daily for clinical signs and mortality. Body weights were recorded on the Day -1, 1, 8 and 15. Post-mortems were carried out on all animals. No organ/tissue samples were taken.

Results

All animals given 5000 mg/kg were found dead within 4 h. The initial effect of clonic convulsions and lethargy were noted in 15 minutes of dosing leading to coma and death.

All females and 1 male given 2000 mg/kg were found dead within 4 h. The remaining males were dead by Day 2. Lethargy and prostration were noted in the first 4 hours of dosing leading to coma and death after showing unconsciousness and clonic convulsions.

At 1000 mg/kg, 2 females were found dead on day 2, and all animals showed unconsciousness and prostration mainly on the day of dosing. Surviving animals resumed normal appearance and behaviour within day 8.

There were no deaths at 500 mg/kg, no body weight changes, and clinical signs were limited to prostration on the day of dosing.

Conclusion

Under the conditions of this study, the maximal non-lethal dose of hydroxybenzomorpholine was 500 mg/kg, The minimal lethal dose was 1000 mg/kg, and the median lethal dose (LD50) was between 1000 and 2000 mg/kg.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

> 1 000 - < 2 000 mg/kg bw

Quality of whole database:

Reliability 1 study (Guideline and GLP compliant study).

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Data waiving:

exposure considerations

Justification for data waiving:

the study does not need to be conducted because exposure of humans via inhalation is not likely taking into account the vapour pressure of the substance and/or the possibility of exposure to aerosols, particles or droplets of an inhalable size

Justification for type of information:

JUSTIFICATION FOR DATA WAIVING
Exposure to Hydroxybenzomorpholine via the inhalation route is unlikely. Indeed, The relatively high melting temperature (113.2 °C) and the low vapour pressure (1.66 x 10 -3 Pa at 20°C), indicate that it is not likely that Hydroxybenzomorpholine vapours will reach the nasopharyngeal region or subsequently the tracheobronchial or pulmonary region. In addition, the relatively high average median particle size (143μm) prevents particles to be inhaled, as these have an aerodynamic diameter higher than 100 μm. Thus, Hydroxybenzomorpholine has a low potential to be inhaled.

Endpoint conclusion

Endpoint conclusion:

no study available

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study does not need to be conducted because the physicochemical and toxicological properties suggest no potential for a significant rate of absorption through the skin

Justification for type of information:

JUSTIFICATION FOR DATA WAIVING
According to Regulation (EC) No 1907/2006, Annex VIII, 8.5.3, column 2 (specific rules for adaptation from column 1), testing by the dermal route is appropriate if:
1) inhalation of the substance is unlikely; and
2) skin contact in production and/or use is likely; and
3) the physicochemical and toxicological properties suggest potential for a significant rate of absorption through the skin.
According to the results of the in vitro dermal absorption study (OECD TG Guideline 428), dermal absorption rate for hydroxybenzomorpholine was considered as 2.2%. Given the low and non-significant rate of absorption through skin of the substance, acute toxicity testing by the dermal route is not considered appropriate. However, skin contact may occur during worker manufacturing, formulation or consumer use.
Hydroxybenzomorpholine exhibited a low order of acute oral toxicity with an acute oral LD50 value between 1000 and 2000 mg/kg bw in male and female rats. Taking into consideration that dermal absorption rate for hydroxybenzomorpholine is 2.2% and by default that oral absorption is 50%, an extrapolation could be done by calculation
50% oral vs. 2.2% dermal = 50 :2.2 = 22.72
The LD50 demal for Hydroxybenzomorpholine could be calculated to be between 22 727 mg/kg bw/d and 45 440 mg/kg bw/d.
• 1000 mg/kg bw/d x 22.72 = 22 727 mg/kg bw/d
• 2000 mg/kg bw/d x 22.72 = 45 440 mg/kg bw/d
Hydroxybenzomorpholine is not expected to be acutely hazardous via the dermal route.
No further testing is required.

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

One key study is available for assessment of acute oral toxicity (Lheritier, OECD TG 401, GLP, 1989, Klimisch 1):

A single oral (gavage) 5000 mg/kg bw dose of the test substance to 5 week old rats (5 per sex) after overnight fasting. Due to the mortalities, 5 days later, further groups were dosed at 2000 and 1000 mg/kg bw as 10 and 5% (w/v) suspension, followed by a final group 14 days later, at 500 mg/kg bw dose as 2.5% (w/v) suspension. Animals were observed at 15 min, 1, 2 and 4h and then daily for clinical signs and mortality. Body weights were recorded on the Day -1, 1, 8 and 15. Post-mortems were carried out on all animals. No organ/tissue samples were taken. All animals given 5000 mg/kg were found dead within 4 h. The initial effect of clonic convulsions and lethargy were noted in 15 minutes of dosing leading to coma and death. All females and 1 male given 2000 mg/kg were found dead within 4 h. The remaining males were dead by Day 2. Lethargy and prostration were noted in the first 4 hours of dosing leading to coma and death after showing unconsciousness and clonic convulsions. At 1000 mg/kg, 2 females were found dead on day 2, and all animals showed unconsciousness and prostration mainly on the day of dosing. Surviving animals resumed normal appearance and behaviour within day 8. There were no deaths at 500 mg/kg, no body weight changes, and clinical signs were limited to prostration on the day of dosing.
Hence, under the conditions of this study, the maximal non-lethal dose of hydroxybenzomorpholine was 500 mg/kg, The minimal lethal dose was 1000 mg/kg, and the median lethal dose (LD50) was between 1000 and 2000 mg/kg.
No inhalation or dermal acute toxicity studies are available for Hydroxybenzomorpholine.
Exposure to Hydroxybenzomorpholine via the inhalation route is unlikely. Indeed, The relatively high melting temperature (113.2 °C) and the low vapour pressure (1.66 x 10 -3 Pa at 20°C), indicate that it is not likely that Hydroxybenzomorpholine vapours will reach the nasopharyngeal region or subsequently the tracheobronchial or pulmonary region. In addition, the relatively high average median particle size (143μm) prevents particles to be inhaled, as these have an aerodynamic diameter higher than 100 μm. Thus, Hydroxybenzomorpholine has a low potential to be inhaled.
According to the results of the in vitro dermal absorption study (OECD TG Guideline 428), dermal absorption rate for hydroxybenzomorpholine was considered as 2.2%. Given the low and non-significant rate of absorption through skin of the substance, acute toxicity testing by the dermal route is not considered appropriate. However, skin contact may occur during worker manufacturing, formulation or consumer use.
Hydroxybenzomorpholine exhibited a low order of acute oral toxicity with an acute oral LD50 value between 1000 and 2000 mg/kg bw in male and female rats. Taking into consideration that dermal absorption rate for hydroxybenzomorpholine is 2.2% and by default that oral absorption is 50%, an extrapolation could be done by calculation
50% oral vs. 2.2% dermal = 50 :2.2 = 22.72
The LD50 demal for Hydroxybenzomorpholine could be calculated to be between 22 727 mg/kg bw/d and 45 440 mg/kg bw/d.
• 1000 mg/kg bw/d x 22.72 = 22 727 mg/kg bw/d
• 2000 mg/kg bw/d x 22.72 = 45 440 mg/kg bw/d
Hydroxybenzomorpholine is not expected to be acutely hazardous via the dermal route.
No further testing is required.

Justification for classification or non-classification

Based on the available key study, the acute oral LD50 value of the test item Hydroxybenzomorpholine was found to be between 1000 and 2000 mg/kg bw in male and female rats. Hence, the substance Hydroxybenzomorpholine requires classification as Category 4 (Acute Tox. 4, H302) for acute oral toxicity according to the CLP criteria.