Dimethylamine-borane (1:1)

Endpoint summary

Description of key information

Two different acute dermal toxicity studies conducted similar to OECD test guideline 402 with the target substance dimethylamine-borane have been reported. Both examined the skin irritation potential of the substance with negative results. The eye irritation potential of the target substance was examined in an in vitro study conducted according to OECD test guideline 437 showing a high irritation potential and based on the results the target substance is classified for irreversible effects on the eye (Category 1, H318).

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Reference 1

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1979-11-14 to 1979-12-26

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Deviations:

yes

Remarks:

see box "principles of method if other than guideline"

Qualifier:

equivalent or similar to guideline

Guideline:

other: OECD 402 "Acute dermal Toxicity"

Deviations:

yes

Remarks:

see box "principles of method if other than guideline"

Principles of method if other than guideline:

Deviations from OECD 404:
- Exposure: 24 h
- Application area: 100 cm²
- Occlusive
- Tested on the intact and abraded skin

Deviations from OECD 402:
- 6 Animals per dose group (abraded and intact skin 3 each)
- Application area: 100 cm²
- Occlusive
- Tested on the intact and abraded skin

GLP compliance:

yes

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No. of test material: dimethylamine-borane, Lot 40-37, was received from Callery Chemical Company on November 14, 1979 as a crystalline powder. The material was mixed with 0.5 mL distilled water to form a thin paste for testing.

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

Eighteen albino rabbits of the New Zealand White variety weighing approximately 2.5 kg were obtained and acclimated to the laboratory for a minimum of seven days. Rabbits were individually housed in suspended cages with feed and water freely available.

Type of coverage:

occlusive

Preparation of test site:

other: clipped and abraded

Vehicle:

water

Remarks:

0.5 mL to form a thin paste

Controls:

no

Amount / concentration applied:

200, 282 and 398 mg/kg body weight

Duration of treatment / exposure:

24 h

Observation period:

14 days

Number of animals:

6 per dose group (3 abraded, 3 with intact skin)

Details on study design:

Prior to application of the test substance, one half of the rabbits received a series of abrasions, with a sterile needle, through the epidermis. The trunk of each rabbit was wrapped with an impervious material taped in place to form a reservoir over the test skin site. The test material was introduced onto the skin of 3 intact and 3 abraded rabbits at dose levels of 200, 282, and 398 mg/kg as a paste. During application, the rabbits were kept in a restrainer device and later were fitted with a collar and returned to their respective cages. Wrappings were removed after 24 hours. Where possible, test material residue was flushed from the skin.
The rabbits were observed immediately after treatment, at four hours and daily for 14 days for signs of illness and mortality. Dermal reactions were scored daily for erythema and edema. Body weights were recorded at dosing and at 14 days for the survivors. Animals found dead or those euthanized were subjected to a macroscopic examination of the viscera. Death per number dosed was plotted and the dermal LD50 estimated.

Irritation parameter:

erythema score

Basis:

mean

Time point:

24/48/72 h

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

mean

Time point:

24/48/72 h

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

There were no significant signs of dermal irritation.

Other effects:

Mortality levels of 33% were noted at 200 and 282 mg/kg bw and of 50% at 398 mg/kg bw. As in the previously reported study, the plotted curve was very steep. Similarly, there was no significant dermal reactions and the surviving rabbits qained weight during the 14 day observation period. Under the conditions of this test, the dermal LD50 can be considered to be 398 mg/kg bw or slightly less toxic than the 210 mg/kg bw (156 - 284 mg/kg) reported earlier.

Interpretation of results:

GHS criteria not met

Conclusions:

In an acute dermal toxicity study conducted in rabbits, no signs of dermal irritation were observed.

Executive summary:

In an acute dermal toxicity study similar to OECD 402, dimethylamine-borane was applied occlusively to the abraded and intact skin of New Zealand White rabbits at concentrations of 200, 282 and 398 mg/kg body weight for 24 h. Afterwards, remaining test material was washed off. The animals were observed for 14 days for clinical signs and mortality including a daily scoring for erythema and edema. Body weight was recorded at dosing and at the end of the study for the survivors. Animals found dead or euthanised at 14 days were subjected to a macroscopic examination of the viscera.

Based on the results, the LD50 can be considered to be 398 mg/kg body weight. No macroscopic changes were noted at necropsy, which could be attributed to the test material. All surviving animals gained body weight during the course of the study. There were no significant signs of dermal irritation noted throughout the study.

Reference 2

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1979-07-31 to 1979-10-03

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Deviations:

yes

Remarks:

see box "principles of method if other than guideline"

Qualifier:

equivalent or similar to guideline

Guideline:

other: OECD 402 "Acute dermal Toxicity"

Deviations:

yes

Remarks:

see box "principles of method if other than guideline"

Principles of method if other than guideline:

Deviations from OECD 404:
- Exposure: 24 h
- Application area: 100 cm²
- Occlusive
- Tested on the intact and abraded skin

Deviations from OECD 402:
- 4 Animals per dose group (abraded and intact skin 2 each)
- Application area: 100 cm²
- Occlusive

GLP compliance:

yes

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No. of test material: dimethylamine-borane, Lot. No. 40-37 was received from Callery Chemical Company.

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

Rabbits of the New Zealand White variety weighing 2.5 kg or greater were obtained and acclimated to the laboratory a minimum of seven days. Rabbits were individually housed in suspended cages with feed and water freely available.

Type of coverage:

occlusive

Preparation of test site:

other: clipped and abraded

Vehicle:

unchanged (no vehicle)

Controls:

no

Amount / concentration applied:

126, 158, 200, 251, 316 and 398 mg/kg body weight

Duration of treatment / exposure:

24 h

Observation period:

14 days

Number of animals:

4 per dose group (2 abraded, 2 with intact skin)

Details on study design:

TREATMENT:
Prior to application of the test substance, one half (2) of the rabbits in each group received a series of abrasions with a sterile needle through the epidermis and two rabbits remained intact. The trunk of each rabbit was wrapped with an impervious material taped in place to form a reservoir over the test skin site. The test material was introduced onto the skin at the dose levels indicated below. During application, the rabbits were kept in a restrainer device and later were fitted with a collar to prevent ingesting the material and returned to their respective cage. Wrappings were removed after 24 hours and the test material was gently washed from the back.

OBSERVATIONS:
The rabbits were observed immediately after treatment, at four hours and daily for 14 days for signs of illness and mortality. Dermal reactions were scored daily for erythema and edema. Body weight was recorded at dosing and at 14 days for the survivors. Animals found dead or euthanized at 14 days were subjected to a macroscopic examination of the viscera.

Irritation parameter:

edema score

Basis:

mean

Time point:

24/48/72 h

Remarks on result:

no indication of irritation

Irritation parameter:

erythema score

Basis:

mean

Time point:

24/48/72 h

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

There were no significant signs of dermal irritation.

Other effects:

The LD50 was calculated to be 210 mg/kg body weight with 95% confidence limits of 156-284 mg/kg. No macroscopic changes were noted at necropsy, which could be attributed to the test material at the highest dose levels. All four rabbits showed uneven pitted appearance in the cortex of the kidney at the lowest level. No effects on body weight gain during the entire course of the study.

Interpretation of results:

GHS criteria not met

Conclusions:

In an acute dermal toxicity study conducted in rabbits, no signs of dermal irritation were observed.

Executive summary:

In an acute dermal toxicity study similar to OECD 402, dimethylamine-borane was applied occlusively to the abraded and intact skin of New Zealand White rabbits at concentrations of 126, 158, 200, 251, 316 and 398 mg/kg body weight for 24 hours. Afterwards, the test material was washed off. The animals were observed for 14 days for clinical signs and mortality including a daily scoring for erythema and edema. Body weight was recorded at dosing and at the end of the study for the survivors. Animals found dead or euthanised at 14 days were subjected to a macroscopic examination of the viscera. The LD50 was calculated to be 210 mg/kg body weight with 95% confidence limits of 156 - 284 mg/kg bw. No macroscopic changes were noted at necropsy, which could be attributed to the test material at the highest dose levels. All four rabbits showed uneven pitted appearance in the cortex of the kidney at the lowest level. All surviving animals gained body weight during the course of the study. There were no significant signs of dermal irritation noted throughout the study.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017-02-24 to 2017-04-25

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: +5 °C, protected from light

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Final preparation of a solid: The test item was suspended with physiological saline 0.9% NaCl (Alleman Pharma, lot no. 609709, expiry date: 2017-03-03) to give a 20% concentration, afterwards it was vortexed prior to administration.

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES:
- The assay uses isolated corneas obtained as a by-product from animals freshly slaughtered at the abattoir A. Moksel AG, Buchloe, Germany. On the test day, fresh eyes were collected from the slaughterhouse and were transported in HBSS containing Pen/Strep on ice to the laboratories.

Vehicle:

physiological saline

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 750 µL
- Concentration (if solution): 20%

VEHICLE
- Amount(s) applied (volume or weight with unit): 750 µL
- Concentration (if solution): physiological saline 0.9% NaCl
- Lot/batch no. (if required): Alleman Pharma, lot no. 609709

Duration of treatment / exposure:

4 hours ± 5 minutes at 32 ± 1 °C

Duration of post- treatment incubation (in vitro):

The optical density at 490 nm was measured upon 90 minutes of incubation with fluorescein after exposure to the test item by using a spectrophotometer.

Number of animals or in vitro replicates:

Three corneas each for the test item, negative control (physiological saline 0.9% NaCl) and positive control (imidazole 20% in physiological saline 0.9% NaCl).

Details on study design:

SELECTION AND PREPARATION OF CORNEAS
- The assay uses isolated corneas obtained as a by-product from animals freshly slaughtered at the abattoir A. Moksel AG, Buchloe, Germany.
On the test day, fresh eyes were collected from the slaughterhouse and were transported in HBSS containing Pen/Strep on ice to the laboratories. Immediately after arrival of the eyes, cornea preparation was initiated.
The eyes were carefully examined for defects and any defective eyes were discarded.
The tissue surrounding the eyeball was carefully pulled away and the cornea was excised leaving a 2 to 3 mm rim of sclera. The isolated corneas were stored in a petri dish containing HBSS. Before the corneas were mounted in corneal holders (Duratec GmbH) with the endothelial side against the O-ring of the posterior chamber, they had been visually examined for defects and any defective cornea had been discarded. The anterior chamber was then positioned on top of the cornea and tightened with screws. The chambers of the corneal holder were then filled with RPMI (without phenol red) containing 1% FBS and 2 mM L-glutamine (complete RPMI). The posterior chamber was always filled first. The corneas were incubated for one hour at 32  1 °C.

TREATMENT METHOD: closed chamber

REMOVAL OF TEST SUBSTANCE
- Number of washing steps after exposure period: Washing with MEM containing phenol red until the medium was free of test substance (at least 3 times), then completely rinsed with RPMI without phenol red.


METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: After the equilibration period, the medium was removed from both chambers and replaced with fresh complete RPMI. An initial measurement was performed on each of the corneas using the opacitometer. Three corneas with illuminance readings approximately equivalent to the median illuminance of all corneas were selected as negative-control corneas. The illuminance of each cornea was read and recorded. Only corneas that had an initial illuminance reading I > I0/1.1651 lux were used for the assay. The medium was removed from the anterior chamber and replaced with the test item or control. After the 4 hours incubation period and subsequent washing the anterior chamber was refilled with complete RPMI and an illuminance measurement was performed. Also, each cornea was observed visually and pertinent observations were recorded. After the illuminance measurement was performed, the medium was removed from both chambers of the holder.
- Corneal permeability: passage of sodium fluorescein dye measured with the aid of UV/VIS spectrophotometry (OD490):
After the illuminance measurement was performed, the medium was removed from both chambers of the holder. The posterior chamber was refilled with fresh complete RPMI. 1 mL of a 5 mg/mL sodium fluorescein solution was added to the anterior chamber and the corneas were incubated for 90 minutes at 32 +/-1 °C. Then the medium from the posterior chamber was removed and its optical density at 490 nm (OD490) was determined, using a spectrophotometer (Jenway 6405 UV/VIS).

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)
IVIS = mean opacity value + (15 x mean permeability OD490 value)
The IVIS cut-off values for identifying test substances as inducing serious eye damage (UN GHS Category 1) and test substances not requiring classification for eye irritation or serious eye damage (UN GHS No Category) are given in Table 1 (see "Any other information on materials and methods").
An identification of test substances that should be classified as irritating to eyes (UN GHS Category 2 or Category 2A) or test substances that should be classified as mildly irritating to eyes (UN GHS Category 2B) cannot be made.
For this purpose further testing with another suitable method is required

Irritation parameter:

in vitro irritation score

Run / experiment:

3

Value:

101.13

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

positive indication of irritation

Other effects / acceptance of results:

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The negative control responses should result in opacity and permeability values that are less than the established upper limits for background bovine corneas treated with the respective negative control.
- Acceptance criteria met for positive control: The in vitro irritation score obtained with the positive control fell within the two standard deviations of the current historical mean and therefore this assay is considered to be valid

Table 2: Opacity

Cornea No.	Test Item	Initial Opacity	Final Opacity	Change of Opacity Value	Corrected Opacity Value
1	Negative Control Physiological Saline	2.01	2.46	0.54
2		2.23	2.62	0.38
3		1.75	3.32	1.58
MV		2.00	2.80	0.80
4	Positive Control 20% Imidazol in physiological saline	2.54	105.91	103.36	102.56
5		2.54	101.87	99.33	98.52
6		2.66	84.64	81.98	81.18
MV		2.58	97.47	94.89	94.09
7	Test Item dimethylamine-borane (20% in physiological saline)	1.60	24.25	22.65	21.84
8		0.32	60.75	60.43	59.63
9		0.88	42.62	41.74	40.94
MV		0.93	42.54	41.61	40.80

Table 3: Permeabilty

Cornea No.	Test Item	OD490	Corrected OD490
1	Negative Control Physiological Saline	0,007
2		0,009
3		0,018
MV		0,011
4	Positive Control 20% Imidazol in physiological saline	3,060	3,049
5		4,660	4,649
6		4,235	4,224
MV		3,985	3,974
7	Test Item dimethylamine-borane (20% in physiological saline)	0,000	-0,011
8		5,610	5,599
9		6,490	6,479
MV		4,033	4,022

Table 4: In vitro Irritation Score (IVIS)

Cornea No.	Test Item	Corrected Opacity Value	Corrected OD490 Value	IVIS
1	Negative Control Physiological Saline	0.45	0.007	0.97
2		0.38	0.009
3		1.58	0.018
MV		0.80	0.011
4	Positive Control 20% Imidazol in physiological saline	102.56	3.049	153.69
5		98.52	4.649
6		81.18	4.224
MV		94.09	3.974
7	Test Item dimethylamine-borane (20% in physiological saline)	21.84	-0.011	101.13
8		59.63	5.599
9		40.94	6.479
MV		40.80	4.022

 

Interpretation of results:

Category 1 (irreversible effects on the eye) based on GHS criteria

Conclusions:

In conclusion, based on the mean in vitro irritation score of 101.13 obtained in the bovine corneal opacity and permeability assay (OECD 437), the target substance is classified for irreversible effects on the eye (Category 1, H318).

Executive summary:

The eye irritation potential of dimethylamine-borane (DMAB) (> 99% purity) was investigated in the bovine corneal opacity and permeability assay (OECD 437). The test item was suspended with physiological saline 0.9 % NaCl to gain a 20 % concentration. A mean in vitro irritation score (IVIS) of 101.13 was determined. The positive control induced the appropriate responses, indicating the validity of the assay. Based on the results, classification for irreversible effects on the eye (Category 1, H318) is warranted.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irreversible damage)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

In an acute dermal toxicity study conducted similar to OECD test guideline 402, dimethylamine borane was applied to the abraded and intact skin of New Zealand White rabbits at concentrations of 126, 158, 200, 251, 316 and 398 mg/kg body weight for 24 h. The animals were observed for a total of 14 days for clinical signs and mortality including a daily scoring for erythema and edema. There were no significant signs of dermal irritation noted throughout the study.

In a second acute dermal toxicity study conducted similar to OECD test guideline 402, dimethylamine borane was applied to the abraded and intact skin of New Zealand White rabbits at concentrations of 200, 282 and 398 mg/kg body weight for 24 h. The animals were observed for a total of 14 days for clinical signs and mortality including a daily scoring for erythema and edema. There were no significant signs of dermal irritation noted throughout the study.

The eye irritation potential of dimethylamine-borane (DMAB) (> 99% purity) was investigated in the bovine corneal opacity and permeability assay (OECD 437). The test item was suspended with physiological saline 0.9 % NaCl to gain a 20 % concentration. A mean in vitro irritation score of 101.13 was determined. The positive control induced the appropriate responses, indicating the validity of the assay. Based on the results, classification for irreversible effects on the eye (Category 1, H318) is warranted.

Additional information for both, eye and skin irritation potential, is available from a review on the toxicity of boranes (Roush, 1959) stating that a 10% aqueous solution is slightly irritating to both rabbit eye and skin. As no information on the study design was given and the experiment was performed with a 10 % solution, this information will not be considered for the hazard assessment.  

Justification for classification or non-classification

Based on the results of two acute dermal toxicity studies conducted similar to OECD 402, the target substance does not warrant classification for skin irritation. Based on the results of an in vitro eye irritation study conducted according to OECD 437, classification for irreversible effects on the eye (Category 1, H318) is warranted.