Ammonium chloride

Endpoint summary

Description of key information

In two in vivo skin sensitisation studies in guinea pigs comparable to OECD guideline 406 (Hoechst, 1986), no skin sensitising properties of the test item were determined.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Reference 1

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

other: EPA 540/9-82-025

Version / remarks:

November 1982, revised at November 1984

Deviations:

not specified

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Another internationally accepted test method was used.

Species:

guinea pig

Strain:

other: Pirbright-White (Hoe: DHPK (SPFLac))

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Age at study initiation: about 8 weeks
- Mean weight at study initiation: within 20 % of mean weight (Range = -13 % to + 10 %), n = 30
- Housing: 5/cage in Type 4 Macrolon cages
- Diet: ad libitum; "ERKA-Mischfutter Nr. 8300 fuer Meerschweinchen und Kaninchen"
- Water: ad libitum; tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 50 ± 20
- Photoperiod (hrs dark / hrs light): 12/12

Route:

other: intradermal and epicutaneous (occlusive)

Vehicle:

physiological saline

Remarks:

(0.9%)

Concentration / amount:

- 1st Induction: 5%
- Epicutaneous Induction 25 %

Route:

epicutaneous, occlusive

Vehicle:

physiological saline

Remarks:

(0.9%)

Concentration / amount:

- Challenge: 10 %.

No. of animals per dose:

- Treatment group: 20
- Control group: 10
- Satellte group: 5

Details on study design:

JUSTIFICATION FOR CHOICE OF CONCENTRATIONS
- Based on results of range finding test (see below)

RANGE FINDING TESTS (RFT):
Intradermal injection
- No. of animals: 3 females/concentration
- Site/Area: back/2 x 4 cm
- Concentrations: 0.2, 1.0 and 5.%
- Vehicle: physiol. saline (0.9 %)
- Volume injected: 0.1 mL
- Number of injection/concentration: 2
- Evaluation (hr after injection): 24 hour post injection


RANGE FINDING TESTS (RFT):
Epicutanous application
- Rationale: determine the highest concentration of the test substance that causes slight to moderate irritation and for the challenge the maximum non-irritant concentration
- No. of animals: 2 females/test concentration
- Site: left flank
- Area of exposure: 2 x 2 cm
- Type of coverage: occlusive
- Concentrations: 1.0, 10.0 and 25% (25% is technically the maximal achievable concentration)
- Vehicle: physiol. saline (0.9%)
- Volume applied: 0.5 mL applied on a test patch
- Duration: 24 hours
- Evaluation (hr after challenge): 24 hours

MAIN STUDY
A INDUCTION EXPOSURE
Intradermal Injection
- Site/Area: Back (near the shoulders) /2 x 4 cm
- No. of injections/animal: 6 injections
- Test substance group (cranial; 2 injections side by side): 50 % Freund's complete adjuvant (FCA) blended (1:1, v/v) with physio. saline (0.9 %)
- Test substance group (middle; 2 injections side by side): The test article (5 %) in vehicle
- Test substance group (caudal; 2 injections side by side): 50 % Freund's complete adjuvant blended (1:1, v/v) with physio. saline (0.9 %) and 5% test article
- Control group (cranial; 2 injections side by side): 50 % Freund's complete adjuvant (FCA) blended (1:1, v/v) with physio. saline (0.9 %)
- Control group (middle; 2 injections side by side): Vehicle
- Control group (caudal; 2 injections side by side): 50 % Freund's complete adjuvant (FCA) blended (1:1, v/v) with physio. saline (0.9 %)
- Volume per injection: 0.1 mL
- Evaluation (hr after injection): from 24 hour till day 7

Epicutaneous induction exposure
- Time schedule: 8 days after intradermal injection
- Site: same as intradermal injection
- Area of application: 2 x 4 cm (cellulose patches)
- Concentrations: 25 % (test group), 0.9 % Physiol. Saline (control group)
- Volume applied: 0.5 mL/animal
- Type of coverage: occlusive (impemeable foil with adhesive fleece)
- Duration: 48 hours
- Evaluation (hr after challenge): directly after removal of patch

B. CHALLENGE EXPOSURE (all animals)
- Time schedule: 13 days after termination of induction exposures
- Site: left flank
- Area of application: 2 x 2 cm (cellulose patches)
- Concentrations: undiluted
- Volume applied: 0.5 mL/animal
- Type of coverage: occlusive (impemeable foil with adhesive fleece)
- Duration: 24 hours
- Evaluation (hr after challenge): 24 and 48 hours after termination of exposure

SCORING SYSTEM: OECD Draize system

Positive control substance(s):

no

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

10 %

No. with + reactions:

2

Total no. in group:

20

Clinical observations:

No clinical signs of toxicity. Skin indicated very slight, hardly perceptible erythema.

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

10 %

No. with + reactions:

2

Total no. in group:

20

Clinical observations:

No clinical signs of toxicity. Skin indicated very slight, hardly perceptible erythema.

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

10%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

No clinical signs of toxicity

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

10%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

No clinical signs of toxicity

Group:

positive control

Remarks on result:

not measured/tested

 RANGE FINDING TESTS (RFT):

Intradermal injection:

0.2 % and 1.0 % of the test substance formulation caused slight edema at the injection sites. 5.0 % of the test substance formulation caused slight erythema and edema at the injection sites. 5.0 % was used in the main study based on the findings decribed above.

 

Epicutaneous induction:

No effects were seen after dermal application of the test substance formulations (1.0, 10.0, and 25.0 % in physiol. Saline (0.9 %). A maximal non-irritating concentration of 25.0 % is thus attained. However, in combination with Freud’s adjuvant, the maximal non irritating concentration can be reduced. For this reason, the satellite animals (5 animals) which had been pretreated with Freud’s adjuvant were exposed to 25.0 %. The treated skin of 3 of the 5 animals exhibited very slight erythema. In addition, the skin at the application site was dry, cracked and scaly. Based on this finding, the maximal non irritating concentration was set at 10.0 % for them main study.

 

MAIN STUDY

Clinical signs of toxicity

The treated animals displayed no signs of intoxication through out the entire study duration 

 

Induction

Intradermal injection with Freud’s adjuvant (with and without the testsubstance) led in the control and the treated animals as well as in animals of the preliminary dose finding experiment to well defined erythema and slight edema. Very slight to slight edema appeared at the application sites injected with the test substance in physiol. Saline (0.9 %). In addition scab formation was noted in all animals. The body weight gain of treated animals was not affected.

 

 

Challenge:

Only two of ten animals (10.0 %) treated with the test substance formulation had a positive reation. A very slight barely noticeable erythema was seen at the application sites of these animals. The remaining animals showed no irritation effects.

Interpretation of results:

GHS criteria not met

Reference 2

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1987

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Qualifier:

according to guideline

Guideline:

other: EPA 540/9-82-025

Deviations:

not specified

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Another internationally accepted test method was used.

Species:

guinea pig

Strain:

other: Pribright-White

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Age at study initiation: About 8 weeks
- Weight at study initiation: Mean 240 g

Route:

intradermal and epicutaneous

Vehicle:

other: 0.9 % NaCl Solution

Concentration / amount:

1st application: Induction 5 % intracutaneous
2nd application: Induction 25 % occlusive epicutaneous

Route:

epicutaneous, occlusive

Vehicle:

other: 0.9 % NaCl Solution

Concentration / amount:

3rd application: Challenge 10 % occlusive epicutaneous

No. of animals per dose:

20

Details on study design:

1st application: Induction 5 % intracutaneous
2nd application: Induction 25 % occlusive epicutaneous
3rd application: Challenge 10 % occlusive epicutaneous

Day 1 : Intradermal induction exposure (Injection). The injection sites were not covered.    
1-7: The application area was investigated.    
9  : Dermal induction exposure. 0.5 mL of the test substance preparation were applied to a cellulose patch of 2x4 cm. This patch covered the area of the intradermal injection sites. An occlusive dressing with impermeable foil and an elastic bandage sealed the application site for 48 hours.    
11 : Removal of the occlusive dressing. Recording of the irritation.    
22 : Dermal challenge exposure 0.5 mL of test substance preparation were applied to a cellulose patch and placed onto the clipped skin of the flank. An occlusive dressing with impermeable foil and an elastic bandage sealed the application site for 24 hours.    
23 : Removal of the occlusive dressing   
24-25: Assessment of the skin

Positive control substance(s):

not specified

Reading:

other: induction

Group:

test chemical

Clinical observations:

very slight edema

Reading:

other: challenge

Hours after challenge:

24

Group:

test chemical

No. with + reactions:

2

Total no. in group:

20

Clinical observations:

slight erythema

1) The treated animals did not show any signs of toxicity throughout the study period.
2) Induction: Very slight to slight edema were observed in the treatment group.
3) Challenge: 24 and 48 hours after removal of the occlusive dressing, a total of 2 animals (in 20) of the treatment group showed very slight, hardly perceptible erythema.

Ten percent of the animals of treatment group demonstrated a positive reaction after the challenge exposure (the criteria: the limit value of 30 percent).

Interpretation of results:

GHS criteria not met

Reference 3

Endpoint:

skin sensitisation: in vitro

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

In a dermal sensitization study with ammonium chloride (99.1% pure), 8 weeks old guinea pigs [Pirbright-White (Hoe: DHPK (SPFLac))] were tested according to EPA 540/9-82-025 (November 1982, revised at November 1984), a method comparable to OECD TG 406 (Hoechst, 1986). The control group contained 20 animals and the test group contained 10 animals. A satellite group containing 5 animals was also included. Intradermal inductions were performed with 5% ammonium chloride in 0.9 % NaCl mixed with Freund's complete adjuvant (FCA). 8 days later, topical inductions were carried out under occlusive conditions for 48 hours with 25 % ammonium chloride in 0.9 % NaCl. Following a 13 day break, challenge exposures were performed epicutaneously (occlusive, 24 hours) with 10 % ammonium chloride in 0.9 % NaCl. 24 and 48 hours post termination of exposure, two of ten animals (10 %) in the treatment group had a positive reaction. A very slight barely noticeable erythema was seen at the application sites of these animals. The remaining animals showed no irritation effects. No sign of skin irritation was seen in control animals. A positive control testing was not included In a supporting skin sensitisation test with guinea pigs (maximisation test), ammonium chloride also appeared to be non skin sensitising (Bayer Crop, 1987). The treated animals did not show signs of toxicity throughout the study period. During induction very slight to slight edema were observed in the treatment group. During challenge 24 and 48 hours after removal of the occlusive dressing, a total of 2 animals (in 20) of the treatment group showed very slight, hardly perceptible erythema. Ten percent of the animals of the treatment group demonstrated a positive reaction after the challenge exposure (the criteria: the limit value of 30 percent).

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Skin sensitization

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008 (CLP). As a result the substance is considered not to be classified for skin sensitisation (UN GHS No Category) under Regulation (EC) No 1272/2008, as amended for the tenth time in Regulation (EU) No 2017/776.