Chlorbutanol

Endpoint summary

Description of key information

Skin irritation

The performed in-vitro test according to OECD guideline 439 addresses the human health endpoint skin irritation shows that the mean value of relative tissue viability was reduced to 3.8 %. This value is below the threshold for skin irritation potential (50%). Test items that induce values below the threshold of 50% are considered at least irritant to skin.

One in vivo supporting study on rabbits concluded that Chlorobutanol is a mild skin irritant, therefore the substance is classified as skin irritatant Cat. 2.

Eye irritation:

In the performed in vitro test according to the OECD Guideline 492, the EpiOcularTMEye Irritation Test, the mean value of relative tissue viability was 1.6 %.  This value is well below the threshold for eye irritation potential (≤ 60%). Test items that induce values below the threshold are considered either eye irritant or inducing serious eye damage. In vivo supporting studies on rabbits concluded that Chlorobutanol is a mild eye irritant, therefore the substance is classified as eye irritant Cat. 2.

This classification is also supported by the use in eye drops, usually in 0.5% concentrations, whithout producing clinically significant ocular disturbance even though applied serveral time per day for several years.

However, certain precautions in use of Chlorobutanol in eyedrops are necessary. Hydrolysis destroys Chlorobutanol in alkaline solution. It is usually employed in solutions of pH 5 or lower, which is stable for at least many months. If unbuffered solutions are autoclaved, or allowed to stand at room temperature for several years, the pH may drop as low as 2.4. Furthermore, although fresh solutions containing 0.5% Chlorobutanol are quite satisfactory and innoculous for the brief applications involved in the ordinary use of eyedrops, they appear to be unsuited for prolonged bathing of the outer surface of the cornea.

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:

supporting study

Study period:

not specified

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Qualifier:

no guideline available

Principles of method if other than guideline:

review data, access to original study not available

GLP compliance:

not specified

Species:

rabbit

Strain:

not specified

Type of coverage:

not specified

Preparation of test site:

not specified

Vehicle:

not specified

Controls:

not specified

Irritation parameter:

other: MLD

Basis:

other: not specified

Time point:

other: not specified

Reversibility:

not specified

Remarks on result:

probability of mild irritation

Remarks:

probability of mild irritation

Irritant / corrosive response data:

skn-rbt 850 µg MLD

Interpretation of results:

Category 3 (mild irritant) based on GHS criteria

Conclusions:

results of a skin irritation test on rabbits available
mild irritation has been observed resulting in a category 3 classification

Reference 2

Endpoint:

skin irritation: in vitro / ex vivo

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 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: he EpiDermTM tissue consists of human-derived epidermal keratinocytes which have been cultured to form a multi-layered, highly differentiated model of the human epidermis

Details on animal used as source of test system:

The EpiDermTM tissue consists of human-derived epidermal keratinocytes which have been cultured to form a multi-layered, highly differentiated model of the human epidermis. It consists of organized basal, spinous and granular layers, and a multi-layered stratum corneum containing intercellular lamellar lipid layers representing main lipid classes analogous to those found in vivo. The EpiDermTM tissues are cultured on specially prepared cell culture inserts.

Vehicle:

unchanged (no vehicle)

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

Tissue Amount
1 25.8 mg
2 25.5 mg
3 24.7 mg

Duration of treatment / exposure:

1 hour

Duration of post-treatment incubation (if applicable):

23 hours 25 minutes

Number of replicates:

3

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Mean

Value:

3.8

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Designation	Chlorobutanol Hemihydrate	Positive Control
% Tissue viability (tissue 1)	4.1%	3.9%
% Tissue viability (tissue 2)	3.7%	4.3%
% Tissue viability (tissue 3)	3.6%	4.3%
% Tissue viability (mean)	3.8%	4.1%
± SD of mean tissue viability (%)	0.3%	0.2%

Interpretation of results:

Category 2 (irritant) based on GHS criteria

Conclusions:

The test item Chlorobutanol Hemihydrate is considered as at least irritant to skin.
After the treatment, the mean value of relative tissue viability was reduced to 3.8%. This value is below the threshold for skin irritation (50%).
The optical density of the negative control was well within the required acceptability criterion of 0.8 ≤ mean OD ≤ 2.8.
The positive control has met the acceptance criterion too, for thus ensuring the validity of the test system.
Variation within replicates was within the accepted range for negative control, positive control and test item (required: ≤ 18%).
For these reasons, the result of the test is considered valid.

Executive summary:

One valid experiment was performed.

Three tissues of the human skin model EpiDerm^TMwere treated with Chlorobutanol Hemihydrate for 60 minutes.

The test item was applied directly to each tissue and spread to match the tissue size (0.63 cm²; as indicated by the supplier).

DPBS-buffer was used as negative control and 5% SDS solution was used as positive control.

After treatment with the negative control, the mean absorbance value was within the required acceptability criterion of 0.8 ≤ mean OD ≤ 2.8, OD was 1.2. The positive control showed clear irritating effects. The mean value of relative tissue viability was reduced to 4.1 % (required:<20%).

The variation within the tissue replicates of negative control, positive control and test item was acceptable (required: ≤ 18%). 

 

After the treatment with the test item, the mean value of relative tissue viability was reduced to 3.8 %. This value is below the threshold for skin irritation potential (50%). Test items that induce values below the threshold of 50% are considered at least irritant to skin. 

 

The OECD guideline 439 addresses the human health endpoint skin irritation and does not allow discrimination between skin irritation (category 2) and skin corrosion (category 1). For these purposes, further testing with other suitable test methods is required.

 

Therefore, Chlorobutanol Hemihydrate is considered at least irritant to skin in the Reconstructed human Epidermis (RhE) Test Method.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation: in vitro / ex vivo

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 492 (Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

human

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

Replicate Amount
Tissue 1 50.2 mg
Tissue 2 51.0 mg

Duration of treatment / exposure:

6 hours

Duration of post- treatment incubation (in vitro):

18 hours

Number of animals or in vitro replicates:

2

Irritation parameter:

in vitro irritation score

Remarks:

% Viability Mean

Run / experiment:

Mean

Value:

1.6

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

Criterion Demanded Found
Mean OD of negative control > 0.8 and < 2.5 1.7
% mean relative viability of positive control < 50% of negative control 40.0%
Variation within replicates < 20% 0.1% (negative control)
4.1% (positive control)
0.1% (test item)

Designation	Positive Control	Chlorobutanol Hemihydrate
% Viability (Tissue 1)	37.9%	1.6%
% Viability (Tissue 2)	42.0%	1.6%
% Viability Mean	40.0%	1.6%

Interpretation of results:

Category 2 (irritating to eyes) based on GHS criteria

Conclusions:

Under the conditions of the test, Chlorobutanol Hemihydrate is considered either eye irritant or inducing serious eye damage in the EpiOcularTM Eye Irritation Test.
After treatment with the test item, the mean value of relative tissue viability was reduced to 1.6 %. This value is well below the threshold for eye irritation potential (≤ 60%).
All validity criteria were met. The criterion for optical density of the negative control was fulfilled: The OD value was 1.7 (> 0.8 and < 2.5).
The positive control induced a decrease in tissue viability as compared to the negative control to 40.0 %. Variation within the replicates was acceptable (< 20%).
For these reasons, the result of the test is considered valid.

Executive summary:

One valid experiment was performed.

The test item Chlorobutanol Hemihydrate was applied to a three-dimensional human cornea tissue model in duplicate for an exposure time of 6 hours.

After treatment, the respective substance was rinsed from the tissue; then, cell viability of the tissues was evaluated by addition of MTT, which can be reduced to formazan. The formazan production was evaluated by measuring the optical density (OD) of the resulting solution.

Demineralised water was used as negative control and methyl acetate was used as positive control.

The controls showed the following results: After treatment with the negative control, the absorbance values were within the required acceptability criterion of mean OD>0.8 and < 2.5, OD was 1.7. The positive control showed clear eye irritating effects, the mean value of the relative tissue viability was 40.0 % (< 50%). 

Variation within tissue replicates was acceptable (< 20%).

 

After treatment with the test item, the mean value of relative tissue viability was 1.6 %. 

This value is well below the threshold for eye irritation potential (≤ 60%). Test items that induce values below the threshold are considered either eye irritant or inducing serious eye damage.

According to the OECD Guideline 492, the EpiOcular^TMEye Irritation Test does not allow discrimination between eye irritation/reversible effects on the eye (Category 2) and serious eye damage/irreversible effects on the eye (Category 1). For these purposes, further testing with other suitable test methods is required.

 

Reference 2

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Qualifier:

no guideline available

Principles of method if other than guideline:

review data, no access to original study report

GLP compliance:

not specified

Species:

rabbit

Strain:

not specified

Vehicle:

not specified

Irritation parameter:

other: MLD

Basis:

other: not specified

Time point:

other: 30S

Reversibility:

not specified

Remarks on result:

probability of mild irritation

Interpretation of results:

Category 2B (mildly irritating to eyes) based on GHS criteria

Conclusions:

eye-rbt 9180 µg/30S MLD

Executive summary:

mild eye irritation observed after application of chlorobutanol. No further information available.

supporting studies on humans with ophthalmic preservative result in slight but reversible irritation.

Reference 3

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

other: review data

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Chlorobutanol has been widely used as a detergent preservative in ophthalmic preservatives. observation from ophthalmic administration are reported and are considered as supporting

Qualifier:

no guideline available

Principles of method if other than guideline:

review data, no access to original study available

GLP compliance:

not specified

Species:

rabbit

Strain:

not specified

Vehicle:

other: aqueous solution containing 1.4% sodium chloride

Controls:

yes, concurrent vehicle

Amount / concentration applied:

0.4%

Duration of treatment / exposure:

several minutes

Irritation parameter:

other: visual fogging, haloes around lights, and a foreign body sensation

Basis:

other:

Time point:

other: several minutes

Reversibility:

fully reversible within: 1-2 days

Remarks on result:

probability of mild irritation

Irritant / corrosive response data:

Gonioscopy solutions containing 0.4% Chlorobutanol, applied under the contact lens, have been associated with visual fogging, haloes around lights, and a foreign body sensation, which intesifies over several hours and recovers spontaneously in 1 or 2 days

Other effects:

Chlorobutanol has been widely employed as a preservative in eye drops, usually in 0.5% concentrations, whithout producing clinically significant ocular disturbance. However, certain precautions in use of Chlorobutanol in eyedrops are necessary. hydrolysis destroys Chlorobutanol in alkaline solution. It is usually employed in solutions of pH 5 or lower, which is stable for at least many months. However, if unbuffered solutions are autoclaved, or allowed to stand at room temperature for several years, the pH may drop as low as 2.4. Furthermore, although fresh solutions containing 0.5% Chlorobutanol are quite satisfactory and innoculous for the brief applications inveolved in the ordinary use of eyedrops, they appear to be unsuited for prolonged bathing of the outer surface of the cornea.

Interpretation of results:

study cannot be used for classification

Conclusions:

Chlorobutanol containing solutions applied under contact lens to humans show mild and reversible irritation

Reference 4

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

other: review

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Chlorobutanol has been widely used as a detergent preservative in ophthalmic preservatives. observation from ophthalmic administration are reported and are considered as supporting

Qualifier:

no guideline available

GLP compliance:

not specified

Species:

human

Strain:

not specified

Vehicle:

other: opthalmic solution cotaining polyvinylalcoghol 1.4%

Controls:

yes, concurrent vehicle

Amount / concentration applied:

0.5%

Duration of treatment / exposure:

several hours

Observation period (in vivo):

several ours

Irritation parameter:

other: conjunctival erythema

Basis:

other: 9 of 16

Time point:

other: not specified

Reversibility:

fully reversible

Remarks on result:

probability of mild irritation

Irritant / corrosive response data:

It was suggested that the irritation was due to crystallisation of Chlorobutanol, which is close saturation at this concentration

Interpretation of results:

study cannot be used for classification

Conclusions:

Ophthalmic administration of Chlorobutanol containing solutions to humans show mild and reversible irritation

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Chlorobutanol is considered as skin and eye irritant, since the key in-vitro studies showed the irritant effects on skin and eye and the supporting in vivo studies also showed

that the substance is a mild eye and skin irritant.