Nonanoic acid

Endpoint summary

Description of key information

Testing for irritation and corrosion of the skin in rabbits in a reliable study (Celanese/RCC, 2001, RL 1) revealed the test substance to be skin irritant. No reliable tests investigating the effects on the eyes are available. Following the harmonised classification for n-nonanoic acid a classification as eye irritant is recommended. According to Commission Regulation (EU) 2016/863 amending Annexes VII and VIII to Regulation (EC) No 1907/2006, Annex VII and VIII, Nr. 8.2, Column 2, eye irritation studies are not required.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 18 SEP 2001 to 16 OCT 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

GLP compliance:

yes (incl. QA statement)

Remarks:

in accordance with SN EN 45001 (Swiss legislation)

Species:

rabbit

Strain:

New Zealand White

Type of coverage:

semiocclusive

Preparation of test site:

shaved

Vehicle:

unchanged (no vehicle)

Controls:

no

Amount / concentration applied:

undiluted

Duration of treatment / exposure:

4 hour(s)

Observation period:

immediately after end of exposure, 24, 48 and 72 h and 7, 10 and 14 days post exposure

Number of animals:

3

Details on study design:

SCORING SYSTEM: according to guideline OECD 404F

Irritation parameter:

erythema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

fully reversible within: 14 days

Irritation parameter:

erythema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

3.33

Max. score:

4

Reversibility:

fully reversible within: 14 days

Irritation parameter:

erythema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

2.33

Max. score:

4

Reversibility:

fully reversible within: 14 days

Irritation parameter:

edema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

2

Max. score:

4

Reversibility:

fully reversible within: 14 days

Irritation parameter:

edema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

2.33

Max. score:

4

Reversibility:

fully reversible within: 14 days

Irritation parameter:

edema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

2

Max. score:

4

Reversibility:

fully reversible within: 14 days

After 1 h post-exposure: Well-defined erythema (score 2/2/2) and very slight to slight edema (score 2/2/1)

After 24 h: Moderate to severe erythema (score 3/3/2) and slight edema(score 2/2/2).
After 48 h: Moderate to severe erythema (score 3/3/3)and slight edema (score 2/2/2)

After 72 h: Well-defined to severe erythema (score 3/4/2)and slight edema (score 2/3/2), scaling in one animal

After 7 days: Very slight to well-defined erythema (score 2/2/1)and very slight edema (score 1/1/1), scaling in all animals

After 10 days: Very slight erythema (score 1/1/1)and very slight edema (score 1/0/1), scaling in all animal

After 14 days: no edema, no erythema, scaling in 2 of 3 animals

The mean scores across the 3 animals were 
       Erythema     Edema
===============================
24 h     2.7           2
48 h     3             2
72 h     3             2.3
================================

The mean scores across three scoring times (24, 48, 72 h) for the individual animals were 3.00, 3.33,  and 2.33 (erythema/eschar) and 2.00, 2.33, and 2.00 (edema),  respectively. Well-defined to moderate/severe erythema in all animals for  72 h, and very slight to moderate edema for up to 10 d, all effects disappearing 14 d after treatment.  

No corrosive effects were noted.

In a pre-study with exposure for 3 min and 1 h in one animal, skin  reactions appeared in the form of well-defined to moderate erythema  within one to 24 h p.a. and were still visible after 7d.

Interpretation of results:

Category 2 (irritant) based on GHS criteria

Conclusions:

Under the conditions of this study, the test substance was irritating.

Executive summary:

3 New Zealand white rabbits were dermally exposed to the undiluted test substance (purity: 97.27%; 0.5 mL fluid were soaked into 6 cm² gauze patch) according to guidelines EU B4 and OECD 404. After 4 h of semiocclusive exposure, the test substance caused well-defined to severe erythema (mean score: 2.89) and very slight to moderate edema (mean score: 2.11). After 72 h, one animal out of three showed scaling of the treated area, after 7 and 10 days all animals and after 14 days 2 of 3 animals. The test item thus revealed irritating effects (Celanese/RCC, 2001).

This study was judged to be reliable (RL1).

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vitro / ex vivo

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study does not need to be conducted because the substance is classified as skin irritation and the available information indicates that it should be classified as eye irritation (Category 2)

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation

Skin irritancy of n-nonanoic acid has been investigated in several guideline studies. In the OECD 404 guideline study performed by Celanese/RCC (2001) three New Zealand white rabbits were dermally exposed to the undiluted test substance (purity: 97.27%; 0.5 mL fluid were soaked into 6 cm² gauze patch). After 4 h of semiocclusive exposure the test substance caused well-defined to severe erythema (mean score: 2.89) and very slight to moderate edema (mean score: 2.11). After 72 h one animal out of three showed scaling of the treated area, after 7 and 10 days all animals and after 14 days 2 of 3 animals. The test item thus revealed irritating effects (Celanese/RCC, 2001). This reliable study with the neat test substance has been selected as key study.

Another study with n-nonanoic acid in rabbits observed skin effects pointing to corrosive properties of n-nonanoic acid. In the study performed by Unichema/Notox (1984) three New Zealand white rabbits were dermally exposed to the undiluted test substance according to guideline OECD 404 (n-nonanoic acid, purity >90%). After 4 h of semiocclusive exposure, the test substance caused severe erythema and eschar formation (mean score 24 - 48 -72 h: 3.22) and moderate edema (mean score 24 - 48 -72 h: 1.56). Primary dermal irritation index was 4. One hour after exposure, the exposed area was light-brown stained. After 48 h, the skin became necrotic with eschar formation in all animals. At the end of the 14-day observation period the skin had almost returned to normal, though with variable hair growth.

Whereas the key study indicates that n-nonanoic acid causes skin irritation the other guideline study points to possible corrosive effects. Effects observed in the study performed by Unichema/Notox (1984) at the end of the 14-day observation period are only partially indicative of corrosive effects: only variable hair growth was observed, but no other signs of corrosion like discoloration and scars. These different outcomes of the two guideline in vivo tests for skin irritancy indicate that the evidence for skin corrosion of n-nonanoic acid is borderline and not clear cut. A further test with the undiluted test item (EMFAC 1202 Pelargonic acid) for irritation of the skin in rabbits, which is only available as short abstract, produced dermal irritation increasing with dosage (Emery, 1969). The CLH-report mentions another in vivo OECD 404 guideline study (Otterdijk, 2001) with rabbits which revealed skin irritation but not skin corrosion with n-nonanoic acid, and thereby supports the findings of the key study (CLH, 2011).

Additionally, in the key study for acute dermal toxicity (Notox, 2001) the following skin reactions were observed after 24 h application of n-nonanoic acid (purity: 93%): general erythema, scales and scabs in all animals, grades 1 and 2 (slight, moderate) prevailed. In 6 of 10 animals, skin reactions persisted until the end of the observation period on day 15. But no skin corrosion was observed.

Information available from human patch tests also support the interpretation that n-nonanoic acid causes skin irritation but not skin corrosion: Testing for irritation of the skin in healthy humans (healthy males, 21 to 55 years old) of the test item (purity not mentioned; 5, 10, 20 and 39.9% in 1 -propanol, 48 h exposure, occlusive) resulted in dermal irritation according to the criteria for topical irritants of Wahlberg and Maibach (1980). Testing for irritation of the skin in healthy humans (both sexes, 20 to 40 years old, with normal healthy skin) of the test item (purity not mentioned, 0.16 M in propylene glycol) produced no increased dermal irritation in comparison to the vehicle control (Boelsma, 1997).

Skin irritation but not corrosion was also observed in further human patch tests reported in the CLH-report (CLH, 2011): n-nonanoic acid (0.1 mL) in concentrations at or above 40% caused increased skin thickness after repeated exposure for 15 days in a human volunteer. Another study with eight human volunteers did not identify signs of skin irritation after 24 h exposure to 2.5% or 5% n-nonanoic acid in propanol twenty minutes after patch removal (evaluation by visual irritation score, skin reflectance spectrophotometer, transepidermal water loss and laser Doppler flowmetry).

 

In vitro testing for irritation on human skin equivalent (donor cells are from healthy adults) of the test item (purity not mentioned, 0.16 M in propylene glycol applied onto 0.78 cm² for 6 respectively 24 hours) revealed some elevated pro-inflammatory parameters (IL-1alpha protein level, uPA activity), but other pro-inflammatory parameters (tissue morphology, IL-1 alpha mRNA levels, IL-8 mRNA and protein levels) were left unchanged in comparison to the vehicle control respectively the untreated cultures. In total no conclusion can be drawn for the irritation potential of the test item based on the investigated parameters (Boelsma, 1997). The CLH-report (2011) documented the results of an EpiDerm protocol which indicated that n-nonanoic acid and decanoid acid are at least skin irritant. But, there were no information whether the substances might also be corrosive. Another test presenting results from an ex vivo transcutaneous electrical resistance test with human skin did not indicate skin corrosion of neat decanoic acid.

In an in vitro study (performed in a validation context 2017  after the in vivo study) with the China EpiSkin™ test method (similar to OECD TG 439) several substances were evaluated for their skin damaging potential. Nonanoic acid was tetsted in three separte experiments by application of 10 µL pure substance (>97%) to a human skin model. Results from all three experiments indicated classifcation of the substance in category 2 for skin irritation.

 

In summary, available information on possible skin reactivity of n-nonanoic acid together with supporting information from the comparable substances octanoic and decanoic acid as well as nonanoic acid potassium salt (for details see RAC opinion on the harmonised classification for n-nonanoic acid) indicates that n-nonanoic acid is irritating to skin. There is only borderline evidence for skin corrosion of n-nonanoic acid. Especially the key study and the OECD 404 guideline study mentioned in the CLH-report (2011) which both included a 14 day recovery period support the interpretation that n-nonanoic acid causes skin irritation. This interpretation is supported by the findings of the acute dermal toxicity study and the different human patch tests. However, the patch tests are sometimes limited by the fact that only dilutions but no neat substance was tested. Based on the findings of the key study and the supporting findings from the acute toxicity study and the human patch tests as well as in accordance with the procedure described in the RAC-opinion proposing harmonised classification and labelling at EU level of nonanoic acid (2013) it is concluded that the evidence for skin corrosion of n-nonanoic acid is borderline. Therefore, n-nonanoic acid was classified as skin irritant (Cat. 2, H315) and added to Annex VI (list of harmonised classification and labelling of hazardous substances) in the seventh adaptation to technical processes (ATP) to the CLP regulation.

 

Eye irritation

There is only one study available investigating eye irritating properites of n-nonanoic acid. Data are only available as summary of company data. The test item was applied to the eyes of albino rabbits (no further details mentioned) and produced "marked irritation, including signs as iritis, conjunctivitis and corneal opacity" (Emery, 1969). Due to insufficient reporting this data cannot be used as basis for an evaluation.

 

In three different in in vitro test for eye irritation (Hen´s Egg Test – Chorioallantoic Membrane (HET-CAM), Neutral Red Release Assay (NRR), and Ocular Irritation Assay – EpiOcular™ Tissue Model) the test substance showed irritant properties (Jiriva, 2014). No details of the tests were provided. While the EpiOcular test can only differentiate between effects or no effects on the eyes (effects were observed in the study), the NRR test showed irritant properties according to UN GHS category 2 (no further differentiation possible). In the HET-CAM test corrosivity according to UN GHS category 1 was detected. Overall, the tests point to irritant properties of the substance, but no final conclusions can be drawn based on the limited presentation of details.  

 

A harmonised classification for n-nonanoic acid is in force, in which the substance is classified as eye irritating (Cat. 2, H319) according to CLP. Therefore, further tests on the eye irriting potential of n-nonanoic acid are not necessary.

 

In its opinion from 2013, RAC based the decision on available data on octanoic and decanoic acid, which are also organic acids with a similar structure and comparable pKa values like n-nonanoic acid. Eye irritancy of these two structural related substances has been investigated in two older, non-GLP compliant studies in rabbits (Smyth et al., 1962 and Briggs et al., 1976). Smyth et al. (1962) observed severe damage of the rabbit’s eyes (n=5 animals, severity grade 9) with both substances. However, no information on the test item concentration or the reversibility of the effects was provided. Briggs et al. (1976) described corneal opacity and moderate conjunctivitis of the rabbit’s eyes, which was not reversible during the 72 h observation period. Information on the number of rabbits and the test item concentration is missing and no scoring has been applied.

Additionally, a newer unpublished study with octanoic acid (Leoni and Riedel, 2011) submitted by industry has been considered by RAC in its opinion on n-nonanoic acid (RAC, 2013). The study by Leoni and Riedel (2011) did not confirm the findings of the older studies. Three rabbits were exposed for 24 hours to 0.1 mL of 70% octanoic acid. The animals were observed over 72 h and at 6, 9, and 11 days after dosing. All animals revealed conjunctival redness, chemosis, and discharge with average scores of 1, 1.67 and 2. Two animals revealed lesions of the iris (average score 1 for both animals) and the cornea (average scores 1.33 and 0.67, respectively). The effects were fully reversible at day 9 of the prolonged observation period. The results of this study would result in a classification as irritating to eyes (Cat. 2). Further the RAC-opinion also mentioned additional data from a non-published in vitro test (BCOP, Bovine Corneal Opacity and Permeability) with n-decanoic acid. A 20% dilution of n-decanoic acid did not induce corrosive effects in this in vitro assay (opacity score 16.83 which is far below the score for corrosive effects of > 55.1).

Taking all into account, the RAC-opinion based its decision on all availble studies for the structural related substances n-octanoic acid and n-decanoic acid and concluded that classification as eye irritant (Cat. 2, H319) is warranted for n-nonanoic acid. This proposed classification was later implemented in the Annex VI (list of harmonised classification and labelling of hazardous substances) in the seventh adaptation to technical processes (ATP) to the CLP regulation. Therefore, further testing regarding the eye irritancy of n-nonanoic acid is not necessary.

 

Respiratory tract irritation

As n-nonanoic acid has skin and eye irritation properties it has to be considered whether also respiratory irritation hazard has to be assumed. Some respiratory irritation (red foci) has been described in the Bio/dynamic study (1990) for acute toxicity, however the relevance of the findings was unclear in the absence of a control group. However, this study shows some methodological deficiencies. No respiratory irritation was observed in the well conducted study for acute inhalation toxicity performed by Emery (2014). As the Regulation (EC) No. 1272/2008 supports respective classification only when largely based on human respiratory data and in the absence of such human data it is concluded that no classification for respiratory tract irritation (STOT – single exposure, Cat.3) is required.

Justification for classification or non-classification

Based on the finding of the key study, supported by the data from human patch tests and according to the Regulation (EC) No. 1272/2008 it is concluded that n-nonanoic acid should be classified as skin irritant (Cat. 2).

Based on the available information for the read-across substances octanoic acid and decanoic acid, which provide the basis for the harmonised classification published in Regulation (EC) No. 1272/2008 a classification of n-nonanoic acid as irritating to eyes (Cat. 2) is recommended.

Based on available animal acute inhalation studies, which did not reveal respiratory irritation, and in the absence of relevant human data no classification for respiratory tract irritation (STOT – single exposure, Cat.3) is required.