[1α(E),2β]-1-(2,6,6-trimethylcyclohex-3-en-1-yl)but-2-en-1-one

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

The NOAEL for systemic toxicity in the oral 28-day repeated dose toxicity study in rats was considered to be 85 mg/kg bw /day. The next higher dose level of 340 mg/kg bw/day was a NOAEL for fertility based on the absence of direct adverse effects on male and female reproductive organs.

Effect on fertility: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

340 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

The available study is considered to be adequate for assessment of the fertility effects after repeated oral exposure.

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Study plan:

An extended OECD TG 422 with Delta Damascone will be performed and the final report is expected to be finished Q4 2022/Q1 2023. Once the study is available, the OECD 407 study will be replaced with the extended OECD 422 study using Delta Damascone.

 

Current risk assessment

28-day repeated dose toxicity study

In a study performed in compliance with the Japanese guideline '28 -day Repeated Dose Toxicity Study in Mammalian Species' (Concerning Testing Methods Relating to the New Chemical Substances) and GLP, the test substance was administered to five male and female Crl:CD(SD) rats per group by oral gavage as a dilution in corn oil at dose levels of 0, 21, 85 and 340 mg/kg/day. Additionally, the study included two recovery groups of 5 rats/sex each which were dosed for 28 days with the vehicle (control) or 340 mg/kg of test substance and then left untreated for 14 days. During the dosing period, general clinical observations, detailed clinical observations, function examinations, body weight measurements and food consumption measurements were performed. On the day after the last dosing, urinalyses, blood examinations and pathological examinations were performed after collecting urine and blood samples. The NOAEL for systemic general toxicity was determined to be 85 mg/kg bw (see IUCLID Chapter 7.5.1).

Regarding fertility, macroscopic examination at necropsy and histopathological examination of reproductive organs did not reveal treatment-related changes. The weights of the female reproductive organs (ovaries and uterus) showed no treatment-related changes either. Male reproductive organ weights showed statistically significant changes at the highest dose level (340 mg/kg), namely an increase in relative testes weight, a decrease in the absolute weight of the epididymides and decreases in the absolute and relative weights of the prostate and seminal vesicles. At the end of the recovery period, statistically significant changes in the 340 mg/kg group were limited to an increase in relative testes weight and decreases in the absolute weights of the epididymides and prostate. The magnitude of the organ weight changes at the end of the recovery period was smaller than that at the end of the treatment period. The changes in male reproductive organ weights were considered to be secondary to the lower body weight in the 340 mg/kg group. Based on these results, it can be concluded that the test substance did not exert adverse effects on male or female reproductive organs up to the highest dose tested (340 m/kg bw).

Other studies

In addition to the 28 -day oral toxicity study, in the 90-day oral toxicity study (IUCLID Chapter 7.5.1), in which rats were administered alpha-iso-methylionone (a structural analogue of delta-damascone) up to 500 mg/kg bw (Safepharm Laboratories 2007), no adverse effects on the reproductive organs were observed. Further, a developmental toxicity test in rats (IUCLID Chapter 7.8.2) was carried out with Alpha-iso-methylionone, and no toxicity was observed up to the highest dose tested (Charles River 2005). The maternal and developmental NOAEL were >30 mg/kg bw. Therefore, the test substance is considered not to be a reproductive toxicant and further testing for reproduction toxicity is considered to be unnecessary.

Effects on developmental toxicity

Description of key information

Under the conditions of the oral developmental toxicity study in rats (OECD 414, GLP), the NOAEL for maternal and developmental toxicity was at least 30 mg/kg bw/day.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

Reliable information is available from an OECD guideline 414 developmental study with Alpha-iso-methylionone is available which is sufficiently adequate for this dossier.

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Study plan:

An OECD TG 414 with Delta Damascone is proposed. Once ECHA approves and the study is performed, the studies with the structural analogue will be replaced with the OECD TG 414 study using Delta Damascone. Until then data on Alpha-iso-methylionone is used for hazard assessment.

Current risk assessment

No developmental toxicity studies were available for delta-damascone. However, Article 13 of REACH states that, in case no appropriate animal studies are available for assessment, information should be generated whenever possible by means other than vertebrate animal tests, i.e. applying alternative methods such as in vitro tests, QSARs, grouping and read-across. A developmental toxicity study is available for a structural analogue of Delta-damascone: Alpha-iso-methylionone. First the executive summary of the analogue study is presented and thereafter the read across justification.

Developmental toxicity study on Alpha-iso-methylionone

In a developmental toxicity study performed according to OECD 414 and GLP, Alpha-iso-methylionone, dissolved in corn oil, was administered to 25 rats per group by gavage at gestation days (GD) 7 through 17 at dosages of 0, 3, 10 and 30 mg/kg bw/day (Charles River, 2005). Viabilities, clinical observations, body weights and feed consumption values were recorded. All rats were sacrificed on GD 21, Caesarean-sectioned and examined for the number and distribution of corpora lutea, implantation sites and uterine contents. A gross necropsy of the thoracic, abdominal and pelvic viscera was performed. Fetuses were weighed and examined for gross external, visceral and skeletal alterations and sex. All female rats survived to scheduled sacrifice. All clinical and necropsy observations were considered to be unrelated to treatment. Maternal body weights, body weight gains and absolute and relative feed consumption values were unaffected at dosages as high as 30 mg/kg bw/day. Pregnancy occurred in 21 to 25 rats in each dosage group. Caesarean-sectioning and litter parameters were not affected by dosages of the test article as high as 30 mg/kg bw/day. No fetal alterations occurred that were considered associated with treatment. On the basis of these data, the maternal and developmental NOAEL of alpha-iso-methylionone is greater than 30 mg/kg bw/day. Based on these data on the read-across candidate alpha-iso-methylionone, delta-damascone should not be identified as developmental toxicant.

In a developmental toxicity study performed according to OECD 414 and GLP, Alpha-iso-methylionone, dissolved in corn oil, was administered to 25 rats per group by gavage at gestation days (GD) 7 through 17 at dosages of 0, 3, 10 and 30 mg/kg bw/day. Viabilities, clinical observations, body weights and feed consumption values were recorded. All rats were sacrificed on GD 21, Caesarean-sectioned and examined for the number and distribution of corpora lutea, implantation sites and uterine contents. A gross necropsy of the thoracic, abdominal and pelvic viscera was performed. Fetuses were weighed and examined for gross external, visceral and skeletal alterations and sex. All female rats survived to scheduled sacrifice. All clinical and necropsy observations were considered to be unrelated to treatment. Maternal body weights, body weight gains and absolute and relative feed consumption values were unaffected at dosages as high as 30 mg/kg bw/day. Pregnancy occurred in 21 to 25 rats in each dosage group. Caesarean-sectioning and litter parameters were not affected by dosages of the test article as high as 30 mg/kg bw/day. No fetal alterations occurred that were considered associated with treatment. On the basis of these data, the maternal and developmental NOAEL is greater than 30 mg/kg bw/day.

Report on assessing repeated dose toxicity and developmental toxicity of delta-Damascone using alpha-Isomethylionone

Delta-Damascone is an ionone, containing an unsaturated hexylring (hexenyl ring) to which an alkenyl chain with an alpha-beta unsaturated and conjugated ketone is attached. In accordance with Article 13 of REACH, lacking information should be generated whenever possible by means other than vertebrate animal tests, i. e. applying alternative methods such as in vitro tests, QSARs, grouping and read-across. For assessing the repeated dose and developmental toxicity of delta-Damascone the analogue approach is selected because for a closely related analogue 90-day repeated dose and developmental toxicity information is available, which can be used for read across to delta-Damascone.

Hypothesis for the analogue approach

Delta-Damascone has similar systemic repeated dose and developmental toxicity as alpha-Isomethylionone based on the similarity in chemical structure, physical chemical properties, reactivity, receptor binding and metabolism between these two substances (see Table 1). Available information: For the target substance delta-Damascone an acute oral toxicity test is available and a 28-day repeated dose toxicity study similar to OECD TG 401 and OECD TG 407, respectively. For the source substance acute toxicity (OECD TG 401), 90-day repeated dose (OECD TG 408) and developmental toxicity information (OECD TG 414) is available.

Target chemical and source chemical(s)

Chemical structures of the target chemical and the source chemicals are shown in the data matrix , including physico-chemical properties and toxicological information, thought relevant for the endpoints that are assessed.

Purity / Impurities

The main impurities are n-alpha-Isomethylionone (CAS no 7779-30-8) (20.9%) and n-betamethylionone (CAS no 127-43-5) (7.9%). Because of the structural similarities of these impurities, it is expected that these impurities do not affect the physico-chemical parameters and human toxicological properties. It is therefore concluded that the impurities will not hinder the read-across (see table 1).

Analogue approach justification

According to Annex XI 1.5 read across can be used to replace testing when the similarity can be based on a common backbone and a common functional group. It can also be used when the analogues used will have the same or similar metabolites. When using read across the result derived should be applicable for C&L and/or risk assessment and it should be presented with adequate and reliable documentation. This application and documentation is presented here.

Structure similarity between delta-Damascone and alpha-Isomethylionone: Delta-Damascone has significant structural homology with alpha-Isomethylionone, both having a cyclohexenyl ring with three methyl groups and a side chain, with a ketone moiety, attached to this ring. The alkenyl side chain of delta-Damascone is a 2-buten-1-one group and of alpha-Isomethylionone a 3-buten-2-one group. Both are alpha-beta unsaturated ketones. Delta-Damascone may exhibit some higher reactivity, on structural grounds, because it does not have the methyl group attached to this alpha-beta unsaturated ketone, which alpha-Isomethylionone has. This methyl group is expected to somewhat hinder the reactivity of the conjugated bond. Another difference between these two chemicals is the switched positions of the alkenyl double bond and the ketone (see table 1).

Toxicokinetic aspects: The physico-chemical properties of delta-Damascone and alpha-Isomethylionone are similar and are therefore expected to have similar absorption and other kinetic behaviour (see data matrix). Both substances have similar water solubilities and vapour pressures, which are relatively low (Table 1). The substances have Log Kow of 4.2 and 4.7, respectively, which illustrates that they are both highly lipophilic.

Metabolism: The metabolic pathway is also expected to be similar (Belsito et al., 2007). The vinyl substituted functional group is expected to be metabolized through reduction to the butenone group to a secondary alcohol, oxidation of the angular methyl groups (becoming alcohols), reduction of the double bond in the exocyclic alkenyl side chain to form dihydro-derivatives and conjugation with glutathione (see metabolic prediction on general ionones, roughly drafted after Belsito et al., 2007). The results of the rat liver S9 simulator of the OECD Toolbox are presented in Appendix 1.

Toxicity aspects: Delta-Damascone and alpha-Isomethylionone have similar toxicity profiles though delta-Damascone may be slightly more toxic for several endpoints as is summarized below.

Acute oral toxicity: Delta-Damascone has a slightly higher acute toxicity (lower LD50) compared to alpha-Isomethylionone and will be classified for acute oral toxicity (see Table 1).

Skin irritation: Delta-Damascone shows skin irritation properties in an in vitro test and need to be classified as such. Alpha-Isomethylionone is considered slightly irritating in an in vivo test, which does not warrant classification and labelling.

Skin sensitization: Delta-Damascone is a strong sensitizer, while the information on alpha-Isomethylionone presents weak sensitization. This difference can be explained by the presence of the methyl group in the alkyl chain in the latter substance, decreasing the reactivity.

Repeated dose toxicity: For delta-Damascone a 28-day study has become available in 2013. In this study the target organ is the liver and a systemic NOAEL of 85 mg/kg bw is derived, though at the highest dose of 340 mg/kg bw also than liver parameters were affected. For alpha-Isomethylionone also the liver is the target organ in the 90-day repeated dose toxicity study. At the highest dose of 500 mg/kg liver effects are seen. At this high dose also an effect on the bone-marrow is seen. At the mid dose of 30 mg/kg bw no effects are observed.

Reproductive toxicity, Fertility: No effects have been seen on fertility (based on the absence of toxicity to the reproductive organs) in the repeated dose toxicity studies of delta-Damascone and alpha-Isomethylionone. Therefore no adverse effects are expected.

Reproductive toxicity, Developmental toxic effects: Alpha-Isomethylionone can be used for read across for this endpoint similarly to what has been presented at repeated dose toxicity. In addition, there are limited indications for delta-Damascone and alpha-Isomethylionone belonging to a group of developmental toxicants as presented in Wu et al. (2013). The only indicator for developmental toxicity one may see is some similarity between the ionones and Vit A, (retinol) of which the chemical structure is presented below. It can be seen that the hexenyl ring of alpha-Isomethylionone shows some similarity with Vit A, though the double bond in the ring is not conjugated with the bonds in the alkenyl side chain, which diminishes the similarity, resulting in < C3 conjugated bonds and is thus not similar to Vit. A (Wu et al., 2013). The 2-position of the ketone group in the side chain is also dissimilar to Vit A. No developmental effects were seen at high doses for alpha-Isomethylionone at doses up to 30 mg/kg bw for which clear effects are seen for Vit A (Hendrikx et al., 2000): doses >= 6 mg/kg bw in monkeys. Delta-Damascone is even more dissimilar to Vit A because of the position of the double bond in this hexenyl ring and the absence of the methyl group attached to this double bond in the ring. Delta -Damascone also has a short alkenyl chain and not a methyl group attached to the alpha, beta-conjugated bond, which indicates even a lower similarity with Vit A.

Uncertainty: Delta-Damascone and alpha-Isomethylionone are similar with regard to structure, physical chemical properties, reactivity, receptor binding, metabolism and toxicity profile. The main difference is the methyl group attached to the alpha-beta conjugated bond in alpha-Isomethylionone, which is expected to diminish its reactivity. This can indeed be seen in the skin irritation and skin sensitization potential being lower. The absence of effects in the gastrointestinal tract of rats treated orally with alpha-Isomethylionone, which are present in rats treated orally with delta-Damascone, may also be an indicator for slightly lower reactivity. In the 28-day repeated dose study with delta-Damascone at 340 mg/kg bw several toxicological effects are seen. These effects other than liver parameters were not seen in the repeated dose toxicity study of the source substance. No effects were seen in the bone-marrow of delta-Damascone treated animals, while some indications were seen during testing with alpha-Isomethylionone, which may be due to the shorter exposure period of the target. For repeated dose an additional assessment factor can be used to account for this expected slightly higher toxicity of delta-Damascone. It has been shown that alpha-Isomethylionone and delta-Damascone are sufficiently similar to use the information for read across.

In addition, the review paper from Belsito et al. (2007) shows that the whole group of ionones and rose ketones have similar toxicological characteristics, indicating that the functional groups are more important for the repeated dose toxicity than their structural dissimilarities. Therefore, the data gaps for delta-Damascone on sub-chronic toxicity and developmental toxicity could be filled in by using the data of alpha-Isomethylionone.

Data matrix

The relevant information on physico-chemical properties and toxicological characteristics are presented in the Data Matrix in Table 1.

Conclusions per endpoint for C&L, PBT/vPvB and dose descriptor

When using read across the result derived should be applicable for C&L and/or risk assessment, cover an exposure period duration comparable or longer than the corresponding method and be presented with adequate and reliable documentation as is presented in the present document. Alpha-Isomethylionone is considered an appropriate read-across candidate to be used to fill in the data gaps on delta-Damascone, which are 90-d repeated dose toxicity and developmental toxicity. The NOAELs for alpha-Isomethylionone for 90-d repeated dose toxicity and developmental toxicity were 30 and >30 mg/kg bw/day, respectively.

Classification and Labelling

Repeated dose toxicity: The effects and NOAEL (85 mg/kg bw) in 28-day repeated dose toxicity study with delta-Damascone do not indicate severe effects and therefore classification and labelling is not needed for this endpoint. Also, the 90-day repeated dose toxicity of alpha-Isomethylionone does not indicate severe repeated dose effects. Though alpha-Isomethylionone has a NOAEL of 30 mg/kg, the effects seen at 500 mg/kg bw (LOAEL) are not considered sufficiently severe to warrant classification and labelling for this endpoint. Therefore delta-Damascone does not need to be classified for 28-day and 90-day repeated dose toxicity in accordance with EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.

Developmental toxicity: The developmental toxicity is based on alpha-Isomethylionone. Since alpha-Isomethylionone need not to be classified and labelled for this endpoint, delta-Damascone is not considered a developmental toxicant in accordance with EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.

DNEL-derivation: For the DNEL-derivation of delta-Damascone, the NOAELs of 30 mg/kg bw/day for 90-d repeated dose toxicity and developmental toxicity will be used as a starting point. For derivation of the DNEL an assessment factor will be included for uncertainty, because delta-Damascone is considered to be slightly more reactive (for details see DNEL derivation).

Table 1: Data Matrix, Analogue Approach for delta-Damascone with alpha-Isomethylionone

Chemical name	delta-Damascone (Target chemical)	Alpha-Isomethylionone (Source chemical)
Structure
CAS no.	57378-68-4	127-51-5
Molecular Weight	192.3	206.3 1)
Purity	95-100%
PHYSICO-CHEMICAL DATA
Physical state at 20 °C and 101.3 kPa	Colorless to pale yellow liquid	Colorless liquid
Melting Point (°C)	10 (33 calculated1) )	45 (calculated) 1)
Boiling point (°C)	Decomposition before boiling	266.2 1)
Partition coefficient (Log Kow)	4.2	4.6 1)
Vapour pressure (Pa) at 20°C	2.7 (IFF internal data) 3.6 (calculated, EpiSuite)	1.2 (IFF internal data) 1.5 (calculated) 1)
Water solubility (mg/L) at 20°C, pH 7.1	77.2 (13.3 calculated) 1)	16 1)
MAMMALIAN TOXICITY
Acute oral toxicity (LD50)	1821 g/kg bw (mice, Moron, 1980))	8714 LD50 mouse (Hoffmann-Larouch, 1967, RIFM database) > 5 g/kg bw (rat) 1)
Skin irritation	Positive (in vitro test, B.36)	Negative 3)
Sensitisation	Positive EC3 = 0.886%	Positive EC3 = 21.8%
28-day repeated dose toxicity (NOAEL)	85 mg/kg bw
90-d repeated dose toxicity (NOAEL)	Read-across	30 mg/kg bw/day 4)
Developmental toxicity test (NOAEL)	Read-across	> 30 mg/kg bw/day 4)

1) http://www.epa.gov/hpv/pubs/summaries/ionederv/c13660tp.pdf

2) http://www.chemicalbook.com/ProductChemicalPropertiesCB5183650_EN.htm

3) RIFM database, performed according to Annex V of EEC Directive 79/831 (OECD TG 414)

4) Belsito et al., 2007

Belsito, D., Bickers, D., Bruze, M., Calow, P., Greim, H., Hanifin, J.M., Rogers, A.E., Saurat, J.H., Sipes, I.G., Tagami, H., 2007, A toxicologic and dermatologic assessment of ionones when used as fragrance ingredients,  Food Chem Toxicol. 2007;45 Suppl 1:S130-67.

Hendrikxs, A.G., Peterson, P., and Hummler, H., 2000, H., Vitamin A teratogenicity and risk assessment in the macaque retinoid model, Reprod. Toxicol., 14, 311-323.

Wu, S., Fisher, J., Naciff, J., Laufersweiler, M., Lester, C., Daston, G. and Blackburn, K., 2013, Framework for Identifying Chemicals with Structural Features Associated with the Potential to Act as Developmental or Reproductive Toxicants, Chem. Res. Toxicol.,26, 1840-61

 

Appendix 1 OECD Toolbox (3.1.0.21) rat liver metabolism simulator.

Justification for classification or non-classification

Based on the absence of adverse effects on reproductive organs or tissues in the 28 -day study with Delta-damascone and in the 90-day study with alpha iso-methylionone classification for fertility is not necessary in accordance with EU Classification, Labeling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.

Based on the absence of adverse effects in the developmental study with Alpha-iso-methylionone, classification is not necessary for developmental toxicity in accordance with EU Classification, Labeling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.