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Effects on fertility

Description of key information

Cyclacet did not show Reproduction/developmental toxicity in an oral gavage OECD TG 421 in which a NOAEL of 1000 mg/kg bw/day was derived (OECD 421, GLP)

Cyclacet did not show effects on Reproductive organs in a dietary 90 -day study according to OECD TG 408, with a NOAEL of 1500 mg/kg bw.

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Study duration:
subacute
Species:
rat
Quality of whole database:
The reproduction/developmental toxicity screening study and available repeated dose toxicity studies are of sufficient quality and are sufficiently adequate for this dossier.
Additional information

Cyclacet and its fertility and developmental toxicity in an OECD TG 421 study

The potential adverse effects of the test material on reproduction including offspring development were studied in an OECD 421 Reproduction/Developmental Toxicity Screening Test following GLP. The test material was administered by gavage to three groups, each of ten male and ten female Wistar rats, for up to forty-six consecutive days (including a two week maturation phase, pairing, gestation and early lactation for females), at dose levels of 100, 300 and 1000 mg/kg bw/day. A control group of ten males and ten females was dosed with vehicle alone (Arachis oil BP). Adult males were terminated on Day 43, and all females and surviving offspring on Day 5 post partum. 

Clinical signs, bodyweight change, dietary intake and water consumption were monitored during the study. Pairing of animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation. During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. All animals were subjected to a gross necropsy examination and histopathological evaluation of reproductive tissues was performed.

Results on systemic toxicity: There were no unscheduled deaths and no clinically observable signs of toxicity detected. No adverse effect on bodyweight change, food consumption or food efficiency was detected for treated animals when compared with controls. No toxicologically significant macroscopic abnormalities were detected, no treatment-related effects were detected in the organ weights measured, and no treatment-related microscopic changes were observed.

Results on fertility: There were no treatment-related effects on mating, conception rates, and gestation lengths. Of the litters born, litter size, bodyweight gain and litter weight at birth and on day 1 and 4 were comparable to controls.

Results on developmental toxicity: No clinically observable signs of toxicity were detected for offspring from all treatment groups. In conclusion, the oral administration of the test substance to rats by gavage, for a period of up to forty six consecutive days at dose levels of 100, 300 and 1000 mg/kg bw/day did not result in any treatment-related systemic or reproductive effects. The NOAEL for systemic and reproductive toxicity was therefore considered to be 1000 mg/kg bw/day.

Cyclacet reproductive parameters in a 90 -day (OECD TG 408) study

No fertility effects were seen in the 90 -day oral repeated dose toxicity studies, further confirming the absence of effects on fertility.

Effects on developmental toxicity

Description of key information

Cyclacet and its developmental toxicity: Absence of developmental toxicity based on read across from Verdox in a dietary OECD TG 414 study in which the NOAEL≥444 mg/kg bw/day was derived

Cyclacet and its developmental toxicity in a reproduction/developmental toxicity screening study (rats, oral gavage, OECD TG 421): No developmental toxicity is seen in this study and the NOAEL was ≥1000 mg/kg bw/day

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Species:
rat
Quality of whole database:
The prenatal developmental toxicity study with Verdox, and the reproduction/developmental toxicity screening study and available repeated dose toxicity studies with Cyclacet are of sufficient quality and are sufficiently adequate for this dossier.
Additional information

For Cyclacet, developmental toxicity is derived from the close structural analogue Verdox. The summaries of the experimental information available are presented first and thereafter the read across rationale.

The Cyclacet experimental information on developmental toxicity (from the OECD TG 421) is included in the fertility section above.

Prenatal developmental toxicity study with Verdox

Introduction: A prenatal developmental toxicity study was performed according to OECD TG 414 and GLP. The substance was administered to female Wistar Han IGS rats (Crl:WI(Han)) at dose levels of 800, 2500 and 7700 mg/kg in diet (corresponding to 55, 166 and 444 mg/kg bw total dose). The dose levels were selected based on the results of a 14 -day dose range finding (DRF) study for a previously performed 70 -90 -days extended OECD TG 422 study. In the DRF the dose of ca 15400 mg/kg diet (nominal 1000 mg/kg bw) was used. In this DRF at this high dose decrease in food consumption and body weight was seen and these effects were related to the palatability of the test item Verdox. Verdox is a fragrance which can smell awkwardly and is therefore not tasty either. At this 15400 mg/kg diet in the DRF, the relative liver weight increased 17% in males and females, which was considered to be due to metabolic saturation. This liver weight increase is also expected in combination with Verdox’s molecular weight (196 g/mol) and physico-characteristics, which indicate full and fast oral absorption. The dietary 80 -90 -day Repeated dose / Reproscreen study with the highest dose of 7500 mg/kg diet (converted to nominal 500 mg/kg bw) showed decrease in body weight gain (ca. 15%), which was considered test item related but not adverse. Based on this, the 7500 mg/kg diet was sufficiently high to present (absence of adverse) effects for the OECD TG 414 study and not muddle the outcome of the study due to potential palatability issues. In this Combined Repeated dose / Reproscreen (OECD TG 422) study the relative liver weights in males were increased around 14%, which we related to increased metabolic capacity and considered this sufficiently high to set the maximum dose levels at 7500 mg/kg diet in the follow up OECD TG 414 study.

Method: The 24 females/ dose level were exposed from gestation day 0 (defined by a sperm-positive vaginal smear) to necropsy at gestation day 21. A control group on plain diet was included. Analysis of the test diets showed that the test substance was homogeneously distributed in the test diets and that the concentrations were close to intended. Based on the limited stability in the test feeds, the feed was replaced daily with fresh portions from the freezer. Clinical observations and feed consumption were measured daily. Body weight was determined on gestation days 0, 6, 9, 12, 15, 18 and 21. At gestation day 21, the animals were sacrificed and cesarean section was performed. On the maternal animals gross pathology was performed and weights of reproductive organs were recorded. In addition, the number and distribution of implantations, fetuses and resorptions was recorded. Fetuses were weighed and examined for external observations and then sacrificed and processed for visceral or skeletal fetal evaluations.

Results: During the study no mortalities or morbidity was observed. Statistically significant decreased mean body weight and body weight gain in the 7700 mg/kg diet group which were related to decreased food intake in the in this dose-group. The test substance is a fragrance and probably reduced palatability of the high-dose diet resulted in a lower food intake. Hence the reduced food intake and the accompanying lower body weights in the high-dose group are considered to be due to reduced palatability rather than to the test substance per se, and are not considered to be adverse. No effects on maternal reproductive organ weights (full and empty uterus, ovary weight) fertility and litter parameters (pre- and post-implantation loss, number of live fetuses and resorptions), placenta weight and fetus sex ratio were observed. No treatment related effects on external, visceral and skeletal observations were observed. In the high dose group, a decrease in mean fetus weight in the 7700 mg/kg diet group was observed. This effect is considered to be related to the decrease of maternal food intake and maternal body weight. In view of the minor effect on fetal body weight as compared to the maternal effects and in absence of a delay in ossification this fetal body weight decrease is not considered to be adverse.

Conclusion: As the maternal effects on body weight and food consumption observed in the high dose group are considered related to the palatability of the test substance and are not considered adverse, the No Observed Adverse Effect Level (NOAEL) for maternal toxicity is ≥ 7700 mg/kg diet (corresponding to a dose level of 444 mg/kg bw/day). In absence of effects on ossification the slight effect on fetus weight in the high dose group was not considered adverse and therefore the NOAEL for developmental toxicity is ≥7700 mg/kg diet (corresponding to a dose level of 444 mg/kg bw/day). The substance is therefore not considered to be a developmental toxicant.

Developmental toxicity of Cyclacet (EC no.911-369-0)using read across from Verdox (Cas no. 20298-69-5)

Introduction and hypothesis for the analogue approach

Cyclacet has a tricyclodecenyl fused ring hydrocarbon structure to which an ethyl (acetic) ester is attached. For this substance a 90-day study and a screening for reproductive toxicity study (OECD TG 421) is available but no information on developmental toxicity in rat. 

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. The analogue approach is selected since for the structurally related analogue Verdox a reliable rat developmental toxicity test is available that can be used for read-across to assess the developmental toxicity of Cyclacet.

Hypothesis: Cyclacet has the same developmental toxicity as Verdox.

Available information: For Cyclacet a 90-day is study is available (OECD TG 408, Rel. 1) in which no toxicological relevant effects were seen including on the reproductive organs (NOAEL ≥1500 mg/kg bw in a dietary study). For Cyclacet also a gavage OECD TG 421 screening study for reproductive toxicity is available, showing absence of fertility and developmental effects1000 mg/kg bw (Rel. 1). The absence of developmental toxicity is derived from the analogue Verdox, which is tested in a dietary OECD TG 414 study (Rel. 1) in which no effects were seen at444 mg/kg bw.

Target chemical and source chemical(s)

Chemical structures of the target chemical and the source chemical(s) are shown in the data matrix, including relevant physico-chemical properties.

Purity / Impurities

Cyclacet is a reaction mass containing two very similar isomers, which differ in the position of the double bond which can be at the 5 or 6-yl position. The impurities are all below 1%.

Analogue approach justification

According to REACH 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. 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, which is presented below.

Analogue selection:For Cyclacet the analogue Verdox is selected being a close analogue for which a developmental toxicity (OECD TG 414) is available.

Structural similarities and differences:Cyclacet and Verdox have a similar hydrocarbon backbone with an unreactive acetic ester bond as functional group. These backbones have the same number of carbons. Cyclacet contains an unsaturated bond which is absent in the structure of Verdox. This double bond is not adjacent to the ester and is therefore not impacting the ester reactivity.

Toxico-kinetic:Absorptionvia all routes will be similar in view of similar appearance, molecular weights and physico-chemical parameters. The lower water solubility and higher log Kow of Verdox compared to Cyclacet will not make a difference in absorption because it is still in the range for good oral absorption (Martinez and Amidon, 2002).

Metabolism: Small chain straight alkyl esters such as Cyclacet and Verdox, which are not hindered by adjacent bulky groups are likely to be fully metabolised in the gut by micro-organisms or in the liver by human carboxylesterase (hCE-2) into a secondary cyclohexyl alcohol and acetic acid (EFSA, 2012, 2008)). These secondary alcohols and the acetic acids are expected to be more water soluble and have lower Log Kow values. The secondary alcohols will be glucuronated and easily be excreted via the kidney, while Acetic acid will be consumed in the Krebs cycle and metabolised into CO2(EFSA, 2012).

Mode of Action: Both substances present absence of adverse effects. The secondary (bi)cyclic alcohols resulting from metabolism will be glucuronated and excreted via the kidneys. The increased relative liver weights in both 90-day studies of Cyclacet and Verdox show this higher metabolic demand. Both Cyclacet and Verdox, effects related to α2u-microglobulin hydrocarbon nephropathy were identified.

For both Cyclacet and Verdox no adverse effects were seen up to the highest dose tested in OECD TG 421 and extended OECD TG 422 (> 80-days) for fertility and developmental toxicity further confirming the similar absence of repeated dose and reproductive toxicity effects of these two substances.

Uncertainty of the prediction:There are no remaining uncertainties other than those addressed above.

Data matrix

The relevant information on physico-chemical properties and toxicological characteristics are presented in the data matrix.

Conclusions on hazard and risk assessment

For Cyclacet reproductive toxicity information is available from a 90-day repeated dose toxicity study (OECD TG 408) and from a Reproduction Developmental Toxicity Screening Test (OECD TG 421), which do not present a concern for fertility and developmental toxicity. For the current Annex IX registration developmental toxicity information is needed. This type of information is available from a structural analogue Verdox, which can be used for read across. When using read across the result should be applicable for classification and labelling and riskassessment as well as presented with reliable and adequate documentation. This documentation is presented in the current document. Verdox does not show adverse effects in a developmental toxicity study (OECD TG 414, Kl. 1) NOAEL is ≥ 444 mg/kg bw, which can be directly used for read-across to Cyclacet.

Final conclusion: Cyclacet does not show developmental toxicity: NOAEL of≥444 mg/kg bw.

Data matrix to support the read across to Cyclacet from Verdox for developmental toxicity

Common names

Cyclacet

Verdox

Chemical structures

 

 

 

Target

Source

CAS no.

2500-83-6 (5-yl)

20298-69-5

EINECS

911-369-0

243-718-1

REACH registration

Registered

Registered

Empirical formula

C12H16O2

C12H22O2

Molecular weight

192.25

198

Phys-chem properties

 

 

Physical state

Liquid

Liquid during handling

Log Kow

3.9

4.75

Water solubility (mg/l)

186

10

Human health

 

 

Repeated dose toxicity (mg/kg bw)

≥1500

(OECD TG 408)

≥423

(OECD TG 408)

Reproductive screening test (mg/kg bw)

≥1000

(OECD TG 421)

≥437

(> 80-day extended OECD TG 422)

Developmental toxicity (mg/kg bw)

≥444

(Read across)

≥444

(OECD TG 414)

 

References

 

EFSA, 2008, Scientific Opinion, Flavouring Group Evaluation 47, (FGE.47), Bicyclic secondary alcohols, ketones and related esters from chemical group 8, Scientific Opinion of the Panel on Food Additives, Flavourings, Processing Aids and Materials in Contact with Food (Question No EFSA-Q-2008-051)

Adopted on 22 May 2008.https://efsa.onlinelibrary.wiley.com/doi/epdf/10.2903/j.efsa.2008.743

 

EFSA, 2012, (FGE.87Rev1): Consideration of bicyclic secondary alcohols, ketones and related esters evaluated by JECFA (63 rd meeting) structurally related to bicyclic secondary alcohols, ketones and related esters evaluated by EFSA in FGE.47 (2008), EFSA Journal 2012, 10(2), 2564

 

Verdox, 2018, ECHA dissemination site, EC no.243-718-1.

Justification for classification or non-classification

Based on the absence of adverse effects on fertility and developmental toxicity the substance does not need to be classified for reproduction toxicity according to EU CLP (EC 1272/2008 and its amendments).