Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information
No other data available
Effect on fertility: via oral route
Dose descriptor:
NOAEL
100 mg/kg bw/day
Additional information

A position paper was written for the toxicity to reproduction: see "Toxicity to reproduction, waiving" (§7.8.1) . The conclusion is indicated following:

The 14-day repeated-dose toxicity studies described in § 7.5.1 showed marked toxicity (both systemic and local effects) at 1000 mg/kg bw/d. Furthermore, the findings observed in the reproduction screening test (similar to OECD TG 421) showed that the highest dose level of 100 mg/kg bw/day can be considered as the maximum tolerated dose (MTD) for the dams. Therefore, both studies can contribute to a decision on further testing requirements, i.e. a two-generation study, based on aWeight of Evidenceassessment. In fact, the purpose of Reproduction/Developmental Toxicity Screening Test provided information on the effects on male and female reproductive performance such as gonadal function, mating behaviour, conception, development of conceptus and parturition. Although the exposure duration of this study may not be sufficient to detect all effects on the spermatogenic cycle, it was assumed that in practice the 2-week exposure period would be sufficient to detect the majority of testicular toxicants (Ulbrich & Palmer, 1995). Furthermore, an evaluation of the OECD TG 421 has confirmed that this type of test was useful for initial hazard assessment and could contribute to decisions on further testing requirements (Reuteret al.2003, Gelbkeet al.2004). Therefore, the absence of adverse effects on reproduction or on reproductive organs up to 100 mg/kg bw/d HCA (considered as MTD) in the reproduction screening test together with the strong toxicity of HCA observed at higher doses in the 14-day repeated dose toxicity study were sufficient to meet the information needs for non-classification for toxicity to reproduction. This being the case, it is not useful to conduct of a two-generation study as required at the Annex IX level.

Additionally, supplementary evidence to support this conclusion, comes from a multi-generation database on 50 substances which was recently analysed for second generation parental (P1) / offspring (F2) compared to parental (P0) and first generation offspring (F1) with regard to type of effects as well as incidence, magnitude and severity, at different dose levels (Rorijeet al., 2011). It was demonstrated that P1/F2 generation findings did not play a crucial role in the classification decisions for any substances except one which already provided abundant alerts for endocrine activity and developmental neurotoxicity.

Short description of key information:
Key study: reproduction/Developmental toxicity screening test (similar to OECD 421): NOAEL >=100 mg/kg bw/day for toxicity to reproduction
See the document "HCA_waiving for the two-generation study"

Effects on developmental toxicity

Description of key information
- WoE (alpha-hexylcynnamaldehyde): reproduction/Developmental toxicity screening test (similar to OECD 421): NOAEL >=100 mg/kg bw/day for developmental toxicity in rats.
- WoE (cynnamaldehyde): short-term in vivo screening test based on proposal by Chernoff & Kavlock (1983): NOAEL >=1200 mg/kg bw/day for developmental toxicity in mice.
- WoE (cynnamaldehyde): pre-natal (segment II) toxicity study following the protocol of the Japanese guidelines: LOAEL = 5 mg/kg bw/day for developmental toxicity in rats.
- WoE (cinnamic acid and cinnamic alcohol). Russian developmental toxicity studies of non-standard design: NOAEL >=50 mg/kg bw/day for developmental toxicity in rats for both substances.
Effect on developmental toxicity: via oral route
Dose descriptor:
NOAEL
100 mg/kg bw/day
Additional information

Regarding the developmental toxicity assessment for alpha-hexylcynnamaldehyde (HCA), several studies performed with HCA (one study) and with analogues (Cynnamaldehyde, cynnamic acid or cynnamic alcohol) were considered collectively using aWeight of Evidenceapproach to establish the most relevant endpoint and its NOAEL.

The analogue approach can be considered as HCA has been evaluated within the "Cinnamyl derivatives" category by the Flavor and Fragrance High Production Volume Consortia in its submission to the US Environmental Protection Agency (Submission dated 05Mar2005, see attached document “FFHPVC” §7.1.1), and by the World Health Organisation in its Cinnamyl Alcohol and Related Substances review presented in the WHO Food Additives Series 46 (see attached document, WHO §7.1.1).

The grouping of Cinnamaldehyde and HCA into the “Cinnamyl derivatives” category is based on their structural relationships and the resulting similarities of their physico-chemical (as described in Table 7.1/1) and toxicological properties (see FFHPVC, 2005). Based on this grouping approach, studies on Cinnamaldehyde, its tautomer alcohol (Cinnamic alcohol) and their corresponding acid (cinnamic acid) were considered reliable to assess the toxicological profile of HCA (see attached figure “Metabolism of cinnamaldehyde derivatives” in §7.1.1).

A summary of the different studies is presented below:

In the dosage-range finding study conducted similarly according to guideline OECD 421 (Lewis 2010), rats were given once daily by gavage HCA diluted in corn oil at 12.5, 25, 50 or 100 mg/kg bw or the vehicle alone. The males were treated from 14 days before cohabitation, through mating (maximum of 7 days), and continuing through the day before sacrifice on day 47. The female rats were administered HCA (same dosage as for males) or vehicle two weeks before cohabitation, through mating, and continuing through the day before sacrifice on day. Female rats were allowed to deliver their litters and were sacrificed on postpartum day 5 (PPD 5). F1 generation pups were also sacrificed on PPD 5 (i.e.5thday of lactation). The following parameters were evaluated: viability, clinical observations, body weights, feed weights, mating and fertility, delivery and litter observations, organ weights (epididymes, ovaries, prostate, seminal vesicles, testes, and uterus with cervix), necropsy observations and histopathology (epididymes, ovaries, prostate, seminal vesicles, testes, and uterus with cervix).

All P generation male and female rats survived to scheduled sacrifice. There were no treatment-related clinical observations or gross lesions in the P generation rats of both sexes at any dosage level tested. In addition, none of the microscopic findings examined were considered related to treatment withHCA. Body weights and body weight gains of the treated P generation male rats were generally comparable among the dosage groups. Absolute and relative feed consumption values in male rats were unaffected by treatment withHCAduring the entire dosage period.

There was no effect of HCA on estrous cycling. There was no effect on mating and fertility (fertility index, gestation index, number of implantation sites) at any dosage level tested. All recorded pregnant rats (7, 8, 7, 8 and 6 females in the five respective dosage groups from the control group to the highest dose group) delivered a litter. Natural delivery and litter observations (duration of gestation, number and sex of offspring per litter, stillbirths, live births, gross alterations, litter size and viability, viability index, lactation index, percent survival, sex ratio, pup body weights) were unaffected by dosages of HCA as high as 100 mg/kg bw/day. No treatment-related clinical or necropsy (including a single cross-section of the head and examination of the cross-sectioned brain for apparent hydrocephaly) observations occurred in the F1 generation pups. As such, no developmental effects were observed.

The NOAEL for developmental toxicity was 100 mg/kg bw/day or higher under the test conditions of this study (Lewis, 2010).

In a preliminary developmental study testing 60 chemicals, a dose of 1200 mg/kg bw/day of cinnamaldehyde was administered once daily by gavage to mice from days 6 -13 of gestation. The selected dose represented the predicted maternal LD10. Dams were allowed to deliver their litter around day 18 of gestation. Maternal body weights were recorded on gestation days 6, 17 and day 3 postpartum. Number of live pups and weight of pups was recorded on postnatal days 1 and 3. Reproductive endpoints were maternal weight change, litter size, birth weight and neonatal growth and survival to postnatal day 3.

Cinnamaldehyde induced no effect on maternal survival, maternal bodyweight gain and viable litters. Neonatal response showed no difference on liveborn / litter and percentage survival. Birth weight and weight gain was were normal for all pups. Following the classification used by Chernoff and Kavlock, cynnamaldehyde was recorded in the group of those that had no effect. Results in this assay and conventional mouse teratology tests were concordant. Conventional data were available for 14 chemicals (ten teratogens, one fetotoxin, three non teratogen) of which 11 (nine teratogens, one fetotoxin and one non teratogen) produced evidence of developmental toxicity. Therefore, cynnamaldehyde was considered as low priority candidates for conventional testing on the basis of results.

Under the test conditions, no maternal toxicity and no development effect was observed at the highest dose tested. The No Observed Effect Level was higher than 1200 mg/kg bw/d for maternal toxicity and development toxicity in mice (Hardin, 1987).

In a study (Mantovani, 1987) conducted following the Japanese protocol for drug toxicity studies (Segment II pre-natal toxicity study), female rats (15 to 17/group) were administered cinnamaldehyde once daily by gavage at 0, 5, 25, and 250 mg/kg bw from days 7 to 17 of gestation and then killed on day 20. Weight and food consumption were recorded on days 0, 7, 18 and 20.

Significantly lower weight gain was observed between days 7 and 20 in the dams at the two highest dose levels without a concurrent decrease in food intake and effect on relative liver or kidney weight. The lowered weight gain was the only sign of maternal toxicity even if it was not dose-dependent and probably not related to a difference in liver metabolism and renal function reducing maternal performance.

No effects on embryolethality or gross abnormalities were observed.

There was a significant increase in pre-implantation loss in the controls in comparison with the treated groups: index of affected corpora lutea on the basis of 'total foetuses' were 35, 11, 8, and 14, for controls, 5, 25, and 250 mg/kg bw/day for cinnamaldehyde, respectively.

Very few gross malformations of foetuses were observed. Examination of the skeletons and soft tissues showed that several parameters were significantly increase in treated groups in comparison with the controls

Increase in incidence of poor cranial ossification, dilated ureters and renal variations observed in the 5 mg/kg bw/day, in contrast to the lowest observed effect level of 25 mg/kg bw/d in the dams, may indicate that the fetus was slightly more sensitive to cynnamaldehyde toxicity than in the adult. However, without evidence of dose-related trend together with the absence of historic control data and the absence of the fetal length these results may not be considered as of toxicological concern.

Under the test conditions, LOAEL of 25 mg/kg bw/day for maternal toxicity and a LOAEL of 5 mg/kg bw/day for developmental toxicity could be concluded in this study. However some important data were not available in the publication such as the fetal length which is necessary to determine the time of ossification taking into account both the fetal weight and the fetal length. Furthermore, historical control data are always necessary to determine the acceptability of the concurrent control group, to establish the incidence ranges for frequently and rare occurring morphological changes, to detect the “clusters” of abnormalities. Therefore, the slight abnormalities observed at the lowest dose in this study which mainly represented delay in development may not be relevant in regard to developmental toxicity assessment.

In a non-standard design study were given cinnamyl alcohol orally at a dose of 53.5 mg/kg bw /day or cinnamic acid at a dose of 0, 5, or 50 mg/kg bw/day throughout gestation. On day 20 of gestation, 50% of the treated and control animals were sacrificed, and their fetuses were removed for examination. Fetal body weight, liver nucleic acids, number of survivors, and bone development did not differ significantly between test and control groups. The remaining females from both groups were allowed to deliver normally. Again, the body weights, number surviving, and size and general development of offspring at birth or at 1 month did not differ significantly between treated and control groups (Zaitsev & Maganova, 1975).

Therefore, information is available from a variety of animal studies summarized above, which give different amounts of direct and indirect information on the potential reproductive toxicity of HCA:

-         screening study similar to OECD TGs 421 ( HCA tested)

-         other short-termin vivo screening test,e.i.similar Chernoff/Kavlock test (cynnamaldehyde tested),

-         prenatal developmental toxicity test: pre-natal (segment II) toxicity study following the protocol of the Japanese Ministry of Health and Welfare, Pharmacological Affairs Bureau, 1984. (cynnamaldehyde tested),

-         russian developmental toxicity studies of non-standard design (cinnamic acid and cinnamic alcohol tested).

 

Interpreted with caution (as described above), the overall results didn’t show any concern for developmental toxicology of HCA at dose levels where maternal toxicity was not observed in both screening studies (similar OECD 421 guideline study and similar Chernoff/Kavlock test) even if these studies don’t provide complete information on all aspects of reproduction and development. However, it is important to report that workshop participants (workshop sponsored by the National Institute for Occupational Safety and Health) viewed the Chernoff/Kavlock test as highly reliable in correctly identifying developmentally toxic chemicals and suggested that a negative finding could be a sufficient basis for regulatory agencies to determine that conventional teratology tests in the same species are not warranted (Hardin et al. 1987). Moreover, the available prenatal developmental toxicity studies, even if not reliable based on the absence of historical control data (Mantovaniet al.1989) or details on protocol (Russian studies), provided an indirect focused evaluation of potential effects of HCA (since cynnamaldehyde, cynnamic acid and cynnamic alcohol were tested) on prenatal development only represented by delay of ossification and renal variations.

In conclusion, the Weight of Evidence assessment involves the consideration of all data that are available and are relevant to demonstrate the absence of developmental toxicity concern for HCA

Moreover, HCA is naturally occurring substances, and it is already used as common component of traditional foods and generally recognized as safe (GRAS) as flavoring substances by the U.S. Food and Drug Administration (US FDA) and as food additives by the World Health Organization (WHO) for years. HCA are also used in fragranced consumer products such as soaps and cosmetics.

Justification for classification or non-classification

No self-classification is proposed according to the Directive 67/548/EEC and the Regulation (EC) No. 1272/2008 (CLP Regulation).