Registration Dossier

Administrative data

Description of key information

One repeated dose toxicity study and several reproductive toxicity studies are available in which repeated dose toxicity has been observed. There is a 28 -day repeated dose study, a one-generation study and two developmental studies. In the one-generation study the lowest NOAEL for repeated dose toxicity is seen and will be the starting point for the derivation of the DNEL.  

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
repeated dose toxicity: oral
Remarks:
other: one-generation study
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP-compliant OECD 415 one-generation study, including information on repeated dose toxicity but not all repeated dose toxicity parameters are included.
Qualifier:
according to
Guideline:
other: OECD Guideline 415 (One-Generation Reproduction Toxicity Study)
GLP compliance:
yes (incl. certificate)
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River (UK) Limited, Margate, Kent, UK
- Age at study initiation: (P) males ca. 6 weeks, females ca. 10 weeks
- Weight at study initiation: (P) Males: 191-255 g; Females: 211-271 g
- Housing: initially, in groups of 4 in polypropylene cages with stainless steel grid floors and tops. During the mating, animals were housed in similar cages on 1:1 male:female basis. After mating males were transferred to the original cages; females were housed individually during gestation and lactation in polypropylene cages with solid floors and stainless steel lids.
- Diet: pelleted diet (Rodent PMI 5002 (certified) diet, ad libitum
- Water: Mains drinking water, ad libitum
- Acclimation period: 13 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2
- Humidity (%): 55 ± 15
- Air changes (per hr): at least 15/hour
- Photoperiod (hrs dark / hrs light): 12 / 12
Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: formulations were prepared weekly

VEHICLE
- Concentration in vehicle: 0, 6.25, 25 and 125 mg/mL
- Amount of vehicle (if gavage): 4 mL/kg bw
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The concentration of the test substance in arachis oil was determined by gas chromatography using an external standard technique.
Duration of treatment / exposure:
76 days pre-mating, maximal 21 days mating, males were killed and examined upon evidence of successful mating, females and offspring were killed and examined on day 21 post-partum. Non-pregnant females were killed and examined after day 25 post-coitum.
Frequency of treatment:
Daily (except for females during littering/parturition)
Remarks:
Doses / Concentrations:
0, 25, 100 and 500 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
24/sex/dose
Control animals:
yes, concurrent vehicle
Details on study design:
Details on mating procedure
- Age at mating of the mated animals in the study: mating was performed on day 76 of treatment.
- M/F ratio per cage: 1:1
- Length of cohabitation: a maximum of 21 days
- Proof of pregnancy: vaginal plug and/or sperm in vaginal smear referred to as day 1 of pregnancy
- After successful mating each pregnant female was caged individually during gestation and lactation in polypropylene cages with solid floors and stainless steel lids, furnished with softwood flakes
Dose selection rationale: based on the results of a 14-day dose-range finding study
Rationale for animal assignment (if not random): the animals were allocated to dose groups using a randomisation procedure based on stratified body weights.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: the animals were examined for overt signs of toxicity, ill-health and behavioral change immediately before and after the dosing, and 1 and 5 hours post-dosing during the working week. Animals were observed immediately before and after dosing and 1 hour post-dosing at the weekends or public holidays.

BODY WEIGHT: Yes
- Time schedule for examinations: day 0, then weekly for males until termination. Females were weighed weekly during maturation and daily during mating. Once mating was evident, body weights were recorded on days 1, 4, 7, 14 and 21 of post-coitum and post-partum.

FOOD CONSUMPTION AND COMPOUND INTAKE: yes
- Time schedule for examinations: During the maturation period, weekly food consumption was recorded for each cage. For females showing evidence of mating, food consumption was recorded for the period covering days 1-7, 7-14 and 14-21 post-coitum. For females with live litters, food consumption was recorded for the period covering days 1-7, 7-14 and 14-21 post-partum.

WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule for examinations: Water intake was observed daily by visual inspection of water bottles for any overt change.

OTHER:
- During mating, a vaginal smear was prepared for each female daily and the stage of the estrous cycle was recorded.
- Parameters examined in P male parental generations: testis weight, epididymis weight, numbers of homogenisation resistant spermatids, sperm motility, sperm morphology
- The following parameters were examined in F1 offspring: number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities, the detachment and unfolding of pinna, incisor eruption and eyelid separation, reflexological response to stimuli by assessing surface righting reflex on Day 1 post partum and air righting reflex on Day 17 post-partum. Pupillary reflex and auditory startle response were performed on day 21 post-partum.
Sacrifice and pathology:
SACRIFICE
- Male animals: All surviving animals; after succesful mating.
- Maternal animals: All surviving animals; on day 21 post-partum; for non-pregnant females on or after day 25 post-coitum.
- The off-spring was sacrificed on day 21 after birth.

GROSS NECROPSY
- Gross necropsy consisted of full external and internal examinations of parental animals.

HISTOPATHOLOGY / ORGAN WEIGHTS
The following tissues of parental animals were prepared for microscopic examination and weighed, respectively: cervix, coagulating gland, epididymides, ovaries, pituitary gland, prostate, seminal vesicles, testes, uterus, vagina.
Other examinations:
- Reproductive indices: Mating index = (number of animals mated/number of animals paired) x 100%, Pregnancy index = (number of pregnant females/number of animals mated) x 100%, Parturition index = (number of females delivering live offspring/number of pregnant females) x 100%.
- Offspring viability indices: Live birth index = (number of offspring alive on day 1/number of offspring born) x 100%, Viability index 1 = (number of offspring alive on day 4/number of offspring alive on day 1) x 100%, Viability index 2 = (number of offspring alive on day 7/number of offspring alive on day 4) x 100%, Viability index 3 = (number of offspring alive on day 14/number of offspring alive on day 7) x 100%, Viability index 4 = (number of offspring alive on day 21/number of offspring alive on day 14) x 100%, Viability index 5 = (number of offspring alive on day 21/number of offspring alive on day 1) x 100%.
Statistics:
Linear regression analysis, followed by ANOVA incorporating Levene's test for homogeneity of variance was used for data on organ weights, weekly body weight, litter weights, offspring body weights. Where variances were shown to be homogenous, pairwise comparisons were conducted using Dunnett's test. Where Levene's test showed unequal variances, the data were analysed using non-parametric methods: Kruskal-Wallis ANOVA and Mann-Whitney "U" test.
The non-parametric methods were also used to analyse implantation loss, offspring sex ratio, litter size and landmark developmental markers.
Chi-squared analysis was used for differences in the incidence of lesions occurring with an overall frequency of 1 or greater.
Kruskal-Wallis one-way non-parametric analysis of variance was used for the comparison of severity grades for the more frequently observed graded conditions.
Details on results:
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS): one male treated with 500 mg/kg bw/day was killed in extremis on day 93. One female from this treatment group was found dead on day 97 and a further two females were killed in extremis on days 99 and 100 following difficulties during partutition. Episodes of hunched posture, pilo-erection and tiptoe gait were evident in 500 mg/kg bw/day females during the final week of gestation.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS): body weight gain for males of 500 mg/kg bw/day group was generally lower (average -22%, range -9% - -43%) than control animals throughout much of the treatment period, with statistical differences observed in Weeks 4, 5, 6 and 10.

FOOD CONSUMPTION (PARENTAL ANIMALS): females treated with 500 mg/kg bw/day showed a notable reduction in food consumption (average -21%, range -17% - -28%) throughout lactation, with statistically significant differences throughout 3 weeks.

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS): there were no treatment-related effets on female estrous cycles or on the type or proportion of females with anomalous estrous cycle.

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS): there were no toxicologically significant effects on the concentration, motility or morphology of samples of epididymal sperm. There were no treatment-related effects on the concentration of homogenisation resistant epididymal or testicular spermatid count.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS): mating performance was good in all groups with the majority of animals mating within the first four days of pairing. Subsequent pregnancy rate was unaffected by treatment with only 1, 1, 1 and 2 females failing to achieve pregnancy in the control, 25, 100 and 500 mg/kg bw/day groups, respectively.

ORGAN WEIGHTS (PARENTAL ANIMALS): there were no treatment-related effects.

GROSS PATHOLOGY (PARENTAL ANIMALS): the adult male treated with 500 mg/kg bw/day that was killed in extremis showed gaseous distension in the gastro-intestinal tract. The female treated with 500 mg/kg bw/day that was found dead around partutition had 21 foetuses in-utero. The two females from this treatment group that were killed in extremis both had dead/inactive fetuses in-utero and red/brown staining aroudn the ano-genital region and dark contents in the stomach or enlarged adrenals and an absent rougae on the non-glandular region of the stomach.

HISTOPATHOLOGY (PARENTAL ANIMALS): no treatment-related microscopic changes were observed in the parental animals.

VIABILITY (OFFSPRING): Life birth index for 500 mg/kg bw/day females was significantly lower (-15%) than control animals with litter size continuing to be statistically lower than control animals throughout lactation. Of the high dose females that gave birth to live litters, six females had a total litter loss, predominantly between birth and day 1.

CLINICAL SIGNS (OFFSPRING): Skin sloughing was detected in offspring during the first week of lactation in all treatment groups (more pronounced in the high dose group) together with multiple ridges along the tail in 500 and 100 mg/kg bw/day litters. Swollen ears became apparent in 500 and 100 mg/kg bw/day litters together with the premature opening of eyes and sparse fur coverage in 500 mg/kg bw/day litters.
A delay in the onset (+ 2.1 days) and completion (+ 2.8 days) of pinna unfolding was evident in 500 mg/kg bw/day offspring together with a reduction in the number of offspring passing surface righting, air righting and pupil reflex. A total of eight 500 mg/kg bw/day litters had not fully completed eye opening by weaning.

BODY WEIGHT (OFFSPRING): Body weight gain at 500 mg/kg bw/day was lower (-58%) than control animals for the first week of age and again from day 14 to weaning (day 21 of age) (-17%). Litter weight, at this treatment group, was notably lower than control animals throughout lactation.

ORGAN WEIGHTS (OFFSPRING): No treatment-related changes were detected.

GROSS PATHOLOGY (OFFSPRING): Offspring from females treated with 500 and 100 mg/kg bw/day showed skin sloughing at necropsy. Offspring from females treated with 500 mg/kg bw/day also showed sparse fur coverage.

HISTOPATHOLOGY (OFFSPRING): Acanthosis and hyperkeratosis were seen in relation to treatment for the skin of male and female F1 generation animals treated with 500 and 100 mg/kg bw/day.
Dose descriptor:
NOAEL
Remarks:
toxicity
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Based on gestation length increase.
Critical effects observed:
not specified
Conclusions:
In the one-generation study with rats, the NOAEL for parental toxicity was set at 25 mg/kg bw/day, based on gestation length increase in 100 and 500 mg/kg bw. The NOAEL for developmental effects was set at 25 mg/kg bw/day, based on the skin effects (skin peeling and flaking, and acanthosis and hyperkeratosis observed at necropsy) in male and female F1 animals.
Executive summary:

Reproductive toxicity of Lyral was studied in a GLP-compliant study with Sprague-Dawley rats, conducted according to OECD Guideline 415. The substance was administered at dose levels of 0, 25, 100 and 500 mg/kg bw/day as a solution in arachis oil to groups of 24 rats/sex/dose for 76 days pre-mating and during maximum 21 days mating, after which males were killed, while females and subsequent offspring were killed and examined on day 21 post-partum. Non-pregnant females were killed and examined after day 25 post-coitum. One male treated with 500 mg/kg bw/day was killed in extremis on day 93. One female from this treatment group was found dead on day 97 and a further two females were killed in extremis on days 99 and 100 following difficulties during parturition. Episodes of hunched posture, pilo-erection and tiptoe gait were evident in 500 mg/kg bw/day females during the final week of gestation. Body weight gain for males of the 500 mg/kg bw/day group was generally lower (average -22%, range -9% - -43%) than control animals throughout much of the treatment period, with statistical differences observed in weeks 4, 5, 6 and 10. Females treated with 500 mg/kg bw/day showed a notable reduction in food consumption (average -21%, range -17% - -28%) throughout lactation, with statistically significant differences throughout 3 weeks. There were no treatment-related effects on female estrous cycles or on the type or proportion of females with an anomalous estrous cycle, or toxicologically significant effects on the concentration, motility or morphology of samples of epididymal sperm. There were no treatment-related effects on the concentration of homogenisation resistant epididymal or testicular spermatid counts. Mating performance was good in all groups with the majority of animals mating within the first four days of pairing. Subsequent pregnancy rate was unaffected by treatment with only 1, 1, 1 and 2 females failing to achieve pregnancy in the control, 25, 100 and 500 mg/kg bw/day groups, respectively. An increased gestation length was observed in the 100 and 500 mg/kg bw dose groups. At necropsy, the adult male treated with 500 mg/kg bw/day that was killed in extremis showed gaseous distension in the gasto-intestinal tract. The female treated with 500 mg/kg bw/day that was found dead around parturition had 21 foetuses in-utero. The two females from this treatment group that were killed in extremis both had dead/inactive fetuses in-utero and red/brown staining around the ano-genital region and dark contents in the stomach or enlarged adrenals and an absent rougae on the non-glandular region of the stomach. No treatment-related microscopic changes were observed in the parental animals. Based on the observed increased gestation length, the NOAEL for parental toxicity was set at 25 mg/kg bw/day.

Life birth index for 500 mg/kg bw/day females was significantly lower (-15%) than control animals with litter size continuing to be statistically lower than control animals throughout lactation. Of the high dose females that gave birth to live litters, six females had a total litter loss, predominantly between birth and day 1. Skin sloughing was detected in offspring during the first week of lactation in all treatment groups (more pronounced in the high dose group) together with multiple ridges along the tail in 500 and 100 mg/kg bw/day litters. Swollen ears became apparent in 500 and 100 mg/kg bw/day litters together with the premature opening of eyes and sparse fur coverage in 500 mg/kg bw/day litters. A delay in the onset (+ 2.1 days) and completion (+ 2.8 days) of pinna unfolding was evident in 500 mg/kg bw/day offspring together with a reduction in the number of offspring passing surface righting, air righting and pupil reflex. A total of eight 500 mg/kg bw/day litters had not fully completed eye opening by weaning. Body weight gain in offspring at 500 mg/kg bw/day was lower (-58%) than in control animals for the first week of age and again from day 14 to weaning (day 21 of age) (-17%). Litter weight, at this treatment group, was notably lower than control animals throughout lactation. Offspring from females treated with 500 and 100 mg/kg bw/day showed skin sloughing at necropsy. Offspring from females treated with 500 mg/kg bw/day also showed sparse fur coverage. Acanthosis and hyperkeratosis were seen in relation to treatment for the skin of male and female F1 generation animals treated with 500 and 100 mg/kg bw/day. Based on the skin effects in F1 offspring, the NOAEL was set at 25 mg/kg bw/day for the F1 generation.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
25 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
The selected study is a GLP compliant guideline study and has klimisch score of 1

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

One-generation study: This study is the key study also for the repeated dose toxicity. Also the reproductive and developmental toxicity assessment is included to keep a full overview of the effects.

Method: Reproductive toxicity of Lyral was studied in a GLP-compliant study with Sprague-Dawley rats, conducted according to OECD Guideline 415 (Safepharm Laboratories Limited 2007). The substance was administered at dose levels of 0, 25, 100 and 500 mg/kg bw/day as a solution in arachis oil to groups of 24 rats/sex/dose for 76 days pre-mating and during maximum 21 days mating, after which males were killed, while females and subsequent offspring were killed and examined on day 21 post-partum. Non-pregnant females were killed and examined after day 25 post-coitum.

Results, General, clinical signs and body weight of the parental animals: One male treated with 500 mg/kg bw/day was killed in extremis on day 93. One female from this treatment group was found dead on day 97 and a further two females were killed in extremis on days 99 and 100 following difficulties during parturition. Episodes of hunched posture, pilo-erection and tiptoe gait were evident in 500 mg/kg bw/day females during the final week of gestation. Body weight gain for males of the 500 mg/kg bw/day group was generally lower (average -22%, range -9% - -43%) than control animals throughout much of the treatment period, with statistical differences observed in weeks 4, 5, 6 and 10. Females treated with 500 mg/kg bw/day showed a notable reduction in food consumption (average -21%, range -17% - -28%) throughout lactation, with statistically significant differences throughout 3 weeks.An increased gestation length was observed in the 100 and 500 mg/kg bw dose groups. At necropsy, the adult male treated with 500 mg/kg bw/day that was killed in extremis showed gaseous distension in the gastro-intestinal tract. The female treated with 500 mg/kg bw/day that was found dead around parturition had 21 foetuses in-utero. The two females from this treatment group that were killed in extremis both had dead/inactive fetuses in-utero and red/brown staining around the ano-genital region and dark contents in the stomach or enlarged adrenals and an absent rougae on the non-glandular region of the stomach. No treatment-related microscopic changes were observed in the parental animals. Based on the observed increased gestation length, the NOAEL for parental toxicity was set at 25 mg/kg bw/day.

Fertility: There were no treatment-related effects on female estrous cycles or on the type or proportion of females with an anomalous estrous cycle, or toxicologically significant effects on the concentration, motility or morphology of samples of epididymal sperm. There were no treatment-related effects on the concentration of homogenisation resistant epididymal or testicular spermatid counts. Mating performance was good in all groups with the majority of animals mating within the first four days of pairing. Subsequent pregnancy rate was unaffected by treatment with only 1, 1, 1 and 2 females failing to achieve pregnancy in the control, 25, 100 and 500 mg/kg bw/day groups, respectively.

Developmental parameters: Life birth index for 500 mg/kg bw/day females was significantly lower (-15%) than control animals with litter size continuing to be statistically lower than control animals throughout lactation. Of the high dose females that gave birth to live litters, six females had a total litter loss, predominantly between birth and day 1. Swollen ears became apparent in 500 and 100 mg/kg bw/day litters together with the premature opening of eyes and sparse fur coverage in 500 mg/kg bw/day litters. A delay in the onset (+ 2.1 days) and completion (+ 2.8 days) of pinna unfolding was evident in 500 mg/kg bw/day offspring together with a reduction in the number of offspring passing surface righting, air righting and pupil reflex. A total of eight 500 mg/kg bw/day litters had not fully completed eye opening by weaning. Body weight gain in offspring at 500 mg/kg bw/day was lower (-58%) than in control animals for the first week of age and again from day 14 to weaning (day 21 of age) (-17%). Litter weight, at this treatment group, was notably lower than control animals throughout lactation. Offspring from females treated with 500 and 100 mg/kg bw/day showed skin sloughing starting at Day 5 of lactation. Offspring from females treated with 500 mg/kg bw/day also showed sparse fur coverage. Acanthosis and hyperkeratosis were seen in relation to treatment for the skin of male and female F1 generation animals treated with 500 and 100 mg/kg bw/day. Based on delayed development and the skin effects in F1 offspring, the NOAEL was set at 25 mg/kg bw/day for the F1 generation.

In view of the effects found in pups in the one-generation study two additional studies were conducted to assess and interpret the skin sloughing effects. These studies are described at the developmental toxicity section including the systemic toxicity. Several systemic effects are seen at 500 mg/kg bw e.g. body weight and liver parameters. These effects have not been repeated here and it is referred to the developmental section.

A 28-day repeated dose toxicity study: Method: In a GLP-compliant OECD Guideline 407 study of Safepharm Laboratories Limited (2006) Lyral was administered at dose levels of 0 (vehicle control), 15, 150 and 1000 mg/kg bw/day to groups of 5 male and female Crl:CD (SD) IGS BR rats by gavage in arachis oil for 28 consecutive days. Animals were observed for mortality and clinical signs, changes in body weight, food and water consumption. On the 28th day, blood was collected from all rats for haematological and clinical chemistry examinations. Prior to the start of treatment and on days 3, 10, 17 and 24 animals were observed for signs of functional/behavioral toxicity. Functional performance tests were also performed on all animals during week 4, together with an assessment of sensory reactivity to different stimuli. Observations were carried out from ca. 2 hours post-dosing on each occasion. At the study termination, all animals were necropsied and subjected to gross pathological and histopathological examinations.

Results: Animals of both sexes treated with 1000 mg/kg bw/day showed transient increased salivation around the time of dosing from day 3 onwards. Isolated incidents of red/brown staining around the mouth and scab formation (males only) were also evident between days 13 and 25. Episodes of respiratory pattern changes and hunched posture were evident in animals of either sex treated with 1000 mg/kg bw/day during the final two weeks of study. Males treated with 1000 mg/kg bw/day showed a reduction in body weight gain during week 1. Females treated with 1000 mg/kg bw/day showed a slight reduction in body weight gain during week 1 and 4 only, whilst males from this treatment group and 150 mg/kg bw/day showed a reduction in body weight gain throughout the remaining treatment period. Males treated with 1000 mg/kg bw/day showed a reduction in food consumption and food efficiency during week 1 only. No toxicologically significant changes in neurobehavior were noted. Animals of both sexes from the 1000 mg/kg bw/day group showed an increase in plasma aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, albumin and albumin/globulin ratio when compared with the controls. In 1000 mg/kg bw/day males statistically significant reductions in plasma cholesterol, total protein and glucose were observed. An increase in alkaline phosphatase and alanine aminotransferase, and an increase in albumin were observed in 150 mg/kg bw/day females and males, respectively. At necropsy, animals of both sexes of 1000 mg/kg bw/day and males of 150 mg/kg bw/day groups showed an increase in absolute and relative liver weight. Males treated with 1000 mg/kg bw/day also showed an increase in absolute and relative kidney weight. Centrilobular or generalised hepatocyte enlargement, frequently with associated focal centrilobular inflammatory cell infiltrates, were observed in animals of both sexes of the 1000 mg/kg bw/day group. In addition, centrilobular hepatocyte necrosis was seen in males treated with 1000 mg/kg bw/day. Three males treated with 150 mg/kg bw/day also showed hepatocyte enlargement. The proximal tubular epithelium of males treated with 1000 mg/kg bw/day was observed to be generally denser than in controls. Changes in cytoplasmic density are occasionally observed as a consequence of test material administration and are frequently adaptive in nature in the absence of associated degenerative changes. Animals from the remaining treatment groups were not similarly affected. Based on the results of the study, the NOAEL was set at 150 mg/kg bw/day, based on an increased relative liver weight and centrilobular or generalised hepatocyte enlargement associated with focal centrilobular inflammatory cell infiltrates in both sexes, and centrilobular hepatocyte necrosis in males at 1000 mg/kg bw/day. The histopathological changes detected at 150 mg/kg bw/day were confined to adaptive liver changes in three males.

In addition to the present REACH evaluation a toxicological assessment has been done by the SCCS and FFHPVC which will be summarised below.

The SCCS also concluded that the NOAEL in the OECD 415 study for maternal toxicity is 25 mg/kg bw based on effects on gestation length at 100 mg/kg bw. The NOAEL of the 28 day study was set at 15 mg/kg bw based on modifications of biochemical parameters observed in males and females at the dose of 150 mg/kg bw/day and the effects on liver (increase in liver weight and hepatocyte enlargement) in the 28-day repeated dose toxicity study of Safepharm Laboratories Limited (2006) . They conclude that the modifications observed at the dose of 150 mg/kg bw/day may be considered as early indicators of liver toxicity observed at higher doses

(http://ec.europa.eu/health/scientific_committees/consumer_safety/docs/sccs_o_074.pdf). In this REACH evaluation we considere the liver effects at 150 mg/kg bw adaptive in nature.

The US-EPA has evaluated the FFHPVC evaluation of Lyral and some analogues. New reproductive toxicity studies have been added to the dossier after submission in 2008 and therefore the US-EPA evaluation has not included all data (those of 2009). The US-EPA comments can be found at http://www.epa.gov/hpvis/hazchar/13006644_%203%20and%204-_4-hydroxy-4-methylpentyl_-3-cyclohexene-1-carboxaldehyde_March%202010.pdf


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Repeated dose toxicity needs to be assessed according to the REACH regulation. Several repeated dose toxicity studies are available. In a one-generation study the lowest NOAEL was determined for systemic toxicity and therefore this is considered the key study.

Justification for classification or non-classification

The NOAEL in the one-generation study is set at 25 mg/kg bw based on increased gestation length at 100 mg/kg bw being the most critical effect. This NOAEL is below the cut off for the absence of STOT 2 RE classification. These systemic effects seen at 100 mg/kg bw do not indicate effects as mentioned at 3.9.2.7.3 of the EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008,

such as morbidity, functional changes in the nervous system, significant changes in haematology changes parameters and/or significant organ damage. Therefore, classification is not warranted in accordance with EU Directive 67/548/EEC (DSD) and EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.