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Administrative data

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Effects on fertility

Description of key information

In a screening test for reproductive/developmental effects (OECD Guideline 421), the NOAEL for reproductive performance and survival of the offspring is 12000 ppm (709 mg/kg bw/day during pre-pairing period) and the NOAEL for parental toxicity is 12000 ppm for males (728 mg/kg bw/day) and 6000 ppm for females (377 mg/kg bw/day during gestation) due to the lower food consumption and lower mean bodyweights and bodyweight gains observed in females at 12000 ppm, especially during gestation and lactation.


Due to the 31-33% reduction in offspring body weight gain in offspring to mothers treated with 12000 ppm, although almost certainly because of the smaller size of the dams and the lower food consumption during lactation due to the low palatability of the diet at this concentration, and not any specific or intrinsic property of (-)-alpha-pinene, the NOAEL for offspring growth and development is 6000 ppm (807 mg/kg bw/day during lactation).

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
377 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
GLP study conducted acording to OECD guideline 421
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

In a screening test for reproductive/developmental effects, and assessment of endocrine disruptor relevant endpoints, conducted according to OECD Guideline 421 and in compliance with GLP, the test item was administered to groups of Crl:CD(SD) rats at dietary concentrations for at least five weeks.


Three groups of ten male and ten female rats received the test item at dietary concentrations of 3000, 6000 or 12000 ppm. Males were treated daily for three weeks before pairing, up to necropsy after a minimum of four consecutive weeks. Females were treated daily for three weeks before pairing, throughout pairing, gestation and until Day 12 of lactation. Females were allowed to litter, rear their offspring and were killed on Day 13 of lactation. The F1 generation received no direct administration of the test item; any exposure was in utero or via the milk. A similarly constituted Control group received untreated diet with corn oil for the same duration as treated animals.
During the study, clinical condition, body weight, food consumption, thyroid hormone analysis, estrous cycles, pre-coital interval, mating performance, fertility, gestation length, sperm analysis (males only), organ weight and macroscopic pathology and histopathology investigations were undertaken. 
The clinical condition, litter size and survival, sex ratio, body weight, ano-genital distance, thyroid hormone analysis and macropathology for all offspring were also assessed. Nipple counts were performed on male offspring on Day 13 of age.
There was no effect of treatment on the circulating levels of thyroxine (T4) in adult males or in offspring on Day 13 of age.
The achieved dosages for animals during treatment and for reproductive phase females before pairing (F0) were 181, 355 and 728 mg/kg bw/day for males and 180, 348 and 690 mg/kg bw/day for females, at dietary concentrations of 3000, 6000 or 12000 ppm, respectively. The achieved dosages for reproductive phase females during gestation were 186, 377 and 711 mg/kg bw/day, and during lactation were 418, 807 and 1494 mg/kg bw/day, at dietary concentrations of 3000, 6000 or 12000 ppm, respectively.
 
Parental toxicity
One control female was euthanized on Day 10 of lactation due to welfare reasons not related to treatment. Administration with test item at dietary concentrations up to and including 12000 ppm was well tolerated and there were no clinical signs that were considered to be treatment related.
Body weight in males was unaffected by treatment, while, initial body weight loss resulted in low body weight gain in all treated female groups before pairing and body weight gain was statistically significantly low during the gestation and lactation phases in females receiving 12000 ppm.
Food consumption in males was slightly but statistically significantly low on the first day of treatment at 12000 ppm. Food consumption was low in all female treated groups between Days 1-4, with a variable food consumption (ranging from 10-19 g/animal/day) between Days 4-22 in females receiving 12000 ppm. Food consumption during gestation and lactation was low in females receiving 12000 ppm.
Group mean kidney weight was high in all male treated groups when compared with controls. Group mean liver weight was high in male and females in all treated groups when compared with controls. Sperm motion, counts and morphology was considered unaffected by treatment. F0 males at 12000 or 6000 ppm, had a high incidence of pale kidneys.  Histologically, accumulation of hyaline droplets and hyaline droplet nephropathy was observed in several males that received 3000, 6000 or 12000 ppm, in a dose dependent manner, indicative of the species- and sex-specific alpha 2μ-globulin nephropathy.
Litter Responses (F1)
All reproductive phase females were pregnant and successfully littered down; litter size, post implantation survival index, live birth index, viability index and sex ratio were all considered to be unaffected by treatment. Ano-genital distance was considered unaffected in male and female offspring, and males did not have nipples at any dose. Body weight and body weight change was low throughout lactation in male and female offspring of maternal females that received 12000 ppm.
There were no macropathology changes considered to be related to treatment in offspring.
 
In conclusion, daily dietary administration of the test item to Sprague-Dawley rats at concentrations of 3000, 6000 or 12000 ppm for three weeks before pairing, up to necropsy after a minimum of four consecutive weeks for males and for three weeks before pairing, throughout pairing, gestation and until Day 12 of lactation for females was well-tolerated in the adult animals, but did elicit histopathological changes in the males kidneys, in a dose dependent manner for all treated groups, indicative of an accumulation of alpha-2 urinary globulin considered as a phenomenon specific to male rats and of no significance to humans. Following the start of treatment, females were more affected by treatment than males: all groups of treated females showed a dose related initial mean body weight loss on Days 1-4 of study, with females receiving 6000 or 12000 ppm having low food consumption during this period. In contrast, body weight gain of males was unaffected following the start of treatment while food intake was only slightly but statistically significantly low on Day 1 of treatment at 12000 ppm. This initial bodyweight loss associated with reduced food consumption is probably the consequence of the low palatability of the diet, especially at high test item concentrations. This lower food intake was confirmed during gestation and lactation periods in the females exposed to 12000 ppm. As a consequence, mean bodyweights of females at 12000 ppm were statistically significantly lower than Controls from Day 7 of gestation to Day 13 of lactation
There was no effect of treatment on estrus cycles, mating performance, fertility, gestation length or index, litter size or offspring survival. At 12000 ppm, offspring body weights on Day 1 of age were slightly low and subsequent weight gain was 31-33% lower than in Control, a magnitude which is considered adverse: this almost certainly reflects the smaller size of the dams and the lower food consumption during lactation, and not any specific or intrinsic property of (-)-alpha-pinene. Offspring body weight gain was also slightly low (<15% lower) at 3000 or 6000 ppm but this magnitude of effect is considered not to be adverse and it was not dose related. There was no effect of treatment on serum T4 levels in F0 males or male and female offspring at Day 13 of age, offspring anogenital distance, nipple counts or external genitalia, or microscopic changes in male and female reproductive organs and thyroids. There is thus no evidence that (-)-alpha-pinene is an endocrine disruptor.



Based on the results of this study, it is concluded that the No-Observed-Adverse-Effect-Level (NOAEL) for reproductive performance and survival of the offspring is 12000 ppm (709 mg/kg bw/day during pre-pairing period) and the NOAEL for parental toxicity is 12000 ppm for males (728 mg/kg bw/day) and 6000 ppm for females (377 mg/kg bw/day) due to the lower food consumption and lower mean bodyweights and bodyweight gains observed in females at 12000 ppm, especially during gestation and lactation.
Due to the 31-33% reduction in offspring body weight gain in offspring to mothers treated with 12000 ppm, although almost certainly because of the smaller size of the dams and the lower food consumption during lactation due to the low palatability of the diet at this concentration, and not any specific or intrinsic property of (-)-alpha-pinene, the NOAEL for offspring growth and development is 6000 ppm (807 mg/kg bw/day during lactation).

Effects on developmental toxicity

Description of key information

Preliminary study OECD 414 in rats: on-going study


Depending on the results of this preliminary study, and more specifically on the similarity of the toxic effects between (-)-alpha-pinene and the mixture of isomers alpha-pinene multiconstituent, either data on alpha-pinene multiconstituent could be used as read-across or an OECD 414 study on the registered substance could be performed.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no study available
Effect on developmental toxicity: via inhalation route
Endpoint conclusion:
no study available
Effect on developmental toxicity: via dermal route
Endpoint conclusion:
no study available

Justification for classification or non-classification

In recent GLP screening test for reproductive/developmental effects, and assessment of endocrine disruptor relevant endpoints (OECD guideline 421):



  • Parental toxicity was observed due to the lower food consumption and lower mean bodyweights and bodyweight gains observed in females at 12000 ppm, especially during gestation and lactation.

  • No effects on fertility and reproductive parameters were observed up to the highest dose tested. Furthermore, decrease in bodyweight and bodyweight gain of offspring were observed at 12000 ppm, probably due to the decrease in food consumption of dams during the lactation period.


Therefore, the registered substance does not need to be classified for reproductive toxicity according to CLP Regulation (EC) n° 1272/2008.

Additional information