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Key value for chemical safety assessment

Effects on fertility

Description of key information

Systemic Toxicity

In a two-generation reproduction toxicity study (OPP 83-4/OECD 416), the unequivocal LOAEL / NOAEL of SYSTEMIC toxicity are 75 and 25 mg/kg bw/day, respectively, based on significantly and dose-related decreased body-weight gains in the mid- and high-dose groups of the second generation (F1/P1-males only) and slightly but significantly reduced food consumption (p<0.05) in both sexes of these two groups in the premating phase, and based on depressed growth of P0- and P1-dams during gestation and lactation. Overall, systemic adverse effects seemed to be mild, but more marked in the second generation (F1).

Reproduction Toxicity

As for reproduction, significant effects were related to pre-and neonatal toxicity: F1 was delivered with reduced number of pups, associated with increased early post-partum mortality/stillborns in the high-dose groups. F2 was delivered with reduced litter size, associated with increased early post-partum mortality/stillborns in the mid- and high-dose group. While the mean neo-natal body-weights (day 0) were normal for both, the F1- and the F2-offspring, there was evidence of growth depression during lactation in dose-related manner, significant in F1-pups of all treated dams (p<0.01), and less marked in the F2-pups at lactation day 14 and 21. At the dose of 25 mg/kg bw/day, delayed F1-pup growth is the only effect that significantly deviated from the control group. Hence, this is not a NOEL. However, although considered treatment-related and also present to a minor extent in the F2-pubs of the low-dose group, this singular significant effect is not considered adverse, and therefore 25 mg/kg/day has been established as the relevant NOAEL for reproduction performance and pre- and post-natal development across all generations and developmental stages.

Link to relevant study records

Referenceopen allclose all

Endpoint:
two-generation reproductive toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
EPA OPP 83-4 (Reproduction and Fertility Effects)
Qualifier:
according to guideline
Guideline:
OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)
Deviations:
yes
Remarks:
minor deviations; see below under "Principles of method ..."
Principles of method if other than guideline:
Deviations from OECD guideline:
- Treatment of P0-generation for 56 days instead of 70 days
– No observation of oestrous cycle
– No documentation of sex ratio prior to culling of litter size
– No assessment of sperm parameters
– No determination of required organ weights
– No histopathology on coagulating gland
GLP compliance:
yes
Limit test:
no
Specific details on test material used for the study:
- Name of test material: Creosote; North American P1/P13 Creosote; North American Creosote Composite Test Material P1/P13
- Composition of test material, percentage of components: see under Test material information
Species:
rat
Strain:
Sprague-Dawley
Remarks:
SD Crl:CD® VAF/Plus®
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Portage, Michigan, USA
- Age at study initiation: P-generation: 46 d
- Weight at study initiation: P-generation: Males: 150-190 g; Females: 119-144-x g
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
VEHICLE
- Justification for use and choice of vehicle (if other than water): solubility of test material
- Concentration in vehicle: 2.5, 7.5, 15.0 mg/mL
- Amount of vehicle (if gavage): 10 mL/kg bw
Details on mating procedure:
Duration of mating: max. 21 days
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentrations, homogeneity, and stability in the vehicle examined (Report (Vol. 1) Chap. 5.1 "Analytical Results")
Duration of treatment / exposure:
Duration of exposure before mating: 56 days (P0), ≥ 113 days (F1).
Duration of exposure in general: from beginning of the study until sacrifice of P0, F1-, and F2-generation (through mating, gestation, and lactation phase of P0 and F1). F1-offspring was not exposed during lactation but after weaning at 35 days of age to reduce gavage-induced injuries.
Duration of test: 10 months
Frequency of treatment:
once daily
Details on study schedule:
Number of generation studied: 2
Dose / conc.:
25 mg/kg bw/day (actual dose received)
Dose / conc.:
75 mg/kg bw/day (actual dose received)
Dose / conc.:
150 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
26 (P0- and F1-parents)
Control animals:
yes, concurrent vehicle
Positive control:
not required
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily, 2x/d, 7 d/wk
- Cage side observations: all animals for overt signs of toxicity and mortality

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly (during each study week, a detailed clinical observation of each animal was performed and the findings recorded daily)
F1 parental animals were observed daily beginning at 22 days of age.

BODY WEIGHT: Yes
- Time schedule for examinations: individual body weights were recorded weekly until evidence of copulation was observed or until euthanasia. Mated females were weighed on gestation days 0, 6, 15, and 20 and on lactation days 0, 7, 14 and 21. Body weights of non-gravid females were recorded on the above presumed gestation days but were not included in summarisation during gestation.

FOOD CONSUMPTION AND COMPOUND INTAKE
- no feeding study, but individual food consumption was recorded weekly for all animals
- not measured during the mating periods, because the animals were cohabited at that time
- following the mating periods: mated males: weekly until euthanasia
mated females: at the days of body weight measurements during the gestation and lactation periods (data of non-gravid females were not included in summarisation during gestation)
females without evidence of copulation: weekly until either delivery or euthanasia

WATER CONSUMPTION AND COMPOUND INTAKE (no drinking water study): No data

OTHER
Animals not surviving to scheduled euthanasia were necropsied and, where practicable, the tissues were preserved in fixative. Any rat showing signs of severe debility or toxicity were euthanized for humane reasons and to prevent loss of tissues. Following parturition for the F1 and F2 litters, all P and F1 males were evaluated for fertility, euthanized and necropsied. Any P male that failed to sire a litter was evaluated for spermatogenesis by examination of the epididymis for the presence of sperm.
Oestrous cyclicity (parental animals):
not performed
Sperm parameters (parental animals):
Parameters examined only in P0 males that failed to sire a litter:
- Testis weight
- Presence of sperm in epididymis
Litter observations:
PARAMETERS EXAMINED
The following parameters were examined in [F1 / F2] offspring:
number and sex of pups, stillbirths, live births, presence of gross anomalies, weight gain, physical or behavioural abnormalities

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities; possible cause of death was not determined for pups stillborn or found dead.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: all P0- and P1-animals
- Maternal animals: all P0- and P1-animals

GROSS NECROPSY
- Gross necropsy of P0 and P1 consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGHTS
Organ weights: testis only
Histopathology of P0 and P1: focused on reproductive organs, vagina, uterus, ovaries; testis, epididymis, seminal vesicle, prostate (see Report, Vol. 1, Table 34 and 35 (male and female), other organs case by case.



Postmortem examinations (offspring):
SACRIFICE
- The F1-offspring, which had not been selected as parental animals, and all F2-offspring were sacrificed.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows:

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGHTS
F1 not selected for mating, F2: Not performed, only gross necropsy with special attention to reproductive organs.
Statistics:
yes; cf. study report for details
Reproductive indices:
yes; cf. table below under 'Any other information on results incl. tables'
(mating index, fertility index (m/f), pregnacy index, number of implantation sites, duration of pregnancy)
Offspring viability indices:
yes; cf. table below under 'Any other information on results incl. tables'
(litter size, live offspring, live birth index, viability index)
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Increased salivation in the high-dose of both sexes, and female mid-dose group;
increased anogenital staining in the high-dose groups of both sexes.
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, non-treatment-related
Description (incidence):
males (P0): females (P0):
===================================================================
Control: 1/26 (day 43) 1/26 (day 79)
25 mg/kg: no death 2/26
75 mg/kg: 1/26 (day 101, last study day) 4/26 (day 3, 6, 11, 83)
150mg/kg: 1/26 (day 78) 4/26 (day 73, 83, 96, latter euthanised)
====================================================================
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
In the male high-dose group, mean body-weight growth was retarded by more than 10% after 15 weeks (termination) (Report, Vol. 1, Fig.1): BW gain (control) = 359 g vs. BW gain (high-dose) = 308 g, resulting in a weight gain of 86% of the control (Data from Report, Tab.6).
For females, significant differences in mean body-weights were not noted across all groups during the pre-mating phase (8 wks) (Report, Vol. 1, Fig.1, cont´d). Dose-related decreases in body-weight gain were noted in the late gestation phase from days 15 - 20, but without significant effect on the terminal (pre-partum) body weights (decreases <<10 %) (Report, Vol. 1, Fig.2).
During lactation, mean body-weight gain in the high-dose group was significantly retarded from post-partum day 7 through 14, while catching up towards day 21, while maternal weights of the other groups (including the control) slightly declined (Report, Vol. 1, Fig.3).
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
The feed intakes of the males were increased in dose-related manner during the pre-mating period, significant for the high-dose group (Report, Vol. 1, Fig.7). A similar but less marked pattern was seen for the female groups (Report, Vol. 1, Fig. 7 cont´d). During lactation, maternal daily food consumption of the high-dose group was much less than in the control, but also slightly significant at the low- and mid dose (Report, Vol. 1, Fig.9). This correlates with the lower mean body weights in the treated female P0 groups (Report, Fig.3).
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
no effects observed
Description (incidence and severity):
Eye/pupil observation was included within clinical inspections, yet not the complete set of endpoints.
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
effects observed, treatment-related
Description (incidence and severity):
Visual urine status was included within clinical inspections, yet not a complete endpoint set:
The incidence in urinary discoloration became slightly increased with dosing in the treated groups of both sexes.
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
No test-article-related microscopic lesions observed in animals that were either euthanised at study termination or died during the study period (see Report, Vol. 1, Table 34 (male and female))
Histopathological findings: neoplastic:
not examined
Other effects:
not examined
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
effects observed, treatment-related
Description (incidence and severity):
No tretment-related effects on male and female fertility indices in P0-generation (mating, fertilisation, pregnancy, duration of pregnancy, implantations),
Increase in post-implantation loss of F1 --> decrease in litter size and live offspring; in addition, post-partum loss, including cannibalism, increased at 150 mg/kg (highest dose tested).
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
P0 generation, MALES
Appearance, behaviour and mortality:
Control group: One male died on study day 43. The animal had no visible ante-mortem observations. Instead, necropsy findings were consistent with gavage injury. Anogenital staining was observed in nine animals. One animal had a stained body surface.
25 mg/kg/day group: No deaths occurred among males. Discoloured urine was seen in one rat, while nine rats displayed anogenital staining.
75 mg/kg/day group: One male died on day 101 (last day of study) with ante-mortem observations, mostly staining and material around orifices. No anomalies were found during necropsy. Anogenital staining was seen in 13 animals; three rats had a stained body surface.
150 mg/kg/day group: One male died on day 78 with numerous clinical signs (material around eye, anogenital staining, soft stool, decreased defecation and unkempt appearance). All signs started at same time as malocclusion (teeth). Increased salivation was seen in six animals, discoloured urine in two, and anogenital staining in 15 cases. Three rats showed stained body surfaces.

P0 generation, FEMALES
Appearance, behaviour and mortality:
Control group: One female died on study day 79. The animal had no visible ante-mortem observations. Instead, necropsy findings were consistent with gavage injury. Anogenital staining was observed in six animals. Three animals had a stained body surface.
25 mg/kg/day group: Two deaths occurred among females. One animal had no visible ante-mortem observations, while the other showed hair loss only. Both had necropsy findings consistent with gavage injuries (fluid-filled thoracic cavity and discolouration of lungs). One female displayed anogenital staining.
75 mg/kg/day group: Four females died in this group. One animal died on day 83. Her ante-mortem observations consisted of hair loss and 6 days of anogenital staining just prior to death; necropsy findings were all within limits. The other 3 females died on study days 3, 11, and 66, respectively. All three had necropsy findings consistent with gavage injuries (clot in thoracic cavity, lung/thoracic abscess, sternum adhesion, and discolouration of the lung). Increased salivation was seen in four animals, discoloured urine in one, and anogenital staining in seven cases. One rat showed a stained body surface.
150 mg/kg/day group: One female died on day 85. She had delivered 7 stillborns and had numerous clinical signs (discoloured urine, reddish fluid in refuse pan and staining). Necropsy revealed a dilated pelvic region of kidney with multiple foci. Two other females (death on day 73 and euthanasia on day 96) had discolouration of the lungs indicating gavage injury. Increased salivation was seen in five animals, discoloured urine in three, and anogenital staining in 23 cases. Eight rats showed stained body surfaces.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
P0 generation, MALES
Mean male body weights for the 150 mg/kg/day group were significantly reduced on study weeks 7-13 and week 15 when compared to controls (p<0.05) (Overall weight gain was 86% of the control). These differences were considered treatment-related. No significant differences in mean body weight between control males and the other treated males were detected (Report, Fig.1).
Food consumption:
Males in the 150 mg/kg/day group generally consumed more food than the control group. During the pre-mating period, this group consumed more food during 6 out of 8 weeks measured and significantly (p≤ 0.05) consumed more in weeks 12 and 14 (after mating). These increases in food consumption were considered treatment-related. No significant differences in food consumption between control males and the other treated males were detected ((Report, Fig.7).

P0 generation, FEMALES
No significant differences in mean body weight between controls and any treatment group were detected during the pre-mating (Report, Fig.1).
Although reported to be statistically significant (p≤ 0.01) during gestation days 15-20 and 0-20, these decreases in body weight gain were less than 10% relative to controls for the 75 and 150 mg/kg/day groups. These differences were considered treatment-related but not adverse.
During lactation, growth in the 150 mg/kg/day group was significantly retarded (p≤ 0.01), mean body weights different from controls on lactation day 7 and 14, respectively (p≤ 0.05 and p≤ 0.01) (Report, Fig.3). However, body weight gain was significantly increased during lactation days 14-21, when compared to controls and the other groups, as growth of the high-dose dams caught up, while decreasing in the other groups. These differences were considered substance-related. No difference in body weights were found for the low- or mid-dose groups during lactation when compared to the control group.
Food consumption:
Females in the 150 mg/kg/day group consumed significantly (p≤ 0.05 and p≤ 0.01) more food than the control group during weeks 3 and 7, respectively (Report, Fig.7). In the 75 mg/kg/day group, food consumption was significantly (p≤ 0.05 and p≤ 0.01) elevated compared to controls during weeks 2 and 3, respectively. There were no significant differences in food consumption between controls and the 25 mg/kg/day group during the pre-mating period. During the 3 weeks of gestation, food consumption was comparable across all groups (Report, Fig.8)..
During the 3 weeks of lactation, food consumption in the female 150 mg/kg/day group was significantly (p≤ 0.01) reduced compared to controls (Report, Fig. 9). This was considered treatment-related and correlated with the lower body-weight gain in this group (Report, Fig.3). In the 25 and 75 mg/kg/day groups, food consumption was significantly (p≤ 0.05) lower than in controls only between lactation days 14-21. Since these reductions were mainly due to one single animal at 25 mg/kg/day and two animals at 75 mg/kg/day, they were not considered an indication of toxicity.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
P0 Females
Intrauterine examination on lactation day 21 revealed no significant differences in the mean number of implantation sites across all groups (Report, Tab.28). However, the mean litter size (delivered offspring F1) was reduced in the 150 mg/kg/day dams (p<0.05), with 9.2 F1-pups vs. 12.5 implantation sites. Furthermore, the neo-natal live F1-offspring (day 0) decreased to 6.8 pups due to elevated early post-partum death (p<0.01). Hence, for the 150 mg/kg/day dams, the number of implantation sites not corresponding to delivered offspring (representing post-implantation loss and/or complete cannibalisation at birth of the F1-generation) increased when compared to controls: These differences were considered treatment-related. No physiologically relevant difference in these parameters was found in the other treatment groups as compared to controls.

ORGAN WEIGHTS (PARENTAL ANIMALS)
P0 generation, Males
Testis weight: No statistical difference observed across treatment groups.


PATHOLOGY (PARENTAL ANIMALS)
P0 generation, males and females
No test-article-related gross lesions were observed. Any isolated lesion observed in the thoracic cavity (clot, fluid, abscess, foreign material), lung (discolouration, abscess, adhesion, foreign material), heart (adhesion, discolouration, swelling, foreign material), oesophagus (perforation), and bone-rib/sternum (adhesion) in any group were considered as induced by the administration procedure (gavage) and not induced by the test article.
No test-article-related microscopic lesions were observed in animals that were either euthanised at study termination or died during the study period. Isolated lesions observed in the lung (haemorrhage, abscess, pleuritis), heart (pericarditis), oesophagus (inflammation), diaphragm (inflammation, abscess), soft tissue-thorax (abscess) and bone-sternum (abscess) were considered gavage-induced.
Key result
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Effect level:
75 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: hypersalivation, pre-mating body-weight loss (males); reduced body-weight of dams during gestation and lactation
Key result
Dose descriptor:
NOAEL
Remarks:
fertility and prenatal development
Effect level:
75 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
reproductive performance
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
150 mg/kg bw/day (actual dose received)
System:
other: unspecific systemic toxicity (hypersalivation; reduced premating body-weight of males; reduced weight in dams during gestation and lactation)
Organ:
not specified
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
150 mg/kg bw/day (actual dose received)
System:
other: reproductive performance (reduced F1-litter size and live-birth index (post-implantation losses)
Organ:
placenta
uterus
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Increased salivation in the mid- and high-dose group of both sexes, more pronounced in females
Increased anogenital staining in the mid- and high-dose groups of both sexes
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, non-treatment-related
Description (incidence):
males (P1): females (P1):
============================================================
Control: 4/26 no death
25 mg/kg: 2/26 1/26
75 mg/kg: 1/26 3/26
150mg/kg: 2/26 5/26 (2 cases probably by gavage injuries)
============================================================
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
In F1-males, mean body growth was markedly depressed in a dose-related manner from wk 4 through wk 22, reaching about 93%, 85%, and 74% of the control in the low-, mid-, and high-dose group, respectively (Report, Vol. 1, Fig. 4/Tab. 9).
Mean body growth of F1-females was significantly different from the control group during the premating phase from wk 4 through about wk 22, but was not strongly dose-dependant, in particular in the late phase before gestation (Report, Vol. 1, Fig. 4, cont´d). Until wk 22, the mean body-weight gains of all exposed female groups remained distinctly above 90% of the control. During gestation, the mean maternal body weights were significantly lower in the treated groups, most pronounced at the high dose; however this was mainly due to the lower weights at the onset of pregnancy. In reality, the mean weight gains were largely similar across all dose-groups until gestation day 15. Only in the late gestation phase from days 15 - 20, body-weight gain was depressed in the maternal high-dose group (Report, Vol. 1, Fig. 5).
During lactation, mean body weights increased, but in the high-dose group remaining on the same low level from post-partum day 7 through 21 (Report, Vol. 1, Fig. 6).
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Note: The food intakes are reported as amount food[g] per day and animal, hence not normalised to kg body-weight.
The feed intakes of the F1-males followed a similar profile over time in the premating period, but mean consumption in the high-dose group was apparently lower from wk 4 through 10, while increasing over the other group levels, including the control (Report, Vol. 1, Fig. 10). A similar but less marked pattern was seen for the F1-female groups (Report, Vol. 1, Fig. 10 cont´d). During gestation, maternal daily food consumption was steadily increasing in all treated groups, while the mean food rations of the control group remained relatively constant. But note: Three time intervals of 5 and 10 weeks are summed up. (Report, Vol. 1, Fig. 11). During lactation, maternal daily food consumption was also steadily increasing in all treated groups over time, but distinctly lower than in the control, the high-dose group showing the lowest intake (Report, Vol. 1, Fig. 12).
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Description (incidence and severity):
Eye/pupil observation was included within clinical inspections, yet not the complete set of endpoints.
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Description (incidence and severity):
Visual urine status was included within clinical inspections, yet not a complete endpoint set.
There were no particular findings in the treated groups.
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
Only testes in P1-animals examined: Significant and dose-dependent decrease in the mean absolute testis weight was seen in the mid- and high-dose males when compared to controls. No difference was detected in the mean relative testis weight across treatment groups (Report, Tab. 32). Consequently, the decrease in mean absolute testis weight is considered secondary to decreases in body weight and not a direct effect of the test article.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Isolated lesions were caused mechanically due to gavage dosing, but not test item-related (in males and females).
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
No test-article-related microscopic lesions observed in animals that were either euthanized at study termination or died during the study period
Histopathological findings: neoplastic:
no effects observed
Other effects:
not specified
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
effects observed, treatment-related
Description (incidence and severity):
Diffuse and irregular rather than evaluable effects observed in the P1-reproductive parameters, including mating, fertility, pregnancy, possibly influenced by treatment but not dose-related, hence not considered useful for toxicity assessment, even though partly distinctly outside historical control values. These observations have to be weighted in relation to "subfertility problems in CD rats" recognised in previous years (note: authors´ statement in Report, 7.1 Conclusions, p. 37).
At 150 mg/kg bw/day, an increase in post-implantation losses is noted: --> decrease in litter size and in neonatal d0 survival of the F1-offspring. The post-implantation loss is an intra-uterine effect but seems partly to be attributable to direct post-partum cannibalism, increased at 150 mg/kg bw/day (highest dose tested).
CLINICAL SIGNS AND MORTALITY (F1 generation)
F1 males
Appearance, behaviour and mortality:
Control group: Four males died on day of age 139, 181, 126, and 170, respectively. Most had several ante-mortem observations, none considered test-article related and all had necropsy findings (lung) that were consistent with gavage injury. Discoloured urine was seen in one animal and anogenital staining in nine cases. Three rats had stained body surfaces.
25 mg/kg bw/day group: Two males died on study days 114 and 193, respectively. Both had several ante-mortem observations. None of which were considered treatment-related. Necropsy findings were consistent with gavage injuries. Increased salivation was seen in one animal, discoloured urine in two cases, and anogenital staining in five cases. Two rats showed a stained body surface.
75 mg/kg bw/day group: No F1 males died in this group. Six animals displayed increased salivation, 17 had anogenital staining, and two had stained body surfaces.
150 mg/kg bw/day group: Two males died on day of age 159 and 50, respectively, with test-article related ante-mortem observations (anogenital staining, body surface staining). Necropsy findings were consistent with gavage injuries. Three animals showed increased salivation, six showed discoloured urine, 23 had anogenital staining, and eleven had stained body surfaces. Food consumption: There were significant reductions (p≤ 0.05 and p≤ 0.01) of food consumption during the first two weeks of the generation for F1 males in the 75 and 150 mg/kg bw/day groups, respectively, that were attributed to the test article.

F1 females
Appearance, behaviour and mortality:
Control group: No mortalities occurred. One rat showed anogenital staining and one had a stained body surface.
25 mg/kg bw/day group: One female died on day of age 160 but had no visible ante-mortem observations. Necropsy findings were consistent with gavage injury. Increased salivation was seen in one animal and anogenital staining in two cases. One rat showed a stained body surface.
75 mg/kg bw/day group: Three females died in this group. One animal died on day of age 195, 194 and 48, respectively. Ante-mortem observations (anogenital staining, increased salivation and /or discoloured urine) were considered treatment related, but all three had necropsy findings consistent with gavage injuries. Increased salivation was seen in seven animals, discoloured urine in one, and anogenital staining in ten cases. Five rats showed a stained body surface.
150 mg/kg bw/day group: Five females died on day of age 118, 187, 187, 134 and 47, respectively. In two of them, necropsy findings were consistent with gavage injuries. In the remaining females, necropsy findings did not determine the cause of death. Increased salivation was seen in ten animals, discoloured urine in one, and anogenital staining in 22 cases. Eight rats showed stained body surfaces.

BODY WEIGHT / FOOD CONSUMPTION
F1 males
Mean weekly male body weights for the 75 and 150 mg/kg bw/day groups were significantly (p≤ 0.01) reduced every week measured from week of age 4 through 22 when compared to control means (Report, Fig. 4, Tab. 9) with mean body-weight gains of 85% and 74% of the control respectively. Weekly body weights were also consistently reduced for the 25 mg/kg bw/day males, but the mean body-weight gain until wk 22 remained above 90% of the control. These differences were considered treatment-related. The depression of growth was distinctly more conspicuous in the male F1- than in the P0-generation.

F1 females
Mean weekly body weights were significantly (p≤ 0.01) reduced relative to controls in the 150 mg/kg bw/day group during the pre-mating period. The 75 mg/kg bw/day group had consistently reduced body weights during the pre-mating period reaching significance (p≤ 0.05 and p≤ 0.01) on week of age 4-9, 20, and 23-24 relative to controls. Weekly body weights in the 25 mg/kg/ bw/day group were significantly (p≤ 0.05 and p≤ 0.01) reduced compared to controls for every week measured from week of age 4 through 24. These differences may be treatment related. Although the depression of growth was distinctly more conspicuous in the female F1- and P1- than in the maternal P0-generation, on the basis of body-weight gain, however, i.e. taking into account the divergent initial mean body weights, there is no evidence of a dose-response, while the weight gains remain above 90% of the control (Report, Fig. 4, Tab. 9).
During gestation, body weights for 150 mg/kg bw/day females relative to controls were significantly reduced (p≤ 0.05 and p≤ 0.01) for all time points measured. Body weight gain was depressed over gestation days 15-20. These reductions were considered test-article related. However, given the distinctly lower initial body weight in particular for the high-dose group (~250 g vs. ~290 g in the control), a physiologically significant impact of treatment on maternal body weight is not obvious in this phase, while the reduction in body-weight gain during the gestation days 15 to 20 seems to be attributable to foetal intra-uterine development.
This is different during lactation, where there is almost no change of the mean body weight in the 150 mg/kg bw/day group over time, body weight significantly (p≤ 0.01) different from control on lactation days 0, 14 and 21 (Report, Fig. 6). Body weights in the other groups were increasing; statistical significances have again to be weighed against the lower initial weights of the treated groups.

Food consumption:
There were significant reductions of food consumption during the first few weeks post-weaning for F1 males and females in the 150 mg/kg bw/day groups and only slightly for the 75 mg/kg bw/day groups in comparison to controls that were attributed to the test article (Report, Fig. 10, Tab. 15). However, it has to be taken into account that these results were related to rat weight rather than normalised to body weight.
During gestation, food consumption was significantly (p≤ 0.05) increased once over the gestational interval 0-6 for the 75 mg/kg bw/day dams. This was considered biological variability. No other differences were noted across groups over the entire gestational period.
During lactation intervals 0-7, 7-14 and 14-21, the 150 mg/kg bw/day dams showed reduced, only slightly increasing food consumption compared to controls. Food consumption distinctly increasing during lactation did not differ significantly across the other treatment groups.
Depression of food consumption was more conspicuous in the male and female F1- and P1-generation than in the parental P0-generation. This appears to directly correlate with growth depression.

REPRODUCTIVE PERFORMANCE
General: Diffuse and irregular rather than evaluable effects were observed in the P1-reproductive parameters, including mating, fertility, pregnancy, possibly influenced by treatment but not dose related hence not considered useful for toxicity assessment. Even though partly distinctly outside historical control values, these inconsistent data were not applicable. These unusual effects may be seen in relation to "subfertility problems in CD rats" recognised in previous years (note: authors´ statement in Report, 7.1 Conclusions, p. 37).

F1 females
The mean number of implantation sites (9.4 vs. 11.5 in the control, p<0.05) and the mean litter-size (delivered F2-offspring) (3.4 vs.9.9 in the control, p<0.01) were significantly reduced for the 150 mg/kg bw/day dams. Including the neo-natal deaths, the number of implantation sites NOT corresponding to live offspring of F2 (representing post-implantation loss also including possible cannibalisation at birth) was highly increased for the 150 mg/kg bw/day dams, i.e. 9.4 - 1.8 = 7.6 losses => ~81%. A similar less pronounced trend is observed only for the post-implantation loss of the F2-generation from the 75 mg/kg bw/day P1-dams, i.e. 12.5 - 7.3 = 5.2 losses => ~42%. These differences were deemed test-article related. No differences was noted in the 25-mg/kg bw/day P1-group.

F1 males
A significant and dose-dependent decrease in the mean ABSOLUTE testis weight was seen in the mid- and high-dose males when compared to controls. No difference was detected in the mean RELATVE testis weight across treatment groups. Consequently, the decrease in mean absolute testis weight is considered secondary to decreases in body weight and not a direct effect of the test article.

PATHOLOGY
F1 males and females
No test-article-related gross lesions were observed. Any isolated lesion observed in the thoracic cavity (clot, fluid, abscess, foreign material), lung (discolouration, abscess, adhesion, foreign material), heart (adhesion, discolouration, swelling, foreign material), oesophagus (perforation), and bone-rib/sternum (adhesion) in any group were considered as induced by the administration procedure (gavage) and not induced by the test article.
No test-article-related microscopic lesions were observed in animals that were either euthanised at study termination or died during the study period. Isolated lesions observed in the lung (haemorrhage, abscess, pleuritis), heart (pericarditis), oesophagus (inflammation), diaphragm (inflammation, abscess), soft tissue-thorax (abscess) and bone-sternum (abscess) were considered gavage-induced.
Key result
Dose descriptor:
NOAEL
Remarks:
systemic toxicity of P1
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
other: reduced food intake (males/females)
Key result
Dose descriptor:
NOAEL
Remarks:
prenatal and neonatal development of F2
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
reproductive performance
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
75 mg/kg bw/day (actual dose received)
System:
other: unspecific systemic toxicity (body weight, weight gain, food consumption)
Organ:
not specified
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
75 mg/kg bw/day (actual dose received)
System:
other: reproductive performance (decrease in live-birth index for F2-neonates and reduced number of live-born F2-pups)
Organ:
placenta
uterus
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Increased salivation in the mid- and high-dose group of both sexes, more pronounced in females;
Increased anogenital staining in the mid- and high-dose groups of both sexes; cases of emaciation in the male and female high-dose groups
Dermal irritation (if dermal study):
not examined
Mortality / viability:
mortality observed, treatment-related
Description (incidence and severity):
In the high-dose group, increased mortality of neonates (= decrease in live offspring/live birth-index on lactation day 0), and on lactation day 4 , significantly decreased viability (see Table under "Any other information on results ...").
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Mean F1 birth-weights were similar across all treated groups, with no statistical difference from the mean body-weight of the control group (Report, Vol. 1, Tab. 20). During lactation, growth became retarded in dose-related manner, already noted in the the F1-pups derived from the maternal low-dose P0-group.
From weaning, mean body growth was markedly depressed in dose-related manner for both sexes, but less pronounced for female F1 offspring (compare results P1, Report, Vol. 1, Fig. 4).
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
During the first weeks after weaning, food consumption was slightly reduced in dose-related manner, significant in the male and female mid- and high-dose groups, while compensated in the later pre-mating phase (Report, Vol. 1, Tab. 15, and Fig. 10).
Food efficiency:
not specified
Description (incidence and severity):
Food consumption was related to rat weight only.
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Sexual maturation:
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
The decrease in mean absolute testis weight is considered secondary to decreases in body weight and not a direct effect of the test article (see also results for P1).
Gross pathological findings:
no effects observed
Histopathological findings:
no effects observed
Other effects:
not specified
Behaviour (functional findings):
not examined
Developmental immunotoxicity:
not examined
see details under P1:
Pre- and neo-natal devolopment of the F1-offspring was significantly affected in high-dose-derived pubs (reduced number of live-born pubs/reduced live birth-index and reduced viability on lactation day 4). During lactation until weaning (days 14 and 21), mean pub growth also in the lower-treated groups as compared to the control.
Key result
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Generation:
F1
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
food consumption and compound intake
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
75 mg/kg bw/day (actual dose received)
System:
other: unspecific systemic toxicity (body weight, weight gain, food consumption)
Organ:
not specified
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Clinical signs:
no effects observed
Mortality / viability:
mortality observed, treatment-related
Description (incidence and severity):
see "Details on Results"
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
see "Details on Results"
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Sexual maturation:
no effects observed
Description (incidence and severity):
Related to sex ratio: no effect, close to 50 : 50 in all groups
Gross pathological findings:
no effects observed
Behaviour (functional findings):
not examined
Developmental immunotoxicity:
not examined
F2 offspring:
Offspring survival:
Total litter size was greatly reduced for the 150 mg/kg bw/day dams compared to control dams (3.4 vs. control mean = 9.9). A high number of stillborns (litter-size minus live offspring) for the 75 and 150 mg/kg bw/day litters (means = 2.7 and 1.6, respectively) resulted in the mean number of viable offspring on lactation day 0 for these two groups being reduced further to 7.3 and 1.8 (p≤ 0.05).
That means, on day 0 (parturition), the live birth-index became significantly reduced for 75 and 150 mg/kg bw/day relative to the controls with p<0.05 and <0.01, respectively.
Viability by day 4 was not significantly different between all groups, although among the 25 mg/kg bw/day pups a somewhat increased loss of 15 was observed.

Offspring growth:
Neonatal mean body-weights were normal across all groups. Reduction in F2-offspring body weights occurred in the 150 mg/kg bw/day pups beginning day 7 until weaning, reaching statistical significance for the females by day 21 (p<0.05). Mean body weights of the 75 mg/kg bw/day pups at weaning were slightly less than controls without statistical significance. These differences were considered test-article related. No weight effect was seen in the low-dose F2-pups.

Offspring assessment:
No differences between groups in test-article-related ante-mortem findings were found during the lactation period.
Key result
Dose descriptor:
NOAEL
Remarks:
developmental / systemic toxicity
Generation:
F2
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
mortality
body weight and weight gain
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
75 mg/kg bw/day (actual dose received)
System:
other: develomental toxicity (reduced neonatal survival / growth depression during lactation)
Organ:
not specified
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Key result
Reproductive effects observed:
yes
Lowest effective dose / conc.:
75 mg/kg bw/day (actual dose received)
Treatment related:
yes
Relation to other toxic effects:
reproductive effects occurring together with other toxic effects, but not as a secondary non-specific consequence of other toxic effects
Dose response relationship:
yes
Relevant for humans:
not specified

Table for reproductive toxicity study (York 1995: Two-Generation Study)

A.   Food consumption vs. body weight: P0 parents vs. F1 offspring

 

 

Control

Low dose

Mid dose

High dose

Generation

m

f

m

f

m

f

m

f

Food consumption
(Tab.12/Fig.7 and Tab.15/Fig.10)

(Weeks indicated represent weeks on study for the P0 and age for F1. For the dams, the time covers premating and mating period until copulation.)

 

Quali-tatively versus control (—)

P0

wk 0-15

wk 0-8

(↑)

↑*

↑*

↑**

F1

wk 4-22

wk 4-22

↓*

↓*

↓*

↓*

Body weight gain
(Tab.6/Fig.1 and Tab.9/Fig.4)

(Explanation of weeks see above)

 

% of control

P0

100

wk 0-15

100

wk 0-8

95

94

94

101

86*

100

F1

100

wk 4-22

100

wk 4-22

93

91

85*

99

74**

92*

B.    Reproductive Performance of first(P0) and second(P1) generation

 

 

Control

Low dose

Mid dose

High dose

Generation

m

f

m

f

m

f

m

f

Mating index

%

P0

96.0

96.2

100

100

100

100

100

100

P1

91.7

92.3

83.3

85.2

60.0*

60.0**

82.6

87.0

Fertility index
(m. fertile/m. paired)
(f. pregnant/f. paired)
(Tab. 18 / 23)

%

P0

88.0

88.5

76.9

76.9

83.3

83.3

84.6

84.6

P1

50.0

61.5

29.2

25.9*

28.0

28.0*

47.8

47.8

Pregnancy index
(f. pregn./f. mated)

(Tab. 18 / 23)

%

P0

--

92

--

76.9

--

83.3

--

84.6

P1

--

66.7

--

30.4*

--

46.7

--

55

Number of implantation sites (Tab. 28 / 29)

Mean

P0

 

14.0

 

13.4

 

13.2

 

12.5

P1

 

11.5

 

12.3

 

12.5

 

9.4

Duration of pregnancy
(Tab. 18 / 23)

Mean (d)

P0

 

22.1

 

22.4

 

22.4

 

22.9

P1

 

22.4

 

22.3

 

22.8

 

23.4

Litter size from P0 + P1 (Tab. 19 + 24, resp.)

Mean

F1

13.2

12.3

11.5

9.2* #

F2

9.9

11.7

10.0

3.4** #

Live offspr. from P0 + P1 (Tab. 19 + 24, resp.)

Mean

F1

13.0

12.2

11.1

6.8**

F2

9.6

11.1

7.3* #

1.8** #

Live birth index (d0)
(Tab. 19 / 24)

%

F1

98.6

99.1

96.1

73.7**

F2

96.8

95.1

73.3*

51.6**

Viability index (d4)
(Tab. 19 / 24)

%

F1

98.5

94.4

85.4

76.9**

F2

85.6

80.8

100

87.5

Weight of live pups on day of age 0 (Tab. 20 / 25)

Mean (g)

F1

6.1

6.0

5.7

5.9

F2

6.2

5.8

6.0

6.2

Weight of live pups on day of age 14 (Tab.20 / 25)

Mean (g)

F1

34.2

31.0**

29.0**

24.8**

F2

31.0

32.6

27.6

23.9*

Weight of live pups on day of age 21 (Tab 20 / 25)

Mean (g)

F1

56.4

53.6

51.4**

49.9*

46.3**

44.7**

39.8**

39.0**

F2

51.1

48.4

53.3

51.1

46.1

44.5

39.3

38.0*

*significantly different from controls, p≤ 0.05 /**significantly different from controls, p≤ 0.01

  Note: in the report, Tab. 19 and 24, no statistical significance stated, and for 1.8 only p<0.05 given. This appears to be erroneous and is assumed to be p<0.05 and 0.01, respectively.

 

Conclusions:
Creosote showed toxic effects on reproduction (F1: reduced number of pups at mid- and high-dose; increased mortality of mid- and high-dose groups. F2: reduced litter size, reduced viability at high-dose) in the presence of mild maternal toxicity. Further, body weight of F2 live pups was dose-dependently reduced at days 14 and 21 post-partum.
Executive summary:

In an OECD 416 guideline study conducted under GLP conditions, US Creosote P1-13 (0, 25, 75, 150 mg/kg bw/day) showed toxic effects on reproduction in rats in the presence of mild maternal toxicity (F1: at the high dose, reduced number of live pups on d0 and d4, reduced litter size of neonates, reduced d4-viability, and in the mid-and high-dose groups, depressed body-weight gain; F2: at the mid- and high dose, reduced number of live pups on d0, and in addition at the high dose, reduced litter size, and depressed body-weight gain during lactation). The NOAELs are as follows:

Parental (P0 generation) NOAEL: 75 mg/kg bw/day, based on decreased body-weight gain in the high-dosed groups (in P1 parents and during gestation in P0 parents).

Pup NOAEL: 25 mg/kg bw/day, based on decreased litter size in the high-dose group, decreased survival and an increase in stillborns, and decreased body-weight of live pups in the two highest dose groups (less than 10 % bw reduction in the low dose group) (York, 1995b).

The study is considered to be acceptable, although some examinations (oestrus cycle, sperm parameters, extensive histopathology) were not performed according to guideline.
Endpoint:
two-generation reproductive toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The source test material creosote (US type P1/P13) consists predominantly of polycyclic aromatic hydrocarbons (PAH) ranging in size from two up to five fused rings. The target substance anthracene oil (anthracene oil with < 50 ppm benzo[a]pyrene (BaP), AOL) is as well composed of a broad range of PAH but predominantly consisting of two to four aromatic rings.
The nature of both substances and their constituents are considered to be sufficiently similar that toxic effects on reproduction following repeated oral exposure are equivalent. Therefore, the source substance is suited as supporting substance with regard to reproduction toxicity and data resulting from the source substance can be used for characterising toxic properties on fertility of the target substance anthracene oil upon oral application.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The source material creosote (US type P1/P13) is a condensation product in the distillation of coal tars that have been obtained in the high temperature carbonisation of bituminous coal. The material is a UVCB substance forming a dark brown oily liquid. It is only partly volatile and consists of a complex mixture of polycyclic aromatic hydrocarbons with no or only a minimal content of other components (phenols). Two- and three-ring aromatics amount to about 40 % (typical concentration) with two-ring aromatics forming the smaller fraction. PAH with four and more rings accumulate to about 15 %. Five-ring PAH are only present in small individual concentration (below 0.3 %). The water solubility of creosote is relatively low. It is determined by the solubility properties of its constituents.
The target material anthracene oil (AOL) is a UVCB substance as well produced by the distillation of coal tars extracting the approximate distillation range from ca. 300 °C to 400 °C. 10 % to 95 % of the total product distil over between ca. 300 °C and 375 °C. The substance is a brown pasty or liquid material consisting of a complex and within limits variable combination of polycyclic aromatic hydrocarbons. The distillation range excludes mostly low molecular weight aromatic hydrocarbons (especially one-ring and to a lower extent two-ring aromatics) as well as polycyclic aromatic hydrocarbons composed of more than four to five rings depending on the respective boiling points of the individual aromatic substances. Two- and three-ring aromatics amount to about 50 % (typical concentration) with two-ring aromatics forming the smaller fraction. PAH with four and more rings accumulate to about 10 % with pyrene and benzofluorenes representing the highest molecular weight PAH found in AOL. The water solubility of AOL is low being limited by the solubility properties of its constituents.

3. ANALOGUE APPROACH JUSTIFICATION
Toxic effects on reproduction after oral application will depend on the pharmacokinetic and the specific toxicological properties of a substance. Systemic toxic effects will be caused after resorption from the gastrointestinal tract, metabolic activation, and distribution within the body. In the case of complex substances like UVCBs, the overall toxicity will result from the effects caused by the individual components of the substance. If substances are similar, the systemic oral toxicity including toxicity to reproduction will be comparable.
The composition of the source and the target substance is somewhat different creosote having a broader spectrum of PAH with a higher concentration of two-ring and a smaller concentration of three-ring PAH compared to AOL. The amount of four-ring PAH is similar for both substances, but creosote contains a broader range of five-ring PAH (low individual concentrations). Nevertheless, the same PAH are basically present in both substances and their consistency is quite similar.
Constituents of both, the source as well as the target substance, are members of the same chemical class (PAH). As illustrated in Chapter 5.1, toxicokinetic properties are very similar for different compounds within this chemical class. Similar PAH have similar metabolic activation pathways resulting in similar final toxicants. Even if the composition of both substances is somewhat different, the overall total effect on toxicity to reproduction are considered to be comparable. Using creosote as source substance may even be a conservative approach because of the broader compositional profile of creosote.
Overall, taking into account the combined information on the properties of creosote and anthracene oil, differences in the PAH composition of both materials are considered not to modify reprotoxic effects of the source and the target material significantly. Within a range, effects are considered to be comparable. Reprotoxic effects observed upon oral application of creosote are assumed to similarly develop from oral exposure to AOL. For these reasons, it is considered justified to use reprotoxicity data of creosote in order to characterise the toxicity to reproduction of anthracene oil.
Reason / purpose for cross-reference:
read-across source
Justification for study design:
Read-across to preceding entry:
Source test material: US Creosote P1/P13;
Reference: York 1995
Key result
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Effect level:
75 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: hypersalivation; pre-mating bodyweight loss (males); reduced body-weight of dams during gestation and lactation
Remarks on result:
other: the test result of the source substance is adopted for the target substance anthracene oil
Key result
Dose descriptor:
NOAEL
Remarks:
fertility and prenatal development
Effect level:
75 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
reproductive performance
Remarks on result:
other: the test result of the source substance is adopted for the target substance anthracene oil
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
150 mg/kg bw/day (actual dose received)
System:
other: unspecific systemic toxicity (hypersalivation; reduced premating body-weight of males; reduced weight in dams during gestation and lactation)
Organ:
not specified
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
150 mg/kg bw/day (actual dose received)
System:
other: reproductive performance (reduced F1-litter size and live-birth index (post-implantation losses)
Organ:
placenta
uterus
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Haematological findings:
no effects observed
Description (incidence and severity):
Isolated lesions were caused mechanically due to gavage dosage.
Key result
Dose descriptor:
NOAEL
Remarks:
systemic toxicity P1
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
other: reduced food intake (males/females)
Remarks on result:
other: the test result of the source substance is adopted for the target substance anthracene oil
Key result
Dose descriptor:
NOAEL
Remarks:
prenatal and neonatal development of F2
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
reproductive performance
Remarks on result:
other: the test result of the source substance is adopted for the target substance anthracene oil
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
75 mg/kg bw/day (actual dose received)
System:
other: unspecific systemic toxicity (body weight, body weight gain, food consumption)
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
75 mg/kg bw/day (actual dose received)
System:
other: reproductive performance (decrease in live-birth index for F2-neonates and reduced number of live-born F2-pups)
Organ:
placenta
uterus
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Key result
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Generation:
F1
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
food consumption and compound intake
Remarks on result:
other: the test result of the source substance is adopted for the target substance anthracene oil
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
75 mg/kg bw/day (actual dose received)
System:
other: unspecific systemic toxicity (body weight, weight gain, food consumption)
Organ:
not specified
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Key result
Dose descriptor:
NOAEL
Remarks:
developmental / systemic toxicity
Generation:
F2
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
mortality
body weight and weight gain
Remarks on result:
other: the test result of the source substance is adopted for the target substance anthracene oil
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
75 mg/kg bw/day (actual dose received)
System:
other: developmental toxicity (reduced neonatal survival / growth depression during lactation)
Organ:
not specified
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified
Key result
Reproductive effects observed:
yes
Lowest effective dose / conc.:
75 mg/kg bw/day (actual dose received)
Treatment related:
yes
Relation to other toxic effects:
reproductive effects occurring together with other toxic effects, but not as a secondary non-specific consequence of other toxic effects
Dose response relationship:
yes
Relevant for humans:
not specified
Conclusions:
As data on toxicity to reproduction for anthracene oil are not available, the structure-related tar oil US creosote P1-13 is used as substitute/supporting substance. In an OPP 83-4/OECD 416 study in rats under GLP conditions with this test material (source substance), toxic effects on reproduction in the presence of mild systemic parental toxicity were observed. Reproductive performance was impeded appearing as reduction in litter size and in live-birth index of the P0 and P1 generation. Systemic parental toxicity showed as reduced body weight and weight gain, and as reduced food consumption. The F2 generation showed similar symptoms. Mortality was increased compared to controls (decrease in live-birth index and in the number of live-born pups) and weight development until weaning was also depressed. NOAELs (reproduction and systemic toxicity) for the P1 generation were 75 mg/kg bw/day, while NOAELs (reproduction and systemic toxicity) for the F1 and F2 generation were 25 mg/kg bw/day.
The test results of the source material are adopted for the target substance anthracene oil.
Effect on fertility: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
25 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
high
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

The results were obtained from a structure-related tar oil (US creosote P1/13). Read-across to this test material is considered to be a conservative approach as compared to the anthracene oil because of the broader compositional profile of the US creosote used as read-across tar oil.

No typical adverse effect on reproductive performance of either sex was noted; the adverse effects observed appear to relate to pre- and post-natal development of the offspring rather than impairment of fertility.

The apparent inconsistencies in the reproduction parameters having emerged in the second generation (P1) (mating, fertility, pregnancy) have to be weighted in relation to "subfertility problems in CD rats" recognised in previous years (note: authors´ statement in Report, 7.1 Conclusions, p. 37): Parameters already poor in the control groups were irregularly decreased in the treated groups frequently outside the historical control data-ranges, however without showing any dose-response, but nonetheless possibly influenced by exposure. This dubious data was not adopted as adverse and skipped for deriving the NOAELs.

Effects on developmental toxicity

Description of key information

In a developmental/teratogenicity study (OPP 83-3/OECD 414), an increase in the incidence of post-implantation losses (early resorptions) and a reduction in the number of live foetuses was seen at the highest dose tested of 175 mg/(kg bw*d) given orally to pregnant rats from gestation day 6 through 15, including three whole-litter losses and one residual litter with only one live foetus left.

No adverse effect was demonstrated for the late pre-natal development of live foetuses at any dose.

Link to relevant study records

Referenceopen allclose all

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
EPA OPP 83-3 (Prenatal Developmental Toxicity Study)
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
as of 12 May 1981
GLP compliance:
yes
Limit test:
no
Specific details on test material used for the study:
- Name of test material: Creosote; North American P1/P13 Creosote; North American Creosote Composite Test Material P1/P13
- Composition of test material, percentage of components: see under Test material information
Species:
rat
Strain:
Sprague-Dawley
Remarks:
SD Crl:CD® VAF/Plus®
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Portage, Michigan, USA
- Age at study initiation: approx. 13 weeks
- Weight at study initiation: 210 – 278 grams
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
VEHICLE
- Justification for use and choice of vehicle (if other than water): solubilty of test material
- Concentration in vehicle: 2.5, 5.0, 17.5 mg/mL
- Amount of vehicle (if gavage): 10 mL/kg bw/day
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentrations, homogeneity, and stability in the vehicle examined (Report, Vol. 1, Chap. 5.1 "Analytical Results" and Tab. 1-3)
Details on mating procedure:
Until detection of copulatory plug (= day 0 of gestation).
Duration of treatment / exposure:
day 6-15 of gestation
Frequency of treatment:
once daily
Duration of test:
Total duration: 20 days
Exposure period: day 6 - 15 of gestation
Postexposure period: day 16 - 20 of gestation
Dose / conc.:
25 mg/kg bw/day (actual dose received)
Dose / conc.:
50 mg/kg bw/day (actual dose received)
Dose / conc.:
175 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
30 mated females; effectively, 27, 20, 23 and 23 pregnant animals were available in the vehicle control and respective treated groups.
Control animals:
yes, concurrent vehicle
Details on study design:
Sex: female
Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily on days 6-20

DETAILED CLINICAL OBSERVATIONS: Yes

BODY WEIGHT: Yes
- Time schedule for examinations: days 0, 6, 9, 12, 16 and 20 (Report, Vol.I, Tab. 5/6, p. 36/37)

FOOD CONSUMPTION: yes (Report, Vol.I, Tab. 7, p. 38)

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20: Caesarean section (Report, Vol.I, Tab. 8, p. 39-41)
- Organs examined: uterus

Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes, Report, Vol.I, Tab. 8, p. 39-41
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
- External examinations: Yes
- Soft tissue examinations: Yes (1/3 per litter)
- Skeletal examinations: Yes (1/3 per litter)
- Head examinations: Yes

Litter size, number of dead foetuses, foetal weight, crown-rump length, sex ratio, external alterations

Histopathological, microscopic examination for malformations was performed on 121 and 116 foetuses from 20 litters at the the top-dose level, 150 and 161 from 23 litters at the mid-dose level (Report, Vol 1, Tab. 9/10, p. 42/43). 

Statistics:
Male and female sex ratios and the proportions of litters with malformations and developmental variations were compared using the Chi-square test-criterion with Yates´ correction for 2x2 contingency tables and/or Fisher´s exact probability test to determine the significance of differences (see further details: Statistical methods summarised in Report, Vol. I, Chap. 4.7.).
Indices:
yes
Historical control data:
yes, comprehensive compilation in Report, Vol. I, Appendix D
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
increased hair loss during gestation days 6-9
Dermal irritation (if dermal study):
not specified
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
slightly depressed growth probably correlated with reduced food consumption. Overall, in the high-dose group, there was a decreasing trend only - without clear statistical significance (Report, Tab. 6, Tab. 7). No evidence of maternal toxicity was observed at the 50 mg/kg/day dosage level or lower.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not specified
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
not specified
Number of abortions:
no effects observed
Pre- and post-implantation loss:
effects observed, treatment-related
Description (incidence and severity):
Increase in embryonal losses, early resorption
Total litter losses by resorption:
effects observed, treatment-related
Description (incidence and severity):
Resorption of 3 whole liters / 20 dams (total) and 1 litter / 20 dams (only one viable foetus left) at the high dose level (175 mg/kg bw/d)
Early or late resorptions:
effects observed, treatment-related
Description (incidence and severity):
increased post-implantation loss (early resorptions) in the maternal high-dose group.
Dead fetuses:
no effects observed
Description (incidence and severity):
There were no dead or non-viable foetuses at Cesaeran section across all groups.
Changes in pregnancy duration:
no effects observed
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed
Changes in number of pregnant:
no effects observed
Other effects:
not specified
Details on maternal toxic effects:
Maternal toxic effects: yes

Systemic toxicity:
Hair loss, body weight loss, reduced food consumption at 175 mg/kg bw/day. But depression of maternal body-weight gain was not statistically significant, and there was no difference across the other groups, when corrections were made for gravid uterine weight.

Developmental toxicity (pre-natal):
At the maternal 175 mg/kg bw/day dosage level, there was clear evidence of a statistically significant increase in the incidence of post-implantation loss (including three whole-litter and one close-to-total resorptions vs. two litters resorbed in the vehicle control and none in the other treated  groups). No evidence of maternal or developmental toxicity was observed at the 50 mg/kg bw/day dosage level or lower (Report, Vol. I, Tab. in Appendix C, p. 79-86).
Key result
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
body weight and weight gain
other: hair loss, reduced food consumption
Key result
Dose descriptor:
NOAEL
Remarks:
pre-natal development
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
early or late resorptions
pre and post implantation loss
total litter losses by resorption
Key result
Abnormalities:
effects observed, treatment-related
Localisation:
other: systemic toxicity
Description (incidence and severity):
hair loss, food consumption, body weight and weight gain
Key result
Abnormalities:
effects observed, treatment-related
Localisation:
placenta
uterus
Description (incidence and severity):
post-implantation loss (early resorptions)
Fetal body weight changes:
no effects observed
Description (incidence and severity):
See Tab. 2 under "Any other information on results ..."
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): not examined
Reduction in number of live offspring:
effects observed, treatment-related
Description (incidence and severity):
In the maternal high-dose group, the number of total/live foetuses was decreased, yet not statistically significant (see Tab. 2 under "Any other information on results ...").
This was attributable to an increase in post-implantation loss (increase in early resorptions), statistically significant (p<0.05).
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
effects observed, treatment-related
Description (incidence and severity):
In the maternal high-dose group, gravid uterine weight was decreased, yet not statistically significant (see Tab. 2 under "Any other information on results ...")
Changes in postnatal survival:
not examined
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Apparent increases in the incidences of malformations were not statistically significant and are considered to have occurred by chance. 
The incidences of morphological variations were not different from those seen in the vehicle control.
See details under "Any other information on results ..."
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
see details under "Any other information on results ..."
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
see details under "Any other information on results ..."
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects: no test-article related effects

Foetotoxicity:
no effects observed

Malformations:
One, six, seven, and seven foetuses showed malformations in one, three, six, and five litters for the control, low-, mid-, and high-dose groups, respectively. The incidences for the mid-, and high-dose levels were significantly higher than for the control group. Most of the observed malformations are fairly common in rats and the values obtained for the eye malformations in this study were within historical control range. After factoring out these common eye abnormalities, each type of the remaining malformations occurred in only one or two instances per group. Thus, the malformations were not considered to be test-article related. (Report, Vol. I, Chap. 5.4).
Key result
Dose descriptor:
NOAEL
Remarks:
highest dose tested
Effect level:
175 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: foetotoxicity
Key result
Dose descriptor:
NOAEL
Remarks:
highest dose tested
Effect level:
175 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
external malformations
skeletal malformations
visceral malformations
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
yes
Lowest effective dose / conc.:
175 mg/kg bw/day
Treatment related:
yes
Relation to maternal toxicity:
developmental effects occurring together with maternal toxicity effects, but not as a secondary non-specific consequence of maternal toxicity effects
Dose response relationship:
yes
Relevant for humans:
not specified

 

 Table 1: Maternal Effects

Parameter

Control data

25 mg/kg bw/day

50 mg/kg bw/day

175 mg/kg kg/day

Dose-response
+ / –

historical

study

Number of dams examined

716

30

30

30

30

 

Pregnant females

 

27

20

23

23

 

Pregnancies [%]

 

90.0

66.0

76.0

76.0

Clinical findings during application of test substance

 

13

15

11

17

Mortality of dams [%]

0

0

0

0

0

Body weight gain

 

day 0-6

 

34

34

32

30

day 6-9

 

2

5

3

-2

day 9-12

 

12

12

13

12

day 12-16

 

28

21

25

21

day 16-20

 

57

65

64

51

day 6-16

 

42

38

40

31

day 0-20

143

133

137

136

112

Final dam weight change minus weight of uterus

62

64

67

66

61

Food consumption [g/animal/day]

 

 

 

 

 

 

day 0-6

 

21.2

21.0

20.2

20.5

day 6-9

 

14.6

16.7

14.4

12.9

day 9-12

 

14.1

15.5

14.1

13.6

day 12-16

 

17.9

15.0

12.6*

9.8**

+

day 16-20

 

25.5

28.6

29.5

30.4

day 6-16

 

15.7

15.7

13.6

11.9**

+

day 0-20

 

19.4

19.9

18.8

18.2

Necropsy findings in dams dead before end of test

 

No deaths occurred before end of test.

* Significantly different from controls, p ≤ 0.05
** Significantly different from controls, p ≤ 0.01

 

Table 2: Developmental Effects

Litter response (Caesarean section data)

Parameter

Control data

25 mg/kg/day

50 mg/kg/day

175 mg/kg/day

Dose- response
+ / -

historical

study

Corpora lutea:mean/dam

16.8

16.6

16.0

16.3

16.0

Implantations:mean/dam

15.0

14.2

13.8

14.6

12.9

Resorptions:total/number of dams

850/708

31/27

17/20*

25/23

59/23*

+

Number of foetuses/dam

 

13.0

13.0

13.5

10.3

Pre-implantation loss [%]

 

11.6

13.8

10.6

12.2

Post-implantation loss [%]

7.3

8.1

6.2

7.4

19.9 *

+

Total number of litters

 

26

20

23

20

Foetuses / litter

 

13.0

13.0

13.5

10.3

Live foetuses / litter ratio

13.9

13.0

13.0

13.5

10.3

Dead foetuses / litter ratio

 

0

0

0

0

Foetus weight (mean) [g]

3.4

3.3

3.5

3.3

3.2

Male foetuses

 

3.4

3.6

3.4

3.3*1)

+

Female foetuses

 

3.3

3.3

3.2

3.2

Uterine weight (mean) [g]

76.3

72.0

69.5

70.0

61.4

Crown-rump length (mean)[mm]

 

35

36

35

35

Foetal sex ratio[state ratio m/f]

0.982

1.108

0.904

1.006

1.135

* Significantly different from controls, p ≤ 0.05

1)questionable, no effect on crown length, see also female values, considered biologically irrelevant.

Table 3: Morphological Effects

Examination of the foetuses (Report, Tab. 9 + 10, p. 42/43)

Parameter

Control data

25 mg/kg/ day

50 mg/kg/ day

175 mg/kg/ day

Dose-response
+ / -

historical

study

External malformations*

[%]

0.20

0.00

0.77

2.891)

1.27

Skeletal malformations*

[%]

0.93

0.00

0.81

4.351)

1.72

Visceral malformations*

[%]

0.41

0.58

2.21

1.33

2.50

Foetuses with malformations [%]#

 

0.58

3.8

3.92) 3)

5.53)

 

 

 

 

 

 

 

External variations*

[%]

0.01

0.00

0.00

0.00

0.00

Skeletal variations*

[%]

34.4

36.7

50.4

49.7

43.1

Visceral variations*

[%]

1.44

10.5

11.0

7.33

1.67

Foetuses with variations [%]#

 

19.0

24.3

24.8

19.4

*Percentages are not given in the original study but calculated from the number of observations and the number of examined foetuses in the respective category.

# Percent foetuses with malformations or variation are calculated from total foetuses with defects from Tab.9 or 10, respectively.

1)The relatively high incidences are only determined by an accumulation of 11 different skeletal and external malformations within one foetus from doe No. 56513 (Report, Appendix C, p. 100).

2)Because of multi-fold malformations observed in two foetuses (one with 4 and a second with 2 defects), the percentage of the number of animals with malformations is lower than the total percentage of malformations (comp. Report,Appendix C, p. 98ff). Four other foetuses carried only one structural defect as in the other dose groups.

3)Statistically significant increases in the number of affected litters as compared to control, no increase in defect incidence per litter (p<0.05) (Report, Vol. 1, Tab.9).

Conclusions:
US creosote P1-13 was embryotoxic (early resorptions) with a NOAEL of 50 mg/kg bw in the presence of mild maternal toxicity (NOAEL also 50 mg/kg bw). It is unlikely that the increase in post-implantation losses are coupled to the decreased maternal food consumption, while there were virtually no other maternal toxic signs. Results indicate classification as reprotoxic cat. 2, "H361: Suspected of damaging fertility or the unborn child."
Executive summary:

The developmental toxicity of US Creosote P1/13 (0, 25, 50, and 175 mg/kg bw/day, gestational day 6 -15) was tested under GLP conditions in the rat according to OECD 414. Weak maternal toxicity and significantly increased early embryo resorption was seen at the top dose. Hence, the NOAEL was 50 mg/kg bw/day for both maternal and embryo toxicity.

Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The source test material creosote (US type P1/P13) consists predominantly of polycyclic aromatic hydrocarbons (PAH) ranging in size from two up to five fused rings. The target substance anthracene oil (anthracene oil with < 50 ppm benzo[a]pyrene (BaP), AOL) is as well composed of a broad range of PAH but predominantly consisting of two to four aromatic rings.
The nature of both substances and their constituents are considered to be sufficiently similar that toxic effects on reproduction (developmental toxicity) following repeated oral exposure during gestation are equivalent. Therefore, the source substance is suited as supporting substance with regard to developmental toxicity and data resulting from the source substance can be used for characterising adverse developmental effects of the target substance anthracene oil.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The source material creosote (US type P1/P13) is a condensation product in the distillation of coal tars that have been obtained in the high temperature carbonisation of bituminous coal. The material is a UVCB substance forming a dark brown oily liquid. It is only partly volatile and consists of a complex mixture of polycyclic aromatic hydrocarbons with no or only a minimal content of other components (phenols). Two- and three-ring aromatics amount to about 40 % (typical concentration) with two-ring aromatics forming the smaller fraction. PAH with four and more rings accumulate to about 15 %. Five-ring PAH are only present in small individual concentration (below 0.3 %). The water solubility of creosote is relatively low. It is determined by the solubility properties of its constituents.
The target material anthracene oil (AOL) is a UVCB substance as well produced by the distillation of coal tars extracting the approximate distillation range from ca. 300 °C to 400 °C. 10 % to 95 % of the total product distil over between ca. 300 °C and 375 °C. The substance is a brown pasty or liquid material consisting of a complex and within limits variable combination of polycyclic aromatic hydrocarbons. The distillation range excludes mostly low molecular weight aromatic hydrocarbons (especially one-ring and to a lower extent two-ring aromatics) as well as polycyclic aromatic hydrocarbons composed of more than four to five rings depending on the respective boiling points of the individual aromatic substances. Two- and three-ring aromatics amount to about 50 % (typical concentration) with two-ring aromatics forming the smaller fraction. PAH with four and more rings accumulate to about 10 % with pyrene and benzofluorenes representing the highest molecular weight PAH found in AOL. The water solubility of AOL is low being limited by the solubility properties of its constituents.

3. ANALOGUE APPROACH JUSTIFICATION
Developmental toxicity after oral application during gestation will depend on the pharmacokinetic and the specific toxicological properties of a substance. Systemic toxic effects will be caused after resorption from the gastrointestinal tract, metabolic activation, and distribution into the relevant organs. In the case of complex substances like UVCBs, the overall toxicity will result from the effects caused by the individual components of the substance. If substances are similar, the systemic toxicity including developmental toxicity will be comparable.
The composition of the source and the target substance is somewhat different creosote having a broader spectrum of PAH with a higher concentration of two-ring and a smaller concentration of three-ring PAH compared to AOL. The amount of four-ring PAH is similar for both substances, but creosote contains a broader range of five-ring PAH (low individual concentrations). Nevertheless, the same PAH are basically present in both substances and their consistency is quite similar.
Constituents of both, the source as well as the target substance, are members of the same chemical class (PAH). As illustrated in Chapter 5.1, toxicokinetic properties are very similar for different compounds within this chemical class. Similar PAH have similar metabolic activation pathways resulting in similar final toxicants. Even if the composition of both substances is somewhat different, the overall total effect on foetal developmental is considered to be comparable. Using creosote as source substance may even be a conservative approach because of the broader compositional profile of creosote.
Overall, taking into account the combined information on the properties of creosote and anthracene oil, differences in the PAH composition of both materials are considered not to modify reprotoxic effects of the source and the target material significantly. Within a range, effects are considered to be comparable. Developmental toxicity observed upon oral application during gestation of creosote is assumed to similarly appear after oral exposure to AOL. For these reasons, it is considered justified to use developmental toxicity data of creosote in order to characterise adverse effects on foetal development/embryotoxicity and teratogenic effects of anthracene oil.
Reason / purpose for cross-reference:
read-across source
Principles of method if other than guideline:
Read-across to preceding entry:
Source test material: US Creosote P1/P13
Reference: York 1995
Key result
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
body weight and weight gain
other: hair loss, reduced food consumption
Remarks on result:
other: the test result of the source substance is adopted for the target substance anthracene oil
Key result
Dose descriptor:
NOAEL
Remarks:
pre-natal development
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
early or late resorptions
pre and post implantation loss
total litter losses by resorption
Remarks on result:
other: the test result of the source substance is adopted for the target substance anthracene oil
Key result
Abnormalities:
effects observed, treatment-related
Localisation:
other: systemic toxicity
Description (incidence and severity):
hair loss, food consumption, body weight and weight gain
Key result
Abnormalities:
effects observed, treatment-related
Localisation:
placenta
uterus
Description (incidence and severity):
post-implantation loss (early resorptions)
Key result
Dose descriptor:
NOAEL
Remarks:
highest dose tested
Effect level:
175 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: foetotoxicity
Remarks on result:
other: the test result of the source substance is adopted for the target substance anthracene oil
Key result
Dose descriptor:
NOAEL
Remarks:
highest dose tested
Effect level:
175 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
external malformations
skeletal malformations
visceral malformations
Remarks on result:
other: the test result of the source substance is adopted for the target substance anthracene oil
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
yes
Lowest effective dose / conc.:
175 mg/kg bw/day
Treatment related:
yes
Relation to maternal toxicity:
developmental effects occurring together with maternal toxicity effects, but not as a secondary non-specific consequence of maternal toxicity effects
Dose response relationship:
yes
Relevant for humans:
not specified
Conclusions:
As data on developmental toxicity for anthracene oil are not available, the structure-related tar oil US creosote P1-13 is used as substitute/supporting substance. In a OECD 414 study with this test material (source substance) under GLP conditions, a clear embryotoxic effect (early resorptions) was observed together with mild maternal toxicity (food consumption). NOAEL for both effects was 50 mg/kg bw/day. It is unlikely that the increase in post-implantation losses are coupled to the decreased maternal food consumption, while there were virtually no other maternal toxic signs. Results indicate classification of the test material as reprotoxic cat. 2, "H361: Suspected of damaging fertility or the unborn child."
The test results of the source material are adopted for the target substance anthrancene oil.
Effect on developmental toxicity: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
50 mg/kg bw/day
Study duration:
subacute
Species:
rat
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

The results were obtained from a structure-related tar oil (US creosote). Read-across to this test material is considered to be a conservative approach as compared to the anthracene oil, because of the broader compositional profile of the US creosote used as read-across tar-oil.

Justification for classification or non-classification

The results are accepted as relevant for the classification of anthracene oil (BaP < 50 ppm). The adverse effects observed require the classification for potential hazards:

According to Regulation (EC) No 1272/2008 as follows:

      for possible impairment of fertility, reproduction and development: Repr. 2, H361: Suspected of damaging fertility or the unborn child.

Additional information