Registration Dossier

Diss Factsheets

Administrative data

Endpoint:
short-term repeated dose toxicity: dermal
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Cross-referenceopen allclose all
Reason / purpose for cross-reference:
data waiving: supporting information
Reference
Endpoint:
short-term repeated dose toxicity: oral
Remarks:
Combined repeated dose toxicity and reproductive and developmental toxicity study
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
2005
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
KL2 due to RA
Justification for type of information:
Refer to section 13 of IUCLID for details on the read-across justification. The study with the read across substance is considered sufficient to fulfil the information requirements as further explained in the provided endpoint summary.
Reason / purpose for cross-reference:
read-across source
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
not specified
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
- Seventy each of 8-week-old male and female Sprague-Dawley SPF rats (Crj;CD (SD) IGS, Charles River Laboratories Japan, Inc., Atsugi Breeding Center) were purchased (number of animals received: 73 each of males and females)
- The animals were quarantined and acclimatized for 14 days. During this period, the general condition, body weight, and estrous cycle (for 9 days after the quarantine period) were examined, and 58 each of males and females without any abnormality of the general condition (males) (or without any abnormality of the general condition or estrous cycle and with good weight increase for females) were selected and administered the test substance at the age of 10 weeks.
- The body weight at the start of the test substance administration ranged from 338 to 395 g for the males and 219 to 256 g for the females.
- Animals were housed individually in a bracket type metal wire cage in an animal breeding room maintained at a temperature of 21°C to 26°C, relative humidity of 37% to 77%, ventilation frequency of 10 to 15 air changes per hour, and illuminated for 12 hours per day (07:00 to 19:00).
- The animals were allowed free access to Solid chow NMF (nonsterilized: Oriental Yeast Co., Ltd., batch numbers 040713, 040806, 040913) from a stainless steel feeder and to tap water (city water of Gotenba, water bottle was used).
Route of administration:
oral: gavage
Vehicle:
olive oil
Details on oral exposure:
The test substance was administered by oral gavage, a method commonly employed for oral administration to rodents. Each animal received 5 mL/kg body weight of the test suspension by forced oral administration via a gastric tube (08:20-14:24 h). Animals of the control group received the vehicle (olive oil) in a similar manner. The volume of the test suspension was calculated based on the latest body weight of each animal.
Analytical verification of doses or concentrations:
yes
Remarks:
HPLC
Details on analytical verification of doses or concentrations:
- An appropriate amount of the test substance for each concentration was weighed and suspended in olive oil in an agate mortar to prescribed concentrations (50, 100, and 200 mg/mL). The test suspensions were prepared at a frequency of at least once every 7 days and stored in a cold dark place (refrigerator, observed temperature: 3°C-5°C) in light-protected containers (brown glass bottles) until use.
- Stability of the test suspensions was confirmed at Bozo Research Center and showed that 50 and 200 mg/mL suspensions of the test substance (vehicle: olive oil) remained stable for 24 hours at room temperature after storage in light-protected containers in a cold dark place (refrigerator) for 7 days.
- Confirmation of the concentrations and uniformity of the test suspensions of each concentration used for administration at week 1 and on the last week of administration were analyzed by HPLC at the Bozo Research Center. The results showed that for all the suspensions tested, the percentage of the test substance relative to the nominal value was in the range of 96.5% to 105.0%, with a C.V. in the range of 1.0% to 5.3%, which were within the acceptable range (concentration, nominal value ± 10%; C.V., ≤ 10%).
- Analytical method:
The test sample (dosing suspension), 1 mL, was diluted with 60 vol% of THF solution to 10 mL and centrifuged (2000 rpm, 1000 × g, 20°C, 5 minutes); then, 1 mL of the lower layer was diluted to 5 mL with the mobile phase of HPLC. The diluted solution was filtered through a Milex HV filter and the filtrate was subjected to measurement by the HPLC system. Single test samples were taken from the upper, middle and lower layers of the dosing suspension.
Duration of treatment / exposure:
The duration of administration was 14 days before mating, 14 days during the mating period, and 14 days after the mating period, that is, 42 days in total, for the males of the main group and the males and females of the recovery group, and 14 days before mating and up to day 4 of lactation throughout the mating and gestation periods, that is, 42 to 45 days in total, for the females of the main group. The recovery period for the males and females of the recovery group was 14 days after the end of administration, during which period the test substance was not given to the animals.
Frequency of treatment:
Daily
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
A total of 4 dose groups were set up: 250, 500, and 1000 mg/kg bw groups and the control group.
Each main group consisted of 12 male and female animals each, and each recovery group consisted of 5 each of males and female animals in the control and high- dose groups.
Control animals:
yes, concurrent vehicle
Details on study design:
- Rationale for selection of doses. In a previous study “14-day repeated-dose oral toxicity study of the test substance in rats (a preliminary study)” (doses: 125, 250, 500, and 1000 mg/kg bw, Bozo Research Center study No.: C-R016), administration of the test substance did not produce any effect even at 1000 mg/kg bw, the level defined as the limiting dose by the OECD Guideline for Testing of Chemicals 422. Therefore, 1000 mg/kg was set as the highest dose, and doses of 500 and 250 mg/kg bw were derived by dividing by a common factor of 2. A total of 3 doses were thus set up.

Details on mating procedure
- After the end of the pre-mating administration period, each pair of male and female animals in the same dose group of the main group was housed in the same cage overnight, and the females were judged to have copulated if the vaginal smear contained sperm or the presence of the vaginal plug was confirmed. Days to copulation was calculated from the day of mating, taken as day 0. From gestation day 17 to day 5 of lactation, the animals were housed individually in a plastic Econ cage with
bedding.
- Observation of mother animals. All female animals confirmed to have copulated were allowed to undergo spontaneous delivery, and observed for the presence/absence of abnormalities in the delivery. Delivery completion was checked twice daily (morning, afternoon) from gestation day 21 to the morning of gestation day 25, from which the gestation period was calculated in units of 0.5 days. If delivery was complete by 5:00 h in the afternoon, that day was regarded as day 0 of lactation.
- Mother animals that completed the delivery were observed for the presence/absence of pup licking and ingestion of the placenta and amnion. They were allowed to suckle pups up to day 4 of lactation (the date of delivery completion was regarded as day 0 of lactation) and observed for lactating behavior, using pup gathering, nest building, and breastfeeding as indices.
Positive control:
No
Observations and examinations performed and frequency:
- Observation of the general condition. All animals were observed for the presence of any abnormality of the general condition, such as in the external appearance, nutritional condition, posture, behavior, and excrements, 3 times everyday (before, immediately after, and 2 hours after the administration) during the administration period and once every morning during the recovery period.
Detailed observation of the general condition, function tests, measurement of the grip strength and spontaneous motor activity. Detailed observation of the general condition was performed once before the start of administration for all animals, once every week during the administration period for the males of the main group, once every week during the pre-mating administration and mating periods, and on predetermined days (gestation days 1, 7, 14 and 20, day 4 of lactation) during the gestation period and the lactation period for the females of the main group, and once every week during the administration and recovery periods for the animals of the recovery group. Function tests, measurement of the grip strength, and measurement of the spontaneous motor activity were performed on the last week of administration (day 39 of administration) for the males of the main group, after F1 necropsy on day 4 of lactation (day 42-45 of administration) for the females of the main group, and on the last week of administration (day 39 of administration) and last week of the recovery period (day 11 of recovery) for the males and females of the recovery group. These tests were performed on 5 animals each per group. - Measurement of body weight. Body weight was measured on days 1, 4, 8, 11, 15, 18, 22, 25, 29, 32, 36, 39 and 42 of administration, and on the day of necropsy for the males of the main group, on days 1, 4, 8, 11 and 14 of the recovery period and on the day of necropsy, in addition to the days of measurement for the males of the main group, for the males and females of the recovery group, and on days 1, 4, 8, 11 and 15 of administration (and days 18, 22 and 25 of the administration period as well as in non-copulated animals), days 0, 4, 7, 11, 14, 17 and 20 of gesta tion, and days 0 and 4 of lactation for the females of the main group. Body weight was measured from 08:06 to 10:45 h, except for the measurement on day 0 of lactation for females whose end of delivery was confirmed during the observation in the afternoon. On the day of necropsy, the body weight was measured after the animals had been denied access to food for approximately 16 hours from the previous day, in order to calculate the relative organ weight.
- Measurement of food consumption. The amount of food remaining relative to that supplied on the previous day was measured on days 1, 4, 8, 11, 15, 32, 36, 39 and 42 of administration for the males of the main group, on days 1, 4, 8, 11 and 14, in addition to the days of measurements for the males of the main group, for the males and females of the recovery group, on days 1, 4, 8, 11 and 15 of administration, days 1, 4, 7, 11, 14, 17 and 20 of gestation, and days 2 and 4 of lactation for the females of the main group. Food consumption per animal was calculated from the data thus obtained. The amount of food supplied and the amount of food remaining were measured from 08:26 to 11:25 h.
- Urinalysis (including measurement of water intake). On the last week of administration (day 36 to 37 of administration) and on the last week of the recovery period (day 8 to 9 of recovery), each of the male animals was individually housed in a cage equipped with a urine collector. Four-hour urine specimens were collected under fasting conditions of the animals with free access to water, followed by 20-hour urine specimen collection under free access to food and water. The parameters tested are
as shown below. The first 4-week urine specimens were subjected to tests from pH up to sediments, as well as measurement of the urine volume, and the subsequent 20-hour urine specimens were subjected to measurement of the osmotic pressure and urine volume. Urine volume was calculated as the sum of the volumes of 4-hour and 20-hour urine specimens. The amount of water intake from the previous day was measured using a water bottle while the animals were housed in the cage
equipped with the urine collector.
Sacrifice and pathology:
After delivery, the mother animals were exsanguinated to death by dissecting the abdominal aorta on Day 5 of lactation, after the animals had been denied access to food overnight (approx. 16-20 h) from Day 4 of lactation: 5 in each group after blood collection for hematological tests and blood chemistry tests, and the remaining animals under ether anesthesia.

- Necropsy and organ weight measurement. All of the 5 male and female animals in each group from which blood samples were collected for the hematology and blood chemistry tests on the day after the last day of administration and on the last day of the recovery period were exsanguinated to death after the blood collection, and all the other animals were exsanguinated to death by dissecting the abdominal aorta under ether anesthesia. They were then subjected to detailed gross pathological e
xamination of the body organs and tissues, including the external surface, head, chest and abdomen, and the results were recorded. In the females (mother animals), the number of corpora lutea and number of implantation sites were counted on day 5 of lactation. Then, in 5 each of the male and female animals from which blood samples were collected for the hematology and blood chemistry tests, the weight (absolute) of the following organs (testes and epididymes of all the animals) was measured
and the relative weight of each organ per 100 g body weight was calculated from the absolute organ weight and the body weight at necropsy. For bilateral organs marked with an asterisk, the weight of each side was measured separately and the sum of the weights was calculated. Brain, thyroid gland* (including parathyroid gland), thymus gland, heart, liver, spleen, kidney*, adrenal*, testis*, and epididymis*.

- Histopathological examination. The following organs and tissues of all the animals were fixed and st ored in 10 vol% formalin solution in phosphate buffer (the testes and epididymes were fixed in Bouin's fluid, followed by storage in 10 vol% formalin solution in phosphate buffer). Then, organs and tissues (see below) were embedded in paraffin, and sections were stained with hematoxylin and eosin (H-E). Specimens obtained from 5 each of the male and female animals of the control and high-dose groups
from which blood specimens were collected for the hematology and blood chemistry tests were subjected to microscopic examination (for bilateral organs, both sides were isolated and one side was subjected to the microscopic examination). The results revealed the effect of the test substance on the stomach. Therefore, specimens from 5 each of the male and female animals of the low- and medium-dose groups were also subjected to microscopic examination. Representative cases of normal
and abnormal findings were photographed.

(Cerebrum, cerebellum, pituitary gland, spinal cord (thoracic), sciatic nerve, thyroid gland, parathyroid gland, adrenal, thymus gland, spleen, submandibular lymph nodes, mesenteric lymph nodes, heart, lung (including bronchus), stomach, duodenum, jejunum, ileum, cecum, colon, rectum, liver, kidney, bladder, testis, epididymis, ovary, uterus, seminal vesicle, sternum (including bone marrow), femur (including bone marrow), and the animal identification site (auricle))
Statistics:
- Bartlett test
- Dunnett’s test
- χ2 test with Yates’ continuity correction
- Fisher’s exact test
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
- In the main group, one female in the 500 mg/kg bw group showed opacity of an eyeball (unilateral) from gestation day 5, which was not related to the dose and was therefore considered to be an incidental change.
- In the recovery group, one male in the 1000 mg/kg bw group showed decreased spontaneous motor activity from day 37 of administration up to day 7 of the recovery period, and wheezing from day 37 of administration until the end of the recovery period.
- No abnormality was observed in the other animals, either in the main or in the recovery groups.
Mortality:
no mortality observed
Description (incidence):
- There were no significant difference in the body weight between males and females of the main group. A significantly greater increase in body weight was observed in the females of the 250 and 1000 mg/kg bw groups during the lactation period, but the increase was not dose-related.
- In the recovery group, males in the 1000 mg/kg bw group showed decreased body weight gain during the administration period and decreased body weight (-21g) during the recovery period. This was caused by the abnormality in 1 out of the 5 animals. This animal showed continued body weight decrease (and also decreased spontaneous activity and wheezing in the observation of the general condition; the body weight was 466g before manifestation of the symptom and 261g on day 14 of the recovery period). The body weights of the other 4 males and 5 females in the same group were similar to those of the animals in the control group, showing no statistically significant differences.
Body weight and weight changes:
effects observed, non-treatment-related
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
Administration of the test substance did not have any effect on the food consumption in the males or females of either the main group or the recovery group. A significant increase was observed on day s 2 and 4 of lactation in the females of the 250 mg/kg dose group in the main group, but this was not dose-related.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
-Tests at the end of the administration period. A significant increase in the serum level of fibrinogen was observed in the females of the 250 mg/kg bw group and a significant decrease in the percentage of lymphocytes and significant increase in the percentage of segmented neutrophils were observed in the females of the 500 mg/kg bw group. However, none of these changes were observed in the 1000 mg/kg bw group, suggesting that they were within the range of physiological variations. No significant differences were observed in the male animals between the control group and any of the treatment groups.
- Tests at the end of recovery period. A significant increase in the mean corpuscular volume of the red blood cells, significant decrease of the platelet count, significant increase in the percentage of lymphocytes, and significant decrease in the percentage of segmented neutrophils were observed in females of the 1000 mg/kg bw group. However, these changes were not observed at the end of the administration period, which suggested that they were within the range of physiological variations. No significant differences were observed in the male animals between the control group and any of the treatment groups.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
- Test at the end of the administration period. A significant increase in the serum level of ALT was observed in the males of the 1000 mg/kg bw group. A significant decrease of the serum level of inorganic phosphorus was observed in the males of the 250 mg/kg bw group. However, since the decrease was not dose-related, it was considered to be within the range of physiological variations.
- Tests at the end of recovery period. A significant increase in the serum level of total protein was observed in the females of the 1000 mg/kg bw group. However, since no such change was observed at the end of the administration period, the increase was considered to be within the range of physiological variations.
Urinalysis findings:
no effects observed
Description (incidence and severity):
Urinalysis (including measurement of water intake) showed no abnormalities in the qualitative parameter values in any of animals in either the main group or in the recovery group. No significant difference was observed in any of the quantitative parameter values between the control group and any of the treatment groups.
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
- Observation of animals in the home cage. No abnormalities were observed in any of the animals in either the main group or the recovery group.
- Observation of the animals while being handled. No abnormality was observed in any of animals in either the main group or the recovery group.
- Observation of animals in the open field. Males of the 1000 mg/kg bw group in the main group showed a significant increase of the defecation frequency during weeks 1 and 2 of administration, which was a very mild transient change and considered to be within normal range. No abnormalities were observed in the other parameters in any of the animals in either the main group or the recovery group. No significant differences were observed in the standing frequency between the control group and any of the treatment groups.
- Function tests. No abnormalities were observed in any of the animals in either the main group or the recovery group. No significant differences were observed in the air righting reflex or landing foot splay between the control group and any of the treatment groups.
- Measurement of the grip strength. No significant differences were observed between the control group and any of the treatment groups in either the main group or the recovery group.
- Measurement of spontaneous motor activity (measured for 10-minute periods and a total of 60 minutes). No significant difference was observed between the control group and any of the treatment groups in either the main group or the recovery group.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
No dose-related changes in either direction (increase or decrease) were observed in either the absolute or the relative weight. Although significant differences in the weights of the following organs were observed, they were considered to be within the range of normal variations, because they were neither dose-related nor were observed at the end of the administration period.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
- Findings at the end of the administration period. Stomach: Indentation of the anterior stomach was observed in 0, 5 and 7 males, and 1, 1 and 3 females of the 250, 500 and 1000 mg/kg bw groups, respectively. White foci were observed in 1 male of the 500 mg/kg bw group. Rough mucosa in the anterior stomach was observed in 5 and 9 males, and 5 and 6 females of the 500 and 1000 mg/kg bw groups, respectively. Indentation of the glandular stomach was observed in 1 female of the 500 mg/kg bw group. All the other findings observed in the organs and tissues were considered to be incidental, as judged from the frequency of their occurrence and the pathological findings (Dark red foci in the glandular stomach were observed in 0, 1, 2 and 1 males and 4, 2, 1 and 1 females of the control
group, 250, 500 and 1000 mg/kg bw groups, respectively. Kidney: Pyelectasis was observed in 2 and 1 males of the 250 and 1000 mg/kg bw groups, respectively. Eyeballs: Corneal opacity (unilateral) was observed in 1 female of the 500 mg/kg bw group).
- Findings at the end of the recovery period. Stomach: Rough mucosa in the anterior stomach was observed in 1 male of the 1000 mg/kg bw group. This animal also showed expansion of the digestive tract from the stomach to the colon due to gas accumulation, and a mild decrease in the size of the testis. Other findings observed in the following organs were considered to be incidental as judged from the frequency of their occurrence and the pathological findings (Lung: Dark red foci were observed in 1 female of the 1000 mg/kg bw group).
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Administration of the test substance had effects on the stomach of the animals in the 250 mg/kg bw and higher dose groups.
- Findings at the end of the administration period.
Stomach: Mild to moderate erosions or ulcers of the anterior stomach were observed in 0, 4 and 4 males and 1, 1 and 1 females of the 250, 500, and 1000 mg/kg bw groups, respectively. Very mild to moderate thickening of the anterior stomach mucosa was observed in 1, 4 and 5 males and 1, 4 and 3 females of the 250, 500, and 1000 mg/kg bw groups, respectively. Very mild to mild edema of the submucosal tissue in the anterior stomach was observed in 1, 5 and 5 males and 0, 4 and 3 females of the 250, 500, and 1000 mg/kg bw groups, respectively. Most of these changes in the anterior stomach were localized findings. All of the other findings observed, as follows, were considered to be incidental changes as judged from the frequency of their occurrence and the histopathological findings
Epididymis: Very mild infiltration by stromal cells was observed in 1 male of the control group. Heart: Very mild localized myocarditis was observed in 4 males of the control group and 1 male of the 1000 mg/kg bw group.
Kidney: Very mild basophilic tubules were observed in 3 males of the control group and 1 male and 1 female in the 1000 mg/kg bw group.
Liver: Very mild, minute granulomas were observed in 3 males of the control group and 1 male of the 1000 mg/kg bw group.
Lung (including bronchi): Very mild mineral deposits in the arterial walls were observed in 1 male of the control group and 1 female of the 1000 mg/kg bw group. Very mild accumulation of foam cells was observed in 2 males and 1 female of the control group, and 1 male and 3 females of the 1000 mg/kg bw group.
Spleen: Very mild to mild extramedullary hematopoiesis was observed in 5 females each in the control group and 1000 mg/kg bwgroup.
Stomach: Inclusion cysts were observed in 1 male of the 500 mg/kg bw group. Very mild to mild erosions in the glandular stomach were observed in 0, 0, 0 and 1 male and 3, 1, 1, and 0 females in the control group, 250, 500, and 1000 mg/kg bw group, respectively).

- Findings at the end of the recovery period. Stomach: Moderate thickening of the anterior stomach mucosa was observed in 1 male of the 1000 mg/kg bw group.
Histopathological findings: neoplastic:
no effects observed
Other effects:
no effects observed
Description (incidence and severity):
Refer to the other related RSS for details on reproductive and development toxicity.
Dose descriptor:
NOAEL
Effect level:
ca. 1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No systemic effects observed up to 1000 mg/kg bw/d; the changes in forestomach are of local nature due to the irritant properties of the substance
Key result
Dose descriptor:
LOAEL
Effect level:
ca. 250 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: effects on forestomach
Key result
Critical effects observed:
no
Conclusions:
Based on the results of the read across study, NOAEL for systemic effects for the test substance, is considered to be 1000 mg/kg bw/day.
Executive summary:

A screening study was conducted to determine the toxicity to reproduction of the read across substance, 'mono- C12 PSE, Na+' (purity: 100%, according to the OECD Guideline 422, in compliance with GLP. The read across substance was administered at 0 (control group), 250, 500 or 1000 mg/kg bw to male Sprague-Dawley SPF rats for 14 days before mating, through the mating period, and up to 1 d before necropsy (42 d in total) and to female Sprague-Dawley SPF rats for 14-d before mating, through the mating period and the gestation period, up to Day 4 of lactation (42 to 45 d in total) to investigate the repeated-dose, reproductive and developmental toxicities. In the 0 and 1000 mg/kg bw groups, a 14-d recovery period was allowed after the 42-d administration period to investigate the reversibility of the toxic changes. Administration of the read across substance had no effect on any of the following: general condition, findings on detailed observation of the general condition, results of function tests, grip strength, spontaneous motor activity, body weight, food consumption, results of urinalysis (including water intake), or in the results of haematological or blood chemistry tests. Administration of the read across substance had no effect on any of the following: general condition, findings on detailed observation of the general condition, results of function tests, grip strength, spontaneous motor activity, body weight, food consumption, results of urinalysis (including water intake), or in the results of haematological or blood chemistry tests. Changes in the forestomach may be considered to be adverse, however, they are also considered to be a result of local irritation of the (irritant) test substance (which is brought directly in contact to the mucosa in a massive amount by gavage application) than a true effect of systemic toxicity. Furthermore, in the light of the absence of a forestomach in humans, observed effects on this tissue are of questionable relevance with reference to the extrapolation of the toxic properties of a read across substance in humans. Under the study conditions, only (local) changes of the forestomach mucosa were observed at 250 mg/kg bw/day and above, which are due to the irritant properties of the substance. Therefore, the NOAEL (systemic) is considered to be 1000 mg/kg bw/day (BRC, 2005). Based on the results of the read across study, a similar systemic NOAEL can be expected for the test substance, 'mono- and di- C16-18 PSE and C16-18 AE10 PSE'.

Reason / purpose for cross-reference:
data waiving: supporting information
Reference
Endpoint:
short-term repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
From May 30, 2008 to February 06, 2009
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
KL2 due to RA
Justification for type of information:
Refer to section 13 of IUCLID for details on the read-across justification. The study with the read across substance is considered sufficient to fulfil the information requirements as further explained in the provided endpoint summary.
Reason / purpose for cross-reference:
read-across source
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Crl:CD(SD)
Sex:
male/female
Details on test animals or test system and environmental conditions:
Test animals
- Source: Charles River Laboratories, Inc., Portage, MI, USA
- Age at study initiation: 72 d (males), 66 d (females)
- Weight at study initiation: 306-344 g (males), 208-239 g (females)
- Housing: individual housing in stainless steel, wire-bottomed cages.
- Diet: Certified Rodent Diet 5002 (PMI Nutrition International, St.Louis, MO, USA), ad libitum
- Water: filtered local water (chlorine addition), ad libitum
- Acclimation period: 7 d

Environmental conditions
- Temperature (°C): 19-25°C
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
Preparation of dosing solutions:
Formulations were prepared at least once daily. Formulations were used within four hours of preparation and were stirred for at least 30 or 60 minutes before dosage administration.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
A weight/weight verification method was employed to verify the test-substance content of the prepared formulations.
Duration of treatment / exposure:
Males: 14 d before cohabitation period through the day before sacrifice (45 dosages)
Females: 14 d before cohabitation period through the day before scheduled sacrifice (Day of lactation (DL) 4 for dams that delivered a litter, Day of gestation (DG) 24 for rats that did not deliver a litter, study Day (DS) 52 for the rat with no confirmed day of mating). Surviving female rats were given a total of 38 to 52 dosages.
Frequency of treatment:
Once daily
Remarks:
Doses / Concentrations:
25, 50, 200, 800 mg/kg bw
Basis: actual ingested
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: In a range-finding study rats were dosed with 100, 300, and 1000 mg/kg bw/day. At Day 6 the high dose of 1000 mg/kg bw was lowered to 750 mg/kg bw/day, due to excessive toxicity. Reductions of the body weight were observed within Days 4-5. In the gestation period body weights were only slightly reduced in the 300 and 750/1000 mg/kg bw/day dose groups. No effects were observed after Caesarean section on Day 21 of gestation up to the highest dose of 750/1000 mg/kg bw/day. The female animal of the 1000 mg/kg bw/day that was sacrificed on study Day 4 had received 3 doses. The animal lost a total of 28 g of body weight and consumed an average of 8 g of food per day. Clinical signs including excess salivation, rales, cold/warm to touch, hunched posture, and scant feces were noted. Necropsy revealed two black ulcerations present in the pyloric region of the stomach, dark red lobes of the liver, red lobes of the lungs, and intestines were distended with gas. Thus, doses of 25, 50, 200, and 800 mg/kg bw/day were chosen for the main study. The low dose was expected to be a NOAEL for maternal and embryo-fetal toxicity, and the 800 mg/kg bw/day dose was expected to produce minimal maternal toxicity and little or no developmental toxicity.
Observations and examinations performed and frequency:
Cage side observations: Yes
- Time schedule: twice daily
- Cage side observations: clinical signs, abortions, premature deliveries, deaths

Detailed clinical observations: Yes
- Time schedule: before the first dosage and weekly thereafter

Body weight: Yes
- Time schedule for examinations: daily

Food consumption:
- Males: weekly, except for the cohabitation period; females: weekly until the cohabitation period, and on gestation Day (DG) 0, 7, 10, 12, 15, 18, 20, DG 24, DG 25, and on Days of lactation (DL) 1 and 4. For the rat with no confirmed mating on study days (DS) 28, 35, 38, 40, 43, 46, 48, and 52.

Haematology: Yes
- Time schedule for collection of blood:
- Anaesthetic used for blood collection: No, rats were sacrificed by carbon dioxide asphyxiation.
- Animals fasted: Yes
- How many animals: 4-5
- Parameters checked: erythrocyte count, haematocrit, mean cell volume, mean corpular volume, haemoglobin, leucocyte count, thrombocyte count, mean platelet volume, activated partial thromboplastin time, prothrombine time.

Clinical chemistry: Yes
- Time schedule for collection of blood: No, rats were sacrificed by carbon dioxide asphyxiation.
- Animals fasted: Yes
- How many animals: 4-5
- Parameters checked: gamma glutamyl transferase, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, sodium, potassium, glucose, urea, albumine, globuline, albumin/globulin ratio, protein, calcium, creatinine, cholesterol, chloride, bilirubin, triglycerides.


Neurobehavioural examination: Yes
- Time schedule for examinations: DS 46 (males), DL 5 and DG 25 (females)
- Dose groups that were examined: 5/sex/group (all dose groups)
- Battery of functions tested: sensory activity / grip strength / motor activity / autonomic functions / abnormal clinical signs
Sacrifice and pathology:
Gross pathology: Yes, whole body

Histopathology: Yes, small intestine, large intestine, brain, urinary bladder, heart, testes, liver, lung, trachea, lymphnodes, mesentery lymphnodes, spleen, epididymides, adrenal gland, peripheral nerve, kidney, ovary, prostate, vesicula seminalis, thyroid gland, oesophagus, thymus, uterus, bone marrow, mammary gland, spinal cord, sternum, stomach, vagina
Statistics:
Proportion data as well as descriptive and quantal data from the functional observational battery (FOB), was analyzed as contingency tables using the variance test for homogeneity of the binominal distribution.
Bartlett´s test of homogeneity of variances were used to estimate the probability that the dosage groups have different variances. Any non-significant result (p>0.001) indicate that an assumption of homogeneity of variance is anappropriate, and the data were compared using the Analysis of Variance. If p≤0.05, the groups given the test substance were compared with the control group using Dunnett´s Test. If Bartlett´s test was significant (p≤0.001), the Analysis of Variance Test was not appropriate (nonparametric). When 75% or fewer of the scores in all the groups were tied, the Kruskal-Wallis Test was used to analyze the data, and in the event of a significant result (p≤0.05), Dunn´s Test was used to compare the groups given the test substance with the control group. When more than 75% of the scores in any dosage group are tied, Fisher´s Exact Test was used to compare the proportion of ties in the dosage group.
Data from the motor activity test, with repeated measurements within a session, was analyzed using an Analysis of Variance with repeated measures. A significant effect (P≤0.05) in that test could have appeared as effect of Concentration (a difference between groups in the total across all measurements in a session) or as an interaction between Concentration and Block (a difference between groups at specific measurement periods). If the Concentration x Block interaction was significant, an Analysis of Variance Test was used to evaluate the data at each measuremnet period, and a significant result (p≤0.05) was followed by a comparison of the groups using Dunnett´s test. Variables with graded count scores, such as litter size, were analyzed using Kruskal-Wallis (if p≤0.05 followed by Dunn´s Test).
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
800 mg/kg bw/day: excess salivation (male and female), chromorhinorrhea, urine-stained abdominal fur, red and/or dried perioral substance and red or clear perinasal substance in males
Mortality:
mortality observed, treatment-related
Description (incidence):
800 mg/kg bw/day: excess salivation (male and female), chromorhinorrhea, urine-stained abdominal fur, red and/or dried perioral substance and red or clear perinasal substance in males
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
800 mg/kg bw/day: reduction of body weight gain
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
800 mg/kg bw/day: reduced food consumption
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
no effects observed
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
800 mg/kg bw/day: damage on the lining of the stomach
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
800 mg/kg bw/day: findings in the non-glandular stomach related to test substance (8/8 males; 4/4 females)
Histopathological findings: neoplastic:
no effects observed
Details on results:
Clinical signs and mortality
There were no substance-related deaths. One female rat of the 800 mg/kg bw/day dose group was sacrificed due to adverse clinical observations and two more female rats were found dead due to an intubation error.
In the 800 mg/kg bw/day group excess salivation, chromorhinorrhea, urine-stained abdominal fur, red and/or dried perioral substance and red or clear perinasal substance occurred in significantly increased numbers of male rats. All other clinical signs were considered unrelated to treatment.
Significantly increased numbers of females in the 800 mg/kg bw/day group had excess salivation.

Body weight and weight gain
800 mg/kg bw/day:
Males: reduction of body weight gain (DSs 1 to 14) and body weight (Day 8). Body weight mean on Day 45 was 94% of the control value.
Females: Precohabitation: no effects; gestation: reduced body weight gain in the 800 mg/kg bw/day group; lactation: no effects

Food consumption:
800 mg/kg bw/day:
Males: reduction from Day 1-14 (absolute and relative) and Day 1-43 (relative)
Females:
Precohabitation: reduction from study Day (DS) 1-8 (absolute and relative)
Gestation: reduction from Days of gestation (DG) 0-7 and 15-18
Lactation: no effects
No effects were observed in the other dose groups.

Haematology
No biologically important differences were observed.
Reductions in APTT for females of the 50 and 800 mg/kg bw/day group reflects an increased value for the vehicle control. ALT and triglycerides were increased at 50/200 and 25/800 mg/kg bw/day groups, respectively. These effects were considered unrelated to treatment as no dose dependency was observed.

Clinical chemistry
No biologically important differences were observed.

Neurobahaviour
No effects were observed.

Organ weights
No treatment-related effects were observed.

Gross pathology
800 mg/kg bw/day:
Males: thickened walls of the cardiac region of the stomach and white areas on the mucosal surface (3/10), ulceration on the mucosal surface of the cardiac region of the stomach (1/10)
Females: lesions of the cardiac region of the stomach (2/10)

Histopathology: Non-neoplastic
800 mg/kg bw/day: findings in the non-glandular stomach related to test substance (8/8 males; 4/4 females)
200 mg/kg bw/day: findings in the non-glandular stomach possibly related to test substance, irritating effects (1/5 males)
25, 50 mg/kg bw/day: no adverse effects observed

Other findings
No clear and consistent treatment related differences in results for the qualitative and quantitative mcroscopic assessment of bone marrow smears were observed.
Key result
Dose descriptor:
NOAEL
Remarks:
systemic
Effect level:
800 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No treatment related systemic effects
Key result
Dose descriptor:
NOAEL
Remarks:
local
Effect level:
200 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No treatment related local effects on forestomach
Key result
Dose descriptor:
LOAEL
Remarks:
local
Effect level:
800 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Irritating effects on the forestomach of the rat. Occurrence of hyperplasia and ulceration.
Key result
Critical effects observed:
no
Conclusions:
Based on the results of the read across study, the test substance, NOAEL for local effects and systemic effects are considered to be 200 mg/kg bw/day and 800 mg/kg bw/day respectively.
Executive summary:

A screening study was conducted to determine the repeated dose and reproduction / developmental toxicity potential of the read across substance, 'C8-10 AE4 PSE' (purity: 100%), according to OECD Guideline 422, in compliance with GLP. In the main test, the test substance was administered at 0 (control group), 25, 50, 200, 800 mg/kg bw/day doses to male Crl:CD(SD) rats (10 animals per dose) 14 d before cohabitation period, through the day before sacrifice (45 dosages) and female rats (per dose 10 animals) 14 d before cohabitation period, through the day before scheduled sacrifice (i.e., Day of lactation (DL) 4 for dams that delivered a litter, Day of gestation (DG) 24 for rats that did not deliver a litter, study Day (DS) 52 for the rat with no confirmed day of mating). Surviving female rats were given a total of 38 to 52 dosages. There were no substance-related deaths observed. In the 800 mg/kg bw/day group excess salivation, chromorhinorrhea, urine-stained abdominal fur, red and/or dried perioral substance and red or clear perinasal substance occurred in significantly increased numbers of male rats. Significantly increased numbers of females in the 800 mg/kg bw/day group had excess salivation. All other clinical signs were considered unrelated to treatment. No treatment related significant change in body weight, weight gain, and organ weight and food consumption were observed. No biologically important differences were observed. Reductions in APTT for females of the 50 and 800 mg/kg bw/day group reflected an increased value for the vehicle control. ALT and triglycerides were increased at 50/200 and 25/800 mg/kg bw/day groups, respectively. These effects were considered unrelated to treatment as no dose dependency was observed. In gross pathology, at 800 mg/kg bw/day dose, thickened walls of the cardiac region of the stomach and white areas on the mucosal surface (3/10), ulceration on the mucosal surface of the cardiac region of the stomach (1/10) were observed in males. In females, lesions of the cardiac region of the stomach (2/10) were observed. In histopathology, at 800 mg/kg bw/day dose, findings in the non-glandular stomach related to test substance (8/8 males; 4/4 females) were noted. Also, at 200 mg/kg bw/day dose, findings in the non-glandular stomach possibly related to test substance, irritating effects (1/5 males) were observed, whereas at 25, 50 mg/kg bw/day dose groups, no adverse effects were noted. No clear and consistent treatment related differences in results for the qualitative and quantitative microscopic assessment of bone marrow smears were observed. Changes in the forestomach may be considered to be adverse, however, they are also considered to be a result of local irritation of the (irritant) test substance (which is brought directly in contact to the mucosa in a massive amount by gavage application) than a true effect of systemic toxicity. Furthermore, in the light of the absence of a forestomach in humans, observed effects on this tissue are of questionable relevance with reference to the extrapolation of the toxic properties of a test substance in humans. Under the study conditions, the NOAEL for local effects and systemic effects were determined to be 200 mg/kg bw/day and 800 mg/kg bw/day respectively (Lech, 2009). Based on the results of the read across study, similar NOAELs for systemic and local effects can be considered for the test substance, 'mono- and di- C16-18 PSE and C16-18 AE10 PSE'.

Reason / purpose for cross-reference:
data waiving: supporting information
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
1982
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Justification for type of information:
Refer to section 13 of IUCLID for details on the read-across justification. The study with the read across substance is considered sufficient to fulfil the information requirements as further explained in the provided endpoint summary.
Reason / purpose for cross-reference:
read-across source
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Deviations:
yes
Remarks:
Urine analysis was not performed
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
As described in guideline
Route of administration:
oral: feed
Vehicle:
other: plain diet
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
90 d
Frequency of treatment:
Daily
Remarks:
Doses / Concentrations: 0, 300, 1000, 3000, 10,000 ppm
Basis: nominal in diet
Remarks:
Doses / Concentrations: 0, 15, 50, 150, 500 mg/kg bw/day
Basis: actual ingested
No. of animals per sex per dose:
12
Control animals:
yes, concurrent vehicle
Details on study design:
As described in guideline
Positive control:
Not required
Observations and examinations performed and frequency:
As described in guideline
Sacrifice and pathology:
As described in guideline
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Details on results:
Body weight
Mean body weights of male rats fed 10,000 ppm and of females fed 10,000 and 3000 ppm test substance were significantly lower than control throughout the 13 week feeding study.

Food consumption
Significantly reduced food intake and increased spillage of the 10,000 ppm diet were observed in both sexes; females fed 3000 ppm diet also had lower food intakes than control but the decrease was significant for only five of the 13 weeks.

Haematology
Haematological alterations observed in 3000 ppm males and in both sexes fed 10,000 ppm test substance were significant increases in total leukocytes and in absolute lymphocyte values. Females fed 10,000 ppm also showed significant depression in numbers of neutrophils (absolute and percent), mean cell volume and mean cell haemoglobin. Males in the upper two treatment groups showed shortened prothrombin times.

Clinical chemistry
Elevated clinical chemical values were, with the exception of plasma urea concentrations, confined to rats in the 10,000 ppm treatment group. Males in this group had significantly higher urea, calcium and potassium values, while females had significantly higher urea, chloride, calcium and cholesterol values. Urea concentrations of females in the 1000 and 3000 ppm groups were also higher than control.

Organ weights
After correction for reduced terminal body weight, there were increases in mean liver weight in the 3000 and 10,000 ppm groups (both sexes) and in 1000 ppm females. Males fed 10,000 ppm test substance had significantly increased spleen weights and females in the 1000 ppm group had significantly heavier kidneys compared to controls.
Key result
Dose descriptor:
NOAEL
Effect level:
>= 500 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
body weight and weight gain
clinical biochemistry
food consumption and compound intake
haematology
Remarks on result:
other: changes reported are considered minor and not of toxicological significance
Key result
Critical effects observed:
no
Conclusions:
Based on the results of the read across study, the test substance, NOAEL for systemic toxicity is considered to be >500 mg/kg bw/day.
Executive summary:

A 13 week study was conducted to determine the sub chronic oral toxicity potential of the read across substance, 'C14-15 AE7' (purity not specified) in rats, according to the method similar to OECD Guideline 408. In this study, the read across substance was administered daily at dietary concentrations of 0, 300, 1000, 3000, 10,000 ppm (corresponding to approximately 0, 15, 50, 150, 500 mg/kg bw/day to male and female Wistar rats (12 animals per sex per dose)) up to 13 weeks. The animals were observed for mortality, clinical signs, body weight, food consumption, haematology and clinical chemistry at termination. Gross and histopathological examinations were also performed.Mean body weights of male rats fed 10,000 ppm and of females fed 10,000 and 3000 ppm test substance were significantly lower than control throughout the 13 week feeding study. Significantly reduced food intake and increased spillage of the 10,000 ppm diet were observed in both sexes; females fed 3000 ppm diet also had lower food intakes than control but the decrease was significant for only five of the 13 weeks. Haematological alterations observed in 3000 ppm males and in both sexes fed 10,000 ppm test substance were significant increases in total leukocytes and in absolute lymphocyte values. Females fed 10,000 ppm also showed significant depression in numbers of neutrophils (absolute and percent), mean cell volume and mean cell haemoglobin. Males in the upper two treatment groups showed shortened prothrombin times. Elevated clinical chemical values were, with the exception of plasma urea concentrations, confined to rats in the 10,000 ppm treatment group. Males in this group had significantly higher urea, calcium and potassium values, while females had significantly higher urea, chloride, calcium and cholesterol values. Urea concentrations of females in the 1000 and 3000 ppm groups were also higher than control. After correction for reduced terminal body weight, there were increases in mean liver weight in the 3000 and 10,000 ppm groups (both sexes) and in 1000 ppm females. Males fed 10,000 ppm test substance had significantly increased spleen weights and females in the 1000 ppm group had significantly heavier kidneys compared to controls. No effects on the general health and behaviour of treated rats were evident. Significant treatment-related effects on body weight, food intake, organ weights, clinical chemistry and haematology were identified in one or both sexes at daily doses of 150 and 500 mg/kg bw/day. Due to the fact that no compound-related gross or histopathological lesions were identified at any dose level, the changes reported are considered minor and not of toxicological significance. Hence the study author concluded that, under the study conditions the NOAEL for systemic toxicity was set to greater than 500 mg/kg bw/day (XXXX, 1982). Based on the results of the read across study, similar systemic NOAEL can be considered for the test substance 'mono- and di- C16-18 PSE and C16-18 AE10 PSE'.

Data source

Materials and methods

Results and discussion

Applicant's summary and conclusion