2-hydroxy-1-(4-(4-(2-hydroxy-2-methylpropionyl)benzyl)phenyl)-2-methylpropan-1-one

Endpoint summary

Description of key information

The study was conducted to a recognised testing guideline with GLP certification.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 July 2011 - 15 May 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Version / remarks:

22 March 1996

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: EPA OPPTS 870.3650 Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test

Version / remarks:

July 2000

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Source: Harlan Laboratories, B.V. Kreuzelweg 53 5961 NM Horst / Netherlands
- Age at study initiation: 11 weeks
- Weight at study initiation: Males: 291 to 371 g, Females: 182 to 213 g
- Housing: singly
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 7d
- Fasting: 16h before administration

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 30-70
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
-The dose formulations were prepared weekly
- Homogeneity of the test item in the vehicle was maintained during the daily administration period using a magnetic stirrer.

VEHICLE
- Justification for use and choice of vehicle (if other than water): As used in previous dose range-finding study

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The samples were analyzed following an analytical procedure provided by the Sponsor and adapted at Harlan Laboratories. The test item was used as the analytical standard. Analyzed samples were not discarded without written consent from the study director. Duplicates were taken of all samples and were stored at Harlan Laboratories Ltd., Füllinsdorf / Switzerland. The samples were not discarded without written consent from the study director.

Duration of treatment / exposure:

Males: 4 weeks
Females: Approximately 7 weeks

Frequency of treatment:

daily

Dose / conc.:

10 mg/kg bw/day (actual dose received)

Dose / conc.:

30 mg/kg bw/day (actual dose received)

Dose / conc.:

90 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The dose levels were selected based on a previous dose range-finding toxicity study in Han Wistar rats

Positive control:

no

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: daily

HAEMATOLOGY: Yes
- Time schedule for collection of blood: on the day of the scheduled necropsy from 5 males from each group, blood samples from 5 lactating females from each group were obtained on day 5 post partum
- Anaesthetic used for blood collection: Yes, isoflurane
- Animals fasted: Yes, 18h
- How many animals: 5 per sex and group
- Parameters checked in table [1] were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: on the day of the scheduled necropsy from 5 males from each group, blood samples from 5 lactating females from each group were obtained on day 5 post partum
- Animals fasted: Yes, 18h
- How many animals: 5 per sex and group
- Parameters checked in table [2] were examined.

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: one time during the study
- Dose groups that were examined: 5 animals per sex and group
- Battery of functions tested: cage side observations, hand-held observations, open field observations, reflexes, hind / fore limb strength

Sacrifice and pathology:

GROSS PATHOLOGY: Yes (see table)
HISTOPATHOLOGY: Yes (see table)

Statistics:

The Dunnett-test (many to one t-test) based on a pooled variance
estimate was applied if the variables could be assumed to follow a normal distribution
for the comparison of the treated groups and the control groups for each sex.
• The Steel-test (many-one rank test) was applied instead of the
Dunnett-test when the data could not be assumed to follow a normal distribution.
• Fisher's exact-test was applied if the variables could be
dichotomized without loss of information.

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

DAILY
Test item-related clinical signs were observed of males and females receiving 90 mg/kg body weight/day of Irgacure 127. Hunched posture, ruffled fur and discoloured feces were seen.

At the dose level of 90 mg/kg body weight/day, the male animal killed in extremies (no. 35) showed clinical signs such as ruffled fur from day 6 of treatment and hunched posture, ruffled fur and yellowish discoloured feces from day 8 of treatment. At necropsy no macroscopical finding was noted for this animal. Female no. 78, treated at the dose level of 90 mg/kg body weight/day and found dead on day 2 of the pairing period, had ruffled fur from day 6 of treatment and yellowish discoloured feces from day 9 of treatment. At necropsy examination, dark red iscoloration of the thymus, enlarged stomach and redish brown firm nodule in the jejunum were found. After histopathological examination it was concluded that the cause of death was likely the thrombosis of jejunum therefore this death was considered to be incidental.

At the dose level of 90 mg/kg body weight/day additional males were noted with ruffled fur and yellowish discoloured feces (no. 34 and 40) in the pre-pairing and pairing period. Male no. 37 had scabs and reddish sore on the right shoulder appearing in the pairing period and the scabs were noted in the after pairing period too.
Three additional females in this group (nos. 72, 73, 74) had ruffled fur starting in the pre-treatment period through pairing period. Female no. 73 had yellowish discoloured feces from the second week of pre-pairing period up to and including the gestation period. Female no. 72 also had huched posture on the first two days of the gestation period.
In groups 3 and 2 no clinical signs were noted in any sex during the study.

In group 1, male no. 10 had reddish sore at the cervical region for a short period in the pairing
period.


WEEKLY
At the dose level of 90 mg/kg body weight/day, the ruffled fur noted during the detailed weekly
clinical observations of males and females indicated a test item-related effect.

No test item-related effect was found in the other dose groups.

Mortality:

mortality observed, treatment-related

Description (incidence):

At 90 mg/kg body weight/day, one male was killed for ethical reason on day 9 of treatment in the pre-pairing period and one female died spontaneously on day 2 of pairing period (on day 16 of treatment). After histopathological examination it was concluded that the cause of death of the female was likely the thrombosis in jejunum and, therefore, this death was considered to be incidental and not test item related. Histopathological examination of the male showed slight diffuse hyperkeratosis in forestomach and minimal erosion in glandular stomach, signs of moderate stress in thymus and adrenal glands as well as slight diffuse atrophy in the prostate, coagulating gland and seminal vesicles. The body weight and food consumption of this male was rapidly decreasing from the second day of treatment onwards probably due to individual sensitivity. Since other males at 90 mg/kg body weight/day, started to show body weight and food consumption reduction later in the treatment period (the body weight started to decrease from the pairing period onwards and the food consumption was decreased in the second week of pre-pairing period), relationship to the treatment could not be excluded.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

BODY WEIGHT MALES
Pre-pairing and Pairing Periods
From day 5 of the pre-pairing period the mean body weight of males receiving 90 mg/kg body weight/day of Irgacure 127 was minimally decreased throughout the study when compared to the control group, although not reaching a level of statistical significance.

The mean body weight gain of males receiving 90 mg/kg body weight/day of Irgacure 127 was lower from day 5 of pre-pairing period when compared to controls and it was statistically significantly reduced between days 8 and 14. In correlation with the low food consumption this was considered to be a test item-related effect. Thereafter mean values similar to the control group were achieved during the pairing and after pairing periods.

In groups 30 and 10 mg/kg body weight/day, mean body weights and mean body weight gains of the males were similar to that of the control group during the study.

BODY WEIGHT FEMALES
Pre-pairing, Pairing, Gestation and Lactation Periods
During the pre-pairing period the mean body weights of females at the dose level of 90 mg/kg body weight/day were minimally below of the control level achiving levels of occasional statistical significance from day 9 onwards. During the pairing, gestation and lactation periods mean body weights comparable to those of the control group were noted.

Significantly lower mean body weight gain was recorded from day 4 of the pre-pairing period was considered to be a test item-related effects. During the gestation period, minimally increased mean body weight gain values were found while there was no difference in mean body weight gain noted during the lactation period.

In groups 30 and 10 mg/kg body weight/day, mean body weights and mean body weight gains of the females were similar to that of the control group during the study.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

FOOD CONSUMPTION MALES
The mean food consumption of males receiving 90 mg/kg body weight/day of Irgacure 127 was statistically significantly lower between days 8 and 14 in the pre-pairing period and it was lower during the whole pre-pairing period. This was considered to be a test item-related effect.

The mean food consumption of males was at control level for each test item-treated groups after the pairing period.

In groups 30 and 10 mg/kg body weight/day, food consumption of males was not affected by the treatment of the test item.

FOOD CONSUMPTION FEMALES
The mean food consumption of females receiving 90 mg/kg body weight/day of Irgacure 127 was statistically significantly lower during the pre-pairing period which was considered to be a test item-related effect.

There was no effect of treatment with Irgacure 127 on the food consumption of females in the gestation or lactation periods.

In groups 30 and 10 mg/kg body weight/day, the mean food consumption of females was not affected by the treatment of the test item.

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Description (incidence and severity):

MALES
No changes to be of toxicological relevance were noted in males in any dose group.

FEMALES
No changes were noted in females in any dose group.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

The assessment of the biochemistry data did not reveal any test item-related effect in any dose
group in any sex.

MALES
No changes were noted in males in any dose group.

FEMALES
The total cholesterol level was statistically significantly higher in females treated at the dose level of 90 mg/kg body weight/day and lying slightly outside the range of historical control data, thus, test item-realted relationship cannot be excluded. The creatinine level was statistically significantly higher in females treated at the dose level of 10 mg/kg body weight/day however the value was within the historical background range. In absence of dose relationship no experimental relevance was attributed to this finding.

At the dose level of 30 mg/kg body weight/day no changes were noted.

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

effects observed, treatment-related

Description (incidence and severity):

FUNCTIONAL OBSERVATIONAL BATTERY
None of the parameters under investigation during the functional observational battery gave an indication of a test item-related effect.
Mean values of grip strength (fore- and hind paws) and landing foot splay gave no indication of
test item-related effects.
BODY TEMPERATURE
Mean body temperature in males was statistically significantly lower at the dose level of 90 mg/kg body weight/day compared to the control group (37.3 °C compared to 38.1 in the control group) and it was marginally lower when compared to historical control data range (37.5 - 39.1 °C). Additionally, slightly lower locomotor activity of this group was seen, thus a treatment-related effect can not be excluded.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Absolute and relative mean liver weights (both relative to body weight and relative to brain weight) were statistically increased in males and females of group 4 (90 mg/kg body weight/day). This finding correlated to diffuse hepatocellular hypertrophy associated with follicular hypertrophy of thyroid glands on some males and females of group 4.

Additionally, absolute and relative mean thymus weights were statistically significantly decreased in females of group 4, correlated with the observation of reduced size of thymus and thymic atrophy noted at histopathological examination.

No experimental relevance was attributed to increased relative mean kidney and heart weights in males and females of group 4 as the mean values were within the range of historical control data and no corroborative histopathological findings were recorded.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

The relevant macroscopical change in the stomach recorded as crateriform retraction was noted in one female at 30 mg/kg body weight/day, correlating with ulceration and submucosal inflammation.

One, three and four females receiving 10, 30 and 90 mg/kg body weight/day of the test item had reduced sized thymus correlating with thymic atrophy and reduced organ weights at 90 mg/kg body weight/day only. This thymus finding was considered to be likely stress-related. Other isolated macroscopical findings were considered to be within the range of normal background alterations.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Under the conditions of this experiment, the following treatment-related local irritative effects were induced:

Forestomach: hyperkeratosis in some females receiving 30 mg/kg body weight/day and all males receiving 90 mg/kg body weight/day and a single female at 90 mg/kg body weight/day, squamous cell hyperplasia with submucosal inflammation in a female at 30 mg/kg body weight/day; glandular stomach: focal erosion in one male at 10 mg/kg body weight/day and one male and female each at 90 mg/kg body weight/day, in the female associated with submucosal inflammation, mucosal atrophy.

Under the conditions of this experiment, the following treatment-related adaptive findings that are commonly seen in rats exposed with xenobiotics were induced:

Minimal to slight diffuse hepatocellular hypertrophy, characterized by an increase in
endoplasmic reticulum, was recorded in liver in all males and females at 90 mg/kg body
weight/day. This finding was considered to be adaptive.

Diffuse follicular cell hypertrophy in thyroid gland was noted in some males and females at 90 mg/kg body weight/day. This finding was considered to be consequent to the hepatocellular hypertrophy.

Histopathological examination of female no. 78 which died spontaneously receiving 90 mg/kg
body weight/day revealed that the cause of death was likely the thrombosis of jejunum and,
therefore, this death was considered to be incidental and not test item related.

Histopathological examination of the male no. 35 in the 90 mg/kg body weight/day group, which was killed in extremis, showed slight diffuse hyperkeratosis in forestomach and minimal erosion in glandular stomach, signs of moderate stress in thymus and adrenal glands as well as slight diffuse atrophy in the prostate, coagulating gland and seminal vesicles. The cause of moribundity could not be determind from the organs and tissues determined.

Histopathological findings: neoplastic:

not examined

Other effects:

effects observed, non-treatment-related

Description (incidence and severity):

LOCOMOTOR ACTIVITY
For females at the dose level of 90 mg/kg body weight/day, total level of locomotor activity was statistically significantly reduced. In males the total level of activity was lower when compared to controls or other test item-treated groups but it did not reach statistical significance. The reduced locomotor activity in animals receiving 90 mg/kg body weight/day of Irgacure 127 was considered to be a test item-related effect.
In females at the dose level of 30 mg/kg body weight/day the low beam count at 30 minutes was
lower when compared to control but the total level of activity was not affected in this dose group.

Key result

Dose descriptor:

NOAEL

Effect level:

30 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical signs

food consumption and compound intake

mortality

organ weights and organ / body weight ratios

Critical effects observed:

yes

Lowest effective dose / conc.:

30 mg/kg bw/day (nominal)

System:

other: not reported

Organ:

liver

thymus

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

not specified

Conclusions:

Based on the observed mortality, clinical symptoms, decreased body weight, decreased food consumption, increased liver and decreased thymus weights at the dose level of 90 mg/kg body weight/day the NOAEL (No Observed Adverse Effect Level) for general toxicity in parental males and females was considered to be 30 mg/kg body weight/day.
A NOAEL (No Observed Adverse Effect Level) for the local irritative effect of the test item in
stomach could not be established for males and was established at 10 mg/kg bw/day for females based on the microscopic findings detected.

Executive summary:

The purpose of this guideline (OECD 422) study conducted with GLP certification, of the test material (EC: 444 -860 -9) was to generate preliminary information concerning the effects of the test item on the possible health hazards likely to arise from repeated exposure over a relatively limited period of time. In addition it provides information on possible effects on male and female reproductive performance such as gonadal function, mating behavior, conception, development of the conceptus and parturition.

The test item was administered orally by gavage to male rats for 28 days and to female rats for 14 days prior to pairing, through the pairing and gestation periods until the F1 generation reached day 4 post partum.

The test was conducted at three dose levels (10, 30, & 90 mg/kg bw/day). The duration of treatment covered a 2-week premating and a mating period in both sexes, approximately 1 week post-mating in males, and the entire gestation period as well as 4 days of lactation and 2 weeks thereafter in females. Treatment related toxicity was observed - the pups were shown to have a lower bodyweight and a decreased viability index. The NOAEL was determined to be 30 mg/kg bw/day.

A standard dose volume of 10 mL/kg body weight with a daily adjustment to the actual body weight was used. Control animals were dosed with the vehicle alone (Carboxymethylcellulose sodium salt 0.5%).

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

30 mg/kg bw/day

Study duration:

subacute

Species:

rat

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Procedure and observations

The test item was administered orally by gavage to groups of 10 male and 10 female Wistar rats (F0 animals) at dose levels of 0 mg/kg body weight/day (CMC as vehicle), 10 mg/kg bw/d, 30 mg/kg bw/d and 90 mg/kg bw/d. The duration of treatment covered a 2-week premating and a mating period in both sexes, approximately 1 week post-mating in males, and the entire gestation period as well as 4 days of lactation and 2 weeks thereafter in females. A detailed clinical observation was performed in all animals. Body weights and food consumption were determined in F0 animals. Clinicochemical and hematological examinations as well as urinalyses and FOB were performed in all animals towards the end of the administration period. All animals were assessed by gross pathology; weights of selected organs were recorded and a histopathological examination was performed.

At 90 mg/kg body weight/day, one male was killed for ethical reason, relationship to the treatment could not be excluded. Test item-related clinical signs, i.e. hunched posture, ruffled fur and discoloured feces, were observed of males and females of the high dose group. Food consumption and body weight gain in high dose males and females was found to be lower when compared to control. The animals showed also a slightly lower motor activity. Liver weights were increased in group 4 (90 mg/kg body weight/day). Additionally, thymus weights were decreased in females of the 90 mg/kg body weight/day group. Treatment-related adverse local irritative effects in the stomach were observed in all test item-treated groups at incidences of 1, 2 and 2 animals at the dose levels 10, 30 and 90 mg/kg body weight/day, respectively. Thymus atrophy and lowered thymus weight as well as liver cell hypertrophy was observed in (mid and) high dose animals.

Discussion

The reduced size in thymus in females, occurring in increased incidences, at ≥ 10 mg/kg body weight/day correlated histopathologically with athropy and reduced absolute and relative thymus weights only at the high dose levels. Although this is considered to be most likely to be stress related a test item-relationship cannot be excluded at the 90 mg/kg body weight/day dose level.

Treatment-related diffuse liver cell hypertrophy at 90 mg/kg body weight/day was considered to be adaptive. This hypertrophy was associated with follicular cell hypertrophy in thyroid glands in some males and females at 90 mg/kg body weight/day. Most probably the enhanced liver cell metabolism resulted in an acceleration of thyroid hormone breakdown with consequent thyroid follicular cell hypertrophy. This finding is commonly seen in rats after being exposed to xenobiotics.

The No-observed-effect-level for systemic toxicity is considered to be 30 mg/kg bw/day.

In addition to the above mentioned OECD 422 study, two range finding studies and a 28 day study were performed. In the course of the subacute study, 5, 50 and 160 mg/kg bw/day were administered by gavage to male and female wistar rats. Due to mortality in the high dose group an additional group was treated with 100 mg/kg bw/day. Similar to the findings of the OECD 422 study, gastric erosions were observed. Additionally, an adverse, nephrotoxic effect could be recorded in animals treated with 50, 100 and

160 mg/kg/day. The NOAEL was established at 5 mg/kg bw/day.

Repeated dose toxicity: via oral route - systemic effects (target organ) digestive: stomach

Justification for classification or non-classification

Classification, Labeling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. As a result the substance is considered to be classified as H373 STOT RE 2 (Kidneys, Thymus) under Regulation (EC) No. 1272/2008.