Biphenyl-2-ol, ethoxylated, esters with acrylic acid

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Remarks:

90D study

Type of information:

experimental study

Adequacy of study:

key study

Study period:

May 2016 - November 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

yes

Remarks:

The order of sacrifice was not decided at random. No statistical analysis was done for food or water consumption due to the small sample size. These deviations were considered not to have affected the integrity or validity of the study.

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

2-phenoxyethyl acrylate

EC Number:

256-360-6

EC Name:

2-phenoxyethyl acrylate

Cas Number:

48145-04-6

Molecular formula:

C11H12O3

IUPAC Name:

2-phenoxyethyl prop-2-enoate

Specific details on test material used for the study:

- Name of test material (as cited in study report): MIRAMER M140
- Physical state: Colorless liquid
- Analytical purity: 84.46 % ( CAS No.: 48145-04-6; EC No.: 256-360-6)
- Impurities (identity and concentrations): cas no. 61630-25-9; 8.67%; cas no. 122-99-6; 1.48 %
- Lot/batch No.: 151014142
- Expiration date of the lot/batch: 31 December 2016
- Storage condition of test material: Under dry conditions, protected from light and at room temperature (20 ± 5 ºC)
Supplier: Miwon

Test animals

Species:

rat

Strain:

Wistar

Remarks:

Wistar Hannover

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Rat, Wistar Hannover
- Females (if applicable) nulliparous and non-pregnant: yes.
- Age at study initiation: 6-9 weeks.
- Weight at study initiation: Males: 231-285 g, Females: 152-199 g.
- Fasting period before study: NO.
- Housing: Same-sex groups of no more than five; housed in Makrolon type IV cages with Lignocel S8-15 sawdust bedding (supplied by J. Rettenmaier & Söhne).
- Diet (e.g. ad libitum): ad libitum.
- Water (e.g. ad libitum): ad libitum.
- Acclimation period: 7 days.

DETAILS OF FOOD AND WATER QUALITY:
Each batch of food is analyzed by the manufacturer for contaminants and to check its composition. Results of analysis for composition and contaminants will be archived at Envigo CRS, S.A.U.
Results of drinking water bacteriological, chemical and contaminant analyses are included in the raw data.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24ºC.
- Humidity (%):30-75%.
- Air changes (per hr): 15-20 air changes per hour.
- Photoperiod (hrs dark / hrs light): 12 hours fluorescent light/12 hours dark.

IN-LIFE DATES: From: To: 04 May 2016 to 10 August 2016.

Administration / exposure

Route of administration:

oral: gavage

Details on route of administration:

Oral, by gastric gavage,
The amount of test item to be administered was calculated according to the most recent body weight recorded.
Control animals were administered with the vehicle following the same dosing regimen as that used for animals treated with the test item.

Vehicle:

corn oil

Remarks:

from Sigma-Aldrich

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:

VEHICLE
- Justification for use and choice of vehicle (if other than water):
The test item is soluble in corn oil that is non-toxic in the volume used.
- Concentration in vehicle: NA
- Amount of vehicle (if gavage): 4 mL/kg
- Lot/batch no. (if required): MKBV2080V & MKBQ9948V
- Purity: NA

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Results obtained in the analysis done during the study showed:
I. An accuracy interval for all formulations of 86-100% and 0.2-2.6 for precision.
II. The analysis of Group 1 formulations showed that no contamination was present.
The results obtained demonstrate that the means of the concentrations deviated by less than 15% from their nominal value (accuracy range: 85.65-99.79%). Precision was below 4%.

Duration of treatment / exposure:

Single gastric gavage

Frequency of treatment:

Once daily seven days per week.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Remarks:

Vehicle only

Doses / concentrations 2

Dose / conc.:

30 mg/kg bw/day (nominal)

Remarks:

2-Phenoxyethyl Acrylate

Doses / concentrations 3

Dose / conc.:

100 mg/kg bw/day (nominal)

Remarks:

2-Phenoxyethyl Acrylate

Doses / concentrations 4

Dose / conc.:

350 mg/kg bw/day (nominal)

Remarks:

2-Phenoxyethyl Acrylate

No. of animals per sex per dose:

Groups 1 to 4: 10 males and 10 females each (Main study).
Groups 1 and 4: 5 males and 5 females each (Recovery).

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale:
- Rationale for animal assignment (if not random): NA
- Rationale for selecting satellite groups: to test the reversibility or progression of any test-item-related changes was assessed in recovery animals (recovery group) after a 4-week treatment-free recovery period.
- Post-exposure recovery period in satellite groups: 4 weeks
- Section schedule rationale (if not random): NA

Positive control:

No

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily, clinical signs daily and
- Cage side observations checked in table were included.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Detailed clinical signs were recorded once weekly.

BODY WEIGHT: Yes
- Time schedule for examinations: Once during acclimatization, twice weekly during treatment and recovery periods and before sacrifice.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined before treatment start and once a week during treatment period

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: Not specified


OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Observations were performed once at acclimatization or pretest and in treatment week 13.
- Dose groups that were examined: Groups 1 and 4

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Week 13 of treatment (Main Study animals) and
Week 4 of recovery (Recovery animals).
- Anaesthetic used for blood collection: Yes (identity) light isoflurane anesthesia.
- Animals fasted: Yes
- How many animals: all animals in the respective groups.
- Parameters checked in table [No.?] were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Week 13 of treatment (Main Study animals) and
Week 4 of recovery (Recovery animals).
- Animals fasted: Yes
- How many animals: all animals in the respective groups.
- Parameters checked in table [No.?] were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: Week 13 of treatment (Main Study animals) and
Week 4 of recovery (Recovery animals).
- Metabolism cages used for collection of urine: Not specified
- Animals fasted: Yes
- Parameters checked in table [No.?] were examined.

FOB (functional observational battery): yes
-sensory reactivity to stimuli of different types was measured for all rats in week 13 and after a 4 week recovery period
-grip strength in week 13
-locomotor acitivity in week 13

IMMUNOLOGY: No
- Time schedule for examinations: NA
- How many animals: NA
- Dose groups that were examined: NA
- Parameters checked in table [No.?] were examined. NA

OTHER:
FOB (functional
observational battery)

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
Sacrifice:
After 13 weeks of treatment: 80 animals (main study)
After 4 weeks of recovery: 20 animals (recovery)
As the total number of animals exceeds the number that can be sacrificed in one day, the necropsies were carried out on three consecutive days. As far as possible, the same number of animals of each group and sex was sacrificed each day. The animals continued receiving the test item or vehicle until the day before sacrifice.
The animals surviving to the end of the observation period were deprived of food and deeply anesthetized with sodium pentobarbital administered intraperitoneally and killed by exsanguination.
A full necropsy was performed on all main study and recovery animals. The necropsy included the examination of the external surface of the body, all orifices, cranial, thoracic and abdominal cavities and the observation of the organs both in situ and after evisceration. Descriptions of all macroscopic abnormalities were recorded.
Samples of tissues and organs were collected and weighed (see list in Annex 5) from all animals at necropsy and fixed, unless otherwise indicated, in neutral phosphate-buffered 4% formaldehyde solution (10% formalin) or the correspond fixative. Animal identification (such as ear tattoo) was retained for identification purposes.


HISTOPATHOLOGY: Yes
All organ and tissue samples were examined by the study pathologist (see Annex 5) processed, embedded and cut at a nominal thickness of 2-4 micrometers and stained with hematoxylin and eosin.
The bone marrow smears were stained using the May Grünwald-Giemsa Method and kept at the test facility for possible further investigation.

Other examinations:

Specialist observations:

-Ophthalmoscopic examination: Observations were performed once at acclimatization or pretest and in treatment week 13 (Groups 1 and 4).
Pupils were dilated by instillation of ®Colircusí Tropicamida eye drops (batch numbers: 5KFP1B, 5LCK1D and 6FSF1B expiry dates: September 2018, November 2018 and January 2019, respectively).
Observation areas included: cornea, lens, conjunctiva, sclera, iris and fundus


-FOB (functional observational battery): In week 13 of treatment: sensory reactivity to stimuli of different types was measured for all rats (main study and recovery). Relevant parameters form a modified Irwin screen test (blink, pinna, iridic light reflex, push-off (hind legs), pain response, startle/hearing and righting reflex).
Observations were based on procedures used for the detailed weekly observations. Any abnormal findings were recorded and rated in severity.
In week 4 of recovery: sensory reactivity was not recorded in all recovery animals, as findings recorded during week 13 were considered not to be treatment related.

-Grip strength: In week 13 of treatment: hind- and forelimb grip strength were measured with a push-pull strain gauge in the main study.

-Locomotor activity: In week 13 of treatment: was measured quantitatively (beam counts, number of light/shadow (changes)) in main and recovery animals. Decreased or increased activity was recorded. Activity was measured with an AMS from Medical Instruments GmbH (FMI) and DeMeTec-Ams version 2.0 GmbH. Activity of the animals was recorded in 10-minute intervals over a 60-minute period with fluorescent light. These data and the total activity over 60 minutes were reported.
In week 4 of recovery: all recovery animals were measured in order to discard any possible treatment-related effect observed during week 13.

Statistics:

All statistical analyses were carried out separately for males and females using the individual animal as the basic experimental unit.
The following data types were analyzed at each time point separately:
Food and water consumptions
Body weight
FOB: grip strength and locomotor activity
Hematology and Coagulation
Blood chemistry
Urinalysis
Macropathology
Organ weights, absolute and adjusted to body weight values

The following comparisons were performed:
Group 1 vs 2, 3 and 4

A parametric analysis was performed if Bartlett's test for variance homogeneity was not significant at the 1% level. The F1 approximate test was applied. If the F1 approximate test was significant, suggesting that the dose response was not monotone, Dunnett's test (Dunnett 1955, 1964) was performed instead.
A non-parametric analysis was performed if Bartlett's test was still significant at the 1% level following both logarithmic and square-root transformations
For clinical pathology data, if 75% of the data (across all groups) were the same value, for example c, Fisher’s exact tests (Fisher, 1973) were performed. Treatment groups were compared using pairwise comparisons of each dose group against the control
For organ weight data, analysis of covariance was performed using terminal body weight as covariate (Angervall and Carlström, 1963), unless non-parametric methods were applied. The treatment comparisons were made on adjusted group means in order to allow for differences in body weight which might influence the organ weights.
Significant differences between the groups compared were expressed at the 5% (p ≤ 0.05) or 1% (p ≤ 0.01) level.
No statistical analysis was done on data obtained in the food / water consumption or during recovery period given the small sample size.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Salivation was observed in Group 4 (350 mg/kg/day), in both sexes, during the course of the study.

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Description (incidence and severity):

There were no differences compared to controls in the overall body-weight gain that could be related to treatment.

Food consumption and compound intake (if feeding study):

no effects observed

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

no effects observed

Ophthalmological findings:

no effects observed

Description (incidence and severity):

The ophthalmic examination in week 13 did not indicate any test-item related finding.

Haematological findings:

no effects observed

Description (incidence and severity):

No alterations in hematology or coagulation were observed.

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

No changes of toxicological relevance were observed in blood biochemistry parameters.
At the end of treatment period, the following statistically significant differences with respect to the Control group (Group 1) were recorded:
I. In males from Group 4, higher triglyceride and potassium and lower phosphorus values.
II. In females from Groups 3 and 4, higher glucose and urea values.
III. In females from Group 2, higher protein and albumin values.

However, these differences were devoid of any toxicological significance and were attributed to the normal biological variability.

Urinalysis findings:

no effects observed

Description (incidence and severity):

No noteworthy changes were recorded in the urinalysis.

Some minor variations among groups, such as a statistically significant higher pH in the females from Groups 3 and 4 with respect to Group 1 were recorded after 13 weeks of treatment. However, these differences were devoid of any toxicological significance

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

There were no noteworthy differences from Group 1 for organ weights that were considered of toxicological relevance.

A slightly higher statistically significant increase in absolute (about 10%) and adjusted liver weight was recorded in males from Group 4 treated at 350 mg/kg/day with respect to the Control group (Group 1).

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

No histopathological changes related with treatment with 2-Phenoxyethyl Acrylate were seen in this study.
At the end of the treatment period, centrilobular hepatocellular hypertrophy was observed in a single female in Group 4 at a minimal level. However, given its low incidence and distribution, it was considered to be incidental and unrelated to treatment.

Histopathological findings: neoplastic:

not examined

Effect levels

Key result

Dose descriptor:

NOAEL

Effect level:

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

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Systemic and local effects

Target system / organ toxicity

Key result

Critical effects observed:

no

Applicant's summary and conclusion

Conclusions:

The study was performed according to OECD Guideline with minor deviations that did not compromising the integrity of the study. A NOAEL was established to 350 mg/kg bw/day for systemic and local effects for both sexes.

Executive summary:

In this OECD Guideline 408 study Wistar rats (10 animals per sex/ dose level) were dosed by gavage during 90 days at levels of 0, 30, 100 and 350 mg/kg/day. No unscheduled deaths occurred, and there were no toxicological findings in relation to clinical observation or pathological findings. Salivation at the dose level of 350 mg/kg/day was considered as a test item palatability response and only slight differences in clinical laboratory investigations and some slight changes in organ weights at the high dose level were noted. Under the conditions of this study the dose of 350 mg/kg/day is to be considered as a NOAEL both in relation to systemic effects as well as local effects.

It is to be noted that 350 mg/kg bw/day was selected as the highest dose level for the study as inflammatory infiltrates in submucosa and forestomach ulcerations was observed at dose levels of 300 mg/kg bw/day and 800 mg/kg bw/day in a dose range finding study for an oral OECD 422 study. Thus, for longer term exposure an adaptation to the gastric irritational effects seems to occur as such effects were not observed in this 90D study.