Methyl (R)-amino(4-hydroxyphenyl)acetate

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

end on 28-NOV-1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: study performed according to OECD guideline and GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Report date:

1998

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Methyl (R)-amino(4-hydroxyphenyl)acetate

EC Number:

253-657-2

EC Name:

Methyl (R)-amino(4-hydroxyphenyl)acetate

Cas Number:

37763-23-8

Molecular formula:

C9H11NO3

IUPAC Name:

methyl (2R)-2-amino-2-(4-hydroxyphenyl)acetate

Details on test material:

- Name of test material (as cited in study report): HPGM
- Molecular formula (if other than submission substance): C9H11NO3
- Structural formula attached as image file (if other than submission substance): see Fig.
- Substance type: monoconstituent substance
- Physical state: powder
- Stability under test conditions: Stable for at least 5 hours in propylene glycol
- Storage condition of test material: at room temperature, dry and in the dark

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source:
- Age at study initiation: Charles River, Sulzfeld, Germany
- Weight at study initiation: males 192-196 g, females 156-161 g (mean values of the groups)
- Fasting period before study: data not available
- Housing: Group housing o f 5 animals per sex per cage in stainless steel suspended cages with wire mesh floors. During activity monitoring, animals were individually housed overnight in Macrolon plastic cages with sterilised sawdust provided as bedding.
- Diet: free access to standard pelleted laboratory animal diet
- Water: Free access to tap-water
- Acclimation period: at least 5 days before start of treatment under laboratory conditions

ENVIRONMENTAL CONDITIONS
- Temperature: 21°C
- Humidity: 50%
- Air changes: approximately 15 air changes per hour
- Photoperiod: 12 hours artificial fluorescent light and 12 hours dark per day

IN-LIFE DATES:
- Treatment : 14 July 1998 to 10 August 1998
- Blood sampling : 11 August 1998
- Necropsy : 11 August 1998

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

propylene glycol

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
Formulations (wlw) were prepared daily within 4 hours prior to dosing. Adjustment was made for specific gravity of vehicle.
Storage conditions: at ambient temperature

VEHICLE
- Justification for use and choice of vehicle: based on trial formulations performed at NOTOX
- Concentration in vehicle: data not available
- Amount of vehicle (if gavage): 5 mL/kg bw
- Purity: data not available

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Samples of week 3 formulations were analysed to check stability, homogeneity (highest and lowest concentration) and accuracy of preparations (all concentrations). Analysis were performed by HPLC.
Test substance formulations in propylene glycol were noted as stable for at least 5 hours and formed a homogeneous suspension at the concentrations tested. Analysis of the accuracy of dose preparations revealed values within the range of 96% to 105% of nominal, which was considered to represent an acceptable level of accuracy for formulations of this type.

Duration of treatment / exposure:

28 days

Frequency of treatment:

Once daily, approximately the same time each day, 7 days per week

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 50, 200 and 1000 mg/kg bw
Basis:
other: nominal

No. of animals per sex per dose:

5 males and 5 females per group

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: dose levels were selected on the basis of a 5-day dose range finding study
- Rationale for animal assignment: randomisation

Positive control:

not applicable

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Once prior to start of treatment and on days 8, 15, 22 and 28, clinical sings were also performed outside the home cage in a standard arena.
- Cage side observations: data not available

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once daily

BODY WEIGHT: Yes
- Time schedule for examinations: on days 1, 8, 15, 22 and 28.

Food consumption: weekly

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: immediately prior to scheduled post mortem examination
- Anaesthetic used for blood collection: Yes (ether)
- Animals fasted: Yes
- How many animals: all rats/sex/group
- Parameters checked in table 1 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: immediately prior to scheduled post mortem examination
- Animals fasted: Yes
- How many animals: all rats/sex/group
- Parameters checked in table 1 were examined.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: during week 4 of treatment
- Dose groups that were examined: all animals
- Battery of functions tested: hearing ability, pupillary reflex, static righting reflex, grip strength

Sacrifice and pathology:

GROSS PATHOLOGY: Yes (see table 2). All animals surviving to the end of the observation period were deeply anaesthetised using ether vapour and subsequently exsanguinated. All animals assigned to the study were necropsied and descriptions of all macroscopic abnormalities recorded.
HISTOPATHOLOGY: Yes (see table 2)

Other examinations:

no

Statistics:

The following statistical methods were used to analyse the data.
Univariate one-way analysis of variance was used to assess the significance of intergroup differences.
If the variables could be assumed to follow a normal distribution, the Dunnett-test (many-to-one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control groups for each sex.
The Steel-test (many-to-one rank test) was applied when the data could not be assumed to follow a normal distribution.
All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance.
Group means were calculated for continuous data and medians were calculated for discrete data (scores) in the summary tables.
Test statistics were calculated on the basis of exact values for means and pooled variances. Individual values, means and standard deviations may have been rounded off before printing. Therefore, two groups may display the same printed means for a given parameter, yet display different test statistics values.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

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:

effects observed, treatment-related

Description (incidence and severity):

see details below

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

see below

Histopathological findings: neoplastic:

not examined

Details on results:

CLINICAL SIGNS AND MORTALITY:
No mortality occurred during the study period.
Hunched posture was noted in one female treated at 1000 mg/kg/day from day 19 onwards. In addition, red staining of the skin on head or neck were noted in this female and three other females of this dose group.
Incidental findings that were noted included salivation, alopecia and brown staining of the skin. These findings were considered to be within the range of biological variation for rats of this age and strain.

BODY WEIGHT AND WEIGHT GAIN:
Body weights and body weight gain of treated animals remained in the same range as controls over the 4-week study period.

HAEMATOLOGY:
Haematological parameters of treated rats were considered not to have been affected by treatment.

CLINICAL CHEMISTRY:
There were no differences noted between control and treated rats that were considered to be related to treatment with HPGM.

NEUROBEHAVIOUR:
No abnormalities were observed in the functional observation tests. The variation in motor activity did not indicate a relation with treatment. The
values for motor activity in control males were slightly low when compared to historical data.
An extremely increased motor activity was noted in one male and six females. In the absence of a dose-dependent relationship this was considered to have occurred by chance without any toxicological relevance.

ORGAN WEIGHTS:
Liver : body weight ratios were increased in males and females treated at 1000 mg/kgd/da y. Kidney weights and kidney : body weight ratios were increased in males of this dose group only.
Statistically significant changes between heart weights of control and 200 mg/kg/day treated females were considered not to be a sign of toxicity since no dose response relationship or any corroborative findings in the other animals were detected.

GROSS PATHOLOGY:
Macroscopic observations at necropsy did not reveal any alterations that were considered to have arisen as a result of treatment.

HISTOPATHOLOGY: NON-NEOPLASTIC
Midzonallcentrilobular hypertrophy was recorded at a minimal or slight degree in four males and one female of the 1000 mg/kg/day dose group. This alteration was accompanied in females by an increased incidence of minimal or slight hepatocellular vacuolation.
The remainder of microscopic findings were within the range of background pathology encountered in Wistar rats of this age and strain and occurred at similar incidences and severity in both control and treated rats.

Effect levels

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: The observed changes were only minimal in severity, did not indicate clear organ dysfunction or can be described as an adaptive response to the test substance.

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

From the results presented in this report a No Observed Adverse Effect Level (NOAEL) of 1000 mg/kg/day was concluded, since the observed changes were only minimal in severity, did not indicate clear organ dysfunction or can be described as an adaptive response to the test substance.

Executive summary:

In a subacute toxicity study (OECD 407, GLP), HPGM was administered to Wistar rats (5/sex/dose level) by gavage at dose levels of 0, 50, 200 and 1000 mg/kg bw/day for 28 days.

The following parameters were evaluated: clinical signs daily; functional observation tests; body weight and food consumption weekly; clinical pathology and macroscopy at termination; organ weights and histopathology on a selection of tissues.

 

Results:

No treatment-related findings noted at 50 and 200 mg/kg bw/day.

At 1000 mg/kg bw/day:

-        Hunched posture and red staining of the skin noted in females,

-        Increased 1iver:body weight ratios noted in males and females. Increased kidney weights and kidney:body weight ratios noted in males,

-        Hepatocellular hypertrophy observed in males and females, accompanied in females by hepatocellular vacuolation.

 

Conclusion:

From the results presented in this report a No Observed Adverse Effect Level (NOAEL) of 1000 mg/kg/day was concluded, since the observed changes were only minimal in severity, did not indicate clear organ dysfunction or can be described as an adaptive response to the test substance.