A mixture of RR and RS isomers of: (2-(2-methoxy-1-methyl)ethoxy)-1-methylethyl acetate; (2-(2-methoxy-2-methyl)ethoxy)-1-methylethyl acetate; (2-(2-methoxy-2-methyl)ethoxy)-2-methylethyl acetate; (2-(2-methoxy-1-methyl)ethoxy)-2-methylethyl 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:

1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-study according to OECD guideline 407.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: Annex V, Published in the official journal of E.C.(No. L25I, 19 September 1984)

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

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

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Details on test material:

- Name of test material (as cited in study report): T-4388
- Physical state: Colorless Liquid
- Lot/batch No.: ( 6 )
- Storage condition of test material: In the dark at ambient temperature.

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River ( UK )
- Age at study initiation: 5-6 week
- Weight at study initiation: Male -144-150 gms and Female - 107 - 111 gms
- Housing: Rats were housed 2 or 3 of one sex per cage in suspended polypropylene cages with stainless steel wire grid tops and bottoms
- Diet ( ad libitum): SQC Expanded rat and mouse maintenance diet No. 1 supplied by special diet services limited,Stepfield, Witham, Essex, CM8 3 AB.
- Water ( ad libitum):Tap water
- Acclimation period: 8 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 °C - 22 °C
- Humidity (%): 45 - 65 %
- Air changes (per hr): 15-20
- Photoperiod : (12hrs dark / 12hrs light)

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: Distilled water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: Dosing solutions were prepared daily using distilled water as vehicle.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

See the attachment - 2

Duration of treatment / exposure:

28 days

Frequency of treatment:

Daily

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
100 mg/kg
Basis:
other: nominal concentration

Doses / concentrations 2

Remarks:

Doses / Concentrations:
250 mg/kg
Basis:
other: nominal concentration

Doses / concentrations 3

Remarks:

Doses / Concentrations:
1000 mg/kg
Basis:
other: nominal concentration

No. of animals per sex per dose:

5 animals per sex per dose

Control animals:

yes, concurrent no treatment

Details on study design:

- Dose selection rationale: Dose levels were selected for this is on the basis of preliminary one week oral range finding study , conducted by Inveresk Research International.

Positive control:

no data

Examinations

Observations and examinations performed and frequency:

-CAGE SIDE OBSERVATIONS: No
-DETAILED CLINICAL OBSERVATIONS: Yes, Weekly
-BODY WEIGHT: Yes
-Water Consumption : Yes, Weekly
-Time schedule for examinations: Weekly
-FOOD EFFICIENCY: Not Examined
-OPHTHALMOSCOPIC EXAMINATION: No
-HAEMATOLOGY: Yes
- Time schedule for collection of blood: During 4th week of dosing
- Anaesthetic used for blood collection: Yes ( Light Ether Anaesthesia )
- Animals fasted: Yes
- How many animals: 40
- Parameters checked : Hb, RBC, HCT, MCH, MCV, MCHC, WBC, DLC
CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood:During 4th week of dosing
- Animals fasted: Yes
- How many animals: 40
- Parameters checked : BUN, Glu, AST, ALT, AP,Na, K, Cl, TP, Alb, AG-R, Crea, Ca, Phos, T. Bl.
URINALYSIS: No
NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

Other examinations:

none

Statistics:

Hematology, Clinical chemistry, organ weight and body weight data were statistically analysed for homogeneity of variance using the F-max test. If the group variances appeared homogenous a parametric ANOVA was used and pairwise comparisons made via student’s T test using Fischer’s F- protected LSD. If the variances were heterogeneous log or square root transformations were used in an attempt to stabilize the variances. If variances remained heterogeneous then a non parametric test such as Kruskal- Wallis ANOVA was used.
Organ weights were also analysed conditional on body weight. Histological data were analysed by Fisher’s Exact Probability test

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):

no effects observed

Food efficiency:

not examined

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

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

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:

CLINICAL SIGNS AND MORTALITY : Increased breathing and subdued behavior were evident in the majority of animals of both sexes receiving T-4388. There were no deaths found during 4 weeks of dosing period.

BODY WEIGHT AND WEIGHT GAIN : There were no notable intergroup differences in either sex during 4 weeks of oral administration of T-4388.

FOOD CONSUMPTION :There were no notable intergroup differences in either sex during 4 weeks of oral administration of T-4388.

FOOD EFFICIENCY : Not examined

HAEMATOLOGY :There were no notable intergroup differences in either sex .

CLINICAL CHEMISTRY : There were no notable intergroup differences in either sex .

ORGAN WEIGHTS : There was a slight increase in liver in high dose females.

GROSS PATHOLOGY : There were no notable intergroup differences in either sex .

HISTOPATHOLOGY: NON-NEOPLASTIC : There were no findings considered to be due to administration of the test material.

Effect levels

1

Dose descriptor:

NOEL

Effect level:

250 mg/kg bw/day (nominal)

Sex:

male/female

2

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (actual dose received)

Sex:

male/female

Basis for effect level:

other: Slight increase in liver weight of females not accompanied by histopathological changes.

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

The only effects observed during this study were increased liver weights and subdue behaviour and increased breathing. The liver weight increase was very slight and only observed in females of the highest dose group. There were no histopathologic changes accompanying this effect. The same effect was observed with other structurally related molecules, e.g. propylene glycol methyl ether has been shown to cause liver weight increases via a phenobarbital-like enzyme induction mode of action and it is highly likely that dipropylene glycol methyl ether acetate liver weight increases occur via the same mode of action. As this is an adaptive effect typical for many glycol ethers, it is not consered as adverse. The toxicological significance of the observed increased breathing and subdue behaviour is unclear. As these observations were not accompanied by any significant effect, they are considered as not relevant.

Applicant's summary and conclusion

Conclusions:

Based on the results of this study a dose of 1000 mg/kg/day was identified as no observed adverse effect level. Based on a slight increase in liver weight which was not accompanied by histopathologic changes and observed in females only. The no observed effect level is 250 mg/kg/day.

dosing Sprague-Dawley rats with dose levels of 100, 250, 1000 mg/kg/day T-4388 produced increased breathing and subdued behavior in the majority of animals of both sexes receiving T-4388 and a slight increase in liver weight at 1000 mg/kg/day in females
Only.

Executive summary:

Four groups of Sprague- Dawley rats each containing 5 males and 5 females received T- 4388 (colorless liquid) via oral route by steel cannula at dose levels of 0,100, 250, 1000 mg/kg/day daily for 28 days. Rats were received from Charles River (UK) and housed 2 or 3 of one sex per cage in suspended polypropylene cages with stainless steel wire grid tops and bottoms. Animals were kept for acclimatization for 8 days and provided adlibitum feed and water. Temperature maintained in animal rooms was 18 °C - 22 °C with relative humidity of 45 - 65 % and 15-20 air changes.12 hour dark and 12 hour light photoperiod was maintained.

All animals were monitored for toxic effects which included clinical observations, body weights, organ weights, hematology, clinical chemistry, macroscopic and microscopic evaluations. Increased breathing and subdued behavior were evident in the majority of animals of both sexes receiving T-4388. There were no deaths found during 4 weeks of dosing period.

In body weights and feed consumption, there were no notable intergroup differences in either sex during 4 weeks of oral administration of T-4388. In hematology and clinical chemistry, there were no notable intergroup differences in either sex. In organ weights there was a slight increase in liver in high dose females.

There were no notable intergroup differences in either sex in gross pathology. There were no microscopic findings considered to be due to administration of the test material T-4388.

Based on the results of this study a dose of 1000 mg/kg/day was identified as no observed adverse effect level. Based on a slight increase in liver weight which was not accompanied by histopathologic changes and observed in females only. The no observed effect level is 250 mg/kg/day.