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Repeated dose toxicity: oral

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Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1997
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study conducted in compliance with GLP regulations

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1997
Report date:
1999

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
1-phenoxypropan-2-ol
EC Number:
212-222-7
EC Name:
1-phenoxypropan-2-ol
Cas Number:
770-35-4
Molecular formula:
C9H12O2
IUPAC Name:
1-phenoxypropan-2-ol
Details on test material:
- Name of test material (as cited in study report): Protectol PP (#97/109)
- Physical state: liquid / colorless
- Analytical purity: mixture (86/14.9%)
- Composition of test material, percentage of components: 84.8% 1-phenoxy-propan-2-ol and 14.7% 2-phenoxy-propan-1-ol
- Lot/batch No.: 14-1395
- Stability under test conditions: verified by reanalysis
- Storage condition of test material: at room temperature

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Wlstar rats [Chbb:THOM (SPF)] were supplied by Boehringer Ingelheim Pharma KG
- Age at study initiation: 35 days
- Age at study initiation: the body weight of the males was in the range of 166 - 193 g (group mean : 183 g) and the body weight of the females was in the range of 135 - 157 g (group mean 146 g)
- Housing: singly in type DK III stainless steel wire mesh cages supplied by Becker & Co., Castrop-Rauxel, FRG (floor area about 800 cm2). Underneath the cages, waste trays were fixEed containing absorbent material (type 3/4 dustfree embedding, supplied by SSNIFF, Soest, FRG).
- Diet: ground Kliba maintenance diet rat/mouse/hamster, meal, supplied by Klingentalmühle AG, Kaiseraugst, Switzerland; ad libitum
- Water; drinking water (from water bottles); ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24
- Humidity (%): 30 - 70
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:
oral: drinking water
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
The test substance was administered as a solution in drinking water. The test substance was weighed in depending on the dose group, then drinking water was filled up to the desired weight, and mixed with a magnetic stirrer for about 5 minutes. The drinking water solutions were prepared at least twice a week.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analyses of the test substance preparations were carried out. The stability of the test substance in drinking water over a period up to 96 hours at room temperature was proven. Samples for concentration control analyses of all doses were drawn at the start of the administration period as well as about 8 weeks thereafter. As the preparations were real solutions, no homogeneity analyses were performed.
Duration of treatment / exposure:
90 days
Frequency of treatment:
continuously in drinking water
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0, 500, 2000, and 6000 ppm
Basis:
nominal in water
Remarks:
Doses / Concentrations:
0, 35/46, 146/177, and 429/486 mg/kg bw in males/females (calculated based on water consumption)
Basis:
actual ingested
No. of animals per sex per dose:
10
Control animals:
other: normal drinking water
Details on study design:
- Dose selection rationale: after a 4 week study at concentrations of 0 ppm (vehicle control), 1000 ppm, 3000 ppm, and 10000/6000 ppm (11 days 10000 ppm, 17 days 6000 ppm).
- Rationale for selecting satellite groups: no satellite group
- Post-exposure recovery period in satellite groups: not applicable
Positive control:
none

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: the animals were examined for evident signs of toxicity or mortality twice a day (in the morning and in the late afternoon) from Mondays to Fridays and once a day (in the morning) on Saturdays, Sundays and public holidays.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: see above; once a week an additional comprehensive clinical examination was carried out.

BODY WEIGHT: Yes
- Time schedule for examinations: Body weight was determined before the start of the administration period in order to randomize the animals. During the administration period, the body weight was determined on day 0; start of administration period) and thereafter at weekly intervals. The difference between the body weight on the respective day of weighing and the body weight on day 0 was calculated as body weight change.

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Food consumption was determined weekly over a period of 7 days and calculated as mean food consumption in grams per animal and day.

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: Yes

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: water consumption was determined weekly over a period of 3 days and calculated as mean water consumption in grams per animal and day. On few occasions, water bottles were leaking and thus some values are missing in the tables. The mean daily intake of test substance (group means) was calculated based upon individual values for body weight and water consumption.

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Four days prior to the start of the administration period and on day 87 in the animals of high dose and control group were examined.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: after sacrifice at the end of the 3-month administration period.
- Anaesthetic used for blood collection: No data
- Animals fasted: 16-20 hours
- How many animals: 10 animals per test group and sex
- Parameters were examined: leukocytes, erythrocytes, hemoglobin, haematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelets and differential blood count

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: see above
- Animals fasted: 16-20 hours
- How many animals: 10
- Parameters examined: alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, serum-y-glutamyltransferase, sodium, potassium, chloride, inorganic phosphate, calcium, urea, creatinine, glucose, total bilirubin, total protein, albumin, globulins, triglycerides, cholesterol, magnesium

URINALYSIS: Yes
- Time schedule for collection of urine: see above
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters were examined: volume, color, turbidity, nitrite, pH, protein, glucose, ketones, urobilinogen, bilirubin, blood, specific gravity, sediment

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes; all animals were sacrificed by decapitation under COZ-anesthesia. The exsanguinated animals were necropsied and assessed by gross pathology. Weight assessment was carried out on all animals sacrificed at scheduled dates. The weights of the following organs were determined: anesthetized animal, liver, kidneys, adrenal glands, testes/ ovaries, epididymides, thymus, spleen, brain, heart.
HISTOPATHOLOGY: Yes; the following organs / tissues were fixed in 4% formaldehyde solution: all gross lesions, brain, pituitary gland, thyroid glands/ parathyroid glands, thymus, trachea, lungs, heart, aorta, salivary glands (mandibular gland, sublingual gland), liver, spleen, kidneys, adrenal glands, pancreas, testes/ ovaries, uterus/ oviducts/ vagina, epididymides, prostate, seminal vesicle, skin, esophagus, stomach (fore- and glandular stomach), duodenum, jejunum, ileum, cecum, colon, rectum, urinary bladder, lymph nodes (mesenteric and mandibular lymph node), female mammary gland, skeletal muscle, sciatic nerve, sternum with sternal marrow, bone marrow (femur), eyes with Harderian gland, femur with knee joint, spinal cord (cervical, thoracic and lumbar cord), extraorbital lacrimal glands. The hematoxylin-eosin stained slides were examined light-microscopically and assessed.
Statistics:
- Food consumption, water consumption, body weight, body weight change, food efficiency: parametric one-way analysis using F-test (ANOVA; two-sided). If the resulting p-value was equal or less 0.05, a comparison of each group with the control group using the DUNNETT’s test (two-sided) was performed for the hypothesis of equal means.
- Clinical pathology parameters, except differential blood count: non-parametric one-way analysis using KRUSKAL-WALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using MANN-WHITNEY U-test (two-sided) for equal medians.
- Urinanalysis, except volume, color, turbidity and specific gravity: pairwise comparison of each dose group with the control group using FISHER’s exact test for the hypothesis of equal proportions.
- Weight parameters: non-parametric one-way analysis using KRUSKAL-WALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using WILCOXON test for the hypothesis of equal medians.

Results and discussion

Results of examinations

Details on results:
CLINICAL SIGNS AND MORTALITY
Discoloration of urine in bedding was seen in all animals of the mid and high dose group.

BODY WEIGHT AND WEIGHT GAIN
Body weight was statistically significantly decreased in high and mid dose males, resulting in reduced values of 13.6% and 5.7% below control on day 91, respectively. In high dose females, the effect was less prominent and statistically not significant. Nevertheless, the impairment of body weight until day 91 (6.8% below control) might be related to treatment. Corresponding body weight change values on day 91 were 21.6% (high dose males), 9.2% (mid dose males), and 14.9% (high dose females) below controls, respectively.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
Food consumption was statistically significantly decreased in both sexes of the high dose group on several days, the values being up to 12.8% below control in males and 12.1 % below control in females. This was assessed as being related to treatment. By contrast, the single statistically significantly lower value of low dose males on day 21 was assessed as being incidental, due to the isolated occurrence and the lack of a dose-response-relationship.

FOOD EFFICIENCY
Statistically significantly lower values were in low and mid dose males on day 14, and in high dose males on day 7 and 49. Due to the isolated occurrence and the lack of a dose-response relationship, this was assessed as being incidental.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study)
Water consumption was statistically significantly decreased in both sexes of the high dose group on several days, the values being up to 17.9% below control in males and 28.6% below control in females. This was assessed as being related to treatment.

OPHTHALMOSCOPIC EXAMINATION
No substance-related effects were obtained. All findings were spontaneous in nature and equally distributed between treated animals and controls.

HAEMATOLOGY

CLINICAL CHEMISTRY
There are no treatment-related changes in serum enzyme parameters measured. Blood chemistry examinations revealed significant increases in serum urea levels in all treated animals of either sex. The other blood chemistry parameters were not affected by the test compound.

URINALYSIS
This discoloration of the urine was also observed in the previously conducted range finding study and is difficult to explain, as no changes in the kidneys were seen histopathologically. Nevertheless, is has to be assessed as being treatment-related. Concerning clinical pathology, statistically significantly increased urea concentrations were found in all treated males and females. However, these findings are not considered to be test substance-related, because the differences in serum urea concentrations between the controls and the respective treatment groups were only slight in the magnitude of change and did not demonstrate a dose-response- relationship although between the dose levels of test group 1 and 3 there was a 12-fold increase. Urea concentrations of the controls and treatment groups of either sex generally also remained within the normal range of the historical control limits for rats of this age, sex and strain (see tables of "Historical Control Data of Clinical Pathology Testing", Supplement). Only the serum urea concentrations of the females of test group 2 (7.95 mmol/L) were marginally beyond the highest value of the historical controls (7.91 mmol/L). However, this increase is not regarded as being biologically important. Moreover, there was no clinical or histomorphological correlate which explains the elevated urea levels. It is, therefore, reasonable to conclude that the isolated findings of increased urea concentrations in the sera of the treated animals are incidental in nature and not of toxicological relevance. In urinalysis high dose males excreted slightly decreased amounts of urine with increased specific gravity. These changes are regarded to be related to the test compound administered and are possibly indicative of a mild impairment of renal function at dose levels beyond 6,000 ppm.

NEUROBEHAVIOUR
Not assessed

ORGAN WEIGHTS, GROSS PATHOLOGY AND HISTOPATHOLOGY: NON-NEOPLASTIC
Absolute weight and treatment-related histopathological changes were missing in the kidneys of the test animals in all treatment groups. Only the relative kidney weights were increased in both, high dose males and females. Taking into consideration the clinical (decreased water consumption) and clinical pathological data (urine colour, volume, specific gravity), the relative weight changes are as well interpreted as a treatment related effect. All other statistically significant relative organ weight changes in the high dose test animals are seen to be a secondary effect to the impairment of body weight. The slight, light-microscopically noted decrease of the alimentary fat storage intensity in the liver also correlates well with the body weight decrease. The statistical significant increase of the mean weights of the spleen and the adrenal glands in the low and medium male dose groups found no histopathological correlate. Additionally, there is no dose-response relationship (no increase in the male high dose group), and the female test animals do not show a comparable weight increase. Therefore, the statistical significant weight changes of adrenal glands and the spleen are not interpreted to be treatment-related but incidental and of no biological relevance.

OTHER FINDINGS
There are additional statistically significant intergroup differences in the results of clinical Pathology, pathology and histopathology which are marginal, incidental or inconsistent, when compared with the other sex, or lack dose-response relationship. Accordingly, these findings are considered to be of no toxicological significance.

Effect levels

open allclose all
Dose descriptor:
NOEL
Effect level:
ca. 35 - ca. 46 mg/kg bw/day (nominal)
Sex:
male/female
Dose descriptor:
NOAEL
Effect level:
146 mg/kg bw/day (nominal)
Sex:
male
Basis for effect level:
other: Based on impairment of body weight (5.7% below control on day 91 being) and body weight change (9.2% below control on day 91) in males, and discoloration of urine in both sexes

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

The following substance-related findings were observed:

6000 ppm (429 mg/kg bw in males, 486 mg/kg bw in females):

- Decreased water consumption in both sexes

- Decreased food consumption in both sexes

- Impairment of body weight in males and (less pronounced) in females, the values on day 91 being 13.6% and 6.8% below controls, respectively

- Impairment of body weight change in males and females, the values on day 91 being 21.6% and 14.9% below controls, respectively

- Discoloration of urine in both sexes

- Decrease in urinary volume in the males

- Increase in urinary specific gravity in the males

- Increase of relative kidney weight in both sexes 

 

2000 ppm (146 mg/kg bw in males, 177 mg/kg bw in females):

- Impairment of body weight in males, the value on day 91 being 5.7% below control

- Impairment of body weight change in males, the value on day 91 being 9.2% below control

- Discoloration of urine in both sexes

 

500 ppm (35 mg/kg bw in males, 46 mg/kg bw in females):

- No substance-related effects

 

Thus, the oral administration of the test substance in drinking water over 3 months caused effects on water and food consumption, body weight and a mild impairment of renal function. The no observed adverse effect level (NOAEL) under the conditions of this study was 500 ppm (35 mg/kg bw in males, 46 mg/kg bw in females).

Applicant's summary and conclusion

Conclusions:
No treatment related effects were observed at the lowest dose level (500 ppm). At 2000 ppm the only effects observed were a slight decrease in body weight (< 10%) and discoloration of urine. Those effects are not considered to be adverse. Hence, the NOAEL is 2000 ppm.