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Toxicological information

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:
12 February 2019 to 23 March 2019 (experimental dates)
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report Date:
2019

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Version / remarks:
2018
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
Test Material: 3-methylpentane-1,5-diol
Source: Kuraray
Description: Clear colourless liquid
Lot/Batch No.: 93023
Purity: 99.5%
CAS No.: 4457-71-0
Stability of test compound: Confirmed stable for the duration of the study (expiry date: 10 July 2020)
Test substance:
- Name of test material (as cited in study report): 3-Methyl-1,5-Pentanediol
- Molecular weight: 118.20
- Physical state: Colorless liquid
- Analytical purity: 99.18%
- Lot/batch No.: 63136
- Expiration date of the lot/batch:
- Storage condition of test material: Nitrogen substituted, and stored sealed at room temperature in a dark place
- Other: Specific gravity: 0.97 (20°20); Boiling point: 270°C; Freezing point: Less than -50°C; Manufacturer: Kuraray Co.; Supplier: Ministry of Health and Welfare Environmental Health Bureau Planning Section Office for Environmental Chemicals Safety; Date of receipt: 1995-09-6;

Medium:
-Name: Japanese Pharmacopoeia water for injection (Otsuka Pharmaceutical Factory Co.,)
-Lot No. 5C82 and 5K81
-Storage: At room temperature

Test animals

Species:
rat
Strain:
Wistar
Details on species / strain selection:
refer below
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
Species: Rat
Strain: Wistar
Age at dosing: 7-8 wks
Weight at dosing: M: 197 – 240g; F: 154-191g
Source: Charles River, Germany
Acclimation period: 5 days

DETAILS OF FOOD AND WATER QUALITY:
Diet: Altromin 1324 maintenance diet for rats and mice, ad libitum
Water: Municipal water, ad libitum (pH adjusted to pH 2-8)
Housing: Housed 5 animals/cage of the same sex

ENVIRONMENTAL CONDITIONS
Temperature: 22 ±3°C
Humidity: 55 ± 10%
Air changes: ca. 10/h
Photoperiod: 12 h light/dark

IN-LIFE DATES: 12 February 2019 to 23 March 2019 (experimental dates)

Administration / exposure

Route of administration:
oral: gavage
Details on route of administration:
animals dosed orally via gavage
Vehicle:
water
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The test article was prepared at concentrations of 0, 20, 60, and 200 mg/mL. A separate study (Eurofins Munich Study No. 187545) concluded that test article concentrations of 15 and 250 mg/mL were stable for =10 days at room temperature, refrigerated (2-8°C) and frozen (-15 to -35°C). These samples were homogenous after 60 minutes (no stirring). The prepared test formulations were prepared and stored in conjunction with the previous stability data generated.

Verification and homogeneity of the diet preparation containing the test article were determined by the analysis of three samples (from upper, middle, lower strata) from each dose level prepared at the start and end of the dosing phase.

Acceptance criteria for concentration analysis:
-Mean concentration of test article formulation 90-110% of nominal

Stability analysis:
- Stability of test article had previously been verified.
Duration of treatment / exposure:
90 days
Frequency of treatment:
once daily
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (nominal)
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
300 mg/kg bw/day (nominal)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
10/sex/gp
Control animals:
yes, concurrent vehicle
Details on study design:
After an acclimatisation period of ca. 5 days, rats were assigned to groups by randomisation based on weight. The test article, 3 methylpentane-1,5-diol was administered once daily orally to groups of rats for a period of 90 d. Animals (10/sex/gp), were administered the test article orally via gavage at concentrations of 0, 100, 300, 1000 mg/kg bw/d, employing a dose volume of 5 mL/kg bw. Following 90 d of treatment animals were subjected to complete necropsy. Body weight and food consumption were measured at regular intervals. Functional observational battery were performed prior to dosing and during the last week of exposure. Clinical pathology evaluations (haematology) were performed with all surviving animals subjected to complete gross necropsy and full histopathology. Fertility parameters (sperm counts) were determined at necropsy.
Positive control:
not applicable

Examinations

Observations and examinations performed and frequency:
Observations:
Animals were inspected at least once daily for signs of toxicity and mortality. Detailed clinical examinations were conducted once before administration (on the day before the administration start day) and once a week during the administration period.

Body weights:
Animals were weighed at initiation of dosing and weekly during administration and at study termination.

Food consumption:
Determined by weighing food supplied and food that remained were calculated as time-weighted averages at weekly intervals.

Water consumption:
Not conducted

Ophthalmological examination:
Conducted on all animals before the first administration and during the final week of the treatment period.

Neurological functional examinations:
The following evaluations (measurements) were performed prior to initiation of dosing and at wk 12: approach response, auditory stimulation, proprioceptive response, grip strength, rearing, and motor activity.

Haematology and clinical chemistry:
Conducted on days 91. Animals were fasted overnight prior to blood sampling.
Haematology: red blood cell parameters (haematocrit (commonly termed PCV), haemoglobin concentration (Hb), mean haemoglobin concentration (MHC), mean cell haemoglobin concentration (MCHC), mean cell volume (MCV), erythrocyte count, platelet count, reticulocyte count), white blood cell parameters (total and differential (neutrophils, lymphocytes, eosinophils, basophils, monocytes) leukocyte count, large unstained cells), coagulation parameters (activated partial thromboplastin time (APTT), prothrombin time (PT)).

Clinical chemistry: electrolytes (sodium, potassium), kidney function test (creatinine, urea), glucose, liver function tests (albumi, alkaline phosphatase (ALP), alanine aminotransferase (ALT [commonly referred to as glutamic pyruvic transaminase (GPT)]), aspartate aminotransferase (AST [commonly referred to as glutamic oxaloacetic transaminase (GOT), gamma-glutamyl transferase (G-GT)]), total bilirubin (T.Bili), total bile acids (TBA), total protein (TP), lipid profile (total triglyceride, total cholesterol, LDL, HDL), thyroid hormones (T3, T4, TSH).

Toxicokinetics:
Blood samples from 5 animals/sex/gp were taken at 30 minutes and 2 h post dosing in week 1, 4 and 13 to assess for possible systemic exposure and accumulation.

Urinalysis:
Conducted on day 91. The following urinary parameters were measured: specific gravity, pH, protein, glucose, ketones, bilirubin, blood, urobilinogen, leukocyte, nitrite

Organ weights:
Adrenal glands, brain, epididymides, heart, kidney, liver, ovary, pituitary gland, prostate (+seminal vesicles, coagulating gland), spleen, thymus, thyroid (+parathyroid), testis, uterus (+cervix).
Sacrifice and pathology:
Conducted on day 91. Gross pathological examination was performed on all animals and included examination of the external surface, all orifices and associated tissues.
The following tissues were preserved in 4% neutral buffered formalin for subsequent histopathological examination (with the exception of eyes, testes and epididymides which were fixed in Modified Davidson's fixative, then transferred to 70% ethanol) and performed on control and high dose group animals:
Accessory sex glands (M: epididymides, prostate (ventral and dorsolateral), seminal vesicle (coagulating gland), testes; F: ovary, uterus +cervix, vagina), cardiovascular/haematological system (aorta, bone (sternum for marrow), heart, lymph nodes (mesenteric, auxiliary), spleen, thymus), gastrointestinal tract (oesophagus, tongue, stomach, intestine (caecum, colon, duodenum, Peyer's patches, ileum, jejunum, rectum), liver, pancreas, salivary glands (submaxillary, sublingual, parotids'), neurological (brain (cerebellum, cerebrum, midbrain), eyes (+optic nerve, Harderian gland), sciatic nerve, spinal cord (cervical, thoracic, lumbar), respiratory system (trachea, lung), urogenital system (kidneys, urinary bladder), other (skeletal muscle, skin, all gross lesions and masses)
Other endocrine producing/sensitive glands (adrenals, mammary gland, pituitary, thyroid (+parathyroid))
Other examinations:
Spermatogenesis:
At necropsy, the left epididymis and left testis were separated and used for the evaluation of sperm parameters. Epididymal sperm motility and testicular sperm counts were undertaken for all males.

Neurohistopathology: No specific neurohistopathology with specific fixatives were performed in addition to the standard histopathology undertaken on neuronal tissues.
Statistics:
Body weight, food consumption, parameters of haematology, blood coagulation and clinical biochemistry and absolute and relative organ weights were undertaken for each gender by comparing values of dosed animals with the respective control group animals using either a parametric one-way ANOVA and a post-hoc Dunnett Test or a non-parametric Kruskal-Wallis Test and a post-hoc Dunn’s Test, based on the results of homogeneity and normality tests.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Description (incidence and severity):
No treatment related effects observed, with effects occurring sporadically, nor dose related and not reflective in both genders.
Mortality:
no mortality observed
Description (incidence):
All animals survived to scheduled necropsy from both sexes
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Body weight and weight gain were unaffected in either males or females treated up to 1000 mg/kg bw/day over the entire administration period.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
There were no treatment-related changes in food consumption in either sex
Food efficiency:
no effects observed
Description (incidence and severity):
There were no treatment-related changes in food efficiency in either sex
Water consumption and compound intake (if drinking water study):
not examined
Description (incidence and severity):
n/a
Ophthalmological findings:
no effects observed
Description (incidence and severity):
No treatment related effects observed in either sex.
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Males:
Haemoglobin was significantly elevated in mid and high dose group.
Neutrophils counts were significantly decreased in the high dose group
Basophil counts were significantly increased in mid dose group
APTT was significantly longer in high dose group

Females:
Prothrombin time in mid and high dose groups and platelet count in the mid dose gp were significantly increased

These changes were not deemed treatment related for the following reasons:
- Changes in the respective parameters, whilst achieving statistical significance were within the historical control range of the individual values reported in the conducting laboratory.
- The changes observed were not replicated in each sex.
- The effects observed were not dose related.
- In terms of coagulation parameters, these values in the treated groups were comparable to controls, with the statistically significant increases observed set against a low concurrent control value.
- In terms of the neutrophil and basophil counts, no other change in WBC parameters were observed, with no histopathological changes observed in associated tissues.
Refer to Table 7.8.2/02-4
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Males:
AST was significantly increased in the high dose group
K was significantly increased in the high dose group

Females:
Creatinine was significantly lower in the high dose group
Total bile acids were significantly higher in the high dose group

These changes were not deemed treatment related for the following reasons:
- Changes in the respective parameters, whilst achieving statistical significance were within the historical control range of the individual values reported in the conducting laboratory.
- The changes observed were not replicated in each sex.
- In terms of coagulation parameters, these values in the treated groups were comparable to controls, with the statistically significant increases observed set against a low concurrent control value.
- In the absence of any histopathological findings to conclude hepatocellular injury, these effects were deemed not test article related.
- For total bile acids, there were no changes in either ALT, AST with no observed effect on hepatic function.
Refer to Table 7.8.2/02-5
Urinalysis findings:
no effects observed
Description (incidence and severity):
There were no treatment related effects observed in either sex.
Behaviour (functional findings):
not examined
Description (incidence and severity):
n/a
Immunological findings:
not examined
Description (incidence and severity):
n/a
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
Males: a statistically significant increase in relative (to body weight) kidney weight was observed in the high dose group.
Females: a statistically significant increase in absolute kidney weight and relative (to body and to brain) weight was seen for the high dose group compared to the respective controls. In female animals a statistically significant higher absolute liver weight, and relative (to body and to brain) weight was seen for the high dose group compared to the respective control. Increase in kidney weight relative to brain weight was also seen at the mid dose group.
- The values for both organs were within the respective historical control ranges
- No histopathological changes were observed in the respective tissues.
- The effect in the liver were not replicated in males.
No other significant changes in organ weights (including thyroid) were observed.

Refer to Table 7.8.2/02-7
Gross pathological findings:
no effects observed
Description (incidence and severity):
No abnormalities were noted in any animal in the control or test article groups in either sex.
Neuropathological findings:
not examined
Description (incidence and severity):
n/a
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
No test article related effects were observed in either sex, with changes deemed representative of normal background variability within the Wistar rat at the conducting laboratory (refer to Table 7.5.2/02-9).
There was no histological evidence of toxicity in the reproductive organs and tissues including testes, epididymides, prostate gland, seminal vesicles, coagulating glands, ovaries, uterus, cervix, and vagina. In addition, in the investigated testes no treatment-related effects on the testicular histopathology were observed and the histological appearance reflected the animal physiology.
Thymus - minimal to slight multifocal hemorrhage within the thymus and moderate multifocal hemorrhages in the connective tissue adjacent to the thymus were observed in several control and treated animals. Further, minimal multifocal hemosiderin deposits mostly associated with the above mentioned hemorrhages were also observed in some control and high dose animals at incidences comparable to published data (McInnes, 2012). In the absence of additional thymic changes (e.g. necrosis, inflammation), all the above mentioned changes in the thymus were considered most likely incidental were deemed to be not test item related [Firth, 2000, McInnes, 2012, 2017; NTP. Sefanski et al 1990).

Mesenteric lymph nodes - minimal to slight multifocal hemorrhages were observed in several animals from the control and high dose group. Further, slight hemosiderin deposits within macrophages was observed in one female from the high dose group only. Haemorrhages and bron pigment deposition (e.g. haemosiderin, ceroid lipofusin) are background lesions commonly observed in lymph nodes of rats (McInnes, 2000, 2017, Stefanski et al 1990). Furthermore, in this present study the incidence and severity of the above mentioned changes was similar to the published data and all observed in the mesenteric lymph nodes changes were considered to be most likely incidental and deemed to be not test item related (McInnes, 2012, 2017).

References:
Firth, C.H., Ward, J.M., Chandra, M., Losco, P.E. 2000. Non-proliferative lesions of the hematopoietic system in rats. In: HL-1 guides for toxicologic pathology. STP/ARP/AFIP, Washington DC
McInnes, E.F. 2012. Wistar and Sprague Dawley rats. In: McInnes, E.F. and Mann, P. (eds. Background lesions in laboratory animals. A colour Atlas. Saunders Elsevier, Toronto, pp. 17-36
McInnes, E.F. 2017. Common spontaneous and background lesions in laboratory animals. Principles and Practices of Laboratory Animal pathology for Study Personnel. Wiley Blackwell, West Sussex, pp. 62-69
NTP: https://ntp.niehs.nih.gov/nnl/immune/thymus/hemorr/index.htm
Stefanski, S.A, Elwell, M.R., Stromberg, P.E. 1990. Spleen, lymph nodes and thymus. In: Pathology of the Fischer Rat: Reference and Atlas (Boorman G.A, Eustis, S.L., Elwell, M.R., Montgomery, C.A., MacKenzie, W.F. eds). Academic Press, San Diego
Histopathological findings: neoplastic:
not examined
Description (incidence and severity):
n/a
Other effects:
no effects observed
Description (incidence and severity):
Thyroid hormones:
T3 was significantly elevated in females at both the mid and high dose groups, exceeding, but were not deemed biologically relevant for the following reasons:
- Due to the lack of historical control data, in this instance, the biological relevance assessment should be made against the concurrent control group which had an individual animal range of 0.73 – 2.01 ng/mL. In the treated groups, 2/10, 5/10 and 5/10 animals exceeded the upper concurrent vehicle control range for the low, mid and high dose groups respectively. No dose response relationship was evident at the individual animal level. These data are consistent with open literature data drawn on a greater number of studies.
- The changes observed were not replicated in each sex.
- No associated histopathological changes in the liver, thyroid or pituitary gland were observed.
- Both T4 and TSH levels were comparable with the concurrent vehicle control.
Refer to Table 7.8.2/02-6

Spermatogenesis:
There were treatment related effects observed in mean testis weights, mean sperm count, mean sperm motility for all dose groups in the treatment period. Mean total number of abnormal and normal sperms/findings and sperm morphology for high dose group animals were comparable with the vehicle control.
Refer to Table 7.8.2/02-8

Effect levels

Dose descriptor:
NOAEL
Effect level:
> 1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects

Target system / organ toxicity

Critical effects observed:
no

Any other information on results incl. tables

Table 7.5.1/02-1: Overview of sub-chronic toxicity study in rats treated orally (via gavage) with 3-methylpentane-1,5-diol: dose formulation analysis

Parameters

Dose (mg/mL)

20

60

200

Week 1

90.8

90.5

98.6

Week 5

99.1

98.2

105.1

Week 9

97.8

98.6

105.7

Week 12

97.0

91.4

96.

Table 7.5.1/02-2: Overview of sub-chronic toxicity study in rats treated orally (via gavage) with 3-methylpentane-1,5-diol: Functional observations

Parameters

 

M (mg/kg bw/d)

F (mg/kg bw/d)

Wk

0

100

300

1000

0

100

300

1000

Open field grooming

-1
13

0.00
0.40

0.10
0.00**

0.00
0.00**

0.00
0.00**

0.20
0.00

0.10
0.00

0.00
0.00

0.00
0.00

Response to handling

-1
13

4.20
4.00

4.30
3.80

4.00
3.70

4.10
3.00**

4.60
4.20

4.40
4.20

4.20
4.10

4.40
4.00

Spontaneous activity

-1
13

4.00
4.00

4.00
4.00

4.00
3.90

3.90
3.50**

4.00
4.00

4.00
4.00

4.00
4.00

4.00
4.00

Rearing supported

-1
13

2.70
4.00

4.50
4.50

3.80
4.10

2.80
2.20*

5.60
4.10

4.60
5.10

5.10
6.00

4.20
4.90

Temperature (°C)

-1
13

37.99
38.440

38.220
37.700*

38.330
37.870*

38.040
38.240

38.130
38.990

38.310
39.00

38.440
38.650

38.300
38.940

Table 7.5.1/02-3: Overview of sub-chronic toxicity study in rats treated orally (via gavage) with3-methylpentane-1,5-diol: body weight effects

Parameters

M (mg/kg bw/d)

F (mg/kg bw/d)

0

100

300

1000

0

100

300

1000

Body wt (g)

Wk 1
Wk 7
Wk 13

217.30
350.60
394.00

219.30
348.10
387.90

217.50
344.50
388.10

216.10
335.10
372.80

174.50
216.80
227.80

171.40
218.80
232.20

170.70
213.10
228.70

171.60
214.70
225.50

Body wt gain (g)

Wk
0-13


176.70


168.60


170.60


156.70


53.30


60.80


58.00


53.90

Table 7.5.1/02-4: Overview of sub-chronic toxicity study in rats treated orally (via gavage) with3-methylpentane-1,5-diol: selected haematological parameters

Parameters

M (mg/kg bw/d)

F (mg/kg bw/d)

0

100

300

1000

0

100

300

1000

Hb (g/dL)

14.744

15.300

15.544*

15.562*

14.040

14.450

14.530

14.640

WBC (109/L)

3.1967

4.2089

5.2000

4.3212

2.0260

2.2060

2.8480

2.2700

Eosinophils (%)

0.5444

0.4222

0.2222

0.9250

0.6800

0.3900

0.3800

0.2600

Lymphocytes (%)

73.744

79.911

78.789

80.112

79.120

79.800

81.730

85.410

Neutrophils (%)

23.522

17.578

19.022

16.275*

17.870

16.980

15.450

11.970

Monocytes (%)

1.800

1.678

1.444

2.238

1.940

2.400

1.990

2.050

Basophils (%)

0.0778

0.1333

0.1667*

0.1625

0.0600

0.1000

0.1200

0.1000

PT (sec)

22.644

22.600

23.080

23.189

24.480

25.300

25.680**

26.470***

APTT (sec)

12.810

13.370

13.260

14.422**

11.610

11.840

12.340

12.530

Parameters

Laboratory historical control data (2011 – 2019) Wistar rat

n

Observed range

Mean ±SD

Mean -/+2SD

n

Observed range

Mean ±SD

Mean -/+2SD

Hb (g/dL)

255

13.7 – 18.5

16.2 ±0.9

14.5 - 17.9

250

12.6 – 17.5

15.2 ±0.9

13.4 – 17.0

WBC (109/L)

256

1.5 – 13.8

4.7 ±1.5

1.6 – 7.8

250

0.3 – 6.8

2.6 ±1.2

0.3 – 5.0

Eosinophils (%)

256

0.4 – 1.9

0.6 ±0.4

-0.1 – 1.3

250

0.0 – 6.9

0.6 ±0.7

-0.7 – 2.0

Lymphocytes (%)

256

27.3 – 88.8

60.3 – 90.2

75.3 ±7.5

250

4.3 – 89.4

77.4 ±10.5

56.3 – 98.4

Neutrophils (%)

256

8.0 – 69.4

20.9 ±7.3

1.6 – 7.8

250

7.2 – 82.0

18.9 ±9.8

-0.8 – 38.6

Monocytes (%)

256

0.7 – 7.2

2.7 ±1.4

0.5 – 74.9

250

0.7 – 13.5

2.6 ±1.4

-0.3 – 5.5

Basophils (%)

256

0.0 – 0.5

0.2 ±0.1

0.0 – 0.4

250

0.0 – 0.9

0.1 ±0.1

-0.1 – 0.3

PT (sec)

200

12.5 – 26.0

21.6 ±1.8

17.9 – 25.3

188

13.9 – 39.7

22.9 ±2.3

18.3 – 27.4

APTT (sec)

201

8.4 – 30.1

14.1 ±3.2

7.7 – 20.6

189

8.7 – 25.5

13.6 ±2.8

8.0 – 19.2

*p=0.05, **p=0.01; ***p=0.001

Hb: haemoglobin

WBC: white blood cell count

APTT: activated partial thromboplastin time

 

PT: prothrombin

Table 7.5.1/02-5: Overview of sub-chronic toxicity study in rats treated orally (via gavage) with3-methylpentane-1,5-diol: selected clinical chemistry parameters

Parameters

M (mg/kg bw/d)

F (mg/kg bw/d)

0

100

300

1000

0

100

300

1000

ALT (U/L)

31.080

34.970

35.370

41.720

27.090

31.750

26.970

31.030

AST (U/L)

88.620

88.270

98.720

110.060*

84.510

80.860

78.080

78.920

ALP (U/L)

88.0270

82.3250

79.7870

79.4200

56.7970

43.6820

48.2260

46.2940

¿GT (U/L)

0.2450

0.2360

2.6957

0.3540

0.7670

0.4275

0.4611

0.4688

Creat. (µmol/L)

22.50

21.10

29.90

23.30

26.60

26.70

23.30

21.90**

K (mmol/L)

4.4820

4.2490

5.0740

7.1070***

4.0130

3.9330

4.7300

4.9610

T.bili (µmol/L)

1.860

1.970

2.110

2.250

2.050

2.080

2.490*

2.340

TBA (µmol/L)

29.5080

24.9550

32.0000

34.0210

19.1920

28.7000

25.8670

37.0730*

Parameters

Laboratory historical control data (2011 – 2019) Wistar rat

n

Observed range

Mean ±SD

Mean -/+2SD

n

Observed range

Mean ±SD

Mean -/+2SD

ALT (U/L)

214

8.1 – 107.9

35.1 ±12.3

10.6 – 59.6

201

3.5 – 84.7

28.5 ±10.3

7.9 – 49.1

AST (U/L)

115

31.3 – 159.8

88.6 ±24.4

39.6 – 137.7

201

12.6 – 211.8

77.8 ±25.3

27.1 – 128.4

ALP (U/L)

214

13.6 – 337.0

115.6 ±51.5

12.7 – 218.6

201

6.6 – 258.0

65.2 ±42.6

-20.1 – 150.4

gGT (U/L)

20

0.0 – 3.7

0.5 ±0.8

-1.1 – 2.1

23

0.0 – 2.3

0.6 ±0.5

-0.5 – 1.6

Creat. (µmol/L)

214

0.0 – 103.0

31.0 ±12.4

6.3 – 55.8

199

2.0 – 103.0

31.1 ±10.7

9.6 – 52.5

K (mmol/L)

214

1.4 – 8.7

3.9 ±0.8

-0.6 – 76.1

201

1.0 – 9.0

3.5 ±0.9

1.7 – 5.3

T.bili (µmol/L)

204

0.9 – 3.7

2.3 ±0.4

1.4 – 3.2

191

1.1 – 5.7

2.7 ±0.7

1.4 – 4.1

TBA (µmol/L)

143

2.1 – 110.0

37.8 ±0.4

-0.6 – 76.1

133

5.4 – 162.7

30.9 ±25.2

-19.5 – 81.4

*p=0.05; **p=0.01; ***p=0.001

ALT: alanine aminotransferase

AST: aspartate transaminase

ALP: alanine phosphatase

gGT: gamma glutamyl transferase

Creat.: creatinine

K: potassium

T.bili.: total bilirubin

TBA: total bile acids

Table 7.5.1/02-6: Overview of sub-chronic toxicity study in rats treated orally (via gavage) with3-methylpentane-1,5-diol: thyroid hormones

Parameters

M (mg/kg bw/d)

F (mg/kg bw/d)

0

100

300

1000

0

100

300

1000

T3 (ng/mL)
[individual animal range]

1.97
[1.59 – 2.50]

2.04
[1.52 – 2.50]

1.68
[0.76 – 2.68]

2.17
[1.66 – 2.50]

1.43
[0.73 – 2.01]

1.63
[0.89 – 2.25]

1.98*
[1.30 – 2.48]

2.04*
[1.33 – 2.91]

T4 (ng/mL)
[individual animal range]

55.56
[41.25 - 68.68]

57.92
[47.70 – 78.47]

50.61
[36.44 – 57.80]

55.87
[45.37 – 65.65]

38.58
[30.22 – 59.35]

38.35
[27.97 – 60.13]

41.28
[31.85 – 53.29]

45.63
[33.79 – 53.76]

TSH (ng/mL)
[individual animal range]

0.57
[0.28 – 0.99]

0.60
[0.14 – 1.01]

0.67
[0.22 - 1.23]

0.96
[0.53 - 1.88]

0.63
[0.24 – 1.55]

0.79
[0.26 – 2.10]

0.75
[0.24 – 1.69]

0.96
[0.30 - 1.93]

*p=0.05;

T3: triiodothyronine

T4: thyroxine

TSH: thyroid stimulating hormone

 

Table 7.5.1/02-7: Overview of sub-chronic toxicity study in rats treated orally (via gavage) with 3-methylpentane-1,5-diol: selected organ weights

Parameters

M (mg/kg bw/d)

F (mg/kg bw/d)

0

100

300

1000

0

100

300

1000

Terminal bwt (g)

380.20

373.90

370.80

358.30

222.40

225.10

223.60

218.40

Liver

Abs (g)
Rel. (g%b wt)
(g%br wt)

9.1128

2.3968
446.57

9.1358

2.4395
435.95

8.9011

2.4010
433.61

9.1312

2.5519
448.39

5..5761

2.171
284.79

5.6454

2.5110
294.70

5.8836

2.6347
298.81

6.1847*

2.8279**
320.833**

Kidney

Abs (g)
Rel. (g%b wt)
(g%br wt)

2.1523

0.5674
105.41

2.2865

0.6123
109.19

2.2975

0.6197
111.77

2.3699

0.6631***
116.21

1.5066

0.6803
76.88

1.5795

0.7027
82.39

1.6301

0.7302
82.88*

1.7181***

0.7868**
89.11***

Thyroid/parathyroid

Abs (g)
Rel. (g%b wt)
(g%br wt)

0.0334

0.0088
1.63

0.0335

0.0090
1.61

0.0312

0.0084
1.52

0.0315

0.0087
1.54

0.0245

0.0110
1.25

0.0273

0.0121
1.42

0.0263

0.0118
1.33

0.0266

0.0121
1.38

Testes

Abs (g)
Rel. (g%b wt)
(g%br wt)

3.6115

0.9546
176.98

3.5371

0.9493
169.59

3.6955

0.9985
179.79

3.7218

1.0431
182.48

-

-

-

-

Epididymides

Abs (g)
Rel. (g%b wt)
(g%br wt)

1.3267

0.3525
65.04

1.2452

0.3340
59.61

1.2710

0.3434
61.84

1.3703

0.3839
67.19

-

-

-

-

Ovaries

Abs (g)
Rel. (g%b wt)
(g%br wt)

-

-

-

-

0.1121

0.0503
5.71

0.1094

0.0487
5.71

0.1107

0.0496
5.62

0.1055

0.0484
5.48

Uterus (+cervix)

Abs (g)
Rel. (g%b wt)
(g%br wt)

-

-

-

-

0.7100

0.3236
39.39

0.7667

0.3405
40.12

1.0725

0.4849
54.30

0.6480

0.2976
33.71

Parameters (absolute weights [g])

Laboratory historical control data (2011 – 2019) Wistar rat

n

Observed range

Mean ±SD

Mean -/+2SD

n

Observed range

Mean ±SD

Mean -/+2SD

Liver

130

6.92 – 12.77

9.12 ±1.14

6.83 – 11.40

130

2.47 – 8.85

5.73 ±1.05

3.63 – 7.83

Kidney

130

1.71 – 3.12

2.33 ±0.26

1.82 – 2.84

130

0.67 – 1.94

1.47 ±0.25

0.97 – 1.96

Thyroid/ parathyroid

89

0.0098 – 0.0790

0.0257 ±0.0088

0.0080 – 0.0433

90

0.0068 – 0.0419

0.0188 ±0.0071

0.0046 – 0.0329

Testes

134

2.34 – 4.47

3.69 ±0.32

3.05 – 4.33

-

-

-

-

Epididymides

134

0.77 – 1.72

1.37 ±0.15

1.07 - 1.67

-

-

-

-

Ovaries

-

-

-

-

135

0.05 – 0.17

0.10 ±0.02

0.05 – 0.15

Uterus (+cervix)

-

-

-

-

134

0.22 – 1.82

0.82 ±0.33

0.17 – 1.48

*p=0.05; **p=0.01; ***p=0.01

Abs.: absolute


Rel (g%b wt): relative to body weight

Rel (g%br wt): relative to brain weight

Table 7.5.1/02-8: Overview of sub-chronic toxicity study in rats treated orally (via gavage) with 3-methylpentane-1,5-diol: spermatogenesis parameters

Parameters

Dose group (mg/kg bw/d)

0

100

300

1000

Mean testes weight

Testes

1.855

1.790

1.864

1.883

Tunica albuginea

0.128

0.113

0.128

0.122

Calculated wt of parenchyma

1.727

1.677

1.736

1.762

Mean sperm motility (%)

Motile count

83.1

85.8

84.2

84.5

Static count

16.9

14.2

15.8

15.5

Rapid

67.3

69.9

66.7

68.4

Testicular sperm count (106/g)

105.2

91.0

100.0

97.4

Mean sperm morphology

Total no. of normal sperm/findings

193.20

-

-

192.50

Total no. of abnormal sperm/findings

6.80

-

-

7.50

% normal

96.60

-

-

96.25

% abnormal

3.40

-

-

3.75

Table 7.5.1/02-8: Overview of sub-chronic toxicity study in rats treated orally (via gavage) with 3-methylpentane-1,5-diol: selected histopathology data

Parameters

M (mg/kg bw/d)

F (mg/kg bw/d)

0

100

300

1000

0

100

300

1000

HEART

- Cart. metaplasia
- Haem. deposits
- Mono. infiltr

3/10
0/10
0/10

-
-
-

-
-
-

2/10
0/10
0/10

2/10
1/10
1/10

-
-
-

-
-
-

1/10
0/10
1/10

TRACHEA

- Gland. ectasia
- Mixed cell infiltr

4/10
0/10

-
-

-
-

8/10
0/10

5/10
1/10

-
-

-
-

5/10
0/10

LIVER

- Haem. deposits
- Necrosis
- Fibrosis
- Mono. Infiltr
- Granuloma
- Haematopoiesis
- Vacuolation

0/10
0/10
0/10
0/10
9/10
1/10
0/10

1/1
1/1
1/1
1/1
0/1
0/1
0/1

-
-
-
-
-
-
-

0/10
0/10
0/10
0/10
7/10
0/10
0/10

0/10
0/10
0/10
0/10
8/10
0/10
0/10

-
-
-
-
-
-
-

-
-
-
-
-
-
-

0/10
0/10
0/10
0/10
5/10
0/10
0/10

SPLEEN

- Haem. deposits
- Incr. haemop.

7/10
10/10

-
-

-
-

9/10
8/10

10/10
10/10

-
-

-
-

10/10
10/10

MESENTERIC LYMPH NODES

- Haemorrhage
- Histiocytosis
- Haem. deposits
- Lymphoid depl.

6/10
1/10
0/10
1/10

-
-
-
-

-
-
-
-

3/10
0/10
0/10
2/10

2/10
0/10
0/10
1/10

-
-
-
-

-
-
-
-

7/10
0/10
1/10
0/10

KIDNEYS

- Hyal. droplets
- Mineralisation
- Mono. Infiltr.
- Pelvic dilation
- Tub. s. dilation

10/10
3/10
2/10
2/10
4/10

-
-
-
-
-

-
-
-
-
-

5/10
0/10
0/10
3/10
4/10

0/10
3/10
0/10
1/10
1/10

-
-
-
-
-

-
-
-
-
-

0/10
2/10
0/10
2/10
2/10

STOMACH

- Hyperkeratosis
- Mix. cell infiltr.
- Mono. infiltr.
- Squ. hyperlasia

2/10
1/10
1/10
2/10

-
-
-
-

-
-
-
-

0/10
1/10
1/10
0/10

3/10
1/10
0/10
3/10

-
-
-
-

-
-
-
-

1/10
0/10
0/10
1/10

LUNG

- Haem. deposits
- Alve. histiocyt.
- Mixed cell infiltr
- Mononuc. Infiltr
- Vasc. minerlisa.

0/10
5/10
1/10
2/10
3/10

1/1
0/1
0/1
0/1
0/1

-
-
-
-
-

-
-
-
-
-

1/10
4/10
1/10
0/10
4/10

-
-
-
-
-

-
-
-
-
-

0/10
8/10
1/10
0/10
9/10

PAROTID GLANDS

- Mononuc.Infiltr

0/10

-

-

0/10

1/10

-

-

0/10

THYMUS

- Haemorrhage
- Haem. deposits

0/10
0/10

2/2
0/2

3/3
0/3

0/10
0/10

2/10
2/10

1/1
0/1

1/2
0/2

5/10
4/10

EPIDIDYMIDES

- Mononuc.infiltr
- Pigmen. macrop.

1/10
1/10

-
-

-
-

2/10
2/10

-

-

-

-

AXILLARY LYMPH NODES

- Haem. deposits
- Haemorrhage
- Lymphoid depl.

0/10
1/10
2/10

-
-
-

-
-
-

0/10
0/10
1/10

1/10
1/10
3/10

-
-
-

-
-
-

0/10
0/10
2/10

ADRENAL CORTICLES

- Vacuolisation

3/10

-

-

3/10

1/10

-

-

0/10

VAGINA: OESTROUS CYCLE

- Proestrus
- Oestrus
- Diestrus

-


-

-

-

1/9
1/9
7/9

-
-
-

-
-
-

0/10
2/10
8/10

Cart. metaplasia: cartilage metaphase

Haem. deposits: Haemosiderin deposits

Mono. Infiltr.: mononuclear infiltrates

Gland. ectasia : Glandular ectasia

Mixed cell infiltr: mixed cell infiltrates

Incr. haematop.: increased haemopoiesis

Lymphoid depl.: lymphoid depletion

Hyal. droplets: hyaline droplets

Tub. s. dilation: tubular simple dilation

Mix. cell infiltr : mixed cell infiltrate
Squ. hyperplasia: Squamous hyperplasia

Alve. histiocyt. :alveolar histiocytosis

Mononuc. infiltr: Mononuclear infiltrate

Vasc. minerlisa.: vascular mineralisation

Pigmen. macrop.: Pigment macrophage

Applicant's summary and conclusion

Conclusions:
Under the conditions of this study, the NOAEL is deemed to be 1000 mg/kg bw/day for males and females based on no adverse effects when tested up to a maximum recommended concentration for repeat dose toxicity studies. 
Executive summary:

In this study, 3-methylpentane-1,5-diol was administered once via oral gavage for 90 days to Wistar rats. Animals (10/sex/group) were administered test article formulations at concentrations of 0 (water for injection), 100, 300, 1000 mg/kg bw/day. Body weight and food consumption were measured at regular intervals with neurological function examination performed. Clinical pathology evaluations (haematology, clinical chemistry (including thyroid hormones) and urinalysis) were performed with all surviving animals subjected to complete gross necropsy. All control and high dose group animals were subjected to a full histopathology.Spermatogenesis assessments were made on all male animals at necropsy.

No treatment-related effects were seen on survival, clinical signs, bloodhaematologicaland clinical chemistry parameters, urinalysis or in neurological parameters (FOBs), organ weight,histopathology or spermatogenesis.

Whilst no toxicologically relevant findings attributed to the test article were observed in either sex, a statistically significant lower mean score for response to open field grooming, handling, spontaneous activity and rearing supported in high dose group males were observed. A statistically significant lower mean body temperature was measured in the low and mid dose groups. These changes were considered unrelated to test article administration and deemed serendipitous because: i) the effects were not replicated in the high dose group (temperature); ii) there were no significant differences in any functional observations; iii) the effects were not replicated in females.

Under the conditions of this study, the NOAEL is deemed to be 1000 mg/kg bw/day for males and females based on no adverse effects when tested up to a maximum recommended concentration for repeat dose toxicity studies.