Butane-1,2-diol

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:

TBC

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

The study was conducted to international guideline with some deviations i.e. only 45 days exposure instead of 63 days, no clarity on measurement of estrous cycle or sperm analysis and no T4/T3 measurements were taken.e

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1992

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Version / remarks:

Not specified

Deviations:

yes

Remarks:

The study was conducted to international guideline with some deviations i.e. only 45 days exposure instead of 63 days, no clarity on measurement of estrous cycle or sperm analysis and no T4/T3 measurements were taken.e

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Butane-1,2-diol

EC Number:

209-527-2

EC Name:

Butane-1,2-diol

Cas Number:

584-03-2

Molecular formula:

C4H10O2

IUPAC Name:

butane-1,2-diol

Test animals

Species:

rat

Strain:

Crj: CD(SD)

Details on species / strain selection:

Not stated

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories Japan, Inc.,
- Age at study initiation: Approximately 8 weeks
- Fasting period before study: no
- Housing: Animals were housed in polycarbonate cages (265 W × 426 D × 200 H mm, Tokiwa Kagaku Kikai Co., Ltd.) in which bedding for experimental animals (Betachip, Charles River Laboratories Japan, Inc.) was laid: 2 animals/sex/cage during quarantine and acclimatization period, 1 animal/cage during the premating treatment period, 1 male and 1 female/cage during the mating period, 1 litter/cage during the lactation period. The animals were reared on steel racks (four-tiered).
- Diet (e.g. ad libitum): Autoclaved solid feed for experimental animals (CRF-1, Oriental Yeast Co., Ltd.) was fed ad libitum. Feed was changed at a frequency of once per week. Feed for which the analytical values of residual agricultural chemicals and contaminants were confirmed to be the internal SOP-defined concentrations or less was used.
- Water (e.g. ad libitum): All animals had free access to tap water. Tap water for drinking was filtrated through a 5 μm filter and irradiated by UV light. Water for drinking was changed at a frequency of once per week. Water quality tests in compliance with the Water Supply Law were conducted regularly and the analysis results were confirmed to be within the acceptance criteria as stipulated by the annex of the Ordinance of the Ministry of Health and Welfare No. 56.
- Acclimation period: At least 6 days prior to start of treatment.
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20°C to 25°C.
- Humidity (%): 40 - 70%
- Air changes (per hr): 12 times/hour
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: Animal arrival August 28, 1991 but date of necropsy was not documented

Administration / exposure

Route of administration:

oral: gavage

Details on route of administration:

Oral gavaged using a syringe attached to a stomach tube.

Vehicle:

water

Remarks:

Purified water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
The test article was dissolved in purified water and dosing solutions at the nominal concentrations were prepared. The preparation was conducted once weekly and the dosing solutions were kept refrigerated and protected from light until immediately before dosing.

For stability of the test article in the dosing solution, the dosing solutions for the low dose group and high dose group were analyzed before the commencement of dosing in a laboratory of Mitsubishi Kasei Institute of Toxicological and Environmental Sciences, Co., Ltd., and the test article was confirmed to be stable for 14 days under the storage condition.

DIET PREPARATION: N/A 
 - Rate of preparation of diet (frequency):
 - Mixing appropriate amounts with (Type of food): 
 - Storage temperature of food:
 
 VEHICLE
 - Justification for use and choice of vehicle (if other than water): N/A
- Concentration in vehicle: The concentrations measured for all dose levels were within ±2% of the nominal concentrations.
- Amount of vehicle (if gavage): The concentrations measured for all dose levels were within ±2% of the nominal concentrations.
- Lot/batch no. (if required): Not stated 

- Purity: Not stated.

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

42 days for male
40 - 45 day for female

Frequency of treatment:

Daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

40 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

200 mg/kg bw/day (nominal)

Doses / concentrations 4

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

Method: Animals were gavaged using a syringe attached to a stomach tube. Dose volume was 5 mL/kg, and individual volumes were calculated based on the bodyweights measured at the most recent day.
Frequency: Once daily
-F0 males:42 consecutive days
-F0 females: 40 – 45 consecutive days.
-F1 animals: Potential indirect exposure in utero and through the milk during lactation
Dose levels: The amount of test item to be administered was calculated according to the most recent body weight recorded.
Group 1: 0 mg/kg/day
Group 2: 40 mg/kg/day
Group 3: 200 mg/kg/day
Group 4: 1000 mg/kg/day
Administration volume 5 mL/kg
Dosing was administered once daily in the morning to males 14 days prior to mating and during the mating period and thereafter until the day before the planned sacrifice for a total of 42 days and to females from 14 days prior to mating and during the period covering completion of mating and delivery until Lactation Day 3 for a total of 40 to 45 days. For notations of days after the commencement of dosing, the day of commencement of dosing was defined as 0 days after the commencement of treatment, the day when mating was confirmed as Gestation Day 0, and the day after delivery as Lactation Day 1.

Positive control:

No

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Not specified

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily

BODY WEIGHT: Yes
or males, body weights were measured on the day of the commencement of dosing and thereafter once per week. For females, bodyweights were measured on the day of the commencement of dosing and thereafter once per week before mating and after the completion of mating on gestation days 0, 7, 14, and 20, and lactation days 1 and 4.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
For males, food consumption was measured once per week from the commencement of dosing, except for the mating period. For females, gross weights of food were measured once per week before mating, and after the completion of mating, on gestation days 0, 7, 14, and 20, and Lactation Days 1 and 4.

FOOD EFFICIENCY: No

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: 21 hours after completion of treatment
- Anaesthetic used for blood collection: Not specified
- How many animals: All surviving males.
- Parameters checked in table: Yes

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: 21 hours after completion of treatment
- Animals fasted: Yes, but details not disclosed.
- How many animals: All surviving males.
- Parameters checked in table: Yes

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No

OTHER: Yes (see below);

ESTROUS CYCLE: Yes
- Dry smears - not specified.
- Wet smears - not specified.

THYROID HORMONE ANALYSIS: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

Statistics:

Bartlett's test for homogeneity of variance, when the variances were heterogeneous, the Kruskal-Wallis test was performed. Dunnett’s method in case of an equal number of animals between the groups or using Scheffe’s method in case of a different number of animals between the groups. For count data, the data were tested by Fisher's exact probability test.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

In females of the 1000 mg/kg group, post-treatment decreases in locomotor activity were observed in more than half of the animals and hypopnea in a few animals. Both signs were transient in nature. The number of affected animals decreased over time, and on day 6 or later, the sign occurred only in one animal until gestation day 5 (at 21 days after the commencement of treatment). No abnormalities occurred in males. In addition, hair loss was observed in one female of the 1000 mg/kg group but the finding is considered a spontaneous finding.

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):

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

For glucose in the 40 mg/kg group and total bilirubin in the 200 mg/kg group, statistical differences were noted. However, no related changes suggestive of the effects of the test article were noted.

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

Bilateral atrophy of the testes was found in one male of the 1000 mg/kg group (animal number: DM09). In the other animals, no abnormalities were found in either males or females of any groups

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

In both the control group and 1000 mg/kg group, focal myocardial degeneration in the heart, microgranuloma in the liver, fatty change of hepatocytes, and brown pigmentation in tubular epithelial cells, cyst, focal basophilic change in renal tubules and fatty change of tubular epithelia in the kidneys were sporadically found. No between-group differences in the incidences of these changes were noted; therefore, these changes were not indicative of the treatment effects. In addition, in females, slight increases of hematopoietic cells in the spleen and diffuse chronic hypertrophy in the fascicular zone of the adrenals were found in almost all animals in the control and 1000 mg/kg groups without any between-group differences.

The atrophy of the testes of the male in the 1000 mg/kg found in necropsy was reflected by extensive atrophy of seminiferous tubules found in histopathology. The change was associated with slight bilateral atrophy of the epididymides. In the testes, degeneration, vacuolation, and multinucleated cell formation were observed in the sperm cells, but no abnormalities were observed in Sertoli cells or interstitial cells. In the other animals, including those of the 40 and 200 mg/kg groups, no abnormalities were found in the testes or epididymides. The other changes included germinal center deficiency in the spleen and unilateral extensive hemorrhagic necrosis in the adrenal cortex in one female of the 1000 mg/kg group (D07).

Histopathological findings: neoplastic:

no effects observed

Other effects:

not specified

Effect levels

Key result

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

Key result

Critical effects observed:

no

Any other information on results incl. tables

Table 2. Total incidence of clinical signs in male

Dose (mg/kg)	No. of animals	Findings	Days after commencement of treatment
			0	2	4	6	8	10	12	14	16	18	20	22	24	26	28	30	32	34	36	38	41
0	10	RSM (+) Hypopnoea	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -
40	10	RSM (+) Hypopnoea	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -
200	10	RSM (+) Hypopnoea	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -
1000	10	RSM (+) Hypopnoea	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -

RSM: Decrease locomotor activity, -: No signs were observed in any animals, +: Slight

Table 3. Total Incidence of Clinical Signs in Female

Dose (mg/kg)	No. of animals	Findings	Days after commencement of treatment								Days of gestation											Days of lactation
			0	2	4	6	8	10	12	14	0	2	4	6	8	10	12	14	16	18	20	0	1	2	3
0	10	RSM (+) Hypopnoea	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -
40	10	RSM (+) Hypopnoea	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -
200	10	RSM (+) Hypopnoea	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -
1000	10	RSM (+) Hypopnoea	6 -	3 1	2 2	2 1	2 -	2 -	1 1	1 1	1 1	1 1	1 1	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -	- -

RSM: Decrease locomotor activity, -: No signs were observed in any animals, +: Slight, 1) two of the animals did not copulate

Table 4. male group mean body weight values and mean body weight gain.

Dose (mg/kg)	Group mean body weight							Mean body weight gain
	Days after commencement of treatment
	0	7	14	21	28	35	41	7	14	21	28	35	41
0	313	360	400	419	463	470	483	47	87	106	130	157	170
40	315	363	401	420	442	465	480	48	87	105	128	150	165
200	314	358	395	416	438	462	479	44	81	102	123	148	164
1000	313	358	397	415	441	463	475	45	84	102	128	150	163

Table 5. Female group mean body weight values and mean body weight gain.

Dose (mg/kg	Group mean body weight									Mean body weight gain
	Days after commencement of treatment			Gestation days				Lactation days		Days after commencement of treatment		Gestation days			Lactation days
	0	7	14	0	7	14	20	1	4	7	14	7	14	20	4
0	203	214	229	240	280	321	396	300	317	11	26	40	81	157	17
40	201	214	226	241	279	316	385	297	315	13	25	38	75	144	18
200	204	218	231	241	280	316	389	296	313	14	27	39	76	149	17
1000	201	210	223	233	269	307	382	290	308	10	22	36	74	149	18

Table 6. Male group mean food consumption values.

Dose (mg/kg	Days after commencement of treatment
	7	14	28	35	41
0	28.1	28.1	27.1	28.5	28.1
40	28.8	28.3	26.4	28.0	27.7
200	27.8	27.8	27.1	27.7	27.5
1000	27.8	27.4	26.6	27.4	26.4

Table 7. Female group mean food consumption values.

Dose (mg/kg	Days after commencement of treatment		Gestation days			Lactation days
	7	14	7	14	20	4
0	17.9	18.4	23.9	26.6	27.1	32.8
40	18.1	18.8	23.5	25.8	25.8	32.4
200	18.2	18.8	24.3	26.2	26.5	32.1
1000	16.7	17.7	22.1	24.9	26.3	31.6

Table 8. Male mean haematology values.

Dose (mg/kg	RBC Count (x10%L)	HT (%)	HB Conc (G/DL)	Reticulocyte Count (%)	MVC (m)	MCH (PG)	MCHC (%)	Platelet Count (x10%L)	WBC Count (x10%L)	Differential count of leukocytes (% of total counted cells)
										Lymphocytes	Neutrophils		Eosinophils	Basophile	Monocytes
											Segmented	Band
0	1001	47.0	15.4	17	46.9	15.4	32.7	74.1	120	93	3	0	1	0	4
40	993	46.6	15.4	12	46.9	15.6	33.0	76.7	107	94	2	0	1	0	3
200	968	45.7	15.0	21	47.2	15.5	32.9	75.1	109	88	5	0	1	0	5
1000	996	47.2	15.5	18	47.4	15.6	32.9	73.3	86	92	4	0	1	0	3

Table 9. Male group mean clinical chemistry values.

Dose (mg/kg	GOT (IU/L)	GPT (IU/L)	GTP (IU/L)	ALP (IU/L)	Total Bilirubin (MG/DL)	Urea Nitrogen(MG/DL)	Creatinin (MG/DL)	Glucose (MG/DL)	Total Chol. (MG/DL)	Triglyceride (MG/DL)	Total Protein (G/DL)	Albumin (G/DL)	A/G Ratio	Calcium (MG/DL)	Inorganic Phos. (MG/DL)	NA (MEQ/L)	K (MEQ/L)	CL (MEQ/L)
0	88	30	0	245	0.11	18.1	0.6	129	60	67	6.76	3.97	1.43	9.2	7.8	143	4.4	103
40	83	28	0	264	0.06	19.5	0.5	149**	68	67	6.77	3.99	1.44	9.1	7.6	143	4.5	103
200	76	26	0	232	0.05*	17.3	0.6	135	64	50	6.63	3.92	1.45	9.0	7.5	143	4.3	103
1000	78	24	0	223	0.10	18.4	0.6	135	54	52	6.66	3.95	1.46	9.0	7.7	143	4.3	103

*: Significantly different from control value, * P<0.05; ** P<0.01

Table 10. Group mean absolute and relative organ weight values.

Dose (mg/kg	Male (group mean absolute organ weight)						Male (group mean relative organ weights)					Female (group mean absolute organ weight)				Female (group mean relative organ weight)
	Final Body weight (g)	Thymus (mg)	Liver (g)	Kidneys (g)	Testes (g)	Epididymides (g)	Thymus (x10^-3)	Liver	Kidneys	Testes	Epididymides	Final Body weight (g)	Thymus (mg)	Liver (g)	Kidneys (g)	Thymus (x10^-3)	Liver	Kidneys
0	461	359	12.57	3.07	3.24	1.21	78	2.72	0.67	0.71	0.26	317	256	12.79	1.98	81	4.04	0.63
40	456	375	12.65	3.04	3.37	1.28	82	2.77	0.67	0.74	0.28	315	279	12.61	2.03	88	4.01	0.65
200	455	327	12.19	2.89	3.18	1.18	72	2.68	0.64	0.70	0.26	313	231	12.48	1.88	73	4.00	0.60
1000	451	366	12.41	2.97	3.14	1.18	80	2.75	0.66	0.70	0.26	308	244	12.91	1.98	79	4.20	0.65

Applicant's summary and conclusion

Conclusions:

Daily repeat oral gavage doses of 1,2-butanediol at 40, 200, and 1000 mg/kg were administered to rats (males) from 14 days prior to mating and during the mating period and thereafter for a total of 42 days and to females from 14 days prior to mating and during the period covering mating, gestation, and delivery until lactation day 3 did not result in adverse effect. Therefore, the no observed effect levels (NOEL) under the experimental conditions used were considered 200 mg/kg for repeat dose toxicity and 1000 mg/kg for reproductive and developmental toxicity while the no observed adverse effect levels of 1000 mg/kg for repeat dose toxicity and > 1000 mg/kg for reproductive and developmental toxicity.

Executive summary:

OECD 422 - Daily repeat oral gavage doses of 1,2-butanediol at 40, 200, and 1000 mg/kg were administered to SD rats. The test article was orally administered once daily to males from 14 days prior to mating and during the mating period and thereafter for a total of 42 days and to females from 14 days prior to mating and during the period covering mating, gestation, and delivery until lactation day 3. Toxicological effects on parent animals and effects on reproductive performance, such as gonadal function, mating behaviour, fertility, delivery, and lactation were investigated.

No mortality was observed, body weight, food consumption, haematology and organ weight were unaffected throughout the study.  For glucose in the 40 mg/kg group and total bilirubin in the 200 mg/kg group, statistical differences were noted. However, no related changes suggestive of the effects of the test article were noted.

In females of the 1000 mg/kg group, post-treatment decreases in locomotor activity were observed in more than half of the animals and hypopnea in a few animals. Both signs were transient in nature. The number of affected animals decreased over time, and on day 6 or later, the sign occurred only in one animal until gestation day 5 (at 21 days after the commencement of treatment). No abnormalities occurred in males. In addition, hair loss was observed in one female of the 1000 mg/kg group, but the finding is considered a spontaneous finding.

Bilateral atrophy of the testes was found in one male of the 1000 mg/kg group (animal number: DM09), this was reflected by extensive atrophy of seminiferous tubules found in histopathology and associated with slight bilateral atrophy of the epididymides. In the testes, degeneration, vacuolation, and multinucleate cell formation were observed in the sperm cells, but no abnormalities were observed in Sertoli cells or interstitial cells. In the other animals, including those of the 40 and 200 mg/kg groups, no abnormalities were found in the testes or epididymides. The other changes included germinal centre deficiency in the spleen and unilateral extensive hemorrhagic necrosis in the adrenal cortex in one female of the 1000 mg/kg group (D07).

In both the control group and 1000 mg/kg group, focal myocardial degeneration in the heart, microgranuloma in the liver, fatty change of hepatocytes, and brown pigmentation in tubular epithelial cells, cyst, focal basophilic change in renal tubules and fatty change of tubular epithelia in the kidneys were sporadically found. No between-group differences in the incidences of these changes were noted; therefore, these changes were not indicative of the treatment effects. In addition, in females, slight increases of hematopoietic cells in the spleen and diffuse chronic hypertrophy in the fascicular zone of the adrenals were found in almost all animals in the control and 1000 mg/kg groups without any between-group differences.

Reproductive performance for both males and females, no abnormality was noted in copulation index or fertility index, and no effect of the test article was noted in gestation length, delivery, or nursing behaviour. Furthermore, the examination on corpora lutea count, implantation site count, implantation index, gestation index, delivery index, offspring count, and viable offspring count revealed no changes suggestive of effects of the test article on ovulation, implantation, or subsequent embryonic development. In addition, no abnormality was noted in the external surface of the offspring, viability index, bodyweight at birth, or bodyweight gain after birth. In necropsy of neonates, thymic remnant in the neck and left umbilical artery were sporadically found. However, these findings were those often spontaneously observed at the end of the fetal stage, and the incidences were not correlated with dose levels. In one female of the 1000 mg/kg group, (a) white spot(s) was(were) found, which was (were) hepatic necrosis and fatty change. These changes are known to be induced by various factors, and it is widely recognised that these changes are also induced by circulatory impairment and metabolic disorder including nutritional disorder. In this study, however, the changes occurred in only one animal and were not found in the other neonates; therefore, they were not considered treatment related.

Therefore, the no observed effect levels (NOEL) under the experimental conditions used were considered 200 mg/kg for repeat dose toxicity and 1000 mg/kg for reproductive and developmental toxicity while the no observed adverse effect levels of 1000 mg/kg for repeat dose toxicity and > 1000 mg/kg for reproductive and developmental toxicity.