Acetone

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study protocol comparable to guideline study with minor deviations, well documented

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference 1

Reference Type:

publication

Title:

Toxicity studies of acetone administered in the drinking water of rodents

Author:

Dietz DD, Leininger JR,Rauckman EJ, Thompson MB, Chapin RE, Morissey RL, Levine BS

Year:

1991

Bibliographic source:

Fund Appl Tox 17: 347-360

Reference 2

Reference Type:

study report

Title:

Unnamed

Year:

1991

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

yes

Remarks:

(no functional observations, no clinical biochemistry determinations performed; additional examination of sperm morphology and vaginal cytology)

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Acetone

EC Number:

200-662-2

EC Name:

Acetone

Cas Number:

67-64-1

Molecular formula:

C3H6O

IUPAC Name:

propan-2-one

Details on test material:

- Substance type: pure active substance
- Analytical purity: > 99%
- Lot/batch No.: AN228
- Stability under test conditions: at least 96 h
- Storage condition of test material: ambient conditions

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Simonsen Laboratories Inc.
- Age at study initiation: 6 w
- Weight at study initiation: males ca. 100-110 g, females ca. 85-100 g
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 12 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 12.2-22.2 °C
- Humidity (%): 20-68 %
- Air changes (per hr): 12
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: drinking water

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

mean: 99.4% of target concentration, SD 3.4%

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

daily

Doses / concentrations

Remarks:

Doses / Concentrations:
2,500, 5,000, 10,000, 20,000, 50,000 ppm in drinking water corresponding to dosages of 200, 400, 900, 1,700, 3,400 mg/kg bw/d in male rats and 300, 600, 1,200, 1,600, 3,100 mg/kg bw/d in female rats (time-weighted average doses)
Basis:
actual ingested

No. of animals per sex per dose:

10

Control animals:

yes, sham-exposed

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: twice weekly

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at termination of study
- Anaesthetic used for blood collection: Yes (carbon dioxide)
- Animals fasted: No data
- How many animals: 10 per dose and sex
- Parameters examined: counts of erythrocytes, leukocytes, platelet morphology, hemoglobin, packed cell volume (hematocrit), number of reticulocytes, mean cell volume, mean corpuscular hemoglobin

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: no data
- Dose groups that were examined: no data

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, all animals

HISTOPATHOLOGY:
Yes, tissues examined in control and high dose group: adrenal glands, brain, cecum, colon, duodenum, male reproductive tissues (epididymis, seminal vesicles, prostate, testes), female reproductive tissues (ovaries, uterus), esophagus, eyes (if grossly abnormal), femur including marrow, gross lesions and tissue masses with regional lymph nodes, heart, ileum, jejunum, kidneys, liver, lungs and main-stem bronchi, mammary gland, mandibular and mesenteric lymph nodes, nasal cavity and turbinates, pancreas, parathyroid glands, pituitary gland, preputial or clitoral gland, rectum, salivary glands, skin, spinal cord, spleen, stomach, thymus, thyroid gland, trachea, urinary bladder;
additionally examined tissues in male rats: 5,000 ppm kidneys; 10,000 ppm heart, kidneys and spleen; 20,000 ppm heart, kidneys, spleen, bone marrow
additionally examined tissues in female rats: nose all dose groups; 20,000 ppm mandibular lymph nodes

Other examinations:

organ weights: liver, right kidney, right testis, heart, thymus, brain, lungs, spleen
sperm morphology investigated in the following dose groups: 0, 2500, 10,000 or 50,000 ppm
vaginal cytology investigated in the following dose groups: 0, 2500, 10,000 or 50,000 ppm

Statistics:

Non-parametric multiple comparison procedures of Dunn or Shirley: comparison of organ weight, hematologic data, male reproductive parameters
Jonkheere's test for analysis of dose response trends

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

effects observed, treatment-related

Ophthalmological findings:

no effects observed

Haematological findings:

effects observed, treatment-related

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

not specified

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Details on results:

BODY WEIGHT GAIN (0, 2,500, 5,000, 10,000, 20,000, 50,000 ppm):
m 229, 242, 242, 239, 227, 166 g;
f 111, 106, 104, 105, 105, 96 g;
depressed weight gain in males and in females at 50,000 pm corresponding to a depression of 27.5 % and 13.5 % compared to controls

WATER CONSUMPTION (0, 2,500, 5,000, 10,000, 20,000, 50,000 ppm):
m 87, 79, 78, 80, 83, 65 ml/kg bw/d
f 107, 110, 121, 112, 75, 58 ml/kg bw/d
depressed water consumption in males at 50,000 ppm, in females at >= 20,000 ppm
Evaluation: no signs of dehydration, indicating a lack of palatability; no toxicologically significant effect

HAEMATOLOGY: (see Table 1 for changes of individual parameters, significance levels and corresponding LOELS)
m: 5,000 ppm and higher : depressed reticulocyte counts, minimally depressed Hb (no dose-dependence)
10,000 ppm and higher: reactive anaemia indicated by marginal but significant increases in MCV and MCH, no effect on MCHC
at 20,000 and 50,000 ppm: mild but significant leukocytosis produced by absolute increase in lymphocytes; mild but significant depression of erythrocyte count and platelet count
50,000 ppm
Evaluation: Overall, changes of haematological parameters were small and partly without dose-relationship. Haematological changes are consistent with a mild macrocytic normochromic anemia with a depressed regenerative response (depression of reticulocytes) with an overall LOAEL of 20,000 ppm for mild leukocytosis and depression of erythrocytes; finding consistent with hemosiderosis in spleen

f: at 20,000 and 50,000 ppm: mild but significant depression of erythrocyte count and platelet count
50,000 ppm: mild but significant leukocytosis produced by absolute increase in lymphocytes; mild but significant increase of MCH and MCV
Evaluation: no indication of anaemia

ORGAN WEIGHTS:
m: at 20,000 ppm increase of relative liver weight (p<0.01), at 50,000 ppm increases of relative weights of liver, kidney, lung, testis (p<0.01) together with significant depression of terminal body weight by 19 %; absolute weights of liver, kidney and lung were also increased, though not significantly;Evaluation: kidney weight changes associated with nephropathy (see Table 2);
liver weight changes were not associated with acetone-mediated histopathologic effects, but may be associated with the ability of acetone to induce hepatic microsomal cytochrome P450;
relative testis weight was increased significantly by 20 % whereas the absolute testis weight was decreased by 5 %, this effect may be biologically not significant as testicular toxicants typically decrease testis weight; however, a mild toxic effect on spermatogenesis was indicated by the evaluation of sperm parameters (see below)

f: at 20,000 ppm increase of relative liver and kidney weight (p<0.01), at 50,000 ppm increases of relative weights of liver (p<0.05>), of absolute and relative kidney weight and of relative lung weight (p<0.01) together with depression of terminal body weights by 7 % at both doses
Evaluation: the toxicological relevance of these increased organ weights is questionable as effects were associated with significant decreases of terminal body weights in the absence of any correlating histopathological changes

HISTOPATHOLOGY:
m: kidney: incidences and severity of nephropathy increased with increasing dose levels, an apparent level of nephropathy was found at 20,000 ppm and 50,000 ppm with 90 % of the rats showing mild nephropathy in contrast to 10 % in the control group (for details see Table 3).
The histological picture of this nephropathy was considered to be the same as the chronic progressive nephropathy of aging rats characterized by foci of regenerating cortical tubules located in or adjacent to areas of interstitial fibrosis with and without inflammatory cell infiltration. Besides, a thickening of tubule basement membranes, of the glomerular mesangium and glomerular capillary basement membranes.
Evaluation: These results suggest that acetone accelerated the occurrence and severity of this spontaneously occurring progressive condition of aging rats. As a possible mechanism, the ability of acetone to depress protein catabolism was discussed.
m: spleen: pigmentation of the splenic red pulp with hemosiderin existed at 20,000 and 50,000 ppm in all males (see Table 3)
Evaluation: indicative of accelerated erythrocyte turnover, thus being consistent with the haematological findings

f: no histological changes reported

REPRODUCTIVE ENDPOINTS:
m: significant depression of sperm motility, cauda epidydimal weight, and epididymal weight, significantly increased incidence of abnormal sperm with a LOAEL of 50,000 ppm (for details see Table 4)
Evaluation: effects are consistent with a mild toxic effect on spermatogenesis, although no effects are visible microscopically

f: no changes of vaginal cytology

Effect levels

1

Dose descriptor:

NOAEL

Effect level:

50 000 ppm

Sex:

female

Basis for effect level:

other: dose: 3,100 mg/kg bw/d

2

Dose descriptor:

LOAEL

Effect level:

20 000 ppm

Sex:

male

Basis for effect level:

other: histopathology (kidney, spleen); hematopoetic system (mild macrocytic normochromic anemia); dose: 1,700 mg/kg bw/d

3

Dose descriptor:

NOAEL

Effect level:

10 000 ppm

Sex:

male

Basis for effect level:

other: dose: 900 mg/kg bw/d

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Table 1: Overview of haematological parameters in male rats

Dose group	Control	2500 ppm	5000 ppm	10,000 ppm	20,000 ppm	50,000 ppm	Remarks
Leukocytes (10^3/µl)	5.5 +- 0.22	6.5 +- 0.51	6.2 +- 0.36	6.4 +- 0.47	6.9 +- 0.40 ^a	7.3 +- 0.38 ^b	LOEL 20,000 ppm ^c
Lymphocytes (10^3/µl)	4.4 +- 0.18	5.0 +- 0.43	4.7 +- 0.43	5.1 +- 0.33	5.4 +- 0.33 ^a	5.9 +- 0.33 ^b	LOEL 20,000 ppm
Hematocrit (%)	43.6 +- 0.57	43.4 +- 0.38	42.1 +- 0.32 ^a	44.0 +- 0.61	42.2 +- 0.49^a	42.7 +- 0.36	no dose-relationship
Hemoglobin (g/dl)	16.1 +- 0.21	16.3 +- 0.19	15.6 +- 0.13 ^a	16.1 +- 0.09	15.6 +- 0.21 ^a	15.6 +- 0.15 ^a	no dose- relationship
Mean corpuscular hemoglobin (pg)	17.3 +- 0.11	17.7 +- 0.13 ^a	17.3 +- 0.10	17.8 +- 0.14 ^a	17.7 +- 0.26^b	18.6 +- 0.15 ^b	LOEL 10,000 ppm ^d
Mean cell volume (fL)	46.8 +- 0.20	47.6 +- 0.16  ^a	47.0 +- 0.21	48.0 +- 0.21  ^b	49.0 +- 0.42 ^b	50.8 +- 0.25  ^b	LOEL 10,000 ppm ^d
Erythrocytes (10^6/ml)	9.3 +- 0.14	9.1 +- 0.09	9.0 +- 0.06 ^a	9.1 +- 0.05	8.6 +- 0.06 ^b	8.4 +- 0.08  ^b	LOEL 20,000 ppm ^d
Reticulocytes (10^3/µl)	225 +- 11.0	195 +- 14.7	171 +- 14.9  ^b	179 +- 15.0 ^a	168 +- 11.0  ^b	152 +- 9.3  ^b	LOEL 5,000 ppm
Platelets (10^3/µl)	651 +- 10.7	641 +- 13.8	654 +- 13.2	617 +- 12.2	605 +- 20.3 ^a	501 +- 23.6  ^b	LOEL 50,000 ppm

significance levels: ^a p<0.05; ^b p< 0.01

Remarks: ^c mild leukocytosis due to increase of lymphocytes; no treatment-related changes in eosinophils, monocytes, or segmented neutrophils

^d indicative of reactive anemia

Table 2: Overview of significant changes of haematological parameters in female rats

Dose group	Control	2500 pmm	5000 ppm	10,000 ppm	20,000 ppm	50,000 ppm	Remark
Leukocytes (10^3/µl)	4.7 +- 0.19	4.5 +- 0.38	5.0 +- 0.30	4.5 +- 0.30	5.2 +- 0.68	5.9 +- 0.26 ^a	LOEL 20,000 ppm ^c
Lymphocytes (10^3/µl)	3.6 +- 0.21	3.5 +- 0.36	4.0 +- 0.22	3.6 +- 0.25	3.9 +- 0.42	4.8 +- 0.20^b	LOEL 50,000 ppm
Mean corpuscular hemoglobin (pg) Mean corpuscular hemoglobin (pg)	18.3 +- 0.29	18.8 +- 0.13	18.7 +- 0.09	18.6 +- 0.08	18.7 +- 0.08	19.0 +- 0.08 ^b	LOEL 50,000 ppm
Mean cell volume (fL)	50.0 +- 0.15	50.0 +- 0.21	50.2 +- 0.13	50.3 +- 0.15	50.4 +- 0.16	51.1 +- 0.18 ^b	LOEL 50,000 ppm
Platelets (10^3/µl)	684 +- 13.0	655 +- 17.4	662 +- 20.0	658 +- 15.5	654 +- 57.2 ^a	528 +- 17.0 ^b	LOEL 20,000 ppm

significance levels: ^a p<0.05; ^b p<0.01

Remarks: ^c mild leukocytosis due to increase of lymphocytes; no treatment-related changes in eosinophils, monocytes, or segmented neutrophils

Table 3: Histopathological changes in male rats

Site / lesion	0 ppm	2,500 ppm	5,000 ppm	10,000 ppm	20,000 ppm	50,000 ppm
Spleen: pigmentation	0/10	0/10	0/10	0/10	10/10	10/10
Kidney: total nephropathy	6/10	8/10	8/10	9/10	10/10	10/10
minimal	5/10	8/10	8/10	9/10	1/10	1/10
mild	1/10	0/10	0/10	0/10	9/10	9/10

Table 4: Reproductive endpoints in male rats

Parameter	0 ppm	2,500 ppm	10,000 ppm	50,000 ppm
Caudal epididymal weight (g)	0.14 +- 0.006	0.14 +- 0.006	0.15 +- 0.005	0.10 +- 0.005^a
Right epididymal weight (g)	0.44 +- 0.009	0.44 +- 0.009	0.45 +- 0.008	0.35 +- 0.008 ^a
Abnormal sperm (%)	0.68 +- 0.061	0.98 +- 0.156	0.92 +- 0.131	3.42 +- 0.532 ^a
Sperm motility (%)	75.7 +- 1.15	71.6 +- 1.44 ^b	72.9 +- 0.80	66.8 +- 2.69 ^a
Sperm density (10^6/g cauda)	571 +- 61.1	564 +- 19.7	570 +- 19.4	424 +- 69.5

significance levels: ^a p<0.05; ^b p<0.01

Applicant's summary and conclusion

Conclusions:

Acetone was found to be mildly toxic to rats when administered in the drinking water for 13 weeks. The minimal toxic dose (LOAEL) was 1,700 mg/kg bw/d for male rats with the testis, kidneys, and hematopoetic system as target organs.

Executive summary:

In a 13-week drinking water study, groups of 10 male and 10 female Fischer 344 rats were exposed to acetone at concentrations of 2,500, 5,000, 10,000, 20,000, 50,000 ppm. The test protocol was similar to OECD Guideline 408, without investigation of biochemical parameters and without functional observations, and additional investigation of sperm parameters and vaginal cytology. All rats survived to the end of the study without clinical signs. At 50,000 ppm final mean body weight gains of males and females were 27.5 % and 13.5 % lower than controls. Reduced water consumption in 50,000 ppm males and 20,000 and 50,000 ppm females resulted in effective dosages of 200, 400, 900, 1,700, 3,400 mg/kg bw/d in male rats and of 300, 600, 1,200, 1,600, 3,100 mg/kg bw/d in female rats.

Increased relative organ weights of liver and kidney at 20,000 ppm, and of lung and testis at 50,000 ppm were associated with decreases of terminal body weights. Correlating histopathological changes were only found in the kidneys of male rats showing increased incidence and severity of nephropathy at >= 20,000 ppm considered to be the most prominent chemically related finding. Increased liver weights may be associated with the ability of acetone to induce hepatic microsomal cytochrome P450. At 50,000 ppm a mild toxic effect on spermatogenesis was indicated by significant depressions of sperm motility, cauda epidydimal weight, and epididymal weight and a significantly increased incidence of abnormal sperm together with a 5 % decrease of absolute testis weight. Overall, changes of haematological parameters were small and partly without dose-relationship. Only in male rats, haematological changes indicated a mild macrocytic normochromic anemia with a depressed regenerative response (depression of reticulocytes) with an overall LOAEL of 20,000 ppm for mild leukocytosis and depression of erythrocytes. This finding was consistent with hemosiderosis in spleen.

For male rats the minimal toxic concentration (LOAEL) was 20,000 ppm corresponding to a dose of 1,700 mg/kg bw/d, and the NOAEL was 10,000 ppm or 900 mg/kg bw/d with the testis, kidneys, and hematopoetic system as target organs. No relevant toxicological findings were observed in female rats up to the highest dose leading to a NOAEL of 50,000 ppm or 3,100 mg/kg bw/d.