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Diss Factsheets

Administrative data

Description of key information

Oral toxicity:
Subchronic neurotoxicity study, Sprague-Dawley rat, 13 w gavage (5 d/w) at doses of 0, 20, 64 and 200 mg/kg bw/d: NOAEL 20 mg/kg bw/d, LOAEL 64 mg/kg bw/d (observation of tremors as clinical signs in male and female rats) (Topping, 1988, 2007).
Combined carcinogenicity and chronic toxicity study, Fischer 344 rat, males, 103 w gavage (5 d/w) at doses of 0, 25 and 50 mg/kg bw/d: NOAEL 25 mg/kg bw/d, LOAEL 50 mg/kg bw/d (decreased body weight, increased severity of chronic progressive nephropathy, significant increase of renal tubuli adenoma; renal findings as sex- and species-specific effect of no toxicological relevance for human risk assessment) (Kari et al., 1992; NTP, 1989).
Dermal toxicity
14 day range finding dermal toxicity study, F344 rats, 12 open applications on 5 d/w of 240 - 3840 mg/kg bw/application: NOAEL 3840 mg/kg bw (Kari et al., 1992; NTP, 1989). No effects were observed in a 90-day dermal study in rats, but HQ was applied at only 0.5% in a cream formulation (i.e. eq. to 74 mg/kg/day), which limited the use of a NOAEL as point of departure for risk assessment.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Referenceopen allclose all

Endpoint:
chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from August 1982 to August 1984
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Study comparable to guideline study with acceptable restrictions: only 2 instead of 3 dose groups, no data on food consumption; hematology and clinical chemistry examinations only at 15 mo, no urinalysis data
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
reference to other study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)
Deviations:
yes
Remarks:
(only 2 instead of 3 dose groups, no data on food consumption; hematology and clinical chemistry examinations only at 15 mo, no urinalysis data)
GLP compliance:
not specified
Limit test:
no
Species:
mouse
Strain:
B6C3F1
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Kingston, NY, USA
- Age at study initiation: males 8-9 w , females 9-10 w
- Fasting period before study: no
- Housing: 5 per cage
- Diet: NIH 07 rat ration (Zeigler Bros. Inc., Gardner, PA, USA) ad libitum
- Water: tap water ad libitum
- Acclimation period: males 18 d, females 26 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 27 °C (65 - 80 °F)
- Humidity (%): 40 - 79
- Air changes (per hr): 6 - 13 room air changes per hr
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: appropriate amount of hydroquinone dissolved in deionised water by stirring with a magnetic stir bar; maximum storage time 21 d at room temperature in the dark in amber serum vials with Teflon-lined seals, sparged with argon or nitrogen before sealing

VEHICLE
- Concentration in vehicle: 5, 10 mg/mL
- Amount of vehicle (if gavage): 10 mL/kg
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
UV spectroscopy of acetonitrile extracts at 295 nm or methanol extracts at 293 nm; analyzed concentrations (data from 10 samples each): mean 4.88, range 4.72 - 5.09 mg/mL; mean 9.78, range 9.39 – 10.25 mg/mL; variation considered to be in acceptable range; stability investigations by HPLC after 21 days of storage of the dosing solution at room temperature in the dark indicated no notable decrease of hydroquinone concentration
Duration of treatment / exposure:
65 or 103 w , sacrifice after ca. 65 and 104 to 105 w
Frequency of treatment:
5 d/w
Remarks:
Doses / Concentrations:
0, 50, 100 mg/kg bw/d
Basis:
actual ingested
No. of animals per sex per dose:
64 or 65 males, 65 females (including 10 for 65 w treatment)
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: based on 13 week range-finding toxicity study (see separate endpoint study record)
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: 2 times per day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: 2 times per day

BODY WEIGHT: Yes
- Time schedule for examinations: at initiation of dosing, weekly thereafter for 13 weeks, and monthly for the rest of the study period

FOOD CONSUMPTION: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes, at 15 mo
Investigated parameters: leukocytes, lymphocytes, segmented neutrophiles, monocytes, eosinophils, atypical lymphocytes, atypical mononuclear cells, bands, hematocrit, hemoglobin, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, mean cell volume, erythrocytes, reticulocytes

CLINICAL CHEMISTRY: Yes, at 15 mo
Investigated parameters: albumin, alkaline phosphatase, alanine aminotransferase, blood urea nitrogen, creatinine, sorbitol dehydrogenase, total protein

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes, all animals

ORGAN WEIGHTS: Yes, at 15 mo and 2 y

HISTOPATHOLOGY: Yes
Tissues examined in high-dose mice and controls: adrenal glands, brain, caecum, colon, duodenum, epididymis/prostrate/testes or ovaries/uterus, esophagus, gallbladder, gross lesions and tissue masses, heart, ileum, jejunum, kidneys, liver, lungs and mainstem bronchi, mammary gland, mandibular or mesenteric lymph nodes, nasal cavity and turbinates, pancreas, parathyroid glands, pituitary gland, rectum, salivary glands, skin, spleen, sternebrae and vertebrae including marrow, stomach, thymus, thyroid gland, trachea, and urinary bladder
Tissues examined in low-dose male mice: adrenal glands, gross lesions, liver, spleen, thyroid gland
Tissues examined in low-dose female mice: gross lesions, liver, lungs, ovaries, salivary glands, thyroid gland
Statistics:
Survival probabilities estimated by product-limit procedure of Kaplan and Meier (1958), statistical analysis by methods of Cox (1972) and Tarone (1975);
Statistical significance of organ weights, hematological and clinical chemistry data analyzed by Dunn's test (Dunn, 1964) or Shirley's test (Shirley, 1977);
Data on tumor incidences analyzed by life-table analysis, logistic regression, Fisher's exact test, and Cochran-Armitage trend test
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):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
changes not biologically relevant
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
changes not biologically relevant
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
no effects observed
Description (incidence and severity):
see discussion in Section 7.7
Details on results:
CLINICAL SIGNS AND MORTALITY
No significant differences in survival were observed between any groups of either sex; no compound-related clinical signs


BODY WEIGHT AND WEIGHT GAIN (for details see Table 1)
100 mg/kg: mean body weights of male mice 5-8% lower between week 93 and 104; mean body weight of female mice 5-8% lower between week 20 to 44 and 10-14% lower between week 45 and 104
Evaluation: In females, the decrease of body weight of 10-14% between week 45 and 104 was considered to represent an adverse treatment-related effect.


HAEMATOLOGY (data from 15 mo interim sacrifice group, for details see Table 2)
100 mg/kg: male mice showed significant increases of hematocrit and erythrocytes of 13% compared to the vehicle control
Evaluation: Slight significant increases of hematocrit and erythrocytes were only found in high-dose males and are therefore considered to be of negligible biological significance.


CLINICAL CHEMISTRY (data from 15 mo interim sacrifice group, for details see Table 2)
100 mg/kg: male mice showed showed significant increases of total protein and serum albumin, of serum alkaline phosphatase and of sorbitol dehydrogenase; female mice showed significant increases of total protein and serum albumin, whereas a significantly lower activity was observed for alanine aminotransferase and sorbitol dehydrogenase
Evaluation: The statistically significant effects in general represented only slight changes. Changes in enzyme activites were contrary in both sexes. Therefore the observed changes of clinical chemistry are considered to be of negligible biological significance.


ORGAN WEIGHTS (data from 15 mo interim and terminal sacrifice group, for details see Table 1)
100 mg/kg: relative liver weights of both sexes showed significant increases after 65 and 103 w of administration, whereas relative brain and kidney weights of females showed significant increases only at interim sacrifice after 65 w
50 mg/kg: relative liver weight of males showed a significant increase after 103 w, relative kidney weight of females showed a significant increase after 65 w only
Evaluation: Increases of relative liver weights in both sexes at both doses correlated with histopathological non-neoplastic and neoplastic liver changes (see below). However, increases of relative weights of brains and kidneys in female mice were only observed at the interim but not at the terminal sacrifice and are therefore considered to be of negligible biological significance.


HISTOPATHOLOGY: NON-NEOPLASTIC (for details see Table 3)
LIVER 65-week interim sacrifice:
50 and 100 mg/kg, male mice: increased incidences of diffuse centrilobular fatty change, diffuse cytomegaly, occasional syncytial cells (multinucleated hepatocytes)
LIVER 105-week sacrifice:
100 mg/kg, male mice: increased incidence of anisokaryosis (variation in size of hepatocyte nuclei), syncitial alteration (hepatocytes with more than 5 nuclei per cell), and foci of cellular alteration (basophilic foci) compared to vehicle controls
Evaluation: Centrilobular fatty change and cytomegaly were observed at the 65-week interim sacrifice but not at the terminal sacrifice. This may be explained by the fact that after the 65-week administration necropsy occurred within 24 hrs after the last dose, whereas dosing was stopped 2 weeks before necropsy in the 2 year study. Presumably, fatty change and cytomegaly were reversible after cessation of hydroquinone administration and are considered not to represent a toxicologically relevant effect.


HISTOPATHOLOGY: NEOPLASTIC
For detailed evaluation see Section 7.7
Dose descriptor:
LOAEL
Remarks:
chronic non-neoplastic and neoplastic effects
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: liver: increase of weights, histopathologal findings (at 65-week interim sacrifice only)
Dose descriptor:
LOAEL
Remarks:
chronic non-neoplastic and neoplastic effects
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: Significant increase of liver adenoma or carcinoma combined
Critical effects observed:
not specified

Table 1: Body weights and relative organ weights after 65 and 103 w of gavage (mean ± standard error)

Application period

Parameter

Dose groups males

Dose groups females

Vehicle control

50 mg/kg

100 mg/kg

Vehicle control

50 mg/kg

100 mg/kg

65 w

Body weight (g)

45.6±1.26

44.9±1.34

46.9±1.01

46.7±2.51

42.4±2.54

40.6±1.17

Brain *

10.6±0.41

11.0±0.33

10.2±0.21

10.7±0.48

12.2±0.74

12.6±0.35a

Kidney *

18.0±0.63

19.6±0.41

19.2±0.57

11.2±0.30

13.1±0.44b

13.3±0.26b

Liver *

44.6±2.32

52.5±5.35

54.8±3.88b

40.5±0.95

40.9±1.15

45.2±1.25b

103 w

Number of animals weighed

33

36

36

37

39

36

Body weight

44.0±0.76

43.0±0.72

42.0 ±0.95

50.7±1.65

51.5±1.46

47.8±1.24

Brain *

11.7±0.21

11.9±0.22

12.1±0.28

10.7±0.40

10.5±0.38

11.0±0.29

Kidney *

11.8±0.38

11.7±0.23§

12.4±0.32

7.5±0.44#

7.4±0.31

7.3±0.22

Liver *

67.2±4.80

76.4±4.82

70.0±3.17a

52.0±3.22#

52.0±2.65

55.1±2.68a

* relative organ weights as mg of organ per g body weight; § N=35 mice weighed; # N=36 mice weighed

Statistically significant increased compared to vehicle control: a p<0.05

Table 2: Overview of statistically significant haematological and clinical chemical parameters in the 65-week gavage studies of hydroquinone

Affected sex

Endpoint

Parameter

Dose group

Vehicle control

50 mg/kg

100 mg/kg

Male

Hematology

Hematocrit (%)

36.5±2.38

39.5±1.27

41.3±1.17a

Erythrocytes (106/µL)

7.3±0.52

8.0±0.29

8.3±0.28a

Clinical chemistry

Total protein (g/dL)

5.2±0.04

5.4±0.15

5.9±0.23b

Albumin (g/dL)

3.3±0.05

3.5±0.08

3.8±0.15b

Alkaline phosphatase (IU/L)

38.4±2.34

38.8±1.41

50.0±3.92a

Sorbitol dehydrogenase (SU/mL)

35.8±1.24

35.6±2.05

43.0±1.79b

Female

Clinical chemistry

Total protein (g/dL)

5.3±0.07

5.5±0.12

5.7±0.05b

Albumin (g/dL)

3.5±0.06

3.6±0.10

3.9±0.04b

Alanine aminotransferase (IU/L)

38.9±6.62

31.6±6.19

23.7±1.33b

Sorbitol dehydrogenase (SU/mL)

35.6±1.09

33.7±1.37

32.4±0.73a

statistically different from vehicle control: a p<0.05, b p<0.0.1

Table 3: Number of mice with selected lesions in the liver and thyroid gland in the 65 w and 103 w gavage study of hydroquinone

Application period

Site of lesion

Type of lesion

Dose groups males

Dose groups females

Vehicle control

50 mg/kg

100 mg/kg

Vehicle control

50 mg/kg

100 mg/kg

65 w

Number of animals examined

10

10

10

10

10

10

Liver

Diffuse centrilobular fatty change

1

0

7

0

0

0

Diffuse fatty change

0

0

0

1

3

0

Diffuse cytomegaly

0

8

10

0

0

0

Syncytial cells

1

1

4

0

0

0

Basophilic focus

0

0

1

0

0

0

Clear cell focus

0

0

0

0

0

1

Hepatocellular adenoma

1

1

4

0

1

0

Hepatocellular carcinoma

2

1

1

0

0

0

Hepatocellular adenoma or carcinoma

3

2

4

0

1

0

Thyroid gland

Hyperplasia

0

0

0

0

0

2

103 w

Liver

Number of animals examined

55

54

55

55

55

55

Anisokaryosis

0/55 (0%)

2/54 (4%)

12/55 (22%)

n.s.

n.s.

n.s.

Syncytial alteration

5/55 (9%)

3/54 (6%)

25/55 (45%)

n.s.

n.s.

n.s.

Basophilic focus

2/55 (4%)

5/54 (9%)

11/55 (20%)

2/55 (4%)

6/55 (11%)

3/55 (5%)

Hepatocellular adenoma

9/55 (16%)
P=0.018

21/54 (39%)
P=0.008

20/55 (36%)
P=0.015

2/55 (4%)
P=0.007

15/55 (27%)
P=0.001

12/55 (22%)
P=0.005

Hepatocellular carcinoma

13/55 (24%)

11/54 (20%)

7/55 (13%)

1/55 (2%)

2/55 (4%)

2/55 (4%)

Hepatocellular adenoma or carcinoma a

20/55 (36%)
P=0.223

29/54 (54%)
P=0.053

25/55 (45%)
P=0.250

3/55 (5%)
P=0.009

16/55 (29%)
P=0.002

13/55 (24%)
P=0.007

Thyroid gland

Number of animals examined

55

53

54

55

55

55

Hyperplasia

5/55 (9%)

15/53 (28%)

19/54 (35%)

13/55 (24%)

47/55 (85%)

45/55 (82%)

Adenoma

2/55 (4%)

1/53 (2%)

2/54 (4%)

3/55 (5%)
P=0.186

5/55 (9%)
P=0.397

6/55 (11%)
P=0.233

Carcinoma

0/55 (0%)

0/53 (0%)

0/54 (0%)

0/55 (0%)

0/55 (0%)

1/55 (2%)

Adenoma or carcinoma b

2/55 (4%)

1/53 (2%)

2/54 (4%)

3/55 (5%)
P=0.115

5/55 (9%)
P=0.397

7/55 (13%)
P=0.152

Historical incidences in control groups (means±SD):

a male mice: water vehicle controls: 106/347 (31±6 %), range 20 to 38%; untreated controls 609/2,032 (30±8%), range 16 to 58%

  female mice: water vehicle controls: 29/348 (8±5 %), range 0 to 14%; untreated controls 184/2,032 (9±5%), range 2 to 20 %

b female mice: water vehicle controls 10/337 (3±2%), range 0 to 6 %; untreated controls 49/1,937 (3±3%), range 0 to 15%

Conclusions:
The LOAEL for chronic toxicity was 50 mg/kg bw/d both in male B6C3F1 mice based on liver weight effects and liver histopathology at the 65-week interim sacrifice, and in female mice based on increased incidences of hepatocellular adenomas and of hepatocellular adenomas or carcinomas combined (detailed evaluation of neoplastic histopathology in Section 7.7). Female mice showed non-neoplastic findings (decrease of body weights) at 100 mg/kg bw/d only.
Executive summary:

HQ was administered to groups of 65 male and 65 female B6C3F1 mice for 103 weeks on 5 days per week via gavage at doses of 0, 50, and 100 mg/kg bw/d (vehicle water). The study protocol was similar to OECD Guideline 453 with several minor deviations. There were only 2 instead of 3 dose groups, and food consumption and urinalysis were not examined. Hematology and clinical chemistry examinations were performed at a single timepoint after 65 w of dosing in subgroups of 10 mice of each sex and dose. Mice were sacrificed for gross and histopathological examination at about 66 and 104 to 105 w after start of dosing.

There were no significant differences in survival between any groups of either sex and no compound-related clinical signs. There were no biologically significant changes of haematological or clinical chemistry parameters. Toxicity was indicated at 100 mg/kg by decreased body weights in males (5-8% lower than those of vehicle controls between weeks 93-104), and in females (5-8% lower between week 20-44, 10-14% lower between week 45-104). Relative brain weights (at 100 mg/kg) and kidney weights (at 50 and 100 mg/kg) showed significant increases in female mice at the interim sacrifice only and were considered to be of negligible biological significance. Significant increases of relative liver weights were found at 100 mg/kg in both sexes after 65 and 103 w of administration, and at 50 mg/kg in males after 103 w. Centrilobular fatty change and cytomegaly were observed in male mice at the 65-week interim sacrifice but not at the terminal sacrifice. This may be explained by the fact that after the 65-week administration necropsy occurred within 24 hrs after the last dose, whereas dosing was stopped 2 weeks before necropsy in the 2 year study. Presumably, fatty change and cytomegaly were reversible after cessation of hydroquinone administration. Findings in livers of male mice at 100 mg/kg after 105 w were characterized by increased incidences of anisokaryosis (variation in size of hepatocyte nuclei), syncitial alteration (hepatocytes with more than 5 nuclei per cell), and foci of cellular alteration (basophilic foci) compared to incidences in vehicle controls. Female mice showed increased incidences of hepatocellular adenomas and of hepatocellular adenomas or carcinomas (combined) at 50 and 100 mg/kg bw/d (for detailed presentation and discussion of neoplastic findings see Section 7.7).

The LOAEL for chronic toxicity was 50 mg/kg bw/d both in male B6C3F1 mice based on liver weight effects and liver histopathology at the 65-week interim sacrifice, and in female mice based on increased incidences of hepatocellular adenomas and of hepatocellular adenomas or carcinomas combined. Female mice showed non-neoplastic findings (decrease of body weights) at 100 mg/kg bw/d only.

Endpoint:
chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
September 1982 to September 1984
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Study comparable to guideline study with acceptable restrictions: only 2 instead of 3 dose groups, no data on food consumption; hematology and clinical chemistry examinations only at 15 mo, no urinalysis data
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
reference to other study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)
Deviations:
yes
Remarks:
(only 2 instead of 3 dose groups, no data on food consumption; hematology and clinical chemistry examinations only at 15 mo, no urinalysis data)
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Fischer 344
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Kingston, NY, USA
- Age at study initiation: males 7-8 w , females 8-9 w
- Fasting period before study: no
- Housing: 5 per cage
- Diet: NIH 07 rat ration (Zeigler Bros. Inc., Gardner, PA, USA) ad libitum
- Water: tap water ad libitum
- Acclimation period: males 21 d, females 28 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 27 °C (65 - 80 °F)
- Humidity (%): 40 - 79
- Air changes (per hr): 6 - 13 room air changes per hr
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: appropriate amount of hydroquinone dissolved in deionised water by stirring with a magnetic stir bar; maximum storage time 21 d at room temperature in the dark in amber serum vials with Teflon-lined seals, sparged with argon or nitrogen before sealing

VEHICLE
- Concentration in vehicle: 5, 10 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
UV spectroscopy of acetonitrile extracts at 295 nm or methanol extracts at 293 nm; analyzed concentrations (data from 10 samples each): mean 4.88, range 4.72 - 5.09 mg/mL; mean 9.78, range 9.39 – 10.25 mg/mL; variation considered to be in acceptable range; stability investigations by HPLC after 21 days of storage of the dosing solution at room temperature in the dark indicated no notable decrease of hydroquinone concentration
Duration of treatment / exposure:
65 or 103 w , sacrifice after ca. 66 and 104 w
Frequency of treatment:
5 d/w
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Dose / conc.:
25 mg/kg bw/day (actual dose received)
Dose / conc.:
50 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
64 or 65 males, 65 females (including 10 for 65 w treatment)
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: based on 13 week range-finding toxicity study (see separate endpoint study record)
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: 2 times per day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: 2 times per day

BODY WEIGHT: Yes
- Time schedule for examinations: at initiation of dosing, weekly thereafter for 13 weeks, and monthly for the rest of the study period

FOOD CONSUMPTION: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes, at 15 mo
Investigated parameters: leukocytes, lymphocytes, segmented neutrophiles, monocytes, eosinophils, atypical lymphocytes, atypical mononuclear cells, bands, hematocrit, hemoglobin, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, mean cell volume, erythrocytes, reticulocytes

CLINICAL CHEMISTRY: Yes, at 15 mo
Investigated parameters: albumin, alkaline phosphatase, alanine aminotransferase, blood urea nitrogen, creatinine, sorbitol dehydrogenase, total protein

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes, all animals

ORGAN WEIGHTS: Yes, at 15 mo and 2 y

HISTOPATHOLOGY: Yes
Tissues examined in all rats (except for preputial gland and thyroid gland for low dose male rats): adrenal glands, brain, caecum, colon, duodenum, epididymis/prostrate/testes or ovaries/uterus, esophagus, gross lesions and tissue masses, heart, ileum, jejunum, kidneys, liver, lungs and mainstem bronchi, mammary gland, mandibular or mesenteric lymph nodes, nasal cavity and turbinates, pancreas, parathyroid glands, pituitary gland, preputial or clitoral gland, rectum, salivary glands, skin, spleen, sternebrae and vertebrae including marrow, stomach, thymus, thyroid gland, trachea, and urinary bladder
Statistics:
Survival probabilities estimated by product-limit procedure of Kaplan and Meier (1958), statistical analysis by methods of Cox (1972) and Tarone (1975);
Statistical significance of organ weights, hematological and clinical chemistry data analyzed by Dunn's test (Dunn, 1964) or Shirley's test (Shirley, 1977);
Data on tumor incidences analyzed by life-table analysis, logistic regression, Fisher's exact test, and Cochran-Armitage trend test
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):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
changes not biologically relevant
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
no effects observed
Description (incidence and severity):
for evaluation see Section 7.7
Details on results:
CLINICAL SIGNS AND MORTALITY
No significant differences in survival were observed between any groups of either sex; no compound-related clinical signs


BODY WEIGHT AND WEIGHT GAIN (for details see Table 1)
50 mg/kg: mean body weights of male rats 5-9% lower between week 73 and 93, and 10-13% lower between week 94 and 104; mean body weight of female rats within 4% of those of the vehicle controls throughout the studies
25 mg/kg: mean body weights of male rats 5-9% lower between week 89 and 104; mean body weight of female rats within 4% of those of the vehicle controls throughout the studies
Evaluation: For male rats the decrease of body weight of at least 10% observed between week 94 and 104 is considered to represent an adverse treatment-related effect.


HAEMATOLOGY (data from 15 mo interim sacrifice group, for details see Table 2)
50 mg/kg: female rats showed slight but significant decreases of hematocrit, hemoglobin concentration and erythrocytes (P<0.05)
25 mg/kg: male rats showed slight but significant increase of lymphocytes
Evaluation: Slight significant decreases of hematocrit, hemoglobin concentration and erythrocytes (90% of values of vehicle control) in high-dose female rats are consistent with mild regenerative anemia. However, with decreases of 10% only, the changes may be in the range of biological variation. Additionally, regenerative anemia was only indicated in female but not in male rats, so that the effect is considered to be of negligible biological relevance. In males, a significant increase of lymphocytes was only observed at the low dose and is also considered to be of negligible biological significance.


CLINICAL CHEMISTRY (data from 15 mo interim sacrifice group, for details see Table 2)
50 mg/kg: male rats showed slight but significant decrease of serum alkaline phosphatase
25 mg/kg: female rats showed significant increase of serum alkaline phosphatase
Evaluation: Statistically significant changes were singular effects in one sex at a single dose and are considered to be of negligible biological significance.


ORGAN WEIGHTS (data from 15 mo interim and terminal sacrifice group, for details see Table 1)
50 mg/kg: male rats showed significant increases of relative brain, kidney, and liver weights after 103 w of administration, and significant increases of relative kidney weights after 65 w of administration; no data on absolute organ weights
Evaluation: The increases in relative kidney weights, both after 65 and 103 w of exposure, existed in combination with increased severity of nephropathy, and proliferative and neoplastic findings in the kidneys. In contrast, increases of relative brain and liver weights were only obvious at the termination of the study combined with decreased body weight, but not at the interim sacrifice when body weight was not yet affected. In addition, in the absence of substance-related histopathological findings in the liver and brain, the biological significance of the increased relative brain and liver weights of male rats is questionable.


HISTOPATHOLOGY: NON-NEOPLASTIC
KIDNEY 66-week interim sacrifice:
25 and 50 mg/kg, male rats: treatment-related increased severity of nephropathy (for details see Table 3)
KIDNEY 104-week sacrifice:
50 mg/kg, male rats: increased severity of nephropathy compared to vehicle controls (for details see Table 3). Advanced renal disease characterized by varied degrees of degeneration and regeneration of tubular epithelium, atrophy and dilatation of some tubules, hyaline casts in the tubular lumina, glomerulosclerosis, interstitial fibrosis, and chronic inflammation, cysts (dilated tubules in the renal cortex), and increased papillary hyperplasia of the transitional epithelium overlying the renal papillae


HISTOPATHOLOGY: NEOPLASTIC
Detailed evaluation presented in Section 7.7

Dose descriptor:
LOAEL
Remarks:
chronic non-neoplastic and neoplastic effects
Effect level:
50 mg/kg bw/day (actual dose received)
Sex:
male
Basis for effect level:
other: body weight reduction, increased severity of CPN, significant increase of renal tubuli adenoma
Dose descriptor:
LOAEL
Remarks:
chronic non-neoplastic and neoplastic effects
Effect level:
25 mg/kg bw/day (actual dose received)
Sex:
female
Basis for effect level:
other: Significant increase of mononuclear leukaemia
Dose descriptor:
NOAEL
Remarks:
chronic toxicity
Effect level:
25 mg/kg bw/day (actual dose received)
Sex:
male
Basis for effect level:
other: no biologically relevant adverse effects
Critical effects observed:
not specified

Table 1: Body weights and relative organ weights after 65 and 103 w of gavage (mean ± standard error)

Application period

Parameter

Dose groups males

Dose groups females

Vehicle control

25 mg/kg

50 mg/kg

Vehicle control

25 mg/kg

50 mg/kg

65 w

Body weight (g)

492±9.6

504±8.1

466±10.3

312±11.6

307±8.1

303±7.9

Brain*

4.4±0.07

4.2±0.10

4.6±0.10

6.3±0.18

6.1±0.17

6.3±0.17

Kidney*

6.2±0.12

6.6±0.20

6.8±0.14b

6.6±0.52

6.0±0.19

6.0±0.12

Liver*

33.6±0.65

33.7±0.78

36.8±1.29

29.8±1.16

30.8±0.79

31.6±0.73

103 w

Number of animals weighed

27

18

18

39

27

31

Body weight (g)

464±9.2

434±8.5a

402±11.9b

337±7.1

360±13.1

330±7.4

Brain*

4.9±0.11

5.2±0.12a

5.6±0.18b

5.9±0.16

5.6±0.17

6.0±0.15

Kidney*

4.5±0.12

4.7±0.14

6.6±0.59b

3.9±0.11

3.7±0.12

4.0±0.10

Liver*

46.6±2.09

47.6±2.76

53.9±3.04a

37.2±0.96

38.1±1.25

41.2±1.71

* relative organ weights as mg of organ per g body weight

Statistically significant increase compared to vehicle control: a p<0.05, b p<0.0.1

Table 2: Overview of statistically significant haematological and clinical chemical parameters in the 65-week gavage studies of hydroquinone

Affected sex

Endpoint

Parameter

Dose group

Vehicle control

25 mg/kg

50 mg/kg

Male

Hematology

Lymphocytes (1,000 µL)

1.2±0.14

1.8±0.15a

1.9±0.37

Clinical chemistry

Alkaline phosphatase (IU/L)

139±4.7

133±5.1

132±17.6a

Female

Hematology

Hematocrit (%)

40.2±0.69

38.9±0.89

36.0±1.70a

Hemoglobin (g/dL)

14.8±0.27

14.3±0.26

13.4±0.62a

Erythrocytes (106/µL)

7.3±0.13

7.1±0.15

6.5±0.40a

Clinical chemistry

Alkaline phosphatase (IU/L)

119±11.2

150±4.6a

142±5.9

a statistically different from vehicle control with P<0.05

Table 3: Number of rats with indicated severity of nephropathy in the 65 w and 103 w gavage study of hydroquinone b

Application period

Severity

Dose groups males

Dose groups females

Vehicle control

25 mg/kg

50 mg/kg

Vehicle control

25 mg/kg

50 mg/kg

65 w

Number of rats examined

10

10

10

10

10

10

No nephropathy

0

0

0

4

5

3

minimal

0

0

0

4

1

3

mild

10

5

4

1

4

4

moderate

0

5

6

1

0

0

103 w

Number of rats examined

55

55

55

55

55

55

No nephropathy

2

3

0

8

8

9

minimal

3

1

3

47 *

47 *

46 *

mild

12

12

5

moderate

26

31

15

marked

12

8

32

* no grading of severity of nephropathy for female rats of the 2 year study

b original diagnoses of NTP report; a reevaluation of renal histopathology was published by Hard et al. (1997)

Conclusions:
The LOAEL for chronic oral toxicity was 50 mg/kg bw/d in male rats based on body weight reduction, increased severity of CPN, and increased incidences of renal adenomas. Female F344 rats were found to be less sensitive as indicated by a NOAEL of 50 mg/kg bw/d for non-neoplastic toxicity. At 25 and 50 mg/kg bw/d, statistically significant increased incidences of mononuclear cell leukaemia were observed in female rats. Both, the findings in the kidneys of male rats as a species-specific effect, and the findings of mononuclear cell leukaemia in female rats which is a lesion occuring at a high spontaneous rate, are considered not to be relevant for human risk assessment (see also Section 7.7 and 7.9.3).
Executive summary:

HQ was administered to groups of 65 male and 65 female F344 rats for 103 weeks on 5 days per week via gavage at doses of 0, 25, and 50 mg/kg bw/d (vehicle water). The study protocol was similar to OECD Guideline 453 with several minor deviations. There were only 2 instead of 3 dose groups, and food consumption and urinalysis were not examined. Hematology and clinical chemistry examinations were performed at a single timepoint after 65 w of dosing in subgroups of 10 rats of each sex and dose. Rats were sacrificed for gross and histopathological examination at about 66 and 104 w after start of dosing.

There were no significant differences in survival between any groups of either sex and no compound-related clinical signs. In male rats, toxicity was indicated at 50 mg/kg by decreases of body weights (5-9% between week 73-93, 10-13% between week 94-104). Mean body weights of low-dose male rats were 5-9% lower between week 89 and 104. Mean body weights of female rats were within 4% of those of the vehicle controls throughout the studies.

Based on haematological analyses, a finding of mild regenerative anaemia was discussed in female rats at 50 mg/kg bw/d, but the significant decreases of hematocrit, hemoglobin concentration and erythrocytes by 10% compared to the vehicle control may be within the range of biological variation. Additionally, male rats showed no treatment related changes of haematological parameters so that the effect in females is considered to be of negligible biological significance. There was no effect of treatment on clinical chemistry parameters in any sex, or on organ weights or non-neoplastic histopathological findings in female rats. Male rats of the 50 mg/kg-group showed significant increases of relative brain and liver weights combined with decreased body weight after 103 w of administration. In absence of histopathological findings these organ weight changes are considered to be of questionable biological significance. Relative kidney weights of the high-dose males were increased after 65 and 103 w of administration. All male rats of all groups (dosed and vehicle control) showed chronic progressive nephropathy (CPN), which is a common effect in ageing F344 rats. Severity of CPN was increased at the 66-week interim sacrifice in both dose groups and at the terminal sacrifice in the high-dose group (5/55 mild, 15/55 moderate, 32/55 marked compared to respective gradings of 12/55, 26/55, 12/55 in the vehicle control). Advanced renal disease was characterized by varied degrees of degeneration and regeneration of tubular epithelium, atrophy and dilatation of some tubules, hyaline casts in the tubular lumina, glomerulosclerosis, interstitial fibrosis, and chronic inflammation, cysts (dilated tubules in the renal cortex), and increased papillary hyperplasia of the transitional epithelium overlying the renal papillae. At 50 mg/kg bw/d, a significant increase of renal tubuli adenoma was observed in male rats. At 25 and 50 mg/kg bw/d, statistically significant increased incidences of mononuclear cell leukaemia were observed in female rats (for detailed discussion of neoplastic histopathology see Section 7.7)

In male F344 rats, the LOAEL for chronic toxicity was 50 mg/kg bw/d, based on body weight reduction, increased severity of CPN as an effect specific for this strain of rats, and the finding of renal adenomas. The NOAEL for male rats was 25 mg/kg bw/d. Female F344 rats were found to be less sensitive as indicated by a NOAEL of 50 mg/kg bw/d for non-neoplastic toxicity. At 25 and 50 mg/kg bw/d, statistically significant increased incidences of mononuclear cell leukaemia were observed in female rats. Both, the findings in the kidneys of male rats as a species-specific effect, and the findings of mononuclear cell leukaemia in female rats which is a lesion occuring at a high spontaneous rate, are considered not to be relevant for human risk assessment (see also Section 7.7 and 7.9.3).

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
October 27, 1987 - October 31, 1988
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Study similar to OECD Guideline 424 (Neurotoxicity study in rodents); with regard to the requests of OECD Guideline 408 investigations focussed on neurotoxicity and nephrotoxicity; study acceptable as key study on repeated dose toxicity
Reason / purpose for cross-reference:
reference to same study
Qualifier:
equivalent or similar to guideline
Guideline:
other: OECD Guideline 424 (Neurotoxicity study in rodents)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: EPA OTS 798.6050 (Neurotoxicity Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: Hydroquinone Final Test Standards and Reporting Requirements (52 FR 19865-19870, May 28, 1987)
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Wilmington, MA, USA
- Age at study initiation: ca. 7 w
- Weight at study initiation: males 194 +- 7 g, females 153 +- 6 g
- Fasting period before study:
- Housing: single, males and females in separate rooms, no other studies housed in these rooms
- Diet: Agway Prolab Animal Diet (RMH 3200) ad libitum
- Water: drinking water ad libitum
- Acclimation period: 12 d


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 - 23 °C (70- 74 °F)
- Humidity (%): 32 - 53
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
other: distilled degassed water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: daily

VEHICLE
- Concentration in vehicle: 5%
- Amount of vehicle (if gavage): 0.4, 1.28, 4.0 mL/kg, weekly adjustment for gain of body weight
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Mean concentration determined by HPLC and UV detection at 286 nm was 5.1 +- 0.3 % in males and 5.1 +- 0.2% in females being within acceptable variance.
Duration of treatment / exposure:
13 w
Frequency of treatment:
5 d/w
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Dose / conc.:
20 mg/kg bw/day (actual dose received)
Dose / conc.:
64 mg/kg bw/day (actual dose received)
Dose / conc.:
200 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: high dose reported to produce clear behavioral effects and life-threatening toxicity during previous studies (NTP, 1986)
Positive control:
Physostigmine (1 mg/kg bw/d s.c. in propylene glycol), 2,5-hexanedione (750 mg/kg bw/d p.o. by gavage), 3,3'-iminodipropionitrile (IDPN) (2000 mg/kg i.p.) administered 5 d/w for 28 d, 14 d and as single dose, respectively (dosing period until signs of subchronic neurotoxicity appeared for the different substances).
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: 3 days prior to treatment (day -3) and at 1, 6 and 24 h, and 7, 14, 30, 60 and 91 d after administration of the initial dose; examinations conducted prior to dosing except on day 0 to avoid confounding with acute effects
- Parameters: home-cage activity / feces amount / feces color / feces consistency / urine amount / urine color (urine stains on papers under the home cages when left to stand overnight) / behavior while removing from cage

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: immediately after each dosing, daily check of mortality
- Physical examinations included hair coat, skin, eyes, mucus membranes, body temperature, respiratory rate, respiratory character, lacrimation, salivation, eye prominence, palpebral closure, convulsions, seizures, length of postictal period, biting, tremors, handling behavior.

BODY WEIGHT: Yes
- Time schedule for examinations: preexposure (day -3), day 0, 1, 4, 7, and at least once weekly thereafter

FOOD CONSUMPTION:
- Food consumption for each animal determined on day 0, 1, 4, 7, and at least once weekly thereafter and mean daily diet consumption calculated as g food/kg body weight/day: Yes

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: 3 days prior to treatment (day -3) and at 1, 6 and 24 h, and 7, 14, 30, 60 and 91 d after administration of the initial dose; examinations conducted prior to dosing except on day 0 to avoid confounding with acute effects
- Dose groups that were examined: all groups
- Battery of functions tested: locomotor activity / rearing activity / tail elevation /static limb position / abnormal gait / hypotonic gait of hindlimb and forelimb / ataxic gait /head position / approach response / touch response / auditory orientation / olfactory orientation / righting reflex / proprioceptive positioning / tail pinch / abdominal tone / bladder tone / muscle tone / passive manipulation / pupillary size / eye position / nystagmus / visual orientation / palpebral reflex / pinna touch response / extensor postural thrust reflex / hopping reflex / visual placing / grip strength (semi-quantitative) / bizarre behavior / urination / defecation / spontaneous vocalizations
forelimb and hindlimb grip strength by procedure of Meyer et al. (1979) and modified by Mattson et al. (1986): determined three times at each time interval and the highest recorded value was used for statistical analysis
Sacrifice and pathology:
- Treatment of animals:
6 animals per sex and dose group, that were designated for neuropathological examination, were anesthesized and perfused through the ascending aorta with 4% paraformaldehyde followed by 5% glutaraldehyde both in 0.1 M phosphate buffer, pH 7.4 at 4 °C. Following perfusion brain, spinal cord and sciatic-tibial nerves were removed. Tissues destined for plastic embedment were fixed for an additional 2 h in 5% glutaraldehyde. Tissues not destined for plastic embedment and all tissues from unperfused animals were immersed in 10% neutral buffered formalin.
The remaining animals were necropsied without perfusion.

GROSS PATHOLOGY: Yes
- Organ weights: brains and kidneys both from perfused and unperfused animals were collected and analyzed separately depending on the method of fixation

HISTOPATHOLOGY: Yes
- Investigations of the nervous system:
Cross sections through the forebrain, cerebrum, midbrain, cerebellum, pons, medulla oblongata, cervical spinal cord swelling, luminal spinal cord swelling, cervical and lumbar spinal cord ganglia, dorsal and ventral spinal roots (cervical and lumbar), sciatic nerve at mid-thigh, and tibial nerve including branches to the calf musculature
Tissues from the high-dose and the control group: processed by standard neuropathological techniques for paraffin embedment, hematoxylin-eosin staining, and light microscopic examination; special stains (combined Luxol Fast Blue Stain-Bodian silver protargol impregnation) performed on perfused neural tissue; left sciatic-tibial nerve and medulla oblongata postfixed in Dalton's chrome-osmium solution, embedded inplastic, sectioned at 1 mm, stained with toluidine blue, and examined by light microscopy.
- Investigation of kidneys: imbedding by standard methods, staining with hematoxylin-eosin and light microscopic examination
Statistics:
Statistical significance of subchronic toxicity study data evaluated by ANOVA (p<= 0.05), Bartlett's test (p<=0.01), Duncan's multiple range test (p<=0.05). FOB results analyzed using contingency table methods and Dunnett's t-test, modified for proportions. Loglinear models were fit to the three-way tables and significant effects were determined. Both godness of fit and statistical significance were detected (p<= 0.05). If significant or near significant dose-behavior interactions were detected, Dunnett's t-test (one-tail) was used to compare incidences at each observation time.
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY
Depression of general motor activity (decreased movement primarily in the home cage) and appearance of tremors with a LOEL of 64 mg/kg; brown discoloration of urine stains with LOEL of 20 mg/kg; no mortality

200 mg/kg:
- minimal to mild tremors in all animals appearing between 30 and 60 min after dosing in males, and between 4 and 25 min in females; statistically significant increase of incidence of tremors during weeks 1 to 13 for both sexes and during week 14 for males only;
- statistically significant increase of incidence of depression of general activity for males (10/10) during week 1 to 13, and for females (5/10) during week 4 to 8
- minimal to moderate brown discoloration of urine stains
64 mg/kg:
- minimal to mild tremors in all females and in 7/10 males; statistically significant increase of incidence of tremors during weeks 6, 7 and 9 to 13 for males, and during weeks 2, 3, 5, 6, and 9 to 13 for the females;
- statistically significant increase of incidence of depression of general activity for males (1/10) during week 6 and (2/10) during week 8, and for females (2/10) during week 4
- minimal to moderate brown discoloration of urine stains
20 mg/kg:
- minimal to minor brown discoloration of urine stains

Evaluation: Tremors were considered to be related to hydroquinone exposure at 64 and 200 mg/kg due to frequent appearance and statistically significance compared to controls. Depression of general activity was considered to represent an adverse effect related to exposure at 200 mg/kg only, because at 64 mg/kg this effect occurred in single animals and at single timepoints only. Brown discoloration of urine indicated systemic absorption and excretion of hydroquinone and its metabolites respectively.


BODY WEIGHT AND WEIGHT GAIN
200 mg/kg, only males: slightly decreased mean body weight compared to controls from day 1 until termination of the study, no significant effect; mean terminal body weight 7% lower than for control group


FOOD CONSUMPTION
200 mg/kg: significantly decreased compared to controls between days 0 to 4; thereafter comparable to controls


NEUROBEHAVIOUR
Statistically significant behavioral changes attributed to hydroquinone exposure observed at the following dose levels primarily during the 1 and 6 h FOB examinations:
200 mg/kg:
- decreased home-cage activity and locomotor activity for males at 6 h FOB
- decreased behavior while removing from the cage for females at 1 h FOB
- increased tremors for females at 1 h FOB
- brown discoloration of urine stains for males at 1 and 6 h, days 1, 30 and 60 FOB, and for females at days 30 and 60 FOB
64 mg/kg:
- brown discoloration of urine stains for males at days 1, 30 and 60 FOB, and for females at days 30 and 60 FOB
20 mg/kg:
- brown discoloration of urine stains for males at days 1 and 60 FOB, and for females at days 30 and 60 FOB

Evaluation: The behavioral changes listed above were attributed to be related to the exposure and represented acutely observable effects. In contrast, no changes attributable to long-term exposure were evident as other statistically significant changes appeared to be random in nature, were not related to dose level, or were not reproducible with time. Changes not considered related to HQ exposure included increased urination, increased or decreased defecation, increased spontaneous vocalizations, increased approach response, increased or decreased auditory orientation, increased olfactory orientation, increased visual orientation, decreased pinna touch response, increased tail pinch response, decreased quantitative grip strength. the positive control substances resulted in characteristic neurobehavioral effects.


ORGAN WEIGHTS
No effect on mean brain or kidney weights


GROSS PATHOLOGY
No treatment-related changes


HISTOPATHOLOGY: NON-NEOPLASTIC
Kidneys: no treatment-related changes
Neuropathologic findings: single findings in 3/20 controls and 2/20 animals in the 200 mg/kg group; effects not related to treatment
Dose descriptor:
NOAEL
Effect level:
20 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects
Dose descriptor:
LOAEL
Effect level:
64 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: acute CNS effects indicated by clinical examination (tremors)
Critical effects observed:
not specified

Number of tremors occurrences in males and females during the study

Doses

Males

Females

 

Males and females

mg/kg/day

N

Number of occurrences

%

Number of occurrences

%

 

N

Number of occurrences

%

0

680

0

0

0

0

1360

0

0

20

680

0

0

0

0

1352

0

0

64

680

74

11

88

13

1360

162

12

200

680

194

29

239

34

1360

461

35

N: number of observations during the study consists of the number of dosing days*(10 animals/sex)

Tremors were observed after each dosing

Conclusions:
Subchronic oral exposure of Sprague-Dawley rats to doses of 20, 64, and 200 mg/kg bw/d did not result in evidence of subchronic neurotoxicity as assessed by functional observation battery examinations, quantitative grip strength measurements, brain weights, or neuropathology examinations. Effects were restricted to acute responses of the central nervous system to dosing of hydroquinone at doses of 64 mg/kg bw and up. In addition, no nephropathy was evident in this rat strain given 200 mg hydroquinone/kg bw/d, a dose level that was nephrotoxic to F-344 rats.
Executive summary:

Groups of Sprague-Dawley rats (CD-SD, 10 males and 10 females/group) were exposed to doses of 0, 20, 64, and 200 mg/kg bw/d by gavage application of 5% aqueous hydroquinone solutions on 5 d/w for 13 w to investigate subchronic toxicity with emphasis on neurobehavioral and neurotoxic effects and on nephrotoxic effects. The study protocol was similar to OECD Guideline 424 (Neurotoxicity study in rodents) except for application on 5 d/w instead of 7 d/w. Investigations included detailed clinical observations, a comprehensive functional observation battery, quantitative measurements of grip strength, and neuropathological examinations of the central and the peripheral nervous system as well as determination of brain and kidney weights.


Brown discoloration of urine stains in all dose groups indicated systemic absorption and urinary excretion of hydroquinone and its metabolites. Effects were restricted to responses of the central nervous system with appearance of tremors at 64 and 200 mg/kg shortly after dosing as acute response. Significant depression of general motor activity (decreased movement primarily in the home cage) was observed at 200 mg/kg in all males during the whole dosing period and in 5/10 females from week 4 to 8. However, subchronic oral exposure did not result in evidence of subchronic neurobehavioral changes or morphologic changes in the CNS or PNS. In addition, no nephropathy was evident in Sprague-Dawley rats given 200 mg hydroquinone/kg bw/d, a dose level that was found to be nephrotoxic to F344 rats in the NTP study (NTP, 1989). Under the conditions of this study, the LOAEL was 64 mg/kg bw/d based on clinical signs indicating effects on the central nervous system and the NOAEL was 20 mg/kg bw/d.

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from June 24 1981 to September 25 1981
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Remarks:
Study meets basic scientific principles as a range-finding subchronic toxicity study, sufficient documentation of test conditions and test result
Reason / purpose for cross-reference:
reference to same study
Qualifier:
no guideline followed
Principles of method if other than guideline:
13-week range-finding toxicity study investigating mortality, clinical signs, body weights and selected organ weights, as well as gross necropsy and histopathology
GLP compliance:
not specified
Limit test:
no
Species:
mouse
Strain:
B6C3F1
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Portage, MI, USA
- Age at study initiation: 8 - 9 w
- Fasting period before study: no
- Housing: 5 per cage
- Diet: Purina Rodent Laboratory Chow Pellets ad libitum
- Water: tap water ad libitum
- Acclimation period: 21 d


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 24 °C (66 - 76 °F)
- Humidity (%): 44 - 84
- Air changes (per hr): 10 - 12 room air changes per hr
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: appropriate amount of hydroquinone mixed with appropriate amount of corn oil; suspensions homogenized in Polytron homogenizer; air removed under vacuum, solutions sealed under argon; maximum storage time 11 d at room temperature in the dark in amber serum vials with Teflon-lined seals

VEHICLE
- Justification for use and choice of vehicle (if other than water): solubility
- Concentration in vehicle: 5, 10, 20, 40, and 80 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Method: UV spectroscopy of acetonitrile extracts at 295 nm or methanol extracts at 293 nm; analyzed concentrations were in the range between 88 and 132 % of the nominal concentration before start of dosing and between 90 and 103 % ca. 8 weeks later; variation of analyzed concentrations dependant on layer (top, middle or bottom) of the dosing mixture where the sample was taken, variation considered to be in acceptable range
Duration of treatment / exposure:
13 w
Frequency of treatment:
5 d/w
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Dose / conc.:
25 mg/kg bw/day (actual dose received)
Dose / conc.:
50 mg/kg bw/day (actual dose received)
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
200 mg/kg bw/day (actual dose received)
Dose / conc.:
400 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: based on 14-day range-finding toxicity study (see separate endpoint study record)
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: 2 times per day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: 2 times per day

BODY WEIGHT: Yes
- Time schedule for examinations: at initiation of dosing, weekly thereafter

FOOD CONSUMPTION: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes, all animals

ORGAN WEIGHTS: Liver

HISTOPATHOLOGY: Yes
Tissues examined in all vehicle controls and dosed rats receiving 200 or 400 mg/kg, and animals dying before end of the study: adrenal glands, brain, colon, esophagus, gall bladder, gross lesions and tissue masses, heart, kidneys, liver, lungs and mainstem bronchi, mammary gland, mandibular or mesenteric lymph nodes, pancreas, parathyroid glands, pituitary gland, prostrate/testes or ovaries/uterus, salivary glands, skin, small intestine, spleen, sternebrae and vertebrae including marrow, stomach, thymus, thyroid gland, trachea, and urinary bladder.
Statistics:
Calculation of statistical significance for body and organ weights by Dunn's test (Dunn, 1964) or Shirley's test (Shirley, 1977)
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Details on results:
CLINICAL SIGNS AND MORTALITY
400 mg/kg: mortality 8/10 males (N=6 in week 1, N=1 each in week 2 and 13), and 8/10 females (N=4 in week 1, N=1 in week 8, 9, 10 and 12); clinical signs: lethargy in all dosed males and females; tremors in all mice immediately after dosing often followed by convulsions
200 mg/kg: mortality 2/10 males in week 1 and 9, and 0/10 females; clinical signs: lethargy in all dosed males and females, tremors in male mice immediately after dosing
25 - 100 mg/kg: lethargy in all dosed males, and in females only at 100 mg/kg
Evaluation: The observation of lethargy only in males at the low dose range of 25 and 50 mg/kg bw/d was not reproducible in the chronic study at doses of 50 and 100 mg/kg bw/d and thus is considered to be of no toxicological relevance

BODY WEIGHT AND WEIGHT GAIN (For details see Table 1)
Mean body weights of dosed and vehicle control mice were similar. However, an unexpected drop in the mean body weight of the male vehicle control group was noted during weeks 12 and 13, resulting in significantly increased necropsy body weights for males of the 25 and 100 mg/kg-dose groups; slight but significant increase of necropsy body weights for females of the 50 mg/kg-dose group
Evaluation: Increases of body weights in the low dose groups not considered to be substance related as there was a drop of the final body weights in the vehicle control group and as there was no dose-response relationship.

ORGAN WEIGHTS (For details see Table 1)
Absolute mean liver weights: significant increases in all dosed males and in females at 100 and 400 mg/kg
Relative mean liver weights: significant increases in all dosed males and in females at 200 and 400 mg/kg
Evaluation: The toxicological significance of the changes of liver weights is considered to be negligible, as effects did not show a clear dose dependance and were observed in the absence of corresponding histopathological findings.

HISTOPATHOLOGY: NON-NEOPLASTIC
400 mg/kg: 3/10 males and 2/10 females with ulceration, inflammation, or epithelial hyperplasia in the forestomach
200 mg/kg: 1/10 females with ulceration, inflammation, or epithelial hyperplasia in the forestomach
up to 100 mg/kg: no findings
Evaluation: The findings in the forestomach are indicative of a local irritant action of HQ in the forestomach due to bolus application via gavage.
Dose descriptor:
NOAEL
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects
Dose descriptor:
LOAEL
Effect level:
100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: clinical sign (lethargy )
Critical effects observed:
not specified

Table 1: Liver weights and terminal body weights after 13-week gavage of hydroquinone (mean±standard error)

Sex

Dose

(mg/kg)

Number weighed

Necropsy body weight (g)

Absolute liver weight

(mg)

Relative liver weight (mg/g)

Male

0

10

30.5 ± 1.33

1,253 ± 56

41.2 ± 1.40

25

10

37.6 ± 1.30b

2,016 ± 79b

54.0 ± 2.38b

50

10

35.7 ± 0.53

1,826 ± 46b

51.2 ± 1.05a

100

9 *

37.5 ± 0.75b

1,818 ± 37b

48.9 ± 1.35b

200

8

35.1 ± 0.67

2,000 ± 112b

57.1 ± 3.37b

400

2

31.8 ± 1.30

1,750 ± 120a

55.0 ± 1.53b

Female

0

10

25.0 ± 0.41

1,230 ± 31

49.1 ± 1.08

25

10

26.5 ± 0.42

1,331 ± 62

50.4 ± 2.47

50

10

26.9 ± 0.51a

1,309 ± 33

48.8 ± 1.02

100

10

26.4 ± 0.55

1,396 ± 60a

52.8 ± 1.39

200

10

25.3 ± 0.38

1,338 ± 47

52.9 ± 1.25a

400

2

26.1 ± 0.35

1,505 ± 85a

57.8 ± 4.04a

Statistically different from control group: a p<0.05; b p<0.01

* One weight was not recorded at necropsy

Conclusions:
When applied on 5 days per week for 13 weeks by gavage, hydroquinone at doses of 100 mg/kg and below had negligible effects in mice. Because of deaths and forestomach lesions at doses of 200 mg/kg bw/d and up, doses selected for the 2-year studies with mice were 50 and 100 mg/kg bw/d.
Executive summary:

Hydroquinone was administered to groups of 10 male and 10 female B6C3F1 mice for 13 weeks on 5 days per week via gavage at doses of 0, 25, 50, 100, 200 and 400 mg/kg bw/d (vehicle corn oil) as a range-finding toxicity study for a subsequent 2-year study. Investigated endpoints were mortality, clinical signs,body weights and selected organ weights, as well as gross necropsy and a comprehensive histopathology of tissues.

Eight from10 mice died from both sexes each at 400 mg/kg, and 2/10 male mice at 200 mg/kg, mostly within the first week of the study. Male mice of all dose groups and female mice at doses ranging from 100 – 400 mg/kg exhibited lethargy. The observation of lethargy only in males at the low dose range of 25 and 50 mg/kg bw/d was not reproducible in the chronic study at doses of 50 and 100 mg/kg bw/d and thus is considered to be of no toxicological relevance. Immediately after dosing tremors were seen in all mice of the 400 mg/kg group and in male mice of the 200 mg/kg group. In the high-dose group tremors were often followed by convulsions. There was no treatment-related effect on body weights or on liver weights. Ulceration, inflammation, or epithelial hyperplasia in the forestomach were found in 3/10 males and 2/10 females at 400 mg/kg, and in 1/10 females at 200 mg/kg, indicating a local irritant action in the forestomach due to bolus application via gavage.Subchronic administration of hydroquinone at doses of 100 mg/kg and below had negligible effects in mice resulting in a NOAEL of 100 mg/kg bw/d and a LOAEL of 200 mg/kg bw/d.

 

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from June 26 1981 to September 28 1981
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Reason / purpose for cross-reference:
reference to same study
Qualifier:
no guideline followed
Principles of method if other than guideline:
13-week range-finding toxicity study investigating mortality, clinical signs and body weights and selected organ weights, as well as gross necropsy and histopathology
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Fischer 344
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Portage, MI, USA
- Age at study initiation: 7-8 w
- Fasting period before study: no
- Housing: 5 per cage
- Diet: NIH 07 rat ration (Zeigler Bros. Inc. , Gardner, PA, USA) ad libitum
- Water: tap water ad libitum
- Acclimation period: 22 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 24 °C (66 - 76 °F)
- Humidity (%): 44 - 84
- Air changes (per hr): 10 - 12 room air changes per hr
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: appropriate amount of hydroquinone mixed with appropriate amount of corn oil; suspensions homogenized in Polytron homogenizer; air removed under vacuum, solutions sealed under argon; maximum storage time 11 d at room temperature in the dark in amber serum vials with Teflon-lined seals

VEHICLE
- Justification for use and choice of vehicle (if other than water): solubility
- Concentration in vehicle: 5, 10, 20, 40, and 80 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Method: UV spectroscopy of acetonitrile extracts at 295 nm or methanol extracts at 293 nm; analyzed concentrations were in the range between 90 and 110 % of the nominal concentration before start of dosing and between 96 and 101 % ca. 8 weeks later; variation of analyzed concentrations dependant on layer (top, middle or bottom) of the dosing mixture where the sample was taken, variation considered to be in acceptable range
Duration of treatment / exposure:
13 w
Frequency of treatment:
5 d/w
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Dose / conc.:
25 mg/kg bw/day (actual dose received)
Dose / conc.:
50 mg/kg bw/day (actual dose received)
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
200 mg/kg bw/day (actual dose received)
Dose / conc.:
400 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: based on 14-day range-finding toxicity study (see separate endpoint study record)
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: 2 times per day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: 2 times per day

BODY WEIGHT: Yes
- Time schedule for examinations: at initiation of dosing, weekly thereafter

FOOD CONSUMPTION: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes, all animals

ORGAN WEIGHTS: Liver

HISTOPATHOLOGY: Yes
Tissues examined in all vehicle controls and dosed rats receiving 200 or 400 mg/kg, and animals dying before end of the study: adrenal glands, brain, colon, esophagus, gross lesions and tissue masses, heart, kidneys, liver, lungs and mainstem bronchi, mammary gland, mandibular or mesenteric lymph nodes, pancreas, parathyroid glands, pituitary gland, prostrate/testes or ovaries/uterus, salivary glands, skin, small intestine, spinal cord, spleen, sternebrae and vertebrae including marrow, stomach, thymus, thyroid gland, trachea, and urinary bladder.
Tissues examined in 100 mg/kg-group: liver, kidneys, and stomach of male rats, and kidneys of female rats
Statistics:
Statistical significance of liver weights by Dunn's test (Dunn, 1964) or Shirley's test (Shirley, 1977)
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
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):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Details on results:
CLINICAL SIGNS AND MORTALITY
400 mg/kg: mortality 10/10 males (N=8 in week 2, N=1 each in week 7 and 13), and 10/10 females (N=2 in week 1 and 2 each, N=1 in week 4 and 6, N=2 in week 5 and 7); tremors and convulsions observed in all rats immediately after dosing and preceding death, clear orange fluid or orange staining around the nose in most cases
200 mg/kg: mortality 0/10 males, and 3/10 females in week 11; clinical signs: lethargy in male rats after 10 w of dosing and in all female rats for the duration of the study, females exhibited tremors and sometimes convulsions immediately after dosing, clear orange fluid or orange staining around the nose in most cases
No effects at doses up to 100 mg/kg

BODY WEIGHT AND WEIGHT GAIN
Females: similar to controls in all dose groups, final body weight 97 to 100 % of control value
Males: final body weight significantly decreased from 50 mg/kg up (no data for food consumption available): 99, 95, 92, and 91% of control value at 25, 50, 100, and 200 mg/kg

ORGAN WEIGHTS (For details see Table 1)
Absolute liver weights: significant decreases in males from 25 to 200 mg/kg in combination with decreased body weight from 50 to 200 mg/kg; significant increase in females from 50 to 200 mg/kg without effect on body weight
Relative liver weights: significant decreases in males from 25 to 100 mg/kg; significant increase in females from 50 to 200 mg/kg
Evaluation: The toxicological significance of the changes of liver weights is questionable, as effects were opposed in male and female rats and there were no corresponding histopathological findings. Additionally, decreased liver weights in male rats of the 50 to 200 mg/kg-groups occurred with a simultaneous decrease of body weight.

GROSS PATHOLOGY
400 mg/kg: 4/10 males and 5/10 females red-to-brown perioral staining; 1/10 males and 2/10 females reddened mucosa in the stomach; 1/10 males meningeal hemorrhage
200 mg/kg: 2/10 males intra-abdominal bleeding; 1/10 females blood in the stomach, 2/10 females perioral staining
up to 100 mg/kg: no findings

HISTOPATHOLOGY: NON-NEOPLASTIC
Inflammation and epithelial hyperplasia of the forestomach: at 200 mg/kg in 4/10 males and 1/10 females, mild to moderate severity
Toxic nephropathy with tubular cell degeneration and regeneration in the renal cortex: at 200 mg/kg in 7/10 males and 6/10 females, at 100 mg/kg in 1/10 females; moderate to marked severity in males and minimal to mild severity in females; no indications for alpha2µ-globulin nephrotoxic syndrome (no hyaline droplet formation, no granular casts, no linear mineralization)
up to 50 mg/kg: no findings
Evaluation: The findings in the forestomach are indicative of a local irritant action in the forestomach due to bolus application via gavage.
Dose descriptor:
NOAEL
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects
Dose descriptor:
LOAEL
Effect level:
100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: kidney lesions in females
Dose descriptor:
LOAEL
Effect level:
100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: reduced body weight
Critical effects observed:
not specified

Table 1: Liver weights and terminal body weights after 13-week gavage of hydroquinone (mean±standard error)

Sex

Dose

(mg/kg)

Number weighed

Necropsy body weight (g)

Absolute liver weight

(mg)

Relative liver weight (mg/g)

Male

0

10

367±7.1

15,708±652

42.9±1.77

25

10

365±3.4

12,319±342b

33.7±0.76b

50

10

347± 5.6b

12,331±490b

35.5±1.41a

100

10

338±6.2b

11,227±271b

33.2±0.69b

200

10

333±5.3b

13,653±456b

40.9±1.06

Female

0

10

201±3.4

6,845±218

34.0±0.68

25

10

202±2.4

6,924±203

34.2± 0.93

50

10

200±2.8

7,611±247a

38.0±1.06b

100

10

195±2.9

7,551±224a

38.8±1.12b

200

7

196±3.1

7,990±110b

40.9±0.89b

Statistically different from control group: a p<0.05; b p<0.01

Conclusions:
When applied on 5 days per week for 13 weeks by gavage, hydroquinone at doses of 50 mg/kg and below had negligible effects in rats. Because of deaths, reductions in body weight gain, and forestomach and kidney lesions at doses of 100 mg/kg bw/d and up, doses selected for the 2-year studies with rats were 25 and 50 mg/kg bw/d.
Executive summary:

Hydroquinone was administered to groups of 10 male and 10 female F344 rats for 13 weeks on 5 days per week via gavage at doses of 0, 25, 50, 100, 200 and 400 mg/kg bw/d (vehicle corn oil) as a range-finding toxicity study for a subsequent 2-year study. Investigated endpoints were mortality, clinical signs, body weights and selected organ weights, as well as gross necropsy and a comprehensive histopathology of tissues.

At 400 mg/kg, all rats died, mostly within the first 7 weeks of the study with tremors and convulsions preceding death. At 200 mg/kg, 3/10 females died in week 11, and lethargy was observed in male rats after 10 w of dosing and in all female rats during the whole dosing period. Female rats exhibited tremors and sometimes convulsions immediately after dosing. Final body weights were similar to controls in all dose groups of females, whereas in males final body weights were decreased by 8% and 9% at 100 and 200 mg/kg, respectively (no data for food consumption available). Gross pathology findings in the 400 and 200 mg/kg-groups were red-to-brown perioral staining in 11/40 rats and single cases of reddened mucosa or blood in the stomach, intra-abdominal bleeding, and meningeal hemorrhage. Inflammation and epithelial hyperplasia of the forestomach were seen at 200 mg/kg in 4/10 males and 1/10 females, indicating a local irritant action of HQ in the forestomach due to bolus application via gavage. Toxic nephropathy, characterized by tubular cell degeneration and regeneration in the renal cortex, was seen at 200 mg/kg in 7/10 males and 6/10 females, and at 100 mg/kg in 1/10 females. Subchronic administration of hydroquinone at doses of 50 mg/kg and below had negligible effects in rats resulting in a NOAEL of 50 mg/kg bw/d and a LOAEL of 100 mg/kg bw/d.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
20 mg/kg bw/day
Study duration:
chronic
Species:
rat

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records

Referenceopen allclose all

Endpoint:
short-term repeated dose toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 1 August to 14 August 1979
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Remarks:
Study meets basic scientific principles as a range-finding toxicity study, sufficient documentation; study acceptable as key study
Reason / purpose for cross-reference:
reference to same study
Principles of method if other than guideline:
14-day range-finding toxicity study investigating mortality, clinical signs and body weights, as well as gross necropsy
GLP compliance:
not specified
Limit test:
no
Species:
mouse
Strain:
B6C3F1
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Portage, MI, USA
- Age at study initiation: 6-8 w
- Fasting period before study: no
- Housing: 5 per cage
- Diet: Purina Rodent Laboratory Chow Pellets ad libitum
- Water: tap water ad libitum
- Acclimation period: 13 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 27 °C (64 - 80 °F)
- Humidity (%): 50 - 65
- Air changes (per hr): 12 - 15 room air changes per hr
- Photoperiod (hrs dark / hrs light): 12/12
Type of coverage:
open
Vehicle:
ethanol
Details on exposure:
TEST SITE
- Area of exposure: clipped scapular area
- % coverage: no data
- Time intervals for shavings or clipplings: no data

REMOVAL OF TEST SUBSTANCE: no

TEST MATERIAL
- Amount(s) applied per mouse (volume or weight with unit): ca. 0.1 mL at 300 to 2400 mg/kg bw; two applications of 0.1 mL/mouse at 4800 mg/kg bw with a 15- to 30-min interval to allow the applied material to dry
- Concentration (if solution): up to 480 mg/mL
- Constant volume or concentration used: yes
- For solids: the dosing solution of the 480 mg/kg dose group was rather a suspension than a true solution and was homogenized in a Warring Blender and continuously stirred during dosing

VEHICLE
- Justification for use and choice of vehicle (if other than water): solubility

USE OF RESTRAINERS FOR PREVENTING INGESTION: no data
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
12 treatments in 14 d
Frequency of treatment:
5 d/w
Remarks:
Doses / Concentrations:
0, 300, 600, 1200, 2400 and 4800 mg/kg bw/d
Basis:
nominal per unit body weight
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once per day

BODY WEIGHT: Yes
- Time schedule for examinations: day 0, 7 and 14

GROSS PATHOLOGY: Yes, all animals
Other examinations:
Preliminary qualitative skin absorption study:
groups of 6 mice were treated with doses of 0, 4 oder 40 mg/animal (ca. 40 or 400 mg/kg bw applied as 0.2 mL of 2% or 20% HQ solutions in 95% ethanol) on the clipped interscapular region for three days. Rats were housed in individual metabolism cages and urine was collected at 2, 8, 24, 48 and 72 hrs after the initial dose. Hydroquinone was detected in ether extracts of original urine samples and urine treated with ß-glucuronidase and aryl sulfatase by thin layer chromatography (Uv detection after iodine treatment).
Clinical signs:
no effects observed
Dermal irritation:
not specified
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY: no signs of intoxication; crystals of the test substance were seen on the skin and fur of the 4800 mg/kg group

BODY WEIGHT AND WEIGHT GAIN
Final body weights were lower than initial weights in all groups including controls, but were comparable among all test groups.

DERMAL ABSORPTION STUDY:
HQ was detected qualitatively in urine as early as 2 hrs after application and during the whole observation period of 72 hrs (up to 24 hr after last treatment). The intensity of the spot on the TLC was higher after enzymatic cleavage of glucuronide and sulfate conjugates. However, contamination of the urine by HQ released from the skin or the fur could not be ruled out.
Dose descriptor:
NOEL
Effect level:
4 800 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects observed
Critical effects observed:
not specified
Conclusions:
In a 14 day range finding dermal toxicity study with male and female B6C3F1 mice, no signs of intoxication were observed up to 4800 mg/kg bw/d. A preliminary qualitative dermal absorption study indicated systemic availability of unknown amounts of HQ as glucuronide and sulfate conjugates were excreted in urine.
Executive summary:

In a 14 day range finding dermal toxicity study with groups of 5 male and 5 female B6C3F1 mice (12 open applications on 5 d/w of 300 to 4800 mg/kg bw/d in 95% ethanol as vehicle), there was no indication of toxicity from survival, clinical signs, body weights and gross necropsy up to 4800 mg/kg bw/application. So the NOEL in B6C3F1 mice is 4800 mg/kg bw/d. Crystals of the test substance were seen on the skin and fur of the 4800 mg/kg group. A preliminary qualitative dermal absorption study indicated systemic availability of unknown amounts of HQ as glucuronide and sulfate conjugates were excreted in urine.

Endpoint:
short-term repeated dose toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 1 August to 14 August 1979
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Remarks:
Study meets basic scientific principles as a range-finding toxicity study, sufficient documentation; study acceptable as key study
Reason / purpose for cross-reference:
reference to same study
Principles of method if other than guideline:
14-day range-finding toxicity study investigating mortality, clinical signs and body weights, as well as gross necropsy
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Fischer 344
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Portage, MI, USA
- Age at study initiation: 6 w
- Fasting period before study: no
- Housing: 5 per cage
- Diet: Purina Rodent Laboratory Chow Pellets ad libitum
- Water: tap water ad libitum
- Acclimation period: 13 d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 27 °C (64 - 80 °F)
- Humidity (%): 50 - 65
- Air changes (per hr): 12 - 15 room air changes per hr
- Photoperiod (hrs dark / hrs light): 12/12
Type of coverage:
open
Vehicle:
ethanol
Details on exposure:
TEST SITE
- Area of exposure: clipped scapular area
- % coverage: no data
- Time intervals for shavings or clipplings: no data

REMOVAL OF TEST SUBSTANCE: no

TEST MATERIAL
- Amount(s) applied per rat (volume or weight with unit): ca. 0.2 mL at 240, 480, and 960 mg/kg; 0.4 mL at 1920 mg/kg; two applications of 0.4 mL/rat with a 15- to 30-min interval to allow the applied material to dry
- Concentration (if solution): 120, 240, 480 mg/mL
- Constant volume or concentration used: no
- For solids: the dosing solution of the 480 mg/kg dose group was rather a suspension than a true solution and was homogenized in a Warring Blender and continuously stirred during dosing

VEHICLE
- Justification for use and choice of vehicle (if other than water): solubility

USE OF RESTRAINERS FOR PREVENTING INGESTION: no data
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
12 treatments in 14 d
Frequency of treatment:
5 d/w
Remarks:
Doses / Concentrations:
0, 240, 480, 960, 1920, and 3840 mg/kg bw/d
Basis:
nominal per unit body weight
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once per day

BODY WEIGHT: Yes
- Time schedule for examinations: day 0, 7 and 14

GROSS PATHOLOGY: Yes, all animals
Other examinations:
Preliminary qualitative skin absorption study:
groups of 6 rats were treated with doses of 0, 4 oder 40 mg/animal (ca. 40 or 400 mg/kg bw applied as 0.2 mL of 2% or 20% HQ solutions in 95% ethanol) on the clipped interscapular region for three days. Rats were housed in individual metabolism cages and urine was collected at 2, 8, 24, 48 and 72 hrs after the initial dose. Hydroquinone was detected in ether extracts of original urine samples and urine treated with ß-glucuronidase and aryl sulfatase by thin layer chromatography (Uv detection after iodine treatment).
Clinical signs:
no effects observed
Dermal irritation:
not specified
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):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY: no signs of intoxication; crystals of the test substance were seen on the skin and fur

FINAL BODY WEIGHT as % of control group at doses of at 240, 480, 960, 1920, and 3840 mg/kg bw/d:
males 99%, 95%, 102%, 103%, and 94%, females 106%, 100%, 103%, 100%, 99%
Evaluation: As reductions of body weight were observed only at 480 and 3840 mg/kg bw/d without a dose-relationship this effect is considered not to be a substance related effect.

DERMAL ABSORPTION STUDY:
HQ was detected qualitatively in urine as early as 2 hrs after application and during the whole observation period of 72 hrs (up to 24 hr after last treatment). The intensity of the spot on the TLC was higher after enzymatic cleavage of glucuronide and sulfate conjugates. However, contamination of the urine by HQ released from the skin or the fur could not be ruled out.
Dose descriptor:
NOEL
Effect level:
3 840 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no effects observed
Critical effects observed:
not specified
Conclusions:
In a 14 day range finding dermal toxicity study with F344 rats (12 treatments in total on 5 d/w), no substance-related effects were observed in male and female rats so that the NOEL is 3840 mg/kg bw/d. A preliminary qualitative dermal absorption study indicated systemic availability of unknown amounts of HQ, as glucuronide and sulfate conjugates were excreted in urine.
Executive summary:

In a 14 day range finding dermal toxicity study with groups of 5 male and 5 female F344 rats (12 open applications on 5 d/w of 240 to 3840 mg/kg bw/d in 95% ethanol as vehicle), no substance-related effects were observed in male and female rats so that the NOEL is 3840 mg/kg bw/d. Crystals of the test substance were seen on the skin and fur. A preliminary qualitative dermal absorption study indicated systemic availability of unknown amounts of HQ, as glucuronide and sulfate conjugates were excreted in urine. Based on the absence of substance-related effects even at the high doses, the dermal route was considered to be inappropriate for investigation of repeated dose systemic toxicity at longer treatment intervals.

Endpoint:
sub-chronic toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Comparable to guideline study with the following deviations: additional investigation of urinalysis and renal cell proliferation, addtional organ weights, histological examination of different tissues; no ophthalmoscopic examination; sufficient documentation of test conditions and test results
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
reference to other study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 411 (Subchronic Dermal Toxicity: 90-Day Study)
Deviations:
yes
Remarks:
(additional investigations: urinalysis, weights of brain, ovaries, lungs, thymus; histology of different tissues, renal cell proliferation; no ophthalmoscopic examination)
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Fischer 344
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Kingston
- Age at study initiation: 8 w
- Weight at study initiation: 175 g (males) or 134 g (females)
- Fasting period before study: no
- Housing: individually
- Diet: Agway Prolab RMH 3200 ad libitum
- Water: tap water ad libitum
- Acclimation period: not specified


ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): no data
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): no data
Type of coverage:
semiocclusive
Vehicle:
other: oil-in-water emulsion cream
Details on exposure:
TEST SITE
- Area of exposure: back skin, 4 x 6 cm x cm
- % coverage: not specified
- Type of wrap if used: semiocclusive, Vetrap (3 M Animal Care Products), secured with hypoallergenic tape
- Time intervals for shavings or clipplings: at least once weekly


REMOVAL OF TEST SUBSTANCE
- Washing: area wiped clean with saline-moistened gauze; no attempt was made to remove the HQ cream beyond wiping the skin surface
- Time after start of exposure: 6 h after daily application


TEST MATERIAL
- Amount(s) applied (volume or weight with unit): the amount applied was adjusted on the basis of the weekly mean body weight of each sex
- Concentration: 0, 2, 3.5 and 5%
- Constant volume or concentration used: no


VEHICLE
- Justification for use and choice of vehicle (if other than water): oil-in-water emulsion cream comparable to cream base of commercial cream preparations with HQ
- Amount(s) applied (volume or weight with unit): dosing volume 1.6 mL/kg bw
- Composition: HQ was applied in oil-in-water emulsion creams consisting of water (71.50%), propylene glycol (10.00%) stearic acid (7.00%), light mineral oil (3.00%), isopropyl myristate (2.50%), diethylaminocetyl phosphate (2.00%), cetyl alcohol (1.50%), Oleth-20 (1.50%), Oleth-2 (0.50%), hydroxyethyl cellulose (0.25%), sodium metabisulfite (0.20%) and disodium EDTA (0.05%); the water content was adjusted to compensate for the amount of hydroquinone; pH values were between 5.30 - 5.65


USE OF RESTRAINERS FOR PREVENTING INGESTION: no; however, ingestion was prevented during the exposure by the semiocclusive wrap on the entire torso, and after removing the wrap by application of an Elizabethan collar
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analysis of the squeeze tubes by HPLC before use and at termination of the study indicated that HQ concentrations were within 10% of the nominal for the duration of the study, which is acceptable.
Duration of treatment / exposure:
13 w
Frequency of treatment:
5 d/w, 6 h/d
Remarks:
Doses / Concentrations:
0, 2.0, 3.5, and 5.0% (w/w)
Basis:
other: nominal concentration in test material
Remarks:
Doses / Concentrations:
29.5 +- 2.4, 51.9 +- 4.3, and 73.9 +- 6.3 mg/kg bw/d in males; 43.8 +- 3.8, 77.0 +- 6.9, 109.6 +- 9.8 mg/kg bw/d in females
Basis:
other: based on concentration in test material, average weekly body weight, and the amount applied to the skin
No. of animals per sex per dose:
20 for toxicity study, additionally 5 for cell proliferation and urinalysis study
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: based on preliminary study to determine the highest concentration of HQ that could be tolerated when administered repeatedly under a semi-occlusive wrap using the criteria of the US EPA guidelines for determining a maximum tolerated dose. Concentrations higher than 5% applied under a semi-occlusive wrap produced severe irritation and parakeratosis after 4 weeks of treatment.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least once daily


DETAILED CLINICAL OBSERVATIONS: Yes


DERMAL IRRITATION: Yes
- Time schedule for grading of the erythema and oedema scores according to Draize (1959): daily during weeks 1, 2, 6, 7, 12, and 13


BODY WEIGHT: Yes
- Time schedule for examinations: weekly


FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes


WATER CONSUMPTION: Yes
- Time schedule for examinations: 3 times per week during week 1, 2, 6, 7, 12, and 13


OPHTHALMOSCOPIC EXAMINATION: No


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: 20 per group and sex
- Parameters examined: haemoglobin concentration, haematocrit, erythrocytes, total leukocytes, differential leukocyte count, platelet count, red blood cell indices, prothrombin time; examination of blood smears for cellular morphology


CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at termination of study
- Animals fasted: No data
- How many animals: 20 per group and sex
- Parameters examined: alanine aminotransferase (ALT), sorbitol dehydrogenase (SDH), alkaline phosphatase, trigycerides, cholesterol, total bilirubin, total protein, albumin, albumin/globulin ratio, creatinine, urea nitrogen, sodium, potassium, chloride, calcium, phosphorus, glucose


URINALYSIS: Yes
- Time schedule for collection of urine: w 3, 6, 13
- Metabolism cages used for collection of urine: Yes
- Animals fasted: No data
- How many animals: 5 per group and sex and timepoint
- Parameters examined: acitivities of n-acetyl glucosaminidase (NAG), gamma-glutamyl transpeptidase (GGT), alanine aminopeptidase (AAP) activities, creatinine concentration, osmolality, specific gravity, colour and appearance; microscopic evaluation for blood cells, epithelial cells, bacteria, crystals, and amorphous sediment


NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes

ORGAN WEIGHTS: Yes
liver. kidneys, brain, testes or ovaries, lungs, thymus, adrenal glands

HISTOPATHOLOGY: Yes, high dose and control groups
After fixation in 10% neutral buffered formalin, staining with haematoxylin and eosin microscopic examination of:
samples of skin from treated and untreated areas on the back, bone marrow, kidneys, liver, lymph nodes, spleen, thymus, thyroid gland, Zymbal's gland
Other examinations:
RENAL CELL PROLIFERATION by BrdU incorporation method (Details taken from English et al. (1994) Fund Appl Toxicol 23: 397 - 406)
- Time schedule for examination: w 3, 6, 13
- How many animals: 5 per group and sex and timepoint
- Procedure: At the beginning of week 3, 6 and 13, osmotic pumps, that were loaded with bromodeoxyuridine (20 mg BrdU/mL) prepared in 0.1 M PBS, were implanted surgically, subcutaneously into the mid-dorsal lumbar area. Animals were dosed with HQ cream as usual (application site moved to the flank) and killed 3 days later. Kidneys were fixed in situ by whole-body perfusion with PBS followed by 2% paraformaldehyde and 1% glutaraldehyde in PBS. After 24 h, kidneys and a 2-inch length of duodenum (as experimental control for cell replication) were embedded and sectioned and were stained by immunohistochemically methods for detection of BrdU incorporation.
- Evaluation: The numbers of labelled cells per 500 renal cells were counted for the P1, P2, P3 and "other tubular cells" of the kidney.
Identification of cells on the basis of their morphology and topographic location:
P1 segment: origin at glomerulus and comprising about one half of the pars convoluta; location within the cortical labyrinth adjacent to the glomeruli; morphology tall with intermediate length microvilli
P2: remainder of the pars convoluta and initial part of the pars recta; location within the cortical labyrinth and at the tips of the medullary rays; morpholgy similar to P1 cells except that they were shorter in height with shorter microvilli
P3: remainder of the pars recta; location at the base of the medullary rays and in the outer stripe of the outer medulla; morphology cells with long microvilli and dark cytoplasm
Statistics:
Homogeneity of variances with a Bartlett's test at P<- 0.01; comparison of groups with analysis of variance followed by Duncan's test at P<-0.05
Clinical signs:
no effects observed
Dermal irritation:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY: No HQ-related changes

BODY WEIGHT AND WEIGHT GAIN: No HQ-related changes

FOOD CONSUMPTION: significant decrease in males at 3.5% and 5.0% test concentration compared to control only on day 4 (P<=0.05)
Evaluation: No toxicologically significant effect on food consumption


WATER CONSUMPTION:
Males: for all dose groups within 10% of the values of the control group, both during dosing period on workdays and on exposure-free weekends
Females: water consumption was comparable in all groups; however, in all groups including controls water consumption was 20-30% higher on weekends compared to workdays
Evaluation: No HQ-related changes


DERMAL EFFECTS:
Males:
- Brown discoloration: in all groups including controls appeared within first days of treatment; intensity of discoloration minimal, but exacerbated by HQ-treatment;
- Scaly skin (minimal ichthyosis): in 70-100% of treated males compared to 20% of controls;
- Brown spots: incidences were higher in HQ-treated groups without dose-related trend
Females:
- Brown discoloration: incidence and intensity was slightly higher at 3.5% and 5.0% test concentration compared to other groups, effect appeared within first days of treatment;
- Scaly skin (minimal ichthyosis): in 92-100% of treated females compared to 6% of controls indicating that females may be more sensitive than males- Brown spots: incidences were slightly higher in HQ-treated groups without dose-related trend
All groups: epidermal hyperplasia at application site of all groups, no difference in incidence or severity among groups; no evidence of exogenous ochronosis
Evaluation: no toxicologically significant skin effects


DERMAL IRRITATION:
Concentration-related increase of erythema in HQ-treated groups with highest score of 2 throughout the course of the study, effects abated during treatment-free weekends
Evaluation: no indication that maximal tolerable dose (MTD) was exceeded


HAEMATOLOGY:
Males: significant changes of leukocytes (decrease at 5% HQ), of platelets (increase at 3.5% HQ), and hemoglobin and hematocrit( both increased at 2 and 5% HQ)
Females: no differences among groups
Evaluation: Significant changes of haematogical parameters were within normal biological range for rats and were not considered to be biologically significant.

CLINICAL CHEMISTRY:
Males: significant increases of total protein, ALT and SDH at 5% HQ
Females: no differences among groups
Evaluation: ALT and SDH activities were increased in individual males from all groups including controls. The magnitude of the group mean enzyme acitivities was not clinically relevant. A possible effect in the liver, indicated by increased ALT and SDH, was not reflected in liver weights or treatment-related histological changes in the liver. Thus, significant changes of serum chemistry parameters were not considered to be an adverse or toxicologically meaningful effect of HQ.

URINALYSIS
3 wk data: significant changes of osmolality and specific gravity at 3.5% HQ, however, with increases in males and decreases in females (volume of urine higher in this group)
6 wk data: all male dose-groups sigificant decreases of GGT, AAP, creatinine; females showed sigificant decreases of GGT and AAP at 2% HQ only
13 wk data: males without significant changes; females showed significant increase of osmolality and specific gravity
Evaluation: There was no consistent trends among groups concerning urinary parameters which might indicate kidney damage.


ORGAN WEIGHTS: no significant differences


HISTOPATHOLOGY: NON-NEOPLASTIC
Kidneys:
Lesions observed after 3, 6 and 13 wk: not considered to be HQ-related, lymphocytic infiltration or tubular regeneration was present in occasional animals from all groups.
Terminal sacrifice: renal tubular regeneration in all control animals, in all high-dose males, and in 15/20 high-dose females; lymphocytic infiltration and degeneration of proximal tubule in few control and high-dose rats, incidence and severity not altered by exposure; mineralization of renal tubular epithelium both in controls and high-dose groups; minimal to minor hydropic degeneration of the proximal tubule epithelium more prominent in control rats than in high-dose rats.


OTHER FINDINGS: RENAL CELL PROLIFERATION ASSAY (Labelling index = LI)
3 wk data: males: significant increase of LI in P1 cells at 5% HQ, slight increases in other dose groups and in P2 and P3 cells
6 wk data: no changes in males or females
13 wk data: males without significant changes; females with significant increases of LI of other tubular cells at 3.5 and 5% HQ
Evaluation: There were no changes indicative of sustained cellular proliferation.
Dose descriptor:
NOAEL
Effect level:
73.9 mg/kg bw/day (actual dose received)
Sex:
male
Basis for effect level:
other: No signs of systemic toxicity or nephrotoxicity / highest tested dose
Dose descriptor:
NOAEL
Effect level:
109.6 mg/kg bw/day (actual dose received)
Sex:
female
Basis for effect level:
other: No signs of systemic toxicity or nephrotoxicity / highest tested dose
Critical effects observed:
not specified
Conclusions:
After dermal application of HQ for up to 13 weeks no signs of treatment-related nephrotoxicity or renal cell proliferation were found in F344 rats after exposure to doses of up to 73.9 mg/kg bw/d in male rats, and 109.6 mg/kg bw/d in female rats.
Executive summary:

Subchronic dermal toxicity with emphasis on aspects of nephrotoxicity and renal cell proliferation was investigated in groups of 20 male and 20 female F344 rats with a test protocol similar to OECD Guideline 411. Dermal application of 0, 2.0, 3.5, and 5.0% hydroquinone in an oil-in-water emulsion cream, comparable to cream base of commercial cream preparations with HQ, was performed on 5 d/w, for 6 h/d during 13 w. Ingestion of test substance from the application site was prevented using a semi-occlusive wrap, and during periods without treatment by application of an Elizabethan collar. Resulting dermal doses were 29.5, 51.9, and 73.9 mg/kg bw/d in male rats, and 43.8, 77.0, and 109.6 mg/kg bw/d in female rats applied on a skin surface area of 24 cm2 (dermal doses of ca. 1 to 4 mg/cm2). In additional groups of 5 rats per sex, that had been treated for 3, 6 and 13 w, urinalysis data were collected and renal cell proliferation in different proximal tubule segments (P1, P2, P3 and other cells) was investigated via a BrdU incorporation assay. Dermal irritation was scored by grading of the erythema and oedema scores according to Draize (1959) daily during weeks 1, 2, 6, 7, 12, and 13. There was a concentration-related increase of erythema in HQ-treated groups with highest score of 2 throughout the course of the study. These effects abated during treatment-free weekends. Consequently, there is no indication that the maximal tolerable dose (MTD) was exceeded.

Dermal application of 5% HQ procuded transient minor irritation of the skin but no evidence of exogenous ochronosis. Up to the highest tested dose, there were no clinical signs of intoxication, and no statistically significant or toxicologically relevant changes of body weights, organ weights, of food and water consumption, of haematological and clinical chemical parameters. Treatment-related nephrotoxicity was not indicated from urinalysis data and from kidney histology. There were no changes indicative of sustained cellular proliferation in renal tubuli. Consequently, NOAELs in this subchronic dermal toxicity study were 73.9 mg/kg bw/d in male rats, and 109.6 mg/kg bw/d in female rats.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
3 840 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
There are limitations in the available studies, with either a duration shorter than standard requirements, or with a formulation containing the substance in concentration lower than the standard requirements that does not allow to fully investigate potential adverse effects.

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed

Additional information

ORAL TOXICITY STUDIES


 


In a 14-day range-finding toxicity study mortalities, clinical signs, and reduced body weight gain indicated toxicity in male and female F344 rats at doses of ≥ 500 mg/kg bw/d and in male and female B6C3F1 mice at ≥ 250 mg/kg bw/d by NOAELs were 250 mg/kg bw/d in rats and 125 mg/kg in mice (Supporting study: Kari et al., 1992; NTP, 1989).


 


In a subchronic range finding toxicity study, HQ was applied on 5 days per week for 13 weeks by gavage (25 - 400 mg/kg bw/d to Fischer 344 rats andB6C3F1 mice.


In Fischer 344 rats, mortalities occurred at 400 and 200 mg/kg bw/d. Often, tremors were observed after dosing, and convulsions preceding death. The most common clinical sign was lethargy in rats from 200 mg/kg up. Final body weights were similar to controls in all dose groups of females, whereas in males final body weights were decreased by 8% and 9% at 100 and 200 mg/kg, respectively (no data for food consumption available). In rats, prominent pathological findings were inflammation and epithelial hyperplasia of the forestomach seen at 200 mg/kg (4/10 males, 1/10 females), and toxic nephropathy, characterized by tubular cell degeneration and regeneration in the renal cortex, seen at 200 mg/kg (7/10 males, 6/10 females), and at 100 mg/kg (1/10 females).


In B6C3F1 mice, mortalities occurred at 400 and 200 mg/kg bw/d. Often, tremors were observed after dosing and convulsions preceding death. The most common clinical sign was lethargy in male mice of all dose groups (from 25 mg/kg up), and female mice from 100 mg/kg up.In mice,ulceration, inflammation, or epithelial hyperplasia in the forestomach were found at 400 mg/kg (3/10 males, 2/10 females), and at 200 mg/kg(1/10 females).


Based on negligible effects at lower doses, the NOAELs are 50 mg/kg bw/d and the LOAELs are 100 mg/kg bw/d, both in rats and mice. The findings in the forestomach are indicative of a local irritant action in the forestomach due to bolus application via gavage. At 100 mg/kg, body weight reduction and toxic nephropathy is the critical effect in the Fischer 344 strain of rats (for further discussion see below: Assessment of specific target organ toxicity). In B6C3F1 mice, lethargy observed in both sexes at 100 mg/kg is considered as a clinical sign indicating an adverse effect on the central nervous system.The observation of lethargy only in males at the low dose range of 25 and 50 mg/kg bw/d was not reproducible in the chronic study at doses of 50 and 100 mg/kg bw/d and thus is considered to be of no toxicological relevance.At the higher dose of 200 mg/kg effects are much more severe with mortality of 30% of female rats and 20% of male mice(Key study: Kari et al., 1992; NTP, 1989).


 


There was no evidence of subchronic neurotoxicity after oral exposure of groups of 10 male and 10 female Sprague-Dawley rats to doses of 20, 64, and 200 mg HQ/kg bw/d by gavage for 13 weeks on 5 d/was assessed by functional observation battery examinations, quantitative grip strength measurements, brain weights, or neuropathology examinations (Study protocol similar to OECD Guideline 424, Neurotoxicity study in rodents). Effects were restricted to clinical signs indicating responses of the central nervous system as depression of general motor activity at 200 mg/kg, and appearance of tremors after dosing of HQ at 64 and 200 mg/kg bw/d. No renal changes were evident in this rat strain given 200 mg HQ/kg bw/d, a dose level that was nephrotoxic to F344 rats.Under the conditions of this study, in Sprague-Dawley rats the LOAEL was 64 mg/kg bw/d based on clinical signs indicating an adverse effect on the central nervous system and the NOAEL was 20 mg/kg bw/d (Key study: Topping et al., 1988, 1991, 2007).


 


HQ was administered to groups of 65 male and 65 female F344 rats or B6C3F1 mice for 103 weeks on 5 days per week via gavage (vehicle water) at doses of 0, 25, and 50 mg/kg bw/d for rats or 0, 50 and 100 mg/kg bw/d for mice. The study protocol was similar to OECD Guideline 453 with several minor deviations. There were only 2 instead of 3 dose groups, and food consumption and urinalysis were not examined. Hematology and clinical chemistry examinations were performed at a single timepoint after 65 w of dosing in subgroups of 10 animals of each sex and dose. Sacrifice for gross and histopathological examination was at about 66 w (N=10) and 104-105 w (N=55) after start of dosing.Both in rats and mice, no significant differences in survival and no compound-related clinical signs were reported.


In male F344 rats, toxicity was indicated at 50 mg/kg by decreased body weights (5-9% between week 73-93, 10-13% between week 94-104) and relative kidney weights were increased after 65 and 103 w of administration. All male rats of all groups (dosed and vehicle control) showed chronic progressive nephropathy (CPN), which is a common effect in ageing F344 rats. Severity of CPN was increased at the 66-week interim sacrifice in both dose groups and at the terminal sacrifice in the high-dose group (gradings of CPN: 5/55 mild, 15/55 moderate, 32/55 marked compared to respective gradings of 12/55, 26/55, 12/55 in the vehicle control). In male F344 rats, the LOAEL for non-neoplastic chronic toxicity was 50 mg/kg bw/d, based on body weight reduction and increased severity of CPN as an effect specific for this strain of rats (for further discussionsee below: Assessment of specific target organ toxicity), and the NOAEL was 25 mg/kg bw/d.. At 50 mg/kg bw/d, a significant increase of renal tubuli adenoma was observed in male rats (for detailed discussion of neoplastic histopathology see Section 7.7).Female F344 rats were found to be less sensitive as indicated by a NOAEL of 50 mg/kg bw/d for non-neoplastic toxicity. At 25 and 50 mg/kg bw/d, statistically significant increased incidences of mononuclear cell leukaemia were observed in female rats (for detailed discussion see Section 7.7). (Key study: Kari et al., 1992; NTP, 1989).


In B6C3F1 mice, toxicity was indicated at 100 mg/kg by decreased body weights in females (5-8% lower between week 20-44, 10-14% lower between week 45-104). Significant increases of relative liver weights were found at 100 mg/kg in both sexes after 65 and 103 w of administration, and at 50 mg/kg in males after 103 w. Centrilobular fatty change and cytomegaly were observed in male mice at the 65-week interim sacrifice but not at the terminal sacrifice. As necropsies at these timepoints were performed within 24 hrs and 14 days after the last dosing, respectively, fatty change and cytomegaly seem to be reversible after cessation of hydroquinone administration. Additionally, at 100 mg/kg findings in livers of male mice after 105 w were characterized by increased incidences of anisokaryosis (variation in size of hepatocyte nuclei), syncitial alteration (hepatocytes with more than 5 nuclei per cell), and foci of cellular alteration (basophilic foci) compared to incidences in vehicle controls. The LOAEL for chronic toxicity was 50 mg/kg bw/d both in male B6C3F1 mice based on liver weight effects and liver histopathology at the 65-week interim sacrifice, and in female mice based on increased incidences of hepatocellular adenomas and of hepatocellular adenomas or carcinomas combined(detailed evaluation of neoplastic histopathology in Section 7.7) (Key study: Kari et al., 1992; NTP, 1989).


 


A reevaluation of the renal histopathology findings of certain dose groups of the NTP cancer bioassay with Fischer 344 rats (all males with terminal sacrifice, and the control groups and 50 mg/kg groups of males with 66 week interim sacrifice and of females with terminal sacrifice) was performed with emphasis on evaluation of incidences, severity and location of chronic progressive nephropathy (CPN) and renal proliferative lesions (for details see Section 7.7). In this detailed reevaluation of the histological sections there was no indication of alpha-2µ-globulin nephropathy or a direct nephrotoxic effect of HQ in high dosed male rats (50 mg/kg bw) after chronic exposure. However, HQ exacerbated the spontaneously occurring chronic progressive nephropathy (CPN) and additionally exerted a stimulatory effect on the proliferative potential of the advanced stages of CPN at both 25 and 50 mg/kg (for details see Section 7.7). Of the rats surviving into the last 10 w of the study 49% had endstage CPN and 40% had severe CPN. Principally, these renal effects are of little relevance for humans, as CPN is a rodent-specific spontaneous renal disease (Key study:Hard et al., 1997).


 


 


DERMAL TOXICITY STUDIES


 


Subchronic dermal toxicity with emphasis on aspects of nephrotoxicity and renal cell proliferation was investigated in groups of 20 male and 20 female F344 rats with a test protocol similar to OECD Guideline 411. Dermal application of 0, 2.0, 3.5, and 5.0% HQ in an oil-in-water emulsion cream, comparable to cream base of commercial cream preparations with HQ, was performed on 5 d/w, for 6 h/d during 13 w. Resulting dermal doses were 29.5, 51.9, and 73.9 mg/kg bw/d in male rats, and 43.8, 77.0, and 109.6 mg/kg bw/d in female ratsapplied on a skin surface area of 24 cm2(dermal doses of ca. 1 to 4 mg/cm2). Dermal application of 5% HQ produced transient minor irritation of the skin (no indication that the maximal tolerable dose (MTD) was exceeded) but no evidence of exogenous ochronosis. Up to the highest tested dose, there were no clinical signs of intoxication, and no statistically significant or toxicologically relevant changes of body weights, organ weights, of food and water consumption, of haematological and clinical chemical parameters. Treatment-related nephrotoxicity was not indicated from urinalysis data and from kidney histology. There were no changes indicative of sustained renal cell proliferation in different proximal tubule segments (P1, P2, P3 and other cells) investigated via a BrdU incorporation assay. Consequently, NOAELs in this subchronic dermal toxicity study were at least 73.9 mg/kg bw/d in male rats, and 109.6 mg/kg bw/d in female rats (no higher doses tested) (Key study: David et al., 1998).


 


In a 14 day range finding dermal toxicity study with F344 rats (12 open applications on 5 d/w of 240 to 3840 mg/kg bw/d in 95% ethanol as vehicle), no toxicologically significant effects were observed up to the highest dose, so that 3840 mg/kg bw/d is the NOAEL for male and female F344 rats. After equal exposure of male and female B6C3F1 mice to doses of 300 to 4800 mg/kg bw/d, there was no indication of toxicity from survival, clinical signs, body weights and gross necropsy up to 4800 mg/kg bw/d, so the NOEL for B6C3F1 mice is 4800 mg/kg bw/d. A preliminary qualitative dermal absorption study in both species indicated systemic availability of unknown amounts of HQ, excreted as glucuronide and sulfate conjugates in urine (Key studies: NTP, 1989).


 


 


INHALATION TOXICITY STUDIES


 


There are no experimental inhalation studies with test animals available. All the information available was considered to assess the potential concerns for the inhalation route and conduct the risk assessment in humans.


 


 


HUMAN EXPERIENCE


 


From health surveys of workmen, exposed to benzoquinone (BQ) vapors and HQ dust during the manufacture of HQ, there was no indication of systemic effects from annual physical examinations and hematological parameters (Sterner et al., 1947). The absence of effects on erythrocyte numbers, hematocrit and hemoglobin concentrations in workers exposed to higher levels of HQ dust before 1946, with reported air concentrations of up to 35 mg/m3, shows that the blood system is not a possible target of an adverse action of HQ in humans. In contrast, local eye injury was found as the prominent effect in these HQ-exposed workers (Key study: Anderson, 1947; Sterner et al., 1947; Anderson and Oglesby, 1958; for details see IUCLID Section 7.10.1 and 7.3).


 


A retrospective study was performed in a well-characterized 1942-1990 cohort of 879 workers (858 men and 21 women) employed principally in the manufacture and use of HQ at a large chemical plant comprising 22,895 person-years of exposure during the 50-year observation period. Mean duration of tenure in a HQ environment was 13.7 years, mean follow-up from first exposure was 26.8 yrs; 2205 person-years (10%) were represented by subjects with ages of 65 and older. Average exposure concentrations, 1949-1990, ranged from 0.1 to 6.0 mg/m3for HQ dust and less than 0.1 to 0.3 mg/m3for benzoquinone vapour. 20% of cohort members had presumably experienced higher exposures to HQ dust, e.g. up to 35 mg/m3in the packaging area, at the period up to 1949, before measures to reduce exposure had been installed. Causes of mortality were followed up to 1991 and were compared with vital statistics from the general population of the State of Tennessee, and to an occupational reference group of 30,000 hourly wage personnel employed at Kodak's Rochester facilities from 1964 to 1992, few of whom had potential HQ exposure.


There was no evidence of excess mortality in the investigated cohort. In contrast, the number of deaths from all causes of death, and from all forms of malignant cancer was statistically significantly below expectation compared with both the general population and the occupational controls. There was also a lower than expected death rate from circulatory diseases. Additionally, there were fewer than expected deaths from respiratory cancer, non-malignant pulmonary diseases, and genitourinary system illness, including nephrotoxicity(Key study: Pifer et al., 1995; for details see IUCLID Section 7.10.2). Consequently, from this study there are no indications that HQ affects the kidneys, the livers, or the blood system of humans with occupational exposures to HQ, even at the elevated exposure levels of the early years of HQ production before 1950.


 


 


ASSESSMENT OF SPECIFIC TARGET ORGAN TOXICITY AND OF SPECIES-SPECIFITY OF EFFECTS


 


Specific investigations of the mechanisms of the nephrotoxic action of HQ (for details see IUCLID Section 7.9.3) show a strain- and species-specific susceptibility of male F344 rats compared to other strains of rats, to mice and other species. Nephrotoxic effects of HQ were found after acute and subchronic oral exposure of F344 rats with LOAELs of 200 and 25 mg/kg bw/d, respectively. Renal lesions are characterized by tubular cell necrosis followed by cell regeneration. HQ-induced renal cell proliferation, presumably is an attempt to compensate for proximal tubular cell lossrather than representing a direct mitogenic effect of HQ.The nephrotoxic action of HQ is dependent on the formation of metabolites that require processing by GGT, that are glutathionyl conjugates of HQ. 2,3,5-(tris-glutathion-S-yl)HQ displayed the highest nephrotoxic potency (about 600-fold that of HQ).Bioactivation steps predominate in rat liver cells leading to higher body burdens of higher substituted GSH HQ conjugates, which are considered to be the key nephrotoxic metabolites, whereas deactivation steps predominate in human liver cells. F344 rats were shown to be especially prone to the formation of these metabolites compared to other strains or animal species. In contrast, humans have an increased capacity for hepatic detoxification, especially for the formation of HQ-glucuronide and the mercapturic acid conjugate (N-AcCysS-HQ), limiting exposure of the human kidney to nephrotoxic metabolites.


Consequently, renal toxic and renal neoplastic effects observed in Fischer 344 rats (Kari et al., 1992; NTP, 1989) but not in Sprague-Dawley rats (Hydroquinone Program Panel, 1998; Topping et al., 1991, 2007) or B6C3F1 mice (Kari et al., 1992; NTP, 1989)are considered not be biologically significant for human exposure (see also discussion in Section 7.7).


A possible hepatotoxic action, indicated by histopathological changes only in male mice after chronic exposure but not in female mice or in rats of either sex (Kari et al., 1992; NTP, 1989), is considered to be of questionable biological significance for human exposure. This is supported by data from a retrospective study with workers exposed to HQ by inhalation (see above: Sterner et al., 1947; Pifer et al., 1995) which gave no indication for specific target organs (especially of the kidneys, livers, or the blood system) even at elevated exposure levels in the early years of HQ production before 1950 with maximum air concentrations of up to 35 mg/m3. At these workplaces, a 8-hour exposure under a condition of light physical activity with an estimated respiratory volume of 10 m3, would account to a maximal dose of 350 mg per person and day or ca. 5 mg/kg bw/d (assumption of 100% absorption from the respiratory tract).


 


 


OVERALL EVALUATION


 


Oral exposure

 


Table: Overview on NOAELS and LOAELsbased on non-neoplastic and neoplastic findings derived from the key studies with repeated oral application of HQ, including critical acute CNS effects.


 























































































Species


Strain


Sex



Dose (mg/kg bw/d)


Application


Duration



NOAEL


(mg/kg bw/d)



LOAEL


(mg/kg bw/d)



Basis for LOAEL



Acute CNS effects


(effects observed rapidly after dosing)



Reference




F344


rat


m



0, 25 or 50


gavage


103 w, 5 d/w



25



50



body weights ↓; severity of chronic progressive nephropathy ↑; renal tubuli adenomas ↑



No indications for CNS effects.


No neurobehavioral examination



Kari et al. (1992); NTP (1989)



F344


Rat


f



0, 25 or 50


Gavage


103 w, 5 d/w



-



25



mononuclear leukaemia ↑



No indications for CNS effects.


No neurobehavioral examination



Kari et al. (1992); NTP (1989)



B6C3F1


mouse


m / f



0, 50 or 100


gavage


103 w, 5 d/w



-



50



m: relative liver weight ↑; non-neoplastic liver changes only at 65 week interim sacrifice


f: combined liver adenomas / carcinomas ↑



No indications for CNS effects.


No neurobehavioral examination



Kari et al. (1992); NTP (1989)



Sprague-Dawley


rat


m / f



0, 20, 64 or 200


gavage


13 w, 5 d/w



20



64



Tremors



>= 64 mg: tremors, depression


200 mg: behavioural changes


Neurobehavioral examination ()



Topping (1988); Topping et al. (1991, 2007)[1]



F344


rat


m / f



0, 25, 50, 100, 200 or 400


gavage


13 w, 5 d/w



50



100



m: body weights ↓;
f: nephrotoxicity ↑



>= 200 mg: lethargy, tremors and/or convulsions.


No neurobehavioral examination



Kari et al. (1992); NTP (1989)



B6C3F1


mouse


m / f



0, 25, 50, 100, 200 or 400


gavage


13 w, 5 d/w



50



100



Lethargy



≤ 100 mg: lethargy in all dosed males and in females only at 100 mg[2]


>= 200 mg: lethargy, tremors and/or convulsions.


No neurobehavioral examination



Kari et al. (1992); NTP (1989)



F344


rat


m / f



0, 63, 125, 250, 500or 1000


gavage


14 d, 5 d/w



250



500



mortality ↑



>= 500 mg: tremors


No neurobehavioral examination



Kari et al. (1992); NTP (1989)



B6C3F1


mouse


m / f



0, 31, 63, 125, 250or 500


gavage


14 d, 5 d/w



125



250



mortality ↑



>= 250 mg: tremors and/or convulsions


No neurobehavioral examination



Kari et al. (1992); NTP (1989)





[1]Study similar to OECD Guideline 424 (Neurotoxicity study in rodents). With regard to the requests of OECD Guideline 408 investigations focussed on neurotoxicity and nephrotoxicity




[2]The observation of lethargy only in males at the low dose range of 25 and 50 mg was not reproducible in the chronic study at doses of 50 and 100 mg and thus is considered to be of no toxicological relevance




 


 


As the overview in the Table shows, both the NOAELs and LOAELs that can be derived from subchronic and chronic toxicity studies including both non-neoplastic and neoplastic findings (for details see Section 7.7) are at similar ranges each, with values ranging generally from 20 to 50 mg/kg bw/d for the NOAEL and 50 to 100 mg/kg bw/d for the LOAEL. The observation of a significant increase of mononuclear leukaemia in female F344 rats at 25 mg/kg bw/d (Kari et al., 1992; NTP, 1989) is a questionable effect based on the high spontaneous incidence of these lesion (for detailed discussion see Section 7.7) and is disregarded for deriving a NOAEL relevant for human exposure as no other adverse effects were observed in dosed female rats up to 50 mg/kg bw/d.


 


Based on the overview of 5 repeated dose toxicity studies by gavage covering 2-week subacute, 13-week subchronic to 103-week chronic studies, effects on the CNS are consistent in rats and mice and occur rapidly after administration and are transient (table 1). In rats, the range of concentrations tested in all studies showed CNS effects at 64 mg/kg/day (13-weeks, rats) and higher, in male and female Sprague-Dawley rats (Topping et al., 1988, 2007, RSS in section 7.9.1), while no CNS effects were observed at 50 mg/kg/day and below, even in chronic studies combined with carcinogenicity (F344 rats, B6C3F1 mice) (Kari et al., 1992; NTP, 1989). The only exception is the report of mild transient tremors in only one Sprague-Dawley male at 50 mg/kg/day in a 2-generation reproductive toxicity study (Blacker et al., section 7.8.1), indicating that this rat strain may display a slightly higher susceptibility. The data of the subchronic and chronic toxicity studies (Kari et al., 1992; NTP, 1989) do not indicate a higher sensitivity of mice compared to rats with regard to toxicologically relevant adverse effects. Based on a weight of evidence considering all the available studies, it can be inferred that there are no CNS effects at 25 mg/kg/day or below. As there was no specific influence of treatment duration, the lowest NOAEL of 20 mg/kg/day from a 13-week study, focused on neurotoxicity in rats, was considered relevant also for long-term exposure and was used as a NOAEL for CNS effects and a possible point of departure for the risk assessment. Benchmark dose modelling from this dataset (see section 7 - DNEL) determined a BMDL10 of 61.2 mg/kg/day.


 


 


Inhalation toxicity


 


There is no repeated dose toxicity study available by the inhalation route. Based on the physico-chemical properties of HQ (vapour pressure, particle size, water solubility), there is a low likelihood that the solid particles reach the deep lung in a significant amount. As a conservative approach, and assuming the absorption is similar by the inhalation route as compared to the oral route, it is usually acceptable to conduct a risk assessment using the oral data and identified adverse effects and route-to route extrapolation to derive a NOAEC for the inhalation route. Based on the discussion above for the oral route, the relevant adverse effect identified in repeated dose toxicity studies considered for the risk assessment is the CNS effects.


 


Dermal toxicity


 


Based on the overall evidence, the NOAEL is 3840 mg/kg bw/d in the Fischer 344 rat for a 14-day dermal exposure. Consequently, the dermal NOAEL is about 15-fold the oral NOAEL of 250 mg/kg after 14-day gavage exposure F344 rats (see Oral toxicity studies; both studies performed as range-finding toxicity studies: NTP, 1989). The difference in dermal and oral repeated dose toxicity is supported by the known low dermal absorption of HQ which amounts to about 10% in comparison to about 100% bioavailability by the oral route. The high dermal NOAEL observed in rats is confirmed by an even higher dermal NOEL of 4800 mg/kg bw/d in the mouse as second species in the 14-day range finding study (NTP, 1989).



Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
Because of the dust explosiveness properties shown with the product as manufactured (particle size > 100µm) as well as under standardised conditions (particle size > 63 µm), producing a micronised sample with the particle size distribution required by the test guidelines for a repeated dose inhalation study was not considered feasible under safe conditions. (Communication with ECHA and MSCA).

Justification for selection of repeated dose toxicity inhalation - local effects endpoint:
Because of the dust explosiveness properties shown with the product as manufactured (particle size > 100µm) as well as under standardised conditions (particle size > 63 µm), producing a micronised sample with the particle size distribution as required by the test guidelines for a repeated dose inhalation study was not considered feasible under safe conditions (Communication with ECHA and MSCA). There are no specific inhalation data or report of local effects on the respiratory tract under the current workplace conditions. Instead, there is a history of eye effects (brown staining of the conjunctiva or cornea, and/or corneal opacities and alterations) reported in workers who had been repeatedly exposed over the years (usually more than 5 years) to very high airborne concentrations of HQ dusts and benzoquinone vapours (section 7.10.1 – Exposure related observations in humans/Health surveillance data, and endpoint summary section 7.5). There are no specific documentation regarding concurrent local respiratory effects but early publications indicated the presence of respiratory tract irritation at the very high airborne concentrations (as high as 20-35 mg/m3) experienced prior to industrial hygiene improvements (Oglesby et al., 1947). These airborne concentrations are no longer relevant to the current workplace conditions since the implementation of pragmatic TWA in many European countries, originating from the ACGIH value set up in the USA to protect against the adverse ocular damage (Oglesby et al., 1947, Pifer et al. 1995) with values all below 2 mg/m3 and most frequently 1 mg/m3 and below. Despite the absence of a clear dose-response and mode of action for the ocular effects, control of workplace airborne concentrations to low levels has also been preventing any potential effects on the respiratory tract.

Justification for selection of repeated dose toxicity dermal - systemic effects endpoint:
No specific findings in a 14-day repeated dose toxicity studies in rats and mice (NTP studies). In addition, no systemic effects were observed in a 90-day dermal studies in rats exposed to a cream formulation containing 0.5% HQ (i.e. approx. 74 mg/kg/day daily dose for males).

Repeated dose toxicity: via oral route - systemic effects (target organ) neurologic: central nervous system; urogenital: kidneys

Justification for classification or non-classification