Sodium benzoate

Repeated dose toxicity: dermal

Administrative data

Endpoint:

short-term repeated dose toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 14 November to 5 December 1979

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study run to a reliable method and to GLP.

Justification for type of information:

See Read Across Justification document in Section 13.2 of IUCLID

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

read-across source

Cross-reference 2

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

short-term repeated dose toxicity: dermal

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

From 14 November to 5 December 1979

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study run to a reliable method and to GLP.

Justification for type of information:

See Read Across Justification document in Section 13.2 of IUCLID

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

EPA OPP 82-2 (Repeated Dose Dermal Toxicity -21/28 Days)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Species:

rabbit

Strain:

New Zealand White

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Langshaw Farms, Augusta, Michigan
- Age at study initiation:
- Weight at study initiation: 2127 to 2891 grams
- Fasting period before study:
- Housing: Individually housed in hanging wire-mesh cages.
- Diet (e.g. ad libitum): Purina Certified Rabbit Chow, ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 13 days

ENVIRONMENTAL CONDITIONS
- Temperature (°F): 70-76 °F
- Humidity (%): 44-57%
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): 12 hours light and 12 hours dark

IN-LIFE DATES: From: 14 November 1979 To: 5 December 1979

Type of coverage:

semiocclusive

Vehicle:

physiological saline

Remarks:

0.9%

Details on exposure:

TEST SITE
- Area of exposure: 100 cm2
- % coverage: 23%, 35% and 100%
- Type of wrap if used: Each site was covered with a 10 x 10-centimeter gauze pad with adhesive tape borders. The entire area was then wrapped with gauze bandaging and Elastoplast tape.
- Time intervals for shavings or clipplings: twice each week prior to test article administration during the three week study period

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The test areas were washed with tepid IRDC tap water and dried with disposable paper towels.
- Time after start of exposure: 6 hours

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 100, 500 and 2500 mg/kg bw
- For solids, paste formed: no

VEHICLE
not applicable

USE OF RESTRAINERS FOR PREVENTING INGESTION: no

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

6 hours, test substance moistened with physiological saline

Frequency of treatment:

Once a day, 5 days per week, for 3 consecutive weeks

Remarks:

Doses / Concentrations:
100, 500, 2500 mg/kg
Basis:
nominal per unit body weight

No. of animals per sex per dose:

4 males and 4 females per dose

Control animals:

yes

Details on study design:

None stated

Positive control:

None stated

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once daily during the 21-day study period
- Cage side observations checked in table [No.?] were included.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once daily

DERMAL IRRITATION (if dermal study): Yes
- Time schedule for examinations: once daily

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were obtained during the pretest period, prior to study initiation and at weekly intervals during the 21-day study period

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data

WATER CONSUMPTION: No data

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: once during the pretest period and at 21 days of study
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: all animals
- Parameters: hematocrit, hemoglobin, erythrocyte count, total leucocyte count, differential leucocyte count, platelets, MCH, MCV and MCHC

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Once during the pretest period and at 21 days of study.
- Animals fasted: Yes / No / No data
- How many animals: all animals
- Parameters:glucose, blood urea nitrogen, lkaline phosphatase, serum glutamic pyruvic transaminase, and serum glutamic oxalacetic transaminase

URINALYSIS: No data

NEUROBEHAVIOURAL EXAMINATION: No data

OTHER:

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
masses and lesions, untreated skin, treated skin, kidney, lungs, pituitary, sciatic nerve, muscle, spleen, urinary bladder, thymus, lumbar spinal cord, pancreas, stomach, duodenum, testes, thyroid, femur, colon, mesenteric lymph nodes, mediastinal lymph nodes, ovaries, parathyroid, liver, gall bladder, adrenals, heart, brain
HISTOPATHOLOGY: Yes
spleen, pancreas, stomach, duodenum, colon, lymph nodes, urinary bladder, heart, lung, brain (3 sections), treated skin (3 sections), untreated skin (3 sections), liver, kidney, gall bladder, adrenals, spinal cord, gonads, nerve (with muscle), bone marrow, thyroid (parathyroid), pituitary, thymus

Other examinations:

None stated

Statistics:

All statistical analyses compared the treatment groups with the control group, by sex.
Body weights (day 21), hematological and biochemical parameters (pretest and day 21) and absolute and relative organ weights (terminal sacrifice) were compared by analysis of variance (one-way classification), Bartlett’s test for homogeneity of variances and the appropriate t-test (for equal or unequal variances) as described by Steel and Torrie using Dunnett’s multiple comparison tables to judge significance of differences.

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):

not specified

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

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

Histopathological findings: neoplastic:

not specified

Details on results:

CLINICAL SIGNS AND MORTALITY
In the control group, a few rabbits exhibited signs of anorexia and purulent nasal discharge. One rabbit also exhibited diarrhea.
In the test groups, a few rabbits exhibited signs of anorexia, purulent nasal discharge, clear nasal discharge, soft stool, diarrhea and ocular discharge.
None of the rabbits died during the 21-day study period.

DERMAL IRRITATION
Very slight dermal irritation was noted for one rabbit at the 2500 mg/kg dosage level.

BODY WEIGHT AND WEIGHT GAIN
There were no statistical differences found between the control and test groups.

HAEMATOLOGY
There were no treatment-related differences noted in the hematology data in this study.

CLINICAL CHEMISTRY
Although there was one statistically significant difference noted, this was not considered physiologically meaningful or treatment-related. The mid-dose female SGOT mean was significantly lower than the control mean, however, there were only four females in each group and the difference was only 5 IU/1. Therefore the statistically significant difference observed does not have any biological significance.

ORGAN WEIGHTS
The comparison of the control and treated group means for body and organ weights did not show any important differences.

GROSS PATHOLOGY
The skin from the application site was described as erythematous in one, two and seven animals of groups I, III and IV, respectively. The lungs presented foci or areas of congestion that were considered unrelated to the application of the test compound.

HISTOPATHOLOGY: NON-NEOPLASTIC
The skin from the application site presented mainly infiltration of inflammatory cells in the dermis with slight severity in both control and treated groups. Slight hyperkeratosis was observed in three males and four females in group IV. These lesions were very slight and represent a minimal reaction to the test compound.
The foci of myocarditis found in some treated animals were considered unrelated to the application of the test compound since they are observed from time to time in control animals.

Dose descriptor:

NOAEL

Effect level:

> 2 500 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: One female in the 2500 mg/kg test group exhibited dermal irritation during the 21-day study period.

Critical effects observed:

not specified

Conclusions:

The NOAEL was considered to be > 2500 mg/kg. Very slight dermal irritation was noted for one rabbit at this dosage level. No compound related changes were seen in general behavior and appearance, body weight, clinical laboratory tests, organ weights or survival.

Executive summary:

In a dermal study in New Zealand White rabbits, the test substance was applied 5 days a week for 3 weeks at dosage levels of 100, 500 and 2500 mg/kg. Four male and four female rabbits were used in each treated group and in a control group. 0.9% physiological saline was used as the control substane. The rabbits were observed daily for signs of dermal irritation and changes in general behavior and appearance. Individual body weights were recorded weekly Hematologic and biochemical studies were conducted once in the pretest period and at 21 days of study.

The NOAEL was considered to be > 2500 mg/kg. Very slight dermal irritation was noted for one rabbit at this dosage level. No compound related changes were seen in general behavior and appearance, body weight, clinical laboratory tests, organ weights or survival.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1981

Report date:

1981

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EPA OPP 82-2 (Repeated Dose Dermal Toxicity -21/28 Days)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Benzoic acid

EC Number:

200-618-2

EC Name:

Benzoic acid

Cas Number:

65-85-0

Molecular formula:

C7H6O2

IUPAC Name:

Benzoic acid

Details on test material:

Batch No.: 52829055
Purity: 99.5%

Test animals

Species:

rabbit

Strain:

New Zealand White

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Langshaw Farms, Augusta, Michigan
- Age at study initiation:
- Weight at study initiation: 2127 to 2891 grams
- Fasting period before study:
- Housing: Individually housed in hanging wire-mesh cages.
- Diet (e.g. ad libitum): Purina Certified Rabbit Chow, ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 13 days

ENVIRONMENTAL CONDITIONS
- Temperature (°F): 70-76 °F
- Humidity (%): 44-57%
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): 12 hours light and 12 hours dark

IN-LIFE DATES: From: 14 November 1979 To: 5 December 1979

Administration / exposure

Type of coverage:

semiocclusive

Vehicle:

physiological saline

Remarks:

0.9%

Details on exposure:

TEST SITE
- Area of exposure: 100 cm2
- % coverage: 23%, 35% and 100%
- Type of wrap if used: Each site was covered with a 10 x 10-centimeter gauze pad with adhesive tape borders. The entire area was then wrapped with gauze bandaging and Elastoplast tape.
- Time intervals for shavings or clipplings: twice each week prior to test article administration during the three week study period

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The test areas were washed with tepid IRDC tap water and dried with disposable paper towels.
- Time after start of exposure: 6 hours

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 100, 500 and 2500 mg/kg bw
- For solids, paste formed: no

VEHICLE
not applicable

USE OF RESTRAINERS FOR PREVENTING INGESTION: no

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

6 hours, test substance moistened with physiological saline

Frequency of treatment:

Once a day, 5 days per week, for 3 consecutive weeks

Doses / concentrations

Remarks:

Doses / Concentrations:
100, 500, 2500 mg/kg
Basis:
nominal per unit body weight

No. of animals per sex per dose:

4 males and 4 females per dose

Control animals:

yes

Details on study design:

None stated

Positive control:

None stated

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once daily during the 21-day study period
- Cage side observations checked in table [No.?] were included.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once daily

DERMAL IRRITATION (if dermal study): Yes
- Time schedule for examinations: once daily

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were obtained during the pretest period, prior to study initiation and at weekly intervals during the 21-day study period

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data

WATER CONSUMPTION: No data

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: once during the pretest period and at 21 days of study
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: all animals
- Parameters: hematocrit, hemoglobin, erythrocyte count, total leucocyte count, differential leucocyte count, platelets, MCH, MCV and MCHC

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Once during the pretest period and at 21 days of study.
- Animals fasted: Yes / No / No data
- How many animals: all animals
- Parameters:glucose, blood urea nitrogen, lkaline phosphatase, serum glutamic pyruvic transaminase, and serum glutamic oxalacetic transaminase

URINALYSIS: No data

NEUROBEHAVIOURAL EXAMINATION: No data

OTHER:

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
masses and lesions, untreated skin, treated skin, kidney, lungs, pituitary, sciatic nerve, muscle, spleen, urinary bladder, thymus, lumbar spinal cord, pancreas, stomach, duodenum, testes, thyroid, femur, colon, mesenteric lymph nodes, mediastinal lymph nodes, ovaries, parathyroid, liver, gall bladder, adrenals, heart, brain
HISTOPATHOLOGY: Yes
spleen, pancreas, stomach, duodenum, colon, lymph nodes, urinary bladder, heart, lung, brain (3 sections), treated skin (3 sections), untreated skin (3 sections), liver, kidney, gall bladder, adrenals, spinal cord, gonads, nerve (with muscle), bone marrow, thyroid (parathyroid), pituitary, thymus

Other examinations:

None stated

Statistics:

All statistical analyses compared the treatment groups with the control group, by sex.
Body weights (day 21), hematological and biochemical parameters (pretest and day 21) and absolute and relative organ weights (terminal sacrifice) were compared by analysis of variance (one-way classification), Bartlett’s test for homogeneity of variances and the appropriate t-test (for equal or unequal variances) as described by Steel and Torrie using Dunnett’s multiple comparison tables to judge significance of differences.

Results and discussion

Results of examinations

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):

not specified

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

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

Histopathological findings: neoplastic:

not specified

Details on results:

CLINICAL SIGNS AND MORTALITY
In the control group, a few rabbits exhibited signs of anorexia and purulent nasal discharge. One rabbit also exhibited diarrhea.
In the test groups, a few rabbits exhibited signs of anorexia, purulent nasal discharge, clear nasal discharge, soft stool, diarrhea and ocular discharge.
None of the rabbits died during the 21-day study period.

DERMAL IRRITATION
Very slight dermal irritation was noted for one rabbit at the 2500 mg/kg dosage level.

BODY WEIGHT AND WEIGHT GAIN
There were no statistical differences found between the control and test groups.

HAEMATOLOGY
There were no treatment-related differences noted in the hematology data in this study.

CLINICAL CHEMISTRY
Although there was one statistically significant difference noted, this was not considered physiologically meaningful or treatment-related. The mid-dose female SGOT mean was significantly lower than the control mean, however, there were only four females in each group and the difference was only 5 IU/1. Therefore the statistically significant difference observed does not have any biological significance.

ORGAN WEIGHTS
The comparison of the control and treated group means for body and organ weights did not show any important differences.

GROSS PATHOLOGY
The skin from the application site was described as erythematous in one, two and seven animals of groups I, III and IV, respectively. The lungs presented foci or areas of congestion that were considered unrelated to the application of the test compound.

HISTOPATHOLOGY: NON-NEOPLASTIC
The skin from the application site presented mainly infiltration of inflammatory cells in the dermis with slight severity in both control and treated groups. Slight hyperkeratosis was observed in three males and four females in group IV. These lesions were very slight and represent a minimal reaction to the test compound.
The foci of myocarditis found in some treated animals were considered unrelated to the application of the test compound since they are observed from time to time in control animals.

Effect levels

Dose descriptor:

NOAEL

Effect level:

> 2 500 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: One female in the 2500 mg/kg test group exhibited dermal irritation during the 21-day study period.

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

The NOAEL was considered to be > 2500 mg/kg. Very slight dermal irritation was noted for one rabbit at this dosage level. No compound related changes were seen in general behavior and appearance, body weight, clinical laboratory tests, organ weights or survival.

Executive summary:

In a dermal study in New Zealand White rabbits, the test substance was applied 5 days a week for 3 weeks at dosage levels of 100, 500 and 2500 mg/kg. Four male and four female rabbits were used in each treated group and in a control group. 0.9% physiological saline was used as the control substane. The rabbits were observed daily for signs of dermal irritation and changes in general behavior and appearance. Individual body weights were recorded weekly Hematologic and biochemical studies were conducted once in the pretest period and at 21 days of study.

The NOAEL was considered to be > 2500 mg/kg. Very slight dermal irritation was noted for one rabbit at this dosage level. No compound related changes were seen in general behavior and appearance, body weight, clinical laboratory tests, organ weights or survival.