N-butylbenzenesulphonamide

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:

27/10/2011-03/01/2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 410 (Repeated Dose Dermal Toxicity: 21/28-Day Study)

Principles of method if other than guideline:

/

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

N-butylbenzenesulphonamide

EC Number:

222-823-6

EC Name:

N-butylbenzenesulphonamide

Cas Number:

3622-84-2

Molecular formula:

C10H15NO2S

IUPAC Name:

N-butylbenzenesulfonamide

Constituent 2

Reference substance name:

N-n-butylbenzenesulphonamide (Proviplast 024)

IUPAC Name:

N-n-butylbenzenesulphonamide (Proviplast 024)

Test material form:

other: liquid

Details on test material:

- Name of test material (as cited in study report): N-n-butylbenzenesulphonamide (Proviplast 024)
- Physical state: liquid
- Analytical purity: 99.9%
- Impurities (identity and concentrations): water: 0.08% want butylaminde content: 0.001%
- Purity test date: 1/9/2011
- Batch/ Lot no.: 201109070021
- Storage condition of test material:Required quantity of the test substance was received from the Test Substance Controller (TSC, IIBAT) and stored at dry cool and well ventilated area in the utility room.
- Expiration date of the lot/batch: two years

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, USA
- Age at study initiation:
- Weight at study initiation: Males: 224-285g, Females: 221-237g
- Fasting period before study: no
- Housing: Standard polypropylene rat cages with stainless steel top grills supplied by M/s. Vishnu Traders, UP, India was used to house the animals. The cages were autoclaved. Gamma irradiated corn cobs supplied by M/s. Ceutics Pharma Pvt. Ltd., Bangalore, India was used as the bedding material.
- Diet (e.g. ad libitum):ad libitum
- Water (e.g. ad libitum):ad libitum
- Acclimation period: Five days prior to the range finding and main study in the test room.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.5-22.9°C
- Humidity (%): 53-65%
- Photoperiod (hrs dark / hrs light): 12hrs dark/12 hrs light

IN-LIFE DATES: From: 17/11/2011 To: 03/01/2012

Administration / exposure

Type of coverage:

semiocclusive

Vehicle:

unchanged (no vehicle)

Details on exposure:

TEST SITE
- Area of exposure: dorsal area
- % coverage: +/- 10%
- Type of wrap if used: a porous gauze dressing and a non irritating tape
- Time intervals for shavings or clipplings: Initially, 24 hours prior to the first application of the test substance, there after clipping was done at weekly intervals, before 24 hours of the application.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): After the exposure period, the residual test substance was wiped gently from the skin using cotton soaked in water.
- Time after start of exposure: 6h

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): not specified
- Constant volume or concentration used: yes


USE OF RESTRAINERS FOR PREVENTING INGESTION: yes

Analytical verification of doses or concentrations:

no

Details on analytical verification of doses or concentrations:

/

Duration of treatment / exposure:

6 hours

Frequency of treatment:

5 days per week

Doses / concentrations

Remarks:

Doses / Concentrations:
250, 500 and 1000 mg/kg bw
Basis:
nominal per unit body weight

No. of animals per sex per dose:

5 males, 5 females

Control animals:

yes, concurrent no treatment

Details on study design:

- Dose selection rationale: Based on a range finding study
- Rationale for selecting satellite groups: not specified
- Post-exposure recovery period in satellite groups: 14 days

Positive control:

/

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Range finding animals were observed for a period of 7 days. In main study, all animals were observed individually, daily, for 28 days
- Cage side observations: various end-points of toxicity

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule:In main study, during the treatment period, all animals were observed daily soon after the application for clinical signs of toxicity. During treatment period all groups were observed twice daily, and during the post-treatment observation period satellite groups of animals were observed once daily.
- Clinical signs of toxicity: changes in skin, fur, eyes and mucous membranes, occurrence of secretions and excretions.

DERMAL IRRITATION (if dermal study): No

BODY WEIGHT: Yes
- Time schedule for examinations:Body weight of each animal was recorded just prior to the application of the test substance (day 0) and then on a weekly basis upto termination of test.

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

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION: No
- Time schedule for examinations:

OPHTHALMOSCOPIC EXAMINATION: No
- Time schedule for examinations:
- Dose groups that were examined:

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Blood was collected through orbital sinus from main group animals (overnight fasted) at the end of treatment period (day 28). Blood was collected through orbital sinus from all the groups of satellite animals (overnight fasted) at the end of treatment period (day 28) and at the end of the post treatment period (day 42).
- Anaesthetic used for blood collection: No
- Animals fasted: Yes, overnight
- How many animals: all
- Parameters checked: Erythrocyte count, Hemoglobin (Hb), Haematocrit (HCT), MCV, MCH, MCHC, Platelet count, Leucocyte count, Differential count (5 part) using Hematology Bayer Advia 120 fully automated analyzer. Prothrombin time was done by Thromborel-S method. Clotting time was done by capillary tube method.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood:Blood was collected through orbital sinus from main group animals (overnight fasted) at the end of treatment period (day 28). Blood was collected through orbital sinus from all the groups of satellite animals (overnight fasted) at the end of treatment period (day 28) and at the end of the post treatment period (day 42).
- Animals fasted: Yes
- How many animals:all
- Parameters checked: Glucose, Total cholesterol, Triglycerides, Total Bilirubin, Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), Alkaline phosphatase (ALP), Gamma glutamyl transpeptidase, Blood urea nitrogen (BUN), Creatinine, Total proteins, Albumin, Globulin, Calcium and Phosphorus using Seimens dimension Xpand plus fully automated biochemistry analyzer.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No


OTHER:

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
At the end of the observation period (day 28/42), all test animals were humanely sacrificed by using CO2 asphyxiation method. A detailed gross necropsy was carried out on all animals of all groups after examination of the external surfaces of the body including all orifices. The cranial, thoracic and abdominal cavities were opened and the appearance of the tissues and organs were observed. All gross pathological findings were recorded individually for each animal.
The following organ wet weights were determined from all the animals during the time of necropsy.
1. Liver 2. Kidneys 3. Adrenals 4. Testes (males)
Normal and treated skin, liver, kidneys were collected and kept in 10% neutral buffered formalin for fixation. Organs with gross pathological lesions were also collected and kept in the appropriate fixative and processed for histopathologic examinations.

HISTOPATHOLOGY: Yes
Histopathological examination was performed on the preserved organs and tissues of the high dose group, control group and on the organs showing gross pathological lesions.

Other examinations:

/

Statistics:

The data was subjected to Modified-levene equal-variance test for homogeneity. Homogenous data was submitted to Analysis of Variance (ANOVA) followed by Student-Newman-Keul’s test for post hoc comparison in NCSS, 2007. In the case of heterogeneous data it was subjected to kruskal-wallis multiple-comparison Z-value test.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Dermal irritation:

not examined

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

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:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY: None of the animals from treated as well as control group showed any clinical signs of toxicity during entire experimental period. No morbidity/mortality was observed in both treated and control groups of rats during the entire experiment period.

BODY WEIGHT AND WEIGHT GAIN:
Day-28
No statistical significant changes were observed in body weights of both sex belong to G2 - G4 groups when compared with G1 (Control) group of animals
Day-42
No statistical significant changes were observed in body weights of both sex belong to G6 group of animals when compared with G5 (Control - satellite) group of animals

FOOD CONSUMPTION:
Day-28
No statistical significant changes were observed in feed weight of both sex belong to G2 - G4 groups when compared with G1 (Control) group of animals
Day-42
No statistical significant changes were observed in feed weight of both sex belong to G6 group of animals when compared with G5 (Control - satellite) group of animals


HAEMATOLOGY
Day-28
Statistical significant difference was exhibited by G3 and G4 group of males in MCV when compared with G1 group of males. G6 males exhibited statistical difference in platelet and basophil count when compared with G5 males. No statistical significant changes were observed in hematology parameters of females of G2 -G4 and G6 group animals when compared with G1 and G5 groups respectively.
Day-42
No statistical significant changes were observed in hematology parameters of G6 group males when compared with G5 group males. Statistical significant difference in basophil count was observed in G6 females when compared with G5 group females

CLINICAL CHEMISTRY
Day- 28
Total Protein in G3 males and albumin in G4 males showed statistical significant difference when compared with G1 group males. Globulin in G3 females exhibited statistical significant difference when compared with G1 group females. No statistical significant changes were observed in males of G6 when compared with G5 group. Statistical significant difference in chlorides was observed in G6 females when compared with G5 group females.
Day-42
No statistical significant changes were observed in G6 males when compared with G5 group males. Statistical significant difference was noted in albumin of G6 females when compared with G5 female group
The above changes (day 28 & 42) were not observed in dose dependent manner, hence these changes could be considered as individual animal normal biological variance and not attributed to the application of test substance.

ORGAN WEIGHTS
Day-28
No statistical significant values were observed in relative as well as absolute organ weights of G2- G4 groups of both sex when compared with G1 group of animals
Day - 42
No statistical significant values were observed in relative as well as absolute organ weights of G6 of both sex when compared with G5 group of animals

GROSS PATHOLOGY: No test substance related gross pathological findings were observed. The gross necropsy findings observed were either related to physiological, agonal, to spontaneous, or were incidental and of the type routinely observed in Wistar rats of this age

HISTOPATHOLOGY: No test substance related histopathological findings were observed. The histopathologic findings observed were either related to agonal, to spontaneous, or were incidental and of the type routinely observed in Wistar rats of this age

Effect levels

Key result

Dose descriptor:

NOEL

Effect level:

> 1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: overall effects

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

/

Applicant's summary and conclusion

Conclusions:

On the basis of the results obtained in the present study, no mortality was observed in both sex of treated as well as control group of animals during entire experiment period. Although few changes occurred in hematology and clinical biochemistry parameters, those changes were not related to test substance and could be considered as individual animal normal biological variance. There was no test substance related macroscopic and microscopic findings observed. In the light of above observations, the NOEL of N-n- Butylbenzenesulphonamide (PROVIPLAST 024) for Wistar rats could be considered as> 1000 mg/kg b.w. under the experimental conditions.

Executive summary:

A repeated dose - 28 day dermal toxicity study was conducted with N-n-butylbenzenesulphonamide in Wistar Rats at dose levels of 0, 250, 500 and 1000 mg/kg bw (semi-occlusive dosing; no vehicle). On the basis of the results obtained in the present study, no mortality was observed in both sex of treated as well as control group of animals during entire experiment period. Although few changes occurred in hematology and clinical biochemistry parameters, those changes were not related to test substance and could be considered as individual animal normal biological variance. There was no test substance related macroscopic and microscopic findings observed. In the light of above observations, the NOEL of N-n- Butylbenzenesulphonamide for Wistar rats could be considered as >1000 mg/kg b.w. under the experimental conditions.