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Administrative data

Description of key information

Repeated Dose (Oral) Toxicity:

Based on all the available data, it was concluded that the test chemical did not cause any toxicity to any organs when exposed to the test animals in sub-acute and sub-chronic durations via oral route. Therefore, it was concluded that the test chemical was not likely to be classified under category STOT-RE 1 or 2 as per the CLP criteria of classification and labeling.

Repeated Dose (Inhalation) Toxicity:

The particle size distribution of the substance methyl 2-naphthyl ether was found to vary in the size of 150 µm to 2000 µm,so the potential for the generation of inhalable vapours of methyl 2-naphthyl ether is low. Moreover the normal conditions of use of this substance will not result in aerosols, particles or droplets of an inhalable size, so exposure to humans via the inhalatory route will be unlikely to occur and therefore repeated dose toxicity via the inhalation route was considered for waiver.

Repeated Dose (Dermal) Toxicity:

The acute toxicity value for test chemical (as provided in section 7.2.3) is >2000 mg/kg body weight. Also, given the use of the chemical; repeated exposure by the dermal route is unlikely since the use of gloves is common practice in industries. Thus, it is expected that the test chemical shall not exhibit 28 day repeated dose toxicity by the dermal route. In addition, there is no data available that suggests that the test chemical shall exhibit repeated dose toxicity by the dermal route. Hence this end point was considered for waiver.

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Referenceopen allclose all

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Data is from a study report
Qualifier:
according to guideline
Guideline:
other: OECD TG 443: Extended One-Generation Reproductive Toxicity Study
Principles of method if other than guideline:
According to OECD TG 443: Extended One-Generation Reproductive Toxicity Study
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Details on species / strain selection:
Rat is the commonly used species for toxicity studies and recommended by the regulatory guidelines and Wistar strain is a commonly used strain to assess the reproduction and developmental toxicity in the reproductive toxicity studies.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: The test animals were procured from a CPCSEA approved vendor.
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: (P) 12-14 weeks at the start of treatment wks; (F1) 11-12 weeks at the start of treatment wks
- Weight at study initiation: (P) Males: 291-413 g; Females: 198-277 g; (F1) Males: x-x g; Females: x-x g
- Fasting period before study: No Data Available
- Housing: Two to four rats were housed in each polycarbonate cage. During mating, one male and one female rat were housed in a single cage. Pregnant and lactating females were housed individually. Sterilized corn-cob produced from pure corn, dried and free from dust, procured from approved supplier, was used as bedding material. It was renewed as often as necessary to keep the animals dry and clean.
- Use of restrainers for preventing ingestion (if dermal): no
- Diet (e.g. ad libitum): A conventional laboratory pellet diet from approved supplier was available ad libitum.
- Water (e.g. ad libitum): Aquaguard™ filtered drinking water was available ad libitum in regularly cleaned bottles.
- Acclimation period: Animals were acclimatized for at least 5 days prior to test item administration.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.60 to 24.80°C
- Humidity (%): 34.40 to 66.70 %
- Air changes (per hr): At least 12 changes per hour
- Photoperiod (hrs dark / hrs light): 12 hrs dark / 12 hrs light

IN-LIFE DATES:
From: August 8, 2019
To: March 1, 2020
Route of administration:
oral: gavage
Details on route of administration:
While concentrations of the formulation were fixed for each experimental group, the dose volume for each animal was calculated on the basis of the most recently recorded body weight. Formulation was administered in a single dose through oral gavage at a constant volume to weight ratio of 4 mL/ kg body weight.
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: The vehicle used for dose administration was corn oil. Required quantity for each concentration level of test chemical was weighed and triturated in a mortar-pestle, first without vehicle and then in small volume of the vehicle. After thorough trituration, the test chemical and vehicle were transferred quantitatively into a calibrated measuring cylinder and diluted using vehicle up to the required volume of the formulation, before being transferred to the final container. Formulations were prepared on the same day or one day prior to dose administration and stored at room temperature until usage as the test item stability in solution was proved up to 24 hours. At the time of dosing, dose formulations were kept on a magnetic stirrer for maintaining homogeneity of test formulation. The dose formulation samples [of doses viz; G1 (Control), G2 (Low dose), G3 (Mid dose), and G4 (High dose)] were analyzed for homogeneity and concentration. The dose formulation analysis was carried out once on Day 1 of dose administration to animals, on day 91 and on day 182.

DIET PREPARATION
- Rate of preparation of diet (frequency): No Data Available
- Mixing appropriate amounts with (Type of food): No Data Available
- Storage temperature of food: No Data Available

VEHICLE
- Justification for use and choice of vehicle (if other than water): Corn Oil was selected as a vehicle because the test chemical was found to be insoluble in water whereas corn oil provided adequate suspensibility and the formulation was found stable in this vehicle. Further, corn oil is widely used as vehicle in oral toxicity study and the selected concentration is well below the acceptable limit.
- Concentration in vehicle: 0 mg/kg bw: 0 mg/ml; 62.5 mg/kg bw: 15.63 mg/ml; 125 mg/kg bw: 31.25 mg/ml and 250 mg/kg bw: 62.5 mg/ml
- Amount of vehicle (if gavage): 4 mL/ kg
- Lot/batch no. (if required): Batch No.A1906001, N2191405, N2192016, O21904007
- Purity: No Data Available
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical method was validated with respect to the parameters covering specificity, linearity, limit of quantification (LOQ), precision (% RSD), Accuracy (% recovery) and Homogeneity. The analytical method was validated with respect to the following parameters as below:

1. Specificity: The result of specificity was reported either no interference observed based on visual comparison or the degree of interference does not contribute more than 20 % peak area of the target analyte at the LOQ Level.

2. Limit of Quantification: The LOQ of the method was the minimum concentration/quantity of a component, which could be quantified precisely and accurately.
The LOQ was carried out at the lowest sensitivity of the device under the same instrumental parameters used for the qualified specificity.
The LOQ of a compound for a specific method was determined by injecting in duplicate the working solution (s) made in suitable solvent fortified with test chemical / reference standard, until it produces a minimum signal to noise ratio ≥ 10:1 for precise quantification.

3. Linearity: The linearity was carried out by preparing and analyzing minimum five linear concentrations of reference standard with at least one determination. The linear calibration curve was established by plotting the analyte peak response against concentration (mg/l). The linearity was given in the form of equation of the resulting curve and correlation coefficient (r). The value of correlation coefficient (r) ≥ 0.99 was considered as an acceptable value.

4. Precision (% RSD), Accuracy (% Recovery) and Homogeneity: Precision, Accuracy was established at the level of test concentration used i.e., at low, mid and high dose level along with control. The active ingredient was diluted using sample diluent and injected onto HPLC using validated HPLC method. Minimum six determinations were made at each fortification level. The homogeneity was performed by analysing Top, Middle and Bottom layers of each fortification level. The active ingredient concentration in each replicate of each fortification level, mean of active ingredient concentration, accuracy (% recovery), SD and % RSD was calculated and reported.
The active ingredient concentration of test item in dose formulation was calculated using the following formula:
Y = bX + a

"AI Concentration (mg/L) = " "Y - a" /"b" " × D"


Where,
Y = Peak area of the sample
a = Intercept
b = Slope of the line
D = Dilution factor (if applicable)
X = A.I. Concentration (mg/L)


The accuracy (% Recovery) will be calculated using the following formula:

"Accuracy " ("% Recovery" )"= " "Obtained Concentration" /"Fortified concentration" " ×100"
The precision (% RSD) will be calculated using the formula provided as below:
"Precision (% RSD)= " "Standard Deviation" /"Mean Recovered Concentration" " ×100"

All the paramaters were according to and within the acceptance criteria of the above test.
Duration of treatment / exposure:
Dosing in all animals was carried out on a 7-days/week basis from the treatment initiation until necropsy. All animals of both sexes were dosed 2 weeks prior to mating. Dosing was continued in both sexes during the mating period. Females were treated throughout gestation and lactation up to the day of termination after weaning. Males were treated in the same manner until termination. The parental animals were dosed until 7 weeks.
Frequency of treatment:
Once Daily
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Remarks:
Control Group (G1)
Dose / conc.:
62.5 mg/kg bw/day (actual dose received)
Remarks:
Low Dose Group (G2)
Dose / conc.:
125 mg/kg bw/day (actual dose received)
Remarks:
Mid Dose Group (G3)
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Remarks:
High Dose Group (G4)
No. of animals per sex per dose:
30 animals per sex per dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Doses were selected on the basis of previously performed GLP compliant repeated dose 28 days oral toxicity study. Two deaths were observed in the highest dose (500 mg/kg bw) unrelated to the gavage error of the OECD 407 study and therefore a subsequent lower dose of 250 mg/kg bw was selected as a highest dose in current study.
- Rationale for animal assignment (if not random): During the acclimatisation phase, male and female animals were randomized into four different experimental groups, based on the most recently recorded body weights, using the “Daniel’s XL Toolbar” (http://xltoolbox.sourceforge.net/) for MS Excel. After randomisation, it was ensured that individual body weights were within ± 20% of the respective group means. Details of the randomization were documented in the study raw data. The mean body weights were analysed by One-way ANOVA and it was found that the group means were statistically comparable to each other within each sex and group.
- Fasting period before blood sampling for clinical biochemistry: Animals were overnight fasted prior to blood collection.
- Rationale for selecting satellite groups: No Data Available
- Post-exposure recovery period in satellite groups: No Data Available
- Section schedule rationale (if not random): No Data Available
- Other: No Data Available
Positive control:
No Data Available
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: All animals were observed at least twice daily for cage size observation, throughout the acclimatization and study periods.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: General clinical observations of rats from all the groups were made once a day, preferably at the same time each day and considering the peak period of anticipated effects after dosing.

BODY WEIGHT: Yes
- Time schedule for examinations: Parental animals were weighed once on receipt, once during randomization, once at the start of treatment and once every week thereafter. In addition, pregnant females were weighed during gestation (GD 0, GD 7, GD 14 and GD 20) and lactation on the same days as the weighing of the pups in their litters.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes, during the study, feed consumption was recorded weekly or on the same days as animal body weights (except during cohabitation).
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Not specified
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Not specified

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: Yes

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Not specified
- Time schedule for examinations:

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

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Prior to termination of the parental animals
- Anaesthetic used for blood collection: Yes (isofluorane)
- Animals fasted: Yes
- How many animals: 10 males and 10 females, randomly selected from each group of parental animals
- Parameters checked in table [1.1] were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Prior to termination of the parental animals
- Animals fasted: Yes (isofluorane)
- How many animals: 10 males and 10 females, randomly selected from each group of parental animals
- Parameters checked in table [1.2] were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: During the last week of the treatment, 10 males and 10 females, randomly selected from each group of parental animals. During the last week of the treatment, 10 males and 10 females, randomly selected from each group of parental animals.
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters checked in table [1.3] were examined.

NEUROBEHAVIOURAL EXAMINATION: No
- Time schedule for examinations: No Data Available
- Dose groups that were examined: No Data Available
- Battery of functions tested: No Data Available

IMMUNOLOGY: Not specified
- Time schedule for examinations: No Data Available
- How many animals: No Data Available
- Dose groups that were examined: No Data Available

OTHER: Hormone Analysis: From 10 males and 10 females, randomly selected from each group of parental animals , blood was collected in the morning; serum was separated and stored under appropriate conditions (-70°C ± 5°C). All collected blood samples were assessed for serum levels of T4, TSH, Estradiol, Progesterone and Testosterone using commercially available Rat ELISA kits.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (see table 1.4)

HISTOPATHOLOGY: Yes (see table 1.4), full histopathology was carried out on the preserved organs and tissues of all parental animals in the control and high dose group of both sexes. Slides for low- and mid-dose groups were processed only if, test-chemical related pathologies were observed in the high-dose group sections.
Other examinations:
Organ Weights: At the time of termination, organs marked with asterisk (*) in table 1.4 from all parental animals were trimmed of adherent tissue/ fat and weighed, prior to preservation in fixative.
Statistics:
The findings of this study were evaluated in terms of the observed effects, necropsy and microscopic findings. The evaluation will included the relationship between the dose of the test substance and the presence or absence, the incidence and severity, of abnormalities. Raw data was processed using statistical software. The mean and standard deviation was calculated using the software and all data was summarized in tabular form. All continuous data were checked for their homogeneity using Shapiro Wilk test. All homogenous data were analyzed using ANOVA and data showing significance in their variances were subjected to Dunnett’s t-test. All heterogeneous data was analysed using Kruskal-Wallis ANOVA on ranks and data showing significance in their variances were subjected to Dunn’s Test. P values of less than or equal to 0.05 were accepted as being statistically significant.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
No treatment-related clinical signs or symptoms were observed in any of the parent animals up to 125 mg/kg body weight of the test chemical. However, mild salivation was observed in total 13 of 30 males of G4 group (250 mg/kg), starting from the 27th day of treatment and continuing till termination. All adult animals were found normal with respect to all the parameters examined during detailed clinical examinations through the course of the study. No treatment-related symptoms were found up to the dose level of 250 mg/kg body weight in either sex.
Mortality:
no mortality observed
Description (incidence):
No morbidities or mortality were recorded in adult animals of parent generation in any of the experimental groups throughout the duration of the experiment.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Mean body weights remained comparable among all the groups for every time point (days 8, 15, 22, 29, 36, 43, 50, 57, 64 and 71) recorded during the 70 days exposure period for males. No significant difference was found for the mean body weights of females during the two weeks pre-mating period, during mating, during gestation or during lactation period. However, the mean percent change in body weight on all the instances (days 8, 15, 22, 29, 36, 43, 50, 57, 64 and 71) with respect to the first day of exposure was significantly reduced for parental males of G4 group (250 mg/kg) (P≤0.05 to P≤0.001) as compared to control animals, whereas mean body weight change for G2 (62.5 mg/kg) and G3 (125 mg/kg) males remained comparable to control animals. Among the females, mean percent change in body weights remained statistically comparable among all groups during pre-mating, mating, gestation and lactation periods.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
The mean feed consumed per animal per day significantly increased on day 8 of treatment for G2 (62.5 mg/kg) (P≤0.01) and G3 (125 mg/kg) (P≤0.05) males as compared to G1 (control) animals. However, it was found to be similar for all other time points across the groups. The mean feed consumption of treatment group (G2, G3 & G4) females remained statistically comparable to control group during pre-mating, mating, gestation and lactation period except for one instance, where mean feed consumption on day 8 of treatment was significantly reduced for G2 (62.5 mg/kg) (P≤0.01) as compared to control females. The differences found in mean feed consumption of male and female animals were neither dose-dependent nor consistent, hence it was deemed to be independent of test-chemical treatment.
Food efficiency:
effects observed, non-treatment-related
Description (incidence and severity):
Feed efficiency for the G4 males had statistically reduced in the first two weeks of treatment, i.e. on days 8 and 15. It was also found to have increased significantly during the penultimate week of treatment, i.e. on day 63. For all other groups among the males, the feed efficiency was statistically comparable across the groups. Among the parental females, no statistical differences were observed for feed efficiency in any of the groups.
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
No significant difference was observed in any of the measured haematology parameters in all the groups of both the sexes, except in the WBC count, which was found to be significantly increased in males of G4 group (P≤0.01) as compared to control males.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Significant increase was observed in Phosphorus, Sodium and Chloride levels in G4 males (P≤0.05) as compared to control males. All the other clinical chemistry parameters in males and females were found comparable in control and treatment groups.
Urinalysis findings:
no effects observed
Description (incidence and severity):
The urine parameters tested did not show up any significant changes among the experimental groups in the parental generation of either sex.
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
No significant difference was observed in absolute organ weight or organ weight relative to body weight for the entire set of measured organs in parent male and female animals. However, significant reduction was observed in absolute and relative liver weight in G4 females (P<0.05) as compared to control animals. In the current study, however, the decreased liver weights with respect to body weights in G4 females does not correlate with any histological observations made, making it inconsequential from the point of view of toxic effects of test-chemical.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
External examination of all male and female rats of control and all treated groups did not reveal any lesion of pathological significance. Internal examination of the rats revealed reduced testicular size in two animals (Male: G2: 1/30, G4:1/30), increased prostate size in one animal (Male: G2: 1/30) and reduced prostate size in one animal (Male: G3: 1/30). Remaining animals of parent generation did not reveal any abnormality of pathological significance.
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
No treatment-related histopathological findings are reported in this study that could arise out of test-item administration. All observed tissues were normal at the level of histology. Minimal to moderate and focal lymphocytic infiltrates were observed in some slides of liver, kidney, lung, heart, colon, thymus and adrenals with mild to moderate cyst in thyroid gland, which were distributed randomly across the experimental groups and did not show any pattern of either dose-dependency or sex-based selectivity.
Histopathological findings: neoplastic:
not specified
Other effects:
effects observed, treatment-related
Description (incidence and severity):
While TSH, Estradiol, Progesterone and Testosterone remained statistically comparable among the groups of parental males, the mean level of T4 was found reduced in G3 males (P<0.01) as compared to control. This is likely an incidental result, given that G4 was statistically comparable to control. The mean T4, TSH, Estradiol and testosterone levels remained comparable among the groups of parental females; whereas, Progesterone was found to be significantly reduced in G4 females (P<0.001). This could be a test-chemical related effect.
Details on results:
No treatment-related clinical signs or symptoms were observed in any of the parent animals up to 125 mg/kg body weight of the test chemical. However, mild salivation was observed in total 13 of 30 males of G4 group (250 mg/kg), starting from the 27th day of treatment and continuing till termination. All adult animals were found normal with respect to all the parameters examined during detailed clinical examinations through the course of the study. No treatment-related symptoms were found up to the dose level of 250 mg/kg body weight in either sex. No morbidities or mortality were recorded in adult animals of parent generation in any of the experimental groups throughout the duration of the experiment. Mean body weights remained comparable among all the groups for every time point (days 8, 15, 22, 29, 36, 43, 50, 57, 64 and 71) recorded during the 70 days exposure period for males. No significant difference was found for the mean body weights of females during the two weeks pre-mating period, during mating, during gestation or during lactation period. However, the mean percent change in body weight on all the instances (days 8, 15, 22, 29, 36, 43, 50, 57, 64 and 71) with respect to the first day of exposure was significantly reduced for parental males of G4 group (250 mg/kg) (P≤0.05 to P≤0.001) as compared to control animals, whereas mean body weight change for G2 (62.5 mg/kg) and G3 (125 mg/kg) males remained comparable to control animals. Among the females, mean percent change in body weights remained statistically comparable among all groups during pre-mating, mating, gestation and lactation periods. The mean feed consumed per animal per day significantly increased on day 8 of treatment for G2 (62.5 mg/kg) (P≤0.01) and G3 (125 mg/kg) (P≤0.05) males as compared to G1 (control) animals. However, it was found to be similar for all other time points across the groups. The mean feed consumption of treatment group (G2, G3 & G4) females remained statistically comparable to control group during pre-mating, mating, gestation and lactation period except for one instance, where mean feed consumption on day 8 of treatment was significantly reduced for G2 (62.5 mg/kg) (P≤0.01) as compared to control females. The differences found in mean feed consumption of male and female animals were neither dose-dependent nor consistent, hence it was deemed to be independent of test-item treatment. Feed efficiency for the G4 males had statistically reduced in the first two weeks of treatment, i.e. on days 8 and 15. It was also found to have increased significantly during the penultimate week of treatment, i.e. on day 63. For all other groups among the males, the feed efficiency was statistically comparable across the groups. Among the parental females, no statistical differences were observed for feed efficiency in any of the groups. No significant difference was observed in any of the measured haematology parameters in all the groups of both the sexes, except in the WBC count, which was found to be significantly increased in males of G4 group (P≤0.01) as compared to control males. Significant increase was observed in Phosphorus, Sodium and Chloride levels in G4 males (P≤0.05) as compared to control males. All the other clinical chemistry parameters in males and females were found comparable in control and treatment groups. The urine parameters tested did not show up any significant changes among the experimental groups in the parental generation of either sex. No significant difference was observed in absolute organ weight or organ weight relative to body weight for the entire set of measured organs in parent male and female animals. However, significant reduction was observed in absolute and relative liver weight in G4 females (P<0.05) as compared to control animals. In the current study, however, the decreased liver weights with respect to body weights in G4 females does not correlate with any histological observations made, making it inconsequential from the point of view of toxic effects of test-chemical. External examination of all male and female rats of control and all treated groups did not reveal any lesion of pathological significance. Internal examination of the rats revealed reduced testicular size in two animals (Male: G2: 1/30, G4:1/30), increased prostate size in one animal (Male: G2: 1/30) and reduced prostate size in one animal (Male: G3: 1/30). Remaining animals of parent generation did not reveal any abnormality of pathological significance. No treatment-related histopathological findings are reported in this study that could arise out of test-item administration. All observed tissues were normal at the level of histology. Minimal to moderate and focal lymphocytic infiltrates were observed in some slides of liver, kidney, lung, heart, colon, thymus and adrenals with mild to moderate cyst in thyroid gland, which were distributed randomly across the experimental groups and did not show any pattern of either dose-dependency or sex-based selectivity. While TSH, Estradiol, Progesterone and Testosterone remained statistically comparable among the groups of parental males, the mean level of T4 was found reduced in G3 males (P<0.01) as compared to control. This is likely an incidental result, given that G4 was statistically comparable to control. The mean T4, TSH, Estradiol and testosterone levels remained comparable among the groups of parental females; whereas, Progesterone was found to be significantly reduced in G4 females (P<0.001). This could be a test-chemical related effect.
Key result
Dose descriptor:
NOAEL
Effect level:
125 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
clinical biochemistry
clinical signs
food consumption and compound intake
food efficiency
gross pathology
haematology
histopathology: non-neoplastic
mortality
organ weights and organ / body weight ratios
urinalysis
other: Hormone Analysis
Remarks on result:
not determinable due to absence of adverse toxic effects
Dose descriptor:
LOAEL
Effect level:
250 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
organ weights and organ / body weight ratios
other: Hormone Analysis
Remarks on result:
other: Please see 'remarks'
Remarks:
There were treatment related changes observed during clinical signs examination, organ weights and progesterone (hormonal) levels.
Critical effects observed:
no

Table 2.1: Mortality and Morbidity Record

 

Sex: Male

Group

Dose (mg/kg)

No. of Animals

Observation

G1

0

30

No mortality or morbidity observed

G2

62.5

30

No mortality or morbidity observed

G3

125

30

No mortality or morbidity observed

G4

250

30

No mortality or morbidity observed

 

 

Sex: Female

Group

Dose (mg/kg)

No. of Animals

Observation

G1

0

30

No mortality or morbidity observed

G2

62.5

30

No mortality or morbidity observed

G3

125

30

No mortality or morbidity observed

G4

250

30

No mortality or morbidity observed

 

Table 2.2: Clinical Signs and Symptoms Record

 

Sex: Male

Group

Dose (mg/kg)

No. of Animals

Observation

G1

0

30

Normal

G2

62.5

30

Normal

G3

125

30

Normal

G4

250

30

Normal

 

Sex: Female

Group

Dose (mg/kg)

No. of Animals

Observation

G1

0

30

Normal

G2

62.5

30

Normal

G3

125

30

Normal

G4

250

30

*Normal

Table 2.3 Mean Body weights (g)

Sex: Male

Group

Dose (mg/Kg)

 

Day of Exposure

1

8

15

22

29

36

43

50

57

64

71

G1

0

Mean

351.73

359.60

371.80

384.63

400.93

419.17

426.90

431.53

441.13

444.70

452.53

SD

26.71

26.09

25.43

27.51

28.22

30.60

31.14

30.71

29.88

33.09

32.31

N

30

30

30

30

30

30

30

30

30

30

30

G2

62.5

Mean

357.10

365.17

377.03

387.97

406.10

426.53

434.03

439.53

448.87

451.17

459.23

SD

26.62

24.64

22.51

23.92

26.57

28.79

29.40

29.89

29.65

31.22

32.62

N

30

30

30

30

30

30

30

30

30

30

30

G3

125

Mean

356.17

363.80

373.40

385.77

402.57

419.73

428.73

435.50

441.70

447.23

454.03

SD

25.19

24.28

24.93

28.35

26.99

30.13

33.92

32.82

31.43

34.62

38.82

N

30

30

30

30

30

30

30

30

30

30

30

G4

250

Mean

356.17

359.17

365.40

377.27

392.43

407.17

413.50

417.67

427.13

435.77

441.33

SD

26.31

25.80

25.49

26.75

31.22

31.45

32.22

34.58

39.78

41.59

42.78

N

30

30

30

30

30

30

30

30

30

30

30

 

Pre-mating

Sex: Female

Group

Dose (mg/Kg)

 

Day of Exposure

1

8

15

22

G1

0

Mean

238.07

242.10

246.67

256.50

SD

17.01

16.63

16.35

14.89

N

30

30

30

4

G2

62.5

Mean

237.60

242.53

248.17

272.50

SD

20.80

20.77

19.25

16.26

N

30

30

30

2

G3

125

Mean

241.83

244.50

247.47

244.25

SD

19.86

20.38

21.86

14.45

N

30

30

30

4

G4

250

Mean

240.87

241.93

245.80

258.00

SD

16.30

15.80

14.70

22.63

N

30

30

30

2

Keys:N: number; SD: standard deviation

 

Gestation

Sex: Female

Group

Dose (mg/Kg)

 

Day of Gestation

0

7

14

20

G1

0

Mean

253.72

280.45

310.76

374.66

SD

17.43

19.74

22.43

29.95

N

29

29

29

29

G2

62.5

Mean

253.36

282.14

312.32

372.75

SD

21.86

23.49

24.03

30.58

N

28

28

28

28

G3

125

Mean

250.93

278.55

310.45

372.38

SD

20.57

20.32

24.60

30.02

N

29

29

29

29

G4

250

Mean

247.04

275.42

306.04

364.27

SD

15.24

17.44

20.50

27.77

N

26

26

26

26

 

Lactation

Sex: Female

Group

Dose (mg/Kg)

 

Day of Lactation

0

4

7

14

21

G1

0

Mean

293.83

294.10

294.90

299.31

296.38

SD

19.69

21.02

21.57

25.65

23.89

N

29

29

29

29

29

G2

62.5

Mean

295.23

300.73

303.13

308.03

307.90

SD

29.28

24.37

22.64

24.24

25.74

N

28

28

28

28

28

G3

125

Mean

293.48

293.21

297.86

305.03

304.83

SD

27.80

26.55

25.42

27.10

26.87

N

29

29

29

29

29

G4

250

Mean

293.88

292.58

299.12

304.77

302.19

SD

26.71

27.01

28.33

26.21

26.98

N

26

26

26

26

26

Keys:N: number; SD: standard deviation

 

Non-Parturated

Sex: Female

Group

Dose (mg/Kg)

 

Day of Gestation

0

7

14

20

27

30

34

G1

0

Mean

260.00

261.00

271.00

274.00

271.00

./.

270.00

SD

./.

./.

./.

./.

./.

./.

./.

N

1

1

1

1

1

0

1

G2

62.5

Mean

271.50

274.50

279.50

278.50

281.50

289.00

264.00

SD

41.72

37.48

12.02

16.26

21.92

./.

./.

N

2

2

2

2

2

1

1

G3

125

Mean

251.00

252.00

257.00

255.00

255.00

./.

258.00

SD

./.

./.

./.

./.

./.

./.

./.

N

1

1

1

1

1

0

1

G4

250

Mean

254.25

268.50

272.25

270.50

274.00

./.

271.00

SD

17.69

12.66

20.85

20.44

21.23

./.

20.85

N

4

4

4

4

4

0

4

Keys:N: number; SD: standard deviation;./.= Not applicable

Table 2.4 Mean Body weight Change (%)

Sex: Male

Group

Dose (mg/Kg)

 

Day of Exposure

8

15

22

29

36

43

50

57

64

71

 

G1

0

Mean

2.28

5.81

9.46

14.11

19.26

21.47

22.83

25.59

26.57

28.83

 

SD

2.19

3.17

3.81

4.23

3.92

4.07

4.89

4.99

5.55

5.66

 

N

30

30

30

30

30

30

30

30

30

30

 

G2

62.5

Mean

2.34

5.75

8.79

13.84

19.57

21.69

23.23

25.88

26.49

28.74

 

SD

2.39

3.62

3.29

3.39

4.00

4.62

4.85

5.24

5.26

5.44

 

N

30

30

30

30

30

30

30

30

30

30

 

G3

125

Mean

2.20

4.92

8.35

13.13

17.89

20.38

22.32

24.10

25.62

27.49

 

SD

2.35

3.59

3.79

4.14

3.85

4.37

4.57

4.89

5.22

6.30

 

N

30

30

30

30

30

30

30

30

30

30

 

G4

250

Mean

0.87↓

2.65↓↓↓

5.99↓↓↓

10.20↓↓↓

14.38↓↓↓

16.17↓↓↓

17.32↓↓↓

19.96↓↓↓

22.36↓↓

23.93↓↓

 

SD

1.38

2.43

2.86

3.68

4.38

4.73

5.21

7.03

7.22

7.63

 

N

30

30

30

30

30

30

30

30

30

30

 

Pre-mating

Sex: Female

 

Group

Dose (mg/Kg)

 

Day of Exposure

 

8

15

22

 

G1

0

Mean

1.73

3.71

8.14

 

SD

2.12

3.96

2.73

 

N

30

30

4

 

G2

62.5

Mean

2.12

4.60

11.64

 

SD

2.46

4.08

4.00

 

N

30

30

2

 

G3

125

Mean

1.11

2.33

7.00

 

SD

1.84

3.07

4.96

 

N

30

30

4

 

G4

250

Mean

0.46

2.12

5.38

 

SD

1.05

2.25

4.44

 

N

30

30

2

 

Key: Body weight change (%) with respect to Day 1 of treatment; ↓: decreased as compared to control (P<0.05); ↓↓: decreased as compared to control (P<0.01); ↓↓↓: decreased as compared to control (P<0.001)

Gestation

Sex: Female

Group

Dose (mg/Kg)

 

Day of Gestation

7

14

20

G1

0

Mean

10.63

22.69

47.92

SD

5.04

7.67

10.38

N

29

29

29

G2

62.5

Mean

11.42

23.41

47.30

SD

2.70

3.34

6.02

N

28

28

28

G3

125

Mean

11.14

23.83

48.53

SD

3.65

4.62

6.04

N

29

29

29

G4

250

Mean

11.50

23.92

47.47

SD

2.09

4.45

7.00

N

26

26

26

 Key: Body weight change (%) with respect to Day 0 of Gestation

Lactation

Sex: Female

Group

Dose (mg/Kg)

 

Day of Lactation

4

7

14

21

G1

0

Mean

0.18

0.46

1.95

1.07

SD

4.9

5.5

6.85

7.89

N

29

29

29

29

G2

62.5

Mean

2.15

3.04

4.73

4.67

SD

4.83

5.44

6.86

6.86

N

28

28

28

28

G3

125

Mean

0.04

1.71

4.17

4.09

SD

4.35

5.34

6.45

6.2

N

29

29

29

29

G4

250

Mean

-0.33

1.93

3.97

3.22

SD

5.12

6.24

6.91

8.61

N

26

26

26

26

Key: Body weight change (%) with respect to Day 0 of Lactation

Non-Parturated

Sex: Female

Group

Dose (mg/Kg)

 

Day of Gestation

7

14

20

27

30

34

G1

0

Mean

0.38

4.23

5.38

4.23

./.

3.85

SD

./.

./.

./.

./.

./.

./.

N

1

1

1

1

0

1

G2

62.5

Mean

1.24

3.83

3.34

4.29

-3.99

9.09

SD

1.75

11.53

9.89

7.95

./.

./.

N

2

2

2

2

1

1

G3

125

Mean

0.40

2.39

1.59

1.59

./.

2.79

SD

./.

./.

./.

./.

./.

./.

N

1

1

1

0

1

0

G4

250

Mean

5.74

7.41

6.49

7.76

./.

6.61

SD

3.00

10.47

5.99

3.05

./.

4.03

N

4

4

4

4

0

4

Table 2.5 Summary of Feed Consumption (gram/animal/day)

Sex: Male

Group

Dose (mg/Kg)

 

Day of Exposure

1 to 8

8 to 15

28 to 36

36 to 43

43 to50

50 to 57

57 to 64

64 to 71

G1

0

Mean

15.82

20.01

19.78

19.10

19.74

19.57

20.54

21.00

SD

2.90

1.52

1.66

1.33

1.47

1.10

1.78

1.58

N

30

30

30

30

30

30

30

30

G2

62.5

Mean

19.09↑↑

19.36

20.10

19.36

19.88

19.81

20.04

21.13

SD

1.89

1.59

1.30

1.27

1.48

1.23

1.92

1.55

N

30

30

30

30

30

30

30

30

G3

125

Mean

18.73↑

19.18

19.71

19.32

20.31

20.31

21.37

21.90

SD

1.48

1.73

1.48

1.53

1.78

1.23

2.11

2.10

N

30

30

30

30

30

30

30

30

G4

250

Mean

18.03

18.88

19.04

19.07

19.59

20.02

20.91

20.08

SD

1.39

1.42

1.73

1.28

1.33

1.76

1.83

2.95

N

30

30

30

30

30

30

30

30

Pre-mating

Sex: Female

 

Group

Dose (mg/Kg)

 

Day of Exposure

1 to 8

8 to 15

G1

0

Mean

15.00

13.89

SD

1.42

1.26

N

30

30

G2

62.5

Mean

13.20↓↓

13.88

SD

2.04

1.72

N

30

30

G3

125

Mean

14.14

14.61

SD

1.47

1.56

N

30

30

G4

250

Mean

13.69

13.68

SD

0.76

1.33

N

30

30

Key: SD: Standard Deviation; N: Number of Animals; ↑: increased as compared to control (P<0.05); ↑↑: increased as compared to control (P<0.01); ↓↓: decreased as compared to control (P<0.01).

Gestation

Sex: Female

Group

Dose (mg/Kg)

 

Day of Gestation

0 to 7

7 to 14

14 to 20

20 to 22

20 to 23

G1

0

Mean

16.71

19.13

22.30

25.43

23.00

SD

2.57

3.95

3.35

9.27

./.

N

29

29

29

28

1

G2

62.5

Mean

15.81

19.32

22.10

21.83

18.00

SD

2.50

1.49

3.84

7.37

1.41

N

28

28

28

26

2

G3

125

Mean

16.93

19.68

21.95

26.39

17.33

SD

1.97

1.98

3.13

11.30

1.41

N

29

29

29

27

2

G4

250

Mean

16.30

19.15

20.62

25.65

./.

SD

2.01

1.76

3.19

7.90

./.

N

26

26

26

26

0

Lactation

Sex: Female

Group

Dose (mg/Kg)

 

Day of Lactation

0 to 4

4 to 7

7 to 14

14 to 21

G1

0

Mean

29.47

38.51

48.31

56.82

SD

9.17

9.11

9.07

9.02

N

29

29

29

29

G2

62.5

Mean

27.44

39.04

47.22

57.88

SD

5.28

7.85

10.01

6.89

N

29

29

29

29

G3

125

Mean

26.88

40.29

47.83

57.73

SD

5.33

10.03

7.88

9.19

N

29

29

29

29

G4

250

Mean

24.72

35.53

47.18

55.68

SD

6.47

8.32

6.59

10.46

N

26

26

26

26

Key: SD: Standard Deviation; N: Number of Animals

Non-Parturated

Sex: Female

Group

Dose (mg/Kg)

 

Day of Gestation

0 to 7

7 to 14

14 to 20

20 to 27

27 to 30

27 to 34

G1

0

Mean

14.86

16.29

18.5

17.71

./.

17.29

SD

./.

./.

./.

./.

./.

./.

N

1

1

1

1

./.

1

G2

62.5

Mean

15.22

12.5

20.42

10.5

14

11.57

SD

7.38

3.34

5.3

9.8

./.

./.

N

2

2

2

2

1

1

G3

125

Mean

10.43

12.57

12.83

11.86

./.

11.57

SD

./.

./.

./.

./.

./.

./.

N

1

1

1

1

./.

1

G4

250

Mean

15.46

15.79

13.96

18.54

./.

16

SD

1.96

1.97

2.87

2.94

./.

1.65

N

4

4

4

4

./.

4

Key: SD: Standard Deviation; N: Number of Animals; ./. Not applicable

Table 2.6 Feed efficiency

Sex: Male


Group

Dose (mg/kg)

 

Feed Efficiency

8

15

36

43

50

57

64

71

G1

0

Mean

0.074

0.088

0.132

0.057

0.033

0.071

0.023

0.053

SD

0.067

0.049

0.046

0.043

0.047

0.047

0.059

0.031

N

30

30

30

30

30

30

30

30

G2

62.5

Mean

0.061

0.086

0.144

0.055

0.039

0.067

0.015

0.054

SD

0.063

0.055

0.047

0.043

0.039

0.040

0.053

0.040

N

30

30

30

30

30

30

30

30

G3

125

Mean

0.058

0.071

0.122

0.065

0.049

0.044

0.036

0.044

SD

0.06

0.051

0.087

0.061

0.037

0.036

0.046

0.066

N

30

30

30

30

30

30

30

30

G4

250

Mean

0.023↓↓

0.047↓↓

0.110

0.047

0.030

0.064

0.059↑

0.039

SD

0.037

0.035

0.066

0.044

0.038

0.088

0.057

0.061

N

30

30

30

30

30

30

30

30

Sex: Female

Group

Dose (mg/kg)

 

Feed Efficiency

0

15

G1

0

Mean

0.039

0.047

SD

0.047

0.056

N

30

30

G2

62.5

Mean

0.057

0.059

SD

0.070

0.057

N

30

30

G3

125

Mean

0.026

0.028

SD

0.044

0.035

N

30

30

G4

250

Mean

0.011

0.039

SD

0.026

0.037

N

30

30

Key: SD: Standard Deviation; N: Number of Animals;: increased as compared to control (P<0.05);↓↓: decreased as compared to control (P<0.01).

Gestation

Sex: Female

Group

Dose (mg/kg)

 

Feed Efficiency

0

15

20

G1

0

Mean

0.229

0.209

0.480

SD

0.102

0.127

0.070

N

30

30

30

G2

62.5

Mean

0.264

0.223

0.465

SD

0.063

0.054

0.095

N

28

28

28

G3

125

Mean

0.236

0.231

0.478

SD

0.078

0.058

0.084

N

28

28

28

G4

250

Mean

0.251

0.226

0.476

SD

0.048

0.066

0.093

N

26

26

26

Non Parturated Dam

Sex: Female

Group

Dose (mg/kg)

Feed Efficiency

0

15

20

27

G1

0

Mean

0.010

0.088

0.027

-0.024

SD

./.

./.

./.

./.

N

1

1

1

1

G2

62.5

Mean

0.043

0.019

-0.004

0.009

SD

0.061

0.286

0.034

0.069

N

2

2

2

2

G3

125

Mean

0.014

0.057

-0.026

0.000

SD

./.

./.

./.

./.

N

1

1

1

1

G4

250

Mean

0.129

0.024

-0.029

0.024

SD

0.045

0.160

0.153

0.069

N

4

4

4

4

Key: SD: Standard Deviation; N: Number of Animals; ./.: not applicable

Table 2.7 Detailed clinical examination

 

 

Sex: Male

 

Group

Dose

(mg/Kg)

Day of Exposure

 

1

8

15

22

29

36

43

50

57

64

71

 

G1

0

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

 

G2

62.5

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

 

G3

125

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

 

G4

250

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

NAD

 

 

 

 

Pre-mating

 

Sex: Female

Group

Dose

(mg/Kg)

Day of Exposure

1

8

15

22

G1

0

NAD

NAD

NAD

NAD

G2

62.5

NAD

NAD

NAD

NAD

G3

125

NAD

NAD

NAD

NAD

G4

250

NAD

NAD

NAD

NAD

 

Gestation

Sex: Female

Group

Dos
  Gestation Sex: Female e

(mg/Kg)

Day of Gestation

0

7

14

20

G1

0

NAD

NAD

NAD

NAD

G2

62.5

NAD

NAD

NAD

NAD

G3

125

NAD

NAD

NAD

NAD

G4

250

NAD

NAD

NAD

NAD

Key: NAD: no abnormality detected

 

Lactation

Sex: Female

Group

Dose

(mg/Kg)

Day of Lactation

0

4

7

14

21

G1

0

NAD

NAD

NAD

NAD

NAD

G2

62.5

NAD

NAD

NAD

NAD

NAD

G3

125

NAD

NAD

NAD

NAD

NAD

G4

250

NAD

NAD

NAD

NAD

NAD

 

Non-Parturated

Sex: Female

Group

Dose

(mg/Kg)

Day of Gestation

0

7

14

20

27

30

34

G1

0

NAD

NAD

NAD

NAD

NAD

NAD

NAD

G2

62.5

NAD

NAD

NAD

NAD

NAD

NAD

NAD

G3

125

NAD

NAD

NAD

NAD

NAD

NAD

NAD

G4

250

NAD

NAD

NAD

NAD

NAD

NAD

NAD

Table 2.8 Mean Hormone levels (T4, TSH, Estradiol, Progesterone, Testesterone)

 

 

Sex: Male

Group

Dose (mg/kg)

 

TSH(UI/ml)

Estradiol (pg/ml)

Progesterone (ng/ml)

Tetsesterone (pg/ml)

T4 (ng/ml)

G1

0

Mean

0.17

114.02

3.74

37.18

81.36

SD

0.06

24.85

0.86

10.58

18.84

N

10

10

10

10

10

G2

62.5

Mean

0.15

100.65

3.46

39.36

64.34

SD

0.03

24.63

0.50

15.81

13.55

N

10

10

10

10

10

G3

125

Mean

0.14

97.43

3.47

32.89

53.49↓↓

SD

0.07

27.86

1.12

12.84

11.01

N

10

10

10

10

10

G4

250

Mean

0.12

104.89

3.97

32.22

69.79

SD

0.03

19.89

1.03

16.61

21.42

N

10

10

10

9

10

 

 

Sex: Female

Group

Dose (mg/kg)

 

TSH(UI/ml)

Estradiol (pg/ml)

Progesterone (ng/ml)

Tetsesterone (pg/ml)

T4 (ng/ml)

G1

0

Mean

0.12

104.81

6.26

52.54

49.27

SD

0.02

27.48

1.34

10.91

8.33

N

10

10

10

10

10

G2

62.5

Mean

0.10

97.80

5.76

65.59

51.60

SD

0.03

24.62

2.07

16.44

6.71

N

10

10

10

10

10

G3

125

Mean

0.12

103.23

5.10

54.71

56.61

SD

0.03

22.84

1.56

16.05

8.09

N

10

10

10

10

10

G4

250

Mean

0.10

84.89

3.99↓↓↓

45.70

47.63

SD

0.02

25.31

1.56

9.51

11.29

N

10

10

10

10

10

Key: N: number of animals, SD: standard deviation; ↓↓: decreased as compared to control (P<0.01); ↓↓↓: decreased as compared to control (P<0.001).

Table 2.9 Hematology

Sex: Male

Group (N)

G1 (10)

G2 (10)

G3 (10)

G4 (10)

Dose (mg/kg)

0

62.5

125

250

Parameters

Mean

SD

Mean

SD

Mean

SD

Mean

SD

 

Total Erythrocyte Count (RBC) (X 1012/L)

8.62

0.49

8.82

0.41

8.47

0.39

8.76

0.48

 

Hematocrit (HCT) (%)

43.61

1.61

44.22

2.05

42.28

2.43

43.41

1.85

 

Mean Corpuscular Volume (MCV) (fL)

50.69

2.78

50.11

1.37

49.90

1.39

49.57

1.97

 

Hemoglobin (HGB) (g/dL)

15.62

0.69

15.84

0.69

15.19

0.90

15.56

0.60

 

Mean Corpuscular Hemoglobin (MCH) (pg)

18.16

0.91

17.95

0.40

17.92

0.47

17.77

0.68

 

Mean Corpuscular Hemoglobin Concentration (MCHC) (g/dL)

35.84

0.45

35.88

0.39

35.93

0.25

35.88

0.66

 

Platelet Count (PLT) (X 109/L)

676.30

63.08

682.20

86.96

686.90

71.09

705.80

49.65

 

Total Leukocyte count (WBC) (X 109/L)

9.33

0.99

10.29

1.84

10.27

3.32

12.50↑↑

1.93

 

Neutrophil (%)

18.20

2.35

18.50

3.24

17.70

2.58

18.00

2.67

 

Lymphocyte (%)

81.30

2.26

81.00

3.16

82.00

2.40

81.10

2.42

 

Monocyte (%)

0.20

0.42

0.30

0.48

0.30

0.48

0.60

0.52

 

Eosinophil (%)

0.30

0.48

0.20

0.42

0.00

0.00

0.30

0.48

 

Basophil (%)

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

 

Prothrombin Time (PT) (Sec.)

22.22

2.43

23.31

1.93

22.98

2.01

22.00

1.11

 

Activated Partial thromboplastin Time (aPTT) (Sec.)

28.45

1.76

27.44

1.12

28.33

1.58

27.07

1.72

 

Reticulocytes (%)

1.00

0.07

1.00

0.05

1.05

0.10

1.00

0.05

 

Keys: SD = Standard Deviation; N = Number of animals; ↑↑: increased as compared to control (P<0.01).

Sex: Female

Group (N)

G1 (10)

G2 (10)

G3 (10)

G4 (10)

Dose (mg/kg)

0

62.5

125

250

Parameters

Mean

SD

Mean

SD

Mean

SD

Mean

SD

 

Total Erythrocyte Count (RBC) (X 1012/L)

7.91

0.52

7.77

0.50

7.99

0.36

7.85

0.22

 

Hematocrit (HCT) (%)

42.17

2.70

41.33

2.51

42.93

1.55

42.41

1.61

 

Mean Corpuscular Volume (MCV) (fL)

53.34

2.41

53.25

1.78

53.71

2.08

54.01

1.54

 

Hemoglobin (HGB) (g/dL)

14.97

0.91

14.79

0.85

15.29

0.43

15.04

0.55

 

Mean Corpuscular Hemoglobin (MCH) (pg)

18.95

0.66

19.06

0.49

19.14

0.61

19.16

0.61

 

Mean Corpuscular Hemoglobin Concentration (MCHC) (g/dL)

35.52

0.71

35.84

0.55

35.67

0.62

35.46

0.27

 

Platelet Count (PLT) (X 109/L)

718.00

82.00

739.20

82.18

718.70

81.04

719.50

58.33

 

Total Leukocyte count (WBC) (X 109/L)

9.36

3.28

11.50

5.01

12.88

3.70

13.14

3.52

 

Neutrophil (%)

18.30

3.30

17.60

2.84

19.40

1.78

16.40

2.01

 

Lymphocyte (%)

81.30

2.98

81.80

2.62

80.00

1.56

83.00

1.83

 

Monocyte (%)

0.20

0.42

0.30

0.48

0.40

0.52

0.40

0.52

 

Eosinophil (%)

0.20

0.42

0.30

0.48

0.20

0.42

0.20

0.42

 

Basophil (%)

0.00

0.00

0.00

0.00

0.00

0.00

0.00

0.00

 

Prothrombin Time (PT) (Sec.)

20.83

1.70

21.08

1.56

21.69

1.53

22.45

2.89

 

Activated Partial thromboplastin Time (aPTT) (Sec.)

28.39

1.53

27.18

1.24

27.17

1.37

28.25

1.42

 

Reticulocytes (%)

1.01

0.10

1.03

0.12

1.08

0.14

1.08

0.12

 

Table 3.1 Clinical Chemistry

Sex: Male

Group (N)

G1 (10)

G2 (10)

G3 (10)

G4 (10)

Dose (mg/kg)

0

62.5

125

250

Parameters

Mean

SD

Mean

SD

Mean

SD

Mean

SD

 

ALT (U/l)

49.95

6.63

44.29

5.47

46.29

7.59

55.30

5.71

 

AST (U/l)

176.42

22.54

171.89

35.76

178.80

34.14

192.26

32.39

 

Albumin (g/dl)

3.85

0.14

4.00

0.17

4.00

0.20

4.07

0.36

 

ALP (U/l)

171.20

15.92

150.88

11.15

201.88

31.82

176.10

21.19

 

T.Protein (g/dl)

6.32

0.63

6.54

0.59

6.45

0.47

6.67

0.52

 

Urea (mg/dl)

49.76

7.90

49.48

3.88

48.13

12.59

51.20

10.64

 

Bile Acid (µmol/l)

21.62

11.11

25.12

10.76

27.38

9.23

23.67

11.07

 

Cholesterol (mg/dl)

53.21

7.92

55.99

7.83

60.08

11.93

58.38

8.37

 

Calcium (mg/dl)

10.11

0.31

10.12

0.60

10.30

0.25

10.37

0.51

 

CK- Nac (U/I)

780.10

183.19

813.90

209.05

803.40

228.85

733.80

177.29

 

Creatinine (mg/dl)

0.53

0.05

0.53

0.02

0.51

0.04

0.51

0.04

 

Glucose (mg/dl)

88.59

11.28

84.55

9.86

88.30

5.54

92.58

12.36

 

GGT(U/I)

4.93

1.81

5.60

2.33

5.12

2.85

5.14

2.60

 

LDH(U/I)

1131.39

225.25

1042.27

286.03

1028.03

238.64

1146.85

248.37

 

Phosphorus(mg/dl)

6.06

0.83

5.62

0.74

6.15

0.65

7.04↑

1.08

 

Triglyceride (mg/dl)

65.59

15.66

65.37

10.46

60.29

6.70

60.00

9.70

 

Total Bilirubin (mg/dl)

0.33

0.16

0.25

0.08

0.29

0.11

0.32

0.05

 

BUN (mg/dl)

23.25

3.69

23.10

1.84

22.50

5.88

23.45

4.92

 

A/G Ratio

1.65

0.39

1.67

0.50

1.68

0.30

1.63

0.42

 

Globulin (g/dl)

2.46

0.57

2.55

0.61

2.45

0.41

2.60

0.50

 

Na (mmol/l)

145.00

2.05

142.40

4.84

147.20

2.44

149.60↑

4.95

 

K (mmol/l)

5.87

0.79

5.99

0.93

5.85

0.52

5.76

0.44

 

Cl (mmol/l)

99.30

2.36

99.10

4.33

101.70

2.45

104.00↑

3.94

 

Sex: Female

Group (N)

G1 (10)

G2 (10)

G3 (10)

G4 (10)

Dose (mg/kg)

0

62.5

125

250

Parameters

Mean

SD

Mean

SD

Mean

SD

Mean

SD

 

ALT (U/l)

80.68

11.99

87.19

18.28

79.58

13.05

88.76

17.95

 

AST (U/l)

184.60

23.52

192.07

21.92

186.66

23.20

192.09

20.06

 

Albumin (g/dl)

3.94

0.35

3.78

0.31

3.88

0.28

3.78

0.33

 

ALP (U/l)

148.25

29.22

132.24

17.25

136.98

20.93

144.91

21.30

 

T.Protein (g/dl)

7.15

0.66

7.02

0.52

6.92

0.67

6.95

0.75

 

Urea (mg/dl)

70.14

7.13

65.41

10.72

64.40

7.58

65.18

12.35

 

Bile Acid (µmol/l)

55.72

13.92

52.76

19.48

50.90

20.35

50.62

17.47

 

Cholesterol (mg/dl)

74.43

10.30

78.96

10.13

80.41

13.27

77.15

12.86

 

Calcium (mg/dl)

9.52

0.30

9.68

0.51

10.02

0.41

9.58

0.51

 

CK- Nac (U/I)

647.00

216.82

660.10

183.83

691.60

180.56

689.30

186.26

 

Creatinine (mg/dl)

1.29

0.26

1.37

0.25

1.31

0.25

1.23

0.26

 

Glucose (mg/dl)

89.82

8.90

96.87

11.33

95.40

12.38

91.79

13.60

 

GGT(U/I)

9.49

2.31

8.08

2.90

8.43

1.69

8.01

1.51

 

LDH(U/I)

1083.00

232.85

988.32

258.29

1108.26

244.02

1120.21

279.06

 

Phosphorus(mg/dl)

8.01

1.03

7.70

1.08

7.87

1.35

7.88

0.58

 

Triglyceride (mg/dl)

61.52

19.01

65.80

12.71

63.31

9.60

67.74

10.02

 

Total Bilirubin (mg/dl)

0.50

0.09

0.64

0.16

0.64

0.26

0.56

0.17

 

BUN (mg/dl)

32.77

3.34

30.58

5.01

30.00

3.49

30.46

5.77

 

A/G Ratio

1.20

0.16

1.19

0.18

1.29

0.15

1.23

0.21

 

Globulin (g/dl)

3.28

0.56

3.23

0.43

3.04

0.45

3.17

0.62

 

Na (mmol/l)

139.80

1.48

139.60

0.70

139.30

2.21

138.70

2.00

 

K (mmol/l)

6.54

0.77

6.21

0.73

6.57

0.79

6.29

0.69

 

Cl (mmol/l)

99.00

2.21

99.50

2.46

100.60

2.01

100.30

2.58

 

Table 3.2 Urinalysis

Sex: Male

Group (N)

G1 (10)

G2 (10)

G3 (10)

G4 (10)

Dose (mg/kg)

0

62.5

125

250

Parameters

Mean

SD

Mean

SD

Mean

SD

Mean

SD

 

Blood / Blood Cell (RBC/µL)

neg

./.

neg

./.

neg

./.

neg

./.

 

Bilirubin (mg/dL)

neg

./.

neg

./.

neg

./.

neg

./.

 

Urobilinogen (mg/dL)

norm

./.

norm

./.

norm

./.

norm

./.

 

Ketone (mg/dL)

neg

./.

neg

./.

neg

./.

neg

./.

 

Protein (mg/dL)

neg

./.

neg

./.

neg

./.

neg

./.

 

Nitrite

neg

./.

neg

./.

neg

./.

neg

./.

 

Glucose (mg/dL)

neg

./.

neg

./.

neg

./.

neg

./.

 

pH

5.90

0.74

5.85

0.47

5.80

0.48

6.00

0.62

 

Specific Gravity

1.022

0.005

1.017

0.006

1.020

0.006

1.019

0.007

 

Leukocytes (WBC/µL)

neg

./.

neg

./.

neg

./.

neg

./.

 

Physical Observations

Volume (mL)

16.40

3.86

16.80

3.05

15.20

2.90

16.80

3.79

 

Colour

Pale Yellow

./.

Pale Yellow

./.

Pale Yellow

./.

Pale Yellow

./.

 

Appearance

Clear

./.

Clear

./.

Clear

./.

Clear

./.

 

Keys: neg:negative; norm: normal

Sex: Female

Group (N)

G1 (10)

G2 (10)

G3 (10)

G4 (10)

Dose (mg/kg)

0

62.5

125

250

Parameters

Mean

SD

Mean

SD

Mean

SD

Mean

SD

 

Blood / Blood Cell (RBC/µL)

neg

./.

neg

./.

neg

./.

neg

./.

 

Bilirubin (mg/dL)

neg

./.

neg

./.

neg

./.

neg

./.

 

Urobilinogen (mg/dL)

norm

./.

norm

./.

norm

./.

norm

./.

 

Ketone (mg/dL)

neg

./.

neg

./.

neg

./.

neg

./.

 

Protein (mg/dL)

neg

./.

neg

./.

neg

./.

neg

./.

 

Nitrite

neg

./.

neg

./.

neg

./.

neg

./.

 

Glucose (mg/dL)

neg

./.

neg

./.

neg

./.

neg

./.

 

pH

5.35

0.47

5.30

0.42

5.45

0.50

5.25

0.42

 

Specific Gravity

1.024

0.002

1.025

0.000

1.024

0.002

1.024

0.002

 

Leukocytes (WBC/µL)

neg

./.

neg

./.

neg

./.

neg

./.

 

Physical Observations

Volume (mL)

14.10

3.60

14.80

2.25

15.60

1.71

16.30

3.16

 

Colour

Pale Yellow

./.

Pale Yellow

./.

Pale Yellow

./.

Pale Yellow

./.

 

Appearance

Clear

./.

Clear

./.

Clear

./.

Clear

./.

 

Keys: neg: negative; norm: normal

Table 3.3 Absolute organ weights

Mean organ weights (g)

Sex: Male

Group

Dose

BW

Brain

Adrenals

P & SV

Testes

Ep

Heart

Liver

Kidneys

Spleen

Thymus

Pituitary

T & PT

G1

0

Mean

432.13

2.1733

0.0653

3.8409

3.2878

1.5106

1.3489

13.2371

2.8793

0.8518

0.3846

0.0148

0.0225

SD

26.85

0.0938

0.0115

0.4526

0.2009

0.1184

0.1111

1.4264

0.4488

0.4390

0.0820

0.0021

0.0031

N

30

30

30

30

30

30

30

30

30

30

30

30

30

G2

62.5

Mean

439.10

2.1894

0.0659

3.7032

3.3069

1.5295

1.3957

13.6968

2.9693

0.7618

0.4097

0.0146

0.0219

SD

27.80

0.1205

0.0104

0.3724

0.2647

0.1288

0.1500

1.7457

0.1780

0.0994

0.0887

0.0017

0.0041

N

30

30

30

30

30

30

30

30

30

30

30

30

30

G3

125

Mean

432.57

2.2122

0.0688

3.5891

3.4090

1.5136

1.3780

13.9272

3.0300

0.8931

0.4029

0.0145

0.0224

SD

32.71

0.0822

0.0088

0.5126

0.2741

0.1457

0.1242

1.3925

0.2507

0.4175

0.0688

0.0020

0.0038

N

30

30

30

30

30

30

30

30

30

30

30

30

30

G4

250

Mean

417.10

2.1821

0.0653

3.5503

3.2489

1.4888

1.3824

13.3019

2.9758

0.7588

0.3749

0.0138

0.0220

SD

36.89

0.0900

0.0101

0.4892

0.4676

0.1595

0.1867

1.7709

0.3089

0.0928

0.0990

0.0016

0.0048

N

30

30

30

30

30

30

30

30

30

30

30

30

30

Mean organ weights (g)

Sex: Female

Group

Dose

BW

Brain

Adrenals

Ovaries

Uterus

Heart

Liver

Kidneys

Spleen

Thymus

Pituitary

T & PT

G1

0

Mean

277.13

2.0061

0.0883

0.1506

0.4639

1.0677

15.8516

2.1307

0.6678

0.2184

0.0137

0.0191

SD

22.36

0.0991

0.0131

0.0368

0.1418

0.1691

3.0913

0.2136

0.1410

0.1030

0.0021

0.0039

N

30

30

30

30

30

30

30

30

30

30

30

30

G2

62.5

Mean

280.60

2.0174

0.0861

0.1476

0.4310

1.0576

16.0515

2.0773

0.6261

0.2262

0.0139

0.0179

SD

22.07

0.1135

0.0114

0.0252

0.1315

0.0815

3.4609

0.1922

0.0792

0.0813

0.0019

0.0042

N

30

30

30

30

30

30

30

30

30

30

30

30

G3

125

Mean

278.37

1.9954

0.0873

0.1484

0.4800

1.0546

16.2290

2.0188

0.7056

0.2027

0.0137

0.0187

SD

24.92

0.1270

0.0106

0.0308

0.1354

0.1175

3.4161

0.3792

0.4207

0.0728

0.0020

0.0036

N

30

30

30

30

30

30

30

30

30

30

30

30

G4

250

Mean

271.83

1.9932

0.0875

0.1493

0.4865

1.0543

13.7392↓

2.0776

0.6942

0.2307

0.0146

0.0194

SD

21.92

0.0826

0.0117

0.0227

0.1652

0.1286

2.8704

0.3441

0.2913

0.0681

0.0023

0.0032

N

30

30

30

30

30

30

30

30

30

30

30

30

Key: BW = Body Weight; Ep = Epididymis; N = Number of samples; P & SV = Prostate and Seminal Vesicle; SD = Standard deviation; T & PT = Thyroid & Parathyroid. Dose levels are in terms of mg/kg body weight; ↓: decreased as compared to control (P<0.05).

Table 3.4 Relative organ weights

Mean organ weight relative body weight (%)

Sex: Male

Group

Dose

Brain

Adrenals

P & SV

Testes

Ep

Heart

Liver

Kidneys

Spleen

Thymus

Pituitary

T & PT

G1

0

Mean

0.5046

0.0152

0.8920

0.7624

0.3504

0.3126

3.0619

0.6669

0.1975

0.0892

0.0034

0.0052

SD

0.0354

0.0028

0.1175

0.0478

0.0291

0.0239

0.2536

0.1008

0.1037

0.0192

0.0004

0.0008

N

30

30

30

30

30

30

30

30

30

30

30

30

G2

62.5

Mean

0.4999

0.0151

0.8449

0.7541

0.3487

0.3178

3.1250

0.6776

0.1740

0.0932

0.0033

0.0050

SD

0.0321

0.0027

0.0845

0.0549

0.0255

0.0270

0.3947

0.0416

0.0239

0.0191

0.0004

0.0010

N

30

30

30

30

30

30

30

30

30

30

30

30

G3

125

Mean

0.5141

0.0160

0.8327

0.7901

0.3509

0.3190

3.2271

0.7014

0.2081

0.0935

0.0034

0.0052

SD

0.0411

0.0022

0.1216

0.0609

0.0336

0.0226

0.3076

0.0434

0.1036

0.0167

0.0005

0.0010

N

30

30

30

30

30

30

30

30

30

30

30

30

G4

250

Mean

0.5266

0.0157

0.8536

0.7834

0.3589

0.3318

3.1888

0.7142

0.1831

0.0903

0.0033

0.0053

SD

0.0451

0.0023

0.1126

0.1228

0.0442

0.0393

0.3091

0.0526

0.0258

0.0245

0.0004

0.0013

N

30

30

30

30

30

30

30

30

30

30

30

30

Mean organ weight relative body weight (%)

Sex: Female

Group

Dose

Brain

Adrenals

Ovaries

Uterus

Heart

Liver

Kidneys

Spleen

Thymus

Pituitary

T & PT

G1

0

Mean

0.7279

0.0321

0.0545

0.1686

0.3855

5.7142

0.7716

0.2431

0.0796

0.0049

0.0069

SD

0.0621

0.0055

0.0136

0.0532

0.0518

0.9615

0.0797

0.0590

0.0388

0.0007

0.0015

N

30

30

30

30

30

30

30

30

30

30

30

G2

62.5

Mean

0.7224

0.0308

0.0525

0.1529

0.3777

5.7142

0.7414

0.2233

0.0809

0.0050

0.0064

SD

0.0596

0.0040

0.0067

0.0424

0.0239

1.0950

0.0554

0.0246

0.0299

0.0008

0.0014

N

30

30

30

30

30

30

30

30

30

30

30

G3

125

Mean

0.7208

0.0315

0.0535

0.1743

0.3793

5.8108

0.7278

0.2525

0.0731

0.0050

0.0067

SD

0.0619

0.0037

0.0109

0.0534

0.0310

1.0410

0.1324

0.1423

0.0277

0.0009

0.0011

N

30

30

30

30

30

30

30

30

30

30

30

G4

250

Mean

0.7371

0.0324

0.0552

0.1794

0.3877

5.0395↓

0.7652

0.2558

0.0848

0.0054

0.0072

SD

0.0590

0.0053

0.0088

0.0600

0.0325

0.8677

0.1217

0.1115

0.0238

0.0008

0.0013

N

30

30

30

30

30

30

30

30

30

30

30

Key: Ep = Epididymis; N = Number of samples; P & SV = Prostate and Seminal Vesicle; SD = Standard deviation; T & PT = Thyroid & Parathyroid. Dose levels are in terms of mg/kg body weight; ↓: decreased as compared to control (P<0.05).

Table 3.5 Gross Pathology

 

Sex: Male

Group

G1

G2

G3

G4

Dose (mg/kg)

0

62.5

125

250

Total No. of Animals Observed

30

30

30

30

No. of animals with Abnormalities

0

2

1

1

No. of animals with No Abnormalities

30

28

29

29

Organ & Lesion

Testes

 

 

 

 

Reduced sizeTotal

0

1

0

1

Mild

0

0

0

1

Moderate

0

1

0

0

Prostate

 

 

 

 

Increased sizeTotal

0

1

0

0

Minimal

0

1

0

0

Reduced sizeTotal

0

0

1

0

Moderate

0

0

1

0

 

Sex: Female

Group

G1

G2

G3

G4

Dose (mg/kg)

0

62.5

125

250

Total No. of Animals Observed

30

30

30

30

No. of animals with Abnormalities

0

0

0

0

No. of animals with No Abnormalities

30

30

30

30

Table 3.6 Individual Animal Microscopic Observations

 

Sex: Male

Group

G1

G2

G3

G4

Dose (mg/kg)

0

62.5

125

250

Total No. of Animals Observed

30

30

30

30

No. of animals with Abnormalities

11

X

X

5

No. of animals with No Abnormalities

19

X

X

25

Organ & Lesion

Liver

 

Lymphocyte infiltrationTotal

2

X

X

1

Focal Minimal

0

X

X

1

Focal Mild

2

X

X

0

Kidneys

 

Tubular dilation             Total

3

X

X

0

Focal Mild

2

X

X

0

Focal Moderate

1

X

X

0

Lymphocyte infiltration    Total

1

X

X

0

Focal Mild

1

X

X

0

Lungs

 

Lymphocyte infiltration    Total

3

X

X

2

Multifocal Mild

2

X

X

1

Multifocal Moderate

1

X

X

1

Aorta

 

Aneurysm                      Total

1

X

X

1

Focal Mild

1

X

X

0

Focal Moderate

0

X

X

1

Colon

 

Lymphocyte infiltration    Total

1

X

X

0

Multifocal Mild

1

X

X

0

Thymus

 

LymphophagocytosisTotal

1

X

X

0

Multifocal Minimal

1

X

X

0

Thyroid

 

Ultimobranchial cystTotal

1

X

X

1

Focal Mild

1

X

X

1

Adrenals

 

Cytoplasmic vacuolationTotal

1

X

X

0

Diffuse Mild

1

X

X

0

Accessory adrenocortical tissueTotal

1

X

X

0

Unilateral

1

X

X

0

 

Sex: Female

Group

G1

G2

G3

G4

Dose (mg/kg)

0

62.5

125

250

Total No. of Animals Observed

30

30

30

30

No. of animals with Abnormalities

12

X

X

8

No. of animals with No Abnormalities

18

X

X

22

Organ & Lesion

Liver

 

Lymphocyte infiltration    Total

0

X

X

1

Focal Minimal

0

X

X

1

Kidneys

 

Tubular dilation             Total

1

X

X

0

Multifocal Mild

1

X

X

0

Lymphocyte infiltration    Total

2

X

X

1

Focal Minimal

0

X

X

1

Focal Mild

1

X

X

0

Multifocal Mild

1

X

X

0

Lungs

 

Lymphocyte infiltration    Total

1

X

X

2

Focal Mild

0

X

X

1

Multifocal Mild

1

X

X

1

Alveolar histiocytosis    Total

1

X

X

1

Focal Mild

0

X

X

1

Multifocal Mild

1

X

X

0

Aorta

 

Aneurysm                      Total

1

X

X

0

Focal Mild

1

X

X

0

Thymus

 

LymphophagocytosisTotal

1

X

X

1

Multifocal Mild

1

X

X

1

Atrophy                     Total

1

X

X

0

Focal Mild

1

X

X

0

Thyroid

 

Ultimobranchial cystTotal

2

X

X

1

Focal Mild

2

X

X

0

Focal Moderate

0

X

X

1

Adrenals

 

 

 

 

Accessory adrenocortical tissueTotal

2

X

X

2

Unilateral

2

X

X

2

Conclusions:
Based on all the available data, it was observed that there were no effects observed on male and female animals up to 125 mg/kg bw (Mid Dose-G3) of the test chemical administration. However, at 250 mg/kg bw (High Dose-G4) dose group, clinical signs, effects on weights of liver and effects on hormonal levels was observed, due to test chemical administration. Therefore, it was concluded that the NOAEL and LOAEL of the test chemical was 125 mg/kg bw and 250 mg/kg bw, respectively.
Executive summary:

An Extended One-Generation Reproductive Toxicity Study was performed according to OECD TG 443 to evaluate the general, reproductive and developmental toxicological endpoints associated with the repeated oral administration of graduated doses of the test chemical in Wistar rats. A total of 240 wistar rats (120 males and 120 females) were randomized into 4 experimental groups each containing 30 animals per sex per group. The groups, viz., G1, G2, G3 and G4 received 0, 62.5, 125 and 250 mg/kg body weight of test chemical respectively. Dose administration was through an oral gavage and the vehicle used in this study was corn oil. All animals of both sexes were dosed 2 weeks prior to mating. Dosing was continued in both sexes during the mating period. Females were treated throughout gestation and lactation up to the day of termination after weaning. Males were treated in the same manner until termination. During observations, it was seen that no treatment-related clinical signs or symptoms were observed in any of the parent animals up to 125 mg/kg body weight of the test chemical. However, mild salivation was observed in total 13 of 30 males of G4 group (250 mg/kg), starting from the 27th day of treatment and continuing till termination. All adult animals were found normal with respect to all the parameters examined during detailed clinical examinations through the course of the study. No treatment-related symptoms were found up to the dose level of 250 mg/kg body weight in either sex. No morbidities or mortality were recorded in adult animals of parent generation in any of the experimental groups throughout the duration of the experiment. Mean body weights remained comparable among all the groups for every time point (days 8, 15, 22, 29, 36, 43, 50, 57, 64 and 71) recorded during the 70 days exposure period for males. No significant difference was found for the mean body weights of females during the two weeks pre-mating period, during mating, during gestation or during lactation period. However, the mean percent change in body weight on all the instances (days 8, 15, 22, 29, 36, 43, 50, 57, 64 and 71) with respect to the first day of exposure was significantly reduced for parental males of G4 group (250 mg/kg) (P≤0.05 to P≤0.001) as compared to control animals, whereas mean body weight change for G2 (62.5 mg/kg) and G3 (125 mg/kg) males remained comparable to control animals. Among the females, mean percent change in body weights remained statistically comparable among all groups during pre-mating, mating, gestation and lactation periods. The mean feed consumed per animal per day significantly increased on day 8 of treatment for G2 (62.5 mg/kg) (P≤0.01) and G3 (125 mg/kg) (P≤0.05) males as compared to G1 (control) animals. However, it was found to be similar for all other time points across the groups. The mean feed consumption of treatment group (G2, G3 & G4) females remained statistically comparable to control group during pre-mating, mating, gestation and lactation period except for one instance, where mean feed consumption on day 8 of treatment was significantly reduced for G2 (62.5 mg/kg) (P≤0.01) as compared to control females. The differences found in mean feed consumption of male and female animals were neither dose-dependent nor consistent, hence it was deemed to be independent of test-item treatment. Feed efficiency for the G4 males had statistically reduced in the first two weeks of treatment, i.e. on days 8 and 15. It was also found to have increased significantly during the penultimate week of treatment, i.e. on day 63. For all other groups among the males, the feed efficiency was statistically comparable across the groups. Among the parental females, no statistical differences were observed for feed efficiency in any of the groups. No significant difference was observed in any of the measured hematology parameters in all the groups of both the sexes, except in the WBC count, which was found to be significantly increased in males of G4 group (P≤0.01) as compared to control males. Significant increase was observed in Phosphorus, Sodium and Chloride levels in G4 males (P≤0.05) as compared to control males. All the other clinical chemistry parameters in males and females were found comparable in control and treatment groups. The urine parameters tested did not show up any significant changes among the experimental groups in the parental generation of either sex. No significant difference was observed in absolute organ weight or organ weight relative to body weight for the entire set of measured organs in parent male and female animals. However, significant reduction was observed in absolute and relative liver weight in G4 females (P<0.05) as compared to control animals. In the current study, however, the decreased liver weights with respect to body weights in G4 females does not correlate with any histological observations made, making it inconsequential from the point of view of toxic effects of test-chemical. External examination of all male and female rats of control and all treated groups did not reveal any lesion of pathological significance. Internal examination of the rats revealed reduced testicular size in two animals (Male: G2: 1/30, G4:1/30), increased prostate size in one animal (Male: G2: 1/30) and reduced prostate size in one animal (Male: G3: 1/30). Remaining animals of parent generation did not reveal any abnormality of pathological significance. No treatment-related histopathological findings are reported in this study that could arise out of test-item administration. All observed tissues were normal at the level of histology. Minimal to moderate and focal lymphocytic infiltrates were observed in some slides of liver, kidney, lung, heart, colon, thymus and adrenals with mild to moderate cyst in thyroid gland, which were distributed randomly across the experimental groups and did not show any pattern of either dose-dependency or sex-based selectivity. While TSH, Estradiol, Progesterone and Testosterone remained statistically comparable among the groups of parental males, the mean level of T4 was found reduced in G3 males (P<0.01) as compared to control. This is likely an incidental result, given that G4 was statistically comparable to control. The mean T4, TSH, Estradiol and testosterone levels remained comparable among the groups of parental females; whereas, Progesterone was found to be significantly reduced in G4 females (P<0.001). This could be a test-chemical related effect. Based on all the available data, it was observed that there were no effects observed on male and female animals up to 125 mg/kg bw (Mid Dose-G3) of the test chemical administration. However, at 250 mg/kg bw (High Dose-G4) dose group, clinical signs, effects on weights of liver and effects on hormonal levels was observed, due to test chemical administration. Therefore, it was concluded that the NOAEL and LOAEL of the test chemical was 125 mg/kg bw and 250 mg/kg bw, respectively.

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Data is form Study report
Qualifier:
according to guideline
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
Principles of method if other than guideline:
According to OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity in Rodents)
GLP compliance:
yes
Limit test:
no
Specific details on test material used for the study:
- Name of test material (as cited in study report): Methyl 2-naphthyl ether (MNE) (Batch no. 0001)
- Molecular formula (if other than submission substance): C11H10O
- Molecular weight (if other than submission substance): 158.19 g/mol
- Substance type: Organic
- Physical state: Solid
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
Details on test animal
TEST ANIMALS
- Source: In-house bred
- Age at study initiation: 7 to 8 weeks old
- Weight at study initiation: Male 184.38-234.56 g, Female 176.90-208.56 g
- Fasting period before study: No data available
- Housing: Four rats per sex per cage were housed in sterilized solid bottom polypropylene cages with stainless steel grill tops with bedding of clean paddy husk. The cages were suspended on stainless steel racks in a controlled environment.
- Diet (e.g. ad libitum): Standard laboratory sterile extruded pelleted rodent feed (Provimi Animal Nutrition India Pvt. Ltd, Bangalore, India), ad libitum
- Water (e.g. ad libitum): Potable tap water filtered through Reviva Reverse Osmosis System (water filter cum purifier), ad libitum.
- Acclimatization period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3°C
- Humidity (%):30-70 %
- Air changes (per hr): 25 ± 5 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours dark/12 hours light

IN-LIFE DATES:
From: 11.04.2014 (Male), 13.04.2014 (Female);
To: 17.05.2014 (Male), 19.05.2014 (Female)
Route of administration:
oral: gavage
Vehicle:
other: Groundnut oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: 125, 250 and 500 mg was weighed and dissolved in 10 ml of groundnut oil by vortexing the resultant (concentration / dose).

DIET PREPARATION
- Rate of preparation of diet (frequency): No data available
- Mixing appropriate amounts with (Type of food): No data available
- Storage temperature of food: No data available

VEHICLE
- Justification for use and choice of vehicle (if other than water): Groundnut oil
- Concentration in vehicle: 0, 125, 250 and 500 mg/kg body weight/day
- Amount of vehicle (if gavage): 5 ml/kg body weight
- Lot/batch no. (if required): No data available
- Purity: No data available
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Doses were analyzed by using HPLC system.
Duration of treatment / exposure:
28 days
Frequency of treatment:
Daily
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Remarks:
Control Group
Dose / conc.:
125 mg/kg bw/day (actual dose received)
Remarks:
Low Dose Group
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Remarks:
Mid Dose Group
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Remarks:
High Dose Group
No. of animals per sex per dose:
Total animals: 56
Control: 7 males, 7 females
125 mg/kg/day: 7males, 7 females
250 mg/kg/day: 7 males, 7 females
500 mg/kg/day: 7 males, 7 females
Control animals:
yes, concurrent vehicle
Details on study design:
Details on study design
- Dose selection rationale: No data available
- Rationale for animal assignment (if not random): Animals were randomized by body weight and sex.
- Rationale for selecting satellite groups: No data available
- Post-exposure recovery period in satellite groups: No data available
- Section schedule rationale (if not random): No data available
Positive control:
No Data Available
Observations and examinations performed and frequency:
Observations and examinations performed & frequency

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily
- Cage side observations performed: All cages were checked twice daily (morning and evening) for dead or moribund animals, in order to allow necropsy examination to be carried out immediately at the same day.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: The rats were daily subjected to general cage side clinical examinations to characterize onset and duration of clinical signs. All signs of behavioral changes or reaction to treatment were recorded for individual animals. However, detailed physical and clinical observations were conducted before the first exposure of test item and at least once a week thereafter till the end of the study. Appearances of clinical signs were recorded on weekly clinical observations and detailed abnormality observation sheets for individual animals.

BODY WEIGHT: Yes
- Time schedule for examinations: Individual body weights was recorded on day 1, 8, 15, 22, 28 (before fasting), and day 29 (before sacrifice) during the entire course of treatment.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Food consumption was recorded weekly once (days 2, 9, 16 & 23) during the entire course of treatment. The measured quantity of food was offered in each cage and the left quantity was measured after 24 hrs and finally food intake per rat was calculated.
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data available
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data available

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 data available

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: Once weekly (on days 2, 9, 16 & 23)

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Fourth week of chemical treatment.
- Dose groups that were examined: All 56 animals were examined.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: On completion of 28 days of treatment and prior to necropsy.
- Anaesthetic used for blood collection: Yes
- Animals fasted: Yes, overnight fasting.
- How many animals: All 56 animals were examined.
- Parameters examined: Haemoglobin (Hb) , RBC Count, Total and differential leucocyte count, Haematocrit (Hct /PCV), Mean corpuscular volume (MCV), Mean corpuscular haemoglobin (MCH), Mean corpuscular haemoglobin concentration (MCHC) and Platelet Count.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: On completion of 28 days of treatment and prior to necropsy.
- Animals fasted: Yes, overnight fasting.
- How many animals: All 56 animals were examined.
- Parameters examined: Sodium, Potassium, Glucose, Total Cholesterol, Blood Urea, Creatinine, Total Protein Albumin, SGPT (Serum glutamic pyruvic transaminase)/ALT, SGOT (Serum glutamic oxaloacetic transaminase)/AST, Hormones analysis (testosterone and estrogen) and Total Bile acids.

URINALYSIS: No data available
- Time schedule for collection of urine: No data available
- Metabolism cages used for collection of urine: No data available
- Animals fasted: No data available
- Parameters examined: No data available

NEUROBEHAVIOURAL EXAMINATION: No data available
- Time schedule for examinations: No data available
- Dose groups that were examined: No data available
- Battery of functions tested: sensory activity / grip strength / motor activity / other: Yes, Locomotor activity was observed.

OTHER:
Organ weight were measured.
Sacrifice and pathology:
Sacrifice and pathology
GROSS PATHOLOGY: Yes
On completion of treatment, all surviving rats were sacrificed and a complete necropsy was carried out on all animals. Tissues were collected from brain, stomach, large and small intestine, liver, kidneys, adrenal gland(s), spleen, heart, thymus, lungs, testis/ovary, uterus, lymph nodes, peripheral nerve (sciatic), bone marrow, and other gross lesions, if any.
Tissues were preserved in 10% formal saline. However testes, ovaries and uterus were first fixed in Bouin’s fixative for two hours then transferred to 10% formal saline.

HISTOPATHOLOGY: Yes
Histological examination was conducted on tissues/organs from the control and the high-dose group animals Organ examined: lung, liver, kidney, heart, spleen, testis, ovaries, adrenal glands, large and small intestine, brain, sciatic nerve, lymph nodes, bone marrow, stomach and thymus were examined.
Other examinations:
The collected organs were also weighed.
Statistics:
Statistical analysis was carried out by using Microsoft Excel and IBM SPSS statistics version -20. All analyses and comparisons were evaluated at the 5 % level, statistically significant differences (p>0.05) indicated by appropriate notation. PAIRED T-Testing procedure was used to check the significance between above mentioned groups. For multiple comparisons Turkey’s HSD test was applied.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Abnormalities such as nasal discharge, red crust around the nostrils was observed in all treatment groups. In addition, a few cases of snuffling, eye lid swelling and hunched back posture were also observed in the treated groups.
Mortality:
mortality observed, treatment-related
Description (incidence):
Two animals at high dose groups died during the course of the study.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Treatment with 500 mg/kg body weight/day showed a significant reduction in body weights in male rats on the first, second and fourth week of treatment. In female rats, a significant decrease in body weight was observed on the first, second and third week of treatment.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
No difference in the feed consumption was observed among the groups.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
no effects observed
Description (incidence and severity):
No difference in water consumption was observed among the groups.
Ophthalmological findings:
no effects observed
Description (incidence and severity):
No difference was observed in opthalmoscopic examination.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
In male rats, significant increase in the platelet count was observed along with decrease in MCV and MCH levels at 125 mg/kg body weight/day.
Treatment with 250 mg/kg body weight/day showed significantly decreased values of hemoglobin and MCHC in male rats. The level of MCHC was also found to be significantly decreased when treated with 500 mg/kg body weight/day.
In female rats, treatment with 125 mg/kg body weight/day showed significant alterations in platelet count and MCV.
At 250 mg/kg body weight/day, a significant decrease was noticed in neutrophils along with altered values of MCV in female rats.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
In female animals, the level of total bile acid was decreased at 250 mg/kg body weight/day.
A significant increase was noticed in the level of estrogen in the same group. A similar trend was also noticed at 125 and 500 mg/kg body weight/day., but was statistically non-significant.
At 250 and 500 mg/kg body weight/day the levels of potassium and albumin were found to be significantly increased. The cholesterol level in the 250 mg/kg body weight/day group was noticed to be significantly elevated in the mid-dose group.
In male rats, the level of testosterone was significantly increased after treatment with 500 mg/kg body weight/day.
Urinalysis findings:
not specified
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
No statistically significant difference was observed in the motor activity scores for any dose group when compared with the control animals.
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
The weight of spleen was decreased in the 250 mg/kg body weight/day groups while treatment with 500 mg/kg body weight/day resulted in the decreased weight of kidney in both female and male rats. However, the absolute organ weight of male animals does not show any significant difference.

In male rats, the relative weight of liver was noticed to be increased at 125 mg/kg body weight/day. Significant increase in the relative weights of heart and testes was also noticed at 500 mg/kg body weight/day.

In female rats, at all dose levels, the relative weight of ovaries was decreased in comparison to the control group animals. At 250 mg/kg body weight/day, the relative weights of brain and heart were significantly decreased. The relative weight of uterus was also decreased in the 125 or 250 mg/kg body weight/day groups.A significant decrease in the relative weight of spleen was observed at250 or 500 mg/kg body weight/day.

The absolute organ weight of ovaries from all treated groups was found to be decreased significantly in female rats.

The weight of uterus was found to be significantly decreased in 250 or 500 mg/kg body weight/day-treated male and female rats.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Nasal discharge, red crust around nostril, perineum wet, bloated stomach, viscous oily secretion was found inside the thoracic cavity.

Small white solid mass slightly firm in consistency floating inside the urinary bladder was observed.

However, due to adaptive metabolic and physiological changes, anoxic/ hypoxic conditions during anesthesia and terminal sacrifice, these findings were considered to be of no toxicological significance.
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
No remarkable changes observed in control and the 500 mg/kg body weight/day-treated animals.

A few microscopic findings observed in 500 mg/kg body weight/day-treated animals included collapsed lung with focal inflammation, focal fatty change and excess of lymphocytes in liver, inflammation in small intestine, reactive spleen and excess of mucous in colon. However, these findings were also observed in the control animals.
Histopathological findings: neoplastic:
not specified
Other effects:
not specified
Dose descriptor:
LOAEL
Effect level:
250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
clinical biochemistry
haematology
organ weights and organ / body weight ratios
other: Effects observed
Dose descriptor:
NOAEL
Effect level:
125 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
clinical biochemistry
clinical signs
food consumption and compound intake
gross pathology
haematology
histopathology: non-neoplastic
mortality
ophthalmological examination
organ weights and organ / body weight ratios
water consumption and compound intake
other: No effect observed
Critical effects observed:
not specified
System:
other: not specified
Organ:
not specified

SUMMARY OF BODY WEIGHTS(g)

Male

 

 

Group

 

Dose mg/kg

 

 

 

Day1

 

 

Day8

 

 

Day15

 

 

Day22

 

 

Day28

 

Terminal

Day

 

 

1

 

 

Vehicle

Mean

211.80

242.34

262.97

283.01

300.76

296.93

S.D.

14.03

14.91

16.29

24.57

26.41

27.19

 

S.E.M.

6.62

6.15

6.20

8.68

8.78

9.16

 

 

2

 

 

125

Mean

218.02

243.85

260.27

279.35

296.61

291.98

S.D.

9.56

11.42

11.59

14.54

14.88

14.86

 

S.E.M.

4.38

4.68

4.45

5.21

5.02

5.09

 

 

3

 

 

250

Mean

210.57

221.01

241.60

267.42

286.08

282.17

S.D.

9.82

26.98

30.11

23.54

25.05

24.52

 

S.E.M.

4.67

12.21

12.46

8.80

8.76

8.69

 

 

4

 

 

500

Mean

214.35

212.06

223.41

240.97

256.81

256.24

S.D.

10.64

10.08

11.64

13.72

6.86

9.27

 

S.E.M.

4.96

4.76

5.21

5.70

     2.67

3.62

 

SUMMARY OF BODYWEIGHTS(g)

Female

 

 

Group

 

Dose mg/kg

 

 

 

Day1

 

 

Day8

 

 

Day15

 

 

Day22

 

 

Day28

 

Terminal

Day

 

 

1

 

 

Vehicle

Mean

191.17

200.82

208.91

215.94

227.15

219.85

S.D.

7.12

12.39

16.56

15.30

17.64

19.32

 

S.E.M.

3.72

6.17

7.93

7.09

7.77

8.79

 

 

2

 

 

125

Mean

192.17

201.14

208.56

214.38

220.79

216.51

S.D.

9.31

10.16

8.58

7.62

10.76

10.09

 

S.E.M.

4.85

5.05

4.11

3.55

4.87

4.66

 

 

3

 

 

250

Mean

194.94

202.08

209.12

216.24

228.85

226.38

S.D.

9.45

8.89

8.24

7.85

7.51

8.75

 

S.E.M.

4.85

4.40

3.94

3.63

3.28

3.87

 

 

4

 

 

500

Mean

190.49

182.52

192.46

205.74

216.14

212.37

S.D.

8.82

9.74

12.73

11.46

11.61

12.10

 

S.E.M.

4.63

5.34

6.61

5.57

5.37

5.70

SUMMARY OF HEMATOLOGY DATA Male

Group/Dose

 

WBC

3

(x10/

mm3)

RBC

6

(x10/

mm3)

PLT (x10/mm3)

Hct

(%)

Hgb

(g/dl)

MCV(fl)

MCH (pg)

MCHC(g/dl)

Lym

%

Mon

%

Nut

%

Eos

%

 

Bas%

 

 

1/0

 

Mean

12.06

69.49

4.34

23.27

1.17

0.99

10.15

52.77

53.54

16.40

31.07

16.67

600.2

9

±S.D.

2.24

9.56

1.11

9.04

1.07

0.33

0.63

1.40

2.92

0.51

0.97

1.00

76.78

± S.E.M.

18.59

13.76

25.46

38.85

91.51

33.35

6.17

2.65

5.45

3.11

3.12

5.99

12.79

 

 

 

2/125mg/kg

 

Mean

12.09

69.73

4.57

22.04

2.19

0.71

10.16

50.24*

51.00

15.49*

30.83

15.73

1118.

00*

 

±S.D.

2.18

5.30

0.71

6.81

2.70

0.17

0.53

1.93

2.36

0.71

0.83

0.80

383.1

9

±

S.E.M.

0.8

2.0

0.3

2.6

1.0

0.1

0.2

0.7

0.9

0.3

0.3

0.3

144.8

 

 

 

3/250mg/kg

 

Mean

14.97

63.19

4.66

28.04

2.36

0.93

9.93

52.76

52.41

15.81

29.99

15.70

614.8

6

 

±S.D.

2.34

11.26

0.75

12.16

2.83

0.20

0.24

2.50

3.03

0.80

0.18

0.96

173.0

5

±

S.E.M.

0.9

4.3

0.3

4.6

1.1

0.1

0.1

0.9

1.1

0.3

0.1

0.4

65.4

 

 

 

4/ 500mg/kg

 

Mean

9.10

53.17

3.28

16.91

2.29

0.60

7.32

38.41

39.25

11.61

21.62

11.87

476.4

9

 

±S.D.

5.76

31.42

2.26

6.52

1.66

0.35

4.89

25.12

25.52

7.57

14.70

7.70

295.4

8

± S.E.M.

2.6

14.1

1.0

2.9

0.7

0.2

2.2

11.2

11.4

3.4

6.6

3.4

132.1

 

 

SUMMARY OF HEMATOLOGY DATA Female

emaleGroup/Dose

 

WBC (x103/mm3)

RBC (x106/mm3)

PLT (x103/mm3)

 

Hct

(%)

 

Hgb

(g/dl)

 

MCV(fl)

 

MCH (pg)

 

MCHC (g/dl)

 

Lym

%

 

Mon

%

 

Nut

%

 

Eos

%

 

Bas%

 

 

Vehicle

Mean

11.66

62.20

4.41

30.31

1.39

0.87

10.18

49.90

50.74

15.44

30.97

15.71

994.57

±S.D.

2.19

8.62

1.09

8.69

1.61

0.17

0.55

2.34

2.56

0.68

0.31

0.84

520.25

± S..E..M.

 

0.8

 

3.3

 

0.4

 

3.3

 

0.6

 

0.1

 

0.2

 

0.9

 

1.0

 

0.3

 

0.1

 

0.3

 

196.6

 

 

2      /

125mg/kg

 

Mean

 

12.28

 

67.97

 

4.34

 

25.10

 

0.94

 

0.90

 

10.13

 

51.66*

 

52.26

 

15.86

 

30.71

 

16.06

518.71

±S.D.

4.51

6.98

0.78

6.78

0.47

0.18

0.47

1.36

1.39

0.73

0.71

0.45

114.08

±S.E.M.

 

1.7

 

2.6

 

0.3

 

2.6

 

0.2

 

0.1

 

0.2

 

0.5

 

0.5

 

0.3

 

0.3

 

0.2

 

43.1

 

 

3    / 250mg/kg

Mean

11.94

69.24

4.13

21.83*

2.93

1.07

10.18

51.76*

52.76

15.71

30.37

15.99

585.00

±S.D.

3.21

7.96

0.54

7.77

2.71

0.24

0.45

1.55

3.36

0.75

1.44

1.17

213.78

±S.E.M.

 

1.2

 

3.0

 

0.2

 

2.9

 

1.0

 

0.1

 

0.2

 

0.6

 

1.3

 

0.3

 

0.5

 

0.4

 

80.8

 

 

4       /

500mg/kg

Mean

11.37

66.26

4.36

26.57

1.06

1.03

10.00

51.77

51.79

15.79

30.47

15.79

614.29

±S.D.

2.10

9.91

0.99

9.88

1.34

0.19

0.73

0.85

3.90

0.57

0.92

1.41

139.13

±S.E

.M.

 

0.8

 

3.7

 

0.4

 

3.7

 

0.5

 

0.1

 

0.3

 

0.3

 

1.5

 

0.2

 

0.3

 

0.5

 

52.6

 

SUMMARY OF CLINICAL BIOCHEMISTRY DATA Male Rats

Group/Dose

 

Sod.(mmol/L)

Pot.(mmol/L)

ALB(g%)

CHOL(mg%)

CRT(mg%)

SGOT(IU/L)

SGPT (IU/L)

GLU(mg%)

TP(g%)

BUN(mg%)

TBA

Test

ng/L

nmol/L

 

 

1   /  0

Mean

131.5

7.0

3.9

83.9

0.9

33.1

32.9

117.0

6.5

12.5

490.1

4.363

±S.D.

8.6

0.5

0.3

18.1

0.2

4.0

5.1

7.1

0.4

3.0

44.82

1.02

±S.E.M.

3.2

0.2

0.1

6.8

0.1

1.5

1.9

2.7

0.1

1.1

16.94

0.387

 

 

 

2  /125 mg/kg

Mean

130.0

7.6

3.7

87.6

0.9

33.9

32.5

119.1

6.5

14.4

485.4

5.421

±S.D.

3.3

0.7

0.3

10.7

0.2

8.2

7.6

4.9

0.5

3.0

36.17

1.03

±S.E.M.

1.2

0.3

0.1

4.0

0.1

3.1

2.9

1.9

0.2

1.1

13.67

0.392

 

 

 

 

 

3 /250 mg/kg

Mean

130.1

7.1

3.7

87.7

0.9

65.4

30.0

121.0

6.7

12.1

465.4

4.716

±S.D.

9.8

1.0

0.2

17.4

0.1

90.0

7.0

9.5

0.5

2.8

35.10

1.28

±S.E.M.

3.7

0.4

0.1

6.6

0.1

34.0

2.6

3.6

0.2

1.1

13.26

0.487

 

 

 

4 / 500 mg/kg

 

Mean

 

124.5

 

11.0

 

3.7

 

96.2

 

0.9

 

31.1

 

30.5

 

121.6

 

6.4

 

13.7

485.2

*

 

6.289*

±S.D.

11.1

8.4

0.3

12.1

0.2

7.5

6.4

13.8

0.5

4.0

23.33

0.398

 

±S.E.M.

 

5.0

 

3.8

 

0.1

 

5.4

 

0.1

 

3.3

 

2.8

 

6.2

 

0.2

 

1.8

 

10.43

 

0.178

 

SUMMARY OF CLINICAL BIOCHEMISTRY DATA - Female

 

Group

/Dose

 

Sod.

Pot.

ALB

CHOL

CRT

SGOT

SGPT

GLU

TP

BUN

TBA

Est

(mmol/L)

(mmol/L)

(g%)

(mg%)

(mg%)

(IU/L)

(IU/L)

(mg%)

(g%)

(mg%)

ng/L

ng/L

 

1        /

0 mg/kg

Mean

139.2

7.2

3.3

93.0

1.1

32.3

39.9

110.4

7.0

12.9

485

9.156

±S.D.

9.3

0.6

0.3

14.5

0.2

15.3

21.7

14.2

0.5

4.3

30.60

1.47

±S.E.M.

3.5

0.2

0.1

5.5

0.1

5.8

8.2

5.4

0.2

1.6

11.56

0.558

 

 

2     /

125 mg/kg

Mean

136.6

7.8

3.4

98.0

1.1

28.8

39.6

100.0

7.3

12.5

475.1

9.869

±S.D.

4.3

0.9

0.1

17.1

0.4

7.4

9.0

8.9

0.5

2.1

30.59

1.27

±S.E.M.

1.6

0.3

0.1

6.5

0.1

2.8

3.4

3.4

0.2

0.8

11.56

0.482

 

 

 

 

3     /

250 mg/kg

Mean

141.6

8.1

3.8

119.9

1.1

27.9

40.0

100.2

7.0

12.5

425.5*

10.478*

±S.D.

7.3

0.9

0.4

24.7

0.2

6.0

8.1

9.0

0.6

2.8

52.38

1.33

±S.E.M.

2.8

0.4

0.2

9.3

0.1

2.3

3.0

3.4

0.2

1.1

19.79

0.505

 

 

4     /

500 mg/kg

Mean

137.0

8.1

3.7

113.4

1.1

32.3

43.5

100.1

7.0

14.1

448.5

9.842

±S.D.

5.9

0.8

0.2

23.6

0.2

8.4

9.8

5.8

0.6

2.3

31.16

1.93

 

±S.E.M.

 

2.2

 

0.3

 

0.1

 

8.9

 

0.1

 

3.2

 

3.7

 

2.2

 

0.2

 

0.9

 

13.93

 

0.73

 

SUMMARY OF ABSOLUTE ORGAN WEIGHTS(g) - Male Rats

Group/ Dose

 

 

Brain

 

Thymus

 

Liver

 

Kidneys

 

Adrenals

 

Ovaries

 

Spleen

 

Heart

 

Epididymides

 

 

1

Vehicle

Mean

1.79

0.30

10.72

1.98

0.03

2.79

1.04

0.55

0.88

±S.D.

0.28

0.06

1.48

0.30

0.01

0.27

0.15

0.11

0.09

±S.E.M.

0.11

0.02

0.56

0.11

0.01

0.10

0.05

0.04

0.03

 

 

2

(125) mg/kg

 

Mean

1.89

0.46

11.54

2.09

0.07

2.75

1.05

0.54

0.89

 

±S.D.

0.12

0.16

0.70

0.11

0.08

0.22

0.06

0.13

0.08

±S.E.M.

0.05

0.06

0.26

0.04

0.03

0.08

0.02

0.05

0.03

 

3

(250)mg/kg

 

Mean

1.87

0.32

11.25

2.04

0.04

2.81

1.10

0.63

0.91

 

±S.D.

0.07

0.06

1.96

0.23

0.01

0.14

0.12

0.19

0.10

±S.E.M.

0.02

0.02

0.74

0.09

0.00

0.05

0.05

0.07

0.04

 

 

4

 (500)

mg/kg

Mean

1.79

0.28

10.61

1.93

0.04

2.87

1.03

0.54

0.88

±S.D.

0.07

0.05

0.51

0.17

0.01

0.13

0.06

0.11

0.05

±S.E.M.

0.03

0.02

0.19

0.06

0.00

0.05

0.02

0.04

0.02

 SUMMARY OF ABSOLUTE ORGAN WEIGHTS(g)

 Femalerats

Group/ Dose

 

 

Brain

 

Thymus

 

Liver

 

Kidneys

 

Adrenals

 

Ovaries

 

Spleen

Heart

 

Uterus

1/0

   (mg/kg)

Mean

1.79639

0.69686

8.13916

1.66367

0.0451

0.14289

0.63041

0.79684

0.46711

±S.D.

0.13458

0.547

0.47047

0.18627

0.01172

0.02302

0.11848

0.11455

0.07792

±S.E.M.

0.05087

0.20675

0.17782

0.0704

0.00443

0.0087

0.04478

0.04329

0.02945

 

2/(120)

mg/kg

Mean

1.78147

0.28884

8.15586

1.55246

0.04734

0.08319*

0.63869

0.77013

0.39303

±S.D.

0.11628

0.1304

1.06437

0.16655

0.00937

0.01815

0.24329

0.06459

0.08246

±S.E.M.

0.04395

0.04929

0.40229

0.06295

0.00354

0.00686

0.09196

0.02441

0.03117

 

2/

 

250 mg/kg

Mean

1.6069

0.3289

8.3645

1.566

0.0395

0.0935*

0.4718

0.7185

0.4667*

 

±S.D.

 

0.27569

 

0.12711

 

0.78925

 

0.15522

 

0.01286

 

0.02423

 

0.11176

 

0.06134

 

0.07708

±S.E.M.

0.1042

0.04804

0.29831

0.05867

0.00486

0.00916

0.04224

0.02319

0.02913

 

4  /

(500) mg/kg

Mean

1.75207

0.42151

7.9445

1.525*

0.04407

0.07481

0.49059*

0.70961

0.34757*

±S.D.

0.12692

0.20921

0.63133

0.13336

0.00665

0.01518

0.09178

0.02657

0.10007

±S.E.M.

0.04797

0.07907

0.23862

0.05041

0.00251

0.00574

0.03469

0.01004

0.03782

SUMMARY OF RELATIVE ORGAN WEIGHTS Male Rats

 

Group/Dose

 

 

Brain

 

Thymus

 

Liver

 

Kidneys

 

Adrenals

 

Testes

 

Splee n

 

Heart

 

epididymide s

 

1/

 

0

Mean

0.6051

0.1009

3.6170

0.6680

0.0110

0.9447

0.1846

0.2970

0.3521

±S.D.

 

0.1132

 

0.0236

 

0.4211

 

0.0880

 

0.0052

 

0.1114

 

0.0407

 

0.0210

 

0.0527

±S.E.M.

 

0.0428

 

0.0089

 

0.1592

 

0.0333

 

0.0020

 

0.0421

 

0.0154

 

0.0079

 

0.0199

 

 

 

 

2/

125 mg/kg

Mean

 

0.6466

 

0.1549

 

3.9569*

 

0.7236

 

0.0243

 

0.9377

 

0.1856

 

0.3060

 

0.3599

±S.D.

 

0.0330

 

0.0515

 

0.2711

 

0.0369

 

0.0274

 

0.0434

 

0.0428

 

0.0262

 

0.0299

±S.E.M.

 

0.0125

 

0.0195

 

0.1025

 

0.0139

 

0.0104

 

0.0164

 

0.0162

 

0.0099

 

0.0113

 

 

 

3  /

250 mg/kg

Mean

 

0.6674

 

0.1140

 

3.9886

 

0.7207

 

0.0143

 

1.0007

 

0.2209

 

0.3220

 

0.3883

±S.D.

 

0.0627

 

0.0252

 

0.6023

 

0.0431

 

0.0021

 

0.0816

 

0.0608

 

0.0367

 

0.0346

±S.E.M.

 

0.0237

 

0.0095

 

0.2277

 

0.0163

 

0.0008

 

0.0309

 

0.0230

 

0.0139

 

0.0131

 

 

 

4   /

500 mg/kg

Mean

 

0.5317

 

0.0868

 

3.1877

 

0.5684

 

0.0110

 

0.8401

 

0.1709

 

0.2618*

 

0.3071*

±S.D.

 

0.3033

 

0.0461

 

1.7161

 

0.3359

 

0.0062

 

0.4873

 

0.0898

 

0.1468

 

0.1775

±S.E.M.

 

0.1357

 

0.0206

 

0.7675

 

0.1502

 

0.0028

 

0.2179

 

0.0402

 

0.0656

 

0.0794

SUMMARY OF RELATIVE ORGAN WEIGHTS- Females

Group/Dose

 

 

Brain

Thymu s

 

Liver

 

Kidneys

Adrenal s

 

Ovaries

 

uterus

 

Spleen

 

Heart

 

 

1/0

Mean

 

0.8181

 

0.3176

 

3.7199

 

0.7573

 

0.0200

 

0.0654

 

0.2107

 

0.2849

 

0.3614

±S.D.

0.0380

0.2566

0.3170

0.0667

0.0045

0.0141

0.0202

0.0354

0.0334

±S.E.M.

0.0144

0.0970

0.1198

0.0252

0.0017

0.0053

0.0076

0.0134

0.0126

 

 

2 / 125 mg/kg

Mean

 

0.8227

 

0.1324

 

3.7541

 

113.7454

 

0.0214

 

*0.0380

 

0.1824

 

0.2926

 

0.3557

±S.D.

0.0439

0.0598

0.3280

299.0822

0.0053

0.0090

0.0439

0.1052

0.0323

±S.E.M.

0.0166

0.0226

0.1240

113.0424

0.0020

0.0034

0.0166

0.0398

0.0122

 

3  /          250 mg/kg

Mean

 

0.7137

 

0.1443

 

3.6900

 

0.6917

 

0.0171

 

0.0406*

 

0.2054

 

0.2077*

 

0.3169*

±S.D.

0.1425

0.0553

0.2348

0.0629

0.0053

0.0089

0.0298

0.0487

0.0212

±S.E.M.

0.0539

0.0209

0.0887

0.0238

0.0020

0.0034

0.0113

0.0184

0.0080

 

4    /  500mg/kg

Mean

0.8253

0.1970

3.7417

0.7173

0.0203

0.0349*

0.1636

0.2304*

0.3341

±S.D.

0.0546

0.0920

0.2484

0.0423

0.0028

0.0080

0.0463

0.0405

0.0130

±S.E.M.

0.0206

0.0348

0.0939

0.0160

0.0011

0.0030

0.0175

0.0153

0.0049

Conclusions:
NOAEL was considered to be 125 mg/kg bw and LOAEL was considered to be 250 mg/kg body weight/day when Sprague Dawley male and female rats were exposed daily to methyl 2-naphthyl ether by oral route for 28 days.
Executive summary:

In a 28 days repeated dose toxicity study, the effect of methyl 2-naphthyl ether was evaluated in male and female Sprague Dawley rats. The test chemical was administered by oral gavage in the concentration of 0, 125, 250 or 500 mg/kg body weight/day. 2 animals died at 500 mg/kg bw as compared to control.Abnormalities such as nasal discharge, red crust around the nostrils was observed in all treatment groups. In addition, a few cases of snuffling, eye lid swelling and hunched back posture were also observed in the treated groups. A significant decrease in body weights in male rats on the first, second and fourth week of treatment and in female rats, a significant decrease in body weight was observed on the first, second and third week of treatment were observed as compared to control. No effects on food and water consumption were observed as compared to control. In addition,In hematology, methyl 2-naphthyl ether resulted in significantly increased platelet count along with decreased MCV, MCH and MCHC levels. Treatment with 250 mg/kg body weight/day resulted in significantly decreased levels of hemoglobin and MCHC in male rats. At 250 mg/kg body weight/day, a significant decrease was noticed in neutrophils along with altered values of MCV in female rats. In clinical chemistry, significantly increased the level of testosterone in the 500 mg/kg body weight/day group in males, as well as it significantly increased the level of estrogen in the 250 mg/kg body weight/day group in females. Significant increased levels in cholesterol, potassium and albumin was also observed. Similarly, Changes in relative and absolute organ weight of spleen, kidney, liver, brain, heart, ovaries and uterus were observed when treated with 125, 250 or 500 mg/kg body weight/day. Nasal discharge, red crust around nostril, perineum wet, bloated stomach, viscous oily secretion was found inside the thoracic cavity. Small white solid mass slightly firm in consistency floating inside the urinary bladder was observed. However, due to adaptive metabolic and physiological changes, anoxic/ hypoxic conditions during anesthesia and terminal sacrifice, these findings were considered to be of no toxicological significance. No remarkable changes observed in control and the 500 mg/kg body weight/day-treated animals. A few microscopic findings observed in 500 mg/kg body weight/day-treated animals included collapsed lung with focal inflammation, focal fatty change and excess of lymphocytes in liver, inflammation in small intestine, reactive spleen and excess of mucous in colon. Since the histopathological changes were not observed in 125 mg/kg bw and these findings were also observed in the control animals. Hence, NOAEL was considered to be 125 mg/kg bw and LOAEL was considered to be 250 mg/kg body weight/day when Sprague Dawley male and female rats were exposed daily to methyl 2-naphthyl ether by oral route for 28 days.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
125 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
Data is from a Klimisch 1 datasource and provides a robust study summary.

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
repeated dose toxicity: inhalation
Data waiving:
exposure considerations
Justification for data waiving:
a short-term toxicity study does not need to be conducted because exposure of humans via inhalation in production and/or use is not likely as based on the provided thorough and rigorous exposure assessment
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Link to relevant study records
Reference
Endpoint:
repeated dose toxicity: inhalation
Data waiving:
exposure considerations
Justification for data waiving:
a short-term toxicity study does not need to be conducted because exposure of humans via inhalation in production and/or use is not likely as based on the provided thorough and rigorous exposure assessment
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: dermal
Data waiving:
exposure considerations
Justification for data waiving:
a short-term toxicity study does not need to be conducted because exposure of humans via the dermal route in production and/or use is not likely as based on the provided thorough and rigorous exposure assessment
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: dermal
Data waiving:
exposure considerations
Justification for data waiving:
a short-term toxicity study does not need to be conducted because exposure of humans via the dermal route in production and/or use is not likely as based on the provided thorough and rigorous exposure assessment
Critical effects observed:
not specified
Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Repeated Dose (Oral) Toxicity:

The available data of the test chemical for the repeated dose toxicity studies via oral route as follows:

Study 1:

An Extended One-Generation Reproductive Toxicity Study was performed according to OECD TG 443 to evaluate the general, reproductive and developmental toxicological endpoints associated with the repeated oral administration of graduated doses of the test chemical in Wistar rats. A total of 240 wistar rats (120 males and 120 females) were randomized into 4 experimental groups each containing 30 animals per sex per group. The groups, viz., G1, G2, G3 and G4 received 0, 62.5, 125 and 250 mg/kg body weight of test chemical respectively. Dose administration was through an oral gavage and the vehicle used in this study was corn oil. All animals of both sexes were dosed 2 weeks prior to mating. Dosing was continued in both sexes during the mating period. Females were treated throughout gestation and lactation up to the day of termination after weaning. Males were treated in the same manner until termination. During observations, it was seen that no treatment-related clinical signs or symptoms were observed in any of the parent animals up to 125 mg/kg body weight of the test chemical. However, mild salivation was observed in total 13 of 30 males of G4 group (250 mg/kg), starting from the 27th day of treatment and continuing till termination. All adult animals were found normal with respect to all the parameters examined during detailed clinical examinations through the course of the study. No treatment-related symptoms were found up to the dose level of 250 mg/kg body weight in either sex. No morbidities or mortality were recorded in adult animals of parent generation in any of the experimental groups throughout the duration of the experiment. Mean body weights remained comparable among all the groups for every time point (days 8, 15, 22, 29, 36, 43, 50, 57, 64 and 71) recorded during the 70 days exposure period for males. No significant difference was found for the mean body weights of females during the two weeks pre-mating period, during mating, during gestation or during lactation period. However, the mean percent change in body weight on all the instances (days 8, 15, 22, 29, 36, 43, 50, 57, 64 and 71) with respect to the first day of exposure was significantly reduced for parental males of G4 group (250 mg/kg) (P≤0.05 to P≤0.001) as compared to control animals, whereas mean body weight change for G2 (62.5 mg/kg) and G3 (125 mg/kg) males remained comparable to control animals. Among the females, mean percent change in body weights remained statistically comparable among all groups during pre-mating, mating, gestation and lactation periods. The mean feed consumed per animal per day significantly increased on day 8 of treatment for G2 (62.5 mg/kg) (P≤0.01) and G3 (125 mg/kg) (P≤0.05) males as compared to G1 (control) animals. However, it was found to be similar for all other time points across the groups. The mean feed consumption of treatment group (G2, G3 & G4) females remained statistically comparable to control group during pre-mating, mating, gestation and lactation period except for one instance, where mean feed consumption on day 8 of treatment was significantly reduced for G2 (62.5 mg/kg) (P≤0.01) as compared to control females. The differences found in mean feed consumption of male and female animals were neither dose-dependent nor consistent, hence it was deemed to be independent of test-item treatment. Feed efficiency for the G4 males had statistically reduced in the first two weeks of treatment, i.e. on days 8 and 15. It was also found to have increased significantly during the penultimate week of treatment, i.e. on day 63. For all other groups among the males, the feed efficiency was statistically comparable across the groups. Among the parental females, no statistical differences were observed for feed efficiency in any of the groups. No significant difference was observed in any of the measured hematology parameters in all the groups of both the sexes, except in the WBC count, which was found to be significantly increased in males of G4 group (P≤0.01) as compared to control males. Significant increase was observed in Phosphorus, Sodium and Chloride levels in G4 males (P≤0.05) as compared to control males. All the other clinical chemistry parameters in males and females were found comparable in control and treatment groups. The urine parameters tested did not show up any significant changes among the experimental groups in the parental generation of either sex. No significant difference was observed in absolute organ weight or organ weight relative to body weight for the entire set of measured organs in parent male and female animals. However, significant reduction was observed in absolute and relative liver weight in G4 females (P<0.05) as compared to control animals. In the current study, however, the decreased liver weights with respect to body weights in G4 females does not correlate with any histological observations made, making it inconsequential from the point of view of toxic effects of test-chemical. External examination of all male and female rats of control and all treated groups did not reveal any lesion of pathological significance. Internal examination of the rats revealed reduced testicular size in two animals (Male: G2: 1/30, G4:1/30), increased prostate size in one animal (Male: G2: 1/30) and reduced prostate size in one animal (Male: G3: 1/30). Remaining animals of parent generation did not reveal any abnormality of pathological significance. No treatment-related histopathological findings are reported in this study that could arise out of test-item administration. All observed tissues were normal at the level of histology. Minimal to moderate and focal lymphocytic infiltrates were observed in some slides of liver, kidney, lung, heart, colon, thymus and adrenals with mild to moderate cyst in thyroid gland, which were distributed randomly across the experimental groups and did not show any pattern of either dose-dependency or sex-based selectivity. While TSH, Estradiol, Progesterone and Testosterone remained statistically comparable among the groups of parental males, the mean level of T4 was found reduced in G3 males (P<0.01) as compared to control. This is likely an incidental result, given that G4 was statistically comparable to control. The mean T4, TSH, Estradiol and testosterone levels remained comparable among the groups of parental females; whereas, Progesterone was found to be significantly reduced in G4 females (P<0.001). This could be a test-chemical related effect. Based on all the available data, it was observed that there were no effects observed on male and female animals up to 125 mg/kg bw (Mid Dose-G3) of the test chemical administration. However, at 250 mg/kg bw (High Dose-G4) dose group, clinical signs, effects on weights of liver and effects on hormonal levels was observed, due to test chemical administration. Therefore, it was concluded that the NOAEL and LOAEL of the test chemical was 125 mg/kg bw and 250 mg/kg bw, respectively.

Study 2:

In a 28 days repeated dose toxicity study, the effect of methyl 2-naphthyl ether was evaluated in male and female Sprague Dawley rats. The test chemical was administered by oral gavage in the concentration of 0, 125, 250 or 500 mg/kg body weight/day. 2 animals died at 500 mg/kg bw as compared to control.Abnormalities such as nasal discharge, red crust around the nostrils was observed in all treatment groups. In addition, a few cases of snuffling, eye lid swelling and hunched back posture were also observed in the treated groups. A significant decrease in body weights in male rats on the first, second and fourth week of treatment and in female rats, a significant decrease in body weight was observed on the first, second and third week of treatment were observed as compared to control. No effects on food and water consumption were observed as compared to control. In addition,In hematology, methyl 2-naphthyl ether resulted in significantly increased platelet count along with decreased MCV, MCH and MCHC levels. Treatment with 250 mg/kg body weight/day resulted in significantly decreased levels of hemoglobin and MCHC in male rats. At 250 mg/kg body weight/day, a significant decrease was noticed in neutrophils along with altered values of MCV in female rats. In clinical chemistry, significantly increased the level of testosterone in the 500 mg/kg body weight/day group in males, as well as it significantly increased the level of estrogen in the 250 mg/kg body weight/day group in females. Significant increased levels in cholesterol, potassium and albumin was also observed. Similarly, Changes in relative and absolute organ weight of spleen, kidney, liver, brain, heart, ovaries and uterus were observed when treated with 125, 250 or 500 mg/kg body weight/day. Nasal discharge, red crust around nostril, perineum wet, bloated stomach, viscous oily secretion was found inside the thoracic cavity. Small white solid mass slightly firm in consistency floating inside the urinary bladder was observed. However, due to adaptive metabolic and physiological changes, anoxic/ hypoxic conditions during anesthesia and terminal sacrifice, these findings were considered to be of no toxicological significance. No remarkable changes observed in control and the 500 mg/kg body weight/day-treated animals. A few microscopic findings observed in 500 mg/kg body weight/day-treated animals included collapsed lung with focal inflammation, focal fatty change and excess of lymphocytes in liver, inflammation in small intestine, reactive spleen and excess of mucous in colon. Since the histopathological changes were not observed in 125 mg/kg bw and these findings were also observed in the control animals. Hence, NOAEL was considered to be 125 mg/kg bw and LOAEL was considered to be 250 mg/kg body weight/day when Sprague Dawley male and female rats were exposed daily to methyl 2-naphthyl ether by oral route for 28 days.

Repeated Dose (Inhalation) Toxicity:

The particle size distribution of the substance methyl 2-naphthyl ether was found to vary in the size of 150 µm to 2000 µm,so the potential for the generation of inhalable vapours of methyl 2-naphthyl ether is low. Moreover the normal conditions of use of this substance will not result in aerosols, particles or droplets of an inhalable size, so exposure to humans via the inhalatory route will be unlikely to occur and therefore repeated dose toxicity via the inhalation route was considered for waiver.

Repeated Dose (Dermal) Toxicity:

The acute toxicity value for test chemical (as provided in section 7.2.3) is >2000 mg/kg body weight. Also, given the use of the chemical; repeated exposure by the dermal route is unlikely since the use of gloves is common practice in industries. Thus, it is expected that the test chemical shall not exhibit 28 day repeated dose toxicity by the dermal route. In addition, there is no data available that suggests that the test chemical shall exhibit repeated dose toxicity by the dermal route. Hence this end point was considered for waiver.

Justification for classification or non-classification

Based on all the available data, it was concluded that the test chemical did not cause any toxicity to any organs when exposed to the test animals in sub-acute and sub-chronic durations via oral route. The exposure of the test chemical via dermal and inhalation route is not likely for the test chemical. Therefore, it was concluded that the test chemical was not likely to be classified under category STOT-RE 1 or 2 as per the CLP criteria of classification and labeling.