Registration Dossier

Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Data is from study report

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Principles of method if other than guideline:
Evaluations of general and reproduction/ developmental toxicity endpoints associated with administration of repeated doses of 2-[7-(diethylamino)-2-oxo-2H-1-benzopyran-3yl]benzoxazole-5-sulphonamide (CAS No.: 68427-35-0) in Wistar rats.
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Details on test material:
- Name of test material (as cited in study report): 2-[7-(diethylamino)-2-oxo-2H-1-benzopyran-3-yl]benzoxazole-5-sulphonamide
- Molecular formula : C20H19N3O5S
- Molecular weight : 413.452 g/mol.
- Smiles :n1c2cc(S(N)(=O)=O)ccc2oc1c1cc2ccc(N(CC)CC)cc2oc1=O
- InChI : 1S/C20H19N3O5S/c1-3-23(4-2)13-6-5-12-9-15(20(24)28-18(12)10-13)19-22-16-11-14(29(21,25)26)7-8-17(16)27-19/h5-11H,3-4H2,1-2H3,(H2,21,25,26)
- Substance type: Organic
- Physical state: Solid
- Analytical purity: 100%
Specific details on test material used for the study:
- Name of test material (as cited in study report): 2-[7-(diethylamino)-2-oxo-2H-1-benzopyran-3-yl]benzoxazole-5-sulphonamide
- Molecular formula : C20H19N3O5S
- Molecular weight : 413.452 g/mol.
- Substance type: Organic
- Physical state: Solid

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Animals were procured from a CPCSEA approved Vendor [Gentox Bio Services Pvt. Ltd. (CPCSEA Registration No.: 1242/RCBT/S/08/CPCSEA)]
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: (P) x wks; (F1) x wks: 12 - 13 weeks
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x g
- Fasting period before study:
- Housing: Before the animals are brought in, the study room and cages were cleaned and disinfected. During the study, the floor of the experimental room and work tops were swept and mopped with disinfectant solution every day or as on requirement. Cages were cleaned at regular intervals.
A total 2-3 rats/sex were housed in Polycarbonate cages (size 37 [cm] x 21 [cm], height 20 [cm]). Cage rotation was carried out weekly during study period except during mating and during gestation and lactation only for females.
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.
Bedding material of batch No. SPAR-33/2016 and SPAR-34/2016 (Sparconn Life Sciences Bangalore) was used in this study.
All the animals were identified by temporary tail marking with indelible ink and cage cards. Following allocation to the study, each animal was uniquely identified by micro toe pad tattooing and colour coded cage cards labelled with study no., study type, test system, sex, dose, group, animal number, dosing start date, experimental start and completion date. Pups were identified with body marking with permanent non-toxic marker pen
- Use of restrainers for preventing ingestion (if dermal): no
- Diet (e.g. ad libitum): conventional laboratory pelleted diet, ad libitum
- Water (e.g. ad libitum): Aqua guard filtered drinking water in bottles was offered ad libitum.
- Acclimation period:7 days and 8 days prior to test item administration for Dose Range Finding Study (DRF) and Main Study respectively.


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.00 to 24.90 °C
- Humidity (%): 39.90 to 65.50%.
- Air changes (per hr):12 hours light and 12 hours dark
- Photoperiod (hrs dark / hrs light):Air changes were about minimum 12 times per hour and filtered adequately.

IN-LIFE DATES: From: To:

Administration / exposure

Route of administration:
oral: gavage
Type of inhalation exposure (if applicable):
not specified
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test item was weighed and dissolved in a vehicle (corn oil) to achieve desired concentration of test item. Dose formulation was freshly prepared daily. At the time of dosing, dose formulation was kept on the magnetic stirrer to maintain the homogeneity of test item.

DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food):
- Storage temperature of food:

VEHICLE
- Justification for use and choice of vehicle (if other than water): corn oil
- Concentration in vehicle: 0, 250, 500 and 1000 mg/kg body weight
- Amount of vehicle (if gavage): 1.0 ml/100g body weight
- Lot/batch no. (if required): Lot nos.: MR020816, A611001 and A1701001
- Purity:
Details on mating procedure:
- M/F ratio per cage:1:1
- Length of cohabitation: until pregnancy occurs or two weeks elapsed.
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy: Sperm positive vaginal smear was considered as day "0" of gestation.
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility.; yes
- Further matings after two unsuccessful attempts: no
- After successful mating each pregnant female was caged (how): : housed individually
- Any other deviations from standard protocol:
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Specificity, Precision (%RSD), Accuracy (% Recovery) and Homogeneity by HPLC-UV
Duration of treatment / exposure:
Approx 63 days
Frequency of treatment:
Daily
Details on study schedule:
not specified
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day
Dose / conc.:
250 mg/kg bw/day
Dose / conc.:
500 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day
No. of animals per sex per dose:
Total:124
0 mg/kg bw: 13 male, 13 female
250 mg/kg bw: 13 male, 13 female
500 mg/kg bw: 13 male, 13 female
1000 mg/kg bw: 13 male, 13 female
Recovery Group
0 mg/kg bw: 5 male, 5 female
1000 mg/kg bw: 5 male, 5 female
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels 250, 500 and 1000 mg/kg body weight were selected for the Main Study based on the results of Dose Range Finding (DRF).
- Rationale for animal assignment (if not random): Randomization was done based on recent body weight, before first dosing.
- Other:
Positive control:
not specified

Examinations

Parental animals: Observations and examinations:
Mortality and Morbidity, General clinical sign, motor Activity and Behavioural, Body Weight and Feed Consumption were examined.
Oestrous cyclicity (parental animals):
Estrous cycle were monitored daily
Sperm parameters (parental animals):
not specified
Litter observations:
Live pups, Number and sex of pups, stillbirths, live births, runts (pups that are significantly smaller than corresponding control pups) and body weight were examined.
Postmortem examinations (parental animals):
Hematology, clinical biochemistry, Organ Weight, gross Pathology and Histopathology were examined.
Postmortem examinations (offspring):
Gross Pathology were examined.
Statistics:
Raw data was analysed using statistical software “Sigma Plot 11.0” (Supplied by Cranes Software International Ltd. Bangalore). The mean and standard deviation was calculated using the software and all data was summarized in tabular form. All continuous data (body weight, feed consumption, Functional Observational Battery parameters, hematology, clinical chemistry, absolute and relative organ weights, maternal and pup parameters etc.) were checked for normality using Shapiro Wilk test. All homogenous data was analysed using ANOVA and data showing significance in their variances was subjected to Dunnett’s t-test. All heterogeneous data was analysed using F test and Student’s t-test, Dunn’s Test, Kruskal-Wallis, ANOVA on ranks.
Reproductive indices:
Gestational length, Litter size, No. of live births, Post-implantation loss and Pregnancy Index (%) were examined.
Offspring viability indices:
Fetal Survival Index at Post-natal Day 1 and 4 were examined.

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
no effects observed
Description (incidence and severity):
Statistically significant decrease were observed in number of rears (250, 500 and 1000 mg/kg bw at pre-treatment and 1000 mg/kg bw at Week 1), number of urine pools (250 mg/kg bw at Week 2) in male as compared to control. In recovery male, statistically significant increase in number of urine pools at week 4 in recovery at 1000 mg/kg bw as compared to recovery control. In female, statistically significant decrease was observed in number of fecal bolus (1000 mg/kg bw recovery at Week 1) as compared to control recovery group.
The above changes observed were inconsistent/ biologically insignificant and not dose dependant hence considered as incidental and not attributed to the effect of test item administration.
Dermal irritation (if dermal study):
not specified
Mortality:
no mortality observed
Description (incidence):
One female was found dead on day 41 at 1000 mg/kg bw due to gavaging error
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Statistically significant decrease was observed in percent body weight change of 250, 500 and 1000 mg/kg bw on day 1-20 as compared to control group.
Body weight and Percent body weight changes in animals of the all other test groups of both the sexes was comparable and did not show any significant difference as compared to the respective control group. These changes observed were inconsistent, hence not considered as effect of the test item administration
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
Statistically significant decrease in feed consumption was observed in 250, 500 and 1000 mg/kg bw female on gestation day 14-20 as compared to the control. Feed consumption in animals of the all other test groups of both the sexes was comparable and did not show any significant difference as compared to the respective control group.
Changes observed in feed consumption were inconsistent, hence not considered as effect of the test item administration.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
no effects observed
Description (incidence and severity):
No effect on Eye were observed in treated rats as compared to control.
Haematological findings:
no effects observed
Description (incidence and severity):
At the end of treatment period revealed statistically significant decreased of aPTT in male rats at 1000 mg/kg, statistically significant increased of aPTT in female rats at 250 mg/kg while statistically significant decreased in Lymphocyte Count in female rats at 500 mg/kg.
At the end of recovery period, all of the above altered parameters had returned to normal level similar to control group rats in both sexes.
During treatment free recovery period, statistically significant increase was observed in PT of recovery male rats treated at 1000 mg/kg and statistically significant decreased was observed Hematocrit and Hemoglobin in recovery male rats treated at 1000 mg/kg, while statistically significant decreased of Monocyte in female recovery rats at 1000 mg/kg when compared to respective control group. The observed variations in PT, Hematocrit, Hemoglobin and monocyte were considered to be of no toxicological significance, minimal in nature and occurred in the absence of clear dose related response
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
At the end of treatment period revealed, statistically significant increase in Chloride of male rats treated at 500 mg/kg, Sodium of male rats treated at 1000 mg/kg. Statistically significant decrease in bile acid of male rats treated at 250 mg/kg and 1000 mg/kg respectively while statistically significant decrease in Triglyceride of female rats treated at 500 mg/kg.
During treatment-free recovery period revealed statistically significant increase in Potassium in female rats treated at 1000 mg/kg while statistically decrease in Bile acid in female rats treated at 1000 mg/kg was observed when compared to respective control group.
The observed variations in Potassium and Bile acid was considered incidental and not test item related as it was observed only in single sex, inconsistent, not dose dependent and further is not evidenced by histopathological observations.
Urinalysis findings:
not specified
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
The sensory reactivity measurements were comparable and no changes were revealed in any of the animals of all treated groups in both the sexes.
Foot splay and fore limb and hind limb grip strength parameters were comparable and no treatment related changes were revealed in any of the animals of all treated groups.
Motor activity measurements were comparable and no changes were revealed in any of the animals from all treated groups of both the sexes as compare to control group.
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
At the end of treatment and recovery period, absolute and relative weight of organs of treated group rats of either sex did not differ significantly when compared to respective control group except, statistical significant increase absolute and relative adrenal weight in male rat at 250 mg/kg bw compared to main control group. Statistical significant decrease relative testes and Epididymis weight in male rat at 1000 mg/kg bw was observed when compared to control rats. Statistical significant increased relative heart weight in male recovery rats was observed when compared to control rats.
Gross pathological findings:
no effects observed
Description (incidence and severity):
External examination of all male and female rats of control and all treated groups including recovery groups did not reveal any abnormality of pathological significance.
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
Visceral examination of the rats of control and other treated groups did not reveal any pathological abnormality.
focal mild to multifocal mild mononuclear cell infiltration in male and focal minimal to multifocal minimal mononuclear cells infiltration and minimal chronic changes and hydropic degeneration in female liver, Unilateral: focal minimal to multifocal mild mononuclear cells infiltration in male and focal minimal to multifocal minimal mononuclear cells infiltration in female Kidneys, focal mild to multifocal mild mononuclear cells infiltration in male and focal minimal to multifocal minimal mononuclear cells infiltration in female Lungs and focal minimal to multifocal mild GALT hyperplasia in male and focal minimal to multifocal minimal GALT hyperplasia in female intestin at 250 and 1000 mg/kg bw, Vacuolar, degeneration multifocal, mild in male and diffuse vacuolations in female Adrenals at 250 mg/kg bw, minimal cyst in male and mild serosanginous fluid cavity in female Pituitary Gland, focal minimal to multifocal moderate Retention of sperm, focal minimal to focal mild Exfoliation of round spermatid, focal minimal to multifocal mild Seminiferous tubules Vacuolar degeneration, marked diffuse Necrosis of Seminiferous tubules, minimal diffuse Atrophy Seminiferous tubules, minimal to mild Vacuolar degeneration of Leydig cell, marked diffuse Necrosis of Epididymis, marked diffuse decrease in Epididymis sperm and Epididymis of Epididymis in male rat and segmental minimal Vacuolar degeneration of Ovary and minimal Perimetrium and myometrium vacuolations of Uterus in female rats were observed at different doses.
Lesions observed in liver, kidneys, lungs, adrenals, intestine, pituitary gland and reproductive organs of high dose treated group rats are well comparable with respective control group rats and exhibited no dose relationship. Further these observed lesions are common in occurrence in rodents during toxicological studies (Boorman et al., 1990, Greaves, 2007, Greaves and Faccini, 1992; Haschek et al., 2002). Hence, occurrence of these lesions could be considered as spontaneous or incidental in nature and not to be attributed because of administration of the Test Item
Histopathological findings: neoplastic:
not specified
Other effects:
not specified

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
One females showed prolonged diestrous i.e more than 3 days with total estrous cycle period of 6 days or more before mating period at 250 and 1000 mg/kg bw.
Reproductive function: sperm measures:
not specified
Reproductive performance:
no effects observed
Description (incidence and severity):
No statistically significant effect on Gestational length, Litter size, No. of live births, Post-implantation loss, Post-natal loss, Pups sex ratio and Pregnancy index of treated rats were observed as compared to control.

Effect levels (P0)

Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
mortality
body weight and weight gain
food consumption and compound intake
haematology
clinical biochemistry
organ weights and organ / body weight ratios
gross pathology
histopathology: non-neoplastic
reproductive function (oestrous cycle)
reproductive performance
Remarks on result:
other: No effect observed

Target system / organ toxicity (P0)

Critical effects observed:
not specified
System:
other: not specified
Organ:
not specified
Treatment related:
not specified
Dose response relationship:
not specified
Relevant for humans:
not specified

Results: F1 generation

General toxicity (F1)

Clinical signs:
not specified
Dermal irritation (if dermal study):
not specified
Mortality / viability:
no mortality observed
Description (incidence and severity):
No effect on Survival of pups were observed as compared to control.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No effect on weight gain for pups at PND4 were observed as compared to control.
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
not specified
Clinical biochemistry findings:
not specified
Urinalysis findings:
not specified
Sexual maturation:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
no effects observed
Description (incidence and severity):
Pups died during course of study revealed various lesions among the control and treated groups.
Cannibalism in male pups at 250 and 500 mg/kg bw and in female at 250 mg/kg bw, absence of milk in stomach in male at 250 and 500 mg/kg bw and in female at 250 mg/kg bw, reddish discoloration of lungs in female at 250 mg/kg bw and brain haemorrhage in male rat at 250 mg/kg bw.
No gross patholoical changes were observed at 1000 mg/kg bw. Hence, occurrence of these lesions could be considered as spontaneous or incidental in nature and not to be attributed because of administration of the Test Item
Histopathological findings:
not specified
Other effects:
not specified

Developmental neurotoxicity (F1)

Behaviour (functional findings):
not specified

Developmental immunotoxicity (F1)

Developmental immunotoxicity:
not specified

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
viability
mortality
body weight and weight gain
gross pathology
Remarks on result:
other: No effect observed

Target system / organ toxicity (F1)

Critical effects observed:
not specified
System:
other: not specified
Organ:
not specified
Treatment related:
not specified
Dose response relationship:
not specified
Relevant for humans:
not specified

Overall reproductive toxicity

Reproductive effects observed:
not specified
Treatment related:
not specified
Relation to other toxic effects:
not specified
Dose response relationship:
not specified
Relevant for humans:
not specified

Any other information on results incl. tables

Mortality and Morbidity

Sex: Male

Group

Treatment

Dose (mg/kg b.wt.)

No. of Animals/

Group

Observation During Study Period

G1

Control

0

13

NMM

G2

Low

250

13

NMM

G3

Mid

500

13

NMM

G4

High

1000

13

NMM

G1-R

Control- Recovery

0

5

NMM

G4-R

High- Recovery

1000

5

NMM

 

Sex: Female

Group

Treatment

Dose (mg/kg b.wt.)

No. of Animals/

Group

Observation During Study Period

G1

Control

0

13

NMM

G2

Low

250

13

NMM

G3

Mid

500

13

NMM

G4

High

1000

13

01/13*

G1-R

Control -Recovery

0

5

NMM

G4-R

High- Recovery

1000

5

NMM

Keys:NMM = No mortality and morbidity observed, No.= Number, *= One female found dead during clinical sign observation on Gestation Day 24.

Summary of Days of Conception and Pregnancy Index (%)

Group(N)

G1(13)

G2(13)

G3(13)

G4(13)

Dose (mg/kg b.wt.)

0

250

500

1000

No. of females showed evidence of copulation

13

12

13

12

No. of females concieved between Days 1-5 of cohabitation

10

7

10

8

No. of females concieved after Day 5 of cohabitation

3

5

3

4

Females achieved pregnancy

13

12

13

12

Pregnancy Index (%)

100.00

92.31

100.00

92.31

Key:N= number of dams in a group, No. = Number

Post-natal Loss (%) and Pups Survival Index (%)

Group(N)

G1(13)

G2(12)

G3(11)

G4(10)

Dose(mg/kg bwt)

0

250

500

1000

Parameter

Mean

SD

Mean

SD

Mean

SD

Mean

SD

No. of Live Births

10.54

1.13

9.67

1.61

10.00

3.00

9.70

1.95

No. of alive pups at Post-natal Day 4

10.31

0.95

8.42

3.15

9.55

2.81

9.50

2.17

Post-natal Loss (%)

1.99

3.80

11.41

29.11

3.99

6.18

2.67

5.84

Fetal Survival Index at Post-natal Day 4 (%)

98.01

3.80

88.59

29.11

96.01

6.18

97.33

5.84

Keys:SD= Standard Deviation, N= number of dams in a group

Note:Group G2 one female was found non-pregnant. Group G3 two females were showed all the pups still births and in Group G4 one female was non-pregnant, one female was found dead and one females showed all the pups still births. 

Mean Gestational Lengthand Litter size

G1(13)

G2(12)

G3(11)

G4(11) 

0

250

500

1000

Mean

SD

Mean

SD

Mean

SD

Mean

SD

22.23

0.60

19.69

6.10

22.00

1.53

18.54

8.33

10.54

1.13

9.92

1.78

9.38

3.33

9.27

2.23

Keys:SD= Standard Deviation, N= number of dams in a group

Note:Group G2 one female was found non-pregnant. Group G3 two females were showed all the pups still births and in Group G4 one female was non-pregnant and one female was found dead.

Mean Pups Body Weight, Sex Ratio and Gross Observation

Group

G1

G2

G3

G4

Dose(mg/kg bwt)

0

250

500

1000

Mean Pups Weight

Mean

SD

N

Mean

SD

N

Mean

SD

N

Mean

SD

N

Day 0/1

5.88

0.25

13

5.68

0.37

12

5.87

0.49

11

5.75

0.25

10

Day 4

8.71

0.41

13

8.48

0.93

11

8.50

1.02

11

8.83

0.78

10

Group(n)

G1(13)

G2(12)

G3(11)

G4(10)

Pups Body Weight gain

(%)

Mean

SD

N

Mean

SD

N

Mean

SD

N

Mean

SD

N

Day 0/1-Day 4

48.09

5.55

13

49.23

15.97

11

44.01

6.89

11

53.52

10.37

10

Group

(Number of Litter size)

G1(137)

G2(119)

G3(122)

G4(97)

 Sex Ratio at birth

(Male/Female)

81/56

56/60

56/54

58/39

 Sex Ratio at Day 4

(Male/Female)

80/54

47/55

52/53

57/38

Gross Observations

NAD

NAD

NAD

NAD

Keys:SD= Standard Deviation, N= number of dams with live pups on Day 0/1, n= number of dams with alive pups on Day 4, NAD = No Abnormality Detected

Applicant's summary and conclusion

Conclusions:
No Observed Adverse Effect Level (NOAEL) of the test item 2-[7-(diethylamino)-2-oxo-2H-1-benzopyran-3yl]benzoxazole-5-sulphonamide (CAS No.: 68427-35-0) is considered 1000 mg/kg body weight when Wistar male and female rats orally by gavage for Approx 63 days.
Executive summary:

In a Repeated Dose Oral Toxicity Study in combination with Reproduction/ Developmental Toxicity study, Wistar male and female rats were treated with 2-[7-(diethylamino)-2-oxo-2H-1-benzopyran-3yl]benzoxazole-5-sulphonamide in the concentration of 0, 250, 500 and 1000 mg/kg bw orally by gavage for aprrox 63 days. Statistically significant decrease were observed in number of rears (250, 500 and 1000 mg/kg bw at pre-treatment and 1000 mg/kg bw at Week 1), number of urine pools (250 mg/kg bw at Week 2) in male as compared to control. In recovery male, statistically significant increase in number of urine pools at week 4 in recovery at 1000 mg/kg bw as compared to recovery control. In female, statistically significant decrease was observed in number of fecal bolus (1000 mg/kg bw recovery at Week 1) as compared to control recovery group. The above changes observed were inconsistent/ biologically insignificant and not dose dependant hence considered as incidental and not attributed to the effect of test item administration. One female was found dead on day 41 at 1000 mg/kg bw due to gavaging error. Statistically significant decrease was observed in percent body weight change of 250, 500 and 1000 mg/kg bw on day 1-20 as compared to control group. Body weight and Percent body weight changes in animals of the all other test groups of both the sexes was comparable and did not show any significant difference as compared to the respective control group. These changes observed were inconsistent, hence not considered as effect of the test item administration. Statistically significant decrease in feed consumption was observed in 250, 500 and 1000 mg/kg bw female on gestation day 14-20 as compared to the control. Feed consumption in animals of the all other test groups of both the sexes was comparable and did not show any significant difference as compared to the respective control group. Changes observed in feed consumption were inconsistent, hence not considered as effect of the test item administration. No effect on Eye was observed in treated rats as compared to control. Similarly, At the end of treatment period revealed statistically significant decreased of aPTT in male rats at 1000 mg/kg, statistically significant increased of aPTT in female rats at 250 mg/kg while statistically significant decreased in Lymphocyte Count in female rats at 500 mg/kg. At the end of recovery period, all of the above altered parameters had returned to normal level similar to control group rats in both sexes. During treatment free recovery period, statistically significant increase was observed in PT of recovery male rats treated at 1000 mg/kg and statistically significant decreased was observed Hematocrit and Hemoglobin in recovery male rats treated at 1000 mg/kg, while statistically significant decreased of Monocyte in female recovery rats at 1000 mg/kg when compared to respective control group. The observed variations in PT, Hematocrit, Hemoglobin and monocyte were considered to be of no toxicological significance, minimal in nature and occurred in the absence of clear dose related response. At the end of treatment period revealed, statistically significant increase in Chloride of male rats treated at 500 mg/kg, Sodium of male rats treated at 1000 mg/kg. Statistically significant decrease in bile acid of male rats treated at 250 mg/kg and 1000 mg/kg respectively while statistically significant decrease in Triglyceride of female rats treated at 500 mg/kg. During treatment-free recovery period revealed statistically significant increase in Potassium in female rats treated at 1000 mg/kg while statistically decrease in Bile acid in female rats treated at 1000 mg/kg was observed when compared to respective control group. The observed variations in Potassium and Bile acid was considered incidental and not test item related as it was observed only in single sex, inconsistent, not dose dependent and further is not evidenced by histopathological observations. The sensory reactivity measurements were comparable and no changes were revealed in any of the animals of all treated groups in both the sexes. Foot splay and fore limb and hind limb grip strength parameters were comparable and no treatment related changes were revealed in any of the animals of all treated groups. Motor activity measurements were comparable and no changes were revealed in any of the animals from all treated groups of both the sexes as compare to control group. The above changes observed were inconsistent, hence considered as incidental and not attributed to the effect of test item administration. At the end of treatment and recovery period, absolute and relative weight of organs of treated group rats of either sex did not differ significantly when compared to respective control group except, statistical significant increase absolute and relative adrenal weight in male rat at 250 mg/kg bw as compared to main control group. Statistical significant decrease relative testes and Epididymis weight in male rat at 1000 mg/kg bw was observed when compared to control rats. Statistical significant increased relative heart weight in male recovery rats was observed when compared to control rats. External examination of all male and female rats of control and all treated groups including recovery groups did not reveal any abnormality of pathological significance. Visceral examination of the rats of control and other treated groups did not reveal any pathological abnormality. Focal mild to multifocal mild mononuclear cell infiltration in male and focal minimal to multifocal minimal mononuclear cells infiltration and minimal chronic changes and hydropic degeneration in female liver, Unilateral: focal minimal to multifocal mild mononuclear cells infiltration in male and focal minimal to multifocal minimal mononuclear cells infiltration in female Kidneys, focal mild to multifocal mild mononuclear cells infiltration in male and focal minimal to multifocal minimal mononuclear cells infiltration in female Lungs and focal minimal to multifocal mild GALT hyperplasia in male and focal minimal to multifocal minimal GALT hyperplasia in female intestine at 250 and 1000 mg/kg bw, Vacuolar, degeneration multifocal, mild in male and diffuse vacuolations in female Adrenals at 250 mg/kg bw, minimal cyst in male and mild serosanginous fluid cavity in female Pituitary Gland, focal minimal to multifocal moderate Retention of sperm, focal minimal to focal mild Exfoliation of round spermatid, focal minimal to multifocal mild Seminiferous tubules Vacuolar degeneration, marked diffuse Necrosis of Seminiferous tubules, minimal diffuse Atrophy Seminiferous tubules, minimal to mild Vacuolar degeneration of Leydig cell, marked diffuse Necrosis of Epididymis, marked diffuse decrease in Epididymis sperm and Epididymis of Epididymis in male rat and segmental minimal Vacuolar degeneration of Ovary and minimal Perimetrium and myometrium vacuolations of Uterus in female rats were observed at different doses. Lesions observed in liver, kidneys, lungs, adrenals, intestine, pituitary gland and reproductive organs of high dose treated group rats are well comparable with respective control group rats and exhibited no dose relationship. Further these observed lesions are common in occurrence in rodents during toxicological studies (Boorman et al., 1990, Greaves, 2007, Greaves and Faccini, 1992; Haschek et al., 2002). Hence, occurrence of these lesions could be considered as spontaneous or incidental in nature and not to be attributed because of administration of the Test Item. In addition, no reproductive toxicity were observed in treated rats such as Gestational length, Litter size, No. of live births, Post-implantation loss, Post-natal loss, Pregnancy index, Pups sex ratio, survival, body weight on PND 4 and gross pathology of treated rats were observed as compared to control. One females showed prolonged diestrous i.e more than 3 days with total estrous cycle period of 6 days or more before mating period at 250 and 1000 mg/kg bw. Therefore, No Observed Adverse Effect Level (NOAEL) of the test item 2-[7-(diethylamino)-2-oxo-2H-1-benzopyran-3yl] benzoxazole-5-sulphonamide (CAS No.: 68427-35-0) is considered 1000 mg/kg body weight when Wistar male and female rats orally by gavage for Approx 63 days.