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  6. Introduction to the Template for Chronic Toxicity Study with an in utero Phase
  1. Ingredients, Additives, GRAS & Packaging Guidance Documents & Regulatory Information

Introduction to the Template for Chronic Toxicity Study with an in utero Phase

April 2005

Return to Toxicology Template Introduction

TABLE OF CONTENTS

  1. IDENTIFICATION OF STUDY
  2. GOOD LABORATORY PRACTICE
  3. EXECUTIVE SUMMARY
  4. MATERIALS AND METHODS
    1. TEST SUBSTANCE
    2. TEST SUBSTANCE AS ADMINISTERED
    3. ANIMAL DIET
    4. TEST ANIMALS
    5. EXPERIMENTAL DESIGN
    6. MATING PROCEDURES FOR IN UTERO PHASE
    7. CULLING PROCEDURES FOR IN UTERO PHASE
    8. PARENTAL BODY WEIGHT AND FEED INTAKE FOR IN UTERO PHASE
    9. PARENTAL CAGE-SIDE OBSERVATIONS FOR IN UTERO PHASE
    10. PARENTAL REPRODUCTIVE PARAMETERS FOR IN UTERO PHASE
    11. OFFSPRING OBSERVATIONS AND BODY WEIGHT FOR IN UTERO PHASE
    12. BODY WEIGHT AND FEED INTAKE FOR CHRONIC PHASE
    13. CAGE-SIDE OBSERVATIONS FOR CHRONIC PHASE
    14. OPTHALMOLOGICAL EXAMINATION
    15. HEMATOLOGY
    16. CLINICAL CHEMISTRY
    17. URINALYSIS
    18. OTHER TESTS
    19. NECROPSY (INTERIM SACRIFICE)
    20. NECROPSY (TERMINAL)
    21. GROSS PATHOLOGY OBSERVATIONS
    22. HISTOPATHOLOGY OBSERVATIONS
    23. STATISTICAL METHODS
  5. Results
    1. DOSE VERIFICATION
    2. INTAKE OF TEST SUBSTANCE
    3. PARENTAL FEED CONSUMPTION CHANGES FOR THE IN UTERO PHASE
    4. FEED EFFICIENCY FOR THE UTERO PHASE
    5. PARENTAL BODY WEIGHT CHANGES FOR THE IN UTERO PHASE
    6. PARENTAL ABNORMAL CAGE-SIDE OBSERVATIONS FOR THE IN UTERO PHASE
    7. MORTALITY IN ADULTS
    8. MATING, FERTILITY, AND REPRODUCTION
    9. OFFSPRING BODY WEIGHT CHANGES FOR IN UTERO PHASE
    10. OFFSPRING OBSERVATIONS FOR IN UTERO PHASE
    11. FEED CONSUMPTION CHANGES FOR THE CHRONIC PHASE
    12. FEED EFFICIENCY FOR THE CHRONIC PHASE
    13. BODY WEIGHT CHANGES FOR THE CHRONIC PHASE
    14. CAGE-SIDE OBSERVATIONS FOR THE CHRONIC PHASE
    15. MORTALITY IN ADULTS
    16. OPTHALMOLOGICAL EXAMINATION
    17. HEMATOLOGY
    18. CLINICAL CHEMISTRY
    19. URINALYSIS
    20. OTHER TESTS
    21. ORGAN WEIGHTS
    22. GROSS PATHOLOGY CHANGES OBSERVED
    23. HISTOPATHOLOGY CHANGES OBSERVED
    24. NEUROTOXICITY
  6. Evaluation and Comments on Study
  7. Summary and Conclusions
    1. BRIEF SUMMARY OF MAJOR FINDINGS FROM THE STUDY
    2. RELATIONSHIP BETWEEN DOSE AND INCIDENCE/ SEVERITY OF LESIONS OR ABNORMALITIES
    3. IS THERE A TARGET ORGAN?
    4. NOEL
  8. REFERENCES
  9. APPENDIX
    1. HEMATOLOGY RESULTS
    2. CLINICAL CHEMISTRY RESULTS
    3. URINALYSIS RESULTS

CHRONIC TOXICITY STUDY WITH AN IN UTERO PHASE2 

DATE OF SUBMISSION:
TITLE OF PETITION OR NOTIFICATION:
NAME AND ADDRESS OF PETITIONER OR NOTIFIER:

I. IDENTIFICATION OF STUDY3 

A.    STUDY FILE LOCATION:
B.    STUDY TITLE/REPORT NUMBER:
C.    NAME AND ADDRESS OF TESTING FACILITY:
D.    DATE OF STUDY REPORT:
E.    DATES STUDY CONDUCTED:
F.    STUDY OBJECTIVE:
G.    COMMENTS:

II. GOOD LABORATORY PRACTICE4 

A.    WAS THERE COMPLIANCE WITH GOOD LABORATORY PRACTICE REGULATIONS?
B.    WAS A QUALITY ASSURANCE (QA) STATEMENT INCLUDEDd?
C.    AVAILABILITY AND LOCATION OF ORIGINAL DATA/SPECIMENS/TEST SUBSTANCE:

III. EXECUTIVE SUMMARY5   

IV. MATERIALS AND METHODS

A. TEST SUBSTANCE6 

1.     CAS NAME:
2.     OTHER NAME(S):
3.     CAS REGISTRY NUMBER:
4.     MOLECULAR STRUCTURE: 7 
5.     PURITY:
6.     IMPURITIES:
7.     STABILITY:
8.     COMMENTS:

B. TEST SUBSTANCE AS ADMINISTERED8 

1.     BATCH/LOT NUMBER:
2.     ROUTE:
3.     VEHICLE USED:
4.     TESTED ADEQUATELY FOR CONCENTRATION?
5.     TESTED FOR HOMOGENEITY?
6.     TESTED FOR STABILITY?
7.     PROBLEMS WITH STORAGE?

C. ANIMAL DIET

1. FEED

  1. TYPE:
  2. NAME:
  3. AVAILABILITY:
  4. ANALYSIS FOR CONTAMINANTS:
  5. COMMENTS:

2. WATER

  1. SOURCE:
  2. AVAILABILITY:
  3. ANALYSIS FOR CONTAMINANTS:
  4. COMMENTS:

D. TEST ANIMALS9 

1.     SPECIES/STRAIN/SUBSTRAIN:
2.     SEX:
3.     AGE RANGE AT INITIATION OF STUDY:
4.     WEIGHT RANGE AT INITIATION OF STUDY:
5.     QUARANTINE/ACCLIMATION?
6.     PHYSICAL EXAMINATION TIMES:
7.     NUMBER PER CAGE:
8.     ENVIRONMENTAL CONDITIONS:

  • TEMPERATURE:
  • HUMIDITY:
  • AIR CHANGES:
  • PHOTOPERIOD:

9.     COMMENTS:

E. EXPERIMENTAL DESIGN10 

1. TARGETED DOSE LEVELS; IN UTERO PHASE FOR PARENTAL ANIMALS.

TABLE # [HEADING]

TEST GROUP CONC. IN DIET
(ppm or mg/kg)		 NUMBER OF MALES DOSE TO MALES
(mg/kg body-weight/day		 NUMBER OF FEMALES  DOSE TO FEMALES
(mg/kg body-weight/day)
CONTROL          
LOW          
MID          
HIGH          

2. TARGETED DOSE LEVELS FOR THE ANIMALS IN THE CHRONIC PHASE:

TABLE # [HEADING]

TEST GROUP CONC. IN DIET
(ppm or mg/kg)		 NUMBER OF MALES DOSE TO MALES
(mg/kg body-weight/day)		 NUMBER OF FEMALES
 		 DOSE TO FEMALES
(MG/KG BODY-WEIGHT/DAY)
CONTROL          
LOW          
MID          
HIGH          

3. TOTAL NUMBER OF ANIMALS:
4. DURATION OF STUDY (INCLUDING RECOVERY PERIOD, IF ANY):
5. LENGTH OF EXPOSURE TO TEST SUBSTANCE:
6. WERE ANIMALS RANDOMIZED
7. RECOVERY PERIOD:
8. COMMENTS:

IVA. MATERIALS AND METHODS - IN UTERO PHASE

F. MATING PROCEDURES FOR IN UTERO PHASE11 

1. DESCRIPTION:
2. COMMENT:

G. CULLING PROCEDURES FOR IN UTERO PHASE12 

1. DESCRIPTION:
2. COMMENT:

H. PARENTAL BODY WEIGHT AND FEED INTAKE FOR IN UTERO PHASE 13

1. PARAMETER EXAMINED:
2. FREQUENCY OF MEASUREMENT:

TABLE # [HEADING]

EXAMINED NOT EXAMINED
  Feed Intake,
Feed Spillage,
Water Intake,
Body Weight,
Body Weight Changes

3. COMMENT:

I. PARENTAL CAGE-SIDE OBSERVATIONS FOR IN UTERO PHASE13

1. PARAMETER EXAMINED:
2. FREQUENCY OF MEASUREMENT:

TABLE # [HEADING]

EXAMINED NOT EXAMINED
  Appearance,
Abnormal Stool,
Deficiencies in care**
Morbidity,
Mortality,
Neurotoxicity Screening (Specify parameters)***

** Deficiencies in care (inadequate construction or cleaning of the nest, pups left scattered and cold, physical abuse of pups, or inadequate lactation or feeding]
*** The parameters for neurotoxicity screening may include, but are not limited to, the following:

  • Changes in skin, fur, eyes, mucous membranes, gut, posture, and response to handling,
  • Occurrence of secretions/excretions or other evidence of autonomic activity such as lacrimation, piloerection, pupil size change, unusual respiratory pattern
  • Presence of clonic or tonic seizure
  • Stereotype behaviors such as excessive grooming and repetitive circling,
  • Bizarre behavior such as self-mutilating and walking backwards,
  • Gross tumor development.

3. COMMENT:

J. PARENTAL REPRODUCTIVE PARAMETERS FOR IN UTERO PHASE13 

1. REPRODUCTIVE PARAMETERS EXAMINED:

TABLE # [HEADING]

EXAMINED NOT EXAMINED
  Female fertility Index,
Gestation Index,
Gestation Length,
Live-born Index, Number (Total and Per Litter) of Stillbirths and Live Births at Day 0

2. INDICES FORMULAS:
3. COMMENTS:

K. OFFSPRING OBSERVATIONS AND BODY WEIGHT FOR IN UTERO PHASE13

1. PARAMETER EXAMINED:
2. FREQUENCY OF MEASUREMENT:

TABLE # [HEADING]

OBSERVATION EXAMINED NOT EXAMINED
CAGE- SIDE   Appearance,
Gross Anomalies,
Morbidity,
Mortality
OTHER   Litter size (Total: Day 0, 4, 7, 14, 21),
Litter Weight (Day 0, 4, 7, 14, 21),
Pup Body Weight and Sex (Day 0, 4, 7, 14, 21),
Sex Ratio,
Viability Index (Day 0-4, 4-7, 7-14, 14-21),
Weaning Index

3. COMMENTS:

IVB. MATERIALS AND METHODS - CHRONIC PHASE

L. BODY WEIGHT AND FEED INTAKE FOR CHRONIC PHASE13

1. PARAMETER EXAMINED:
2. FREQUENCY OF MEASUREMENT:

TABLE # [HEADING]

EXAMINED* NOT EXAMINED*
  Feed Intake,
Feed Spillage,
Water Intake,
Body Weight,
Body Weight Changes

*These parameters are recommended in Toxicological Principles for the Safety of Food Ingredients: Redbook 2000 for chronic toxicity studies.

3. COMMENTS:

M. CAGE-SIDE OBSERVATIONS FOR CHRONIC PHASE13 

1. PARAMETER EXAMINED:
2. FREQUENCY OF MEASUREMENT:

TABLE # [HEADING]

EXAMINED* NOT EXAMINED*
  Appearance,
Abnormal Stool,
Morbidity,
Mortality,
Neurotoxicity Screening (Specify parameters)**

* These parameters are recommended in Toxicological Principles for the Safety of Food Ingredients: Redbook 2000 for chronic toxicity studies.
** The parameters for neurotoxicity screening may include, but are not limited to, the following:

  • Changes in skin, fur, eyes, mucous membranes, gait, posture, and response to handling,
  • Occurrence of secretions/excretions or other evidence of autonomic activity such as lacrimation, piloerection, pupil size change, unusual respiratory pattern,
  • Presence of clonic or tonic seizure,
  • Stereotype behaviors such as excessive grooming and repetitive circling,
  • Bizarre behavior such as self-mutilating and walking backwards,
  • Gross tumor development.

3. COMMENTS:

N. OPTHALMOLOGICAL EXAMINATION

1. PARAMETER EXAMINED:
2. FREQUENCY OF MEASUREMENT:
3. COMMENTS:

O. HEMATOLOGY13 

1. FASTING DURATION PRIOR TO BLOOD COLLECTION:
2. SCHEDULE INTERVAL FOR BLOOD SAMPLE COLLECTION:
3. METHOD OF BLOOD COLLECTION:
4. DOSE GROUPS AND NUMBER OF ANIMALS TESTED:
5. PARAMETER EXAMINED:

TABLE # [HEADING]

MEASUREMENT
RELATED TO

 EXAMINED* NOT EXAMINED*
RED BLOOD CELLS
   Hematocrit (Hct),
Hemoglobin Conc. (Hb),
Mean Corp. Hb. (MCH),
Mean Corp. Hb. Conc. (MCHC),
Mean Corp. Volume (MCV),
Total Erythrocyte Count (RBC)
WHITE BLOOD CELLS
   Basophils,
Eosinophils,
Lymphocytes,
Macrophage/Monocytes,
Neutrophils,
Total Leukocytes (WBC)
CLOTTING POTENTIAL 
   Activated Partial-Thromboplastin Time,
Clotting Time,
Platelet Count Prothrombin Time
OTHERS 
   Bone marrow cytology,
Reticulocyte counts

* These parameters are recommended in Toxicological Principles for the Safety of Food Ingredients: Redbook 2000 for chronic toxicity studies.

6. Comment:

P. CLINICAL CHEMISTRY13 

1.     FASTING DURATION PRIOR TO BLOOD COLLECTION:
2.     SCHEDULE INTERVAL FOR BLOOD SAMPLE COLLECTION:
3.     METHOD OF BLOOD COLLECTION:
4.     DOSE GROUPS AND NUMBER OF ANIMALS TESTED:
5.     PARAMETER EXAMINED:

TABLE # [HEADING]

MEASUREMENT RELATED TO

 EXAMINED NOT EXAMINED
ELECTROLYTE BALANCE
   Calcium*,
Chloride*,**,
Phosphorus*,
Potassium*,**,
Sodium*,**
BLOOD SUGAR
   Glucose*,**

liver function:
A) HEPATO-CELLULAR (RECOMMEND AT LEAST 3 OUT OF 5)

B) HEPATOBILIARY (RECOMMEND AT LEAST 3 OUT OF 5)

   Alanine Aminotransferase (ALT or SGPT)*,**
,Aspartate Aminotransferase (AST or SGOT)*,
Glutamate Dehydrogenase*,
Sorbitol Dehydrogenase*,
Total Bile Acids*
  Alkaline Phosphatase (ALP)*,**,
Gamma-Glutamyl Transferase (GGT)*,**,
Total Bile Acids*,
Total Bilirubin*,
5' Nucleotidase*
KIDNEY FUNCTION
   Creatinine*,**,
Urea Nitrogen*,**
OTHERS (ACID/BASE BALANCE, CHOLINESTERASES, HORMONES, LIPIDS, METHEMOGLOBIN, AND PROTEINS)
   Albumin (A)*,
Globulin* (G, calculated) or A/G Ratio*,
Total Cholesterol*,
Cholinesterase*,
Total protein*,**,
Fasting Triglycerides*

* These parameters are recommended in Toxicological Principles for the Safety of Food Ingredients: Redbook 2000 for chronic toxicity studies.
** These parameters should generally be given priority when adequate volumes of blood samples can not be obtained from test animals.

6.     COMMENTS:

Q. URINALYSIS14 

1.     Describe the frequency of urine sample collection:
2.     Parameter examined:

TABLE # [HEADING]

examined not examined
  Glucose*,
Microscopic Evaluation for Sediment and presence of blood/blood cells*,
pH*,
Protein*,
Specific Gravity*,
Volume*

* These parameters are recommended in Toxicological Principles for the Safety of Food Ingredients: Redbook 2000 for chronic toxicity studies.

3.     COMMENTS:

R. OTHER TESTS15 

1.     Observations:
2.     COMMENTS:

S. NECROPSY (INTERIM SACRIFICE)16 

1.     Was there an interim sacrifice?
2.     Dose groups and number of animals:
3.     Organs/Tissues weighed:

TABLE # [HEADING]

examined not examined
  Adrenals*,
Brain*,
Epididymides*,
Heart*,
Kidneys*,
Liver*, Spleen*,
Testes*,
Thyroid/parathyroid*,
Thymus*,
Ovaries*,
Uterus*

* These parameters are recommended in Toxicological Principles for the Safety of Food Ingredients: Redbook 2000 for chronic toxicity studies.

4.     COMMENTS:

T. NECROPSY (TERMINAL)

1.     Organs/Tissues weighed:

TABLE # [HEADING]

examined not examined
  Adrenals*, Brain*, Epididymides*,
Heart*, Kidneys*, Liver*, Spleen*,
Testes*, Thyroid/parathyroid*,
Thymus*, Ovaries*, Uterus*

* These parameters are recommended in Toxicological Principles for the Safety of Food Ingredients: Redbook 2000 for chronic toxicity studies.

2.     COMMENTS:

U. GROSS PATHOLOGY OBSERVATIONS

1.     Organs/Tissues Examined:
2.     COMMENTS:

V. HISTOPATHOLOGY OBSERVATIONS

1.     Organs/Tissues were collected from which dose groups?
2.     Organs/Tissues were examined from which dose groups?
3.     How were the organs/tissues prepared for histopathology observation?
4.     Organs/Tissues collected:

TABLE # [HEADING]

system

 examined not examined

digestive

   Salivary Gland*, Esophagus*,
Stomach*, Duodenum*,
Jejunum*, Ileum*, Cecum*,
Colon, Rectum*, Gall Bladder* (in case of mice),
Liver* (middle, left and triangular lobes), Pancreas*
respiratory
   Nasal Turbinates*, Trachea*,
Lung* (with main-stem bronchi)
cardio-vascular
   Aorta*, Heart*  
reticulo- endothelial/hematopoietic 
   Bone Marrow *(sternum),
Lymph Nodes* (1 related to route of administration, and 1 from a distant location),
Spleen*, Thymus*  
urogenital 
   Kidneys*, Ovaries* and fallopian tubes*,
Corpus Uteri*, Cervix Uteri*,
Prostate*, Seminal Vesicle* (if present),
Testes*, Urinary Bladder*, Vagina* 
neurologic
   Brain* (at least 3 different levels),
Spinal-Cervical*,
Spinal-Lumbar*,
Spinal-Midthoracic*,
Sciatic Nerve*,
Harderian Gland* (if present)
glandular
   Adrenals*, Mammary Glands*,
Pituitary Glands*,
Thyroid/Parathyroid Glands*,
Thymus*,
Zymbal's Gland* (if present)
other
   Bone (Femur)*, Eyes*, Skeletal Muscle*,
Skin*, Epididymis*

* These parameters are recommended in Toxicological Principles for the Safety of Food Ingredients: Redbook 2000 for chronic toxicity studies.

5.     COMMENTS:

W. STATISTICAL METHODS

1.     Methods of statistical analysis:

TABLE # [HEADING]

methods of statistical analysis parameters tested
   
   
   
   

2.     COMMENTS:

V. Results

A. DOSE VERIFICATION17 

1.     Were doses verified?

TABLE # [HEADING]

dose group targeted concentration
(ppm or mg/kg)		  concentrations found in feed
(ppm or mg/kg)		 standard
Deviation		 N* 
low
        
mid
        
high
        

* Number of measurements (N)

2.     Verified by:
3.     COMMENTS:

B. INTAKE OF TEST SUBSTANCE18

1.     Intake during in utero phase:

TABLE # [HEADING]

dose group daily dose
(mg/kg body-weight/day)
control
0
low  
mid  
high  

2.     Intake during the chronic phase:

Table # [Heading]

dose group daily dose
(mg/kg body-weight/day)
control
0
low  
mid  
high  

3.     COMMENTS:

Va. Results - In Utero Phase

C. PARENTAL FEED CONSUMPTION CHANGES FOR THE IN UTERO PHASE19 

1.     Observations during Pre-mating and Post-mating:
2.     Observations during Gestation:
3.     Observations during Lactation:
4.     COMMENTS:

D. FEED EFFICIENCY FOR THE UTERO PHASE20

1. Was feed efficiency calculated?
2. COMMENTS:

E. PARENTAL BODY WEIGHT CHANGES FOR THE IN UTERO PHASE21 

1. Observations during Premating and Postmating:
2. Observations during Gestation:
3. Observations during Lactation:
4. COMMENTS:

F. PARENTAL ABNORMAL CAGE-SIDE OBSERVATIONS FOR THE IN UTERO PHASE22 

1. Observations during Premating and Postmating:
2. Observations during Gestation:
3. Observations during Lactation:
4. COMMENTS:

G. MORTALITY IN ADULTS 23 

1.     Observations:
2.     COMMENTS:

H. MATING, FERTILITY, AND REPRODUCTION

1. Observations:

TABLE # [HEADING]

PREGNANCY
  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		      
Number of Females Paired
        
Number of Females Achieving Pregnancy
        
Female Fertility Index (%)
        
MATERNAL WASTAGE
  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		      
Number Died
        
Number Died Pregnant
        
Number Died Nonpregnant
        
Number Aborted
        
Number Premature Delivery
        
GESTATION
  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		      
Gestation Length
Mean
(S.D.)
      
Gestation Index
        
CORPORA LUTEA
  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		      
Total Number Corpora Lutea
        
Corpora Lutea/Dam
Mean
(S.D.)
      
IMPLANTATIONS
  Females
0
(Control)		      
Total Number Implantations
        
Implantations/Dam
Mean
(S.D.)
      
BIRTHS - DAY 0
  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		      
Number Stillborn - Total Per Litter
Mean
(SD)
N
      
Number Live-Born - Total Per Litter
Mean
(SD)
N
      
Live-Born Index (%)        

(Specify a method of statistical analysis): * p<0.05, **=""><>

2. COMMENTS:

I. OFFSPRING BODY WEIGHT CHANGES FOR IN UTERO PHASE24

1. Observations:

 

TABLE # [HEADING]

Pup Weight (gm)
 Sex  Males  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		       0
(Control)		      
Day 0
Mean
(SD)
N
              
Day 4
Mean
(SD)
N
              
Day 7
Mean
(SD)
N
              
Day 14
Mean
(SD)
N
              
Day 21
Mean
(SD)
N
              

(Specify a method of statistical analysis): * p<0.05, **=""><>

2. COMMENTS:

J. OFFSPRING OBSERVATIONS FOR IN UTERO PHASE 25 

1. Observations:

TABLE # [HEADING]

DAILY DOSE
(mg/kg body-weight/day)
 0
(Control)		      
LITTER SIZE
Number Born - Total Per Litter
Mean
(SD)
N
      
Day 0 - Total Per Litter
Mean
(SD)
N
      
Day 4 -Total Per Litter
Mean
(SD)
N
      
Day 7 -Total Per Litter
Mean
(SD)
N
      
Day 14 - Total Per Litter
Mean
(SD)
N
      
Day 21 - Total Per Litter
Mean
(SD)
N
      
LITTER WEIGHT (G)
Day 0
Mean
(SD)
N
      
Day 4
Mean
(SD)
N
      
Day 7
Mean
(SD)
N
      
Day 14
Mean
(SD)
N
      
Day 21
Mean
(SD)
N
      
VIABILITY INDICES
Day 0-4
Mean
(SD)
N
      
Day 4-7
Mean
(SD)
N
      
Day 7-14
Mean
(SD)
N
      
Day 14-21
Mean
(SD)
N
      
WEANING INDEX
        
SEX RATIO
        
Day 0
        
Day 21
        

(Specify a method of statistical analysis): * p<0.05, **=""><>

3.     COMMENTS:

Vb. Results - Chronic Phase

K. FEED CONSUMPTION CHANGES FOR THE CHRONIC PHASE26

1. Observations:
2. feed consumption table (and/or graph)27 :

TABLE # [HEADING]

 FEED CONSUMPTION (gm feed/ kg body-weight/day)
 Sex  Males  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		     0
(Control)		      
Day 0
Mean
(SD)
N
              
Day A
Mean
(SD)
N
              
Day B
Mean
(SD)
N
              
Day C
Mean
(SD)
N
              
Day D
Mean
(SD)
N
              

(Specify a method of statistical analysis): * p<0.05, **=""><>

3. COMMENTS:

L. FEED EFFICIENCY FOR THE CHRONIC PHASE28 

1. Was feed efficiency calculated?
2. COMMENTS:

M. BODY WEIGHT CHANGES FOR THE CHRONIC PHASE29 

1. Observations:
2. body weight table (and/or graph)30 :

TABLE # [HEADING]

 body weight (gm/day)
 Sex  Males  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		       0
(Control)		      
Day 0
Mean
(SD)
N
              
Day A
Mean
(SD)
N
              
Day B
Mean
(SD)
N
              
Day C
Mean
(SD)
N
              
Day D
Mean
(SD)
N
              

(Specify a method of statistical analysis): * p<0.05, **=""><>

3. COMMENTS:

N. CAGE-SIDE OBSERVATIONS FOR THE CHRONIC PHASE

1. observation31 :

TABLE # [HEADING]

 cage-side observation (specify parameters32 )
 Sex  Males  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		       0
(Control)		      
Day 0
N
              
Day A
N
              
Day B
N
              
Day C
N
              
Day D
N
              

N: Number of Animals with abnormal observation

(Specify a method of statistical analysis): * p<0.05, **=""><>

2. COMMENTS:

O. MORTALITY IN ADULTS33

1. Observations:
2. mortality table (and/or graph)34 :

TABLE # [HEADING]

 mortality (%)
 Sex  Males  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		       0
(Control)		      
Day 0                
Day A                
Day B                
Day C                
Day D                

(Specify a method of statistical analysis): * p<0.05, **=""><>

3. COMMENTS:

P. OPTHALMOLOGICAL EXAMINATION

1. Observations:
2. COMMENTS:

Q. HEMATOLOGY

1. Observations35 :

TABLE # [HEADING]36 

(specify parameter)
 Sex  Males  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		       0
(Control)		      
Day 0
Mean
(SD)
N
              
Day A
Mean
(SD)
N
              
Day B
Mean
(SD)
N
              
Day C
Mean
(SD)
N
              
Day D
Mean
(SD)
N
              

(Specify a method of statistical analysis): * p<0.05, **=""><>

3. COMMENTS:

R. CLINICAL CHEMISTRY37 

1. Observations:

TABLE # [HEADING]38 

 hematology results (specify parameter)
 Sex  Males  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		       0
(Control)		      
Day 0
Mean
(SD)
N
              
Day A
Mean
(SD)
N
              
Day B
Mean
(SD)
N
              
Day C
Mean
(SD)
N
              
Day D
Mean
(SD)
N
              

(Specify a method of statistical analysis): * p<0.05, **=""><>

2. COMMENTS:

S. URINALYSIS39 

1. Observations:

TABLE # [HEADING]40 

 urinalysis results (specify parameter)
 Sex  Males  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		       0
(Control)		      
Day 0
Mean
(SD)
N
              
Day A
Mean
(SD)
N
              
Day B
Mean
(SD)
N
              
Day C
Mean
(SD)
N
              
Day D
Mean
(SD)
N
              

(Specify a method of statistical analysis): * p<0.05, **=""><>

2. COMMENTS:

T. OTHER TESTS41 

1. Observations:

TABLE # [HEADING]42 

 specify parameter
 Sex  Males  Females
DAILY DOSE
(mg/kg body-weight/day)		 0
(Control)		       0
(Control)		      
Day 0
Mean
(SD)
N
              
Day A
Mean
(SD)
N
              
Day B
Mean
(SD)
N
              
Day C
Mean
(SD)
N
              
Day D
Mean
(SD)
N
              

(Specify a method of statistical analysis): * p<0.05, **=""><>

2. COMMENTS:

U. ORGAN WEIGHTS43 

1.     Observations:

TABLE # [HEADING]

 sex  males  females
DAILY DOSE
(mg/kg body-weight/day)
 0
(CONTROL)
 
 
 
 0
(CONTROL)
 
 
 
NUMBER OF ANIMALS
 
 
 
 
 
 
 
 
BODY WEIGHT (gram)a
 
 
 
 
 
 
 
 
BRAIN
Absolute Weighta
gram
                
Per Body Weighta
%
                
ADRENALS
Absolute Weighta
gram
                
Per Body Weighta
%
                
Per Brain Weighta
%
                
EPIDIDYMIDES
  
Absolute Weighta
gram
        
Per Body Weighta
%
        
Per Brain Weighta
%
        
HEART
Absolute Weighta
gram
                
Per Body Weighta
%
                
Per Brain Weighta
%
                
KIDNEYS
Absolute Weighta
gram
                
Per Body Weighta
%
                
Per Brain Weighta
%
                
LIVER
Absolute Weighta
gram
                
Per Body Weighta
%
                
Per Brain Weighta
%
                
SPLEEN
Absolute Weighta
gram
                
Per Body Weighta
%
                
Per Brain Weighta
%
                
TESTES
  
Absolute Weighta
gram
        
Per Body Weighta
%
        
Per Brain Weighta
%
        
THYROID and PARATHYROID
Absolute Weighta
gram
                
Per Body Weighta
%
                
Per Brain Weighta
%
                
THYMUS
Absolute Weighta
gram
                
Per Body Weighta
%
                
Per Brain Weighta
%
                
OVARIES
    
Absolute Weighta
gram
        
Per Body Weighta
%
        
Per Brain Weighta
%
        
UTERUS
    
Absolute Weighta
gram
        
Per Body Weighta
%
        
Per Brain Weighta
%
        

a: Group means at the end of terminal necropsy are shown.

(Specify methods of statistical analysis): * p<0.05, **=""><>

2.     COMMENTS:

V. GROSS PATHOLOGY CHANGES OBSERVED44 

1.     Observations:
2.     COMMENTS:

W. HISTOPATHOLOGY CHANGES OBSERVED45 

1.     Observations:

Table # [Heading)

NUMBER OF ANIMALS WITH, GROSS, NON-NEOPLASTIC, OR NEOPLASTIC LESIONS
SEX
  males  females
DAILY DOSE
(mg/kg body-weight/day)
 0
(CONTROL)		       0
(CONTROL)		      
NUMBER OF ANIMALS EXAMINED
                
DIGESTIVE SYSTEM
ORGAN/TISSUE #
                
GROSS LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
                 
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
                 
RESPIRATORY SYSTEM
ORGAN/TISSUE #
                
GROSS LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
                 
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
                 
CARDIOVASCULAR SYSTEM
ORGAN/TISSUE #
                
GROSS LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
                 
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
                 
RETICULO-ENDOTHELIAL /HEMATOPOIETIC SYSTEM
ORGAN/TISSUE #
                
GROSS LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
                 
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
                 
UROGENITAL SYSTEM
ORGAN/TISSUE #
                
GROSS LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
                 
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
                 
GLANDULAR SYSTEM
ORGAN/TISSUE #
                
GROSS LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
                 
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                

(Specify methods of statistical analysis): * p<0.05, **=""><>

# Organs/tissues listed under section IV.V.

In general, data at end of dosing period can be shown; however, if there were additional noteworthy findings at earlier timepoints, these should be included. Note severity of lesions as needed.

2.     COMMENTS:

X. NEUROTOXICITY46 

1.     Observations:

TABLE # [HEADING]

   NUMBER OF ANIMALS
 
SEX
  males  females
DAILY DOSE
(mg/kg body-weight/day)
 0
(CONTROL)		       0
(CONTROL)		      
NUMBER OF ANIMALS EXAMINED
                
OBSERVATIONS OF NERVOUS SYSTEM TOXICITY +
OBSERVATION
                
OBSERVATION
                
OBSERVATION
                
                 
GROSS- AND HISTO-PATHOLOGY CHANGES IN THE NEUROLOGIC SYSTEM #
ORGAN/TISSUE
                
GROSS LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
NON-NEOPLASTIC LESION
                
                 
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                
NEOPLASTIC LESIONS
                

+ See under section IV.M for the types of observation for nervous system toxicity: List noteworthy findings. If additional parameters (other than those in the Template) showed noteworthy changes, these should be added to the tables. In general, include data at the end of the dosing period. However, if one observes additional noteworthy findings at earlier timepoints, these should be included. Footnotes should be used as needed to provide additional information about the tests or the results. Note the severity of the abnormal observations using the following scales; + Mild, ++ Moderate, and +++ Marked or other scales, as appropriate.

# Organs/tissues listed under section IV.V

2.     COMMENTS:

VI. Evaluation and Comment on Study47 

VII. Summary and Conclusions48

A. BRIEF SUMMARY OF MAJOR FINDINGS FROM THE STUDY

B. RELATIONSHIP BETWEEN DOSE AND INCIDENCE/ SEVERITY OF LESIONS OR ABNORMALITIES

C. IS THERE A TARGET ORGAN?

D. NOEL

1.     Was there a no observed effect level?
2.     COMMENTS:

VIII. References

IX. Appendix

Vb.Q. HEMATOLOGY RESULTS

TABLE # [HEADING]

day of Sampling49
  SEX  males  females
DAILY DOSE
(mg/kg body-weight/day)
 0
(Control)		       0
(CONTROL)		       
NUMBER OF ANIMALS
                
red blood cells
Hematocrit (Hct)
%
                
Hemoglobin Conc. (Hb)
g/L
                
Mean Corp. Hb. (MCH)
 
                
Mean Corp. Hb. Conc. (MCHC)
 
                
Mean Corp. Volume (MCV)
L/L
                
Total Erythrocyte Count (RBC)
1012/L
                
white blood cells
Basophils
                  
Eosinophils
                  
Lymphocytes
109/L
                
Macrophage/Monocytes
                  
Neutrophils
109/L
                
Total Leukocytes (WBC)
109/L
                
clotting potential
Activated Partial-Thromboplastin Time
                  
Clotting Time
                  
Platelet Count
109/L
                
Prothrombin Time
                  
others
Bone marrow cytology
                  
Reticulocyte counts
1012/L
                

(Specify a method of statistical analysis): * p<0.05, **=""><>

Vb.R. CLINICAL CHEMISTRY RESULTS

TABLE # [HEADING]

day of Sampling49
  SEX  males  females
DAILY DOSE
(mg/kg body-weight/day)
 0
(CONTROL)		       0
(CONTROL)		      
NUMBER OF ANIMALS
                
electrolyte balance
Calcium
mmol/L
                
Chloride
mmol/L
                
Phosphorus
mmol/L
                
Potassium
mmol/L
                
Sodium
mmol/L
                
carbohydrate metabolism
Glucose
mmol/L
                
liver function: A) hepatocellular
Alanine Amino-transferase (ALT or SGPT)
U/L
                
Aspartate Amino-transferase (AST or SGOT)
U/L
                
Glutamate Dehydrogenase
U/L
                
Sorbitol Dehydrogenase
U/L
                
liver function: B) hepatobiliary
Alkaline Phosphatase (ALP)
U/L
                
Gamma-Glutamyl Transferase (GGT)
U/L
                
Total Bile Acids
mmol/L
                
Total Bilirubin
mmol/L
                
5' Nucleotidase
U/L
                
kidney function
Creatinine
mmol/L
                
Urea Nitrogen
mg/dL
                
others
Albumin (A)
g/L
                
Globulin (G, calculated)
g/L
                
A/G Ratio
                  
Total protein
g/L
                
Total Cholesterol
mmol/L
                
Fasting Triglycerides
mmol/L
                
Cholinesterase
U/L
                

(Specify statistical method of analysis): * p<0.05, **=""><>

Vb.S. URINALYSIS RESULTS

TABLE # [HEADING]

day of Sampling49
SEX males females
DAILY DOSE
(mg/kg body-weight/day)
 0
(CONTROL)		       0
(CONTROL)		      
NUMBER OF ANIMALS
                
Glucose
mmol/L
                
Microscopic evaluation for sediment and presence of blood/blood cells
                  
pH
                  
Protein
g/L
                
Specific Gravity
                  
Volume
L/time
                

(Specify statistical method of analysis): * p<0.05, **=""><>

End Notes

This section includes some comments that are only relevant when using the Word template.

 1 Please refer to the Introduction to the Template for general instructions before you begin using this form

 2 Please refer to the Introduction to the Template for general instructions before you begin using this form

 3Make note of: study file location (Volume, pages), study title/report #, testing facility name, publication dates of study, study objective, and comments, if needed.

 4Indicate Yes or No for Questions A and B. However, you may want to elaborate on the type of GLP compliance (USFDA, OECD, etc.) or make other notations.

 5The executive summary should be a brief, concise, stand alone summary describing the test system, the test protocol and the results. It contains three paragraphs: the first describing the test system, the second reporting and discussing the results and the third stating whether the study is acceptable or not. If not acceptable, state why not and what information could be supplied to make it acceptable. A typical executive summary should be less than one page long.

 6Description of the test substance should be given, including purity, any possible contaminants or impurities, and any properties of the test substance that could affect its integrity. Were there factors that have affected the actual administered dose, as opposed to the intended dose?

 7After you put a blinking cursor on the http address, (CTRL +) left-click once to follow the link and wait. Enter a chemical name, CAS Number, or molecular formula inside the search window and click 'search' button. Right-click inside the molecular structure and select 'copy' submenu. Return to your document and put the cursor underneath item #4 (molecular structure). Right click to open up the menu options and select 'paste' submenu. You can drag the structure to any position you want and resize.

If preferred, the reviewer is free to use other methods of depicting the molecular structure.

 8Indicate how the test substance was given and whether any vehicle was used to dissolve/suspend the test substance (e.g., dissolved in corn oil and mixed into the feed). Also note if there was adequate testing for concentration and homogeneity (appropriate tests with replicates) and whether there were stability tests done. Was the strength of the test material checked over the period and under the conditions that the feed mix, test article in vehicle, etc., would be stored? Note anything that was not normal or routine.

 9Note anything that might have affected the study. Use the comments to indicate extremes or other factors that might have impacted the study.

 10Provide adequate details. Use the comments to indicate additional information about the experimental design. To change the table, place cursor in the row or column that you wish to modify, then from the 'Table' menu, use 'Select Row' or 'Select Column' to highlight the row or column that you wish to change, and use the 'delete' or 'insert' functions to make your changes.

 11Describe how animals were mated. Indicate when and how females were examined for pregnancy. Note if sibling mating was avoided.

 12Standardization of the number of pups per litter is optional. If performed, describe how the litters were standardized (on what day, standardized to how many, if standardized in a random manner, etc.).

 13Indicate the measurements or observations that were made by highlighting the parameter and dragging the term into the "Examined" side. Those parameters not examined will remain in the "Not Examined" side. Use the comments to indicate any other notable fact.

 14Note when urine samples were collected for testing, the groups that were affected, and drag and drop the parameters that were tested. Make comments on anything significant or treatment-related.

 15When other tests are conducted, make note of the tests and any significant treatment-related effects.

 16If there was an interim sacrifice (e.g., at 3 month intervals), make notes on which groups were affected, and the number and sex of the animals that were tested.

 17Note here whether there was a check or verification of the doses being administered (by what analytical methodology and in which laboratory). If the test substance was mixed in the feed, was there a verification for substance concentration in the feed? Was this verification done more than once? Were there adequate data to permit the calculation of the actual dose that was administered?

 18Knowing what the intake of feed was and the level of test substance in the feed, what was the dose actually being delivered to the animals?

 19Note any statistically or biologically significant feed consumption changes. Note feed/water consumption changes that were not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend.

 20Note in this section if feed efficiency was determined; when in the study determinations were made; if any changes were observed; if feed efficiency was calculated for all groups or only selected groups; and if calculations were made per group, per cage, or per animal.

 21Note any statistically or biologically significant body weight changes. Note body weight changes that are not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend. You may wish to insert graphs or tables in this section.

 22List significant, dose-related abnormal cage-side observations reported. This may include neurotoxicity endpoints.

 23For each animal with an unscheduled death, note the group, date of death, and any observations, including lesions observed at necropsy.

 24Note any statistically or biologically significant body weight changes. Also note body weight changes that were not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend. The comment section is available to explain data findings, if needed.

25List significant, dose-related abnormal observations reported.

26Note any statistically or biologically significant feed consumption changes. Also note feed consumption changes that were not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend.

27To change the table, place cursor in the row or column that you wish to modify, then from the 'Table' menu, use 'Select Row' or 'Select Column' to highlight the row or column that you wish to change, and use the 'delete' or 'insert' functions to make your changes.

28Note in this section if feed efficiency was determined; when in the study determinations were made; if any changes were observed; if feed efficiency was calculated for all groups or only selected groups; and if calculations were made per group, per cage, or per animal.

 29Note any statistically or biologically significant body weight changes. Also note body weight changes that were not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend. You may wish to insert graphs or tables in this section.

 30To change the table, place cursor in the row or column that you wish to modify, then from the 'Table' menu, use 'Select Row' or 'Select Column' to highlight the row or column that you wish to change, and use the 'delete' or 'insert' functions to make your changes.

 31To change the table, place cursor in the row or column that you wish to modify, then from the 'Table' menu, use 'Select Row' or 'Select Column' to highlight the row or column that you wish to change, and use the 'delete' or 'insert' functions to make your changes.

 32List significant, dose-related abnormal cage-side observations reported. For neurotoxicological observations, also use a Table in Section V. X. (see Section IV. M. for parameters).

 33For each animal with an unscheduled death, note the group, date of death, and any observations, including lesions observed at necropsy.

 34To change the table, place cursor in the row or column that you wish to modify, then from the 'Table' menu, use 'Select Row' or 'Select Column' to highlight the row or column that you wish to change, and use the 'delete' or 'insert' functions to make your changes.

 35Note findings that were statistically or biologically treatment-related. Also note changes that were not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend. The comment section is available to explain data findings, if needed. Note detailed results in Appendix.

 36To change the table, place cursor in the row or column that you wish to modify, then from the 'Table' menu, use 'Select Row' or 'Select Column' to highlight the row or column that you wish to change, and use the 'delete' or 'insert' functions to make your changes.

 37Note findings that were statistically or biologically treatment-related. Also note changes that were not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend. The comment section is available to explain data findings, if needed. Note detailed results in Appendix.

 38To change the table, place cursor in the row or column that you wish to modify, then from the 'Table' menu, use 'Select Row' or 'Select Column' to highlight the row or column that you wish to change, and use the 'delete' or 'insert' functions to make your changes.

 39Note treatment-related findings that were statistically or biologically significant. Also note changes that were not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend. The comment section is available to explain data findings, if needed. Note detailed results in Appendix.

 40To change the table, place cursor in the row or column that you wish to modify, then from the 'Table' menu, use 'Select Row' or 'Select Column' to highlight the row or column that you wish to change, and use the 'delete' or 'insert' functions to make your changes.

 41When other tests are conducted, make note of the tests and any significant treatment-related effects.

 42To change the table, place cursor in the row or column that you wish to modify, then from the 'Table' menu, use 'Select Row' or 'Select Column' to highlight the row or column that you wish to change, and use the 'delete' or 'insert' functions to make your changes.

 43Note treatment-related findings that were statistically or biologically significant. Also note changes that were not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend. The comment section is available to explain data findings, if needed.

 44Note treatment-related findings that were statistically or biologically significant. Also note changes that were not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend. The type of lesion should be noted. The comment section is available to explain data findings, if needed. Also note the findings in Histopathology Section V.W. below.

 45Note treatment-related findings that were statistically or biologically significant. Also note changes that were not statistically significant (e.g., at low or mid dose) but could indicate a treatment-related trend. The type of lesion should be noted. The comment section is available to explain data findings, if needed.

 46 Summary tables of all positive effects should be prescribed. In addition, provide an explicit statement as to whether or not the test substance represents a potential neurotoxic hazard.

 47Give your evaluation of the study: what was done well; what the problems were; did the study accomplish what it was supposed to do?

 48Summarize the key findings from the study. Was there a dose-effect relationship? Were target organs identified? Was a NOEL achieved for each significant effect/observation? What was the NOEL? The whole study should be summarized in this section.

 49Specify day of sampling (e.g., day 0, day 90). And, create separate tables for each interval.

(Return to Toxicology Template Introduction)

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