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UNITED STATES ENVIRONMENTAL PROTECTION AGENCY 

WASHINGTON D.C., 20460 

VIA FACSIMILE: 201-437-4801 

April 19,2012 

Attention: Glenn Merritt (201-437-4000) 

OFFICE OF 

PREVENTION, PESTICIDES AND 

TOXIC SUBSTANCES 

Subject: Follow-up to fax transmission of 4/13/12 (ePMA removal ofCBI claims regarding 

identified studies) 

Dear Mr. Merritt: 

This is sent to you pursuant our conversation of several minutes ago. In Larry Robinson's fax of 

April 13th while there is a waiver of confidential business information (CBI) claims in the 

identified studies there is a an assertion of copyright protections and some additional text that 

provides in part ... "Reproduction, reference, summarization, retransmission or other use of this 

copyrighted document. .. including electronic transmission, using the internet or email .... is 

strictly prohibited .... " 

While we understand the basis for the copyright protections, we have a concern that the language 

in the fax might be interpreted to mean that ePMA is asserting that the studies submitted per the 

TSCA activity are not to be made available in an Agency e-docket (electronic docket) or be 

included in EPA websites among the collection of TSCA health and safety studies. 

We are thinking that the identified limitations noted in the fax are not intended to apply to EPA's 

normal procedures for the handling of non-CBI materials submitted by entities subject to TSCA 

regulations (including the posting of such materials on government websites) and submitted by 

such entities in order to comply with a TSCA requirement, but are rather intended to limit 

entities outside the government from the unauthorized use or appropriation of the studies. 

Can you please confirm CPMA's intent? Many thanks. 

Scott M. Sherlock, Attorney Advisor 

Environmental Assistance Division 

Office of Pollution Prevention and Toxics 


CONFIDENTIAL BUSINESS INFORMATION

APPROVAL STATEMENT

PRELIMINARY FINAL REPORT

Study Title A combined 28-day repeated dose toxicity study with the

reproduction/developmental toxicity screening test of ClI. Pigment

Blue 61 in rats by oral gavage

NOTOX Substance 183591/A

NOTOX Project 488231

STUDY DIRECTOR STATEMENT

The undersigned declares that this preliminary final report constitutes a true account of the

procedures adopted and the results obtained in the performance of this study.

This report has not yet been inspected by the NOTOX Quality Assurance Unit to confirm that

the methods and results accurately and completely reflect the raw data. The audit is planned for

Weeks 47 and 48 2008. Any comments will be incorporated in the final report.

Study director

.. .. . ... . . .. .. . . . . . . . . . .

MWE W. Beekhuijzen, M.Sc. date:

APPROVAL

Management

D~rs. A.W.M. van Rozendaal d ate:
CONFIDENTIAL BUSINESS INFORMATION

PRELIMINARY FINAL REPORT

Study Title

A COMBINED 28-DAY REPEATED DOSE TOXICITY STUDY WITH

THE REPRODUCTIONIDEVELOPMENTAL TOXICITY

SCREENING TEST OF C.I. PIGMENT BLUE 61 IN RATS BY

ORAL GAVAGE

Author

KE.W Beekhuijzen, M.Sc.

Study completion date

Test Facility

NOTOX B.

Hambakenwetering 7

5231 DD's-Hertogenbosch

The Netherlands

Laboratory Study Identification

NOTOX Project 488231 (488232)

NOTOX Substance 183591

-Page 1 of 249 -
C.I. PIGMENT BLUE 61 NOTOX Project 488231

1. CONTENTS

I CONTENTS...........................2

2. STATEMENT OF GLP COMPLIANCE................................................ 4

3. LEAD QUALITY ASSURANCE STATEMENT................................................... 5

4. SUMMARY ....................................................................................... 6

5. INTRODUCTION............................................................ ...................... 7

5.1. Preface ...................................................................................... 7

5.2. Study schedule ............................................................................... 8

5.3. Project numbers .............................................................................. 8

5.4. Aims of study................................................................................. 8

5.5. Guidelines................................................................................... 9

5.6. Storage and retention of records and materials............................................ 9

6. MATERIALS AND METHODS................................................................... 10

6.1. Test substance .............................- ............................................... 10

6. 1.1. Test substance information (170676/A)............................................ 10

6.1.2. Study specific test substance information.......................................... 10

6.1.3. Test substance formulation ......................................................... 10

6.1.4. Chemical analysis of dose preparations ........................................... 11

6.2. Test system................................................................................. 11

6.3. Allocation................................................................................... 12

6.4. Animal husbandry........................................................................... 12

6.5. Treatment.................................................................................. 13

6.5.1. Parental animals..................................................................... 13

6.5.2. Offspring.............................................................................. 13

6.6. Observations ................................................................................ 14

6.6.1. Parental animals ..................................................................... 14

6.6.2. Offspring .................................................................... ......... 15

6.7. Clinical laboratory investigations........................................................... 16

6.8. Pathology Parental Animals................................................................ 17

6.8.1 . Termination........................................................................... 17

6.8.2. Macroscopic examination ........................................................... 18

6.8.3. Organ weights........................................................................ 19

6.8.4. Histotechnology...................................................................... 19

6.8 5. Histopathology ....................................................................... 19

6.9. Pathology Offspring ......................................................................... 20

6.9.1. Termination........................................................................... 20

6.9.2. Macroscopic examination ........................................................... 20

6.10. Calculations .......................................................................... 20

6.11. Electronic data capture .......................................................... ... 21

6.12. Interpretation ......................................................................... 21

6.13. List of deviations ..................................................................... 21

6.13.1. List of protocol deviations ........................................................ 21

6.13.2. List of standard operating procedure deviations ............................... 21

7. RESULTS ......................................................................................... 22

7.1. Analysis of Dose Preparations (see also Appendix 3) ................................... 22

7.2. Observations ................................................................................ 22

7.2.1, Mortality .............................................................................. 22

7.2.2. Clinical Signs......................................................................... 22

7.2.3. Body Weights......................................................................... 22

7.2.4. Food Consumption................................................................... 22

7.2.5. Functional observations ............................................................. 23

7.3. Clinical Laboratory Investigations.......................................................... 23

7.3.1. Haematology ......................................................................... 23

7.3.2. Clinical Biochemistry................................................................. 23

7.4, Pathology ................................................................................... 23

7.4.1. Macroscopic Examination ........................................................... 23

7.4.2. Organ Weights ....................................................................... 24

Page 2
C.I. PIGMENT BLUE 61 NOTOX Project 488231

7.4.3. Microscopic Examination (see also Appendix 4).................................. 24

7.5. Reproduction ................................................................................ 24

7.6. Breeding Data............................................................................... 25

7.7. Pup Development........................................................................... 25

8. DISCUSSION AND CONCLUSION............................................................. 26

9. REFERENCES .................................................................................. 27

APPENDICES

APPENDIX 1 SUMMARY TABLES ............................................................... 1-27

Clinical Signs ...................................................................... 2

Body Weight.............I........................................................... 5

Body Weight Gain ................................................................. 7

Food Consumption ........ ... .................. 9

Relative Food Consumption.................................................. 11

Functional Observations......................................................... 13

Motor Activity test................................................................ 14

Haemnatology ..................................................................... 15

Clinical Biochemistry............................................................. 17

Macroscopic Findings............................................................ 19

Organ weights!/ Organ to Body Weight Ratios ................................ 20

Reproduction Processes ........................................................ 24

Mating Performance ............................................................. 25

Breeding Data per Group ....................................................... 26

Mean Body Weights of Pups.................................................... 27

APPENDIX 2 INDIVIDUAL TABLES ............................................................... 1-85

Mortality data ...................................................................... 2

Clinical Signs ...................................................................... 4

Body Weight ...................................................................... 12

Body Weight Gain................................................................ 15

Food Consumption ........................ 18

Relative Food Consumption..................................................... 20

Functional Observations......................................................... 22

Motor Activity test................................................................ 24

Haemnatology ..................................................................... 26

Clinical Biochemistry............................................................. 31

Macroscopic Findings............................................................ 35

Organ weights / Organ to Body Weight Ratios ................................ 38

Breeding Data per Litter......................................................... 46

Mean Body Weights of Pups per Litter ......................................... 50

Body Weights of Pups........................................................... 53

Individual Pup Data.............................................................. 61

Corpora Lutea .................................................................... 83

Implantation Sites................................................................ 84

Key to Missing Values/Remarks Clinical Laboratory Investigations......... 85

APPENDIX 3 FORMULATION ANALYSIS ........................................................ 1-11

APPENDIX 4 PHASE REPORT HISTOPATHOLOGY ........................................... 1-97

APPENDIX 5 SUMMARY DOSE RANGE FINDING............................................... 1-2

TOTAL NUMBER OF PAGES ............ .......................................................... 249

Page 3
C.I. PIGMENT BLUE 61 NOTOX Project 488231

2. STATEMENT OF GLP COMPLIANCE

NOTOX B.V., 's-Hertogenbosch, The Netherlands

The study described in this report has been correctly reported and was conducted in compliance

with:

The Organization for Economic Cooperation and Development (OECD) Good Laboratory

Practice Guidelines (1997).

Which essentially conform to:

The United States Food and Drug Administration Good Laboratory Practice Regulations.

The United States Environmental Protection Agency Good Laboratory Practice Regulations

The sponsor is responsible for Good Laboratory Practice (GLP) compliance for all test

substance information unless determined by NOTOX

The GLP statement for the delegated phase histopathology is included in Appendix 4 of this

report.

NOTOX B.V.

M.E.W. Beekhuijzen, M.Sc. Drs. A.W.M. van Rozendaal

Study Director Section Head Toxicology

Date:........................................ Date:........................................

Page 4
C.I PIGMENT BLUE 61 NOTOX Project 488231

3. LEAD QUALITY ASSURANCE STATEMENT

NOTOX B. 's-Hertogenbosch, The Netherlands

This report was inspected by the NOTOX Quality Assurance Unit to confirm that the methods

and results accurately and completely reflect the raw data.

The dates of Quality Assurance inspections are given below.

During the on-site process inspections procedures applicable to this type of study were

inspected.

The reporting date is the date of reporting to the Study Director. The QAU report was then

forwarded to the Test Facility Management.

Type of Phase I Section Start End Reporting date

inspections Inspection Inspection

date(s) date(s)

Protocol (Study) DD\MMM\JJ DD\MMM\JJ DD\MMM\JJ

On-site (Process) DD\MMM\JJ DD\MMM\JJ DD\MMM\JJ

On-site (Study) DD\MMM\JJ DD\MMM\JJ DD\MMM\JJ

Report (Study) DD\MM M\JJ DD\MMM\JJ DD\MMM\JJ

The Quality Assurance programme for the delegated phase histopathology was performed by

the Quality Assurance appointed by the test site management and a Quality Assurance

statement is included in Appendix 4 of this report.

Head of Quality Assurance

C.J Mitchell B.Sc.

Date: ...................

Page 5
C.I. PIGMENT BLUE 61 NOTOX Project 488231

4. SUMMARY

Combined repeated dose toxicity study with reproduction / developmental toxicity screening test

with ClI. Pigment Blue 61 administered by oral gavage in Wistar rats.

The purpose of this study was to evaluate the potential toxic effects of 0.1. Pigment Blue 61

when administered to rats for a minimum of 28 days and to evaluate the potential of the test

substance to affect male and female reproductive performance such as gonadal function,

mating behaviour, conception, parturition and early postnatal development.

The study was based on the following guideline:

Organisation of Economic Co-operation and Development Guidelines (OECD) for testing of

Chemicals Guideline 422, Combined Repeated Dose Toxicity Study with the

Reproduction/Developmental Toxicity Screening Test, March 1996.

After acclimatisation, four groups of ten male and ten female Wistar Han rats were exposed by

oral gavage to the test substance at 0, 50, 250 and 1000 mg/kg/day.

Males were exposed for 30 days, i.e. 2 weeks prior to mating, during mating, and up to

termination. Females were exposed for 41 to 45 days, i.e. during 2 weeks prior to mating, during

mating, during post-coitum, and during at least 4 days of lactation.

The following parameters were evaluated: clinical signs, functional observations, body weights,

food consumption, reproduction processes, observations offspring, clinical pathology,

macroscopy, organ weights, and histopathology. Chemical analyses of formulations were

conducted once during the study to assess accuracy, homogeneity and stability.

RESULTS

Accuracy, homogeneity and stability of formulations were demonstrated by analyses.

No treatment-related toxicologically significant changes were noted in any of the parameters

investigated in this study (i.e. mortality, clinical appearance, functional observations, body

weight, food consumption, clinical laboratory investigations, macroscopic examination, organ

weights, microscopic examination, reproduction, breeding and pup development).

CONCLUSION

From the results presented in this report, the parental, reproduction, breeding and

developmental No Observed Adverse Effect Level (NOAEL) for 0.1. PIGMENT BLUE 61 of at

least 1000 mg/kg/day was established.

Page 6
C.I. PIGMENT BLUE 61 NOTOX Project 488231

5. INTRODUCTION

5.1. Preface

This is a multi-site study. The study was performed according to the OECD Consensus

document Number 13 The Application of the OECD Principles of GLP to the Organization and

Management of Multi-Site studies.

Information concerning the delegated phase is given in the applicable appendix.

Phase:

Histopathology Principal Investigator: Dr. A. Pilling, ToxPath Consultancy Ltd,

UK (see Appendix 4)

Sponsor Color Pigments Manufacturers Association, Inc.

300 N. Washington St. Ste. 102,

ALEXANDRIA, VA 22314

USA

Study Monitor Mr. L. Robinson

jlr@cpma.com

Study Director M.EW. Beekhuijzen, M.Sc.

NOTOX B.V.

Hambakenwetering 7

Is-Hertogenbosch

The Netherlands

Tel: +31.73 640 6700

Fax: +31.73 640 6799

Email: manon.beekhuiizen~cnotox.nI

Coordinating Biotechnician M.M.A. Rijkers (NOTOX ByV.)

Clinical Pathology J.E. van Kesteren (NOTOX B.V.)

Analytical Chemistry Ir. M.J.C. Brekelmans (Principal Scientist, NOTOX B.V.)

Histotechnology Ing T.A. Mijnders (NOTOX ByV.)

Necropsy J.H. van den Brink, DVM (NOTOX BYV)

Lead QA C.J. Mitchell, B.Sc. (NOTOX B.V.):

christine.mitchell(ftotox.nl

Test facility Management H.H. Emmen, M.Sc. (NOTOX ByV.):

harrv.emmena-notox. ni

Test Facility NOTOX B.V.

Hambakenwetering 7

5231 DD s-Hertogenbosch

The Netherlands

Page 7
C.I. PIGMENT BLUE 61 NOTOX Project 488231

5.2. Study schedule

Delivery of animals 11 June 2008

Experimental start date 12 June 2008 (allocation)

Start treatment 17 June 2008

Start mating 01 July 2008

Necropsy of males 17 July 2008

Delivery of litters 23 to 27 July 2008

Necropsy of females and pups 28 July to 01 August 2008

Experimental completion date 01 August 2008 (end in-life)

5.3. Project numbers

Two project numbers were used to collect online data. Eventually, all data were reported under

NOTOX Project 488231

Project number Online data

488236 Parental animals:

Clinical signs

Functional observations

Body weights and food consumption pre-mating

Body weights mating period

Clinical laboratory investigations

Macroscopic findings and organ weights

488231 All other data

5.4. Alms of study

The purpose of this study was to evaluate the potential toxic effects of the test substance when

administered to rats for a minimum of 28 days and to evaluate the potential of the test

k substance to affect male and female reproductive performance such as gonadal function,

mating behaviour, conception, parturition and early postnatal development.

A No Observed Adverse Effect Level (NOAEL) was evaluated.

This study should provide part of a rational basis for toxicological risk assessment in man.

Page 8
C.l. PIGMENT BLUE 61 NOTOX Project 488231

5.5. Guidelines

The protocol was reviewed and agreed by the Laboratory Animal Welfare Officer and the Ethical

Committee of NOTOX (DEC NOTOX 07-13) as required by the Dutch Act on Animal

Experimentation (February 1997).

The study procedures described in this report were based on the following guideline:

1) Organisation of Economic Co-operation and Development Guidelines (OECD) for testing of

Chemicals Guideline 422, Combined Repeated Dose Toxicity Study with the

Reproduction/Developmental Toxicity Screening Test, March 1996.

5.6. Storage and retention of records and materials

Records and materials pertaining to the study, including protocol, raw data, specimens (except

specimens requiring refrigeration or freezing) and the final report, will be retained in the NOTOX

archives for a period of at least 2 years after finalization of the report. After this period, the

sponsor will be contacted to determine whether raw data and specimens should be returned to

them, retained or destroyed on their behalf.

Those specimens requiring refrigeration or freezing will be retained by NOTOX for as long as

the quality of the specimens permits evaluation but no longer than three months after finalization

of the report.

NOTOX will retain a test substance sample until the expiry date, but no longer than 10 years

after finalization of the report. After this period the sample will be destroyed.

Storage and retention of records and materials by the test sites for the delegated phases are

described in the applicable appendices.

Page 9
ClI. PIGMENT BLUE 61 NOTOX Project 488231

6. MATERIALS AND METHODS

6.1. Test substance

6.1.1. Test substance information (170676/A)

Identification C.l. Pigment Blue 61

Structure

N0

S0 3

H

Molecular formula C37H29N303S

Molecular weight 596

GAS Number 1324-76-1

Description Dark blue powder with granules

Batch 2084701

Composition 96.830%

Test substance storage At room temperature in the dark

Stability under storage conditions Stable

Expiry date 02 January 2018

6.1.2. Study specific test substance information

pH 4.3 at concentration of >1 %

Solubility in vehicle:

0Propylene glycol Yes, <1%

6.1.3. Test substance formulation

Vehicle Propylene glycol, specific gravity 1.036 (Merck, Darmstadt,

Germany).

Rationale for vehicle Based on trial formulations performed at NOTOX

Stability of test substance

in vehicle Formulations at 10 and 200 mg/mI were stable when stored at

room temperature for at least 6 hours (determined during this

project; see Appendix 3).

Method of formulation Formulations (wlw) were prepared daily within 6 hours prior to

dosing and were homogenised to a visually acceptable level.

Adjustment was made for specific gravity of the vehicle (1.036).

Storage conditions At ambient temperature.

Page 10
C.I. PIGMENT BLUE 61 NOTOX Project 488231

6.1.4. Chemical analysis of dose preparations

Analyses were performed on a single occasion during the treatment phase (30 June 2008)

according to a validated method (NOTOX Project 488235) as specified below:

Group Analysis (type of sample)

1 acc (M)

2 acc + hom + stabt.0 (TMB), stabt-6, RT (S)

3 acc (M)

4 acc + homn + stabt 0 (TMB), stabt-6. RT (S)

Duplicate samples were analysed

acc=accuracy, hom=homogeneity, stab=stability (hours), T=top, MWmiddle, B=bottom position of container

S=stability sample taken at middle position of container

RT=room temperature

The accuracy of preparation was considered acceptable if the mean measured concentrations

were 90-1 10% of the target concentration for solutions, or 85-1 15% for suspensions.

Homogeneity was demonstrated if the coefficient of variation was :5 10%. Formulations were

considered stable if the relative difference before and after storage was maximally 10%.

6.2. Test system

Test System Rat: Cr1 WI(Han) (outbred, SPE-Quality). Untreated animals and

virgin females were used at initiation of the study.

Rationale This species and strain of rat has been recognized as appropriate

for general and reproductive toxicity studies. NOTOX BV has

general and reproductive historical data in this species from the

same strain and source. This animal model has been proven to be

susceptible to the effects of reproductive toxicants.

Source Charles River Deutschland, Sulzfeld, Germany.

Age at start Fo-treatment Approximately 10 weeks.

Number of F0-animals 40 females and 40 males.

Acclimatisation FO At least 5 days prior to start of treatment.

Health check F0 A health inspection was performed prior to commencement of

treatment to ensure that the animals were in a good state of health.

Randomisation F0 Prior to commencement of treatment, by computer-generated random algorithm according to body weight, with all animals within

± 20% of the sex mean.

Identification F0 Earmark and tattoo.

Page 11
ClI. PIGMENT BLUE 61 NOTOX Project 488231

Breeding procedures Following a minimum of 14 days of exposure for the males and

females, one female was cohabitated with one male of the same

treatment group, avoiding sibling mating (Charles River supplied

non-litter mates). Detection of mating was confirmed by evidence

of sperm in the vaginal lavage or by the appearance of an

intravaginal copulatory plug. This day was designated Day 0 post- coitum. Once mating had occurred, the males and females were

separated. A maximum of 14 days was allowed for mating.

Parturition F0 The females were allowed to litter normally. Day 1 of lactation was

defined as the day when a litter was found completed (i.e.

membranes, placentas cleaned up, nest build up and/or feeding of

pups started). Females that were littering were left undisturbed.

Lactation F0 Deficiencies in maternal care, such as inadequate construction or

cleaning of the nest, pups left scattered and cold, physical abuse of

pups or apparently inadequate lactation or feeding, were recorded.

Number of pups Approximately 480 pups (40 litters x 12 pups).

Identification offspring On Day 1 of lactation, all pups were individually identified by

means of intracutaneous injection of Indian ink.

6.3. Allocation

Group Dose level Number of animals Animals numbers

mg/kg b.w./day F0 males IF 0 females males females

1 0 10 10 01-10 41-50

2 I 50 I 10 I 10 11-20 51-60

3 250 10 10 21-30 61-70

4 1000 1 10 10 1 31-40 71-80

*Dose levels following were based on the dose range finding study (NOTOX Project 488232;

see Appendix 5).

6.4. Animal husbandry

Animals were housed in room 16.

Conditions

Animals were housed in a controlled environment, in which optimal conditions were considered

to be approximately 15 air changes per hour, a temperature of 21 ± 3

0C (actual range: 19.8 -

22.40C), a relative humidity of 30 - 70% (actual range: 41 - 87%) and 12 hours artificial light and

12 hours darkness per day. Temporary fluctuations from the light/dark cycle (with a maximum of

1 hour) occurred due to performance of functional observations in the room. Cleaning

procedures in the room might have caused the temporary fluctuations above the optimal level of

70% for relative humidity. Based on laboratory historical data these conditions were considered

not to have affected the study integrity.

Page 12
C.I. PIGMENT BLUE 61 NOTOX Project 488231

Accommodation

Pre-mating Animals were housed in groups of 5 animals/sex/cage in Macrolon cages (MIV

type, height 18 cm).

Mating Females were caged together with males on a one-to-one-basis in Macrolon

cages (Mill type, height 18 cm).

Post-mating Males were housed in their home cage (Macrolon cages, MIV type, height 18

cm) with a maximum of 5 animals/sex/cage. Females were individually housed

in Macrolon cages (Mill type, height 18 cm).

Lactation Offspring was kept with the dam until termination.

General Sterilised sawdust as bedding material (Litalabo, S.P.P.S., Argenteuil, France)

and paper as cage-enrichment (Enviro-dri, Win. Lillico & Son (Wonham Mill

Ltd), Surrey, United Kingdom) were supplied. Certificates of analysis were

examined and then retained in the NOTOX archives. During overnight activity

monitoring, animals were housed individually in Macrolon cages (MiIl type;

height 15 cm) with sterilised sawdust as bedding material. No cage-enrichment

was provided during ovemnight activity monitoring.

Diet

Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiaten GmbH, Soest,

Germany). Results of analyses for nutrients and contaminants of each batch were examined

and archived.

Water

Free access to tap-water. Certificates of analysis (performed quarterly) were examined and

archived.

Analysis of bedding, paper, diet and water did not reveal any findings that were considered to

have affected study integrity.

6.5. Treatment

6.5.1. Parental animals

Method Oral gavage, using a plastic feeding tube.

Formulations were placed on a magnetic stirrer during dosing.

Frequency Once daily for 7 days per week, approximately the same time each

day with a maximum of 6 hours difference between the earliest and

latest dose. Animals were dosed up to the day prior to scheduled

necropsy.

Exposure period Males were exposed for 30 days, i.e. 2 weeks prior to mating, during mating, and up to termination. Females were exposed for

41-45 days, i.e. during 2 weeks prior to mating, during mating,

during post-coitum, and during at least 4 days of lactation.

Female 78 was not dosed for one occasion as this female was

littering at the time of dosing.

Dose volume 5 mL/kg body weight. Actual dose volumes were calculated

according to the latest body weight.

6.5.2. Offspring

Offspring was not treated.

Page 13
0.1. PIGMENT BLUE 61 NOTOX Project 488231

6.6. Observations

The following animals were selected (first 5 mated males and first 5 females with live offspring)

for functional observations, clinical laboratory investigations, macroscopic examination and

determination of organ weights.

GroupAnimals numbers

SMales (n=5) Females (n=5)

1 3 7 10 42 1;43 4 48 50

2 11 12 114 1 15~ 16 52 155 56157-1-58

3 1 2 31 41 5 1 63 64 65 67

4 132 34 135 136 37 73 174 75 76 77*

6.6.1. Parental animals

Mortality / Viability At least twice daily.

Clinical signs At least once daily, detailed clinical observations were made in all

animals. Once prior to start of treatment and at weekly intervals

this was also performed outside the home cage in a standard

arena. Arena observations were not performed when the animals

were mating, or housed individually. The time of onset, degree and

duration was recorded. All symptoms were recorded and graded

according to fixed scales:

Maximum grade 1: grade 0 =absent, grade 1 = present

Maximum grade 3 or 4: grade 1 = slight, grade 2 = moderate,

grade 3 = severe, grade 4 = very severe

Cage debris of pregnant females were examined to detect abortion

or premature birth, if applicable. Signs of difficult or prolonged

parturition were recorded, if applicable.

Functional Observations The following tests were performed in the first 5 mated males and

the first 5 females with live offspring, from each group:

- hearing ability, pupillary reflex, static righting reflex and grip

strength (Score 0 = normal/present, score 1

abnormal/absent).

- motor activity test (recording period: 12 hours during overnight

for individual animals, using a computerised monitoring

system, Pearson Technical Services, Debenham, Stowmarket,

England).

During the motor activity test, males were caged individually and

females were caged with their offspring.

The assigned males were tested during week 4 of treatment and

the assigned females were tested during lactation (all before blood

sampling).

In order to avoid hypothermia of pups, dams were removed from

the pups for not more than 30-40 minutes.

Body weights Males and females were weighed on the first day of exposure and

weekly thereafter. Mated females were weighed on Days 0, 4, 7,

11, 14, 17 and 20 of gestation, and during lactation on Days 1 and

4.

Page 14
C.I. PIGMENT BLUE 61 NOTOX Project 488231

Food consumption Weekly, for males and females. Food consumption was not

recorded during the breeding period. Food consumption of mated

females was measured on Days 0, 4, 7, 11, 14, 17 and 20 postcoitum and after delivery on Days 1 and 4 post-partum.

Water consumption Subjective appraisal was maintained during the study, but no

quantitative investigation introduced as no effect was suspected.

Reproduction processes Male number paired with, mating date, confirmation of pregnancy,

and delivery day was recorded.

6.6.2. Offspring

Each litter was examined to determine the following, if practically possible:

Mortality / Viability The numbers of live and dead pups at the First Litter Check (

check at Day 1 of lactation) and daily thereafter were determined. If

possible, defects or cause of death were evaluated.

Clinical signs At least once daily, detailed clinical observations were made in all

animals.

Body weights Live pups were weighed during lactation on Days 1 and 4.

Sex Was determined for all pups on Days 1 and 4 of lactation (by

assessment of the ano-genital distance).

Page 15
C.I. PIGMENT BLUE 61 NOTOX Project 488231

6.7. Clinical laboratory investigations

Blood samples were collected from the first 5 mated males and the first 5 females with live

offspring from each group under iso-flurane (Abbott Laboratories Ltd., Zwolle, The Netherlands)

anaesthesia immediately prior to scheduled post mortem examination, between 7.00 and 10.30

am. The animals were fasted overnight (with a maximum of 20 hours) before blood sampling,

but water was provided. Blood samples were drawn from the retro-orbital sinus and collected

into tubes (Greiner Bio-One, Bad Hailer, Austria) prepared with EDTA for haematological

parameters (0.5 ml), with citrate for clotting tests (0.9 ml) and Li-heparin treated tubes for clinical

biochemistry parameters (0.5 ml). The following parameters were determined:

Parameter Abbreviation Unit

Haematology a

White blood cells WBC 109

/LDifferential leucocyte count %WBC

neutroph is, lymphocytes, monocytes,

eosinophils, basophils

Red blood cells 1 0' 2

/L

Reticulocytes %RBC

Red blood cell distribution width RDW%

Haemoglobin mmol/L

Haematocrit L/L

Mean corpuscular volume MCV flMean corpuscular haemnoglobin MCH find

Mean corpuscular haemoglobin MCHC mmol/L

concentration

Platelets 109

'/L

Clotting Poeta

Prothrombin time PT s

Activated Partial thromboplastin time APTT s

Clinical Biochemistry c

Alanine aminotransferase ALAT U/L

Aspartate ami notra nsfe rase ASAT U/L

Alkaline phosphatase ALP UIL

Total Protein gIL

Albumin g/L

Total Bilirubin pinolIL

Urea minol/L

Creatinine pinol/L

Glucose minolL

Cholesterol minoilL

Sodium inmolIL

Potassium minol/L

Chloride inmol/L

Calcium mmol/L

Inorganic Phosphate lnorg. Phos inmol/L

'Instrumentation: AOVIA 120 (Bayer Diagnostics)

'Instrumentation: STA Compact (Roche Diagnostics).

'Instrumentation: Olympus AU400 (Goffin Meyvis)

Page 16
0.1. PIGMENT BLUE 61 NOTOX Project 488231

6.8. Pathology Parental Animals

6.8.1. Termination

All animals were fasted overnight (with a maximum of 20 hours) prior to planned necropsy, but

water was provided.

Females Which Deliver

On lactation Day 5, the Fo-females were anaesthetised using iso-flurane (Abbott Laboratories

Ltd., Zwolle, The Netherlands) and subsequently exsanguinated. Organ weights were collected

and tissues were preserved for possible future histopathological examination as described in

the following paragraphs. The number of former implantation sites were counted and recorded.

Corpora lutea were also counted and recorded. Gross lesions were saved for possible future

histopathological examination.

Females Which Failed to Deliver

On post-mating Day 25-27 (females with evidence of mating), the F0- females which had not

delivered were anaesthetised using iso-flurane (Abbott Laboratories Ltd., Zwolle, The

Netherlands) and subsequently exsanguinated. No organ weights were collected. Tissues were

preserved for possible future histopathological examination as described in the following

paragraphs, with the following exceptions: Nongravid uteri were stained using the Salewski

technique (Salewski, 1964) in order to determine any former implantation sites (Salewski

staining prepared at NOTOX using Ammoniumsulflde-solution 20% (Merck, Darmstadt,

Germany) and Milli-Ro water (Millipore Corporation, Bedford, USA)). If evidence of

macroscopic implantations was present, the number of implantation sites and corpora lutea

was recorded. Gross lesions were saved for possible future histopathological examination.

Termination Procedures for Fo0-Males

Following completion of the mating period (29 days of dose administration), the F0-males were

anaesthetised using iso-flurane (Abbott Laboratories Ltd., Zwolle, The Netherlands) and

subsequently exsanguinated. Organ weights were collected and tissues were preserved for

possible future histopathological examination as described in the following paragraphs.

Page 17
C.I. PIGMENT BLUE 61 NOTOX Project 488231

6.8.2. Macroscopic examination

After sacrifice all parental animals were subjected to macroscopic examination of the cranial,

thoracic and abdominal tissues and organs, with special attention being paid to the reproductive

organs. Descriptions of all macroscopic abnormalities was recorded. Samples of the following

tissues and organs were collected and fixed in neutral phosphate buffered 4% formaldehyde

solution (Klinipath, Duiven, The Netherlands):

From the first 5 males per group and the first 5 females with live offspring per group:

Identification marks: not processed Ovaries

Adrenal glands Pancreas

Aorta Peyer's patches (jejunum, ileum) if detectable

Brain (cerebellum, mid-brain, cortex) Pituitary gland

Caecum Preputial gland

Cervix Prostate gland

Clitoral gland Rectum

Colon (Salivary glands - mandibular, sublingual)

Coagulation gland Sciatic nerve

Duodenum Seminal vesicles

Epididymides* (Skeletal muscle)

(Eyes* with optic nerve (if detectable) and (Skin)

Harderian gland) Spinal cord -cervical, midthoracic, lumbar

(Female mammary gland area) Spleen

(Femur including joint) Sternum with bone marrow

Heart Stomach

Ileum Testes*

Jejunum Thymus

Kidneys Thyroid including parathyroid (if detectable)

(Larynx) (Tongue)

(Lacrimal gland, exorbital) Trachea

Liver Urinary bladder

Lung, infused with formalin Uterus

Lymph nodes - mandibular, mesenteric Vagina

(Nasopharynx) All gross lesions

Oesophagus

From all remaining animals:

Cervix Prostate gland

Clitoral gland Seminal vesicles

Coagulation gland Testes*

Epididymides* Uterus

Ovaries Vagina

Preputial gland All gross lesions

Identification marks: not processed

*Fixed in modified Davidson's solution (prepared at NOTOX using Formaldehyde 37.40%, Ethanol, Acetic acid (glecial)(al

Merck, Darmstadt, Germany) and Milli-Ro water (Millipore Corporation, Bedford, USA)) and transferred to formalln after

fixation for at least 24 hours.

Tissues/organs mentioned in parentheses were not examined by the pathologist, since no signs of toxicity were noted

at macroscopic examination.

Page 18
C.I. PIGMENT BLUE 61 NOTOX Project 488231

6.8.3. Organ weights

The following organ weights (and terminal body weight) was recorded:

From the first 5 males per group and the first 5 females with live offspring per group:

Adrenal glands Liver

Brain Spleen

Epididymides Testes

Heart Thymus

Kidneys

From all remaining males:

Epididymides

Testes

6.8.4. Histatechnology

All organ and tissue samples. as defined under Histopathology (following), were processed,

embedded and cut at a thickness of 2-4 micrometers and stained with haemnatoxylin and eosin

(Klinipath, Duiven, The Netherlands).

Of the selected 5 males/group of the control and high dose group, additional slides of the testes

were prepared to examine staging of spermatogenesis. The testes was processed, sectioned at

3-4 microns, and stained with PAS/haematoxylin (Klinipath, Duiven, The Netherlands).

6.8.5. Histopathology

The delegated phase was performed by the Principal Investigator for Histopathology. See

Appendix 4.

The following slides were examined by a pathologist:

-The preserved organs and tissues of the selected animals of Groups 1 and 4.

-The additional slides of the testes of the selected 5 males/group of Groups 1 and 4 to

examine staging of spermatogenesis

-All gross lesions of all animals (all dose groups)

-The reproductive organs* of all animals that failed to mate, conceive, sire or deliver healthy

offspring. These animals included:

- Group 1 Male 1 and Female 41 (non-pregnant).

- Group 2 Male 14 and Female 54 (non-pregnant).

- Group 3 Male 26 and Female 66 (non-pregnant). - Group 4 Male 32 and Female 72 (implantation sites only).

Histological examination was extended to the mesenteric lymph nodes of the selected males

and females of Group 2 based on possible treatment-related changes noted for mid and high

dose animals.

All abnormalities were described and included in the report. An attempt was made to correlate

gross observations with microscopic findings.

* Reproductive organs included cervix, clitoral gland, coagulation gland, epididymides, ovaries,

preputial gland, prostate gland, seminal vesicles, testis, uterus, and vagina.

Page 19
C.I. PIGMENT BLUE 61 NOTOX Project 488231

6.9. Pathology Offspring

6.9.1. Termination

Pups were killed by decapitation on Day 5 of lactation or shortly thereafter.

6.9.2. Macroscopic examination

All offspring was sexed and externally examined if practically possible. The stomach was

examined for the presence of milk.

Descriptions of all macroscopic abnormalities were recorded.

If possible, defects or cause of death was evaluated. Any abnormal pup, organ or tissue was

preserved in 10% buffered formalin, for possible further examination.

6.10. Calculations

For each dose group reproduction parameters were expressed in two ways:

- As a mean (with standard deviation) of the number observed for each litter

- Relative to a second parameter and calculated on a total group basis

For each group the following calculations were performed:

Percentage mating Number of females mated Ix 100

Number of females paired

Fertility index Number of pregnant females x 100

Number of females paired

Conception rate Number of oreanant females x 100

Number of females mated

Gestation index Number of females bearing~ live Pups x 100

Number of pregnant femal es

Duration of gestation Number of days between confirmation of mating

and the beginning of parturition

Percentage live males Number of live male pups at First Litter Check x 100

at First Litter Check Number of live pups at First Litter Check

Percentage live females Number of live female pups at First Litter Check x 100

at First Litter Check Number of live pups at First Litter Check

Percentage of postnatal Number of dead ou Ps on Day 4 ipost partum x 100

loss Days 0-4 postpartum Number of live pups at First Litter Check

Viability index Number of live pups on Day 4 Postpcartum x 100

Number of pups born alive

Page 20
C.I. PIGMENT BLUE 61 NOTOX Project 488231

6.11. Electronic data capture

Observations/measurements in the study were recorded electronically using the following

programme(s):

- REES version 1.5 (REES Scientific, Trenton, NJ, USA): Environmental monitoring.

- TOXOATA version 8.0 (NOTOX B.V., 's-Hertogenbosch, The Netherlands): Mortality / Clinical

signs / Body weights / Food consumption / Reproduction processes / Observations offspringI

Organ weights. -Advia 120 Version V.3.1.8.0.MS (Siemens Medical Solution Diagnostics, Mijdrecht, The

Netherlands): Haemnatology. - Sta Compact version 1.06.06 (Stago Instruments, Gennevilliers, France): Clotting parameters

- Olympus AU 400 version 8.2A (Goffin-Meyvis, Etten-Leur, The Netherlands): Clinical

laboratory investigations.

- MAMS version 6.5 (Pearson Technical Services, Suffolk, Great Britain): Motor activity

measurement.

-Empower version 5.00 (Waters, Milford, MA, USA): System control, data acquisition and data

processing for analytical chemistry.

6.12. Interpretation

The following statistical methods will be used to analyse the data: - If the variables could be assumed to follow a normal distribution, the Dunnett-test (Dunnett,

1955) (many-to-one t-test) based on a pooled variance estimate was applied for the

comparison of the treated groups and the control groups for each sex. - The Steel-test (Miller, 1981) (many-to-one rank test) was applied if the data could not be

assumed to follow a normal distribution.

- The Fisher Exact-test (Fisher, 1950) was applied to frequency data.

All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of

significance.

No statistical analysis was performed on histopathology findings.

6.13. List of deviations

6.13.1. List of protocol deviations

1 No organ weight was determined of the testes and epididymides of animal 30.

Evaluation: Sufficient data were available for evaluation.

2 A few tissues were not available for histopathology. Reasons for this included that

these tissues were not discernable at necropsy or trimming, or were erroneously not

collected at necropsy. Tissues are listed in raw data and pathology report.

Evaluation: Sufficient data was available for evaluation.

3 Tissues for histopathology were preserved and organ weights were determined for

another subset of animals (out of 20 selected males, 5 were different: Group 1 males 4

and 5 instead of 7 and 10, Group 2 male 13 instead of 16, and Group 4 males 31 and

33 instead of 36 and 37) than tested for functional observations and clinical laboratory

investigations.

Evaluation: No toxicity was noted up to the high dose level for all parameters; therefore

it was not essential to have results of these parameters on the same subset of

animals.

The study integrity was not adversely affected by the deviations.

6.13.2. List of standard operating procedure deviations

There were no deviations from standard operating procedures.

Page 21
C.I. PIGMENT BLUE 61 NOTOX Project 488231

7. RESULTS

7.1. Analysis of Dose Preparations (see also Appendix 3)

The concentrations analysed in the formulations of Group 2, Group 3 and Group 4 were in

agreement with target concentrations (i.e. mean accuracies between 85% and 115%). No test

substance was detected in the Group 1 formulations. The formulations of Group 2 and Group 4

were homogeneous (i.e. coefficient of variation :5 10%). Formulations of Group 2 and Group 4

were stable when stored at room temperature for at least 6 hours.

7.2. Observations

7.2.1. Mortality

No mortality occurred during the study period.

7.2.2. Clinical Signs

There were no clinical signs of toxicity noted during the observation period.

Blue discolouration of the faeces was noted in all animals of both sexes at 50, 250, and 1000

mg/kg. Blue staining of several body parts (tail, back, mouth, snout, abdomen, forelegs, flank)

was also observed at these dose levels but at a lower incidence. These findings were

considered due to the staining properties of the test substance and not to be toxicologically

relevant.

Slight salivation was noted for one low dose male (1 day), five mid dose males (1 -3 days), six

high dose males (2-18 days) and two high dose females (1 day). Salivation was considered to

be a physiological response to the taste of the formulation rather than a sign of systemic toxicity,

considering the nature, minor severity of the effect and its time of occurrence (i.e. after dosing).

Therefore, this was considered not toxicologically relevant.

Incidental findings that were noted included alopecia and scabbing on several body parts, rales,

piloerection and a broken tail apex. These findings are commonly noted in rats of this age and

strain which are housed and treated under the conditions in this study. At the incidence

observed, these were considered signs of no toxicological significance.

7.2.3. Body Weights

Body weights and body weight gain of treated animals remained in the same range as controls

over the study period.

7.2.4. Food Consumption

Food consumption (absolute and relative) of treated animals in all groups remained in the same

range as controls over the study period.

Two female rats (numbers 55 and 60) of the low dose group showed very low food consumption

on Days 11-14 post-coitum. As body weight gains for these dams were normal, these low

values were probably caused by a weighing error rather than being true values and were

therefore considered not toxicologically relevant.

Page 22
C.I. PIGMENT BLUE 61 NOTOX Project 488231

7.2.5. Functional observations

No changes were observed in hearing ability, pupillary reflex, static righting reflex and grip

strength in the animals treated with 0.1. Pigment Blue 61, when compared to control animals.

The variation in motor activity did not indicate a relation with treatment.

7.3. Clinical Laboratory Investigations

7.3.1. Haematology

Haemnatological parameters of treated rats were not affected by treatment up to 1000 mg/kg.

Females treated with 1000 mg/kg showed statistically significantly lower prothrombin time than

controls. This finding is considered to be of no toxicological significance since the value was

within normal limits and no corroborative findings were noted. In addition, the statistically

significantly increased haemnatocrit value for low dose females was not considered treatment

related as no dose response relationship was noted.

7.3.2. Clinical Biochemistry

No toxicologically relevant changes occurred in clinical biochemistry parameters of treated rats.

Males treated with 1000 mg/kg had statistically significantly higher cholesterol measures than

controls. This was considered to be of no toxicological significance as the value was within

normal limits and no corroborative findings were noted.

7.4. Pathology

7.4.1. Macroscopic Examination

No toxicologically relevant macroscopic findings were noted up to 1000 mg/kg body weight/day.

Greenish or bluish discolouration of the mesenteric lymph nodes was noted for three low dose

animals, seventeen mid dose animals and all high dose animals. Seven high dose females had

bluish contents in the gastro intestinal tract. Bluish discolouration of the tail was observed for

two males. These findings were considered due to the staining properties of the test substance

and not to be toxicologically relevant.

Incidental findings included bent tail, hard yellowish nodule on the tail of the epididymidis,

alopecia at several body parts, reduced seminal vesicle size, dark red or reddish discolouration

of the mandibular lymph nodes, reddish discolouration of the pancreas, and red-brown

discolouration of the adrenal glands. These findings are occasionally seen among rats used in

these types of studies and in the absence of correlated microscopic findings they were not

considered changes of toxicological significance.

Page 23
C.l. PIGMENT BLUE 61 NOTOX Project 488231

7.4.2. Organ Weights

No toxicologically relevant organ weight changes were noted for animals treated up to 1000

mg/kg body weight/day.

The statistically significantly increased relative spleen weight noted for high dose males was

considered not to be a sign of toxicity in the absence of corroborative (microscopic) findings.

7.4.3. Microscopic Examination (see also Appendix 4)

No treatment related histopathological changes were observed in the tissues of any animal

receiving 0.1. Pigment Blue 61.

No abnormalities were noted on evaluation of PAS stained sections of testes of F0 males. All

seminiferous tubules were normal with respect to their stage in the spermatogenic cycle and the

integrity of the various cell types present within the different stages.

No pathological findings were present in the reproductive organs of male and female animals

that failed to produce offspring.

Small amounts of blue material were observed in the lumen of the duodenum, ileum, jejunum,

colon or rectum in a few group 4 animals. Blue material was also noted in the superficial skin

(keratin layer) of the tail in one group 4 male. These findings are considered to represent the

presence of the test substance and in some cases correlated with the macroscopic

observations.

No microscopic findings were observed in the mesenteric lymph nodes to account for the

discolouration of these tissues recorded at necropsy. It is considered that the discolouration is

due to the presence of the test substance and that the pigment was lost from the tissues in the

processing or staining procedures.

All other microscopic findings were considered to be incidental and unrelated to treatment with

the test substance.

7.5. Reproduction

Reproduction parameters are described in tables I and 11:

Table 1. Reproduction Data.

Number of females Group 1 Group 2 Group 3 Group 4

Control 50 mg/kg 250 mg/kg 1000 mg/kg

Paired 10 10 10 10

Mated 10 10 10 10

Non-pregnant 1 1 1 0

Pregnant 9 9 9 10

Implantations sites only 0 0 0 1 (F72)

Number of females with 9 9 9 9

living pups at first litter check

Page 24
CAI. PIGMENT BLUE 61 NOTOX Project 488231

Table i1. Fertility FO-generation.

Group 1 Group 2 Group 3 Group 4

Control 50 mg/kg 250 mg/kg 1000 mg/kg

Percentage mating 100 100 100 100

(Females mated / Females paired) *10

Fertility index 90 90 90 100

(Females achieving a pregnancy / Females paired) *100

Conception rate 90 90 90 100

(Females achieving a pregnancy / Females mated) *100

Gestation index 100 100 100 90

(Number of females with living pups at first litter check/

Number of females pregnant) * 100

Reproduction parameters were unaffected by treatment up to 1000 mg/kg body weight/day.

No treatment related findings were observed for mating performance, fertility parameters,

duration of gestation, number of dead and living pups at first litter check, number of implantation

sites and number of corpora lutea.

7.6. Breeding Data

Breeding parameters were unaffected by treatment up to 1000 mg/kg body weight/day.

Postnatal loss and viability index were similar for the control and treated groups.

7.7. Pup Development

Development of pups was unaffected by treatment up to 1000 mg/kg body weight/day.

(Mean) body weights were similar for the control and treated groups.

Incidental clinical symptoms consisted of small size of the pup, red spot on the nose or head,

and pale appearance. At macroscopic examination, autolysis and absence of milk in the

stomach was noted for a few dead pups. No relationship with treatment was established for

these observations and they were considered to be within the normal biological variation for rats

of this age and strain.

Page 25
C.I. PIGMENT BLUE 61 NOTOX Project 488231

8. DISCUSSION AND CONCLUSION

C.I. PIGMENT BLUE 61 was administered by daily oral gavage to male and female Wistar Han

rats at dose levels of 50, 250 and 1000 mg/kg/day. The males were exposed for 2 weeks prior

to mating, during mating, and up to termination (for 30 days). The females were exposed for 2

weeks prior to mating, during mating, during post-coitum, and at least 4 days of lactation (for 41-

45 days).

Formulation analysis showed that the formulations were prepared accurately, were

homogeneous and were stable for at least 6 hours at room temperature.

No treatment-related toxicologically significant changes were noted in any of the parameters

investigated in this study (i.e. mortality, clinical appearance, functional observations, body

weight, food consumption, clinical laboratory investigations, macroscopic examination, organ

weights, microscopic examination, reproduction, breeding and pup development).

From the results presented in this report, the parental, reproduction, breeding and

developmental No Observed Adverse Effect Level (NOAEL) for ClI. PIGMENT BLUE 61 of at

least 1000 mg/kg/day was established.

Page 26
0.1. PIGMENT BLUE 61 NOTOX Project 488231

9. REFERENCES

Dunnett C.W., A Multiple Comparison Procedure for Comparing Several Treatments with a

Control, J. Amer. Stat. Assoc. 50, 1096-1121 (1955).

Fisher R.A., Statistical Methods for Research Workers, Oliver and Boyd, Edinburgh (1950).

Miller R.G., Simultaneous Statistical Inference, Springer Verlag, New York (1981).

Salewski, E. Farbemethode zum makroskopischen Nachweis von lmplantationsstellen am

Uterus der Ratte. [Staining method for a macroscopic test for implantation sites in the uterus of

the rat]. Naunyn - Schmiedebergs Archly for Experimentelle Pathologie und Pharmakologie,

247, 367 (1964).

Page 27
C.I. PIGMENT BLUE 61 NTXPoet483

APPENDIX 1IOO rjet483

APPENDIX 1 SUMMARY TABLES

-Page 1 -
C.!. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

CLINICAL SIGNS SUMMARY,WEEKLY

MALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: 1. 1.. 4.

(LOCATION)

GROUP II(CONTROL)

Various

Broken (1) G: . 1

(Tail apex) %,1

GROUP 2 (50 MG/KG)

Skin /fur

Scabs (3) G: . 1

(Neck) %: . 1

Blue staining (1) G:11

Blue discolouration (1) G: 11 ill

(Faeces) %: PAA

Secretion / excretion

Salivation (3) 0: ... 1

%:. 1

GROUP 3 (250 MG/KG)

Skin /fur

Alopecla (3) 0:...

(Forelegs) %/: .... 1

Scabs (3) G: . 1

(Neck) %: .. 1.

Blue staining (1) G: .1.

(Back) %: .. 1

Blue staining (1) G: ....

(Tail) %: ..

Blue discolouration (1) G: I I11

(Faeces) %: PAA

Secretion / excretion

Salivation (3) G: .. 1

%: .24

GROUP 4 (1000 MG/KG)

Skin /fur

Alopecia (3) G: . .11

(Foreleg right) %: . .11

Alopecia (3) G: .. 121

(Forelegs) %: .. 11

Scabs (3) G: ... 1

(Foreleg right) %: ... 1

Blue staining (1) G: 1 . ..

(Back) %: 1'. ..

Blue staining (1) G: .. 11

Blue discolouration (1) G: 11 ill

(Faeces) %: PA

Secretion / excretion

Salivation (3) G: I 1

%: .6 653

G: Median value of the highest Individual weekly grades

%: Percent of affected animals (OlIess than 5%, 1 =between 5% and 15%,.,A=more than 95%)

Observation performed, sign not present

- Page 2 - fr24Oct08 15h45
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

CLINICAL SIGNS SUMMARY,WEEKLY

FEMALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: 1. 1-..

(LOCATION)

GROUP I(CONTROL)

Skin /fur

Piloerection (1) G: ... 1

Alopecia (3) G: 11 1..

(Cheek left) %: 11 11 ...

Alopecia (3) G: .1 1..

(Cheek right) %: .1 1..

Alopecia (3) G: 11 11.

(Back) %: 23 32 ...

(Neck) %: 43 4441.

.Alopecia (3) G: . . . 22

(Abdomen) % . ."..22

Alopecia (3) G: ..... 11

(Flank left) % . ... 12

Alopecia (3) G: ..... 11

(Thigh hind left) % . ... 12

Alopecia (3) G: .. .

(Foreleg left) % . .11.

Alopecia (3) G: .. ... 33

(Forelegs) %: 12

Scabs (3) G: . . 1

(Mouth) %. ... 1.

Scabs (3) G: . 1.

(Snout) % ...

GROUP 2 (50 MG/KG) Skin /fur

Piioerection (1) G. .

%.1

Alopecia (3) G: 111..

(Neck) %: 12 1...

Alopecia (3) G: ... 1

(Abdomen) %: ... 1.

Blue staining (1) G: .1

(Tail) % . 1. . .

Blue discolouration (1) G: 11 11111

(Faeces) %: A AAAAA

GROUP 3 (250 MG/KG)

Breathing

Rales (3) G: . .12.

%11

Skin / fur

Piloerectlon (1) G: ... ..

%1.

Alopecia (3) G, 111..

(Cheekrfight) %: 121..

Blue staining (1) G: ... 1

(Mouth) %: ..... 1

Blue staining (1) G: 1....

(Snout) %: 1....

Blue staining (1) G: 111..

(Back) %: 111..

Blue staining (1) G: 1

(Abdomen) %: .... 1

Blue staining (1) G: . .. 1

(Forelegs) %:...1

Blue discolourstion (1) G: 11111

(Faeces) %: AA AAAAM

G- Median value of the highest individual weekiy grades

%: Percent of affected animais (O=iess than 5%, 1 =between 5% and 15%..Amore than 95%)

Observation performed, sign not present

- Page 3 24Oct08 1.5h45
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

CLINICAL SIGNS SUMMARY,WEEKLY

FEMALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: 1.1.4

(LOCATION)

GROUP 4 (1000 MG/KG)

Breathing

Rales (3) G: I.

%: 1.

Skin /fur

Alopecia (3) G: ... 1.

(Back) % . . 1.

Alopecia (3) G: 1.

(Neck) %: 1.

Alopecia (3) G: .*.1

(Abdomen) 1.

Alopecia (3) G: .. .. 333

(Forelegs) % . . .112

Alopecia (3) G: .. .. 22

(Hindlegs) %..12

Scabs (3) G: .1I 

(Mouth) % .. 1.

Blue staining (1) G: .1 1111.

(Tail) % .1 1111.

Blue staining (1) G: ... 1 ...

(Flank ight) % . 1 ...

Blue d~scolouration (1) G: 11 11111

(Faeces) %: AA AAA

Secretion / excretion

Salivation (3) G: 1 1.

%: . 1.

G: Median value of the highest individual weekly grades

%: Percent of effected animals (O=Iess than 5%, I between 5% and 15%..Amore than 95%)

Observation performed, sign not present

-Page 4 - 100- 24Oct08 15h45
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX I

BODY WEIGHTS (GRAM) SUMMARY

MALES

GROUIP1 GROUP 2 GROUP 3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

PRE MATING

DAY 1 MEAN 302 301 303 303

WEEK I ST.DEV 15.1 13.9 8.4 9.2

N 10 10 10 10

DAY 8 MEAN 325 322 328 321

WEEK 2 ST.DEV 18.5 19.6 12.0 10.9

N 10 10 10 10

MATING PERIOD

DAY I MEAN 346 345 352 340

WEEK 1 ST.DEV 21.0 23.1 14.6 10.6

N 10 10 10 10

DAY 8 MEAN 360 349 366 352

WEEK 2 ST.DEV 25.2 18.7 13.2 9.5

N 10 10 10 10

DAY 15 MEAN 378 372 383 369

WEEK 3 ST.DEV 26.9 30.3 16.8 9.3

N 10 10 10 10

FEMALES

GROUIP1 GROUP2 GROUP3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

PRE MATING

DAYl MEAN 193 194 193 192

WEEK 1 ST.DEV 8.1 7.5 6.9 6.2

N 10 10 10 10

DAY8 MEAN 198 203 199 198

WEEK 2 ST.DEV 8.5 9.1 6.4 5.6

N 10 10 10 10

MATING PERIOD

DAYl MEAN 208 212 211 208

WEEK 1 ST.DEV 7.6 6.5 7.1 10.1

N 10 10 10 10

*r Dunnett-test based on pooled variance significant at 5% ()or 1 % (-) level

- Pages5 - JV~ 29Sep08 09h35
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

BODY WEIGHTS (GRAM) SUMMARY

FEMALES

FO-GENERATION

GROUP 1 GROUP2 GROUP3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

POST COITUM

DAY 0 MEAN 209 214 212 210

ST.DEV. 9.7 7.3 4.4 6.9

N 9 9 9 10

DAY 4 MEAN 226 230 230 228

ST.DEV. 11.9 10.2 7.0 5.8

N 9 9 9 10

DAY 7 MEAN 233 238 238 236

ST.DEV. 10.7 13.3 7.8 67

N 9 9 9 10

DAY 11 MEAN 246 251 253 249

ST.DEV 13.1 14.1 8.5 7.0

N 9 9 9 10

DAY 14 MEAN 258 263 265 259

ST.DEV. 18.5 15.8 10.1 9.1

N 9 9 9 10

DAY 17 MEAN 281 285 287 280

ST.DEV. 17.8 18.4 12.7 15.5

N 9 9 9 10

DAY 20 MEAN 314 317 319 312

ST.DEV. 26.3 21.7 17.8 28.9

N 9 9 9 10

LACTATION

DAY I MEAN 241 245 248 238

ST.DEV. 14.5 14.7 8.0 7 1

N 9 9 9 9

DAY 4 MEAN 252 250 256 251

ST.DEV. 15.3 17.6 6.8 10.6

N 9 9 9 9

/D unnet-test based on pooled variance significant at 5% ()or 1% (**) level

Explanations for excluded data are listed in the tables of the individual values

- Page 6 - ON 23Oct05 16h07
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

BODY WEIGHT GAIN (%) SUMMARY

MALES

GROUPi GROUP2 GROUP3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

PRE MATING

DAY1 MEAN 0 0 0 0

WEEK 1 ST.OEV 0.0 0.0 0.0 0.0

N 10 10 10 10

DAY 8 MEAN 8 7 8 6

WEEK 2 ST.DEV 2.5 2.3 1.7 2.0

N 10 10 10 10

MATING PERIOD

DAYl MEAN 15 14 16 12

WEEK 1 ST.DEV 4.3 2.9 24 3.2

N 10 10 10 10

DAYB8 MEAN 19 16 21 16

WEEK 2 ST.OEV 5.9 4.6 2.1 4.1

N 10 10 10 10

DAY 15 MEAN 25 24 26 22

WEEK 3 ST.DEV 6.4 4.9 3.2 3.1

N 10 10 10 10

FEMALES

GROUP 1 GROUP2 GROUP 3 GROUP4

CONTROL 60 MG/KG 250 MG/KG 1000 MG/KG

PRE MATING

DAYl MEAN 0 0 0 0

WEEK 1 ST.DEV 0.0 0.0 0.0 0.0

N 10 10 10 10

DAY 8 MEAN 3 4 4 3

WEEK2 ST.DEV 1.8 1.8 30 1.9

N 10 10 10 10

MATING PERIOD

DAYl MEAN 7 9 10 8

WEEK 1 ST.DEV 22 3.0 5.0 4.4

N 10 10 10 10

*1' Dunnett-test based on pooled variance significant at 5% ()or 1% ()level

- Page 7 - 29Sep08 091136
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

BODY WEIGHT GAIN (%) SUMMARY

FEMALES

FO-GENERZATION

GROUP 1 GROUP2 GROUP 3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

POST COITUM

DAYO0 MEAN 0 0 0 0

ST.DEV. 0.0 0.0 0.0 0.0

N 9 9 9 10

DAY4 MEAN 8 7 8 9

ST.DEV. 2.0 1.9 2-3 1.8

N 9 9 9 10

DAY 7 MEAN 11 11 12 13

ST.DEV. 2.8 3.0 2.6 1.8

N 9 9 9 10

DAY11 MEAN 18 17 19 19

ST.DEV. 3.2 34 2.9 2.8

N 9 9 9 10

DAY14 MEAN 23 23 25 23

ST 0EV. 5.9 4.2 4.2 4.9

N 9 9 9 10

DAY17 MEAN 34 33 35 34

ST.DEV. 5.8 5.1 64 8.2

N 9 9 9 10

DAY 20 MEAN 50 48 50 49

ST.DEV. 10.4 6.5 9.1 14.6

N 9 9 9 10

LACTATION

DAYl MEAN 0 0 0 0

ST.DEV. 0.0 0.0 00 0.0

N 9 9 9 9

DAY 4 MEAN 5 2 3 5

ST.DEV. 3.0 2.5 2.8 2.8

N 9 9 9 9

IDunnett-test based on pooled vaniance significant at 5% ()or 1 % (**) level

Explanations for excluded data are listed in the tables of the individual values

- Page 8 - 23Oct08 16h07
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

FOOD CONSUMPTION (GIANIMAL/DAY) SUMMARY

MALES

GROUP 1 GROUP 2 GROUP 3 GROUP 4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

PRE MATING

DAYS 1-8 MEAN 21 21 22 21

WEEKS 1-2 ST.DEV 2.2 0.0 0.1 0.4

N(CAGE) 2 2 2 2

DAYS 8-15 MEAN 22 22 22 22

WEEKS 2-3 ST.DEV 2.7 0.5 0.2 0.2

N (CAGE) 2 2 2 2

MEAN OF MEANS

OVER PRE MATING MEAN 22 21 22 22

FEMALES

GROUP I GROUP2 GROUP3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

PRE MATING

DAYS 1-83 MEAN 14 14 14 14

WEEKS 1-2 ST.DEV 0.6 0.4 0.5 0.3

N(CAGE) 2 2 2 2

DAYS 8-15 MEAN 15 14 15 15

WEEKS 2-3 ST.DEV 0.5 0.2 0.5 0.4

N (CAGE) 2 2 2 2

MEAN OF MEANS

OVER PRE MATING MEAN 14 14 14 15

/~Dunnett-test based on pooled variance significant at 5% ()or 1% ()level

- Page 9 - '29Sep08 09h35
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

FOOD CONSUMPTION (G/ANIMALIDAY) SUMMARY

FEMALES

FO-GENERATION

GROUP 1 GROUP2 GROUP 3 GROUIP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

DAYS 0-4 MEAN 17 17 17 17

ST DEV. 1.4 2.5 1.9 1.3

N 9 9 9 10

DAYS 4-7 MEAN 18 18 19 19

ST.DEV. 1.9 3.0 2.1 0.8

N 9 9 9 10

DAYS 7-1l MEAN 19 18 20 19

ST.DEV. 14 1.7 2.0 1.7

N 9 9 9 10

DAYS 11-14 MEAN 20 17 23 20

ST.DEV. 3.2 7.7 5.1 1.8

N 9 9 9 10

DAYS 14-17 MEAN 20 21 22 20

ST.DEV. 2.9 3.6 1 8 2.6

N 9 9 9 10

DAYS 17-20 MEAN 22 22 22 21

ST.DEV. 3.3 3.3 1 7 2.8

N 9 9 9 10

MEAN OF MEANS 20 19 20 19

LACTATION

DAYS 1-4 MEAN 31 27 26 31

ST DEV. 7.1 8.5 5.6 10.7

N 9 9 9 8

/Dunnett-test based on pooled variance significant at 5% ()or 1 % ()level

Explanations for excluded data are listed in the tables of the individual values

-Page 10 -h 23Oct08 16h07
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

RELATIVE FOOD CONSUMPTION (GIKG BODY WEIGHTIDAY) SUMMARY

MALES

GROUP 1 GROUP 2 GROUP3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

PRE MATING

DAYS 1-8 MEAN 66 65 66 66

WEEKS 1-2 ST.DEV 3.5 1.7 1.3 0.9

N(CAGE) 2 2 2 2

DAYS 8-15 MEAN 69 68 68 68

WEEKS 2-3 ST.DEV 5.0 0.2 1.5 0.3

N(CAGE) 2 2 2 2

MEAN OF MEANS

OVER PRE MATING MEAN 67 66 67 67

FEMALES

GROUP 1 GROUP 2 GROUP 3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

PRE MATING

DAYS 1-8 MEAN 71 71 71 72

WEEKS 1-2 ST.DEV 4.3 1.9 3.1 0.3

N(CAGE) 2 2 2 2

DAYS 8-15 MEAN 74 71 73 75

WEEKS 2-3 ST.DEV 4.0 1.1 3.3 1.1

N(CAGE) 2 2 2 2

MEAN OF MEANS

OVER PRE MATING MEAN 73 71 72 73

/~Dunnett-test based on pooled variance significant at 5% ()or I% ()level

-Page 11- 29Sep08 09h35
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

RELATIVE FOOD CONSUMPTION (GIKG BODY WEIGHT/DAY) SUMMARY

FEMALES

FO-GENERATION

GROUP 1 GROUP2 GROUP3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

POST COITUM

DAYS 0-4 MEAN 75 74 75 74

ST.DEV. 5.1 8.3 6.4 5.5

N 9 9 9 10

DAYS 4-7 MEAN 79 76 78 79

ST.DEV. 7.7 8.5 7.1 3.2

N 9 9 9 10

DAYS 7-11 MEAN 78 73 77 77

ST.DEV. 4.6 7.6 59 57

N 9 9 9 10

DAYS 11-14 MEAN 79 64 85 78

ST.DEV 9.3 27.5 17.9 6.7

N 9 9 9 10

DAYS 14-17 MEAN 72 74 75 72

ST DEV. 9.6 7.9 5.1 6.7

N 9 9 9 10

DAYS 17-20 MEAN 70 69 69 67

ST.DEV. 7.5 5.8 2.6 5.1

N 9 9 9 10

MEAN OF MEANS 76 72 77 74

LACTATION

DAYS 1-4 MEAN 122 105 102 124

ST.DEV. 24.8 263 21.7 39.1

N 9 9 98

SDunnett-test based on pooled variance significant at 5% ()or 1% ()level

Explanations for excluded data are listed in the tables of the individual values

- Page 12 - 23Oct08 16h07
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

FUNCTIONAL OBSERVATIONS SUMMARY

MALES

GROUP 1 GROUP 2 GROUP3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

AT WEEK 4

HEARING MEDIAN 0 0 0 0

SCORE 0/1 N 5 5 5 5

PUPIL L MEDIAN 0 0 0 0

SCOREO/1 N 5 5 5 5

PUPIL R MEDIAN 0 0 0 0

SCOREO/1 N 5 5 5

STATIC R MEDIAN 0 0 0 0

SCOREO011 N 5 5 5 5

GRIP MEDIAN 0 0 0 0

SCOREO/1 N 5 5 5 5

FEMALES

GROUP 1 GROUP 2 GROUP 3 GROUP 4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

AT WEEK 4

HEARING MEDIAN 0 0 0 0

SCOREO/1 N 5 5 5 5

PUPIL L MEDIAN 0 0 0 0

SCORE 0/1 N 5 5 5 5

PUPIL R MEDIAN 0 0 0 0

SCOREO/1 N 5 5 5 5

STATIC R MEDIAN 0 0 0 0

SCOREO/1 N 5 5 5 5

GRIP MEDIAN 0 0 0 0

SCOREO/1 N 5 55

+/++ Steel-test significant at 5% (4)or 1% (++) level

- Page 13 - 08Oct08 12h2l
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

MOTOR ACTIVITY MEASUREMENTS SUMMARY

MALES

WEEK 4

GROUP 1 GROUP 2 GROUP 3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

total high MEAN 3328 2361 2081 1777

sensor count ST. DEV 1736 578 573 581

N 5 5 5 5

total low MEAN 4533 4690 4204 4759

sensor count ST. DEV 1009 929 266 1030

N 5 5 5 5

FEMALES

LACTATION

GROUP 1 GROUP 2 GROUP 3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

total high MEAN 2238 1917 2124 1422

sensor count ST.DEV 617 1072 1239 644

N 5 5 55

total low MEAN 16563 6827 18471 18503

sensor count ST.DEV 3736 4455 6655 6789

N 5 5 55

I~Dunnett-test based on pooled variance significant at 5% ()or 1 % ()level

- Page 14-
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX I

HAEMATOLOGY SUMMARY

MALES

GROUP 1 GROUP 2 GROUP 3 GROUP 4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

END OF TREATMENT

WBC MEAN 7.1 8.5 8.3 7.6

10E9/L ST.DEV 1.4 1.1 1.6 1.4

N 5 5 5 5

Neutrophils MEAN 15.4 13.6 15.9 12.1

%WBC ST.DEV 2.7 2.3 3.4 3.8

N 5 5 5 5

Lymphocytes MEAN 80.7 83.1 80.5 84 8

%WVBC ST.DEV 3.2 2.5 4.2 3.8

N 5 5 5 5

Monocytes MEAN 2.2 2.2 2.3 1.9

%WBC ST.DEV 0.3 0.5 0.7 0.6

N 5 5 5 5

Eosinophils MEAN 1.3 0.8 0.9 0.8

%WBC ST.DEV 0.6 0.3 0.2 0.4

N 5 5 5 5

Basophils MEAN 0.4 0.4 0.4 0.4

%WBC ST.DEV 0.1 0.1 0.1 0.2

N 5 5 5 5

Red blood cells MEAN 8.11 8.67 8.76 8.69

10E12/L ST.DEV 0.44 0.53 0.51 0.31

N 5 5 5 5

Reticulocytes MEAN 2.8 2.7 2.6 2.4

%RBC ST.DEV 0.3 0.7 0.3 0.3

N 5 5 5 5

ROW MEAN 12.7 13.6 12.8 15.0

%ST.DEV 0.9 2.1 0.4 5.0

N 5 5 5 5

Haemoglobin MEAN 9.7 9.9 10.1 9.9

mmolIL ST.DEV 0.4 0.6 0.4 0.2

N 5 5 5 5

Haematocrit MEAN 0.424 0.444 0455 0.448

L/L ST.DEV 0.025 0029 0.020 0.010

N 5 5 5 5

MCV MEAN 52.4 51.2 51 9 51.6

f'L ST.DEV 0.9 1.7 0.9 1.0

N 5 5 5 5

MCH MEAN 1.20 1.14 1.15 1.14

fmol ST.DEV 0.04 0 07 0.03 0.034

N 5 5 5 5

MCHC MEAN 22.91 22.27 22.12 22.09

mmol/L ST.DEV 0.68 0 66 0.30 0.48

N 5 5 5 5

Platelets MEAN 805 862 873 981

10E9/L ST.DEV 179 93 90 71

IN 5 5 5 5

PT MEAN 17.9 18.1 17.4 17.6

s ST.DEV 1.4 0.9 0.5 0.6

N 4 5 5 5

APTT MEAN 18.9 20.7 19.2 19.4 s ST.DEV 1.1 1.7 2.8 2.4

IN 4 5 5 5

+/++ Steel-test significant at 5% (+) or 1 % (++) level

/~Dunnett-test based on pooled variance signlificanlt at 5% ()or 1 % ()level

- Page 15 - 29Sep08 09h39
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

HAEMATOLOGY SUMMARY

FEMALES

GROUP 1 GROUP 2 GROUP 3 GROUP 4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

END OF TREATMENT

WBC MEAN 4.0 4.0 4.5 5.7

10E9/L ST.DEV 0.5 1.4 1.7 1.1

N 5 4 5 5

Neutrophils MEAN 21.9 26.1 28.3 24.8

%WBC ST.DEV 6.3 15.6 30 9.8

N 5 5 5 5

Lymphocytes MEAN 74.6 71.4 68.6 73.2

%WBC ST.DEV 5.9 15.5 3.1 8.7

N 5 5 5 5

Monocytes MEAN 2.6 2.0 2.1 1.4

%WVBC ST.DEV 0.9 0.65 0.6 1.0

N 5 5 5 5

Eosinophils MEAN 0.7 0.3 0.6 0.5

%WGC ST.DEV 0.6 0.4 0.5 0.5

N 5 5 55

Basophils MEAN 0.2 0.1 0.4 0.1

%WBC ST.DEV 0.2 0.2 0.4 0.1

N 6 5 5 5

Red blood cells MEAN 7.29 7,59 7.32 7.62

10E12/L ST.DEV 0.17 0.47 0.28 0.23

N 5 5 5 5

Reticulocytes MEAN 4.8 4.8 5.7 5.3

%RBC ST.DEV 1.0 1.9 1.9 0.9

N 5 5 5 5

ROW MEAN 14.4 15.6 158 14.0

%ST.DEV 1.8 3.7 1.5 1.2

N 5 5 5 5

Haemoglobin MEAN 689 9.3 8.8 9.2

mmol/L ST.OEV 0.3 0.1 04 0.2

N 5 5 5 5

Haemnatocrit MEAN 0.386 0.405 0.386 0.405

LUL ST.DEV 0.008 0.009 0.014 0.014

N 5 5 5 5

MCV MEAN 52.9 535 52.7 53.1

fL_ ST.DEV 1.2 2,2 04 1.1

N 5 555

MCII MEAN 122 1.23 120 1.20

fmol ST.DEV 0.04 0.08 0.03 0.05

N 5 5 5 5

MOHO MEAN 2309 22.94 2276 22.64

mrnol/L ST.OEV 0.56 0.55 0.64 0.58

N 5 5 5 5

Platelets MEAN 1117 1320 1385 1195

10E9/L ST.DEV 315 95 274 322

N 5 5 5 5

PT MEAN 17.1 16.5 16.3 15.6

sST.DEV 0.5 06 0.7 0.5

N 4 5 5 5

APTT MEAN 17.7 18.4 16.4 15.9

s ST.DEV 23 3.4 1.5 3.4

N 4 5 55

+/++ Steel-test significant at 5% (+) or 1% (++) level

SDunnett-test based on pooled variance significant at 5% ()or 1% (**) level

-Page 16 - N10- 29Sep08 09h39
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

CLINICAL BIOCHEMISTRY SUMMARY

MALES

GROUPI1 GROUP2 GROUP3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

END OF TREATMENT

ALAT MEAN 44.7 52.0 46.8 47.8

UIL ST.OEV 7.5 14.1 6.7 6.2

N 5 5 5 5

ASAT MEAN 81.9 82.9 79.7 82.6

UIL ST.DEV 17.0 10.1 5.3 5.5

N 5 5 5 5

ALP MEAN 154 173 150 143 U/L ST.DEV 40 28 23 33

N 5 5 5 5

Total protein MEAN 63.3 65.2 65.4 66.2

gIL ST.DEV 2.8 2.3 2.1 1.3

N 5 5 5 5

Albumin MEAN 33.4 33.7 34.0 34.3

gIL ST.DEV 1.3 0.8 0.5 0.4

N 5 5 5 5

Total bilirubin MEAN 2.2 2.4 2.2 2.0

umol/L ST.DEV 0.3 0.3 0.3 0.2

N 5 5 5 5

Urea MEAN 8.8 9.5 8.6 8.4

mmol/L ST.DEV 2.3 1.3 0.6 1.1

N 5 5 5 5

Creatinine MEAN 42.8 46.0 46.3 43.0

umol/L ST.DEV 1.7 6.8 4.7 1.5

N 5 5 5 5

Glucose MEAN 8.07 7.49 7.32 7.20

mmol/L ST.OEV 1.49 1.70 0.98 1.56

N 5 5 5 5

Cholesterol MEAN 1.65 1.97 1.88 2.13

mmol/L ST.DEV 0.31 0.20 0.27 0.33

N 5 5 5 5

Sodium MEAN 142.3 142.0 1426 143.4

mmolIL ST.DEV 0.4 0.7 0.8 1.7

N 5 5 5 5

Potassium MEAN 3.77 4.03 3.81 4.03

mmolIL ST.DEV 0.23 0.16 0.07 0.26

N 5 5 5 5

Chloride MEAN 104 104 105 105

mmol/L. ST.IJEV 1 0 1 1

N 5 5 5 5

Calcium MEAN 2.78 2.71 2.73 2.73

mmolIL ST.DEV 0.04 0.05 0.08 0.04

N 5 5 5 5

Inorg.Phos MEAN 2.07 2.05 2.18 2.04

rnmol/L ST.OEV 0.21 0.25 0.13 0.13

N 5 5 5 5

D unnet-test based on pooled variance significant at 5% ()or I% (**) level

- Page 17 - * 29Sep08 09h39
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

CLINICAL BIOCHEMISTRY SUMMARY

FEMALES

GROUP 1 GROUP 2 GROUP 3 GROUP 4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

END OF TREATMENT

ALAT MEAN 58.1 67.8 666 46.2

U/L ST.DEV 13.0 19.6 8.2 6.3

N 5 5 5 5

ASAT MEAN 80.9 75.1 80.1 70.2

U/L ST.OEV 11.7 12.0 10.2 5.9

N 5 5 5 5

ALP MEAN 65 72 70 69

U/L STODEV 22 19 20 44

N 5 5 5 5

Total protein MEAN 65.8 65.3 67.7 65.6

gIL ST.OEV 1.1 2.9 2.2 2.2

N 5 5 5 5

Albumin MEAN 32.6 32.1 33.8 32.7

g/L ST.DEV 0.9 2.2 1.2 1.0

N 5 5 5 5

Total bilirubin MEAN 2.4 2.0 2.5 1.8

umol/L ST.OEV 0.4 0.4 0.6 0.3

N 5 5 5 5

Urea MEAN 9.4 10.1 10.0 9.7

mmoll ST.DEV 2.0 1.2 0.9 1.2

N 5 5 5 5

Creatinine MEAN 46.7 47.2 46.6 48.0

umol/L ST.DEV 3.5 1.6 5.0 3.0

N 5 5 5 5

Glucose MEAN 6.48 5.92 5.47 5.81

mmol/L ST.DEV 1.28 1.23 0.82 0.76

N 5 5 5 5

Cholesterol MEAN 1.45 1.69 1.69 1.75

mmol/L ST.DEV 0.21 0.32 0.15 0.33

N 5 5 5 5

Sodium MEAN 141.5 141.0 141.2 141.2

mmol/L ST.DEV 1.9 0.9 1.2 0.5

N 5 5 5 5

Potassium MEAN 3.58 3.79 3.67 3.61

mmol/L ST.DEV 0.24 0.16 0.19 0.46

N 5 5 5 5

Chloride MEAN 99 99 97 97

mmol/L ST.DEV 2 2 1 2

N 5 5 5 5

Calcium MEAN 2.78 2.75 2.81 2.74

mmol/L ST.DEV 0.03 0.06 0.09 0.04

N 5 8 5 5

Inorg.Phos MEAN 2.27 2.23 2.59 2.31

mmol/L ST.DEV 0.34 0.32 0.26 0.19

N 5 5 5 5

*r. Ounnetl-test based on pooled variance significant at 5% ()or 1% ()level

-Page 18 - 29Sep08 09h39
ClI. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

MACROSCOPIC FINDINGS SUMMARY

MALES

GROUP 1 GROUP 2 GROUP 3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

NECROPSY

Animals examined 10 10 10 10

Animals without findings 9 7 1 0

Animals affected 1 3 9 10

Epididymides Nodule(s) 0 0 2 0

Seminal vesicies

Reduced in size 0 0 0 1

Mesenteric Inode

Discolouration 0 2 9 ## 10 ##

Skin

Alopecia 0 0 0 1

Discolouration 0 1 0 1

Bone

Tail bent 1 0 0 0

FEMALES

GROUP 1 GROUP 2 GROUP 3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

NECROPSY

Animals examined 10 10 10 10

Animals without findings 9 8 2 0

Animals affected 1 2 8 10

General observations

Contents: 0 0 07#

Pancreas

Discolouration 0 1 0 0

Adrenal glands Discolouration 0 0 0 1

Mesenteric Lnode

Discolouration 0 1 8 ## 9 #

Mandibular Lnode

Discolouration 1 2 0 1

Skin

Alopecia 1 0 0 1

## Fisher's Exact test significant at 5% #)or 1 % (#level

- Page 19 00- 29Sep08 09h39
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

ORGAN WEIGHTS (GRAM) SUMMARY

MALES

GROUP 1 GROUP 2 GROUP 3 GROUP 4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

NECROPSY

BODY W, MEAN 355 349 358 348

(GRAM) ST.DEV 26 27 17 7

N 10 10 10 10

BRAIN MEAN 2.04 2.02 2.02 2.08

(GRAM) ST.DEV 0.06 0.04 0,07 0.04

N 5 5 5 5

HEART MEAN 0.909 0.943 0.991 1.005

(GRAM) ST.DEV 0.035 0.053 0083 0.083

N 5 5 5 5

LIVER MEAN 8.44 8.22 9.03 9.23

(GRAM) ST.DEV 0.92 1.03 1.04 0.85

N 5 5 5 5

THYMUS MEAN 0.355 0.369 0.394 0.341

(GRAM) ST 0EV 0.062 0.091 0.038 0.043

N 5 5 5 5

KIDNEYS MEAN 2.43 2.40 2.35 2.41

(GRAM) ST DEV 0.05 0.12 0.16 0.20

N 5 5 5 5

ADRENALS MEAN 0.069 0.058 0.085 0.075

(GRAM) ST.DEV 0.012 0.010 0.005 0.013

N 5 5 5 5

SPLEEN MEAN 0.559 0.590 0.627 0.672

(GRAM) ST DEV 0.114 0.080 0,077 0.032

N 5 5 5 5

TESTES MEAN 3.36 3.28 3.30 3.39

(GRAM) ST.DEV 0.15 0.42 0.38 0.34

N 10 10 9 10

EPIDIDYMIDES MEAN 1.139 1.138 1.207 1 182

(GRAM) ST.DEV 0.086 0.114 0.136 0.124

N 10 10 9 10

/~Dunnett-test based on pooled variance significant at 5% ()or 1 % ()level

- Page 20 - 29Sep08 09h39
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

ORGANIBODY WEIGHT RATIOS I(%) SUMMARY

MALES

GROUP 1 GROUP2 GROUP3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

NECROPSY

BODY W. MEAN 355 349 358 348

(GRAM) ST.DEV 26 27 1 7 7

N 10 10 10 10

BRAIN MEAN 0.60 0.60 0.58 0.60

()ST.DEV 0.03 0.03 0.03 0.01

N 5 5 5 5

HEART MEAN 0.268 0.278 0.283 0.288 ()ST.DEV 0.014 0.017 0.012 0.020

N 5 5 5 5

LIVER MEAN 2.48 2.42 2.57 2.64 ()ST.DEV 0.17 0.20 0.16 0.20

IN 5 5 5 5

THYMUS MEAN 0.104 0.108 0.112 0.098 ()ST.DEV 0.016 0.024 0.006 0.012

N 5 5 5 5

KIDNEYS MEAN 0.72 0.71 0.67 0.69 %)ST.OEV 0.03 0.03 0 02 0.04

N 5 5 5 5

ADRENALS MEAN 0.020 0.020 0 019 0.021

%)ST.DEV 0.003 0.003 0.002 0.003

N 5 5 5 5

SPLEEN MEAN 0.164 0.174 0.178 0.193

(%) ST 0EV 0026 0.016 0015 0.007

N 5 5 5 5

TESTES MEAN 0.95 094 0.92 0.98 ()ST.OEV 0.09 0.14 0.08 0.10

N 10 10 9 10

EPIDIDYMIDES MEAN 0.323 0.327 0.339 0.340 ()ST.DEV 0.040 0.036 0.035 0.033

IN 10 10 9 10

/ unnett-test based on pooled variance significant at 5% ()or 1 % ()level

- Page 21 - NIP 29Sep08 09h39
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

ORGAN WEIGHTS (GRAM) SUMMARY

FEMALES

GROUP 1 GROUP 2 GROUP 3 GROUP 4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

NECROPSY

BODY W. MEAN 225 220 228 221

(GRAM) ST.DEV 17 18 2 15

N 5 5 5 5

BRAIN MEAN 1.84 1.79 1.83 1.86

(GRAM) ST.DEV 0.05 0.04 0.06 0.01

N 5 5 5 5

HEART MEAN 0 733 0.725 0.785 0.716

(GRAM) ST.DEV 0.073 0 055 0.041 0 043

N 5 5 5 5

LIVER MEAN 7,23 7.07 781 7.11

(GRAM) ST.DEV 0.52 0.78 0.32 0.63

N 5 5 5 5

THYMUS MEAN 0.160 0.170 0.171 0.179

(GRAM) ST.DEV 0.055 0.022 0.022 0.026

N 5 5 5 5

KIDNEYS MEAN 1.64 1.62 1.74 1.63

(GRAM) ST 0EV 0.21 008 0.08 0.16

N 5 5 55

ADRENALS MEAN 0.079 0.084 0.082 0.088

(GRAM) ST.DEV 0.005 0.009 0.008 0.014

N 5 6 5 5

SPLEEN MEAN 0.457 0.462 0.538 0.501

(GRAM) ST.DEV 0.088 0.057 0.091 0.061

N 5 5 5 5

IDunnett-test based on pooled variance significant at 5% ()or 1 % ()level

- Page 22 - *I" 29Sep08 09h39
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

ORGANIBODY WEIGHT RATIOS (%) SUMMARY

FEMALES

GROUP 1 GROUP 2 GROUP 3 GROUP 4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

NECROPSY

BODY W MEAN 225 220 228 221

(GRAM) ST.DEV 17 18 2 15

N 55 5 5

BRAIN MEAN 0.82 0.82 0.80 0.84 ()ST.DEV 0 08 0.07 0.03 0.06

N 5 5 5 5

HEART MEAN 0.327 0.331 0.344 0.325

()ST.DEV 0.035 0,026 0 016 0.021

N 5 5 5 5

LIVER MEAN 3.22 3.21 3.42 3.23

()ST.DEV 0.27 013 0.15 0.28

N 5 5 5 5

THYMUS MEAN 0071 0,077 0.075 0.081

()ST.DEV 0023 0.010 0.010 0.014

N 5 5 5 5

KIDNEYS MEAN 0.73 0.74 0.76 0.74 ()ST.DEV 0.11 005 0.03 0.10

N 5 5 5 5

ADRENALS MEAN 0035 0.038 0.036 0040 ()ST.DEV 0.003 0.003 0.003 0,005

N 5 5 5 5

SPLEEN MEAN 0.202 0.211 0.236 0.227

(

0 )ST.DEV 0.035 0.031 0.042 0.029

N 5 5 5 5

*r* Dunnett-test based on pooled variance significant at 5% ()or 1 % (**) level

-Page 23 - 00- 29Sep08 09h39
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

MOTOR ACTIVITY TEST

MALES

WEEK 4

Counts per sample period (hour)

Animal Location of

number sensor 1 2 3 4 5 6 7 8 9 10 11 12 Total

GROUP I(CONTROL)

1 High 172 330 112 410 368 110 202 35 182 130 148 54 2253

1 Low 410 465 224 702 625 440 292 283 445 430 549 238 5103

2 High 1919 189 1117 556 768 63 354 276 186 405 213 363 6407

2 Low 457 336 441 505 446 156 245 468 325 245 348 120 4092

3 High 224 138 409 565 330 263 67 247 198 142 179 120 2882

3 Low 425 328 616 1125 488 645 176 372 301 625 590 335 6026

7 Hiugh 85 166 278 499 339 217 169 55 112 143 216 222 2501

7 Low 346 248 483 591 226 199 343 86 198 330 425 313 3788

10 High 305 209 226 320 189 259 207 53 250 160 305 115 2598

10 Low 139 370 513 537 236 310 355 66 300 120 338 373 3657

GROUP 2 (50 MG/KG)

11 High 94 225 369 186 319 240 191 237 2 70 268 172 2373

11 Low 339 455 503 339 765 805 251 690 29 95 481 391 5143

12 High 140 134 98 246 216 ill 0 47 242 108 132 242 1716

12 Low 137 277 169 471 795 236 26 120 398 248 329 634 3840

14 High 109 183 309 424 227 33 101 123 175 90 110 167 2051

14 Low 359 559 404 587 115 258 159 176 422 308 177 246 3770

15 High 107 513 313 690 463 445 8 37 190 194 200 109 3269

15 Low 161 742 557 596 561 812 131 261 340 554 589 680 5984

16 High 65 126 315 332 284 179 148 44 149 243 316 194 2395

16 Low 436 385 330 673 564 130 536 202 291 640 425 103 4715

GROUP 3 (150 MG/KG)

21 High 225 119 70 159 146 330 196 7 19 227 245 73 1816

21 Low 418 195 158 466 314 468 273 129 295 943 562 306 4527

22 High 74 110 284 383 333 0 42 454 108 0 17 6 1811

22 Low 217 295 681 574 570 2 345 870 146 9 138 35 3882

23 High 16 208 125 184 132 50 96 219 96 100 248 100 1574

23 Low 217 361 430 447 331 84 661 243 432 386 503 310 4405

24 High 308 235 286 252 397 197 287 23 297 120 433 202 3037

24 Low 365 283 185 386 490 482 415 102 186 423 396 305 4018

25 High 161 138 287 313 258 182 221 104 93 139 110 81 2067

25 LOW 341 439 402 513 397 223 512 251 211 326 354 218 4187

GROUP 4 (500 MG/KG)

32 High 89 11 5 66 137 29 1 237 1 79 119 112 886

32 Low 331 119 116 461 436 140 73 699 43 280 3715 345 3418

34 High 258 297 297 252 106 204 322 9 90 100 90 147 2172

34 Low 582 857 655 404 484 790 601 172 132 213 285 323 5498

35 High 300 23 216 449 721 28 283 1 22 15 126 171 2355

35 -Low 632 313 350 750 881 223 672 154 381 339 741 611 6047

36 High 68 130 102 170 205 340 168 65 218 0 50 44 1560

36 Low 470 494 131 417 430 687 439 200 609 85 229 269 4460

37 High 216 25 226 185 243 140 384 73 113 38 148 123 1914

37 Low 578 159 455 422 456 249 718 293 245 86 386 327 4374

-Page 24-
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

REPRODUCTION PROCESSES

FO-GENERATION - POST COITUM

FEMALE MALE MATING PREGNANT SCHEDULE DELIVERY NECROPSY

NUMBER NUMBER DATE RECORDED DATE

GROUP 1 (CONTROL)

41 <NP> 1 02JUL2008 NO ---- 29JUL2008

42 2 02JUL2008 YES BREEDING 23JUL2008 28JUL2008

43 3 02JUL2008 YES BREEDING 23JUL2008 28JUL2008

44 4 05JUL2008 YES BREEDING 27JUL2008 OIAUG2008

45 5 OSJUL2008 YES BREEDING 27JUL2008 CiAUG2008

46 6 05JUL2008 YES BREEDING 27JUL2008 OIAUG2008

47 7 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

48 8 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

49 9 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

50 10 02JUL2008 YES BREEDING 23JUL2008 28JUL2008

GROUP 2 (50 MG/KG)

51 11 04JUL2008 YES BREEDING 26JUL2008 31JUL2008

52 12 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

53 13 05JUL2008 YES BREEDING 27JUL2008 01AUG2008

54 <NP> 14 02JUL2008 NO ---- 29JUL2008

55 15 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

56 16 02JUL2008 YES BREEDING 23JUL2008 28.JUL2008

57 17 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

56 18 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

59 19 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

60 20 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

GROUP 3 (250 MG/KG)

61 21 02JUL2008 YES BREEDING 23JUL2008 28JUL2008

62 22 04JUL2008 YES BREEDING 26JUL2008 31JUL2008

63 23 02JUL2008 YES BREEDING 23JUL2008 28JUL2008

64 24 02JUL2008 YES BREEDING 23JUL2008 28JUL2008

65 25 02JUL2008 YES BREEDING 23JUL2008 28JUL2008

66 <NP> 26 04JUL2008 NO ---- 29JUL2008

67 27 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

68 28 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

69 29 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

70 30 05JUL2008 YES BREEDING 26JUL2008 31JUL2008

GROUP 4 (1000 MGKG)

71 31 05.JUL2008 YES BREEDING 27JUL2008 01AUG2008

72 <10> 32 02JUL2008 YES -- -- 29JUL2008

73 33 04JUL2008 YES BREEDING 25JUL2008 30JUL2008

74 34 02JUL2008 YES BREEDING 23JUL2008 28JUL2008

75 35 02JUL2008 YES BREEDING 23JUL2008 28JUL2008

76 36 03JUL2008 YES BREEDING 24JUL2008 29JUL2008

77 37 02JUL200B YES BREEDING 23JUL2008 28JUL2008

78 38 05JUL2008 YES BREEDING 27JUL2008 01AUG2008

79 39 05JUL2008 YES BREEDING 26JUL2008 31JUL2008

80 40 05JUL2008 YES BREEDING 26JUL2008 31JUL2008

<10> implantation sites only

< N P > N o n - p re g n a n t P g 42lc 

O 6 O
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

MATING PERFORMANCE

FO-GENERATION - POST COITUM

DAY OF THE GROUP 1 GROUP2 GROUP3 GROUP4

PAIRING PERIOD CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

NUMBER OF FEMALES MATED DURING THE FIRST PAIRING PERIOD

1 4 2 4 4

2 1

3 3 7 5 1

4 3 1 1 4

MEDIAN PRECOITAL TIME 3 3 3 3

MEAN PRECOITAL TIME 2.5 2.7 2.3 2.5

N 10 10 10 10

+/++ Steel-test significant at 5% ()or 1 % (++) level

- Page 25 - 23Oct08 10h08
C.L PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 1

BREEDING DATA PER GROUP

FO-GENERATION - LACTATION

GROUPi GROUP2 GROUP3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

LITTERS

TOTAL 9 9 9 9

DURATION OF GESTATION

MEAN (+) 21.3 21.2 21.1 21.2

ST.DEV. 0.5 0.4 0.3 0.4

N 9 9 9 9

DEAD PUPS AT FIRST LITTER CHECK

LITTERS AFFECTED ()0 0 1 2

TOTAL 0 0 1 2

MEAN (+) 0.0 0.0 0.1 0.2

ST DEV. 0.0 0.0 0.3 0.4

N 9 9 9 9

LIVING PUPS AT FIRST LITTER CHECK

% OF MALES /FEMALES(# 44/56 47/53 50/50 50/50 TOTAL 104 103 101 114

MEAN (+) 11.6 11.4 11.2 12.7

ST.DEV. 3.3 1.6 3.9 1.7

N 9 9 9 9

POSTNATAL LOSS

% OF LIVING PUPS 1.0 1.0 0.0 1.8

LITTERS AFFECTED (#) I 1 0 2

TOTAL (#) 1 1 0 2

MEAN (+) 0.1 0.1 0.0 0.2

ST DEV. 0.3 0.3 0.0 0.4

N 9 9 9 9

VIABILITY INDEX (#) 99.0 99.0 100.0 98.2

Viability index =(Number of alive pups before planned necropsy /Number of pups born alive) *100

-+/++ Steel-test significant at 5% (+) or 1 % (++) level

#1 Fisher's Exact test significant at 5% (#) or 1 % (##) level

- Page 26 -23Oct08 16h08
CI. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX I

MEAN BODY WEIGHTS OF PUPS PER GROUP (GRAM)

FO-GENERATION - LACTATION

DAY SEX GROUP 1 GROUP 2 GROUP 3 GROUP 4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

1 M MEAN 6.0 6.1 6.1 5.9

ST.DEV. 0.6 0.7 0.4 0.4

N 9 9 9 9

F MEAN 5.8 5.7 5.6 5.6

ST.DEV. 0.5 0.7 0.4 0.4

N 9 9 8 9

M+F MEAN 5.9 5.9 5.9 5.8

ST.DEV. 0.5 0.7 04 0.4

N 9 9 9 9

4 M MEAN 8.6 9.1 9.0 8.5

ST.DEV. 0.9 1.1 0.9 0.6

N 9 9 9 9

F MEAN 8.4 8.6 8.3 8.2

ST.DEV. 1.0 1.0 0.9 0.8

N 9 9 8 9

M+F MEAN 8.5 8.8 87 8.3

ST.DEV 09 1.1 1.0 0.7

N 9 9 9 9

D unnett-test based on pooled variance significant at 5% ()or 1 % ()level

- Page 27 - 23Oct08 16h08
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

APPENDIX 2 INDIVIDUAL TABLES

-Page 1 -
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

MORTALITY DATA

MALES

ANIMAL SCHEDULED TREATMENT

SACRIFICE FROM TO

GROUP 1 (CONTROL)

1 17JUL08 17JUNOB 16JUL08

2 17JUL08 17JUN08 16JUL08

3 17JUL08 17JUN08 16JUL08

4 17JUL08 17JUN08 16JUL08

5 17JUL08 17JUN08 16JUL08

6 17JUL08 17JUN08 16JUL08

7 17JUL08 17JUN08 16JUL08

8 17JUL08 17JUN08 16JUL08

9 17JUL08 17JUN08 16JUL08

10 17JUL08 17JUN08 I6JUL08

GROUP 2 (50 MGIKG) 11 17JUL08 17JUN08 16JUL08

12 17JUL08 17JUN08 16JUL08

1 3 17JUL08 17JUN08 16JUL08

k14 17JUL08 17JUN08 16JUL08

15 17JUL08 17JUN08 16JUL08

16 17JUL08 17JUN08 16JUL08

17 17JUL08 17JUN08 16JUL08

18 17JUL08 17JUN08 16JUL08

19 17JUL08 17JUN08 16JUL08

20 17JUL08 17JUN08 16JUL08

GROUP 3 (260 MG/KG)

21 17JUL08 17JUN08 16JUL08

22 17JUL08 17JUN08 16JUL08

23 17JUL08 17JUN08 16JUL08

24 17JUL08 17JUN08 16JUL06

25 17JUL08 17JUN08 16JUL08

26 17JUL08 17JUN08 16JUL08

27 17JUL08 17JUN08 16JUL08

28 17JUL08 17JUN08 16JUL08

29 17JUL08 17JUN08 16JUL08

30 17JUL08 17JUN08 16JUL08

GROUP 4 (1000 MG/KG)

31 17JUL08 17JUN08 16JUL08

32 17JUL08 17JUN08 16JUL08

33 17JUL08 17JUN08 16JUL08

34 17JUL08 17JUN08 16JUL08

36 17JUL08 17JUN08 16JUL08

36 17JUL08 17JUN08 16JUL08

37 17JUL08 17JUN08 16JUL08

38 17JUL08 17JUN08 16JUL08

39 17JUL08 17JUN08 16JUL08

40 17JUL08 17JUN08 16JUL08

- Page 2 - 29SepO8lOh2l
ClI. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

MORTALITY DATA

FEMALES

ANIMAL SCHEDULED TREATMENT

SACRIFICE FROM TO

GROUP I (CONTROL)

41 29JUL08 17JUN08 28JUL08

42 28JUL08 17JUN08 27JUL08

43 28JUL08 17JUN08 27JUL08

44 01AUG08 17JUN08 31JUL08

46 01AUG08 17JUN08 31JUL08

46 01AUG08 17JUN08 31JUL08

47 30JUL08 17JUN08 29JUL08

48 30JUL08 17JUN08 29JUL08

49 30JUL08 17JUN08 29JUL08

50 28JUL08 17JUN08 27JUL08

GROUP 2 (50 MG/KG)

51 31JUL08 17JUN08 30JUL08

52 30JUL08 17JUN08 29JUL08

53 01AUG08 17JUN08 31JUL08

54 29JUL08 17JUN08 28JUL08

55 30JUL08 17JUN08 29JUL08

56 28JUL08 17JUN08 27JUL08

57 30JUL08 17JUN08 29JUL08

58 30JUL08 17JUN08 29JUL08

59 30JUL08 17JUN08 29JUL08

60 30JUL08 17JUN08 29JUL08

GROUP 3 (250 MG/KG)

61 28JUL08 17JUN08 27JUL08

62 31JUL08 17JUN08 30JUL08

63 28JUL08 17JUN08 27JUL08

64 28JUL08 17JUN08 27JUL08

65 28JUL08 17JUN08 27JUL08

66 29JUL08 17JUN08 28JUL08

67 30JUL08 17JUN08 29JUL08

68 30JUL08 17JUN08 29JUL08

69 30JUL08 17JUN08 29JUL08

70 31JUL08 17JUN08 30JUL08

GROUP 4 (1000 MG/KG)

71 01AUG08 17JUN08 31JUL08

72 29JUL08 17JUN08 28JUL08

73 30JUL08 17JUN08 29JUL08

74 28JUL08 17JUN08 27JUL08

75 28JUL08 17JUN08 27JUL08

76 29JUL08 17JUN08 28JUL08

77 28JUL08 17JUN08 27JUL08

78 01AUG08 17JUN08 31JUL08

79 31JUL08 17JUN08 30JUL08

80 31JUL08 17JUN08 30JUL08

-Page 3 - 29Sep08 10h21
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL SIGNS

MALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: 1 ............................ ... 4....

(LOCATION) DAY: 123456712345671234567123456712345671234567123

GROUP I (CONTROL)

ANIMAL I

No clinical signs noted

ANIMAL 2

No clinical signs noted

ANIMAL 3

Various

Broken (1) G:.................. ....... 111

(Tell apex)

ANIMAL 4

No clinical signs noted

ANIMAL 5

No clinical signs noted

ANIMAL 6

No clinical signs noted

ANIMAL 7

No clinical signs noted

ANIMAL 8

No clinical signs noted

ANIMAL 9

No clinical signs noted

ANIMAL 10

No clinical signs noted

GROUP 2 (50 MG/KG)

ANIMAL 11

Skin /fur

Blue discolouration (1) G: .... 111111111111111111111111

(Faeces)

Secretion / excretion

Salivation (3) G:............. I........... 1...

ANIMAL 12

Skin /fur

Blue discolouration (1) G: .... 111111111111111111i111111

(Faeces)

ANIMAL 13

Skin / fur

Blue discolouration (1) G: .... 1111111111111111111111111

(Faeces)

ANIMAL 14

Skin / fur

Blue discolouration (1) G: .... 111111111111111111111111 (Faeces)

ANIMAL 15

Skin / fur

Blue staining (1) G:................ 1111111111111111

(Tail)

Blue discolouration (1) G: . 111111111111111111111111

(Faeces)

ANIMAL 16

Skin I fur

Blue discolouration (1) G: 111111111111111111111111

(Faeces)

ANIMAL 17

Skin / fur

Blue discolouration (1) G: .... 1111111111111111111111111

(Faeces)

G0 Highest daily grades

Observation performed, sign not present

- Page 4 -fr24Oct08 15h46
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL SIGNS

MALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: I.............I .................... ...

(LOCATION) DAY: 123456712345671234667123456712345671234567123

GROUP 2 (50 MG/KG)

ANIMAL 16

Skin / fur

Scabs (3) G:..........I.............. 11111111

(Neck)

Blue discolouration (1) G: 111. II111111111111111111111

(Faeces)

ANIMAL 19

Skin / fur

Blue discolouration (1) G: .... 111111111111111111111111

(Faeces)

ANIMAL 20

Skin /fur

Blue discolouration (1) G: .... iiiiiiiiiiiiiiiiiiiiiii

(Faeces)

GROUP 3 (250 MG/KG)

ANIMAL 21

Skin /fur

Blue discolouration (1) G: ... .11111111111111111111111111

(Faeces)

Secretion / excretion

Salivation (3) G......................

ANIMAL 22

Skin / fur

Blue discolouration (1) G: ... 111111111111111111111111111l

(Faeces)

ANIMAL 23

Skin / fur

Alopecia (3) G:.......... .......... I.........11

(Forelegs)

Blue discolouration (1) G: . .. iiiiiiiiiiiiiiiiiii'liiii

(Faeces)

Secretion / excretion

Salivation (3) G:......I.............. ........ 1

ANIMAL 24

Skin /fur

Blue discolouration (1) G: ... .1111111111111111111111111111

(Faeces)

ANIMAL 25

Skin / fur

Blue dscolouration (1) G: ... 111111111111111111111111111

(Faeces)

Secretion / excretion

Salivation (3) G:.............. I.............. 1.

ANIMAL 26

Skin / fur

Blue discolouration (1) G: ... .111111111111111111111111111

(Faeces)

ANIMAL 27

Skin /fur

Blue discolouration (1) G: ... 1111111111111

(Faeces)

Secretion / excretion

Salivation (3) G:..............................1

G: Highest daily grades

Observation performed, sign not present

-PageS 5 24Oct08 15h46
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL SIGNS

MALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: 1 ............ 1...................4.....

(LOCATION) DAY: 123456712345671234567123456712345671234567123

GROUP 3 (250 MG/KG)

ANIMAL 28

Skin/Ifur

Scabs (3) G:..........................1

(Neck)

Blue discolouratIon (1) G: . 111111111111111111111111111

(Faeces)

Secretion / excretion

Salivation (3) G:.......I................ .... 1.

ANI MAL 29

Skn / fur

Blue discolouration (1) G: . . .111111111111111111111111.

(Faeces)

ANIMAL 30

Skin/Ifur

Blue staining (1) G: . . . . . . . . . . .

(Back)

Blue staining (1) G..........................1I

(Tail)

(Faeces)

GROUP 4 (1000 MG/KG)

ANIMAL 31

Skin /fur

Alopecia (3) G:............... ... .. I.......11

(Foreleg right)

Scabs(3) 0:.......................111....

(Foreleg ight)

Blue discolouratlon (1) G: 1111111111111111111111111111ii

(Faeces)

ANIMAL 32

Skin/Ifur

Blue discolouration (1) G: .. 111111111111111111111111111

(Faeces)

ANIMAL 33

Skin / fur

Blue discolouration (1) G .. 111111111111111

(Faeces)

Secretion / excretion

Salivation (3) G:..........11111111111111111.... I1

ANIMAL 34

Skin/Ifur

Alopecia (3) G:.................... 111112111111

(Forelegs)

Blue discolouration (1) G: .. 1111111111 1111111111111111111

(Faeces)

Secretion / excretion

Salivation (3) G:...............1I...11...

ANIMAL 35

Skin /fur

Blue discolouration (1) G: .. iiiiiiiiiiiiii

(Faeces)

ANIMAL 36

Skin /fur

Blue discolouration (1) G .. 11111111111111

(Faeces)

Secretion / excretion

Salivation (3) G:............111l,... 1.. 11. . 1 ..

G: Highest daily grades

Observation performed, sign not present

- Page 6 -fr24Oct08 15h46
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL SIGNS

MALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: 1.............1 .... ............ .......

(LOCATION) DAY: 123456712345671234567123456712345671234567123

GROUP 4 (1000 MG/KG)

ANIMAL 37

Skin / fur

Blue discolouration (1) G, .1111111111111111111111111111

(Faeces)

ANIMAL 38

Skin / fur

Blue discolouration (1) G: . . 11111111111111111111111111111

(Faeces)

Secretion / excretion

Salivation (3) G:............... 1 . ...

ANIMAL 39

Skin /fur

Blue staining (1) G:....................111111111111

(Tail)

Blue discolouration (1) G: . . 11111111111111111111111111111

(Faeces)

Secretion / excretion

Salivation (3) G- ..... 1.111...11. 11..1

ANIMAL 40

Skin / fur

Blue staining (1) G: .. III1......................

(Back)

Blue discolouration (1) G, .. 11111111111111

(Faeces)

Secretion / excretion

Salivation (3) G:.......... 1.... 1 ... 11111 .1.1

FEMALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: 1I.............I ................. .. ...

(LOCATION) DAY: 123456712345671234567123456712345671234567123

GROUP I(CONTROL)

ANIMAL 41

Skin / fur

Alopecia (3) G:.......... I.......I.......................1

(Abdomen)

ANIMAL 42

Skin / fur

Scabs (3) G:............................. ...... 11111i..

(Mouth)

Scabs (3) G:........................1ill...............

(Snout)

ANIMAL 43

No clinical signs noted

ANIMAL 44

No clinical signs noted

ANIMAL 45

No clinical signs noted

ANIMAL 46

Skin /fur

Alopecia (3) G:............... 11111111 .. .....................

(Back)

(Neck)

G: Highest daily grades

Observation performed, sign not present

- Page 7 - ~24Oct08 l5h46
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL SIGNS

FEMALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: I.............I ................... 4 ....

(LOCATION) DAY: 123456712345671234567123456712345671234567123

GROUP 1 (CONTROL)

ANIMAL 47

Skin /fur

Piloerection (1) G:.... .......... I............. ..... 1..

Alopecia (3) G: .11111 ............ .......................

(Back)

Alopecia (3) G:......................... 12222222

(Abdomen)

Alopecia (3) G:............................... 1111111

(Flank left)

Alopecia (3) G:.................... ................ 111

(Thigh hind left)

Alopecia (3) G:... ............................ 1111..

(Foreleg left)

Alopecie (3) G:........................ ........... .2222233

(Forelegs)

ANIMAL 48

Skin / fur

Alopecia (3) G: 1111111111111111111111111111111 ............

(Neck)

ANIMAL 49

Skin /fur

Alopecia (3) G, .1111111111111..............

(Back)

(Neck)

ANIMAL 50

Skin /fur

(Cheek left)

Alopecia (3) G:........... 11111111 ......................

(Cheek right)

(Back)

Alopecia (3) G: 111111 .... 11111111111111111111111111.

(Neck)

GROUP 2 (50 MG/KG)

ANIMAL 51

Skin /fur

Alopecia (3) G: ... .111111........... .......... (Neck)

Blue discolouration (1) G.........11111111111111111111111111111111111111i

(Faeces)

ANIMAL 52

Skin /fur

Blue discolouration (1) G: ...... 1111111111111111

(Faeces)

ANIMAL 53

Skin /fur

Piloerection (1) G: .. . . .I. . .. . . . . . .

Blue discolouration (1) G: ..... iiiiiiiliiiii iiiiiii iiilii

(Faeces)

ANIMAL 54

Skin /fur

Alopecia (3) G:.......... ....... ....... .... 11111111111111

(Abdomen)

(Faeces)

G: Highest daily grades

Observation performed, sign not present

- Page 8 - ~24Oct08 15h46
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL SIGNS

FEMALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: 1............ 1...................4....

(LOCATION) DAY: 123456712345671234567123456712345671234567123

GROUP 2 (50 MG/KG)

ANIMAL 55

Skin / fur

Blue discolouration (1) G: 1111111111111111111111

(Faeces)

ANIMAL 56

Skin /fur

Alopecia (3) G:..........1111111. ... ...................

(Neck)

Blue discolouration (1) G: ...... 111111111111111111111111111111lil

(Faeces)

ANIMAL 57

Skin /fur

Blue staining (1) G:.........111111 ......... I................ ...

(Tail)

Blue discolouration (1) G- .. 111111111111111111111111111111111111 il

(Faeces)

ANIMAL 58

SkiI/fur

Blue discolouration (1) G: ...... 1111111111111111

(Faeces)

ANIMAL 59

Skin / fur

(Faeces)

ANIMAL 60

Skin /fur

(Faeces)

GROUP 3 (250 MG/KG)

ANIMAL 61

Skin /fur

Blue discolouration (1) G: . .. 11111111111111111111111111111111111111

(Faeces)

ANIMAL 62

Breathing

Rales (3) G:....................... ............ 12...

Skin /fur

Blue staining (1) G:...............1111l..... .....................

(Back)

Blue discolouration (1) G: ... .11111111111111111111111111111111111111111

(Faeces)

ANIMAL 63

Skin /fur

Alopecia (3) G:...........1111111111111i. .................

(Cheek right)

Blue staining (1) G:.....................................1 .... (Mouth)

Blue staining (1) G:.... ............................. 1.

(Abdomen)

Blue staining (1) G: .. . . . . . . . . . . . . . . .. .1 . .

(Forelegs)

(Faeces)

ANIMAL 64

Skin /fur

Alopecia (3) G:........111................. ...

(Cheek right)

(Faeces)

G: Highest daily grades

Observation performed, sign not present

- Page 9 - 24Oct08 15h46
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL SIGNS

FEMALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: 1.................. ............. 4 ...

(LOCATION) DAY, 123456712345671234567123456712345671234567123

GROUP 3 (250 MG/KG)

ANIMAL 65

Skin / fur

Blue discolouration (1) G: .11111111111111111111111111111111111111

(Faeces)

ANIMAL 66

Skin / fur

Piloerection (1) G:......... .. .......................... 1.

(Faeces)

ANIMAL 67

Skin /fur

Blue staining (1) G: .. I ...................................

(Back)

(Faeces)

ANIMAL 68

Skin / fur

Blue staining (1) G: ... .111...... ...................... .

(Snout)

(Faeces)

ANIMAL 69

Skin / fur

(Faeces)

ANIMAL 70

Skin / fur

(Faeces)

GROUP 4 (1000 MG/KG)

ANIMAL 71

Skin / fur

Alopecia (3) G: 111111......................................

(Neck)

Alopecia (3) G:........................................11l...

(Abdomen)

Alopecia (3) G:.................._............ 2222333333333333

(Forelegs)

Alopecia (3) G: .................................. 11222222

( (H-indlegs)

(Faeces)

ANIMAL 72

Skin I fur

Blue discolouration (1) G: .11111111111111111111

(Faeces)

ANIMAL 73

Skin/Ifur

(Tail)

(Faeces)

ANIMAL 74

Skin /fur

(Faeces)

Secretion / excretion

Salivation (3) G:..............1...... ....... .........

G: Highest daily grades

Observation performed, sign not present

- Page 10 - 00 24Oct08 15h46
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL SIGNS

FEMALES

PRE MATING REPRO PHASE

SIGN (MAX. GRADE) WEEK: 1 ............. 1 ................... 4....

(LOCATION) DAY: 123456712345671234567123456712345671234567123

GROUP 4 (1000 MG/KG)

ANIMAL 75

Skin /fur

Blue discolouration (1) G: . .1111111111i111111111111111111111111iii

(Faeces)

ANIMAL 76

Skin /fur

(Faeces)

ANIMAL 77

Skin /Ifur

Alopecia (3) G:........ ............................. 11111i

(Back)

(Faeces)

ANIMAL 78

Skin I fur

(Faeces)

ANIMAL 79

Breathing

Rales (3) G: 1 ........................................

Skin /fur

Scabs(3) G:....................................1 1..

(Mouth)

(Faeces)

Secretion / excretion

Salivation (3) G:................. .... 1 .......... .........

ANIMAL 80

Skin /Ifur

Blue staining (1) G:.........................1.1..................

(Flank right)

(Faeces)

G: Highest daily grades

Observation performed, sign not present

-Page 11 - 24Oct08 15h46
C.I PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS (GRAM)

MALES

PRE MATING MATING PERIOD

DAYS 1 8 1 8 15 22 29 36

WEEKS 1 2 1 2 3 4 5 6

ANIMAL

GROUP1I(CONTROL)

1 293 322 351 366 386

2 274 288 306 322 339

3 292 320 343 355 373

4 303 313 323 325 342

5 306 325 339 348 366

6 313 339 371 397 421

7 295 323 348 357 368

8 329 350 369 382 398

9 296 315 337 356 372

10 314 350 369 391 411

GROUP 2 (50 MG/KG)

11 308 323 347 355 371

12 279 292 317 333 343

13 298 319 336 345 359

14 288 305 323 331 339

15 306 342 362 369 389

16 292 314 335 343 361

17 322 350 385 339 420

18 296 314 330 340 357

19 297 311 333 341 357

20 322 352 378 392 427

GROUP 3 (250 MGIKG)

21 319 344 371 388 409

22 302 335 353 367 378

23 291 309 327 349 363

24 305 328 350 362 372

25 292 309 330 342 355

26 302 334 360 374 392

27 307 334 357 372 390

28 312 340 388 373 395

29 300 321 345 362 375

30 301 329 359 372 397

GROUP 4 (1000 MGIKG)

31 314 340 359 368 388

32 290 304 327 348 366

33 312 322 340 351 369

34 304 320 332 346 360

35 291 311 334 346 360

36 301 326 352 368 379

37 294 320 343 354 364

38 307 323 340 351 374

39 314 332 343 344 370

40 298 308 325 340 359

-Page 12 - 29SeP08 101121
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS (GRAM)

FEMALES

PRE MATING MATING PERIOD

DAYS 1 8 1 8 15 22 29 36

WEEKS 1 2 1 2 3 4 5 6

ANIMAL

GROUP I (CONTROL)

41 206 211 217

42 182 185 201

43 199 196 218

44 204 212 212

45 194 199 204

46 186 193 200

47 185 191 198

48 193 203 210

49 189 195 202

50 196 198 215

GROUP 2 (50 MG/KG)

51 195 204 209

52 184 189 210

53 198 210 211

54 183 196 206

55 202 214 220

56 191 193 216

57 197 202 210

58 207 217 225

59 190 197 205

60 196 205 207

GROUP 3 1250 MG/KG)

61 196 200 216

62 203 210 215

63 187 195 214

64 194 198 222

65 191 189 211

66 198 204 205

67 185 204 216

68 196 203 213

69 180 191 202

70 196 200 199

GROUP 4 (1000 MG/KG)

71 198 205 203

72 201 203 221

73 186 196 201

74 196 204 220

75 190 193 214

76 183 195 207

78 195 200 203

79 184 187 190

80 195 201 201

- Page 13 - ~29Sep08 10h21
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS (GRAM) FEMALES

FO-GENEIRATION

POST COITUM LACTATION

DAYS 0 4 7 11 14 17 20 1 4

ANIMAL

GROUP I(CONTROL)

41 <NP> 222 223 220 225 237 240 245 ---

42 194 216 226 239 250 277 315 235 243

43 220 239 247 267 280 297 332 270 280

44 222 244 243 257 269 296 330 242 261

45 213 228 233 250 266 287 322 239 261

46 209 221 224 238 244 263 272 235 248

47 200 213 219 230 224 252 280 225 227

48 210 225 238 248 272 291 335 246 256

49 199 210 220 230 243 263 292 223 239

50 215 235 243 259 273 301 346 254 256

GROUP 2 (50 MGIKG)

51 212 229 234 247 262 279 300 243 247

52 206 218 224 238 247 270 298 225 229

53 223 249 262 277 290 316 352 269 276

54 <NP> 207 211 209 209 210 214 222 ---

55 217 232 242 254 264 288 324 239 242

56 208 226 234 251 265 288 325 253 250

57 210 222 227 240 250 271 304 234 230

58 227 243 257 271 287 314 349 265 279

59 207 222 229 239 250 270 299 234 245

60 215 227 232 244 253 271 301 241 253

GROUP 3 (250 MGIKG)

61 213 224 229 244 251 275 306 239 247

62 220 235 244 259 265 272 287 257 260

63 211 235 242 257 270 296 339 252 259

64 208 228 234 245 264 287 318 237 252

65 211 232 243 259 270 297 330 254 261

66 <NP> 217 221 225 221 225 222 225 ---

67 217 239 249 262 280 298 334 260 263

68 210 229 236 253 266 299 324 242 259

69 206 216 225 237 247 266 299 248 243

70 214 234 243 257 269 292 332 246 257

GROUP 4 (1000 MGIKG)

71 213 232 241 258 271 301 338 246 264

72 216 236 245 243 242 243 237 --- 73 198 218 229 238 251 267 295 230 233

74 217 234 242 255 265 283 309 238 247

75 209 222 230 244 263 286 325 232 248

76 205 223 228 244 250 283 316 232 238

77 214 228 238 253 268 288 332 250 260

78 217 231 241 257 259 286 325 233 256

79 200 224 227 245 256 281 317 241 249

80 209 229 239 253 264 286 324 243 261

<NP> Non-pregnant

-Page 14 -I 23Oct08 16IWO
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHT GAIN(%

MALES

PRE MATING MATING PERIOD

DAYS 1 8 1 8 15 2229 36

WEEKS 1 2 1 2 3 4 5 6

ANIMAL

GROUP1I(CONTROL)

1 0 10 20 25 32

2 0 5 12 1824

3 0 10 17 22 28

4 0 3 7 7 13

5 D 6 11 14 20

6 0 8 19 2735

7 0 9 1 821 25

8 0 6 12 1621

9 0 6 14 20 26

10 0 11 18 25 31

GROUP 2 (50 MG/KG)

11 0 5 13 1520

12 0 5 14 1923

13 0 7 13 1620

14 0 6 12 15 18

15 0 12 18 21 27

16 0 8 15 1724

17 0 9 20 5 30

18 0 6 11 15 21

19 0 5 12 15 20

20 0 9 17 22 33

GROUP 3 (250 MG/KG)

21 0 8 16 22 26

22 0 11 17 22 25

23 0 6 12 20 25

24 0 8 15 19 22

25 0 6 13 17 22

26 0 11 19 24 30

27 0 9 16 21 27

28 0 9 18 2027

29 0 7 15 2125

30 0 9 19 24 32

GROUP 4 (1000 MG/KG)

31 0 8 14 1724

32 0 5 13 2026

33 0 3 9 13 16

34 0 5 9 14 18

35 0 7 15 1924

36 0 8 17 22 26

37 0 9 17 20 24

38 0 5 11 14 22

39 0 6 9 10 18

40 0 3 9 14 20

-Page 15 - ~29Sep08 10h2l
C.l. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHT GAIN I(%)

FEMALES

PRE MATING MATING PERIOD

DAYS 1 8 1 8 15 2229 36

WEEKS 1 2 1 2 3 4 5 6

ANIMAL

GROUP I (CONTROL)

41 0 2 5

42 0 2 10

43 0 -2 10

44 0 4 4

45 0 3 5

46 0 4 8

47 0 3 7

48 0 5 9

49 0 3 7

50 0 1 10

GROUP 2 (50 MG/KG)

51 0 5 7

52 0 3 14

53 0 6 7

54 0 7 13

55 0 6 9

56 0 1 13

57 0 3 7

58 0 5 9

59 0 4 8

60 0 5 6

GROUP 3 (250 MG/KG)

61 0 2 10

62 0 3 6

63 0 4 14

64 0 2 14

65 0 -1 10

66 0 3 4

67 0 10 17

68 0 4 9

69 0 6 12

70 0 2 2

GROUP 4 (1000 MG/KG)

71 0 4 3

72 0 1 10

73 0 5 8

74 0 4 12

75 0 2 13

76 0 7 13

77 0 1 11

78 0 3 4

79 0 2 3

80 0 3 3

-Page 16 -I 29Sep08 10h21
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHT GAIN(%

FEMALES

FO-GENERATION

POST COITUM LACTATION

DAYS 0 4 7 11 14 17 20 1 4

ANIMAL

GROUP I (CONTROL)

41 -NP> 0 0 -1 1 7 8 10 ---

42 0 11 16 23 29 43 62 0 3

43 0 9 12 21 27 35 51 0 4

44 0 10 9 16 21 33 49 0 8

45 0 7 9 17 25 35 51 0 9

46 0 6 7 1 4 17 26 30 0 6

47 0 7 10 15 12 26 40 0 1

48 0 7 13 18 30 39 60 0 4

49 0 6 11 16 22 32 47 0 7

50 0 9 13 20 27 40 61 0 1

GROUP 2 (50 MG/KG)

51 0 8 10 17 24 32 42 0 2

52 0 6 9 16 20 31 45 0 2

53 0 12 17 24 30 42 58 0 3

54 <NP> 0 2 1 1 1 3 7 ----

55 0 7 12 17 22 33 49 0 1

56 0 9 13 21 27 38 56 0 -1

57 0 6 8 14 19 29 45 0 -2

56 0 7 13 19 26 38 54 0 5

59 0 7 11 15 21 30 44 0 5

60 0 6 8 13 18 26 40 0 5

GROUP 3 (250 MG/KG)

61 0 5 8 15 18 29 44 0 3

62 0 7 11 18 20 24 30 0 1

63 0 11 15 22 28 40 61 0 3

64 0 10 13 18 27 38 53 0 6 65 0 10 15 23 26 41 56 0 3

66 <NP> 0 2 4 2 4 2 4 ----

67 0 10 15 21 29 37 54 0 1

68 0 9 12 20 27 42 54 0 7

69 0 5 9 15 20 29 45 0 -2

70 0 9 14 20 26 36 55 0 4

GROUP 4 (1000 MGIKG)

71 0 9 13 21 27 41 59 0 7

72 0 9 13 13 12 13 10 ---

73 0 10 16 20 27 35 49 0 1 74 0 8 12 18 22 30 42 0 4

75 0 6 10 17 26 37 56 0 7

76 0 9 11 19 22 38 54 0 3

77 0 7 11 18 25 35 55 0 4

78 0 6 11 18 19 32 50 0 10

79 0 12 14 23 28 41 59 0 3

80 0 10 14 21 26 37 55 0 7

<NP> Non-pregnant

-Page 17- 230ct08 16hl4
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

FOOD CONSUMPTION (G/ANIMAL/DAY) MALES

PRE MATING

DAYS 1-8 8-15

WEEKS 1-2 2-3

CAGE

GROUP I(CONTROL)

1 20 20

2 23 24

GROUP 2 (50 MG/KG)

3 21 22

4 21 22

GROUP 3 (250 MG/KG)

5 22 22

6 22 22

GROUP 4 (1000 MG/KG)

7 21 22

8 22 22

FEMALES

PRE MATING

DAYS 1-8 8-15

WEEKS 1-2 2-3

CAGE

GROUP I(CONTROL)

9 14 14

10 15 15

GROUP 2 (50 MG/KG)

1 1 15 15

12 14 1 4

GROUP 3 (250 MG/KG)

13 14 15

1 4 14 1 4

GROUP 4 (1000 MG/KG)

1 5 15 15

16 14 14

-Page 18- W 29Sep08 10h21
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

FOOD CONSUIMPTION (G/ANIMALJDAY) FEMALES

FO-GENERATION

POST COITUM LACTATION

DAYS 0-4 4-7 7-11 11-14 14-17 17-20 1-4

ANIMAL

GROUP I (CONTROL)

41 <NP' 14 16 16 18 15 17 --

42 1B 20 20 23 21 24 27

43 18 18 21 22 15 26 29

44 18 17 19 18 21 18 41

45 16 17 19 20 21 21 42

46 18 17 19 19 22 19 34

47 15 18 18 15 16 20 20

48 19 22 22 24 22 25 29

49 15 17 17 19 19 19 26

50 18 20 20 24 24 25 29

GROUP 2 (50 MG/KG)

51 16 16 17 20 19 20 21

52 17 16 20 17 19 19 21

53 22 24 15 25 26 29 47

54 <NP> 17 18 16 18 17 21 --

55 15 18 19 4 21 22 23

56 17 19 19 20 19 23 25

57 17 16 18 19 19 19 20

58 20 22 22 25 27 26 33

59 15 15 18 18 20 21 24

60 16 18 18 4 19 20 25

GROUP 3 (250 MG/KG)

61 16 18 18 20 20 21 27

62 16 18 19 19 19 20 13

63 19 21 20 22 21 24 29

64 16 18 18 20 20 21 30

05 17 21 20 25 22 24 30

66 NP> 18 15 17 15 16 15 --

67 20 19 22 23 22 24 29

68 17 17 20 21 24 21 29

69 15 15 17 18 22 21 22

70 20 21 23 35 24 23 28

GROUP 4 (1000 MG/KG)

71 19 19 22 21 24 23 45

72 17 19 18 20 16 15 --

73 16 17 17 19 19 20 20

74 15 19 18 21 17 19 77(3)

75 16 20 21 22 23 23 31

76 17 18 20 20 21 20 23

77 15 16 19 21 19 23 29

78 18 19 21 16 21 20 50

79 18 18 20 21 22 25 24

80 17 18 20 20 21 22 28

(3) Tare error

<NP> Non-pregnant

-Page 19- ' 23Oct08 16h14
C.l. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

RELATIVE FOOD CONSUMPTION (GIKG BODY WEIGHT/DAY)

MALES

PRE MATING

DAYS 1-8 8-15

WEEKS 1-2 2-3

CAGE

GROUP I (CONTROL)

1 63 65

2 68 72

GROUP 2 (50 MG/KG)

3 66 68

4 64 68

GROUP 3 (250 MG/KG)

5 67 69

6 65 67

GROUP 4 (1000 MG/KG)

7 66 68

8 67 68

FEMALES

PRE MATING

DAYS 1-8 8-15

WEEKS 1-2 2-3

CAGE

GROUP I(CONTROL)

9 68 71

10 74 77

GROUP 2 (50 MG/KG)

11 73 72

12 70 70

GROUP 3 (250 MG/KG)

13 73 76

14 68 71

GROUP 4 (1000 MG/KG)

15 73 75

16 72 74

-Page 20 -I'29Sep08 I01121
CI. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

RELATIVE FOOD CONSUMPTION (GIKG BODY WEIGHT/DAY) FEMALES

FO-GENERATION

POST COITUM LACTATION

DAYS 0-4 4-7 7-11 11-14 14-17 17-20 1-4

ANIMAL

GROUP I (CONTROL)

41 <NP> 63 71 69 75 61 71 --

42 83 88 85 92 77 76 il1

43 74 73 79 79 52 79 102

44 73 70 74 67 71 56 158

45 70 73 74 75 72 64 160

46 79 74 78 77 85 69 138

47 70 82 77 67 65 70 90

48 82 92 87 89 77 76 112

49 70 76 75 77 72 64 109

50 76 82 75 89 80 73 113

GROUP 2 (50 MG/KG)

51 68 67 69 78 67 67 84

52 79 70 82 69 72 65 93

53 89 92 55 86 88 81 170

54 <NP> 79 65 78 86 79 93 --

55 65 73 74 16 73 67 95

56 73 80 74 74 67 70 99

57 77 72 73 76 69 64 87

58 81 86 79 88 87 75 117

59 65 67 73 72 75 69 98

60 69 76 74 17 70 65 100

GROUP 3 (250 MG/KG)

61 70 80 73 81 74 69 109

62 68 74 73 70 69 69 49

63 80 87 78 83 72 72 ill

64 70 77 71 77 69 65 119

65 73 85 75 91 75 72 114

66 <NP> 79 65 75 67 71 68 --

67 85 78 85 81 74 71 109

68 74 72 79 79 79 65 112

69 68 65 73 73 81 70 89

70 83 85 89 130 82 69 109

GROUP 4 (1000 MG/KG)

71 82 79 84 79 79 67 169

72 71 78 72 83 64 62 _

73 75 76 69 76 70 68 86

74 65 79 70 81 60 63 312(3) 75 70 87 84 84 79 70 125

76 75 79 80 81 75 63 98

77 67 77 74 77 67 68 112

78 79 79 82 60 73 61 197

79 79 78 81 82 79 78 95

80 75 75 78 77 75 69 107

(3) Tare error

<NP> Non-pregnant

-Page 21 -ON- 23Oct08 l6h14
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

FUNCTIONAL OBSERVATIONS

MALES

AT WEEK 4

ANIMAL HEARING PUPIL L PUPIL R. STATIC R GRIP

SCORE 0/1 SCORE 0/1 SCORE 0/1 SCORE 0/1 SCORE 0/1

GROUP I (CONTROL)

1 0 0 0 0 0

2 0 0 0 0 0

3 0 0 0 0 0

7 0 0 0 0 0

10 0 0 0 0 0

GROUP 2 (50 MG/KG)

1 1 0 0 0 0 0

12 0 0 0 0 0

14 0 0 0 0 0

15 0 0 0 0 0

16 0 0 0 0 0

GROUP 3 (250 MG/KG)

21 0 0 0 0 0

22 0 0 0 0 0

23 0 0 0 0 0

24 0 0 0 0 0

25 0 0 0 0 0

GROUP 4 (1000 MG/KG)

32 0 0 0 0 0

34 0 0 0 0 0

35 0 0 0 0 0

36 0 0 0 0 0

37 0 0 0 0 0

FEMALES

AT WEEK 4

ANIMAL HEARING PUPIL L PUPIL R. STATIC R GRIP

SCORE 0/1 SCORE 0/1 SCORE 0/1 SCORE 0/1 SCORE 0/1

GROUP I(CONTROL)

42 0 0 0 0 0

43 0 0 0 0 0

47 0 0 0 0 0

48 0 0 0 0 0

50 0 0 0 0 0

GROUP 2 (50 MG/KG)

52 0 0 0 0 0

55 0 0 0 0 0

56 0 0 0 0 0

57 0 0 0 0 0

58 0 0 0 0 0

GROUP 3 (250 MG/KG)

61 0 0 0 0 0

63 0 0 0 0 0

64 0 0 0 0 0

65 0 0 0 0 0

67 0 0 0 0 0

- Page 22 -I 08Oct08 12h22
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

FUNCTIONAL OBSERVATIONS

FEMALES

AT WEEK 4

ANIMAL HEARING PUPIL L PUPIL R STATIC R GRIP

SCORE 011 SCORE 011 SCORE 01i SCORE 0/1 SCORE (3/1

GROUP 4 (1000 MGIKG)

73 0 0 0 0 0

74 0 0 0 0 0

75 0 0 0 0 0

76 0 0 0 0 0

77 0 0 0 0 0

-Page 23 00 I O 8Oct08 121122
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

MOTOR ACTIVITY TEST

FEMALES

LACTATION

Counts per sample period (hour)

Animal Location of

number sensor 1 2 3 4 5 6 7 8 9 10 11 12 Total

GROUPI1(CONTROL)

42 High 141 103 228 462 320 46 184 51 29 0 69 285 1918

42 Low 334 472 1615 2749 2437 327 1930 365 406 21 424 2012 13092

43 High 146 123 174 91 183 115 208 96 268 88 131 112 1733

43 Low 1748 3127 3380 1558 1001 1599 2702 669 2753 1002 470 961 20968

47 High 156 246 130 202 524 479 433 119 244 368 222 151 3274

47 Low 1374 1728 648 2188 3095 2183 2361 134 1088 1392 1158 547 17898

48 High 32 401 180 344 385 254 105 19 101 40 22 60 1943

48 Low 258 2528 1325 2677 2385 2915 1823 444 2128 808 537 767 18595

50 High 407 299 272 96 131 303 378 0 112 218 60 46 2322

50 Low 1573 1790 1316 379 583 1780 2408 6 659 1057 439 276 12266

GROUP 2 (50 MG/KG)

52 High 204 210 69 248 285 86 119 56 73 136 11 8 1505

52 Low 970 808 423 1171 1485 1022 627 525 72 421 41 63 7628

55 High 167 132 93 203 331 54 404 24 197 72 136 96 1909

55 Low 1916 1199 776 1136 2139 431 1740 24 1826 767 1469 793 14216

56 High 57 51 1 256 262 532 182 302 61 47 25 1911 3687

56 Low 297 228 10 537 216 1152 72 686 228 235 76 363 4080

57 High 79 218 154 145 54 19 33 56 2 42 0 1 803

57 Low 912 2301 2682 2079 1203 465 866 1216 59 841 0 101 12725

58 High 273 33 228 139 145 232 1 71 104 133 113 211 1683

58 Low 1042 91 694 315 427 981 7 244 118 354 760 552 5485

GROUP 3 (150 MG/KG)

61 High 134 72 110 136 168 44 145 55 0 58 0 36 958

61 Low 1125 686 905 963 1682 1058 1946 438 0 689 0 416 9908

63 High 375 425 228 292 419 194 140 194 182 10 261 84 2804

63 Low 2660 3379 1624 2570 3178 2539 982 1637 985 0 2088 838 22480

64 High 302 65 483 475 271 317 85 32 72 0 0 48 2150

64 Low 1427 676 2245 2209 2020 2461 2276 295 1123 0 19 182 14933

65 High 185 90 184 18 180 79 54 40 0 30 2 43 905

65 Low 259 615 1961 340 2115 1261 1299 1187 5 636 115 889 10682

67 High 317 161 800 512 458 135 519 251 33 348 121 151 3806

67 LOW 1348 821 4852 3478 3423 926 3940 2300 108 1911 748 496 24351

GROUP 4 (500 MG/KG)

73 High 50 288 39 295 156 292 117 64 272 161 115 20 1869

73 Low 1046 1975 649 2403 1090 2282 156 573 1742 492 471 115 12994

74 High 424 117 375 217 168 214 153 94 183 57 167 144 2313

74 Low 2226 1285 3293 3601 1168 1103 1204 411 2937 460 1553 426 '19667

75 High 107 97 0 134 57 191 37 54 54 40 0 4 775

75 Low 1580 1472 22 1700 924 2659 599 1163 1561 1140 0 197 13017

77 High 235 61 71 171 129 126 0 77 47 103 0 122 1142

77 Low 2140 383 1240 2531 2841 2629 0 1547 670 1865 1 1480 17327

76 High 263 97 55 119 102 27 64 63 80 102 26 16 1014

76 Low 3683 3707 2398 3739 2836 1415 2978 2844 2778 1245 1505 383 29511

- Page 25-
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

HAEMATOLOGY

MALES

END OF TREATMENT

ANIMAL WBC Neutrophils Lymphocytes Monocytes Easinophils

10E9/L %WBC %WBC %WBC %WBC

GROUP I (CONTROL)

1 5.2 19.0 76.1 2.2 2.3

2 69 14.1 82.8 1.9 0.7

3 9. 2 13.3 82.6 2.8 1.0

7 6.9 13.2 83.5 2.1 0.9

10 7.4 17.5 78.5 2.1 1.4

GROUP 2 (50 MGIKG)

11 7.3 14.0 83.0 1.6 0.8

12 10.3 11.8 85.6 1.7 0.6

14 8.1 16.0 81.0 2.2 0.5

15 7.9 15.7 802 27 1.2

16 6.7 10.7 85.6 2.6 0.7

GROUP 3 (250 MG1KG)

21 9.5 20.7 74.7 32 1.1

22 7.7 13.6 83.3 2.1 0.8

23 6.5 18.1 77.7 2.7 1.1

24 7.4 14.9 81.9 1 7 0.9

25 10.4 12.4 84.8 1.7 0.7

GROUP 4 (1000 MGIKG) 32 10.0 8.0 89.6 1.6 0.5

34 7.3 12.2 83.7 28 1.0

35 6.3 12.2 83.6 20 1.5

36 7.8 18.2 79.8 1.3 0.6

37 6.7 9.9 876 1.7 0.6

MALES

END OF TREATMENT

ANIMAL Basophils Red blood cells Reticulocytes ROW Haemoglobin

%WBC 10E12/L %RBC %mmol/L

GROUP I (CONTROL)

1 0.3 7.43 2.8 13.2 9.0

2 0.5 8.42 3.3 136 10.1

3 0.2 8.67 2.6 11.8 9.7

7 0.3 8.06 2.7 13.0 9.7

10 0.6 8.06 2.8 11.7 10.1

GROUP 2 (50 MG/KG)

11 0.6 8.09 2.7 13.2 9.6

12 0.4 9.04 2.6 12.7 10.2

14 0.3 9.36 1.9 12.1 10.6

15 0.3 8.24 2.6 12.7 '10.0

16 0.3 8.61 3.8 17.2 9.0

GROUP 3 (250 MG/KG)

21 0.3 9.05 2.6 13.2 10.3

22 0.3 7.93 3.0 13.2 9.6

23 0.4 9.04 2.6 12.6 10.4

24 0.5 8.62 2.3 12.6 9.8

25 0.4 9.17 2.3 12.2 10.3

-Page 26 -29Sep08 llh10
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

HAEMATOLOGY

MALES

END OF TREATMENT

ANIMAL Basophils Red blood cells Reticulocytes ROW Haemoglobin

%WBC 10E12/L %/RBC %mmol/L

GROUP 4 (1000 MG/KG)

32 0.4 8.48 2.8 12.4 9.5

34 0.4 8. 83 2.5 12,9 10.1

35 0.8 8.89 2.3 12.8 9.9

36 0,3 8.25 2.2 23.9 9.9

37 0.1 8.99 2.0 12.9 9.9

MALES

END OF TREATMENT

ANIMAL Haematocril MCV MCH MCHC Platelets

L/L fL. final mmol/L IO0E9IL

GROUP I(CONTROL)

1 0.382 51.5 1.21 23.44 699

2 0.438 52.0 1.20 23.03 985

3 0447 52.2 1.14 21.75 990

7 0421 52.3 1.20 23.00 770

10 0.434 53.8 1.26 23.33 583

GROUP 2 (50 MGIKG)

11 0.418 51.6 1.19 23.10 750

12 0.465 51.4 1.13 21.96 925

14 0.482 51.5 1.13 21.91 906

15 0.437 53.0 1.21 22.84 775

16 0.417 48.4 1.04 21.55 955

GROUP 3 (250 MG/KG)

21 0.468 51.7 1.14 21.95 972

22 0.425 53.5 1.21 22.53 954

23 0.467 51.7 1.15 22.21 792

24 0.442 51.3 1.14 22.17 774

25 0.472 51.5 1.12 21,74 873

GROUP 4 (1000 MGJKG)

32 0.441 52.0 1.12 21.61 988

34 0.460 52.1 1.14 21.92 964

35 0.451 50.7 1.12 22.07 1008

36 0.434 52.6 1.20 22 89 875

37 0.453 50.4 1.11 21.96 1069

MALES

END OF TREATMENT

ANIMAL PT APTT

s s

GROUP I(CONTROL)

2 17.5 19.9

3 19.5 19.4

7 183 188

10 16.2 17.4

-Page 27 -M 29SepO8 IWhO
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

HAEMATOLOGY

MALES

END OF TREATMENT

ANIMAL PT AP'Fr

$ $

GROUP 2 (50 1861KG)

11 1637 18.3

12 1 8.4 21.4

14 19.1 22.6

15 18.4 21.5

16 17.9 19.81

GROUP 3 (250 MG/KG)

21 16.7 16.2

22 17.9 20.7

23 17.6 22.3

24 17.0 16.2

25 17.6 20.7

GROUP 4 (1000 MG/KG)

32 17.4 19.5

34 18.1 17.7

35 18.2 21.8

36 17.0 2 1.7

37 17.1 16.4

FEMALES

END OF TREATMENT

ANIMAL WBC Neutrophils Lymphocytes Monocytes Eosinophils

10E9/L %WBC %WBC %WBC %WBC

GROUP 1 (CONTROL)

42 3.3 29.2 68.2 1.8 0.3

43 4.7 12.0 84.0 3.0 1.0

47 4.3 22.8 73.5 2.0 1.6

48 3.7 21.6 75.3 2.4 0.4

50 3.9 24.0 72.0 4.0 0.0

GROUP 2 (50 MG/KG)

52 3.4 16.3 81.4 1.3 0.8

55 6.1 15.0 82.9 1.6 0.3

56 --- 53.0 450 2.0 0.0

57 3,5 21.0 77.0 2.0 0.0

58 29 25.4 70,9 2.9 0.6

GROUP 3 (250 MG/KG)

61 5.9 30.2 67.5 1.8 0.2

63 5.9 30.6 66.4 2.3 0.3

64 3.3 27.3 69.8 1.7 0.2

65 5.3 23,5 73.4 1.6 1.4

67 2.2 30,0 66.0 3.0 1.0

GROUP 4 (1000 MG/KG)

73 5.2 15.5 81.1 2.5 0.7

74 7.5 39.0 61.0 0.0 0.0

75 5.6 27.0 71.0 1.0 1.0

76 4.6 15.5 82.1 1.3 0.8

77 5.6 27.0 71.0 2.0 0.0

-Page 28 - Wii 29Sep08 lihG
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

HAEMATOLOGY

FEMALES

END OF TREATMENT

ANIMAL Basophils Red blood cells Reticulocytes RIDW Haemoglobin

%WBC IOE12IL %RBC %mmol/L

GROUP I(CONTROL)

42 0.5 7.13 4.2 16.7 9.1

43 0.0 7.34 5.0 14.0 9.1

47 0.2 7.10 4.2 12.3 8.8

48 0,3 7.37 6.4 15.7 8.5

50 0.0 7,50 4.0 13.1 9.0

GROUP 2 (50 MG/KG)

52 0.3 7.42 4.1 12.6 93

55 0.1 7.81 3.6 12.3 9.2

56 00 7.06 7.7 17.8 9.2

57 0.0 7.36 5.9 21.0 9.4

58 0.3 8.29 2.9 14.2 9.3

GROUP 3 (250 MG/KG)

61 0.3 6.85 7.6 17.2 8.4

63 0.4 7.58 4.3 13.7 8.2

64 1.1 7.32 7.9 16.9 8.4

65 0.1 7.39 4.8 14.8 8.9

67 0.0 7.48 3.9 16.3 9.0

GROUP 4 (1000 MGIKG)

73 0.2 7.91 4.3 12.1 9.3

74 0.0 7.66 5.7 14.9 8.8

75 0.0 750 5.2 14.9 9.1

76 0.2 7.30 4.7 13.5 9.3

77 0.0 7.75 6.6 14.8 9.3

FEMALES

END OF TREATMENT

ANIMAL Haematocrit MGV MCH MCHC Platelets

L/L fL fmol mmol/L I10E9/L

GROUP I(CONTROL)

42 0.377 529 1.27 24.01 1102

43 0.396 539 1.24 22.97 1127

47 0.386 54.4 1.24 22.83 652

48 0.380 51.6 1.16 22.45 1540

s0 0.389 51 9 1.21 23.21 1166

GROUP 2 (50 MG/KG)

52 0.397 53.6 1.26 23.50 1372

55 0.414 530 1.18 22.28 1328

56 0.395 55.9 1.30 23.30 1443

67 0405 55.1 1.28 23.17 1197

58 0.415 50.1 1.12 22.43 1262

GROUP 3 (250 MG/KG)

61 0.361 52.7 1.22 23.18 1660

63 0.399 52.7 1.21 22.93 1312

64 0.389 53.2 1.15 21.64 1670

65 0.388 52.5 1.21 22.94 1273

67 0.391 52.2 1.21 23.12 1022

-Page 29 - .29Sep08 111110
C.l. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

HAEMATOLOGY

FEMALES

END OF TREATMENT

ANIMAL Haematocrit MCV MCH MCHC Platelets

UJL fl. find mmoIIL 1OE9IL

GROUP 4 (1000 MG/KG)

73 0.425 53.7 1.18 21.99 1399

74 0.393 51.3 1.14 22.32 861

75 0.395 52.6 1.21 23.02 1363

76 0.396 54.2 1.27 23.44 1517

77 0.414 53.5 1.20 22.42 835

FEMALES

END OF TREATMENT

ANIMAL PT APTT

S

GROUP I(CONTROL)

42 17.7 19.4

43 16.8 16.0

47----

48 16.7 19.9

50 17.2 15.4

GROUP 2 (50 MVG/KG)

52 16.8 22.0

55 17.4 21.9

56 16.1 14.8

57 15.9 16.9

58 16.4 162

GROUP 3 (260 MG/KG)

61 16.6 18.8

63 16.2 16.0

64 17.3 17.0

65 15.8 14.9

67 15.5 15.5

GROUP 4 (1000 MG/KG)

73 16.4 21.9

74 15.6 15.5

75 15.6 14.9

76 15.3 13,3

77 15.1 14.0

-Page 30 - 29Sep08 IlhIO
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL BIOCHEMISTRY

MALES

END OF TREATMENT

ANIMAL ALAT ASAT ALP Total protein Albumin

U/L U/L U/L g/L g/L

GROUP 1 (CONTROL)

1 57.1 71.1 203 65.9 34.4

2 39.8 79.7 140 64.6 34.4

3 45.0 72.8 135 62.2 34.0

7 38.2 74.1 185 64.7 32.8

10 43.3 111.7 105 58.9 31.3

GROUP 2 (50 MG/KG)

11 40.9 85.1 197 61.7 32.6

12 56.3 79.3 187 68.0 34.5

14 65.5 77.7 195 65.3 33.6

15 33.6 73.2 142 65.6 33.5

16 63.6 99.4 142 65.3 34.3

GROUP 3 (250 MG/KG)

21 40.8 78.3 185 68.3 34.1

22 46.2 72.7 151 65.5 33.9

23 58.2 87.4 126 65.6 34.2

24 43,4 78.8 154 624 33.3

25 45.6 81.4 135 65.4 34.7

GROUP 4 (1000 MG/KG)

32 47.1 82.2 120 64.9 33.8

34 43.5 85.7 116 65.8 34.4

35 44.5 84.0 188 65.5 34.0

36 45.5 73.5 120 66.4 34.5

37 58.6 87.8 169 68.2 34.8

MALES

END OF TREATMENT

ANIMAL Total bitirubin Urea Creatinine Glucose Cholesterol

umol/L mmol/L umol/L mmolIL mmol/L

GROUP I(CONTROL)

1 2.6 6.7 42.1 7.05 1.48

2 1.9 6.4 40.6 10.52 1.54

3 2.3 11.9 43.6 8.38 1.44

7 2.1 9.0 45.1 7.50 2.20

10 2.0 9.9 42.8 6.89 1.59

GROUP 2 (50 MG/KG)

11 2.0 8.4 47.3 6.23 1.84

12 2.1 10.2 39.8 8.03 2.16

14 2.6 11.2 48.1 10.20 1.83

15 2.4 8.2 39.1 6.93 2.22

16 2.8 9.3 55.6 6.07 1.81

GROUP 3 (250 MG/KG)

21 1.8 8.7 43.6 6.84 2.12

22 2.6 8.1 42.1 7.29 2.21

23 2.4 9.4 46.6 8.31 1.60

24 2.2 7.9 45.1 5.96 1.75

25 2.1 8.7 54.1 8.19 1.73

-Page 31 ~ 29SepO81lhIO
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL BIOCHEMISTRY

MALES

END OF TREATMENT

ANIMAL Total bilirubin Urea Creatinine Glucose Cholesterol

umol/L mmol/L umol/L mmol/L mmol/L

GROUP 4 (11000 MG/KG)

32 2.1 7.8 43.6 9.76 2.10

34 2.1 8.3 44.3 6.36 1.72

35 2.0 8.2 41.3 761 1.92

36 1.8 10.3 44.3 6.15 2.33

37 1.8 7.6 41.3 6.11 2.57

MALES

END OF TREATMENT

ANIMAL Sodium Potassium Chloride Calcium lnorg.Phos

mmol/L mmol/L mmol/L mmol/L mmol/L

GROUP 1 (CONTROL) 1 1 42.1 3.94 104 2.80 2.05

2 142.0 3.39 105 2.81 1.75

3 142.0 3.82 104 2.73 2.18

7 142.9 3.74 104 2.83 2.03

10 142.4 3.94 105 2.75 2.32

GROUP 2 (50 MG/KG)

11 142.0 388 104 2.64 1.94

12 142.1 4.22 104 2.76 2.08

14 142.5 4.18 104 2.66 1.71

15 142.6 3.88 104 2.73 210

16 140.9 3.97 103 2.74 2.40

GROUP 3 (250 MG/KG)

21 143.4 3.91 104 2.82 2.40

22 1420 3.83 103 2.69 216

23 142.1 3.71 105 2.77 2.08

24 1421 3.80 105 2.61 2.10

25 143.5 379 106 2.76 2.15

GROUP 4 (1000 MG/KG)

32 142.6 4.25 104 2.69 1.94

34 143.5 3.88 104 2.77 2.08

35 148.2 3.90 104 2.72 2.21

36 141.9 4.37 108 2.77 2.09

37 142.6 3.77 105 2.72 1.88

FEMALES

END OF TREATMENT

ANIMAL ALAT ASAT ALP Total protein Albumin

U/L U/L U/L g1L g/L

GROUP I (CONTROL)

42 62.8 85.7 56 64.6 32.3

43 45.6 85.0 42 65.0 32.2

47 64.8 95.1 102 66.6 32.7

48 43.5 73.9 60 67.1 34.0

50 73.6 64.8 64 657 31.7

- Page 32- 29Sep08 11h10
C.I PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL BIOCHEMISTRY

FEMALES

END OF TREATMENT

ANIMAL ALAT ASAT ALP Total protein Albumin

U/L U/L U/L g/L gIL

GROUP 2 (50 MG/KG)

52 67.2 78.5 47 69.1 33.6

55 45.0 58.4 92 63.7 30.2

56 68.3 76.0 70 64.0 32.6

57 59.7 71.3 61 67.5 34.7

58 98.7 91.5 91 62.1 295

GROUP 3 (250 MG/KG)

61 72.4 74.6 86 69.2 35.4

63 587 83.2 61 682 34.1

64 77.5 762 87 67.3 32.5

65 65.1 70.1 76 64.1 32.8

67 59.4 96.3 39 69.5 34.1

GROUP 4 (1000 MGKG)

73 36.8 62.5 36 643 31.7

74 42.3 76.9 113 62.6 31.8

75 55.4 72.1 120 665 32.6

76 46.0 73.6 44 68.1 34 0

77 48.5 65.9 31 66.5 33.2

FEMALES

END OF TREATMENT

ANIMAL Total bilirubin Urea Creatinine Glucose Cholesterol

umol/L mmolIL umolIL mmol/l. mmolIL

GROUP I(CONTROL)

42 3.0 7.8 46.5 6.56 1.35

43 2.1 9.3 50.0 495 1.14

47 20 11.3 45.4 8.26 1.58

48 2.3 7.1 41.7 5.63 1.48

50 24 11.4 50.0 7.02 1.69

GROUP 2 (60 MG/KG)

52 2.4 9.7 47.6 6.46 2.09

55 15 95 48.3 4.65 1.48

56 2.1 10.1 45.8 5.69 1.40

57 1.9 9.2 45.4 5.06 1.49

58 2.3 12.2 49.0 7.74 1.98

GROUP 3 (250 MG/KG)

61 2.8 9.8 45.1 5.24 1.67

63 2.0 9.4 40.2 5.79 1.51

64 2.8 11.5 46.5 5.16 1 60

65 3.1 9.5 47.2 4.49 1.87

67 1.7 9.7 54.2 6.69 1.59

GROUP 4 (1000 MG/KG)

73 1.6 11.4 49.8 5.10 1.65

74 1.5 9.9 50.0 5.94 1.33

75 1.7 9.3 50.0 5.23 2.24

76 2.3 9.8 47.4 7.02 1.80

77 2.0 8.0 43.0 5.77 1.71

-Page 33 - 29Sep08 llhlO
ClI. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CLINICAL BIOCHEMISTRY

FEMALES

END OF TREATMENT

ANIMAL Sodium Potassium Chloride Calcium lnorg.Phos

mnmol/L mmol/L mmol/L mmol/L mmol/L

GROUP I(CONTROL)

42 142.2 364 96 2.76 264

43 144.3 356 100 2.73 2 .35

47 139.2 3.95 102 2.80 1.91

48 140.5 3.42 98 2.78 1.92

50 141.5 3.34 98 2.82 2.54

GROUP 2 (50 MG/KG)

52 141.2 363 98 2.70 1.71

55 141.9 400 101 2.82 2.61

56 141.2 368 100 272 2.27

57 141.0 3.72 97 2.82 2.29

58 139.5 3.93 99 2.69 2.26

GROUP 3 (250 MG/KG)

61 140.3 3.49 96 2.88 2.63

63 140.1 3.65 97 291 247

64 142.5 3.57 98 2.81 3.02

65 140.4 3.98 98 2.71 2.49

67 142.5 3.66 97 2.72 2.33

GROUP 4 (1000 MG/KG)

73 141.0 3.73 99 2.75 2.61

74 141.2 3.14 98 2.67 2.31

75 141.7 3.38 95 2.76 2.33

76 140.4 4.34 98 2.75 2.10

77 141.7 3.45 95 2.77 2.18

-Page 34 - ~29Sep08 llh10
ClI. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

MACROSCOPIC FINDINGS

MALES

ALL NECROPSIES

ANIMAL ORGAN FINDING DAY OF DEATH

GROUP I(CONTROL) 1 No findings noted Scheduled sacrifice, 17Jul2008

2 No findings noted Scheduled sacrifice. 17JuI2008

3 Bone Tail: tail bent. Scheduled sacrifice, 17Jul2008

4 No findings noted Scheduled sacrifice. 17JuI2008

6 No findings noted Scheduled sacrifice, M1u2008

6 No findings noted Scheduled sacrifice. 17Jul2008

7 No findings noted Scheduled sacrifice, 17Jul2008

8 No findings noted Scheduled sacrifice, 17Jul2008

g No findings noted Scheduled sacrifice, 17,u02008

10 No findings noted Scheduled sacrifice, 17Jul2008

GROUP 2 (50 MG/KG)

I1I No findings noted Scheduled sacrifice, 1 7Jul2008

12 Mesenteric Inode Discolouration, greenish. Scheduled sacrifice, 17Jul2008

13 No findings noted Scheduled sacrifice, 1 7Jul2008

14 No findings noted Scheduled sacrifice, 17Jul2008

15 Skin Tail: discolouration, bluish. Scheduled sacrifice, 17Jul2008

16 Mesenteric Lnode Discolouration, greenish. Scheduled sacrifice, 17Jul2008

17 No findings noted Scheduled sacrifice, 17Jul2008

18 No findings noted Scheduled sacrifice, 17,102008

19 No findings noted Scheduled sacrifice, 17Jul2008

20 No findings noted Scheduled sacrifice, 1 7Jul2008

GROUP 3 (250 MG/KG)

21 Mesenteric Inode Discolouration, greenish. Scheduled sacrifice, 17Jul2008

22 Epididymides Right side, tall: nodule(s), d=7x5 mm, Scheduled sacrifice, 17Jul2008

Yellowish, hard.

Mesentenc Inode Discolouration, greenish.

23 Mesenteric Lnode Discolouralion, greenish. Scheduled sacrifice, 17Jul2008

24 Mesenteric Inode Discolouration, greenish. Scheduled sacrifice, 1 7JuI2008

25 Mesenteric Inode Discolouration, greenish. Scheduled sacrifice, 17Jul2008

26 No findings noted Scheduled sacrifice. 17Jul2008

27 Epidldymides Right side, tall: nodule(s), d=2x2 mm, Scheduled sacrifice, 17Jul2008

Yellowish, hard.

Mesenteric Inode Discolouration, greenish.

28 Mesenteric Inode Discolouration, greenish. Scheduled sacrifice, 1 7Jui2008

29 Mesenteric Inode Discolouration, greenish. Scheduled sacrifice, 17Jul2008

30 Mesenteric Inode Discolouration, greenish. Scheduled sacrifice, 17Jul2008

GROUP 4 (1000 MG/KG)

31 Mesenteric Lnode Discolouration, greenish. Scheduled sacrifice, 17Jul2008

32 Mesenteric Lnode Discolouration, greenish. Scheduled sacrifice, 17,102008

33 Mesenteric Lnode Discolouration, greenish. Scheduled sacrifice, 1 7Jul2008

34 Mesenteric I.node Discolouration, greenish. Scheduled sacrifice, 17Jul2008

Skin Foreleg, left side: alopecia

35 Mesenteric Inode Discolouration, greenish. Scheduled sacrifice, 17Jul2008

36 Mesenteric Lnode Discolouralion, greenish. Scheduled sacrifice, 1 7Jul2008

37 Mesenteric Inode Discolouration, greenish. Scheduled sacrifice, 17Jut2008

38 Mesenteric Lnode Discolouration, greenish. Scheduled sacrifice, 17Jul2008

39 Mesenteric Inode Discolouration, greenish. Scheduled sacrifice, 17Jul2008

Skin Tall: dIscolouration, bluish.

40 Seminal vesicles Left side: reduced in size. Scheduled sacrifice, 1 7Jul2008

Mesenteric Lnode Discolouralion, greenish.

-Page 35 011- 29Sep08 11110
02. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

MACROSCOPIC FINDINGS

FEMALES

ALL NECROPSIES

ANIMAL ORGAN FINDING DAY OF DEATH

GROUP I (CONTROL)

41 No findings noted Scheduled sacrifice, 29Jul2008

42 No findings noted Scheduled sacrifice, 28Jul2008

43 No findings noted Scheduled sacrifice, 28Jul2006

44 No findings noted Scheduled sacrifice, OlAug2008 45 No findings noted Scheduled sacrifice, OlAug2008

46 No findings noted Scheduled sacrifice, OlAug2008

47 Mandibular node Both sides: discolouration, dark red. Scheduled sacrifice, 30Jul2008

Skin Foreleg, left side: alopecia.

Abdominal region: alopecia.

Foreleg, right side: alopecia.

Flank, right side: alopecia.

Flank, left side. alopecia.

48 No findings noted Scheduled sacrifice, 3OJuI2008

49 No findings noted Scheduled sacrifice, 30Jul2008

50 No findings noted Scheduled sacrifice, 28Jul2008

GROUP 2 (So MG/KG)

51 No findings noted Scheduled sacrifice, 31Jul2008

52 Mandibular inode Both sides: discolouration, reddish. Scheduled sacrifice, 30Jul2008

53 No findings noted Scheduled sacrifice, OlAug2008

54 No findings noted Scheduled sacrifice, 29Jut2008

55 No findings noted Scheduled sacrifice, 30JuI2008

56 Pancreas Discolouration, reddish. Scheduled sacrifice, 28Jul2008

Mesenteric Inode Discolouration, bluish

Mandibular Lnode Both sides: discolouration, dark red.

57 No findings noted Scheduled sacrifice, 30Jul2008

58 No findings noted Scheduled sacrifice, 30Jul2008

59 No findings noted Scheduled sacrifice, 30Jul2008

60 No findings noted Scheduled sacrifice, 30Jul2008

GROUP 3 (250 MG/KG) 61 Mesenteric tLnode Discolouration, bluish. Scheduled sacrifice, 28Jul2008

62 No findings noted Scheduled sacrifice 31Jul2008

63 Mesenteric Inode Discolouration, bluish. Scheduled sacrifice, 28Jut2008

64 Mesenteric Inode Discolouration, bluish. Scheduled sacrifice, 28Jul2008

65 Mesenteric Lnode Discolouration, bluish. Scheduled sacrifice, 28JuI2008

66 No findings noted Scheduled sacrifice, 29Jul2008

67 Mesenteric Inode Discolouration, bluish. Scheduled sacrifice, 30JuI2008

68 Mesenteric Inode Discolouration, bluish. Scheduled sacrifice, 30Jut2008

69 Mesenteric Inode Discolouration, bluish. Scheduled sacrifice, 30Jul2008

70 Mesenteric Inode Discolouration, bluish. Scheduled sacrifice, 31Jul2008

GROUP 4 (1000 MG/KG)

71 General observationsG~i-tractus, contents: bluish. Scheduled sacrifice, OlAug2008

Mesenteric Lnode Discolouration, bluish.

Skin Foreleg, right side: alopecia. Hindleg, left side: alopecia.

Hindleg, right side: alopecla.

Foreleg, left side. alopecia.

72 General observations Gl-tractus, contents: bluish. Scheduled sacrifice, 29Jul2008

73 General observations Gi-tractus, contents: bluish. Scheduled sacrifice, 30Jul2008

Mesenteric Lnode Discolouration, bluish.

74 General observations Gi-tractus, contents: bluish. Scheduled sacrifice, 28Jut2008

Adrenal glands Both sides: discolouration, red-brown.

Mesenteric Lnode Discolouration, bluish.

Mandibular Lnode Both sides: discolouration, reddish.

75 Mesenteric Inode Discolouration, bluish. Scheduled sacrifice, 28Jut2008

76 General observations Gi-tra ctus, contents: bluish. Scheduled sacrifice, 29JuI2008

Mesenteric Inode Discolouration, bluish.

77 Mesenteric Inode Discolouration, bluish. Scheduled sacrifice, 28JuI2008

78 General observations G. i-tra ctus, contents: bluish. Scheduled sacrifice, 01Aug2008 Mesenteric Inode Discolouration, bluish.

79 Mesenteric Inode Discolouration, bluish. Scheduled sacrifice, 31Jul2008

-Page 36 -I'29Sep08 lihiC
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

MACROSCOPIC FINDINGS

FEMALES

ALL NECROPSIES

ANIMAL ORGAN FINDING DAY OF DEATH

GROUP 4 (1000 MGIKG) 80 General observatlonsGi-tractus, contents: bluish. Scheduled sacrifice, 31 Jul2008

Mesenteric Inode Discolouration, bluish.

-Page 37 111110 29Sep081lIMO
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

ORGAN WEIGHTS (GRAM)

MALES

NECROPSY

ANIMAL BODY W. BRAIN HEART LIVER THYMUS

(GRAM) (GRAM) (GRAM) (GRAM) (GRAM)

GROUPlI(CONTROL)

1 361 2.06 0.918 9.21 0.361

2 316 2.01 0.914 8.02 0.370

3 354 2.07 0,961 9.51 0.435

4 323 1.95 0.872 725 0.262

5 344 2.09 0.881 8.23 0.346

6 401----- -

7 345------- -

8 373------- -

9 346 ---------

10 383 ---------

GROUP 2 (50 MG/KG)

11 341 1.99 1.019 7.54 0.279

12 332 2.04 0.966 781 0.444

13 337 1.97 0.918 8.47 0.361

14 320 2.03 0.880 7.38 0.283

15 365 2.06 0.930 9.90 0.477

16 335----- -

17 398----- -

18 336----- -

19 335 ---------

20 395 ------ ---

GROUP 3 (260 MG/KG)

21 379 2.11 1 091 10.70 0.425

22 361 1.95 1.055 9.34 0434

23 340 2.08 0889 8.66 0.370

24 347 1.98 0.987 8.33 0.400

25 325 1.97 0.933 8.14 0.343

26 364----

27 369------- -

28 371 -- - ---

29 351------- -

30 371------- -

GROUP 4 (1000 MG/KG)

31 363 2.13 1.066 10.23 0.327

32 348 2.08 1.119 9.92 0.407

33 351 2.04 0.962 8.74 C.337

34 340 2.10 0.936 8.16 0.287

35 342 2.03 0.941 9.10 0.347

36 352 ---------

37 343------- -

38 350 --- ------

39 348------- -

40 341 --- --- ---

-Page 38 -. 29Sep08 IlhIO
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

ORGAN WEIGHTS (GRAM)

MALES

NECROPSY

ANIMAL KIDNEYS ADRENALS SPLEEN TESTES EPIDIDYMIDES

(GRAM) (GRAM) (GRAM) (GRAM) (GRAM)

GROUP I (CONTROL)

1 2.48 0.069 0.647 3.42 1.165

2 2.43 0.061 0.504 3.32 1.254

3 2.46 0.077 0.709 3.43 1.238

4 2.34 0.082 0.434 3.45 1.163

5 2.43 0.066 0.503 3.67 1.153

6 - -- -- -3.30 1.001

7 - -- -- -3.08 1.027

8 -- - -- 3.36 1.187

9 - -- -- -3.29 1.050

10 - -- -- -3.29 1.153

GROUP 2 (50 MG/KG)

11 2.54 0.079 0.626 3.88 1.193

12 2.39 0,064 0586 3.47 1.067

13 246 0.071 0.540 2.60 0.969

14 2.21 0.062 0.495 3.40 1.206

15 2.42 0.073 0.703 324 1 095

16 --- -- -2.66 0.951

17 - -- -- -3.67 1.177

18 - -- -- -3.21 1.183

19 - -- --- 364 1.281

20 - -- -- -3.06 1.261

GROUP 3 (250 MG/KG)

21 2.54 0.065 0.695 3.71 1.219

22 2.36 0.059 0.663 3.54 1.443

23 2.36 0.070 0.647 3.35 1.232

24 2.39 0.070 0.632 3.24 1.011

25 2.10 0.063 0.496 236 1.079

26 - -- --- 3.33 1.078

27 - -- -- -3.29 1.323

28 - -- -- -3.40 1.195

29 -- 3----.47 1.283

30----------

GROUP 4 (1000 MG/KG)

31 265 0.097 0.710 3.70 1.418

32 2.33 0.067 0.659 3.16 1.061

33 2.60 0.068 0.688 3.70 1.148

34 2.32 0.065 0.677 3.24 1.190

35 2.17 0.077 0.625 3.38 1.165

36 - -- -- -3.47 1.195

37 - -- -- -3.78 1.266

38 - -- -- -2.61 0.959

39 - -- -- -3.46 1.273

40 - -- -- -3.44 1.143

Page 39- 29Sep08illI
o i. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

ORGANIBODY WEIGHT RATIOS I(%)

MALES

NECROPSY

ANIMAL BODY W. BRAIN HEART LIVER THYMUS

(GRAM) (%) (%)(%()

GROUP I (CONTROL)

1 361 0.57 0.254 2.55 0.100

2 316 0.64 0.289 2.54 0.117

3 354 0.58 0.271 2.69 0.123

4 323 0.60 0.270 2.24 0.081

5 344 0.61 0.256 2.39 0.101

6 401--- -

7 345------- -

8 373------- -

9 346------- -

10 363------- -

GROUP 2 (60 MG/KG)

11 341 0.58 0.299 2.21 0.082

12 332 0.61 0.291 2.35 0.134

1 3 337 0.58 0.272 2 51 0.107

14 320 0.63 0.275 2 .31 0.088

15 365 0.56 0.255 2.71 0.131

16 335----- -

17 398----- -

18 336------- -

19 335------- -

20 395 - - - -

GROUP 3 (250 MG/KG)

21 379 056 0.288 2.82 0.112

22 361 0.54 0.292 2.59 0.120

23 340 0.61 0.261 2.55 0.109

24 347 0.57 0.284 2.40 0.115

25 325 0.60 0.287 2.50 0.106

26 364----- -

27 369------- -

28 371------- -

29 351------- -

30 371------- -

GROUP 4 (1000 MG/KG)

31 363 0.59 0294 2 82 0.090

32 348 060 0.322 2.85 0.117

33 351 0.58 0.274 2.49 0.096

34 340 0.62 0.275 2.40 0.084

35 342 0.59 0.275 2.66 0.101

36 352------- -

37 343------- -

38 350----- -

39 348------- -

40 341----- -

-Page 40 - lI 29Sep08 llhIO
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

ORGANIBODY WEIGHT RATIOS(% MALES

NECROPSY

ANIMAL KIDNEYS ADRENALS SPLEEN TESTES EPIDIDYMIDES

GROUPI1(CONTROL)

1 0.69 0.019 0.179 0.95 0.323

2 0.77 0.0 16 0.1659 1.05 0.397

3 0.70 0.022 0.200 0.97 0.350

4 0.72 0.025 0.134 1.07 0.360

5 0.71 0.019 0.146 1.07 0.335

6 - -- -- -0.82 0.250

7 - -- -- -0.89 0.298

8 - -- -- -0.90 0.318

9 ---- 0.95 0.303

10 - -- -- -0.86 0.301

GROUP 2 (50 MG/KG)

11 0.74 0.023 0.184 1.14 0.350

12 0.72 0.019 0.177 1.05 0321

13 0.73 0.021 0.160 0.77 0.288

14 0.69 0.016 0 155 1.06 0.377

15 0.66 0.020 0.193 0.89 0300

16 - -- -- -0.79 0.284

17 - ---- -0.92 0.296

18 - -- -- -0.95 0.352

19 - -- -- -1.09 0.382

20 - ----- 0.77 0.319

GROUP 3,(250 MG/KG)

21 0.67 0.017 0.183 0.98 0.322

22 0.65 0.018 0.184 0.98 0.400

23 0.69 0,021 0.190 0.99 0.362

24 0.69 0,020 0.182 093 0.291

25 065 0.019 0.153 0.73 0.332

26 - ----- 0.92 0.296

27 - ----- 0.89 0.359

28 ----- -0.92 0.322

29 ------ 0.99 0.366

30----------

GROUP 4 (1000 MG/KG)

31 0.73 0.027 0.196 1.02 0.391

32 0.67 0.019 0.189 0.91 0.305

33 0.74 0.019 0.196 1.06 0.327

34 0.68 0.019 0.199 0.95 0.350

35 0.63 0.023 0.183 0.99 0.341

36 ------ 0.98 0.339

37 ------ 1.10 0.369

38 ----- -0.74 0.274

39 ------ 099 0.366

40 - ----- 1.01 0.335

-Page 41 295ep08llh10
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

ORGAN WEIGHTS (GRAM)

FEMALES

NECROPSY

ANIMAL BODY W. BRAIN HEART LIVER THYMUS

(GRAM) (GRAM) (GRAM) (GRAM) (GRAM)

GROUP1I(CONTROL)

41 ------ - -

42 243 1.82 0.661 7.00 0.118

43 240 1,81 0.786 7.25 0.237

44--------

45----------

46----------

47 201 1.86 0.650 6.48 0.133

48 220 1.80 0.806 7.66 0,198

49----

50 222 1.93 0.762 7.77 0.115

GROUP 2 (50 MGIKG)

51 ---------- -

52 202 1.76 0,755 6.29 0.151

53 --

54 ---------- -

55 220 1.79 0.731 673 0161

56 223 1.85 0.727 7.54 0.207

57 207 1 83 0.634 6.59 0.162

58 248 1,76 0.780 8 18 0.167

59 --- ------- -

60 --- ------- -

GROUP 3 (250 MGIKG)

61 225 1.85 0.749 8.17 0.202

62----------

63 230 1.87 0,752 8.12 0.145

64 227 1.78 0.774 7.46 0.168

65 230 1.76 0.847 7.72 0.180

66----------

67 230 1.90 0.802 7.59 0.158

68 --- ---------

69----------

70 --- ----

GROUP 4 (1000 MG/KG)

71----------

72--------

73 211 1.87 0.667 6.14 0.182

74 247 1.84 0.740 7.32 0.175

75 214 1.86 0.756 7.19 0.152

76 212 1.87 0.672 7.01 0.221

77 220 1.85 0.745 7.89 0.165

78 - -- --- --- -- -

79----------

80 - -- -- -- --

-Page 42 -29Sep08 IlhlO
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

ORGAN WEIGHTS (GRAM)

FEMALES

NECROPSY

ANIMAL KIDNEYS ADRENALS SPLEEN

(GRAM) (GRAM) (GRAM)

GROUP I(CONTROL) 41------

42 1.58 0.078 0.452

43 1.50 0.077 0.496

44---- -

45----

46---- -

47 1.49 0.072 0.312

48 2.01 0.084 0.548

49 -----

50 1.64 0.084 0.477

GROUP 2 (50 MG/KG)

51 --- -

52 1.65 0.075 0391

53------

54----

55 1.62 0.083 0.475

56 1.62 0.097 0.526

57 1.53 0.076 0.502

58 1.70 0.088 0.415

59

60------

GROUP 3 (250 MG/KG)

61 1.69 0.077 0,693

62------

63 1.88 0.096 0.479

64 1 70 0.080 0.543

65 1.71 0076 0.497

66 -- - -

67 1.71 0.079 0.478

68------

69------

70------

GROUP 4 (1000 MG/KG)

71----

72------

73 1.44 0.065 0400

74 1.48 0.100 0.501

75 1.72 0.088 0535

76 1.68 0.089 0.510

77 1.81 0.098 0.559

78 -----

79------

80------

-Page 43 - 29Sep08 llhlO
C.!. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

ORGAN/BODY WEIGHT RATIOS I(%)

FEMALES

NECROPSY

ANIMAL BODY W. BRAIN HEART LIVER THYMUS

(GRAM) (%(%(%()

GROUP I (CONTROL)

41--------

42 243 0.75 0.272 2.88 0.049

43 240 0.75 0.328 3.02 0.099

44 --- ------- -

45----------

46 --- ---------

47 201 0.93 0.323 3.22 0.066

48 220 0.82 0.366 3.48 0.090

49 --- ----- -

50 222 0.87 0,343 3.50 0.052

GROUP 2 (50 MG/KG)

53 -----

54 --- ---------

55 220 0.81 0.332 3.06 0.073

56 223 0.83 0.326 3.38 0.093

57 207 0.88 0.306 3.18 0.078

58 248 0.71 0.315 3.30 0.067

59 ---------- -

60----------

GROUP 3 (250 MG/KG)

61 225 0.82 0.333 3.63 0.090

62 ---------- -

63 230 0.81 0.327 3.63 0.063

64 227 0.78 0.341 3.29 0.074

65 230 0.77 0.368 3.36 0.078

66--------

67 230 083 0.349 3.30 0.069

68 ---------- -

69 -------- -

70----------

GROUP 4 (1000 MG/KG)

71 ---------- -

72----------

73 211 0.89 0.316 2Z91 0.086

74 247 0.75 0.300 2.96 0.071

75 214 0.87 0.353 3.36 0.071

76 2 12 0.88 0.317 3.31 0104

77 220 0.84 0.339 3.59 0075

78 ---------- -

79 ---------- -

80 -------- -

-Page 44 - *I- 2 9SepOS llhlO
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

ORGAN/BODY WEIGHT RATIOS(%

FEMALES

NECROPSY

ANIMAL KIDNEYS ADRENALS SPLEEN

GROUP 1 (CONTROL)

41------

42 0.65 0.032 0.186

43 0.63 0.032 0.207

44------

45--

46------

47 0.74 0.035 0.155

48 0.91 0.038 0.249

49 -- - -

50 0.74 0.038 0.215

GROUP 2 (50 MG/KG)

51------

52 0.82 0.037 0.194

53 --

54 --

55 0.74 0.038 0.216

56 0.73 0.043 0 236

57 0.74 0 037 0.243

58 0.66 0,035 0.167

59------

60----

GROUP 3 (250 MG/KG)

61 0.75 0.034 0 308

62------

63 0.82 0.042 0.208

64 0.75 0,035 0.239

65 0.74 0.033 0.216

66 --- ----

67 0.74 0.034 0.208

68------

69------

70 -------

GROUP 4 (1000 MG/KG)

71 --

72------

73 0.68 0.031 0.190

74 0.60 0.040 0.203

75 0.81 0.041 0.250

76 0.79 0.042 0.241

77 0.82 0.045 0.254

78------

79 --- ---

80 -------

-Page 45 - 29Sep08 llhJ0
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BREEDING DATA PER LITTER

FO-GENERATION - LACTATION

-- FIRST LITTER CHECKLITTER DURATION OF DEAD PUPS LIVING PUPS P.NATAL LOSS

GESTATION M F M F TOT M F

GROUP 1 (CONTROL)

42 21 0 0 2 10 12 0 0

43 21 0 0 4 6 10 0 0

44 22 0 0 10 5 15 0 0

45 22 0 0 4 9 13 0 0

46 22 0 0 1 3 4 0 0

47 21 0 0 3 7 10 0 0

48 2 1 0 0 6 8 1 4 0 0

49 21 0 0 5 7 12 0 1

50 21 0 0 11 3 14 0 0

TOTAL 0 0 46 58 104 0 1

N 9 9 9 9 9 9 9 9

MEAN 21.3 0.0 0.0 5.1 6.4 11.6 0.0 0.1

ST.DEV. 0.5 0.0 0.0 3.4 2.5 3.3 0.0 0.3

-Page 46 - -23Oct08 10h12
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BREEDING DATA PER LITTER

FO-GENERATION - LACTATION

-- FIRST LITTER CHECK

LITTER DURATION OF DEAD PUPS LIVING PUPS P.NATAL LOSS

GESTATION M F M F TOT. M F

GROUP 2 (50 MG/KG)

51 22 0 0 5 4 9 0 1

52 21 0 0 4 7 11 0 0

53 22 0 0 5 6 11 0 0

55 21 0 0 7 6 13 0 0

56 21 0 0 8 6 14 0 0

57 21 0 0 4 7 11 0 0

58 21 0 0 7 6 13 0 0

59 21 0 0 4 6 10 0 0

60 21 0 0 4 7 11 0 0

TOTAL 0 0 48 55 103 0 1

N 9 9 9 9 9 9 9 9

MEAN 21.2 0.0 0.0 53 6.1 11.4 0.0 0.1

ST.DEV. 0.4 0.0 0.0 1.6 0 9 1 6 0.0 0.3

-Page 47 -'"23Oct08 161112
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BREEDING DATA PER LITTER

FO-GENERATION - LACTATION

-- FIRST LITTER CHECKLITTER DURATION OF DEAD PUPS LIVING PUPS P.NATAL LOSS

GESTATION M F M F TOT. M F

GROUP 3 (250 MGIKG)

61 21 0 0 5 6 11 0 0

62 22 0 0 2 0 2 0 0

63 21 0 0 9 6 15 0 0

64 21 0 0 4 9 13 0 0

65 21 0 0 7 7 14 0 0

67 21 0 0 6 5 11 0 0

68 21 0 1 6 6 12 0 0

69 21 0 0 4 5 9 0 0

70 21 0 0 7 7 14 0 0

TOTAL 0 1 50 51 101 0 0

N 9 9 9 9 9 9 9 9

MEAN 21.1 0.0 01 56 5.7 11.2 0.0 0.0

ST.DEV 0.3 00 0.3 2.1 2.4 39 0.0 0.0

-Page 46 8" 23Oct08 161112
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BREEDING DATA PER LITTER

FO-GENERATION - LACTATION

-- FIRST LITTER CHECKLITTER DURATION OF DEAD PUPS LIVING PUPS P.NATAL LOSS

GESTATION M F M F TOT. M F

GROUP 4 (1000 MG/KG)

71 22 0 0 7 7 14 0 0

73 21 1 0 3 6 9 0 0

74 21 0 0 6 6 12 0 0

75 21 0 0 5 10 15 0 0

76 21 0 0 6 7 13 1 0

77 21 0 0 8 4 12 0 0

78 22 1 0 9 5 1 4 0 0

79 21 0 0 5 8 13 0 1

80 21 0 0 8 4 1 2 0 0

TOTAL 2 0 57 57 114 1 1

N 9 9 9 9 9 9 9 9

MEAN 21.2 0.2 0.0 6.3 6.3 12.7 0.1 0.1

ST.DEV. 0.4 0.4 0.0 1.9 1.9 1.7 0.3 0.3

-Page 49 -23Oct08 10h12
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

MEAN BODY WEIGHTS OF PUPS PER LITTER (GRAM)

FO-GENERATION - LACTATION

LITTER SEX DAYl1 DAY 4

GROUP I(CONTROL)

42 M 6.7 8.6

F 6.0 8.2

M+F 6.1 8.2

43 M 6.3 9.3

F 5.9 8.6

M+F 6.1 89

44 M 5.6 7.9

F 5.7 7.8

M+ F 5.6 7.9

45 M 6.1 8.5

F 6.0 8.7

M+F 6.0 8.7

46 M 6.9 10.7

F 6.9 10.6

M+F 6.9 10.7

47 M 5.3 7.5

F 51 7.3

M+F 5.1 7.4

48 M 5.8 8.3

F 5.5 7.9

M+F 5.6 8.0

49 M 5.2 8.1

F 5.2 8.2

M+F 5.2 8.2

50 M 62 8.5

F 5.9 8.0

M+F 6.1 8.4

GROUP 2 (50 MG/KG)

51 M 7.0 11.1

F 6.1 10.2

M+F 6.6 10.7

52 M 5.8 8.4

F 5.7 8.3

M+F 5.8 8.3

53 M 7.6 10.8

F 7.3 1 0.3

M+F 7.5 10.5

55 M 5.6 8.5

F 5.5 8.5

M+F 5.6 8.5

56 M 6.3 8.1

F 5.9 7.5

M+F 6.1 7.8

57 M 5.8 8.6

F 5.4 8.2

M+F 5.5 8.3

58 M 5.8 8.7

F 5.3 8.1

M+F 5.6 8.4

59 M 5.8 8.7

F 5.4 82

M+F 5.5 8.4

-Pages 50~ 29Sop08l1h3i
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

MEAN BODY WEIGHTS OF PUPS PER LITTER (GRAM)

FO-GENERATION - LACTATION

LITTER SEX DAYlI DAY 4

GROUP 2 (50 MG/KG)

60 M 56 8.6

F 5.0 7.8

M+F 5.2 8.1

GROUP 3 (250 MGIKG)

61 M 5.9 8.3

F 5.5 7.6

M+F 5.7 7.9

62 M 6.6 10.3

M+F 8.6 10.3

63 M 5.9 8.3

F 5.4 7.5

M+tF 5.7 8.0

64 M 5.8 7.9

F 5.0 7.2

M+F 5.2 7.4

65 M 5.6 8.0

F 5.6 8.2

M+F 5.6 8.1

67 M 6.8 10.3

F 6.2 9.8

M+F 65 10.1

68 M 6.0 9.4

F 5.5 9.3

M+F 59 9.3

69 M 6.1 9.5

F 5.7 9.0

M+F 5.9 9.2

70 M 6.0 8.8

F 5.4 7.9

M+F 5.7 8.3

GROUP 4 (1000 MG/KG)

71 M 6.4 9.1

F 6.2 8.7

M+F 6.3 8.9

73 M 6.0 9.1

fF 5.8 9.1

M+F 5.9 9.1

74 M 5.4 7.5

F 5.1 7.1

M+F 5.2 7.3

75 M 5.5 7.7

F 5.3 7.4

M+F 5.3 7.5

76 M 5.8 8.1

F 5.4 7.8

M+F 5.6 8.0

77 M 6.1 8.6

F 5.8 8.3

M+F 6.0 8.5

78 M 6.4 9.2

F 64 9.2

M+F 6.4 9.2

-Page 51 - ~29Sep08 11h31
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

MEAN BODY WEIGHTS OF PUPS PER LITTER (GRAM)

FO-GENERATION - LACTATION

LITTER SEX DAYl1 DAY 4

GROUP 4 (1000 MG/KG)

79 M 5.6 80

F 5.2 7 .5

M+F 5.4 7.,7

80 m 5.8 8.9

F 5.8 8.5

M+F 5.7 5.8

-Page 52 00- 29Sep08 11h31
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS OF PUPS (GRAM)

FO-GENERZATION - LACTATION

LITTER PUP SEX DAY 1 DAY 4

GROUP 1 (CONTROL)

42 1 M 67 8.6

2 M 6.7 8.7

3 F 5.7 7.9

4 F 6.0 8.3

5 F 6.1 8.4

6 F 5.5 7.4

7 F 6.2 86

8 F 6.9 9.2

9 F 6.0 8.1

10 F 6.0 8.1

11 F 5.6 7.6

12 F 6.0 8.0

43 1 M 6.6 9.6

2 M 6.2 9,1

3 M 6.4 9.5

4 M 6.0 8.6

5 F 6.0 9.0

6 F 5.5 7.9

7 F 6.1 8.6

a F 6.7 8.4

9 F 6.3 9.3

10 F 5.8 8.6

44 1 M 6.0 60

2 M 6.1 8. 6

3 M 6.0 8.2

4 M 5.3 7.7

5 M 5.6 8.1

6 M 5.5 7.8

7 M 5.3 7.6

8 M 5.6 8.3

9 M 4.8 6.6

10o M 5.7 8.3

11 F 5.4 7.5

12 F 6.0 8.3

13 F 5.7 7.4

14 F 5.7 7.7

15 F 5.6 7.9

45 1 M 5.6 7.8

2 M 6.6 8.9

3 M 5.7 8.3

4 M 6.5 9.0

5 F 6.5 9.3

6 F 6.0 8.7

7 F 5.8 8.5

8 F 6.0 9.1

9 F 65 9.3

10 F 6.1 8.6

11 F 6.3 8.6

12 F 5.5 8.2

13 F 5.6 7.9

46 1 M 6,9 10.7

2 F 6.6 10.9

3 F 69 10.8

4 F 6.9 10.3

- Page 53-- 29Sep08 11h31
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS OF PUPS (GRAM)

FO-GENERATION - LACTATION

LITTER PUP SEX DAY 1 DAY 4

GROUP I (CONTROL)

47 1 M 5.4 7.8

2 M 5.1 7.3

3 M 5.3 7.3

4 F 5.3 7.8

5 F 4.7 67

6 F 6.1 7.3

7 F 5.0 7.5

8 F 5.2 7.5

9 F 5.4 7.7

10 F 4.8 6.9

48 1 M 5.8 8.3

2 M 6.2 8.7

3 M 6.1 8.8

4 M 5.7 7.9

5 M 5.4 7.9

6 M 5.5 8.2

7 F 5.5 7.9

8 F 5.3 7.2

9 F 5.8 7.9

10 F 5.6 8.2

11 F 5.2 7.7

12 F 5.6 7.9

13 F 5.6 7.8

14 F 5.5 8.2

49 1 M 5.4 8.8

2 M 4.4 7.0

3 M 5.5 8.3

4 M 5.4 8.5

5 M 5.3 7.8

6 F 45 --

7 F 5.3 8.4

8 F 5.2 8.5

9 F 5.3 7.8

10 F 5.4 8.3

11 F 5.1 8.2

12 F 5.4 8.1

50 1 M 6.1 8.6

2 M 5.8 8.0

3 M 6.4 8.8

4 M 6.9 9.8

5 M 6.2 8.5

6 M 62 8.2

7 M 6.2 8.4

8 M 5.9 8.0

9 M 5.7 7.4

1 0 M 6.6 8.6

11 M 6.0 7.8

12 F 6.5 8.8

13 F 8.4 8.6

14 F 5.0 6.7

GROUP 2 150 MGIKG)

51 1 M 7.1 11.3

2 M 6.9 10.9

3 M 7.6 11.6

4 M 6.7 10.6

5 M 7.0 11.0

6 F 6.1 9.2

7 F 6.8 10.8

8 F 4.8 --

9 F 6.7 10.5

-Page 54 - -29SepO8 I Ih3l
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS OF PUPS (GRAM)

FO-GENERATION - LACTATION

LITTER PUP SEX DAY I DAY 4

GROUP 2 (50 MGIKG)

52 1 M 5.9 8.7

2 M 5.6 8.3

3 M 6.2 8.7

4 M 5.6 6.0

5 F 5.7 8.4

6 F 5.9 83

7 F 58 8. 4

8 F 5 .7 8 .3

9 F 5 .3 8 .1

10 F 5.8 8.3

11 F 5.8 8.4

53 1 M 7.5 10.5

2 M 8.0 116

3 M 7.0 99

4 M 7.8 10.9

5 M 7.7 1 1.1

6 F 7.8 11.0

7 F 7.7 10.3

8 F 7.1 10.3

9 F 7.2 10.4

10 F 7.3 10.0

11 F 7.0 9.9

55 1 M 5.5 8.6

2 M 6.2 9.4

3 M 5.9 8.8

4 M 5.3 7.9

6 M 5.4 8.4

6 M 5.4 8.2

7 M 5.4 8.1

8 F 5.3 8.5

9 F 5.8 8.5

10 F 5.3 8.5

11 F 5.6 8.4

12 F 5.6 8.5

13 F 56 8.6

56 1 M 62 8.3

2 M 6.3 7.9

3 M 6.2 7.9

4 M 6.3 80

5 M 6.1 83

6 M 6.4 8.1

7 M 6.0 8.1

8 M 6.5 8.1

9 F 5.9 8.0

10 F 6.2 7.9

11 F 62 7.7

12 F 5.9 7.3

13 F 5.7 7.4

14 F 5.4 6.6

57 1 M 5.2 80

2 M 6.2 9.1

3 F 5.6 8.2

4 F 4.7 7.0

5 F 5.7 8.6

6 F 5.9 9.0

7 F 5.0 7.8

8 F 5.6 8.4

9 F 5.3 8.1

10 M 61 8.8

11 M 57 8.4

- Page 55- 29Sep08 11ih31
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS OF PUPS (GRAM)

FO-GENERATION - LACTATION

LITTER PUP SEX DAYl1 DAY 4

GROUP 2 (50 MGIKG)

58 1 M 6.0 9.1

2 M 5.7 8.8

3 M 5.9 8.8

4 M 51 8.0

5 M 6.2 9.2

6 M 5.8 8.3

7 M 6.0 9.0

8 F 5.7 8.8

9 F 5.4 8.4

10 F 5.2 7.6

11 F 5.2 7.9

1 2 F 5.4 8.2

13 F 5,2 7.6

59 1 M 5.7 8.8

2 M 6.0 8.8

3 M 5.6 8.4

4 M 6.0 9.0

5 F 5.9 90

6 F 5.2 8.0

7 F 4.9 7. 7

8 F 5.4 8. 2

9 F 5.8 8.8

10 F 5.2 7.8

60 1 M 6.2 9.2

2 M 5.6 8.9

3 M 5.1 7.9

4 M 5.5 8.5

5 F 5.6 8.8

6 F 5.2 8.1

7 F 5.3 8.2

8 F 4.8 7.3

9 F 4.1 6.4

10 F 4.5 7.5

11 F 5.3 8.4

GROUP 3 (250 MOIKG)

61 1 M 5.8 8.4

2 M 5.9 7.8

3 M 5.8 8.1

4 M 58 8.2

5 M 64 8.7

6 F 6.1 8.0

7 F 5.5 7.6

8 F 5.3 7.4

9 F 53 7.4

10 F 5.6 7.9

11 F 5.2 7.3

62 1 M 6.5 10.4

2 M 6.6 10.3

63 1 M 5.9 8.8

2 M 6.1 7.7

3 M 6.4 8.6

4 M 6.0 8.7

5 M 5.6 7.7

6 M 5.9 8.2

7 M 5.9 8.3

8 M 5.7 8.6

9 M 5.8 6.2

10 F 5.6 8.7

1 1 F 5.3 71

12 F 5.5 7. 7

13 F 4.9 6.4

-Page 56- 29Sep05 11h31
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS OF PUPS (GRAM)

FO-GENERATION - LACTATION

LITTER PUP SEX DAY I DAY 4

GROUP 3 (250 MG/KG)

14 F 5.5 74

15 F 5.4 7.5

64 1 M 5.7 8.4

2 M 5.4 7.2

3 M 5.9 8.0

4 M 6.1 8.1

5 F 4.1 5.3

6 F 4.9 7.0

7 F 5.1 6.8

8 F 5.6 8.0

9 F 5.2 7.6

10 F 4.9 7.2

I1I F 5.8 8.5

12 F 4.9 7.0

13 F 4.5 6.4

65 1 M 5.5 81

2 M 5.0 7.1

3 M 4.6 63

4 M 5.9 8.7

5 M 6.0 8.6

6 M 6.0 8.3

7 M 6.2 8.9

8 F 5.5 8.4

9 F 5.8 8.6

10 F 5.4 8.0

11 F 5.9 8.6

12 F 5.5 8.1

13 F 5.2 7.7

14 F 5.7 8.2

67 1 M 7.0 10.6

2 M 6.9 10.4

3 M 6.7 10.3

4 M 7.0 10.5

5 M 6.5 10.1

6 M 64 99

7 F 6.5 10.3

8 F 5.9 9.4

9 F 6.0 9.5

10 F 65 10.0

11 F 6.1 9.7

68 1 M 6.2 9.7

2 M 5.9 9.5

3 M 6.2 9.7

4 M 6.0 9.0

5 M 6.0 9.4

6 M 5.8 89

7 F 6.8 9.2

8 F 5.8 9.3

9 F 5.8 9.2

10 F 5.7 9.3

11 F 6.2 9.8

12 F 5.8 8.8

13 F -- -

-Page 57 - 29Sep08 111131
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS OF PUPS (GRAM)

FO-GENERATION - LACTATION

LITTER PUP SEX DAYl1 DAY 4

GROUP 3 (250 MG/KG)

69 1 M 6.0 9.5

2 M 5.8 94

3 M 6.5 9.5

4 M 6.1 9.5

5 F 5.8 9.0

6 F 5.7 9.1

7 F 5.8 9.1

8 F 6.1 9.7

9 F 5.2 8.2

70 1 M 6.1 9.4

2 M 6.0 8.2

3 M 6.1 8.7

4 M 6.1 6.8

5 M 5.6 6.9

6 M 5.7 8.5

7 M 6.2 8.9

8 F 5.8 8.5

9 F 5.7 8.1

10 F 5.5 8.3

11 F 5.0 7.4

12 F 5.7 8.3

13 F 5.1 7.6

14 F 4.8 6.9

GROUP 4 (1000 MG/KG)

71 1 M 6.5 9.4

2 M 6.3 9.1

3 M 6.2 8.9

4 M 6.5 91

5 M 6.6 91

6 M 6.7 9.4

7 M 6.1 8.9

8 F 6.0 6.3

9 F 6.5 9.2

10 F 6.5 9.3

11 F 5.4 7.7

12 F 6.8 9.3

13 F 6.1 8.5

14 F 6.4 8.9

73 1 M 6.1 9.9

2 M 6.1 8.9

3 M 5.7 6.5

5 F 5.6 8.7

6 F 5.8 9.5

7 F 5.6 8.5

a F 5.9 9.4

9 F 5.9 9,4

10 F 6.0 9.4

74 1 M 5.4 8.0

2 M 4.8 7.1

3 M 5.8 7.7

4 M 5.6 7.7

5 M 5.1 7.3

6 M 5.5 7.4

7 F 5.7 7.8

6 F 5.2 72

9 F 5.5 8.0

1 0 F 5.3 7.1

11 F 4.0 5.6

12 F 5.1 7.0

-Page 58 -l* M 9ep06 11h31
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS OF PUPS (GRAM)

FO-GENERATION - LACTATION

LITTER PUP SEX DAYl1 DAY 4

GROUP 4 (1000 MGIKG)

75 1 M 5.8 7.9

2 M 4.9 7.0

3 M 50 7.2

4 M 59 8.0

5 M 6.1 8.4

6 F 4 .9 6.2

7 F 5.3 7.8

8 F 5.6 7.7

9 F 5.6 7.9

10 F 55 7.8

1 1 F 5.1 73

12 F 5.3 7.5

13 F 5.3 78

14 F 4.9 6 .5

15 F 54 7.3

76 1 M 62 8.4

2 M 61 87

3 M 5.7 --

4 M 5.4 7.9

5 M 5.3 6.8

6 M 6.3 8.9

7 F 5.2 7.5

8 F 5.3 7.8

9 F 5.3 7.6

10 F 5.5 8.1

11 F 5.7 8.6

12 F 5.5 7.5

13 F 5.2 7.6

77 1 M 5.9 8.9

2 M 6.0 ,8.2

3 M 5.7 8.2

4 M 6.1 8.9

5 M 5.9 6.4

6 M 6.4 8.8

7 M 6.3 8.8

8 M 64 9.0

9 F 5.6 82

10 F 5.8 8.4

1 1 F 5.9 8.4

12 F 5.7 8.0

78 1 M 7.0 10.1

2 M 6.4 9.3

4 M 6.2 9.0

5 M 6.4 9.4

6 M 5.9 8.5

7 M 6.3 8.7

8 M 5.8 8.9

9 M 66 9.7

1 0 M -- -

11 F 6.5 9.2

12 F 6.7 9.7

13 F 6.1 8.6

14 F 6.4 9.6

15 F 6.3 8.8

79 1 M 5.5 8.2

2 M 5.7 7.8

3 M 5.4 7.8

4 M 5.7 7.8

5 M 5.6 8.2

6 F 5.1 7.0

7 F 5.1 7.8

8 F 5.6 8.1

- Page 59 29Sep08 llh~l
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

BODY WEIGHTS OF PUPS (GRAM)

FO-GENERATION - LACTATION

LITTER PUP SEX DAY I DAY 4

GROUP 41(1000 MGIKG)

9 F 5.1 7.4

10 F 5.0 7.1

1 1 F 5.2 7.3

12 F 5.5 7.8

13 F 5.1 --

80 1 M 6.3 9.6

2 M 5.5 8.2

3 M 6.2 9.5

4 M 5.4 8.2

5 M 5.7 9.3

6 M 6.0 9.1

7 M 6.0 8.9

8 M 5.4 8.3

9 F 5.5 8.7

10 F 5.6 8.6

1 1 F 5.8 8.4

12 F 5.6 8.4

-Page 60 - - ep0811h3l
C.!. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP I (CONTROL)

LITTER 42 1 M DAY 5 Planned Necropsy FLC No findings

23JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings 6 F DAY 5 Planned Necropsy FLC No findings

LLC No findings MACRO No findings

7 F DAY 5 Planned Necropsy FILC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

g F DAY 5 Planned Necropsy FLC No findings

LLC No findings

10 F MACRO No findings

10FDAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

LITTER 43 1 M DAY 5 Planned Necropsy FLC No findings

23JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 F DAY 5 Planned Necropsy FLC No findings

LLC No findings MACRO No findings

6 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 F DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK, DAY P P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

-Page 61 - "24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP I (CONTROL)

9 F DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

LITTER 44 1 M DAY 5 Planned Necropsy FLC No findings

27JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

S M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

1D M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

13 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

14 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

15 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

LITTER 45 1 M DAY 5 Planned Necropsy PLC No findings

27JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 62 - ON- 24Oct08 15h47
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP I (CONTROL) 4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 P DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

10 P DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

12 P DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

13 P DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 46 1 M DAY 5 Planned Necropsy PLC No findings

27JUL2008 LLC No findings

MACRO No findings

2 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 F DAY 5 Planned Necropsy PLC No findings

LLC No findings MACRO No findings

4 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 47 1 M DAY 5 Planned Necropsy PLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

5 P DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 P DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

PLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 63 - 24O0t08 15h47
ClI. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP I (CONTROL)

7 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY S Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

LITTER 48 1 M DAY 5 Planned Necropsy FLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRD No findings

5 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 M DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 P DAY 5 Planned Necropsy PLC No findinga

LLC No findings

MACRO No findings

8 F DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

I1 P DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

12 P DAY 5 Planned Necropsy PLC No findings

LLC No findings MACRO No findings

13 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

14 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

LITTER 49 1 M DAY 5 Planned Necropsy PLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

PLC - PIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 64 - 24Oct08 15h47
ClI. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P P. PHASE FINDINGS

DELIVERY

GROUP I (CONTROL) 3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 F DAY 2 Missing FLC No findings

7 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAYS5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

11 F DAYS5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

LITTER 50 1 M DAY 5 Planned Necropsy FLC No findings

23JUL2008 LLC No findings MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRD No findings

10 M DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 65 -. 240cl08 15h47
C. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP1I(CONTROL)

12 F DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

13 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

14 F DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

GROUP 2 (50 MG/KG)

LITTER 51 1 M DAY 5 Planned Necropsy PLC No findings

26JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

8 F DAY 2 Missing PLC Small

9 F DAY 5 Planned Necropsy PLC No findings

LLC No findings MACRO Ho findings

LITTER 52 1 M DAY 5 Planned Necropsy PLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings 5 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

PLC - FIRST LITTER CHECK, DAY P.P, - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 66 - 0111 24O0t08 15h47
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 2 (50 MG/KG)

9 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 53 1 M DAY S Planned Necropsy PLC No findings

27JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings 3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

S M DAY S Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 P DAY S Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 F DAY S Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

9 P DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

10 P DAY S Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

11 F DAY S Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 55 1 M DAYS5 Planned Necropsy PLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

S M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 M DAYS Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

PLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS ,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

-Page 67 - ~I 24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 2 (50 MG/KG)

7 M DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

13 F DAY 5 Planned Necropsy FL-C No findings

LLC No findings

MACRO No findings

LITTER 56 1 M DAY 5 Planned Necropsy FLC No findings

23JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 M DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

8 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy PLC No findings

LLC: No findings

MACRO No findings

10 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

11 P DAY 5 Planned Necropsy PLC; No findings

LLC No findings

MACRO No findings

12 P DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

13 P DAY 5 Planned Necropsy PLC No findings

LLC: No findings

MACRO No findings

PLC - FIRST LITTER CHECK. DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK. MACRO - MACROSCOPIC FINDINGS

- Page 68 - *- 24Oct08 15h47
ClI. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 2 (50 MG)KG)

14 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 57 1 M DAY 5 Planned Necropsy FLC Red spot nose

25JUL2008 DAY 2 Red spot nose

LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 P DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

8 F DAY S Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

10 M DAYS5 Planned Necropsy PLC No findings

LLC No findings MACRO No findings

11 M DAY 5 Planned Necropsy PLC Red spot nose

DAY 2 Red spot nose

LLC No findings

MACRO No findings

LITTER 58 1 M DAYS5 Planned Necropsy PLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings MACRO No findings

4 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

PLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 69 - b-24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 2 (50 MGIKG) 8 F DAY 5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

9 F DAY 5 Planned Nerropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

13 F DAYS5 Planned Necropsy FLC .No findings

LLC No findings

MACRO No findings

LITTER 59 1 M DAY 5 Planned Necropsy FLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 F DAY 5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

6 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings 9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 60 1 M DAYS5 Planned Necropsy FLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 70 - ~"24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

P0-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 2 (50 MG/KG)

5 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 P DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 F DAYS5 Planned Necropsy FLC No findings

LLC No findings MACRO No findings

9 F DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

GROUP 3 (250 MG/KG)

LITTER 61 1 M DAY 5 Planned Necropsy PLC No findings

23JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings 4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 P DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRD No findings

8 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 F DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

11 F DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 62 1 M DAY 5 Planned Necropsy PLC No findings

26JUL2008 LLC No findings

MACRO No findings

PLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 71 - Ir24Oct08 15h47
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF PP. PHASE FINDINGS

DELIVERY

GROUP 3 (250 MG/KG)

2 M DAY 5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

LITTER 63 1 M DAY 5 Planned Necropsy PLC; No findings

23JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC: No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 M DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

g M DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLC Na findings

LLC No findings

MACRO No findings

13 F DAYS5 Planned Necropsy FLC No findings

LLC Na findings

MACRO No findings

14 F DAYS5 Planned Necropsy FLC Na findings

LLC No findings

MACRO No findings

15 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

LITTER 64 1 M DAY 5 Planned Necropsy FLC No findings

23JUL2008 LLC Na findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC Na findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy PLC Na findings

LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 72 -fr24Oct08 15h,47
ClI. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 3 (250 MG/KG)

5 F DAY 5 Planned Necropsy FLC Small

DAY 2 Sma

DAY 3 Smal

DAY 4 Small

LLC Small

MACRO No findings

6 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Nec-ropsy FLC No findings

LLC No findings MACRO No findings

8 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

13 F DAY 5 Planned Necropsy FLC No findings

LLC Na findings

MACRO No findings

LITTER 65 1 M DAY 5 Planned Necropsy FLC No findings

23JUL2008 LLC No findings

MACRO No findings

2 M DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

PLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 73 - 0- 24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

UITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 3 (250 MG/KG)

11 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

13 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

14 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 67 1 M DAY 5 Planned Necropsy PLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy PLC No findings

LLC No findings MACRO No findings

10 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 68 1 M DAY 5 Planned Necropsy PLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

PLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 74 - '24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P PHASE FINDINGS

DELIVERY

GROUP 3 (250 MGIKG)

6 MDAY Plnne Neropy LLC No findings

6 MDAYS Pannd Ncrosy LC No findings

MACRO No findings

7 P DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11I F DAY 5 Planned Nacropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

13 F DAY 1 Dead at FLC PLC Dead

MACRO Autolysis, no milk

LITTER 69 1 M DAY 5 Planned Necropsy FLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy PLC: No findings

LLC No findings

MACRO No findings

5 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 F DAY 5 Planned Necropsy PLC No findings LLC No findings

MACRD No findings

7 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 P DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 70 1 M DAY 5 Planned Necropsy FLC No findings

26JUL2008 LLC: No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

-Page 75 - 11111- 24Oct05 15h47
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 3 (250 MG/KG)

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy FLC; No findings

LLC No findings MACRO No findings

6 M DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 M DAY 5 Planned Necropsy FLC: No findings

LLC No findings

MACRO No findings

8 F DAY S Planned Necropsy FLC No findings

LLC No findings

MACRO No findings 9 F DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAYS Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

13 F DAYS Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

14 F DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

GROUP 4 (1000 MG/KG)

LITTER 71 1 M DAY 5 Planned Necropsy FLC No findings

27JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy FLC No findings

LLC No findings MACRO No findings

6 M DAY S Planned Necropsy FLC No findings

LLC No findings MACRO No findings

7 M DAY 5 Planned Necropsy FLO: No findings

LLC No findings

MACRO No findings

8 F DAY S Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS.

LLC - LAST LITTER CHECK, MACRD - MACROSCOPIC FINDINGS

- Page 76 - -24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 4 (1000 MGIKG)

9 F DAY 5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAYS5 Planned Necropsy FLC: No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

13 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

14 F DAYS5 Planned Necropsy FLC Pale

LLC No findings

MACRO No findings

LITTER 73 1 M DAYS5 Planned Necropsy PLC No findings

25JUL2008 LLC No findings

MACRO No findings

2 M DAYS Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAY 1 Dead at FLC PLC Dead

MACRO Autolysis, no milk

5 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Necropsy FLC: No findings

LLC No findings

MACRO No findings

8 F DAYS5 Planned Necropsy FLC; No findings

LLC No findings

MACRO No findings

g F DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

LITTER 74 1 M DAY S Planned Necropsy FLC No findings

23JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAYS5 Planned Necropsy FLC No findings LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 77 - ~-24Oct08 15h47
0,1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 4 (1000 MG)KG) 5 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAYS5 Planned Necropey PLC No findings

LLC No findings

MACRO No findings

11 F DAY 5 Planned Necropsy FLC Small

DAY 2 Small

DAY 3 Small

DAY 4 Smell

LLC Small

MACRO No findings

12 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 75 1 M DAY 5 Planned Necropsy PLC No findings

23JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings 5 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy PLC No findings LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy PLC No findings

LLC No findings MACRO No findings

10 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

11 F DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

PLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK. MACRO - MACROSCOPIC FINDINGS

- Page 78 - 24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 4 (1000 MGIKG)

12 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

13 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

14 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

15 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

LITTER 76 1 M DAY 5 Planned Necropsy PLC No findings

24JUL2008 LLC No findings MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 2 Missing PLC No findings

4 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

5 M4 DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

7 F DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

10 F DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

11 P DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

13 P DAYS5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

LITTER 77 1 M DAY 5 Planned Necropsy PLC No findings

23JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy PLC No findings

LLC No findings

MACRO No findings

PLC - FIRST LITTER CHECK, DAY P P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

-Page 79 - No 24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO0-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

G ROU P 4 (1000 M GIKG)

5 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings 6 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 M DAY 5 Planned Necropsy FLC Red spot head

DAY 2 Red spot head

LLC Red spot head

MACRO No Findings

8 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings MACRO No findings

11 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

LITTER 78 1 M DAY 5 Planned Necropsy FLC No findings

27JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy FLC No Findings

LLC No findings

MACRO No findings

7 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 M DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

10 M DAYl1 Dead at FLC FLC Dead

MACRO Autolysis, no milk

11 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 50 - '24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 4 (1000 MG/KG) 13 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

14 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

15 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

LITTER 79 1 M DAY 5 Planned Necropsy FLO No findings

26JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

5 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

7 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAYS5 Planned Necropsy FLC No findings

LLG No findings

MACRO No findings

11 F DAY 5 Planned Necropsy FLO No findings

LLC No findings

MACRO No findings

12 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

13 F DAY 2 Spontaneous death FLC No findings

LLC Dead

MACRO Autolyse

LITTER 80 1 M DAY 5 Planned Necropsy FLO No findings

26JUL2008 LLC No findings

MACRO No findings

2 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

3 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

4 M DAY 5 Planned Nacropsy FLC No findings

LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK. DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

-Page 81 - 24Oct08 15h47
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

INDIVIDUAL PUP DATA

FO-GENERATION - LACTATION

LITTER PUP END OF P.P. PHASE FINDINGS

DELIVERY

GROUP 4 (1000 MG/KG) 5 M DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

6 M DAY 5 Planned Necropsy FLC No findings

LLC No findings MACRO No findings

7 M DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

8 M DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

9 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

10 F DAY 5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

11 F DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

12 F DAYS5 Planned Necropsy FLC No findings

LLC No findings

MACRO No findings

FLC - FIRST LITTER CHECK, DAY P.P. - CLINICAL SIGNS,

LLC - LAST LITTER CHECK, MACRO - MACROSCOPIC FINDINGS

- Page 82 ~-24Oct08 15h47
0.1. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

CORPORA LUTEA

FO-GENERATION

GROUP 1 GROUP2 GROUP 3 GROUP4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

Female Corpora Female Corpora Female Corpora Female Corpora

Number Lutea Number Lutea Number Lutea Number Luta

41 <NP> -- 51 15 61 16 71 22

42 15 52 21 62 11 72 2

43 11 53 15 63 17 73 14

44 15 54<NP> -- 64 15 74 15

45 22 55 19 65 15 75 19

46 22 56 15 66<NP> -- 76 15

47 20 57 12 67 21 77 12

48 23 58 18 68 20 78 16

49 19 59 19 69 13 79 13

50 15 60 20 70 13 80 15

Mean 18.0 17.1 15.7 14.3

St.dev. 4.15 2.96 3.28 5.21

N 9 9 9 10

<NP> non-pregnant

DI unnett-test based on pooled variance significant at 5% ()or 1% ()level

- Page 83 -
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

IMPLANTATION SITES

FO-GENERATION

GROUP 1 GROUP 2 GROUP 3 GROUP 4

CONTROL 50 MG/KG 250 MG/KG 1000 MG/KG

Female Implantation Female Implantation Female Implantation Female Implantation

Number Sites Number Sites Number Sites Number Sites

41 <NP> -- 51 8 61 12 71 16

42 12 52 10 62 2 72 1

43 11 53 11 63 16 73 13

4415 54 <NP> --- 64 14 74 10

45 14 55 12 65 13 75 15

46 7 56 14 66<NP> --- 76 14

47 14 57 12 67 12 77 11

48 14 58 13 68 12 78 16

49 12 59 10 69 11 79 13

50 14 60 11 70 13 80 12

Mean 12.6 11.2 11.7 12.1

St.dev. 2.46 1.79 3.91 4.38

N 9 9 9 10

<NP> non-pregnant

1Dunnett-test based on pooled variance significant at 5% ()or 1 % ()level

- Page 84 -
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 2

KEY TO MISSING VALUES AND REMARKS CLINICAL LABORATORY INVESTIGATIONS

56 = Technical error for WBC

_______ Citate sample clotted

43, 50, 56, 57, 67, 74. Differential leucocyte count was also performed manually because

75, 77 of a technical error/Ian abnormal plot in the automated count and these results are

56 ~~~~Manual differential leucocyte count revealed that 1 % of netohl eand cels

*Page 85 -
APPENDIX 3

ANALYTICAL REPORT

Author

Ir. M.J.C. Brekelmans

Completion date analytical study

Laboratory Project Identification

NOTOX Project 488231

NOTOX Substance 183591/A

-Page 1 ofl 11
0.1. PIGMENT BLUE 61 NOTOX Project 488231

1. CONTENTS

1. CONTENTS........................................................................................... 2

2. REPORT APPROVAL.............................................................................. 3

3. SUMMARY ......................................................................................... 4

4. INTRODUCTION .................................................................................. 5

4.1. Preface...................................................................................... 5

4.2. Aim of the study .............................................................................. 5

5. MATERIALS AND METHODS .................................................................... 5

5.1. Reagents................................................................................... 5

5.2. Samples..................................................................................... 5

5.3. Analytical method............................................................................. 6

5.3.1. Analytical conditions.................................................................. 6

5.3.2. Preparation of the calibration solutions.............................................. 6

5.3.3. Sample injections ..................................................................... 6

5.4. Electronic data capture ...................................................................... 7

5.5. Formulas.................................................................................... 7

5.6. Specifications................................................................................. 7

6. RESULTS .......................................................................................... 8

6.1. Calibration curves ............................................................................ 8

6.2. Samples..................................................................................... 8

6.2.1. Procedural recovery samples ........................................................ 8

6.2.2. Test samples........................................................ ............. ...8

TABLES

Table 1 Procedural recovery samples............................................................ 9

Table 2 Accuracy and homogeneity test for the formulations prepared for use during

treatment ................................................................................. 9

Table 3 Stability test for the formulations prepared for use during treatment ............... 10

FIGURES

Figure 1 Chromatograms of a 15.1 mg/I test substance solution [top; res. id. 1261] and

corresponding blank [bottom; res. id. 1282] ........................................... 11

-Page 2 -
C.I. PIGMENT BLUE 61 NOTOX Project 488231

2. REPORT APPROVAL

PRINCIPAL SCIENTIST: Ir. M.J.C. Brekelmans

(Analytical Chemistry)

Date:

-Page 3 -
0.1. PIGMENT BLUE 61 NOTOX Project 488231

3. SUMMARY

The purpose of this part of the study was to determine the accuracy of preparation, homogeneity

and stability of the test substance in formulations.

Accuracy of preparation

The concentrations analysed in the formulations of Group 2, Group 3 and Group 4 were in

agreement with target concentrations (i.e. mean accuracies between 85% and 115%). No test

substance was detected in the Group 1 formulations.

Homogeneity

The formulations of Group 2 and Group 4 were homogeneous (i.e. coefficient of variation 5 10%).

Stability

Formulations of Group 2 and Group 4 were stable when stored at room temperature for at least

6 hours.

-Page 4 -
C.I. PIGMENT BLUE 61 NOTOX Project 488231

4. INTRODUCTION

4.1. Preface

Study plan Start 27 June 200B

(analytical study) Completion 01 July 2008

4.2. Aim of the study

The purpose of the analytical study was to determine the accuracy of preparation, homogeneity

and stability of the test substance in formulations.

5. MATERIALS AND METHODS

5.1. Reagents

Water Tap water purified by a Milli-Q water purification system

(Millipore, Bedford, MA, USA)

Methanol VWR Intemnational, Leuven, Belgium

Sodium 1-heptanesulfonate Fluka Chemie, Buchs, Switzerland

monohydrate (HSA)

Acetic acid Merck, Djarmstadt, Germany

Dimethyl sulfoxide (DMSO) Merck

All reagents were of analytical grade, unless specified otherwise.

5.2. Samples

Accuracy, homogeneity and stability were determined for formulations prepared for use during

treatment.

Duplicate samples (approximately 500 mg), which were taken from the formulations using a

pipette, were accurately weighed into volumetric flasks of 5, 25 or 50 ml. For determination of

accuracy, samples were taken at 50% height or at 90%, 50% and 10% height. The latter set of

samples was also used for the determination of the homogeneity of the formulations. For

determination of stability, additional samples were taken at 50% height and stored at room

temperature for 6 hours.

The volumetric flasks were filled up to the mark with DMVSO. The solutions were further diluted

with methanol by a factor of at least 100, to obtain concentrations within the calibration range.

-Page 5 -
CJI. PIGMENT BLUE 61 NOTOX Project 488231

5.3. Analytical method

5.3.1. Analytical conditions

Quantitative analysis was based on the analytical method validated for the test substance in

NOTOX project 488235.

Analytical conditions:

Instrument Alliance Separation Module 2695

(Waters, Milford, MA, USA)

Detector Dual A Absorbance Detector 2487 (Waters)

Column Symmetry Shield Ci1B, 150 mm x 4.8 mm i.d., dp =5 pm

(Waters)

Column temperature 50*C ± I *C

Injection volume 10 P1

Mobile phase A - 5/0.5/94.5 (v/vlv) water/acetic acid/methanol

containing 1.2 g/l HSA

B - 5/0.5/94.5 (v/v/v) water/acetic acid/[50/50 (v/v)

methanol/water] containing 1.2 g/l HSA

GradientC - methanol

Time %A %B %

[minutes)

0 0 100 0

20 100 0 0

20.1 0 100 0

23 0 100 0

23.1 0 0 100

28 0 0 100

28.1 0 100 0

35 0 100 0

Flow 2 mI/mmn

UV detection 313 nm

5.3.2. Preparation of the calibration solutions

Stock- and spiking solutions

Stock solutions of the test substance were prepared in DMSO at concentrations of

1108 and 1236 mg/I. In order to dissolve the test substance the solutions were ultrasonicated for

10 minutes.

Calibration solutions

Calibration solutions in the concentration range 0.997 - 15. 1mg/I were prepared from two stock

solutions, The end solution of the calibration solutions was methanol.

Procedural recovery samples

Approximately 500 mg blank vehicle was spiked with the test substance at a target concentration

of 9.66 or 194 mg/g. The accuracy samples were treated similarly as the test samples (see

paragraph 5.2 'Samples').

5.3.3. Sample Injections

Calibration solutions were injected in duplicate. Test samples and procedural recovery samples

were analysed by single injection.

- Page 6 -
ClI. PIGMENT BLUE 61 NOTOX Project 488231

5.4. Electronic data capture

System control, data acquisition and data processing were performed using the following

programme:

- Empower version 5.00 (Waters, Milford, MA, USA).

5.5. Formulas

Response (R) Peak area test substance [units]

Calibration curve R=aCN +b

where:

CN = nominal concentration [mg/I]

a = slope [units x I/mg]

b = intercept [units]

Analysed concentration (CA) CA = ( b)x [ mg/g] a w

where:

w = weight sample [mg]

V = volume volumetric flask [ml]

d = dilution factor

Recovery A X10 0% CN

where:

CN = nominal concentration [mg/g]

Accuracy EA X10O% CT

where:

CT = target concentration [mg/g]

Relative difference (relative duff.) Ct -C0 x1100% CO

where:

Ct= mean concentration of stored samples [mg/g]

C0 = mean concentration of non-stored samples fmg/g]

5.6. Specifications

Preparation of formulations was considered acceptable if the mean accuracy was in the range

85% - 115% of the target concentration and if the coeff icient of variation was 10%. Formulations

were considered stable if the relative difference between the stored and freshly taken samples

was 5 10%.

- Page 7 -
0.1. PIGMENT BLUE 61 NOTOX Project 488231

6. RESULTS

6.1. Calibration curves

A calibration curve was constructed using four concentrations. For each concentration, two

responses were used. 2Linear regression analysis was performed using the least squares method

with a 1/concentration weighting factor. The coefficient of correlation (r) was > 0.99.

Representative chromatograms of a test substance solution and blank solution are shown in

Figure 1.

5.2. Samples

6.2.1. Procedural recovery samples

The results for the procedural recovery samples are given in Table 1.

Mean recoveries of the procedural recovery samples were 100% and 101 %. Because the

criterion that mean recoveries should be between 85% and 115% was met, the results for the test

samples were accepted.

6.2.2. Test samples

The results of the test samples are given in Tables 2 and 3.

Accuracy of preparation

In the Group 1 formulations, no test substance was detected.

The concentrations analysed in the formulations of Group 2, Group 3 and Group 4 were in

agreement with target concentrations (i.e. mean accuracies between 99% and 102%).

Homogeneity

The formulations of Group 2 and Group 4 were homogeneous (i.e. coefficient of variation of 2.4%

and 0.92%, respectively).

Stability

Analysis of Group 2 and Group 4 formulations after storage yielded a relative difference of 0.79%

and 0.35%, respectively. Based on this, the formulations were found to be stable during storage

at room temperature for at least 6 hours.

-Page 8 -
0.1. PIGMENT BLUE 61 NOTOX Project 488231

TABLES

Table 1 Procedural recovery samples

Date of Date of Target Nominal Analysed Recovery Mean recovery

preparation analysis concentration concentration concentration (coefficient of

[dd-mm-yy] [dd-mm-yy] [mg/g] [mg/g] [mg/g] [%] variation)

30-06-08 30-06-08 9.66 9.81 9.86 100 101

9.65 9.63 100 (1.9)

10.4 10.3 99

9.66 9.63 100

9.68 10.1 104

30-06-08 30-06-08 194 194 193 100 100

197 198 100

Table 2 Accuracy and homogeneity test for the formulations prepared for use

during treatment

Group Date of Sample Concentration Accuracy Homogeneity

analysis position (coefficient of

I Jg/....... variation)

[dd-mm-yy] Target Analysed Individual Mean %

1 30-06-08 50% height 0.00 n.d. n.a.

0.00 n.d. n.a.

2 30-06-08 90% height 9.66 10.0 104 102 2.4

9.66 10.3 106

50% height 9.66 9.79 101

9.66 9.87 102

10% height 9.66 9.54 99

9.66 9.82 102

3 30-06-08 50% height 48.3 48.1 99 100

48.3 48.3 100

4 30-06-08 90% height 194 190 98 99 0.92

194 191 98

50% height 194 194 100

194 190 97

10% height 194 192 99

194 193 99

n.d. Not detected.

n. a. Not applicable.

- Page 9 -
0.1. PIGMENT BLUE 61 NOTOX Project 488231

Table 3 Stability test for the formulations prepared for use during treatment

Group Date of analysis Analysed concentration Relative duff.

[M /g]

[dd-mm-yy] t=0 1t=6 hours (%J

2 30-06-08 9.88 9.962 0.79

4 30-06-08 192 192 3 0.35

Mean of six samples at t=0 taken at 10%, 50% and 90% height (see Table 2).

2 Menotwsapeatt6hustkna50hehtInviulrslswr9.0ad1.mgg

Mean of two samples at t=6 hours taken at 50% height. Individual results were 929 and 1930 mg/g.

-Page 10 -
C.I. PIGMENT BLUE 61 NOTOX Project 488231

FIGURES

Figure 1 Chromatograms of a 15.1 mgI test substance solution [top: res. Id. 1261] and

corresponding blank [bottom; res. id. 1297].

-Page 11 -
ToxPath Consultancy Ltd

APPENDIX 4

PATHOLOGY REPORT

A Combined 28-Day Repeated Dose Toxicity Study with the

Reproduction/Developmental Toxicity Screening Test of C.I. Pigment Blue 61

in Rats by Oral Gavage

NOTOX Project: 488231

NOTOX Substance 183591/A

Dr AM Pilling

Po Box 16

Sandy

UK SG19l1WJ

Tel: +44 (0)1767 681546

arnpillinp0abtinternet.corn
Page 2 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

Table of Contents

Statement of Compliance ............................................................ 3

Quality Assurance Statement ........................................................ 4

Summary .............................................................................. 5

Materials and Methods .................................................................... 6

Results ................................................................................ 8

Conclusions ................................................................................ 9

Table I Incidence all Macroscopic Findings ........................................... 10

Table 2 Incidence all Microscopic Findings............................................ 11

Table 3 Individual Animal Data ......................................................... 18
Page 3 of 97

PATHOLOGY REPORT

Test substance: C.l. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

Statement of Compliance

The histopathology findings in this report were compiled by the undersigned pathologist and

accurately reflect the raw data.

The phase of work in this report was carried out in compliance with the OECD Principles of

Good Laboratory Practice, 1997 Revision (ENV/MC/CHEM (98) 17, Paris 1998 and the UK

Good Laboratory Practice Regulations 1999 (Statutory Instrument 1999 No 3106) as

amended by the Good Laboratory Practice (Codification Amendments etc) Regulations 2004

(Statutory Instrument 2004 No 994).

Dr AM Pilling BVSc PhD MRCVS FRCPath

Principal Investigator

ToxPath Consultancy Ltd

PO Box 156

Sandy

UK SG19I1WJ
Page 4 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

Quality Assurance Statement

Quality Assurance surveillance of this study phase was carried out in compliance with the

OECD Principles of Good Laboratory Practice, 1997 Revision (ENVIMCICHEM (98) 17, Paris

1998 and the UK Good Laboratory Practice Regulations 1999 (Statutory Instrument 1999 No

3106) as amended by the Good Laboratory Practice (Codification Amendments Etc)

Regulations 2004 (Statutory Instrument 2004 No 994).

The programme includes the following inspections:

Study Specific Inspections:

Study plan and report for the above study phase are reviewed and it is confirmed that the

report is a true and accurate reflection of the raw data. Findings are reported to PI, SD, Lead

QAU and test facility management.

Process Inspections:

Processes are inspected at approximately three monthly intervals, irrespective of the study

being dealt with at the time. Findings are reported to Pl.

Facility Inspections:

The facilities and operations of ToxPath Ltd are audited at approximately six monthly

intervals. Findings are reported to Pl.

Dates of Inspections

The following inspections have been carried out over the period of this study phase:

Aspect inspected Date of inspection Date of report

Study Specific

Study Plan

Report

Process

Slide reading 18 July 2008 124 July 2008

Slide recei ptides patch 18 July 2008 24 July 2008

Facility 18 July 2008 24 July 2008

David J Ford, PhD, FRQA

Quality Assurance Consultant

David Ford Consultancy

Goodwyn House

Bank Street

Puihamn Market

Diss Norfolk IP21 4TG

ENGLAND
Page 5 of 97

PATHOLOGY REPORT

Test substance: C. . Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study Ini the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

Summary

Histopathological examinations were performed on tissues from Han Wistar rats which had

received C.I. Pigment Blue 61 orally for a minimum of 28 days.

Rats were assigned to four dose groups each containing ten F0 males and ten F0 females.

The test substance was administered by oral gavage once daily for a minimum of 28 days at

doses of 50, 250 or 1000 mg/kg. The control group received the vehicle (propylene glycol)

alone. A full necropsy was performed on five rats/sex/group. A limited necropsy was

performed on the remaining animals. Histopathological evaluation was performed on tissues

from the first five males, and the first five females with live offspring, in groups 1 and 4 and on

macroscopic abnormalities from rats in all groups. In addition, the reproductive organs from

all animals that failed to mate, conceive, sire or deliver healthy offspring were examined

histologically. PAS stained sections of testis from the five selected F0 males in groups 1 and

4 were examined for abnormalities in spermatogenesis. Histological examination was

extended to the mesenteric lymph nodes of the selected males and females of Group 2.

No animals died, or were killed, during the treatment period.

Gross findings included blue/green discolouration of the mesenteric lymph nodes in rats in all

dose groups and the presence of blue gastrointestinal contents in high dose females.

No histopathological changes were observed in association with the gross findings or in any

other tissues of rats receiving the test substance.

On the basis of the pathological findings the NOAEL may be regarded as 1000 mg/kg/day.
Page 6 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.- 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

Materials and Methods

Study Design

Group Dose level Number of Animals Animal Numbers

(mglkg) F0 Males F0 Females Males Females

1 0 10 10 1-10 41-50

2 50 10 10 11-20 51-60

3 250 10 10 21-30 61-70

4 1000 10 10 31-40 71-80

Administration of the Test Substance

The test substance was administered orally once daily for a minimum of 28 days at doses of

0, 50, 250 or 1000 mg/kg. The control group received the vehicle (propylene glycol) alone.

Necropsy and Histopatho/ogy

At the end of the study period, the rats were killed by exsanguination following anaesthesia by

isoflurane. A full necropsy was performed on five rats/sex/group. A limited necropsy was

performed on the remaining animals. Organs were preserved in 10% neutral buffered

formalin (testes, epididymides and eyes were initially fixed in modified Davidson's solution

before being transferred to formalin).

Histopathological evaluation was performed on representative samples of the following

tissues from the first five males, and the first five females with live offspring, in groups 1 and

4: adrenal glands; aorta; brain - cerebellum, midbrain, cortex; cervix; epididymides; heart,

kidneys; large intestine - caecum, clitoral gland: coagulating gland; colon, rectum; liver; lung;

lymph nodes - mandibular, mesenteric; oesophagus; ovaries; pancreas; peyer's patches;

pituitary; preputial gland; prostate gland; sciatic nerve; seminal vesicles; small intestine -

duodenum, jejunum, ileum; spinal cord - cervical, mid-thoracic, lumbar; spleen; sternum with

bone marrow; stomach; te stes; thymus; thyroid with parathyroid; trachea; urinary bladder,

uterus; vagina: all gross lesions (examined from all F0 rats on study). The reproductive

organs (testis, epididymides, coagulating gland, seminal vesicles, prostate, preputial gland,

ovaries, uterus, cervix, vagina, clitoral gland) from all animals that failed to mate, conceive,

sire or deliver healthy offspring were also examined. Histological examination was extended

to the mesenteric lymph nodes of the selected males and females of Group 2.

Following fixation, tissues were trimmed, processed and embedded in paraffin wax. Sections

2-4 tm in thickness were prepared and stained with haemnatoxylin and eosin for routine

evaluation. Additional sections of testis from the five selected F0 males in groups 1 and 4

were stained with PAS and examined for abnormalities in spermatogenesis.
Page 7 of 97

PATHOLOGY REPORT

Test substance: C.i. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct.08

Sponsor: Color Pigments Manufacturers Association Inc.

Data Compilation

Details of study design and macroscopic findings were transferred electronically from the

necropsy raw data files of NOTOX By,. into the computer system of ToxPath Consultancy Ltd

where the microscopic findings were recorded using on-line input.

Macroscopic findings are summarised in Table 1 (Incidence of Macroscopic Findings) and

listed in Table 3 (Individual AnimallData). As far as possible, macroscopic findings were

correlated with a microscopic finding.

Microscopic findings are listed for each animal, along with the severity grades, in Table 3 and

summarised in Table 2 (incidence of Microscopic Findings). Histological changes were

described according to their distribution and morphological character. Findings were also

graded for severity (or degree) on a scale of 1-5 and these are described in Table 3.

Archiving

The final pathology report (original hardcopy), all glass microscope slides, slide inventory

sheets and a copy of the electronic records (including final pathology report) as captured by

the appropriate software on CDROM will be sent to NOTOX BV for archiving. Any other

non-study related documents an~d electronic data, covered by GLP, will be archived by

ToxPath Consultancy Ltd according to company standard operating procedures.
Page 8 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

Results

Mortality

All F0 rats survived the scheduled duration of the study.

Macroscopic findings

Green or blue discolouration of the mesenteric lymph nodes was observed in a few group 2

rats and in the majority of rats of both sexes in groups 3 and 4. Blue gastrointestinal tract

contents were present in several group 4 females. Blue discolouration of the tail skin was

noted in one male group 4 rat.

No other findings were recorded at necropsy that were considered to be due to the test

substance.

Microscopic findings

No treatment related histopathological changes were observed in the tissues of any animal

receiving 0.1. Pigment Blue 61.

No abnormalities were noted on evaluation of PAS stained sections of testes of F0 males. All

seminiferous tubules were normal with respect to their stage in the spermatogenic cycle and

the integrity of the various cell types present within the different stages.

No pathological findings were present in the reproductive organs of male and female animals

that failed to produce offspring.

Small amounts of blue material were observed in the lumen of the duodenum, ileum, jejunum,

colon or rectum in a few group 4 animals. Blue material was also noted in the superficial skin

(keratin layer) of the tail in one group 4 male. These findings are considered to represent the

presence of the test substance and in some cases correlated with the macroscopic

observations.

No microscopic findings were observed in the mesenteric lymph nodes to account for the

discolouration of these tissues recorded at necropsy. It is considered that the discolouration

is due to the presence of the test substance and that the pigment was lost from the tissues in

the processing or staining procedures.

All other microscopic findings were considered to be incidental and unrelated to treatment

with the test substance.
Page 9 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system; 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

Conclusions

Administration of ClI. Pigment Blue 61 by oral gavage to Han Wistar rats at doses of 50, 250

or 1000 mg/kg/day was associated with blue gastrointestinal contents in high dose females

and blue/green discolouration of the mesenteric lymph nodes in rats in all dose groups.

No histopathological changes were observed in association with the gross findings or in any

other tissues of rats receiving the test substance.

On the basis of the pathological findings the NOAEL may be regarded as 1000 mg/kg/day.
PATHOLOGY REPORTPae1of9

Test substance: C. 1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 1 - INCIDENCE OF MACROSCOPIC FINDINGS

SEX: Male Female

GROUP: 1 2 3 4 1 2 3 4

number of animals 10 10 10 10 10 10 10 10

ADRENAL GLANDS

DISCOLOURATION 0 0 0 0 0 0 0 1

EPIDIDYMIDES

NODULE(S)

tail 0 0 2 0 - - - -

GENERAL OBSERVATIONS

BLUISH

Gltract contenlts 0 0 0 0 0 0 0 7

MANDIBULAR LYMPH NODES

DISCOLOURATION 0 0 0 0 1 2 0 1

MESENT. LYMPH NODE

DISCOLOURATION 0 2 9 10 0 1 8 9

PANCREAS

DISCOLOURATION 0 0 0 0 0 1 0 0

SEMINAL VESICLES

REDUCED INSIZE 0 0 0 1 - - - -

SKIM

DISCOLOURATION

tail 0 1 0 1 0 0 0 0

ALOPECIA

abdominal region 0 0 0 0 1 0 0 0

flank 0 0 0 0 1 0 0 0

foreleg 0 0 0 1 1 0 0 1

hindleg 0 0 0 0 0 0 0 1

TAIL

TAIL BENT 1 0 0 0 0 0 0 0
Page 11 of 97

PATHOLOGY REPORT

Test substance- C.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 2 - INCIDENCE OF MICROSCOPIC FINDINGS

The grading systems used are as follows:

1 (minimal)

2 (alight)

3 (moderate)

4 (marked)

5 (severe)
Page 12 of 97

PATHOLOGY REPORT

Test substance: C.l. Pigment Blue 81 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 2 - INCIDENCE OF MICROSCOPIC FINDINGS

SEX: Male Female

GROUP: 1 2 3 4 1 2 3 4

number of animals 10 10 10 10 10 10 10 10

ADRENAL

number examined 5 - - S 5 - - 5

AORTA

number examined 5 - 55 - -

BRAIN

number examined 5 - 5 5 - -

CASCUM

number examined 5 - 5 5 - - 5

CERVIX

number examined - - - 6 1 1 6

CLITORAL GLAND

number examined - - - -6 1 1 6

interstitial

inflammatory cell

infiltration

(minimal) - - -1 0 0 0

COAGULATING GLAND

number examined 5 1 1 5 --

COLON

number examined 5 - - 5 5 5

blue material in

lumen

present 0 - - 1 0 - - 1

DUJODENUM4

number examined 5 5

blue material in

lumen

present 0 - - 1 0 - - 0

EPIDIDYMIS

number examined 5 1 3 5 - - -

sperm granuloma

(slight) 0 0 1 0 - - -

(moderate) 0 0 1 0- - -

Total 0 0 2 0 - - -

HEART

number examined 5 - - 5 5 - - 5

epicardial fibrosis

(minimal) 0 - - 0 0 - - 1
Page 13 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 2 - ZNCIDENCE OF MICROSCOPIC FINDINGS

SEX: Male Female

GROUP: 1 2 3 4 1 2 3 4

number of animals 10 10 10 10 10 10 10 10

ILEUM

number examined 5 -5 5 

blue material in

lumen

present 0 - - 0 0 - - 1

JEJUNUM

number examined 5 - - 5 5 - - 5

blue eaterial in

lumen

present 0 - - 1 o0 1

KIDNEYS

number examined 5 - - 5 5 - - 5

basophilic tubules

(minimal) 1 - - 0 0 - - 1

interstitial

inflammatory call

infiltration

(minimal) 1 - 0 0 - - 0

tubular hyaline

droplets

(minimal) 1 - - 3 0 - - 0

tubular

mineralization

(minimal) 0 - - 0 2 - - 1

pelvic dilatation

(minimal) 2 - - 1 0 - - 0

LIVER

number examined 5 5 5 -

parenchymal

inflammatory cell

infiltration

(minimal) 3 - - 2 1 - - 0

periportal

inflammatory call

infiltration

(minimal) 0 - - 1 0 - - 0
Page 14 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor Color Pigments Manufacturers Association Inc.

TABLE 2 - INCIDENCE OF MICROSCOPIC FINDINGS

SEX: Male Female

GROUP: 1 2 3 4 1 2 3 4

number of animals 10 10 10 10 10 10 10 10

LUNG

number examined 5 - 5

perivascular

inflammatory cell

infiltration

(minimal) 2 - - 1 0 - - 1

alveolitis

(minimal) 1 - - 0 0 - - 0

alveolar macrophage

infiltration

(minimal) 0 - - 1 0 - - 1

LYMPH NODE MESENTERIC

number examined 5 5 9 9 5 5 8 9

macrophage aggregates

(minimal) 1 1 3 2 0 0 1 0

sinus histiocytosia

(minimal) 0 0 0 0 0 0 1 0

(slight) 0 0 0 0 0 1 0 0

Total 0 0 0 0 0 1 1 0

MANDIBULAR LYMPH NODE

number examined 5 - - 5 5 2 - 5

plasma cell

infiltration

(minimal) 3 - - 4 0 - 2 (slight) 0 - - 3 0 0 - 1

Total 3 - - 4 4 0 - 3

congestion

present 0 - - 0 1 2 - 1

OESOPHAGUS

number examined 5 - - 5 5 - - 5

muscularis

inflammation

(minimal) 0 - - 0 2 - - 0

OVARIES

number examined - - - -6 1 1 6

PANCREAS

number examined 5 5 5 1 - 5

acinar vacuolation

(minimal) 1 - - 0 0 0 - 0

focal acinar atrophy

(minimal) 0 - - 0 1 0 - 0
PATHOLOGY REPORT Pglog

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 2 - INCIDENCE OF MICROSCOPIC FINDINGS

SEX: Male Female

GROUP: 1 2 3 4 1 2 3 4

number of animals 10 10 10 10 10 10 10 10

PARATHYROID

number examined 5 - 4 5 -

PEYERS PATCHES

number examined 5 - - 5 5 - 5

PITUITARY

number examined 5 5 5 5

cyst(s)

present 0 - - 1 0 - - 0

PREPIIAL GLAND

number examined 5 1 1 5- - -

PROSTATE

number examined 5 1 1 5- - -

interstitial

inflammatory cell

infiltration

(minimal) 3 0 1 2- - - -

(slight) 1 0 0 0- - - -

Total 4 0 1 2- - -

RECTUM

number examined 5 - 5 - -

blue material in

lumen

present 0 - - 1 0 - - 1

SCIATIC NERVE

number examined 5 - - 4 5 - - 5

SEMINAL VESICLES

number examined 5 1 1 6- - -

reduced luminal

secretory contents

present 0 0 0 1-- -
Page 16 of 97

PATHOLOGY REPORT

Test substance: C. 1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor. Color Pigments Manufacturers Association Inc.

TABLE 2 - INCIDENCE OF MICROSCOPIC FINDINGS

SEX: Male Female

GROUP: 1 2 3 4 1 2 3 4

number of animals 10 10 10 10 10 10 10 10

SKIN

number examined - I - 2 1 - - I

epidermal hyperplasia

(slight) 0 - 1 0 - - 0

abdomen: epidermal

hyperplasia

(minimal) - 0 - 0 1 - - 0

foreleg right side:

epidermal

hyperplasia

(minimal) 0 - 0 1 - -

flank left aide:

epidermal

hyperplasia

(minimal) - 0 - 0 1 - - 0

blue material in

surface keratin

present - 0 - 1 0 - - 0

foreleg left side:

epidermal

hyperplesia

(minimal) - 0 - 0 0 - - 1

SPINAL CORD

number examined 5 - - 5 5 - - 5

SPLEEN

number examined 5 - - 5 5 - - 5

extramedullary

haematopoiesis

(slight) 0 - - 0 0 - -

STER1-UM

number examined - - - 5

STOMACH

number examined 5 - 5 

TAIL

number examined 1I -- - - -

TESTIS

number examined 5 1 1 5 - - -

PAS section: no

abnormalities

present 5 0 0 4 - - -

hypospermatogenesis

(minimal) 0 0 0 1- - -
Page 17 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-OS

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 2 - INCIDENCE OF MICROSCOPIC FINDINGS

SEX: Male Female

GROUP: 1 2 3 4 1 2 3 4

number of animals 10 10 10 10 10 10 10 10

THYMUS

number examined 5 - - s 5 - - 5

involution/atrophy

(minimal) 0 - - 0 1 - - 1

THYROID

number examined 5 - - 5 5 - - 5

TRACHEA

number examined 5 - - 5 5 - 5

URINARY BLADDER

number examined 5 - - 5 5 - - 5

UTERUS

number examined - - - - 6 1 1 6

VAGINA

number examined - - - 6 1 1 6

oestrus

present - - - - 1 0 0 0

metoestrus

present - - 0 0 1 1

dioestrua

present - - -5 1 0 5

epi thelial1

vacuoletion

(minimal) - 3 0 0 1

(slight) - 1 0 0 3

(moderate) - - -1 0 0 0

Total - - - -5 0 0 4
Page 18 of 97

PATHOLOGY REPORT

Test substance: C.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.:

SEX: male

DOSE LEVEL: 1

M A C R O S C O P I C F I N DlI N G S d a s o t r t m n 3 0

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy 17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

LUNG perivascular inflammatory call infiltration (ainimal)

MANDIBULAR plasma cell infiltration (minimal)

LYMPH NODE

PROSTATE interstitial inflammatory cell infiltration (minimal)

TESTIS PAS section: no abnormalities present

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, COAGULATING

GLAND, COLON, DUODENUM, EPIDIDYMIS, HEART, ILEUM, JEJUNUM, KIDNEYS,

LIVER, LYMPH NODE MESENTERIC, OESOPHAGUS, PANCREAS, PARATHYROID, PEYERS

PATCHES, PITUITARY, PREPUTIAL GLAND, RECTUM, SCIATIC NERVE, SEMINAL

VESICLES, SPINAL CORD, SPLEEN, STERNUM, STOMACH, THYMUS, THYROID,

TRACHEA, URINARY BLADDER.
PATHOLOGY REPORTPae1of9

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 2

SEX; Male

DOSE LEVEL: 1

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatent: 16/07/08

date of necropsy :17/07/Os

Mo abnormality detected

MICROSCOPIC FINDINGS

KIDNEYS pelvic dilatation (minimal)

LUNG perivascular inflammatory cell infiltration (minimal)

MANDIBULAR plasma cell infiltration (minimal)

LYMPM NODE

PROSTATE interstitial inflammatory cell infiltration (minimal)

TESTIS PAS section: no abnormalities present

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, COAGULATING

GLAND, COLON, DUODENUM, EPIDIDYMIS, HEART, ILEUM, JEJUNUM, LIVER, LYMPH

MODE MESENTERIC, OESDPHAGUS, PANCREAS, PARATHYROID, PEYERS PATCHES,

PITUITARY, PREPUTIAL GLAND, RECTUM, SCIATIC NERVE, SEMINAL VESICLES,

SPINAL CORD, SPLEEN, STERNUM, STOMACH, THYMUS, THYROID, TRACHEA, URINARY

BLADDER.
Page 20 of 97

PATHOLOGY REPORT

Test substance: C.l. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.:i 3

SEX: Male

DOSS LEVEL: 1

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

data of start of treatment: 17/06/08

date of end of treetment: 16/07/08

date of necropsy :17/07/08

TAIL BENT

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

KIDNEYS basophilic tubules (minimal)

interstitial inflammatory cell infiltration (minimal)

tubular hyaline droplets (minimal)

pelvic dilatation (minimal)

LIVER parenchymal inflammatory cell infiltration (minimal)

MANDIBULAR plasma cell infiltration (minimal)

LYMPH NODE

PROSTATE interstitial inflammatory cell infiltration (slight)

TAIL to0 lesion to account for gross finding(s)

TESTIS PAS section: no abnormalities present

Ho abnormalities found in: ADRENAL, AORTA, DRAIN, CAECUM, COAGULATING

GLAND, COLON, DUODENUM, EPIDIDYMIS, HEART, ILEUM, JEJUNUM, LUNG, LYMPH

MODE MESENTERIC, OESOPHAGUS, PANCREAS, PARATHYROID, PEYERS PATCHES,

PITUITARY, PREPUTIAL GLAND, RECTUM, SCIATIC NERVE, SEMINAL VESICLES,

SPINAL CORD, SPLEEN, STERNUM, STOMACH, THYMUS, THYROID, TRACHEA, URINARY

BLADDER.
Page 21 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 4

SEX: Male

DOSE LEVEL: 1

MACROSCOPIC FINDlINGS

daya of treatment :30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

data of end of treatment: 16/07/08

date of necropsy :17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

LIVER parenchymal inflammatory cell infiltration (minimal)

LUNG alveolitis (minimal)

PANCREAS acinar vacuolation (minimal)

TESTIS PAS section: no abnormalities present

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, COAGULATING

GLAND, COLON, DUODENUM, EPIDIDYNIS, HEART, ILEUM, JEJUNUM, KIDNEYS, LYMPH NODE MESENTERIC, MANDIBULAR LYMPH NODE, OESOPHAGUS, PARATHYROID, PEYERS

PATCHES, PITUITARY, PREPUTIAL GLAND, PROSTATE, RECTUM, SCIATIC NERVE, SEMINAL VESICLES, SPINAL CORD, SPLEEN, STERNUM, STOMACH, THYMUS, THYROID,

TRACHEA, URINARY BLADDER.
Page 22 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor. Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 5

SEX: Male

DOSE LEVEL: 1

MACROSCOPIC FINDINGS

days of treatment :30

sacrif ice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

data of end of treatment: 16/07/08

data of necropsy :17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

LIVER parenchymal inflammatory cell infiltration (minimal)

LYMPH NODE macrophage aggregates (minimal)

MESENTERIC

PROSTATE interstitial inflammatory cell infiltration (minimal)

TESTIS PAS section: no abnormalities present

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, COAGULATING

GLAND, COLON, DUODENUM, EPIDIDYMIS, HEART, ILEUM, JEJUNUM, KIDNEYS, LUNG,

MANDIBULAR LYMPH NODE, OESOPHAGUS, PANCREAS, PARATHYROID, PEYERS PATCHES,

PITUITARY, PREPUTIAL GLAND, RECTUM, SCIATIC NERVE, SEMINAL VESICLES,

SPINAL CORD, SPLEEN, STERNUM, STOMACH, THYMUS, THYROID, TRACHEA, URINARY

BLADDER.
Page 23 of 97

PATHOLOGY REPORT

Test substance: C.!. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: S

SEX: Male

DOSE LEVEL: I

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy :17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 24 of 97

PATHOLOGY REPORT

Test substance: C. 1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 7

SEX: Male

DOSE LEVEL: 1

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group :terminal sacrifice

necropay status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

data of necropsy : 17/07/08

No abnormality detacted

MICROSCOPIC FINDINGS

No tissues examined
Page 25 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 26-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. : 8

SEX; Male

DOSE LEVEL: I

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/O6/OS

date of end of treatment: 16/07/08

date of necropsy :17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
PATHOLOGY REPORTPae2of9

Test substance: C.1. Pigment Blue 61 Notox No-. 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 9

SEX: Male

DOSE LEVEL: I

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment! 16/07/08

date of necropsy : 17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
PATHOLOGY REPORTPae2of9

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 10

SEX: Male

DOSE LEVEL: 1

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment; 16/07/08

date of necropsy 17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 28 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24.Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 11

SEX: Male

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group terminal sacrifice necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy :17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No abnormality detected

No abnormalities found in: LYMPH NODE MESENTERIC,
PATHOLOGY REPORTPae2of9

Test substance: C.l. Pigment Blue 81 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 12

SEX: Male

DOSE LEVEL: 2

M4ACROSCOPIC FINDINGS

days of treatment 3 0

sacrifice group terminal sacrifice

necropsy atatua planned terminal

date of atart of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropay : 17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tiasues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding Is)

MESENTERIC
Page 30 of 97

PATHOLOGY REPORT

Test substance: C. 1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 13

SEX; Male

DOSE LEVEL; 2

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group terminal sacrif ice

necropsy statue planned terminal

date of start of treatment; 17/06/08

date of end of treatment: 16/07/08

date of necropsy 17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
PATHOLOGY REPORTPae3of9

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-OS

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 14

SEX: Male

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment 3 0

sacrifice group terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/Os

date of necropsy :17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

LYMPH MODE macrophage aggregates (minimal)

MESENTERIC

No abnormalities found in: COAGULATING GLAND, EPIDIDYMIS, PREPUTIAL GLAND,

PROSTATE, SEMINAL VESICLES, TESTIS.
Page 32 of 97

PATHOLOGY REPORT

Test substance: C. 1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. is1

SEX H ale

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/ce

date of end of treatment: 16/07/08

date of necropsy ;17/07/08

SKIN tail: DISCOLOURATION BLUISH

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

SKIN no lesion to account for gross finding Is)

No abnormalities found in: LYMPN NODE MESENTERIC.
Page 33 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 26-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 16

SEX: Male

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group :terminal sacrifice

nacropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy : 17/07/08

MESENT. LYMPH DIScOLOURATION GREENISH

NODE

Ho abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding(s)

MESENTERIC
Page 34 of 97

PATHOLOGY REPORT

Test substance: C.l. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. : 17

SEX : Male

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

data Of start of treatment: 17/06/08

data of end of treatment: 16/07/08

date of necropsy : 17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
PATHOLOGY REPORTPae3of9

Test substance: C. . Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 18

SEX: Male

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/05

date of end of treatment: 16/07/08

date of necropsy : 17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tisaues examined
Page 36 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 19

SEX; Male

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment 30

sacrif ice group :terminal sacrif ice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy 17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 37 of 97

PATHOLOGY REPORT

Test substance: C.!. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 20

SEX: Male

DOSE LEVEL; 2

MACROSCOPIC FINDINGS

days of treatment :30

eacrifice group :terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy :17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 38 of 97

PATHOLOGY REPORT

Test substance. CtI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 21

SEX: Male

DOSE LEVEL: 3

MACROSCOPIC FINDlINGS

days of treatment 30

sacrifice group terminal sacrif ice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropay :17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding Is)

MESENTERIC
Page 39 of 97

PATHOLOGY REPORT

Test substance: 0.1I. Pigment Blue 61 Notox No.: 468231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 22

SEX: Male

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 3 0

sacrifice group terminal sacrifice

necropsy status planned terminal

date of start of treatment; 17/06/0e

date of end of treatment: 16/07/08

date of necropsy 17/07/08

EPIDIDYNIDES tail: right side NODULE(S) D=7X5 NM, YELLOWISH, HARD

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

EPIDIDYMIS sperm granuloma (moderate) (correlates to GROSS finding)

LYMPH NODE no lesion to account for gross finding(s)

MESENTERIC
Page 40 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No,: 23

SEX: Male

DOSE LEVEL; 3

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy 17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

MODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE macrophage aggregates (minimal)

MESENTERIC no lesion to account for gross finding 1s1
Page 41 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-OctO8

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.; 24

SEX: Male

DOSE LEVELi 3

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group terminal sacrifice

necropsy status planned terminal

date of start of treatment; 17/06/08

date of end of treatment: 16/07/08

date of necropsy 17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross findingis)

MESENTERIC
Page 42 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 25

SEX: Male

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment :30

sacrif ice group terminal sacrifice

necropsy status planned terminal

date of start of treatment; 17/06/08

date of end of treatment: 16/07/08

date of necropsy : 17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding Is)

MESENTERIC
Page 43 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Ora( Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 26

SEX: Male

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 3 0 sacrifice group :terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy :17/07/08

No abnormality detected

MICROSCOPIC FINDINGS

PROSTATE interstitial inflammatory cell infiltration (minimal)

No abnormalities found in: COAGULATING GLAND, EPIDIDYMIS, PREPUTIAL GLAND,

SEMINAL VESICLES, TESTIS.
Page 44 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 27

SEX: Male

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group :terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy : 17/07/08

EPIDIDYMIDES tail: right side NODTULE(S) D=2X2 MM, YELLOWISH, HARD

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

EPIDIDYMIS sperm granuloma (alight) (correlates to GROSS finding)

LYMPH NODE no lesion to account for gross findinglal

MESENTERIC
Page 45 of 97

PATHOLOGY REPORT

Test substance: C.l. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 28

SEX: Male

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

daya of treatment :30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of atart of treatment; 17/06/08

date of end of treatment: 16/07/08

date of necropsy :17/07/08

mEsENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE macrophage aggregates (minimal)

MESENTERIC no lesion to account for gross finding(s)
Page 46 of 97

PATHOLOGY REPORT

Test substance: ClI. Figment Blue 61 Notox No.: 488231

Test system: 26-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.; 29

SEX: Male

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group terminal sacrifice

necropsy atatus :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy ;17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE macrophage aggregates iminimal)

MESENTERIC no lesion to account for gross findingis)
Page 47 of 97

PATHOLOGY REPORT

Test substance: C. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 30

SEX: Male

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy :17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding(s)

MESENTERIC
Page 48 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 81 Notox No.: 488231

Test system: 28-Day Owal Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 31

SEX: Male

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group :terminal saecrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

data of end of treatment: 16/07/08

date of necropsy 17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

DUODENUM blue material in lumen present

JEJUNUN blue material in lumen present

KIDNEYS pelvic dilatation (minimal)

LYMPH NODE macrophage aggregates (minimal)

MESENTERIC no lesion to account for gross finding(s)

MANDIBULAR plasma cell infiltration (slight)

LYMPH NODE

TESTIS PAS section: no abnormalities present

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, COAGULATING

GLAND, COLON, EPIDIDYMIS, HEART, ILEUM, LIVER, LUNG, CESOPHAGUS,

PANCREAS, PEYERS PATCHES, PITUITARY, PREPUTIAL GLAND, PROSTATE, RECTUM,

SCIATIC NERVE, SEMINAL VESICLES, SPINAL CORD, SPLEEN, STERNUM, STOMACH,

THYMUS, THYROID, TRACHEA, URINARY BLADDER.
Page 49 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-OS

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 -INDIVIDUAL ANIMAL DATA

Animal No.; 32

SEX; Male

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group terminal sacrifice

nacropay status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy : 17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

COLON blue material in lumen present

KIDNEYS tubular hyaline droplets (minimal)

LIVER parenchymal inflammatory cell infiltration (minimal)

LUNG alveolar macrophage infiltration (minimal)

LYMPH NODE no lesion to account for gross finding(s)

MESENTERIC

No abnormalities found in: ADRENAL, AORTA, BRAIN, CASCUN, COAGULATING

GLAND, DUODENUM, EPIDIDYMIS, HEART, ILEUM, JEJUJNUM, MANDIBULAR LYMPH

NODE, OESOPHAGUS, PANCREAS, PARATHYROID, PEYERS PATCHES, PITUITARY,

PREPUTIAL GLAND, PROSTATE, RECTUM, SCIATIC NERVE, SEMINAL VESICLES,

SPINAL CORD, SPLEEN, STERNUM, STOMACH, TESTIS, THYMUS, THYROID, TRACHEA,

URINARY BLADDER.
Page 50 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 33

SEX: Male

DOSE LEVEL: 4

MACROSCOPIC FIND)INGS

days of treatment :30

sacrifice group :terminal sacrif ice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy :17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding(s)

MESENTERIC

MANDIBULAR plasma cell infiltration (slight)

LYMPH NODE

PROSTATE interstitial inflammatory cell infiltration (minimal)

TESTIS PAS section: no abnormalities present

hypospermatogenesis (minimal)

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, COAGULATING

GLAND, COLON, DUODENUM, EPIDIDYMIS, HEART, ILEUM, JEJUNUM, KIDNEYS,

LIVER, LUNG, OESOPHAGUS, PANCREAS, PARATHYROID, PEYERS PATCHES,

PITUITARY, PREPUTIAL GLAND, RECTUM, SCIATIC NERVE, SEMINAL VESICLES,

SPINAL CORD, SPLEEN, STERNUM, STOMACH, THYMUS, THYROID, TRACHEA, URINARY

BLADDER.
Page 51 of197

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 34

SEX: Hale

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :30

aacrif ice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy 17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

SKIN FORELEG: left side ALOPECIA

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

KIDNEYS tubular hyaline droplets (minimal)

LIVER periportal inflammatory cell infiltration (minimal)

LYMPH NODE no lesion to account for gross findingis)

MESENTERIC

MANDIBULAR plasma cell infiltration (minimal)

LYMPH NODE

PITUITARY cyst Is) present

PROSTATE interstitial inflammatory call infiltration (minimal)

RECTUM blue meaterial in lumen present

SKIN epidermal hyperplasia (slight) (correlates to GROSS finding)

TESTIS PAS section: no abnormalities present

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUN, COAGULATING

GLAND, COLON, DUODENUM, EPrDIDYNIS, HEART, ILEUN, JIEJUNUM, LUNG,

OESOPHAGUS, PANCREAS, PARATHYROID, PEYERS PATCHES, PREPUTIAL GLAND,

SEMINAL VESICLES, SPINAL CORD, SPLEEN, STERNUM, STOMACH, THYMUS, THYROID,

TRACHEA, URINARY BLADDER.
Page 52 of 97

PATHOLOGY REPORT

Test substance: C.l. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 35

SEX: Male

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group :terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy i17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

KIDNEYS tubular hyaline droplets (minimal)

LIVER parenchymal inflammatory cell infiltration (minimal)

LUNG perivasculer inflammatory cell infiltration (minimal)

LYMPH HODE no lesion to account for gross finding Is)

NEsENTERIC

MANDIBULAR plasma cell infiltration (slight)

LYMPH NODE

TESTIS PAS section: no abnormalities present

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, COAGULATING

GLAND, COLON, DUODENUM, EPIDIDYNIS, HEART, ILEUM, JEJUNUM, ORSOPHAGUS,

PANCREAS, PARATHYROID, PEYERS PATCHES, PITUITARY, PREPUTIAL GLAND,

PROSTATE, RECTUN, SCIATIC NERVE, SEMINAL VESICLES, SPINAL CORD, SPLEEN,

STERNUM, STOMACH, THYMUS, THYROID, TRACHEA, URINARY BLADDER.
PATHOLOGY REPORTPae5of9

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 36

SEX; Male

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment 30

sacrifice group :terminal sacrifice

necropsy status planned terminal

date of start of treatment; 17/06/08

date of end of treatment: 16/07/08

date of necropsy : 17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE macrophage aggregates (minimal)

MESENTERIC no lesion to account for gross finding Is)
Page 54 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Biue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc,

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 37

SEX: Male

DOSE LEVEL: 4

MACROSCOPIC FINDlINGS

days of treatment :30

sacrifice group terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 1s/07/08

date of necropsy :17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

No tissues examined
Page 55 of 97

PATHOLOGY REPORT

Test substance: 0. 1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 38

SEX: Male

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment .30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy ; 17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

No abnormalities ware found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding(s)

MESENTERIC
Page 56 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 39

SEX: male

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :30

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy :17/07/0s

MESENT. LYNPH DISCOLOURATION GREEMISM

NODE

SKIN tail: DISCOLOURATION BLUISH

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding(s)

MESENTERIC

SKIN blue material in Surface keratin present (correlates to GROSS finding)
Page 57 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.! 40

SEX: Male

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment : 30

sacrifice group : terminal sacrifice

necropsy status : planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 16/07/08

date of necropsy :17/07/08

MESENT. LYMPH DISCOLOURATION GREENISH

NODE

SEMINAL left side REDUCED IN SIZE

VESICLES

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross findingis)

MESENTERIC

SEMINAL reduced luminal secretory contents present (correlates to GROSS finding)

VESICLES
Page 58 of 97

PATHOLOGY REPORT

Test substance: Ctf. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 41

SEX; Female

DOSE LEVEL; 1

MACROSCOPIC FINDINGS

days of treatment 42

aacrifice group terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment; 28/07/08

date of necropsy :29/07/08

No abnormality detected

MICROSCOPIC FINDINGS

CLITORAL GLAND interstitial inflammatory cell infiltration (minimal)

VAGINA oestrus present

No abnormalities found in: CERVIX, OVARIES, UTERUS.
Page 59 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Anlimal No.: 42

SEX: Female

DOSE LEVEL: 1

MACROSCOPIC FINDINGS

days of treatment 41

sacrifice group :terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 27/07/08

date of necropsy ;28/07/08

No abnormality detected

MICROSCOPIC FINDINGS

KIDNEYS tubular mineralization (minimal)

MANDIBUJLAR plasma cell Infiltration (minimal)

LYMPH NODE

THYMUS involution/atrophy (minimal)

VAGINA dioestrus present

epithel~ial vacuolation (slight)

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, CERVIX, CLITORAL

GLAND, COLON, DUODENUM, HEART, ILEUM, JEJUNUM, LIVER, LUNG, LYMPH NODE

MESENTERIC, OESOPHAGUS, OVARIES, PANCREAS, PARATHYROID, PEYERS PATCHES,

PITUITARY, RECTUM, SCIATIC NERVE, SPINAL CORD, SPLEEN, STERNUM, STOMACH,

THYROID, TRACHEA, URINARY BLADDER, UTERUS.
Page 60 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 43

SEX: Female

DOSE LEVEL: 1

MACROSCOPIC FINDlINGS

days of treatment 41

sacrifice group terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 27/07/08

date of necropsy :28/07/08

No abnormality detected

MICROSCOPIC FINDINGS

MANDIBULAR plasma cell infiltration (minimal)

LYMPH NODE

OESOPHAGUS muscularis inflammation (minimal)

VAGINA dioestrus present

epithelial vacuolation (moderate)

Ho abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUH, CERVIX, CLITORAL

GLAND, COLON, DUODENUM, HEART, ILEUM, JEJUNUM, KIDNEYS, LIVER, LUNG,

LYMPH NODE MESENTERIC, OVARIES, PANCREAS, PARATHYROID, PEYERS PATCHES,

PITUITARY, RECTUM, SCIATIC NERVE, SPINAL CORD, SPLEEN, STERNUM, STOMACH,

THYMUS, THYROID), TRACHEA, URINARY BLADDER, UTERUS.
Page 61 of 97

PATHOLOGY REPORT

Test substance: C.l. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-O8

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 44

SEX: Female

DOSE LEVEL; I

MACROSCOPIC FINDINGS

days of treatment :45

sacrifice group :terminal sacrifice necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 31/07/08

date of necropsy : 01/08/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 62 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.; 45

SEX: Female

DOSE LEVEL: I

MACROSCOPIC FINDINGS

days of treatment :45

sacrif ice group :terminal sacrif ice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 31/07/08

date of necropsy :01/08/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 63 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 46

SEX: Female

DOSE LEVEL: I

MACROSCOPIC FINDINGS

days of treatment :45

sacrifice group terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment; 31/07/08

date of necropsy : 01/08/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 64 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 47

SEX: Female

DOSE LEVEL: 1

MACROSCOPIC FINDINGS

days of treatment 4 43

sacrifice group terminal sacrifice

necropsy status planned terminal

data of start of treatment: 17/06/08

date of end of treatment: 29/07/ca

date of nacropsy : 30/07/08

MANDIBULAR both sides DISCOLOURATION DARK RED

LYMPH NODES

SKIN foreleg: left side ALOPECIA

abdominal region: ALOPECIA

foreleg: right side ALOPECIA

flank; right side ALOPECIA

flank; left side ALOPECIA

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

MANDIBULAR plasma cell infiltration (minimal)

LYMPH NODE congestion present (correlates to GROSS finding)

SKIN abdomen: epidermal hyperplasia (minimal) (correlates to GROSS finding)

foreleg right side: epidermal hyperplasia (minimal) (correlates to GROSS

finding)

flank left side: epidermal hyperplasia (minimal) (correlates to GROSS

finding)

VAGINA dioestrus present

epithelial vacuolation (minimal)

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, CERVIX, CLITORAL

GLAND, COLON, DUODENUM, HEART, ILEUM, JEJUNUM, KIDNEYS, LIVER, LUNG, LYMPH NODE MESENTERIC, OESOPHAGUS, OVARIES, PANCREAS, PARATHYROID, PEYERS

PATCHES, PITUITARY, RECTUM, SCIATIC NERVE, SPINAL CORD, SPLEEN, STERNUM,

STOMACH, THYMUS, THYROID, TRACHEA, URINARY BLADDER, UTERUS.
Page 65 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Aniimal No.: 4

SEX: Female

DOSE LEVEL: 1

MACROSCOPIC FINDINGS

days of treatment :43

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 29/07/08

date of necropsy : 30/07/08

No ebnormality detected

MICROSCOPIC FINDINGS

KIDNEYS tubular mineralization (minimal)

MANDIBULAR plasma cell infiltration (minimal)

LYMPH NODE

VAGINA dinestrus present

epithelial. vacuolation (minimal)

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, CERVIX, CLITORAL

GLAND, COLON, DUODENUM, HEART, ILEUM, JEJUNUM, LIVER, LUNG, LYMPH NODE

MESENTERIC, OESOPHAGUS, OVARIES, PANCREAS, PARATHYROID, PEYERS PATCHES,

PITUITARY, RECTUM, SCIATIC NERVE, SPINAL CORD, SPLEEN, STERNUM, STOMACH,

THYMUS, THYROID, TRACHEA, URINARY BLADDER, UTERUS.
Page 66 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-06

Sponsor: Color Pigments Manufacturers Association tnc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 49

SEX: Female

DOSE LEVEL: I

MACROSCOPIC FINDINGS

days of treatment 43

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment; 29/07/08

date of necropsy : 30/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 67 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 81 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 50

SEX: Female

DOSE LEVEL: 1

MACROSCOPIC FINDINGS

days of treatment 41

aacrif ice group terminal sacrif ice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 27/07/08

data of necropsy : 28/07/08

No abnormality detected

MICROSCOPIC FINDINGS

LIVER parenchymal inflammatory cell infiltration (minimal)

OESOPHAGUS muscularis inflammation (minimal)

PANCREAS focal acinar atrophy (minimal)

VAGINA dioestrus present

epithelial vacuolation (minimal)

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, CERVIX, CLITORAL

GLAND, COLON, DUODENUM, HEART, ILEUM, JEJUNUM, KIDNEYS, LUNG, LYMPH NODE

MESENTERIC, MANDIBULAR LYMPH NODE, OVARIES, PARATNYROID, PEYERS PATCHES,

PITUITARY, RECTUM, SCIATIC NERVE, SPINAL CORD, SPLEEN, STERNUM, STOMACH,

THYMUS, THYROID, TRACHEA, URINARY BLADDER, UTERUS.
Page 68 of 97

PATHOLOGY REPORT

Test substance: 0.5. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct.08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 51

SEX: Female

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment 44

sacrifice group terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 30/07/08

date of necropsy :31/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 69 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 52

SEX: Female

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment 43

sacrifice group :terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 29/07/08

date of necropsy :30/07/08

MANDIBULAR both sides DISCOLOURATION REDDISH

LYMPH NODES

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

MANDIBULAR congestion present (correlates to GROSS finding)

LYMPH NODE

No abnormalities found in: LYMPH NODE MESENTERIC,
PATHOLOGY REPORTPae7of9

Test substance: C.I. Pigment Blue 81 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 53

SEX: Female

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment :45

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 31/07/08

date of necropsy ; 01/08/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 71 of 97

PATHOLOGY REPORT

Test substance: C.l. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 54

SEX: Female

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment t 42

sacrifice group terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 28/07/08

date of necropsy 29/07/01s

No abnormality detected

MICROSCOPIC FINDINGS

VAGINA dioeatrua present

No abnormalities found in: CERVIX, CLITORAL GLAND, OVARIES, UTERUS.
Page 72 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 55

SEX: Female

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment :43

sacrifice group terminal sacrifice

necropsy status i planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 29/07/08

date of necropsy :30/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No abnormality detected

No abnormalities found in: LYMPH NODE MESENTERIC.
Page 73 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 56

SEX: Female

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment :41

sacrifice group terminal sacrifice

necropsy status :planned terminal

date of start of treatment; 17/06/08

date of end of treatment: 27/07/08

date of necropsy 28/07/08

MANDIBULAR both aides DISCOLOURATION DARK RED

LYMPH NODES

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

PANCREAS DISCOLOURATION REDDISH

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE sinus histiocytosis (slight)

MESENTERIC

MANDIBUL.AR congestion present (correlates to GROSS finding)

LYMPH NODE

PANCREAS no lesion to account for gross finding Is)
Page 74 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.! 57

SEX: Female

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment 43

sacrif ice group terminal sacrifice

necropsy status planned terminal

date of start of treatment; 17/06/0a

date of end of treatment: 29/07/08

date of necropsy :30/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No abnormality detected

Mo abnormalities found in: LYMPH NODE MESENTERIC.
Page 75 of 97

PATHOLOGY REPORT

Test substance: C.?. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 58

SEX: Female

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment 43 eacrifice group :terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 29/07/08

date of necropay 30/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No abnormality detected

No abnormalities found in: LYMPH NODE MESENTERIC.
Page 76 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 59

SEX: Female

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment :43

sacrifice group terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 29/07/08

date of necropsy :30/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues exami.ned
Page 77 of 97

PATHOLOGY REPORT

Test substance: C.l. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor~ Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 60

SEX: Female

DOSE LEVEL: 2

MACROSCOPIC FINDINGS

days of treatment :43

sacrifice group terminal sacrifice

necropsy status planned terminal

date of start of treatment: 27/06/08

date of end of treatment; 29/07/08

date of necropsy :30/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 78 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.; 61

SEX: Female

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 41

sacrifice group terminal aacrif ice

necropay status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 27/07/08

date of necropsy :28/07/08

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding Is) MESENTERIC
Page 79 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. : 62

SEX: Female

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 4 4

sacrifice group terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of and of treatment; 30/07/08

date of necropsy :31/07/08

No abnormality detected

MICROSCOPIC FINDINGS

No tissues examined
Page 80 of 97

PATHOLOGY REPORT

Test substance: C.t. Pigment Blue 81 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 63

SEX: Female

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment :41

sacrifice group :terminal sacrifice

necropsy statues planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 27/07/08

date of necropsy :28/07/08

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

Ho abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding Is)

MESENTERIC
Page 81 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 64

SEX: Female

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 41

sacrif ice group :terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 27/07/08

date of necropsy : 28/07/08

MESENT. LYMPH DISCOLOURATION BLUJISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE macrophage aggregates (minimal)

MESENTERIC no lesion to account for gross finding Is)
Page 82 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 65

SEX: Female

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment :41

sacrifice group :terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 27/07/08

date of necropsy 28/07/08

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding(s)

MESENTERIC
Page 83 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 66

SEX: Female

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 42

sacrifice group terminal sacrifice

necropsy status planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 28/07/08

date of necropsy :2 9/07/08

No abnormality detected

MICROSCOPIC FINDINGS

VAGINA metoestrus present

No abnormalities found in: CERVIX, CLITORAL GLAND, OVARIES, UTERUS.
Page 84 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 67

SEX: Female

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 43

sacrifice group terminal sacrifice

necropsy status planned terminal

date of start of treatmenti 17/06/08

data of end of treatment: 29/07/08

date of necropsy :30/07/08

MESENT. LYMPH DISCOLOURATION BLUISH NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding ls)

MESENTERIC
Page 85 of 97

PATHOLOGY REPORT

Test substance; C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. 68

SEX! Female

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment :43

sacrifice group terminal sacrifice

necropay atatus planned terminal

date of start of treatment: 17/06/08

date of end of treatment; 29/07/08

date of necropsy :30/07/08

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE sinus histiocytosia (minimal)

MESENTERIC no lesion to account for gross finding Is)
Page 86 of 97

PATHOLOGY REPORT

Test substance: C. LPigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.! 69

SEX: Female

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 43

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 29/07/08

date of necropsy 30/07/08

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross findingls)

MESENTERIC
Page 87 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 70

SEX: Female

DOSE LEVEL: 3

MACROSCOPIC FINDINGS

days of treatment 44

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 30/07/08

date of necropsy :31/07/08

MESENT. LYMPH DISCOLOU1RATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding Is)

MESENTERIC
Page 88 of 97

PATHOLOGY REPORT

Test substance: ClI. Pigment Blue 81 Notox No.: 488231

Test system: 25-Day Oral Toxicity Study in the Rat 24-Oct-OS

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.; 71

SEX: Female

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :45

sacrifice group :terminal sacrifice

necropey status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 31/07/08

date of necropsy :01/08/08

GENERAL GI-TRACT, CONTENTS BLUISH

OBSERVATIONS

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

SKIN FORELEG: right side ALOPECIA

HINDLEG: left side ALOPECIA

FORELEG: left side ALOPECIA

HINDLEG: right side ALOPECIA

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding Is)

MESENTERIC

SKIN foreleg right side: epidermal hyperplasia (minimal) (correlates to GROSS

finding)

foreleg left side: epidermal hyperplasia (minimal) (correlates to GROSS

finding)
Page 89 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 72

SEX: Female

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :42

sacrifice group :terminal sacrif ice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 28/07/08

date of necropay :29/07/08

GENERAL GI-TRACT, CONTENTS BLUISH

OBSERVATIONS

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

VAGINA mtoestrue preaent

No abnormalities found in: CERVIX, CLITORAL GLAND, OVARIES, UTERUS.
Page 90 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-OS

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 73

SEX: Female

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :43

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 29/07/08

date of necropsy :30/07/08

GENERAL 01 TRACT, CONTENTS BLUISH

OBSERVATIONS

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

JEJUNUM blue material in lumen present (GENERAL OBSERVATIONS) (correlates to GROSS

finding)

LYMPH NODE no lesion to account for gross finding Is)

MESENTERIC

MANDIBULAR plasma cell infiltration (minimal)

LYMPH NODE

VAGINA dioestrus present

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, CERVIX, CLITORAL

GLAND, COLON, DUODENUM, HEART, ILEUM, KIDNEYS, LIVER, LUNG, OESOPHAGUS,

OVARIES, PANCREAS, PARATNYROID, PEYERS PATCHES, PITUITARY, RECTUM,

SCIATIC HERVE, SPINAL CORD, SPLEEN, STERNUM, STOMACH, THYMUS, THYROID,

TRACHEA, URINARY BLADDER, UTERUS.
Page 91 of 97

PATHOLOGY REPORT

Test substance: C.l. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 74

SEX: Female

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

daya of treataent :41

sacrifice group i terminal sacrifice

necropay status :planned terminal

date of atart of treatment: 17/06/08

date of end of treatment; 27/07/08

date of necropay 28/07/08

ADRENAL GLANDS both aides DISCOLOURATION RED-BROWN

GENERAL GI TRACT, CONTENTS BLUISH

OBSERVATIONS

MANDIBULAR both sides DISCOLOURATION REDDISH

LYMPH NODES

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

Ho abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

ADRENAL no lesion to account for gross finding~s)

HEART epicardial fibrosis (minimal)

ILEUM blue material in lumen present (GENERAL OBSERVATIONS) (correlates to GROSS

finding)

LYMPH NODE no lesion to account for gross finding (s)

MESENTERIC

MANDIBULAR congestion present (Correlates to GROSS finding)

LYMPH NUDE

RECTUM blue material in lumen present (GENERAL OBSERVATIONS) (correlates to GROSS

finding)

SPLEEN extramedullary haematopoiesis (slight)

THYMUS involution/atrophy (minimal)

VAGINA dioeatrus present

epithelial vacuolation (alight)

No abnormalities found in: AORTA, BRAIN, CAECUM, CERVIX, CLITORAL GLAND,

COLON, DUODENUM, JEJUNUM, KIDNEYS, LIVER, LUNG, OESOPHAGUS, OVARIES,

PANCREAS, PARATHYROID, PEYERS PATCHES, PITUITARY, SCIATIC NERVE, SPINAL

CORD, STERNUM, STOMACH, THYROID, TRACHEA, URINARY BLADDER, UTERUS.
Page 92 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 75

SEX: Female

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment 41

sacrifice group :terminal secrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 27/07/08

date of necropsy :28/07/08

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

KIDNEYS basophilic tubules (minimal)

LYMPH MODE no lesion to account for gross finding Is)

MESENTERIC

MANDIBULAR plasma cell infiltration (slight)

LYMPH NODE

VAGINA dioestrus present

epithelial vacuolation (slight)

No abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, CERVIX, CLITORAL

GLAND, COLON, DUODENUM, HEART, ILEUM, JEJUNUM, LIVER, LUNG, OESOPHAGUS,

OVARIES, PANCREAS, PARATHYROID, PEYERS PATCHES, PITUITARY, RECTUM,

SCIATIC NERVE, SPINAL CORD, SPLEEN, STERNUM, STOMACH, THYMUS, THYROID,

TRACHEA, URINARY BLADDER, UTERUS.
Page 93 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study In the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 76

SEX: Female

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :42

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 28/07/08

date of necropsy :29/07/08

GENERAL 01 -TRACT, CONTENTS BLUISH

OBSERVATIONS

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

COLON blue material in lumen present (GENERAL OBSERVATIONS) (correlates to GROSS

finding)

LYMPH NODS no lesion to account for gross finding Is)

MESENTERIC

MANDIBULAR plasma cell infiltration (minimal)

LYMPH NODE

VAGINA dioestrus present

epithelial vacuclation (slight)

Ho abnormalities found in: ADRENAL, AORTA, BRAIN, CAECUM, CERVIX, CLITORAL

GLAND, DUODENUM, HEART, ILEUN, JEJUNUM, KIDNEYS, LIVER, LUNG, OESOPHAGUS,

OVARIES, PANCREAS, PARATHYROID, PEYERS PATCHES, PITUITARY, RECTUM,

SCIATIC NERVE, SPINAL CORD, SPLEEN, STERNUM, STOMACH, THYMUS, THYROID,

TRACHEA, URINARY BLADDER, UTERUS.
Page 94 of 97

PATHOLOGY REPORT

Test substance: 0.1. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No. : 77

SEX: Female

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :41

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 27/07/08

date of necropay :28/07/08

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

KIDNEYS tubular mineralization (minimal)

LUNG perivescular inflammatory cell infiltration (minimal)

alveolar macrophage infiltration (minimal)

LYMPH NODE no lesion to account for gross finding(s)

MESENTERIC

VAGINA dioestrus present

epithelial vacuolation (minimal)

No abnormalities found in: ADRENAL, AORTA, BRAIN, CARCUN, CERVIX, CLITORAL

GLAND, COLON, DUODENUM, HEART, ILEUM, JEJUNUM, LIVER, MANDIBULAR LYMPH

NODE, OESOPHAGUS, OVARIES, PANCREAS, PARATHYROID, PEYERS PATCHES,

PITUITARY, RECTUM, SCIATIC NERVE, SPINAL CORD, SPLEEN, STERNUM, STOMACH,

'THYMUS, THYROID, TRACHEA, URINARY BLADDER, UTERUS.
Page 95 of 97

PATHOLOGY REPORT

Test substance: C. LPigment Blue 81 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 78

SEX: Female

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :45

sacrifice group :terminal sacrifice

necropsy status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 31/07/08

date of necropsy :01/08/08

GENERAL GE-TRACT, CONTENTS BLUISH

OBSERVATIONS

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding ls)

MESENTERIC
Page 96 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 79

SEX: Female

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :44

aacrifice group :terminal sacrifice

necropsy atatus planned terminal

date of atart of treatment: 17/06/08

date of end of treatment: 30/07/08

date of necropay :31/07/08

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tiasues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding 1s)

MESENTERIC
Page 97 of 97

PATHOLOGY REPORT

Test substance: C.I. Pigment Blue 61 Notox No.: 488231

Test system: 28-Day Oral Toxicity Study in the Rat 24-Oct-08

Sponsor: Color Pigments Manufacturers Association Inc.

TABLE 3 - INDIVIDUAL ANIMAL DATA

Animal No.: 80

SEX: Female

DOSE LEVEL: 4

MACROSCOPIC FINDINGS

days of treatment :44

sacrifice group :terminal sacrifice

necropay status :planned terminal

date of start of treatment: 17/06/08

date of end of treatment: 30/07/Ga

date of necropsy 31/07/0a

GENERAL GI TRACT, CONTENTS BLUISH

OBSERVATIONS

MESENT. LYMPH DISCOLOURATION BLUISH

NODE

No abnormalities were found in any of the other tissues examined

MICROSCOPIC FINDINGS

LYMPH NODE no lesion to account for gross finding is)

MESENTERIC
C.I. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 5

APPENDIX 5 SUMMARY DOSE RANGE FINDING STUDY

Page 1 of 2
C. PIGMENT BLUE 61 NOTOX Project 488231

APPENDIX 5

SUMMARY OF DOSE RANGE FINDING STUDY

In order to set the dose levels for the main study (NOTOX Project 488231), a pilot study was

performed.

Two groups of 3 females (10 weeks old) were dosed at 500 or 1000 mg/kg body weight/day

(Groups 1 and 2) for 5 days by oral gavage with a dose volume of 5 mI/kg body weight. Dose

levels were based on three non-GLP acute oral studies in which the LD50 values were > 15,000

mg/kg (SPF Wistar rats), >15,000 mg/kg (SPF Wistar rats) and >5,000 mg/kg (Sprague Dawley rats). In a non-GLP repeat dose study, mixed race Albino rats were exposed for 14 days at 500

mg/kg: no toxicity was noted.

All animals were housed 3 per cage, identified by ear- and tailmark and allocation was

performed at random. If not mentioned otherwise, test system and techniques were identical to

those used during the main study.

Data was collected under NOTOX Project 488232.

Planning Delivery: 28 May 2008

Treatment: 02 to 06 June 2008

Terminal procedures: 06 June 2008

Animal husbandry Animals were housed in a controlled environment, in which optimal conditions were considered to be approximately 15 air changes per

hour, a temperature of 21 ± 30C (actual range: 19.8 - 2 1.1 *0), a

relative humidity of 30 - 70% (actual range: 50 - 81 %) and 12

hours artificial light and 12 hours darkness per day. Cleaning

procedures in the room might have caused the temporary

fluctuations above the optimal level of 70% for relative humidity.

Based on laboratory historical data these conditions were

considered not to have affected the study integrity.

Chemical analysis Not performed during the pilot study.

Observations Clinical signs: At least once daily.

Mortality: At least twice daily.

Body weights: Daily.

Food consumption: Daily.

Pathology Necropsy: On Day 5

No organs were fixed.

Terminal body weight, organ weights: kidney and liver.

RESULTS

No mortality was observed in the pilot study, and there were no differences in body weights

(absolute or relative), food consumption (absolute or relative), or organ weights (absolute or

relative). For animals treated with 500 and 1000 mg/kg of 0.1. Pigment Blue 61, blue coloration

of the faeces and bluish contents in the GI tract were noted.

CONCLUSION

There were no toxicologically-significant results for treatment up to 1000 mg/kg; based on the

results of the pilot study the doses selected for the main study are: 50, 250, and 1000 mg/kg.

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