### DISCLAIMER This report is the property ofClariant. Its content is confidential and not for general release. It is not allowed to cite and reveal the report or its content to a third party without prior written approval of Clariant. 

## REPORT 

## 31-15153 

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Leuco Sulfur Black 1: Determination ofthe Water Solubility

DataRequirements I Test Guidelines:

StudyDirector: Test Facility:

Sponsor: OECDNo.l05 EEC Directive 92/69, A.6 Council Regulation (EC) 44012008, A.6

Dr Martin Kiihne Harlan Laboratories Ltd. Zelgliweg 1 4452 Itingen I Switzerland

Clariant Products (Switzerland) Ltd. Rothausstrasse 61 4132 Muttenz 1 I Switzerland StudyIdentification: Harlan Laboratories Study C75560
Version: Final Study Completion Date: March 11, 2010 Page 1of 16Harlan Laboratories Study C75560 Leuco Sulfur Black I 

## CONTENTS Report Page2

CONTENTS ....................................................................................................................................2 LIST OF TABLES ..........................................................................................................................2 GLP CERTIFICATE....................................................................................................................... 3 STATEMENT OF COMPLIANCE................................................................................................4 QUALITY ASSURANCE STATEMENT......................................................................................5 PREFACE .......................................................................................................................................6 General Information...............................................................................................................6 Responsibilities......................................................................................................................6 Schedule.................................................................................................................................6 Data Requirements ITest Guidelines ....................................................................................7 Archiving ...............................................................................................................................7 1 SUMMARY........................................................................................................................... 8 2 PURPOSE..............................................................................................................................9 3 MATERIALS AND METHODS......................................................................................... 10 3.1 Test Item ................................................................................................................ 10 3.2 Test System............................................................................................................ 10 3.3 Reagents and Apparatus ........................................................................................ 11 3.4 Principle................................................................................................................. 11 3.5 Performance ofthe Test......................................................................................... 12 3.6 Analytical Method ................................................................................................. 12 3.6.1 System Suitability check ......................................................................... 12 3.7 Data Handling........................................................................................................ 13 4 RESULTS AND DISCUSSION.......................................................................................... 14 5 CONCLUSION.................................................................................................................... 15 6 TABLES .............................................................................................................................. 16 

## LIST OFTABLES Table 1 Data ofthe Performance ofthe Water Solubility Test................................................ 16 Table 2 Results ofthe Water Solubility ofLeuco Sulfur Black 1 ........................................... 16**-~v·1( V~t·!·~** **.ioJ~ ~1**  '¥· ·.'Harlan Laboratories Study C75560 Leuco Sulfur Black 1 

## GLPCERTIFICATE

" S<-oscho !ill-scllall V CoftfHHatioft wiue ConfHif.UiOM S•tn•• Conftdlf.,_. mu• Report



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# __ Statement ofGLP Compliance Acoonling to Miele 14 paragraph 3Ordinance on Good Laboratory Pradlce (OGLP. SR 813.112.11 The nolillcltion IIUihorily for chemicals confilml that till following test facility was inspected with respect to tillcomplilnce with tillSwisS Ordinance on Good L.aborllory Pradlce, adopted on 18th May 2005 [OGLP. SR 813.112.11. This Ordinance is based on till

### oeco Principles of Good Laboralary Practice, as reviled in 1997 and adopt8d on 26th November 1997 by decision of lhe OECD Council [C(97)1861finaq. Unequivocal name ll1d add,_ oftillliltfacility: Harlan Llboratorles Ltd. Zelgliweg 1 4452 ltingen, Switzelllnd. Atusofexpertise ICCOrding to lfllcle 3**par~graph** 1leu.d OGLP: 1JPhysicak:hernlcal testing, 2J T oxldty studies, 41 Environmental toxicity studies on aquatic and terrestrial organilms, 51 Studies on behaviourin water. 101 and air; bioaccumulltlon, 61 Relldue studies, 7.IStudies on etr1c:1s on I'II8IIOCOIIIW and naturllecosylllml, 61 Analytic:IIIIICI clinical c:hemist1y tedng, 9./Otherstudies (ur.ty pl\8miiCCIIclgy and animll melabollsm). Inspection authority: Federal Olllce fortillEnvironment (FOEN) I Federal Olllce ofPublic Heelltl (FOPH) I SwissAtJencyfor Thenlpeutic Products (Swiarnedic) Date of inspection: 05th to 09lh and 26th to 30th November 2007 Date ofdecision: 30th Aplil2008 Based on the above mentionad decision it can be confirmed that till above mentioned test facitity is able to conduct studies acc:ording to the aforementioned area of expertise in compliance wilh the principles of GLP. The above mentioned test facility is listed In till register and GLP list acc:ording to the Miele 14 OGLP and is inspected on a regular basis according to Article 6 paragraph 2 OGLP.**TN~.....,_totCIWII-•** ..**~INllilliiiiDn.._.,..,** ...........,.....IOL")IIII'
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### GOOD LABORATORY PRACTICE Report



## STATEMENT OF COMPLIANCE Harlan Laboratories Study: Test Item: Study Director: C75560 Leuco SulfurBlack I DrMartin Kiihne Page4

Study Title: Leuco Sulfur Black 1: Determination of the Water Solubility This study has beenperformed in compliance withthe: Swiss Ordinance relating to Good Laboratory Practice adopted May 

### 18th, 2005 [SR 813.112.1]. This Ordinance is based on the OECD Principles of Good Laboratory Practice, as revised in 1997 and adopted on November 26th, 1997 bydecision ofthe OECD Council [C (97)186/Final]. These principles are compatible with Good Laboratory Practice regulations specified by regulatory authorities throughout the European Community, the United States (EPA and FDA), and Japan (MHW, MAFF and MET!). There were no circumstances that mayhave affected the qualityorintegrity ofthe data. StudyDirector: DrMartin Kiihne

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## 2o 10 IHarlan Laboratories Study C75560 Leuco SulfurBlack l Report Page 5 

## QUALITYASSURANCE STATEMENT Harlan Laboratories Ltd., Zelgliweg 1, 4452 ltingen I Switzerland Harlan Laboratories Study: Test Item: StudyDirector: Study Title: C75560 Leuco Sulfur Black 1 DrMartin KUhne Leuco Sulfur Black 1: Determination of the Water Solubility The general facilities and activities are inspected at least once a year and the results are reported to the responsible person and the management. Studyprocedures wereperiodically inspected. The study plan and this report were auditedbythe Quality Assurance. The dates are givenbelow. Dates ofReports to the Study Dates and Types ofQAInspections Director and Test Facility Management February 10, 2010 Studyplan February 10, 2010 February 12, 2010 Processbased February 12, 2010 (Work-up) March 09,2010 Report March 09,2010 This statement also confirms that this final report reflects the raw data. QualityAssurance:

# )~ Ms Margot Richter-AuerHarlan Laboratories Study C75560 Leuco Sulfur Black l 

## PREFACE General Information Test Item: Study Title: Sponsor: Study Monitor: Test Facility: QA:



### Responsibilities Study Director: Deputy StudyDirector: Reporting: Technical Coordinators: Head ofQA: 

### Schedule Experimental Starting Date: Experimental Completion Date: Study Completion Date: Report

Leuco SulfurBlack 1 Leuco Sulfur Black 1: Determination ofthe Water Solubility Clariant Products (Switzerland) Ltd. Rothausstrasse 61 4132 Muttenz 1 I Switzerland DrBruno Kunz Harlan Laboratories Ltd. Zelgliweg 

### 1 4452 Itingen I Switzerland HarlanLaboratories Ltd. Quality Assurance GLP Zelgliweg 1 4452 Itingen I Switzerland

DrMartin Kiihne DrCarsten Habeck Ms Christiane Nestler Ms Sabine Schlachter, MrJohann Martens and Ms BarbaraFliikiger MrTobias Fink

February 15, 2010 February 19,2010 March 11, 2010 Page6Harlan Laboratories Study C75560 Leuco Sulfur Black I Report


### Data Requirements 

### ITest Guidelines Page 7

The study procedures indicated in this report meet or exceed the requirements ofthe following guidelines:
• OECD Guideline for Testing of Chemicals, No. 105, "Water solubility"; adopted July 27, 1995. • EEC Directive 92/69, Part A, Methods for the determination of physico-chemical properties, A.6: "Water solubility", EEC Publication No. L383, December 1992.
• Council Regulation (EC) No. 440/2008, 30 May 2008, Part A, Methods for the determination ofphysico-chemical properties, A.6 "Water Solubility" 

### Archiving Harlan Laboratories Ltd. (4452 Itingen I Switzerland) will retain the study plan, all raw data, a sample oftest item(s), and the finalized report ofthe present study for at least ten years. No data will be discarded without the. Sponsor's written consent.Harlan Laboratories Study C75560 Leuco SulfurBlack I 

## 1 SUMMARY Report Page 8

During this study the water solubility of Leuco Sulfur Black 1 was determined. Vessels containing water and test item were shaken for 24 hours, 48 hours and 72 hours, respectively, at 30 °C. Afterwards the samples were equilibrated for another 24 hours at 20.0 °C. After dilution with water the concentrations in the aqueous solutions were determined bya TC-method. The water solubility ofLeuco Sulfur Black 1was determined to be 62.4 mg/L at 20.0 °C and a pH of7.9 based on a measured concentration of11.5 mg/L ofcarbon using the flask shaking method.Harlan Laboratories Study C75560 Leuco Sulfur Black 1 

## 2 PURPOSE Report Page 9

The purpose ofthis study was to evaluate the solubility ofLeuco Sulfur Black 1 in water, which is specified by the saturation mass concentration ofthe test item in water at a given temperature.Harlan Laboratories Study C75560 Leuco Sulfur Black I Report Page 10 

## 3 MATERIALS AND METHODS 3.1 Test Item Data oftest item as supplied by the Sponsor. Identification: CASNr.: Description: Batch Number: Composition:

Stability ofthe test item: Stability ofthe test item dilution: Expiry date: Storage Conditions: 

### 3.2 Test System Leuco Sulfur Black 1 66241-11-0 Thixotrophic, black, wet presscake ESA0007595 Leuco Sulfur Black 1: 52.75% (calculated by amount of by-compounds) 2,4-Dinitrochlorobenzene: < 20 ppm 2,4-Dinitrophenole: < 20 ppm Sulfate: 970 ppm Chloride: 750 ppm Sodium: 1.3% (w/w) Water: 45.75% (w/w) by Karl-Fischer titration Stable under storage conditions > 4 h in water July 11, 20 11 At room temperature (range 20 

### +!- 5 °C), light protected

The test system consists ofdouble distilled water. Rationale: The test systems and test methods are recommended by international guidelines.Harlan Laboratories Study C75560 Leuco Sulfur Black I 

### 3.3 Reagents and Apparatus 1,4-Benzochinone Saccharose Water (used for dilution) Water (used for sample preparation) FilterNylon (0.2 f.tm) Balances 

### TC pH-meter ShakingWater Bath


### 3.4 Principle Report

Fluka Riedel de Haen Harlan Laboratories Harlan Laboratories BGB Mettler Toledo Shimadzu Metrohm 654 Haake Infors AG Art. No. 12309 Art. No. 16104 Page II

from Milli-Q water supply double distilled Art. No. F2503-1 XP205DR Analyzer 5000A Metrohm electrode 6.0232.100 SWB25 Aquatron

An excess amount of the test item is brought into contact with water. When saturation is achieved, the mass concentration of the test item in the aqueous solution is determined by a suitable analytical method. To determine the water solubility ofthe test item, two different methods can be used:

- the column elution method

- the flask method Due to the properties ofthe test item the flask method was used during this study.Harlan Laboratories Study C75560 Leuco Sulfur Black I 

### 3.5 Performance ofthe Test Test Solutions Report Page 12

About 1.5
- 1.7 g oftest item were weighed into each of 6 Erlenmeyer flasks and 50 mL of double distilled water were added to each flask. The tightly closed flasks were shaken at about 30 °C for 24 hours, 48 hours and 72 hours, respectively. Afterwards, the flasks were equilibrated for another 24 hours at 20.0 °C. The pH was measured to be about 7.9. Afterwards, the supernatant solutions were centrifuged**(~2900** g for 10 minutes), filtered (0.2 Jlm, Nylon) and 5 mL ofthis solution was diluted up to 50 mL with water prior to analysis. The quantification was performed using TC measurement as described below. Details ofsample treatment are given in Table 1. 

### 3.6 Analytical Method The concentration ofthe test item was analyzed using TC measurement. The test samples were quantified with reference to a calibration curve, which was generated automatically. The last calibration was performed on January 06, 2010. 3.6.1 System Suitability check1 A system suitability check was performed before sample measurement in order to check the reliability ofthe measurement. Preparation of 1,4-Benzochinone Solutions 75.02 mg of 1,4-benzochinone were filled up to a volume of 1 L with Milli-Q water to get a stock solution ofnominal concentration of 50 ppm of carbon. 2 mL ofthis stock solution was diluted to a volume of200 mL with Milli-Q water to get a solution ofa nominal concentration of 0.5 ppm of carbon. A second solution of a nominal concentration of 10 ppm of carbon was prepared by diluting 20 mL ofthe stock solution up to 100 mL with Milli-Q water. For both solutions, the TOC was measured. The system suitability check was successfully as all TOC values were within the specifications. Preparation ofSaccharose Solutions 118.99 mg of saccharose were filled up to a volume of 1 L with Milli-Q water to get a stock solution ofnominal concentration of50 ppm ofcarbon. 2 mL ofthis stock solution was diluted to a volume of 200 mL with Milli-Q water to get a solution of a nominal concentration of 0.5 ppm of carbon. Afterwards, the TOC was measured. The system suitability check was successfully as the TOC value was within the specifications.

1 Was performed in Study C79744Harlan Laboratories Study C75560 Leuco Sulfur Black 1


### 3.7 Data Handling Report Page 13

The samples were quantified by Total Carbon (TC) measurement with reference to a calibration curve. The calibration curve was calculated using the software. The concentration ofcarbon of the injected samples was quantified by peak area with reference to the calibration curve. From the results obtained by the software the concentration ofcarbon in the analyzed solutions was calculatedbythe following equation: (1) Where C 1 Measured concentration ofcarbon in the sample solution [ppm= mg/L] 

### Cz Concentration ofcarbon in the test item solution before dilution [mg/L] f Dilution factor From the concentration of carbon the water solubility of the test item was calculated by the following equation: (2) Where C3 Water solubility ofthe test item [mg/L] Cc Percentage of carbon in the test item as obtained by elementar analysis (here: 18.5%)Harlan Laboratories Study C75560 Leuco SulfurBlack I Report 

## 4 RESULTS AND DISCUSSION Page 14

The determination ofthe water solubility ofLeuco Sulfur Black 1 was performed according to the EEC Directive 92/69, A.6 "Water Solubility" (1992) and the OECD guideline No. 105, "Water Solubility" (1995) using the flask shaking method. After a shakingtime of24 hours, 48 hours and 72 hours, respectively, at 30 °C, the samples were equilibrated for another 24 hours at 20 °C. Afterwards, the samples were centrifuged, filtered, diluted and analyzed by TC measurement. The water solubility ofLeuco Sulfur Black 1 at 20 °C was determined to be 62.4 mg/L. The measured results differ by less than 15%. Therefore, the study is considered to be valid. The results ofthe individual measurements ofthe main test are summarized in Table 2. The tabulated values represent rounded results, which were obtained by calculation using the exact raw data.Harlan Laboratories Study C75560 Leuco Sulfur Black I 

## 5 CONCLUSION Report Page 15

During this study the water solubility ofLeuco Sulfur Black 1was determined and found to be 62.4 mg/L at 20 °C and a pH of7.9 based on a measured concentration of11.5 mg/L ofcarbon using the flask shaking method.Harlan Laboratories Study C75560 Leuco Sulfur Black I 

## 6 TABLES Report

Table 1 Data ofthe Performance ofthe Water Solubility Test Sample Solution No. 1 2 3 Page 16

4 5 6 Weighed sample ofthe test 1482.52 1674.48 1473.65 1621.87 1734.69 1583.06 item [mg] Added volume ofwater [mL] 50 50 50 50 50 50 Agitation time at 30

## oc 72 72 48 48 24 24 [hours] Equilibration time at 20 °C 24 24 24 24 24 24 [hours] pH value oftest item solution 7.60 7.52 7.68 7.76 8.32 8.29 Dilution factor before 10 10 10 10 10 10 quantification
Table 2 Results ofthe Water Solubility ofLeuco Sulfur Black 1 Concentra- Concentra- Concentra- Water Sample No. Shaking tion of tion of tion oftest solubility time at 30 °C carbon, carbon, item, calculated analyzed calculated calculated (rounded)

[mg!L] [mg/L] [mg/L] [mg/L] 1 72 1.094 10.94 59.14 2 1.400 14.00 75.68 3 1.147 11.47 62.00 62.35 48 SD = 6.90 4 1.104 11.04 59.68 5 24 1.143 11.43 61.78 6 1.033 10.33 55.84 The saturation mass concentration is calculated from the bold values SD = standard deviation 

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