TSCA NON-CONFIDENTIAL BUSINESS INFORMATION

DOCUMENT DESCRIPTION DOCUMENT CONTROL NUMBER DATE RECEIVED

COMMENTS: DEC LASS

DOUS NOT CONTAIN CBI
UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTON D.C., 20460

OFFICE OF

~ PRO~CHEMICAL SAFETY AND

POLLUTION PREVENTION

Notice of Change in CBI Chemical Identity Status

Pre-Manufacture Notice (PMN) P-99-0669

Notice of commencement (NOC) received: 12/22/1999

Please be advised that the Confidential Business Information (CBI) claim for the

above referenced substance was dropped by the information submitter in the

NOC.

The identified non-confidential chemical name for this substance as provided in

the NOC is:

OXIRANE, METHYL-, POLYMER WITH OXIRANE, MONO(3,5,5-

TRIMETHYLHEXYL)ETHER

The current TSCA Inventory name is

Oxirane, 2-methyl-, polymer with oxirane, mono(3,5,5-trimethylhexyl) ether

CASRN: 204336-40-3

This notice as submitted to the NCIC files on 9/1/2011

Name of staffer: Jeffrey Santacroce

Position: Asst. NOC Coordinator

Branch: RDMB/IMD/OPPT/OCSPP

CONTAINS NO CB1
____________________________________________________ Form Approved. 0.MB. No. 2070-0012. Approval Expires 10-31-96.

U.S. ENVIRONMENTAL PROTECTION AGENCY AGENCY USE ONLY Date of receipt

~ EPA FORPREMANUFACTURE SntzdTI75

FRNEW CHEMICAL SUBSTANCES

DOCUMENT CONTROL OFFICER CompanySaize OFFICE OF POLLUTION PREVENTION

AND TOXIC SUBSTANCES, 7407

U.S. E.P A. 401 M STREET, SW ii i ~~k WASHINGTON, D.C.,06,

Enter the total number of pa~ges 175 Docu. et control number PA case numb r

in the Premanufacture Notice I O--r P 7

GENERAL INSTRUCTIONS TS -j"1jCLjC~jS j0 ' " You must provide all information requested in this form to the extent that it is known to or reasonably ascertainable by you. Make reasonable estimates if you do not

have actual data.

" Before you complete this form, you should read the "Instructions Manual for Premtanufacture Notification" (the Instructions Manual is available from the Toxic

Substances Control Act (TSCA) Information Service by calling 202-554-1404, or faxing 202-554-5603).

* If a user fee has been remitted for this notice (40 CFR 700.45), indicate in the boxes above the TS-user fee identification number you have generated. Remember,

your user fee ID number must also appear on your corresponding fee remittance, which is sent to EPA, HQ Accounting Operations Branch (PM-226), P.O. 360399M,

Pittsburgh, PA 1525 1-6399, Attn. TSCA User fee.

Part I - GENERAL INFORMATION TEST DATA AND OTHER DATA

You must provide the currently correct Chemical Abstracts (CA) Name of the new You are required to submit all test data in your possession or control and to provide a chemical substance, even if you claim the identity as confidential You may authorize description of all other data known to or reasonably ascertainable by you, if these

another person to submit chemical identity information for you, but your submission will data arc related to the health and environmental effects on the manufacture, not be complete and the review will not begin until EPA receives this information. A processing, distribution in commerce, use, or disposal of the new chemical

letter in support of your submission should reference your TS user fee identification substance. Standard literature citations may be submitted for data in the open number. You must submit an original and two copies of this notice including all test data.

If you clainmed any information s confidential, a single sanitized copy must also be scientific literature. Complete test data (written in English), not summaries of data,

submitted. must be submitted if they do not aooear in the open literature. You should clearly

identify whether test data is on the substance or on an analog. Also, the chemical

composition of the tested material should be characterized Following are examples

Part RI - HUMAN EXPOSURE AND ENVIRONMENTAL RELEASE of test data and other data. Data should be subitted according to the requirements

of §720.50 of the Premanufacture Notification Rule (40 CFR Part 720).

If there are several manufacture, processing, or use operations to be described in Part 11,

sections A and B of this notice, reproduce the sections as needed.

Part IlIl- LIST OF ATITACHMENTS Test Data (Check Below any included in this notice)

Attach additional sheets if there is not enough space to answer a question fully. Label *Environmental fate data 0 Yes *Other data [: Yes

each continuation sheet with the corresponding section heading. In Part III, tist these

attachments, any test data or other data and any optional information included in the *Health effects data 0 Yes Risk assessments

notice.

*Environmental effects data 0 Yes Structure/activity relationships

* Physi cal/Chiemical Properties* 0 Yes Test data not in the possession

OPTIONAL INFORMATION or control of the submitter

You may include any information that you want EPA to consider in evaluating the new *A physical and chemical properties worksheet is located on the last page of this form.

substance. On page I I of this form, space has been provided for you to described

pollution prevention and recycling information you may have regarding the new TYPE OF NOTICE (Check Only One)

substance.

So-called "binding" boxes are included throughout this form for you to indicate your PMN (Premanufacture Notice)

willingness to be bound to certain statements you make in this section, such as use, production volume, protective equipment .. . This option is intended to reduce delays Li INTERMEDIATE PMN (submitted in sequence with final product PMN)

that routinely accompany the development of content orders or Significant New Use

Rules. Except in the case of exemption applications (such as TMEA, LyE, LOREX) r

where certain information provided in such notification is binding on the submitter when SNUN (Significant New Use Notice)

the Agency approves the exemption application, checking a binding box in this notice t.0~

does not by itself prohibit the submitter from later deviating from the information (except L TMEA (Test Marketing Exemption Application)

chemical identity) reported in the form

ED LVE (Low Volume Exemption) @4 40 CFR 723.50(c)(1) j~

CONFIDENTIALITY CLAIMS D LOREX (Low Release/Low Exposure Exemption) @ 40 CFR 723.5d (2) You may claim any information in this notice as confidential. To assert a claim on the

form mark (X) the confidential box next to the information that you claim as mr

confidential. To assert a claim in an attachment, circle or bracket the information you L.J LVE Modification ~ j LOREX Modification<

claim as confidential. If you claim information in the notices as confidential. you must

also provide a sanitized version of the notice. (includiniz attachments). For additional IS THIS A CONSOLIDATED PMN? L Yes

instructions on claiming information as confidential, read the Instructions Manual.

# of chemicals _________

Z Mark (x) if any information in this notice is claimed as confidential. (Prenotice Communication # required, enter # on page 3)

EPA FORM 7710-25 (Rev. 5-95) Replaces previous editions of EPA Form 7710-25.
Public reporting burden for this collection of information is estimated to average 110 hours per response, including time for reviewing instructions,

searching existing data sources, gathering and maintaining the data needed, and completing and reviewing the collection of information. Send

comments regarding the burden estimate or any other aspect of this collection of information, including suggestions for reducing this burden, to

Chief, Information Policy Branch, PM-223, U.S. Environmental Protection Agency. 401 M. St.. S.W., Washington, D.C. 20460; and to the Office

of Management and Budget, Paperwork Reduction Act (2070-00 12), Washington, D.C. 20503.

CERTIFICATION

I certify that to the best of my knowledge and belief:

1 . The company named in Part 1, section A, subsection I a of this notice form intends to manufacture or import for a

commercial purpose, other than in small quantities solely for research and development, the substance identified in Part 1,

Section B.

2. All information provided in this notice is complete and truthful as of the date of submission.

3. 1 am submitting with this notice all test data in my possession or control and a description of all other data known to or

reasonably ascertainable by me as required by §720.50 of the Premanufacture Notification Rule.

Additional Certification Statements:

If you are submitting a PMN, Intermediate PMN, Consolidated PMN, or SNUTN, check the following user fee certification

statement that applies:

Z The Company named in Part 1, Section A has remitted the fee of $2500 specified in 40 CFR 700.45(b), or

LIThe Company named in Part 1, Section A has remitted the fee of $1000 for an Intermediate PMN (defined@ @40 CFR

700.43) in accordance with 40 CFR 700.45(b), or

LiThe Company named in Part I Section A is a small business concern under 40 CFR 700.43 and has remitted a fee of $100 in

accordance with 40 CFR 700.45(b).

If you are submitting a low volume exemption (LVE) application in accordance with 40 CFR 723.50(c)(1) or a Low release

and low exposure exemption (LoRex) application in accordance with 40 CFR 723.50(c)(2), check the following certification

statements:

LIThe manufacturer submitting this notice intends to manufacture or import the new chemical substance for commercial

purposes, other than in small quantities solely for research and development, under the terms of 40 CFR 723.50.

LIThe manufacturer is familiar with the terms of this section and will comply with those terms; and

LIThe new chemical substance for which the notice is submitted meets all applicable exemption conditions.

LIIf this application is for an LVE in accordance with 40 CFR 723.50(c)(1), the manufacturer intends to commence

manufacture of the exempted substance for commercial purposes within I year of the date of the expiration of the 30 day

review period.

The accuracy of the statements you make in this notice should reflect your best prediction of the anticipated facts regarding the chemical substance

described herein. Aqxkowing and willfullmisinterpretation is subject to criminal penalty pursuant to 18 USC 1001. Confidential

Signature and Wttf tho ffc ~al au equired) Date

R s4 ait an e ft Etnvironmental Compliance

Signature of agent (if appicable) Date

EPA FORM 7710O25 (Rev. 5-95) Page 2
Part I -- GENERAL INFORMATION

Section A -- SUBMITTER IDENTIFICATION ConfiMark () the "Confidential" box next to any subsection you claim as confidential dential

l a Person Name of authorized official Position

Submitting

Notice (in U.S.) Russell B. Mait Manager, Environmental & Safety Compliance

Company

Goldschmidt Chemical Corporation

Mailing address (number and street)

P.O. Box 1299, 914 E. Randolph Rd.

City, State, ZIP Code

Hopewell, VA 23860

b. Agent (if Name of authorized official Position

applicable)

Company

Mailing address (number and street)

City, State, ZIP Code Telephone Area Code Number

c. If you are submitting this notice as part of a joint submission, mark (X) this box ) I

Joint Submitter (if Name of authorized official Position

applicable)

Company

Mailing address (number and street)

City, State, ZIP Code Telephone Area Code Number

2. Technical Name of authorized official Position

Contact (inI

U.S.) Russell B. Mait Manager, Environmental & Safety Compliance

Company

Goldschmidt Chemical Corporation

Mailing address (number and street)

P.O. Box 1299, 914 E. Randolph Road

City, State, ZIP Code Telephone Area Code Number

Hopewell, VA 23860 800 446-1809

3. If you have had a prenotice communication (PC) concerning this notice

and EPA assigned a PC Number to the notice, enter the number Mark (X) -0

if none

4. If you previously submitted an exemption application for the chemical

substance covered by this notice, enter the exemption number assigned by Mark (X) - *

EPA. If you previously submitted a PMN for this substance enter the inn

PMN number assigned by EPA (i.e withdrawn or incomple-i nn

5. If you have submitted a notice of Bona fide intent to manufacture or import

for the chemical substance covered by this notice, enter the notice number Mark (X) . - *

assigned by EPA. if none

1. E: Manufacture 2 Import 6. Type of Notice - Mark (X) Only Only 3. Both

D] Binding Option D Binding Option

Mark WX

EPA FORM 7710-25 (Rev. 5-95) Page 3
Part I -- GENERAL INFORMATION -- Continued

Section B -- CHEMICAL IDENTITY INFORMATION: You must provide a currently correct Chemical Abstracts (CA) name of the substance based on the ninth Collective Index (9Cl) of CA nomenclature rules and conventions

Mark (X) the "Confidential" box next to any item you claim as confidential

Complete either item I (Class I or 2 substances) or 2 (Polymers) as appropriate. Complete allI other items.

If another person will submit chemical identity information for you (for either Item I or 2), mark (X) the box at the right. ConfiIdentify' the name, company, and address of that person in a continuation sheet. ~J dential

I.Class 1 or 2 chemical substances (for definitions of class I and class 2 substances, see the Instructions Manual)

a. Class of substance - Mark (X) 1 0 Class I or 2 Z Class 2 X

b. Chemical name (Currently correct Chemical Abstracts (CA) Name that is consistent with TSCA Inventory listings for similar substances.

For Class I substances a CA Index Name must be provided. For Class 2 substances either a CA tndex Name or CA Preferred Name must x

be provided, which ever is appropriate based on CA 9C1 nomenclature rules and conventions).

c Please identify which method you used to develop or obtain the specified chemical identity information reported in this notice: (check one).

Z Method 1 (CAS Inventory Expert Service - a copy of the tdentification Method 2 (Other Source)

report obtained from the CAS Inventory Expert Services must be

submitted as an attachment to this notice)

d. Molecular formula and CAS Registry Number (if a number already exists for the substance)

x

CAS4

x

e. For a class I substance, provide a complete and correct chemical structure diagram. For a class 2 substance - (1) List the immediate

precursor substances with their respective CAS Registry Numbers. (2) Describe the nature of the reaction or process (3) Indicate the x

range of composition and the typical composition (where appropriate). (4) Provide a correct representative or partial chemical structure

diagram, as complete as can be known, if one can be reasonably ascertained.

DMark (X) this box if you attach a continuation sheet.

EPA FORM 7710-25 (Rev. 5-95) Page 4
Part I -- GENERAL INFORMATION -- Continued

Section B -- CHEMICAL IDENTITY INFORMATION -- Continued

2. Polymers (For a definition of polymer, see the Instructions Manual.) Confidential

a. Indicate the number-average weight of the lowest molecular weight composition of the polymer you intend to manufacture.

Indicate maximum weight percent of low molecular weight species (not including residual monomers, reactants, or solvents) below 500 and x

below 1,000 absolute molecular weight of that composition.

Describe the methods of measurement or the basis for your estimates: GPC Other E]: (Specify) ____________

i) lowest number average molecular weight:___________

ii) maximum weight % below 500 molecular weight. _________

iii) maximum weight % below 1000 molecular weight-_________

Mark (X) this box if you attach a continuation sheet.

b. You must make separate confidentiality claims for monomer or other reactant identity, composition information, and residual information. Mark (X) the

"Confidential" box next to any item you claim as confidential

(1) - Provide the specific chemical name and CAS Registry Number (if a number exists) of each monomer or other reactant used in the manufacture of

the polymer.

(2) - Mark (X) this column if entry in column (1) is confidential.

(3) - Indicate the typical weight percent of each monomer or other reactant in the polymer

(4) - Mark (X) the identity column if you want a monomer or other reactant used at two weight percent or less to be listed as part of the polymer

description on the TSCA Chemical Substance tnventory.

(5) - Mark (X) this column if entries in columns (3) and (4) are confidential.

(6) - tndicate the maximum weight percent of each monomer or other reactant that may be present as a residual in the polymer as manufactured for

commercial purposes.

(7) - Mark (X) this column if entry in column (6) is confidential.

Monomer or other reactant and CAS Registry Number Confi- Typical Identity Confi- Maximum Confidential composition Mark (X) dential residual dential

(1) (2) (3) (4) (5) (6) (7)

X %X % X

X %X % X

X %X % X

E] Mark (X) this box if you attach a continuation sheet.

c. Please identify which method you used to develop or obtain the specified chemical identity information reported in this notice (check one).

Method 1 (CAS Inventory Expert Service - a copy of the identification report F- Method 2 (other source)

obtained from CAS Inventory Expert Service must be submitted as

as attachment to this notice)

d. The currently correct Chemical Abstracts (CA) name for the polymer that is consistent with TSCA Inventory listings for similar polymers.X

e. Provide a correct representative or partial chemical structure diagram, as complete as can be known, if one can be reasonably ascertained.

SMark (X) this boxc if you attach a continutation sheet.

EPA FORM 7710-25 (Rev. 5-95) Page 5
Contains TSCA Confidential Business Information

page 5a
Part I -- GENERAL INFORMATION -- Continued

Section B -- CHEMICAL IDENTITY INFORMATION -- Continued

3. Impurities

(a) - Identify each impurity that may be reasonably anticipated to be present in the chemical substance as manufactured for commercial purpose. Provide the

CAS Registry Number if available. If there are unidentified impurities, enter "unidentified."

(b) - Estimate the maximum weight % of each impurity. If there are unidentified impurities, estimate their total weight %.

Impurity and CAS Registry Number Maximum Confipercent dential

(a) (b)

% X

[] Mark (X) this box if you attach a continuation sheet.

4. Synonyms - Enter any chemical synonyms for the new chemical identified in subsection I or 2. Confidential

None

D Mark (X) this box if you attach a continuation sheet.

5. Trade identification - List trade names for the new chemical substance identified in subsection I or 2.

x

D Mark (X) this box if you attach a continuation sheet.

6. Generic chemical name - If you claim chemical identify as confidential, you must provide a generic name for your substance that reveals

the specific chemical identity of the new chemical substance to the maximum extent possible. Refer to the

TSCA Chemical Substance Inventory, 1985 Edition, Appendix B for guidance on developing generic names.

Alcohol, alkoxylated

jZ Mark (X) this box if you attach a continuation sheet.

7. Byproducts - Describe any byproducts resulting from the manufacture, processing, use, or disposal of the new chemical substance. Provide the CAS Registry

Number if available.

Byproduct CAS Registry Number Con fi -

(1) (2) dential

x

Dl Mark (X) this box if you attach a continuation sheet.

EPA FORM 7710-25 (Rev. 5-95) Page 6
Part I -- GENERAL INFORMATION -- Continued

Section C -- PRODUCTION, IMPORT, AND USE INFORMATION:

Mark (X) the "Confidential" box next to any item you claim as confidential.

1. Production volume -- Estimate the maximum production volume during the first 12 months of production. Also estimate the maximum

production volume for any consecutive 12-month period during the first three years of production. Estimates should be on 100% new chemical

substance basis. For a Low Volume Exemption application, if you choose to have your notice reviewed at a lower production volume than

10,000 kg/yr. specify the volume and mark (x) in the binding box. If granted, you are bound to this volume

Maximum first 12-month production (kg/yr) Maximum 12-month production (kg/yr) Confi- Binding

(100% new chemical substance basis) (100% new chemical substance basis) dential Option

___________________________________________ ____________________________________ _MarkMar((x

2. Use Information -- You must make separate confidentiality claims for the description of the category of use, the percent of production volume

devoted to each category, the formulation of the new substance, and other use information. Mark (X) the "Confidential" Box next to any item

you claim as confidential.

a. (1) -- Describe each intended category of use of the new chemical substance by function and application..

(2) -- Mark (X) this column if entry column (1) is confidential business information (CBI).

(3) -- Indicate your willingness to have the information provided in column (1) binding.

(4) -- Estimate the percent of total production for the first three years devoted to each category of use.

(5) -- Mark (X) this column if entry in column (4) is confidential business information (CBI).

(6) -- Estimate the percent of the new substance as formulated in mixtures, suspensions, emulsions, solutions, or gels as manufactured for

commercial purposes at sites under your control associated with each category of use.

(7) -- Mark (X) this column if entry in column (6) is confidential business information (CBI).

(8) -- Indicate % of product volume expected for the listed "use" sectors. Mark more than one box if appropriate. Mark (X) to indicate

your willingness to have the use type provided in (8) binding.

(9) -- Mark (X) this column if entry(ies) in column (8) is (are) confidential business information (CBI).

Category of use (1) CI Binding Produc- CBt % in CBI % of substance expected per use CBI

Option tion % Form- (8)

(by fuinction and application i.e. a dispersive dye Mark (x) ulation Site- Con-* tndus- Coin- Binding

for finishing polyester fibers) (2) (3) (4) (5) (6) (7) limited sumer trial mercial Option (9)

x x Xx

%4 %

____ ____ __% %

*If you have identified a "consumer" use, please provide on a continuation sheet adetailed description of the use(s) of this chemical substance in consumer products.

tn addition include estimates of the concentration of the new chemical substance as expected in consumer products and describe the chemical reactions by which this

substance loses its identity in the consumer product.

D Mark (X) this box if you attach a continuation sheet.

b. Generic If you claim any category of use description in subsection 2a as confidential, enter a generic description of that category. Read the

use Instructions Manual for examples of generic use descriptions.

description

Wetting agent

DMark (X) this box if you attach a continuation sheet.

3. Hazard Information -- Include in the notice a copy of reasonable facsimile of any hazard warning statement, label, material safety data sheet, or other Binding

information which will be provided to any person who is reasonably likely to be exposed to this substance regarding protective equipment or practices Option

for the safe handing, transport, use, or disposal of the new substance. List in part III hazard information you include. Mr x

Z Mark (X) this box if you attach hazard information.

EPA FORM 77 10-25 (Rev. 5-95) Page 7
Part 11-- HUMAN EXPOSURE AND ENVIRONMENTAL RELEASE

Section A -- INDUSTRIAL SITES CONTROLLED BY THE SUBMITTER Mark (X) the "Confidential" box next to any item you

claim as confidential

Complete section A for each type of manufacture, processing, or use operation involving the new chemical substance at industrial sites you

control. Importers do not have to complete this section for operations outside the U.S.; however, you may still have reporting requirements if

there are further industrial processing or use operations after import. You must describe these operations. See instructions manual

1. Operation description Confia. Identity -- Enter the identity of the site at which the operation will occur. dential

Name

Not applicable as new substance will be imported

Site address (number and street)

City, County, State, ZIP code

If the same operation will occur at more than one site, enter the number of sites. Identify the

additional sites on a continuation sheet, and if any of the sites have significantly different

production rates or operations, include all the information requested in this section for those

sites as attachments.

11Mark (X) this box if you attach a continuation sheet.

b. Type --

Mark (X) EManufacturing EProcessing D]Use____

c. Amount and Duratio -- Complete I or 2 as appropriate_______________ ______________

Maximum kg/batch (100% new chemical Hours/batch Batches/year

substance)

1. Batch

Maximum kg/batch (100% new chemnical Hours/batch Batches/year I substance)

2. Continuous

d. Process description [I] Mark (X) to indicate your willingness to have your process description binding.

(1) Diagram the major unit operation steps and chemical conversions. Include interim storage and transport containers (specify- e.g. 5 gallon pails, 55 gallon

drum, rail car, tank truck, etc.).

(2) Provide the identity , the approximate weight (by kg/day or kg/batch on a 100% new chemical substance basis), and entry point of all starting materials and

feedstocks (including reactants, solvents, catalysts, etc.), and of all products, recycle streams, and wastes. Include cleaning chemicals (note frequency if not

used daily or per batch.).

(3) Identify by number the points of release, including small or intenmittent releases, to the environment of the new chemical substance.

Not applicable

SMark (X) this box if you attach a continuation sheet.

EPA FORM 7710-25 (Rev. 5-95) Page 8
Part 11-- HUMAN EXPOSURE AND ENVIRONMENTAL RELEASE -- Continued

Section A -- INDUSTRIAL SITES CONTROLLED BY THE SUBMITTER -- Continued

2. Occupational Exposure -You must make separate confidentiality claims for the description of worker activity, physical form of the new chemica substance,

number of works exposed, and duration of activity. Mark (X) the "Confidential" box next to any item you claim as confidential.

(1) --Describe the activities (i.e. bag dumping, tote filling, unloading drums, sampling, cleaning, etc.) in which workers may be exposed to the substance.

(2) -- Mark (X) this column if entry in column (1) is confidential business information (CBI).

(3) --Describe any protective equipment and engineering controls used to protect workers.

(4) and (6) -- Indicate your willingness to have the information provided in column (3) or (5) binding.

(5) --Indicate the physical form(s) of the new chemical substance (e.g., solid: crystal, granule, powder, or dust) and % new chemical substance (if part of a mixture)

at the time of exposure.

(7) --Mark (X) this column if entry in column (5) is confidential business information (CBI).

(8) -- Estimate the maximum number of workers involved in each activity for all sites combined.

(9) --Mark (X) this column if entry in column (8) is confidential business information (C131).

(10) and (11) -- Estimate the maximum duration of the activity for any worker in hours per day and days per year.

(12) --Mark (X) this column if entries in columns (10) and (t1) are confidential business information (CBI).___________

Worker activity CBl Protective Equipment/ Binding Physical forns(s) Binding C131 # of CBIl Maximur duration CBI

mn

(i.e., bag dumping, filling drums) Engineering Controls Option and % new Option Worker Uirn/day Days/yr

Mark (x) substance Mark (x)

(1) (2) (3) (4) (5) (6) (7) Expose (9) (10) (It) (12)

d

___________________________________(8) _________

Not applicable

DMark (X) this box if you attach a continuation sheet.

3. Environmental Release and Disposal -- You must make separate confidentiality claims for the release number and the amount of the new chemical substance

released and other release and disposal information. Mark (X) the "Confidential" box next to each item you claim as confidential.

(1) --Enter the number of each release point identified in the process description, part 11, section A, subsection Ild(3).

(2) -Estimate the amount of the new substance released (a) directly to the environment or (b) into control technology (in kg/day or kg/batch)

(3) -Mark (X) this column if entries in columns (1) and (2) are confidential business information (CBI).

(4) -- Identify' the media (stack air, fu~gitive air (optional-see Instruction Manual), surface water, on-site or off-site land or incineration, POTW, or other (specify)) to

which the new substance will be released from that release point.

(5) -a. Describe control technology, if any, and control efficiency that will be used to limit the release of the new substance to the environment. For releases

disposed of on land, characterize the disposal method and state whether it is approved for disposal of RCRA hazardous waste. On a continuation sheet, for

each site describe any additional disposal methods that will be used and whether the waste is subject to secondary or tertiary on-site treatment. b. Estimate

the amount released to the environment after control technology (in kg/day).

(6) --Mark (X) this column if entries in columns (4) and (5) are confidential business information (C131).

(7) -- Identify' the destination(s) of releases to water. Please supply NPDES (National Pollutant Discharge Elimination System) numbers for direct discharges or

NPDES numbers of the POTW (Publicly Owned Treatment Works). Mark (X) if the POTW name or NPDES # is confidential business information (CBI).

Release Amount of new substance CBII Media of Control technology and efficiency (you may wish to optionally attach efficiency data) CB1

Number released release

(I) (2a) (2b) e g.aack air Binding

_______ (3) (4) (5a) Mark (x) (5b) (6)

not

applicable

(7) Mark (X) the D POTW provide name(s) below: CBI 0 Navigable E]Other - Specify provide NPDES # CII

destination(s) of wtra

releases to water.

1D Mark (X) this box if you attach a continuation sheet.

EPA FORM 7710-25 (Rev. 5-95) Page 9
Part 11-- HUMAN EXPOSURE AND ENVIRONMENTAL RELEASE -- Continued

Section B -- INDUSTRIL SITES CONTROLLED BY OTHERS

Complete section B for typical processing or use operations involving the new chemical substance at sites you do not control. Importers do not have to complete this

section for operations outside the U.S., however, you must report any processing or use activities after import. See the Instructions Manual. Complete a separate

section Bfor each type of processing, or use operation involving the new chemical substance If the same operation is performed at more than one site describe the

typical operation common to these sites. Identify additional sites on a continuation sheet.

1. Operation Description -- To claim information in this section as confidential, circle or bracket the specific information that you claim as confidential.

(1) -- Diagram the miajor unit operation steps and chemical conversions, including interim storage and transport containers (specify - e.g. 5 gallon pails, 55 gallon

drums, rail cars, tank trucks, etc). On the diagram, identify by letter and briefly descrnbe each worker activity (2) -- Provide the identity, the approximate weight

(by kg/day or kg/batch, on an 100% new chemical substance basis), and entry point of all feedstocks (including reactants, solvents and catalysts, etc) and all

products, recycle streams, and wastes. Include cleaning chemicals (note frequency if not used daily or per batch). (3) -- Identify by number the points of release,

including small or intermittent releases, to the environment of the new chemical substance. (4) Please enter the # of sites (remember to identify the locations of

these sites on a continuation sheet)

# of sites

DMark (X) this box if you attach a continuation sheet.

2. Worker Exposure/Environmental Release

(1) -From the diagram above, provide the letter for each worker activity. Complete 2-8 for each worker activity descrnbed.

(2) -- Estimate the number of workers exposed for all sites combined

(4) -Estimate the typical duration of exposure per worker in (a) hours per day and (b) days per year.

(6) -- Descnibe physical form of exposure and % new chemical substance (if in mixture), and any protective equipment and engineering controls, if any, used to

protect workers.

(7) -Estimate the percent of the new substance as formulated when packaged or used as a final product.

(9) --From the process diagram above, enter the number of each release point Complete 9-13 for each release point identified.

(10) -- Estimate the amount of the new substance released (a) directly to the environment or (b) into control technology to the environment (in kg/day or

kg/batch).

(12) -- Descnibe media of release i.e stack air, fugitive air (optional -see Ins tructions Manual), surface water, on-site or off-site land or incineration, POTW, or

other (specify) and control technology, if any, that will be used to limit the release of the new substance to the environment.

(14) -- Identify byproducts which may result from the operation.

(3), (5), (8), (11), 13) and (15) -- Mark Xthis colum~n if any of the proceeding entries are confidential business inform-ation (CBI. ________

Letter # of CBI Duration CB1 Protective Equip. / % in CBI Release Amount of CBI Medis of Release CBI

of Workers of Engineening Controls/ Form- Number New & Control

Act- Exposed Exposure Physical Form and % ulation Substance Technology

ivity new substance Released

(1) (2) (3) (4 (5) (6) (7) (8) (9) (lOs) -7(lob) (11) (12) (13)

x -__

(14)- Byproducts: (15)

D: Mark (X) this box if you attach a continuation sheet,

EPA FORM 77 10-25 (Rev. 5-95) Page 10
OPTIONAL POLLUTION PREVENTION INFORMATION

To claim information in this section as confidential circle or bracket the specific information that you claim as confidential.

In this section you may provide information not reported elsewhere in this form regarding your efforts to reduce

or minimize potential risks associated with activities surrounding manufacturing, processing, use and disposal

of the PMN substance. Please include new information pertinent to pollution prevention, including source

reduction, recycling activities and safer processes or products available due to the new chemical substance.

Source reduction includes the reduction in the amount or toxicity of chemical wastes by technological

modification, process and procedure modification, product reform-ulation, raw materials substitution, and/or

inventory control. Recycling refers to the reclamation of useful chemical components from wastes that would

otherwise be treated or released as air emissions or water discharges, or land disposal. Descriptions of pollution

prevention, source reduction and recycling should emphasize potential risk reduction subsequent to compliance

with existing regulatory requirements and can be either quantitative or qualitative. The EPA is interested in the

information to assess overall net reductions in toxicity or environmental releases and exposures, not the shifting

of risks to other environmental media or non-environmental areas (e.g., occupational or consumer exposure). In

addition, information on the relative cost or performance characteristics of the PM1N substance to potential

alternatives may be provided.

All information provided in this section will be taken into consideration during the review of this

substance. See Instructions Manual and Pollution Prevention Guidance manual for guidance and

examples.

Describe the expected net benefits, such as (1) an overall reduction in risk to human health or the environent; (2) a reduction in the

volume manufactured; (3) a reduction in the generation of waste materials through recycling, source reduction or other means; (4) a

reduction in potential toxicity or human exposure and/or environmental release; (5) an increase in product performance, a decrease in

the cost of production and/or improved operation efficiency of the new chemical substance in comparison to existing chemical

substances used in similar application; or (6) the extent to which the new chemical substance may be a substitute for an existing

substance that poses a greater overall risk to human health or the environment.

LIMark (X) this box if you attach a continuation sheet.

EPA FORM 77 10-25 (Rev. 5-95) Page 11I
Part III -- LIST OF ATTACHMENTS

Attach continuation sheets for sections of the form and test data and other data (including physical/chemical properties and

structure/activity information), and optional information after this page. Clearly identify the attachment and the section of the form to

which it relates, if appropriate. Number consecutively the pages of the attachments. In the column below, enter the inclusive page

numbers of each attachment.

Mark (X) the "Confidential" box next to any attachment name you claim as confidential. Read the Instructions Manual for guidance on

how to claim any information in an attachment as confidential. You must include with the sanitized copy of the notice form a sanitized

version of any attachment in which you claim information as confidential.

Attachment name Attachment Confipage number(s) dential

14-18

Material Safety Data Sheet (MSDS)

19-21 X

22-25 X

28-29 X

30 X

CASIES Report 172

CMVA electronic PMVN documentation 173

PMVN fee documentation 174

5a X

26-27 X

Acute Oral Toxicity Test in Rats 83-1 04

Acute Dermal Irritation/CorrosionTest 60-82

Acute Eye Irritation/Corrosion Test 34-59

Activated sludge, respiration inhibition test 131-153

Acute toxicity fish test 154-171

31-33 X

Determination of ready biodegradability 105-1 30

DMark (X) this box if you attach a continuation sheet. Enter the attachment name and number.

EPA FORM 7710-25 (Rev. 5-95) Page 12
PHYSICAL AND CHEMICAL PROPERTIES WORKSHEET

To assist EPA's review of physical and chemical properties data, please complete the following worksheet for data you provide and

include it in the notice. Identify the property measured, the page of the notice on which the property appears, the value of the

property, the units in which the property is measured (as necessary), and whether or not the property is claimed as confidential The

physical state of the neat substance should be provided. These measured properties should be for the neat (100% pure) chemical

substance. Properties that are measured for mixtures or formulations should be so noted (% PMIN substance in 9). You are not

required to submit this worksheet; however, EPA strongly recommends that you do so, as it will simplify review and ensure that

confidential information is properly protected You should submit this worksheet as a supplement to your submission of test data.

This worksheet is not a substitute for submission of test data.

Property Mark Page Value Measured or Confi-

(X) if number Estimate dential

providd (c)Mark (X)

(a) prvdd (b) ()(M orE) (d)

X 16 M

Physical state of neat substance __(s) X2 (1) _(g) ____

Vapor pressure

@ Temperature ________ Torr

X 16 M

Density/relative DIN 51757 0.96 g/cm3

Solubility

@ Temperature _________

Solvent:

X 16

Solubility in water @ Temperature 25 0C <_ <0.5 gI M

Melting temperature 0

Boiling / sublimation temperature@_ to__rr pressure _________________

X M X

Spectra: IR, H-NMR, 13C-NMR, MALDI 26-33 _________ __

Dissociation constant

Particle size distribution

Octanol / water parttion coefficient

Henry's Law constant

Volitalization from water

Volitalization from soil

_pH@ concentration

Flammability DIN EN 22719 X 6 lshPin=>IO

-Explodability __

-Adsorption / coefficient

Other - Specify_____ _______

X 16 Approx. 150 mPas M

-Viscosity 25'C DIN 53015 __ __________ __

ATTACHMENT

PAGE 13
Contains TSCA Confidential Businec.': 11ormZation

SAFETY DATA SHEET (91/155/EEC)

Version 2 Revision date Zb.U1.98-

Date :30.03.98

Page :1

1. Identification of the substance/preparation and company

Trade name

Information on manufacturer/supplier

Telephone no. i 'Telefax:fIn formation provided by

Elehn o Teepoe o.^IjTelefax :L7 J

Emergency information

Telephone no. -. -' Telefax :

* 2. Composition/information on ingredients

Chemical characterization

Aloo, alkoxylated

* 3. Hazards identification

Specific hazards for human health and environment

No particular hazards known.

*4. First aid measures

General information

Remove contaminated clothing.

After inhalation

Ensure supply of fresh air.

After contact with skin

In case of contact with skin wash off immediately with soap and

water

After contact with eyes

Flush eye immediately and thoroughly whilst protecting the unhurted

eye.

if irritation continues, consult a physician.

After ingestion

In the event of symptoms seek medical advice.

Page: 2
Contains TSCA Confidenti-Il Cusiness 1Ilorimtion

SAFETY DATA SHEET (91/155/EEC)

Version :2 Revision date 26.01 .98

Date :30.03.98

Page: 2

5. Fire-fighting measures

Suitable extinguishing media

foam, carbon dioxide, dry powder, water spray.

Extinguishing media that must not be used for safety reasons

not applicable

Special hazards from the substance itself, its combustion products

or from its vapours

In the event of fire the following can be released: -

- carbon dioxide, carbon monoxide

Special protective equipment for firefighting

Do not inhale explosion and/or combustion gases

6. Accidental release measures

Personal precautions

Use personal protective equipment.

Environmental precautions

Do not allow to enter drains or waterways

Do not discharge into the subsoil/soil.

Methods for cleaning up/taking up

Take up with absorbent material (eg sand, kieselguhr, universal

binder)

Dispose of absorbed material in accordance with the regulations.

7. Handling and storage

Handling

Advice on protection against fire and explosion

No special measures required.

Storage

Further information on storage conditions

Keep container closed.

Storage class 10

8. Exposure controls/personal protection

Personal protective equipment

General protective measures

Avoid contact with eyes and skin

Page :3
Contains TSCA Confidenit'l Cusnor-~orat

SAFETY DATA SHEET (91/155/EEC)

Version : 2 Revision date 26.01 .98-

Date 30.03.98

Page: 3

Hygiene measures

Do not eat, drink or smoke when working.

Wash hands before breaks and after work.

Remove soiled or soaked clothing immediately.

Respiratory protection None required

Hand protection PVC gloves

Eye protection tightly fitting safety glasses

Body protection light protective clothing

9. Physical and chemical properties

Appearance

Form : Liquid

Colour : colourless, clear

Odour : mild

Data relevant to safety

Changes in physical state

Melting temperature

not measured

Boiling temperature

not measured

Flash point

> 100 c

Method :DIN EN 22719 (DIN 51758)

Ignition temperature

not measured

Density

0.96 g/cm'

at 20 O

Method :DIN 51757

solubility in water

< 0.5 g/l

at 25 O

pH value

not measured

Viscosi.ty

approx. 150 mPa*s

at 25 O

Method : DIN 53015 (H~ppler)

Page: 4
Contains TSCA Confidential Businocs Infor niti/T J

SAFETY DATA SHEET (91/155/EEC)

Version :2 Revision date 26.01.98 iiDate :30.03.98 Pae]

10. Stability and reactivity

Thermal decomposition

Not measured

Hazardous reactions

No hazardous reactions with proper storage and handling.

Hazardous decomposition products

None with proper storage and handling.

*11. Toxicological information

Acute oral toxicity ( LD50

> 2000 mg/kg

Species rat

Method OECD 401

Irritant effect on skin

non- irritant

Species :rabbit

Method :OECD 404

Irritant effect on eyes

slight irritant effectdoes not require labelling

Species rabbit

Method :OECD 405

Remarks

Due to the composition of the product it cannot be excluded:

Irritant to eyes.

12. Ecological information

Data on elimination ( persistence and degradability)

Biodegradability

not readily degradable

Method : OECD 301 D

Remarks

The product is considered to be a weak water pollutant (German law)

Do not allow to enter soil, waterways or waste water canal.

13. Disposal considerations

Page :5

Ar o
Contains TSCA Confi6didtal Business Informatio

SAFETY DATA SHEET (91/155/EEC)

Version 2 Revision date :26.01.98

Date :30.03.98

Page: 5

Product

In accordance with local authority regulations, take to special

waste incineration plant

Uncleaned packaging

If empty contaminated containers are recycled or disposed of, the

receiver must be informed about possible hazards.

14. Transport information

Not classified as hazardous for transport

*15. Regulatory information

Labelling in accordance with GefStoffVIEC

The product does not require a hazard warning label in accordance with

EC directives/German regulations on dangerous substances.

National regulations

German regulations on flammable substances: is not subjected to the

regulations on flammable substances.

TA-Luft (Ger.) Class III Paragraph 3.1.7

Water Hazard Class (Germany) :1 Cself -classification (Germany)

Other regulations

none

16. Other information

none

This information is based on our present state of knowledge.

It should not therefore be construed as guaranteeing specific

properties of the products described or their suitability for

a particular application.

*Changes compared to the previous version are marked with an

asterisk before the section number!I

Date :30.03.98
19: -~ 1204 Contains TSCA Confidential Business Information/
Contains ISCA Confidential Business Information
Contains TSCA Confidential Business Information
Contains TSCA Confidential Business Information
Contains TSCA Confidential Business Information
I I Contains TSCA Confidential Business Information
Contains TSCA Confidential Business Information
Contains TSCA Confidential Business Information
Contains TSCA Confidential Business Information
Contains TSCA Confidential Business Information
/c

/.

-t:

I In
//

I 

/

//

Contains TSCA Confidential Business Information 30
Contains TSCA Confidential Business Information
Contains TSCA Confidential Business Information '
Contains TSCA Confidential Business Information
Daimlerstral~e 

Tel. 07.21/97 37-0 

5 b 

-Fax 

-D-76185 

07 21/97 

Karlsrhe 

37-222 "BioChem

E-mail: biochem@biochem.de Labor fOr biologische und

Internet: httO/ww.biochern.de e chemische Analytik GmbH

REPORT 97 10 42 412 C

AcuTE EYE IIuuTATioN/CoRROSION

Project No.: 971042412

Report No.: 97 10 42 412 C

Test substance: [I1

Batch No: not stated

BioChem sample designation: 137291 3001

Date of arrival: November 05, 1997

Sponsor.I

Monitor[2

Test facility: BioChem GmbH

Daimlerstr. 5 b

D-76185 Karlsruhe

Germany

Management: Dr. W. Massmann

Dr. M. Miller

Study Director. Dr. H.-J. Kramer

Quality Assurance Manager. Dr. H. Merten

Karlsruhe, December 23, 1997

BioChem GmbH

Page 1 of 26 pages

R""", t7 10 42 412 C

)I Acute Eye Irritation/Corrosion

Contains TSCA Confidential Business Information
Page 2 of 26 pages

Fteport 97 10 42 412 C

JAcute Eye hrritation/Cqrrosion

Statement of GLP compliance

We declare that all tests are made in accordance with the protocol and with the regulations of the

OECD-Principles of the "Good Laboratory Practice" (GLP).

GLP: "Good Laboratory Practice" Regulations of the European Communities, EC, enacted in

Germany in the " Cheniikaliengesetz" (Chemicals Act), 2nd Amendment, dated July 25, 1994,

BGBI I, pp. 1703, 1994 and "AndVO", dated May 14, 1997, BGBl. IL pp. 1060, 1997.

The documented data correspond to the measured values.

Management

Dr. Manfred Miller Date

Study Director

Dr. Hans-Joachim Kramer Date

Quality Assurance Manager

Dr. Hans Merten Date

Contains TSCA Confidential Business Information
Page 3 of 26 pages

Report 97 10 42 412 C

Y Acute Eye Irritation/Corrosion

Umlweltminiserium

Baden-Wirttemberg

GLP-Bescheinigunig

Bescheinigung Certificate

Hiermit wird best~tgt, daB die It is hereby certified that the testPrllftpgeinrichtung filility

in 76185SKarlsruhe in 76185 Kadsruhe

der BioChern, Labor Mr biologische und of BioChem, Labor fir biologische und

chemische Analyti GmbH, - cbemi che Analytik G~mbH,

Daimlerstr. 5b . Daimierstr. 5b

am 19J20.06.95, 27.09.95 und 26.01.96 on 19120. Tune 95, 27. September 95

and 26. January 96

von der far die Oberwachung zust~digen wa s inspected by the competent

Behorde Uber die Einl~tung der Grund- authority regarding compliance with the

sluze der Guten Laborpraids iaspi~iert .Principles of Good Laboratory Practice.

worden ist.

Es wird hiermit bestafigt, daB3 folgende -It is hereby certified that studies in this test

Prafingen in dieser Pruifeinrchtung nach facility are conducted in compliance with the

den Gnindsitzen der (3uten Laborpra~tis Principles of Good Laboratory Practice.

durchgefilhrt werden.

BADEN,

Stuttgart, den 6 qW)ERG

Seite 1/2 page 112

Contains ISCA Confidential Business Information k~/ Y
* Page 4of 26 pages Reoort 97 10 42 412 C

L Acute Eye Irritation/Corrosion

* S cite 2/2 zur GL.P-Bescheinigung

Priifungskategorie I Priifungen auf physikalischchemische Eigenschzften and

Gehatsbestimmungen

Prfifungskategorie 2 PriUhngen auf toxikologische

Eigenschaften

Priifungskategorie 3 Prilfungen auf mutagene

Eigenschaften (in vitro, in vivo)

Priifungskategorfe 4 Umtwelttoikologische Prilfng~en

zu Auswfrktungen auif aquatische

and terrestrische Organismen.

Priifungskategorie 5 Prilfungen xam Verhalten im

Boden, im Wasser and inder

Luft; Bioakkuymulation;

Metabotismus

Prilfungskategorie 6 Prilfungen auf Rfickstinde

Dies entspricht den Kategorien nach der Ailgemi Verv;wtngsvarscbrift mmn

Verfahren der beh~rdlichen (Yberwachung der Einhaltung, der Guten Laborpaxs

* (ChemVwV-4J1.P) vom 29. Oktober 1990:

'Physikalisch-chemische Eigenschaften unci

Gehaltsbestimmungen"

"Toxikologische Eigenschaften"

"Okotoxikologische Eigenschaften"

"Verhaten im Boden, im Wasser and in der Luft't

IfRickst~iude"

Stuttgart, den 26.02.1996

A A(Xd c S4 V I.-1RTTEM8 ERG

Dr. AlrechtContains TSCA Confidential Business InformatioSeite 2/2 page 2/2
Page 5 of 26 pages Report 97 10 42 412 C

( 1 Acute Eye I-itation/Corrosion

Translation of page 2 of the GLP-Certificate by BioChem.

Test category 1: Tests on physico-chemical

properties and content

determinations

Test category 2: Tests on toxicological

properties

Test category 3: -Tests on mutagenic properties

(in vitro, in vivo)

Test category 4: Ecotoxicological tests on*

effects on aquatic and

terrestrial organisms

Test category 5: Tests on behaviour in soil, in water

and in the air;

bioaccumulation; metabolism

Test category 6: Tests on residues

This corresponds to the categories according to the General Administration

Regulation of the Procedure of the Official Supervision of the Compliance with Good

Laboratory Practice (Chem VwV-GLP) dated October 29, 1990:

"Physico-chemical Properties and Content Determinations"

'Toxicological Properties"

"Ecotoxicological Properties"

"Behaviour in Soil, in Water and in the Air"

"Residues"

Stuttgart, February 26, 1996

Baden- Wirttemberg Contains TSCA Confidential Business Informa 3Y

Department of the Environmnent
Page 6 of 26 pages

Report 97 10 424120C

( _iAcute Eye Irritation/Corrosion

Quality Assurance Unit Statement

Title: Final Report

Acute Eye Irritation / Corrosion

project number: 971042412

protocol number: 97 10 42 412 C

Guideline: OECD-Guideline for Testing of Chemicals No. 405 "Acute

Eye Irritation /Corrosion"

Study I test system: Determination of the irritant and/or corrosive effects on eyes

of mammals / albino rabbits

Test substance:

Batch No.: not stated

Sample designation: 137291 3001

Date of arrival: November 05, 1997

Study Director: Dr. H.-J. Kramer

Contains TSCA Confidential Business Information
Page 7 of 26 pages

Report 97 10 42 412 C

I/Acute Eye Irritation/Corrosion

The following checks were effected:

reports to the Study

Director and to the

date verified procedure Management

November 14, 1997 protocol November 14, 1997

December 10, 1997 audit 1 December 10, 1997

December 16, 1997 audit 2 December 16, 1997

December 22, 1997 final report in English December 22, 1997

The data which were taken as a basis for this report were verified by the Quality Assurance Unit.

The accordance of the study with its protocol, the OECD-Guideline for Testing of Chemicals No.

405 and the Principles of Good Laboratory Practice (GLP) is guaranteed.

The methods, procedures and observations are exactly described in the report. The Quality

Assurance Unit ensures that the reported results exactly describe the raw data of the study.

The Study Director and the Management are generally informed of any findings of the Quality

Assurance Unit.

As for this study no departures from the prescribed course of the test were ascertained.

Karlsruhe, December 23, 1997 Dr. H. Merten

(Quality Asuwincc Manager)

Contains TSCA Confidential Business Information
Page 8 of 26 pages

Report 97 10 424120C

')t Acute Eye Irritation/Corrosiona

CONTENTrS

page

1 Preface 9

2 Summary 10

3 Guideline 11

4 Protocol I I

5 Test Substance 11

6 Method "Acute Eye Irritation/Corrosion" 12

6. 1 Animals and Laboratory Conditions 12

6.2 Composition of Diet and Water 13

6.3 Acclimatization/Preparations 16

6.4 Study Procedures 17

6.5 Evaluation of the Ocular Reaction 18

7 Findings "Acute Eye Irritation/Corrosion" 20

7.1 Mortality/Toxicological Symptoms 20

7.2 Behaviour 20

7.3 Body Weight 20

7.4 Eye Reaction 21

8 Archiving 26

Contains TSCA Confidential Business Information 41'
Page 9 of 26 pages

Repor 97 10 42 412 C

I/Acute Eye Irritation/Corrosion

PREFACE

Test system: Acute Eye Irritation /Corrosion

Laboratory animals: Albino rabbits

Guideline: OECD-Guideline for Testing of Chemicals

Section 4, 405 "Acute Eye Irritation/

Corrosion" adopted February 24, 1987

GLP: Gesetz zumn Schutz vor gefdhrlichen

Stoffen (Chemikaliengesetz ChemG, § 19 a - d

vom 25. Juli 1994)

Project No.: 97 1042412

Report No.: 97 1042 412 C

Test substance:L

Sponsor.

Monitor:

Test Facility: BioChem GmbH

Daimlerstra~e 5 b

D-76 185 Karlsruhe

Germany

Management: Dr. W. Massmann

Dr. M. MIler

Study Director. Dr. H.-J. Kramer

Quality Assurance Manager. Dr. H. Merten

Contains TSCA Confidential Business Information4?
Page 10 of 26 pages

Report 97 10 42 412 C

( )IAcute Eye Irritation/Corrosion i

2 SUMMARY

Report: Acute Eye Irritation/Corrosion study of i

compliance with OECD-Guideline 405

Sponsor:

Test Facility: BioChem, GmbH

Daimlers-tra~e 5 b

D-76185 Karlsruhe

Germany

Project No.: 971042412

Protocol No.: 97 10 42 412 C

Report No.: 97 10 42 412 C

Findings:

The test substance was applied in a single dose of 0.1 ml into the conjunctival sac of the

right eye to each of 3 albino rabbits.

Under the conditions of this experimnent >ld not cause any acute systemic

toxicological signs or mortality./

After evaluation of the eye reaction according to table I of the OECD-Guideline 405, all

three rabbits developed hyperaemnic blood vessels. This effect was estimated at grade

0.5 to 2.5 (maximum possible 3). Hyperaemnic blood vessels were not observed any

more on day 5. All three rabbits showed swelling of lids. This effect was estimated at

grade I to 3 (maximum possible 4). Swelling of lids was not observed any more on day

5. Two rabbits showed opacity of the cornea, 48 hours after application. This effect was

estimated at grade I (maximum possible 4). Opacity was not observed any more on day

3. Ocular lesions of iris could not be observed during the study. All lesions turned out to

be completely reversible within 4 days.

In the course of this experiment the body weight of all three rabbits increased.

Nntains TSCA Confidential Business Information
Page 11 of 26 pages

Report 97 10 42 412 C

( __- _ct Eye Irritation/Corrosion

3 GuEDELINE

The study procedures were performed in compliance with the OECD-Guideline for

Testing of Chemicals 405 "Acute Eye Irritation / Corrosion", adopted February 24,

1987.

4. Protocol

Project No.: 97 1042412

Protocol No.: 97 10 42 412 C

Date of Protocol: November 14, 1997

Ratification: -November 20, 1997

Monitor:)

5. Test Substance

Name:

Characterization of

the test substance: alcohol alkoxylate

Main ingredients:(

Date of arrval: 'Nvember 05, 1997

Sample designation: 137291 3001

Purity: not stated

Physical state: liquid

Colour. colourless

Homogeneity: homogeneous

Stability: not stated

Expiry date: not stated

Storage conditions: not stated, storage class 10 (VCI)

In compliance with OECD-Guideline 405, a reference substance was not tested.

Contains ISCA Confidential Business Information 44
Page 12 of 26 pages

t 97 10 42 412 C

tAcute Eye Irrntation/Corrosion

6 METHOD "ACUTE EYE mRRATION/CoRROSION" IN COM~PLIANCE WITH

OECD-GUEUENE 405

6.1 ANIMALS AND LABORATORY CONDITIONS

Species: Albino rabbit

Identification numbers: 152, 153 and 154 (all female)

Breeding: Dr. Zimmermann

Schweizer Hof

D-73453 Untergrdningen/Aalen

Veterinary preliminary

examination: Without morbid signs

Diet: Raiffeisen Ringfutter Kaninchen

Trockenfutter 52/P Raiffeisen

Kraftfutterwerk Kehl, ad libitum.

Water: Community tap water from Karlsruhe; free

access for the animals by daily changing of

the watering-bottles.

Husbandry: The rabbits housed individually in wire

grating cages (area 2450 cmz, height 42 cm)

without bedding.

Temperature: 20 -23 "C

Relative humidity: 30-70 %

Light -dark -rhythm: 12:12

Contains TSCA Confidential Business Information
Page 13 of 26 pages

Report 97 10 42 412 C

L ) Acutt Eye Irritation/Corrosion

6.2 Composition of Diet and Water

Composition of diet "Raiffeisen Ringfutter Kaninchen".

Manufacturer. Raiffeisen Krallfutterwerke Said GmbH, Werk Kehl

Manufacturer specification of lot No. 5089 1/039

INGREDIENTS:

crude protein 17 %

crude fat 3 %

crude fibres 17.5 %

crude ash 8.5 %

calcium 1 %

phosphorus 0.6 %

sodium 0.2 %

ADDED VITAMINS PER KG DIET

vitamin A 12.000 I.E.

vitamin D3 1.000 I.E.

vitamiin E 50 mg

flavophospholipol 4 mg

BHT

Analysis performed on lot No. 5089 1/039 by Dr. Nedvidek /BioChem GmbH in

December 1994/January 1995.

INGREDIENTS:

moisture method: 115*C/ 5h 9.0%

ash method: 550*C / 16 h 9.5%

crude protein Kjeldahl method according

to BioChem SOP L 10 15.3%

crude fat Weibull Stoldt method in analogy

to § 35 LM]BG L 01.00-20 3.1 %

Contains ISCA Confidential Business Information
Page 14 of 26 pages

Report.97 10 42 412 C

J~ Acute Eye Irritation/Corrosion -l

CONTAMINANTS:Is

a-HCH < 0.01 mg/kg

B-HCH < 0.01 mg/kg

y-HCH < 0.01 mg/kg

8-HCH < 0.01 mg/kg

pentachlorobenzene, < 0.01 mg/kg

hexachlorobenzene < 0.01 mg/kg

heptachlor < 0.01 mg/kg

aldrin < 0.01 mng/kg

bromnophos < 0.01 mg/kg

heptachiorepoxide A + B < 0.01 mg/kg

cc-endosulfan < 0.01 mg/kg

dieldrin < 0.01 mg/kg

o,p-DDT < 0.01 mg/kg

p,p'-DDT < 0.01 mg/kg

o,p-DDE < 0.01 mg/kg

p,p'-DDE < 0.01 mg/kg

o,p-DDD < 0.01 mg/kg

p,p'-DDD < 0.01 mg/kg

endrin < 0.01 mg/kg

1-endosulfan < 0.01 mg/kg

methoxychlor < 0.01 mg/kg

rairex < 0.01 mg/kg

propazine < 0.01 mg/kg

atrazine < 0.01 mng/kg

atrazine-desethyl < 0.01 mg/kg

terbuthylazine < 0.01 mg/kg

prometryn < 0.01 mg/kg

amnetryn < 0.01 mg/kg

simetryn < 0.01 mng/kg

terbutryn < 0.01 mng/kg

malathion < 0.01 mg/kg

parathion-ethyl < 0.01 mg/kg

cyanazine < 0.01 mg/kg

simazine < 0.01 mg/kg

promecton < 0.01 mng/kg

quintozene < 0.01 mg

Contains TSCA Confidential Business Information 4~7
Page 15 of 26 pages

~Report 97 10 42 412 C

yAcute Eye Irritation/Corrosiona

COMPOSITON OF DRINKIJNG WATER:

according to the latest available information of Stadtwerke Karlsruhe (1995):

average of determinations

aluminium < 0.03 mg/i

aqmmonium < 0.05 mg/i

antimony < 0.01 mg/i

arsenic < 0.001 mg/I

barium 0.09 mg/I

boron 0.01 mg/i

cadmium < 0.0001 mg/i

calcium 119 mg/I

chromium < 0.001 mg/i

copper < 0.01 mg/i

iron < 0.01 mg/i

lead < 0.001 mg/i

magnesium 10.6 mg/i

manganese < 0.01 mg/i

mercury < 0.00004 mg/i

nickel < 0.003 mg/i

potassium 1.7 mg/i

selenium < 0.01 mg/I

silicon 5.7 mg/i

silver < 0.0005 mg/I1

sodium 11.3 mg/i

zinc < 0.02 mg/i

chlorides 25 mg/i

cyanides < 0.01 mg/I

fluorides 0.08 mg/i

nitrites < 0.01 mg/I

nitrates 6.0 mg/i

phosphates < 0.02 mg/i

sulfates 71 mg/i

Contains TSCA Confidential Business Information Z,
Page 16 of 26 pages

Report 97 10 42 412 C

( 1/ Acute Eye Irritation/Corrosion

dissolved organic carbon 1.4 mg/i

pesticides not verificable

polychlorinated biphenyls not venificable

polycyclic aromatic hydrocarbons < 0.00005 mg/i1

trihalogenmethane < 0.0001 mg/i

halogenated hydrocarbons < 0.01 mg/i

organic chlorinated compounds < 0.0005 mg/i

surfactants < 0.01 mg/i

6.3. ACCLIATIZATION / PREPARATIONS

The animals were acclimated to the test conditions for 5 days prior to the

administration.

24 h prior to the administration all rabbits were checked to detect eye lesions. No

defects were determined.

Contains TSCA Confidential Business Information 4
Page 17 of 26 pages

Reoort 97 10 42 4120C

t ' Acute Eye Irritation/co:rrosion

6.4 STUDY PROCEDURES

Acclimatization: December 05, 1997 - December 09, 1997

Administration: December 10, 1997

The test substance was applied in a single dose of 0. 1

ml into the right eye to each of 3 albino rabbits.

[ ')was placed into the conjunctival sac of the

right eye after gently pulling the lower lid away from

the eyeball. The lids were then gently hold together

for about one second in order to prevent loss of the

material.

24 h after administration the eye was carefulfly rinsed

with NaCI 0.9 % in order to remove the residual test

substance. The left eye was not treated and was used

for control.

Observation: December 10, 1997 - December 18, 1997

The study was finished on day 8 after the application

because no toxicological symptoms could be noted.

Judgement: Mortality, toxicological symptoms, behaviour, body

weight, eye reaction according to OECD-Guideline

405. The first examination was performed 1 hour

after administration.

On the first day after exposure the cornea was tested

with fluorescein solution to detect defects.

(Fluorescein SE Dr. Thilo & Co. GmbH; I ml

contains 1.7 mg of the disodium salt of fluorescein).

Contains TSCA Confidential Business Information34
Page 18 of 26 pages

Report 97 10 42 412 C

C _~'Acute Eye Irritation/Corrosion

6.5 EVALUATION OF THE OCULAR REACTION ACCORDING TO TABLE 1 OF

OECD-GUIDELINE 405

Table I

CONJUNCTIVAE

Redness (refers to palpebral and bulbar

conjunctivae, cornea and iris).

Blood vessels normal 0

Some blood vessels definitely

hyperaemnic (injected) 1

Diffuse, crimson colour, individual

vessels not easily discernible 2

Diffuse beefy red 3

Chemosis (Lids and/or nictatmng membranes)

No swelling 0

Any swelling above normal (includes

nictating membranes) I

Obvious swelling with partial

eversion of lids 2

Swelling with lids about half closed 3

Swelling with lids more than half closed 4

Contains TSCA Confidential Business Information
Page 19 of 26 pages

Report 97 10 42 412 C

J/ Acute Eye Irritation/Corrosion

CORNEA

Opacity: degree of density (area most dense taken for reading).

No ulceration or opacity 0

Scattered or diffuse areas of opacity (other

than slight dulling of normal lustre),

details of iris clearly visible 1

Easily discernible translucent area,

details of iris slightly obscured 2

Nacreous area, no details of iris visible,

size of pupil barely discernible 3

Opaque cornea, iris not discernible

through the opacity 4

IRIS

Normal 0

Markedly deepened rugae, congestion,

swelling, moderate circumcorneal hyperaemia,

or injection; any of these or combination

of any thereof; iris is still reacting to

light (sluggish reaction is positive)1

No reaction to light, haemnorrhage, gross

destruction (any or all of these) 2

Contains TSCA Confidential Business Information
Page 20 of 26 pages

Report 97 10 42 412 C

( iAcute Eye IrritationtCorrosion

7 FINDINGS "ACUTE EYE IRUITATION/CORROSION"

7.1 MORTALITY/TOXICOLOGICAL SYMPTOMS

After the administration of 0.1 ml test substance into the right eye none of the treated

rabbits died during the experiment and observation period. Systemic toxicological

symptoms could not be observed.

7.2 BEHAVIOUR

The general behaviour of the rabbits treated was quiet and watchful. The behavioural

pattern was not different to that of other rabbits which were not in the test.

7-3 BODY WEIGHT

During the experiment the body weight of all three rabbits increased. The weight of

each animal is specified in table 11.

Table 11: Body weight (statements in g):

Phase of test day Rabbit No.

152 153 154

Acclimatization -5 2627 2817 2832

-4 2643 2818 2894

-3

-2 2733 2860 2922

-1 2778 2924 3002

Administration 0 2835 2901 2995

Observation 1 2857 2889 3047

2 2926 2949 3109

3 2951 2974 3095

4-

5 2989 3012 3135

6 3041 3057 3140

7 3085 3073 3270

_________ 8 3108 3079 3281

Contains TSCA Confidential Business Information
Page 21 of 26 pages

Report 97 10 42 412 C

(_( Acute Eye Irritation/Corrosion

7.4 EYE REACTON

The test substance was applied in a single dose of 0. 1 ml into the right eye to each of 3

albino rabbits.

After evaluation of the eye reaction according to table 1 of the OECD-Guideline 405, all

three rabbits developed hyperaemnic blood vessels. This effect was estimated at grade

0.5 to 2.5 (maximum possible 3). Hyperaemnic blood vessels were not observed any

more on day 5.

All three rabbits showed swelling of lids. This effect was estimated at grade I to 3

(maximum possible 4). Swelling of lids was not observed any more on day 5.

Two rabbits showed opacity of the cornea 48 hours after application. This effect was

estimated at grade 1 (maximum possible 4). Opacity was not observ ed any more on day

3.

Ocular lesions of iris could not be observed during the study.

Exsudation was observed in the eyes of all three rabbits after instillation of the test

substance till day 1.

The effects attributed to the administration of~ )Were completely reversible in the

eyes of all three~ rabbits which was shown durinig thle iast four days of the study.

In the untreated left eye which was used as control pathological symptoms could not be

noted.

The individual findings of each rabbit are specified in table 11-V. The tables VI and VII

indicate the grade summary and the mean value of grades of ocular lesions of the

animals.

Contains TSCA Confidential Business Information 6
Page 22 of 26 pages Report 97 10 42 412 C

11 Acute Eye Irritation/Corrosion

Table Ml: Findings, rabbit No. 152, female

Grades of ocular lesions according to OECD-table

Administration: test substance in the right eye

Control: untreated left eye

conjunctivae

time after cornea iris redness chemosis

admiinistration

right left right left right left right left

administration 0 0 0 0 0.5 0 0 0

I St hour 0 0 0 0 2 0 3 0

day 1 00- 00- 0 0 2 0 2 0

day 2 1 0 0 0 2 0 1 0

day 3 0 0 0 0 1 0 1 0

day 4 - - - - - - - -

day 5 0 0 0 0 0 0 0 0

day 6 0 0 0 0 0 0 0 0

day 7 0 0 0 0 0 0 0 0

day 8 0 0 0 0 0 0 0 0

*A fluorescein - test was performed in order to demonstrate defects of the cornea; in this

case:

day 1: right eye (test substance) negative()

left eye (control) negative (-)

Exsudation was observed in the eyes of the rabbit till day 1 of the observation period.

Contains TSCA Confidential Business Information 6
Page 23 of 26 pages

Report 97 10 42 412 C

C cut Eye Irnitation/Corrosion

Table IV: Findings, rabbit No. 153, female

Grades of ocular lesions according to OECD-table

Administration: test substance in the right eye

Control: untreated left eye

conjunctivae

time after cornea iris redness chemosis

administration

right left right left right left right left

administration 0 0 0 0 0.5 0 0 0

I sthour 0 0 0 0 1.5 0 2.5 0

day 1 00- 00- 0 0 2 0 2 0

day 2 0 0 0 0 1.5 0 1 0

day 3 0 0 0 0 1 0 1 0

day 4 - - - - - - - -

day 5 0 0 0 0 0 0 0 0

day 6 0 0 0 0 0 0 0 0

day 7 0 0 0 0 0 0 0 0

day 8 0 0 0 0 0 0 0 0 _

'A fluorescein - test was performed in order to demonstrate defects of the cornea; in this

case:

day 1: right eye (test substance) negative()

left eye (control) negative(-

Exsudation was observed in the eyes of the rabbit till day 1 of the observation period.

Contains TSCA Confidential Business Information
Page 24 of 26 pages

Report 97 10 42 412 C

_) Acute Eye irritationicorrosion

Table V: Findings, rabbit No. 154, female

Grades of ocular lesions according to OECD-table

Administration: test substance in the right eye

Control: untreated left eye

conjunctivae

time after cornea iris redness chemosis

administration

right left right left right left right left

administration 0 0 0 0 0.5 0 0 0

1St hour 0 0 0 0 2.5 0 3 0

dayl 1(00 00- 0 0 2 0 2.5 0

day 2 1 0 0 0 2 0 2.5 0

day 3 0 0 0 0 1 0 2 0

day 4 - - - - - - - -

day 5 0 0 0 0 0 0 0 0

day 6 0 0 0 0 0 0 0 0

day 7 0 0 0 0 0 0 0 0

day 8 0 0 0 0 0 0 0 0

* A fluorescein - test was performed in order to demonstrate defects of the cornea; in this

case:

day 1: right eye (test substance) negative(-

left eye (control) negative(-

Exsudation was observed in the eyes of the rabbit till day 1 of the observation period.

Contains ISCA Confideantial Business Information

S7/
Page 25 of 26 pages

Report 97 10 42 412 C

( -Acute Eye Irritation/Corrosion

Table VI: Findings, rabbit No. 152, 153 and 154

Grade summary of ocular lesions

Administration: test sustance in the right eye

Control: untreated left eye

conjunctivae

time after cornea iris redness chemosis

administration

right left right left right left right left

administration 0 0 0 0 1.5 0 0 0

isthour 0 0 0 0 6 0 8.5 0

dayl1 0 0 0 0 6 0 6.5 0

day 2 2 0 0 0 5.5 0 4.5 0

day 3 0 0 0 0 3 0 4 0

day 4 - - - - - - - -

day 5 0 0 0 0 0 0 0O 0

day 6 0 0 0 0 0 0 0 0

day 7 0 0 0 0 0 0 0 0

day 8 0 0 0 0 0 0 0 0

Table VII: Findings, rabbit No. 152, 153 and 154

Mean value of grades of ocular lesions

Administration: test substance in the right eye

Control: untreated left eye

conjunctivae

time after cornea iris redness chemosis

administration

right left right left right left right left

administration 0 0 0 0 0.5 0 0 0

ist hour 0 0 0 0 2 0 2.8 0

dayl1 0 0 0 0 2 0 2.2 0

day 2 0.7 0 0 0 1.8 0 1.5 0

day 3 0 0 0 0 1 0 1.3 0

day 4 - - - - - - - -

dayS5 0 0 0 0 0 0 0 0

day 6 0 0 0 0 0 0 0 0

day 7 0 0 0 0 0 0 0 0

da8 0 0 0 0 0 0 0 0

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Page 26 of 26 pages

Report 97 10 42 412 C

1Acute Eye Irrtation/Corrosion

8 ARCHIVING

BioChemn GmbH, 76185 Karlsruhe, Germany, will archive raw data, the protocol, a

copy of the report and the test article archive sample for the stipulated period of time

according to GLP regulations.

Contains TSCA ConfideritiaI Busine~s Information
autacnment 9

Daimlerstral~e 

Tel. 07.21/97 37-0 

5 b 

:Fax07 

-U-76185 

21/97 

Kartsnhe 

37-222 BioChem

E-mail: biochem@biochem.de Labor fOr biologische und

Internet: http:/Avwwbiochem.de chemische Analytk GmbH

REPoRT 97 10 42 412 B

AcuTE DERMAL IRRIATION/CORROSION

[E

Project No.: 971042412

Report No.: 97 10 42 412 B

Test substance:()

Batch No: not stated

BioChemn sample designation: 137291 2001

Date of arrival: November 05, 1997

Sponsor:

Monitor.

Test facility: BioChem GmibH

Daimlerstr. 5 b

D-76 185 Karlsruhe

Germany

Management: Dr. W. Massmann

Dr. M. MUller

Study Director. Dr. H.-J. Kramer

Quality Assurance Manager. Dr. H. Merten

Karlsruhe, December 22, 1997

BioChem, GmbH

Page 1 of 23 pages

Report 97 10 42 412 B

YAcute Dermal Irritation/Corrosion

Contains TSCA Confidential Business Information c
Page 2 of 23 pages Report ?7 1042 412 B

Acute Dermal Irritation/Corrosion

Statement of GLP compliance

We declare that all tests are made in accordance with the protocol and with the regulations of the

OECD-Principles of the "Good Laboratory Practice" (GLP).

GLP: "Good Laboratory Practice" Regulations of the European Communities, EC, enacted in

Germany in the "Chemikaliengesetz" (Chemicals Act), 2nd Amendment, dated July 25, 1994,

BGBI I, pp. 1703, 1994 and "AndVO" dated May 14, 1997, BGBI1. 1, pp. 1060, 1997.

The documented data correspond to the measured values.

Management

Dr. Manfred Miller Date

Study Director

Dr. Hans-Joachim Kramer Date

Quality Assurance Manager

Dr. Hans Merten Date

Contains TSCA Confidential Business Information

61
Page 3 of 23 pages

Report 97 10 42 412 B ( - Acute Dermal Irritation/Corrosion

TUmweltmii terium

Baden-Wuarttemberg

GLP-Bescheinigung

Bescheinigu ng Certificate

~Flerm wird bestaWgt, daB die It is hereby certified tha the test

Pnohzngseinrichtung fiacility

in 76185Kar~sruhe in 76185Karlsruhe

der BioChem, Labor fllrbiologische und of BioChe2, Labor ffir biologische umd

chemische Analytik GimbH, chemische Analytik GmbH,

Daimlerst. .5b . Daiinlerstr. .5b

am 19J20.06.95, 27.09.95 und 26.01.96 on 19120. June 95, 27. September 95

and 26. Januiary 96

von der fir die 10bei-wachung zustgndigen was inspected by the competent

Beharde Ober die Einhaltung der (3rund- authority regarding compliance with the

s~tze der Guten Laborpra-,ds inspiziert -Principles of Good Laboratory Practice.

worden ist.

Es wird biermit bestifigt, daB folgende It is hereby certified that studies in this test

Prflmigen in dieser Pnafeinzichtung nach facility are conducted in compliance with the

den Girundsitzen der Guten Laborpra~ds Principles of Good Laboratory Practice.

durchgefMllrt werden.

Sttgrden " R

o

Seite 1/2 page 1/2

Contains ISCA Confidential BQsinoss Information
Tage 4 of 23 pages

Report 97 10 42 412 B

Acute Dermal Irritation/Corrosion

S cite =/ zur GLP-Bescheinigung

Prluhngskategorie 1 Prifungen auf physikalischchemische Eigenschzften und

Gehaltsbestinzmungen

Priifungskategorie 2 Prilungen auf toxikologische

Eigenschaften

Priifungskategorie 3 Pz:Afungen auf mutagene

Eigenschaften (in vitro, in vivo)

Priifuugskatcgorie 4 Umwelttoxikologische Pr~Un gen

zu Auswirknngen auf aquatische

and terrestrische Organismen

Priifungskategorie 5 Prilfungen zum Verhalten im

Boden, im Wasser und in der

Luft; Bioakkumulation;

Metabolismus

Priifungskateg-orie 6 Prilfungen auf Riickstinde

Dies entspricht den Kategorien nach der Ailgemeinen Verwaltungsvorschrift mim

Verfahren der behdrdlichen O0berwacbnng der ihaltung der Guten Laborpraxds

(ChemVwV-GLP) vom 29. Oktober 1990:

"Physikalisch-chemische Eigenschaften unci

Gehaltsbestimmungen"

"Toxikologische Eigenschaften"

"0 kotoxikologiche Eigeuschafteu"

"Verhalten im Boden, im Wasser und in der Luft"

"RrIckstfinde"

Stuttgart, den 26.02.1996

14 .A- 4 WORTM~BErG

Dr. Albrecht

Contains TSQA Confidential Business Information

Seite 2/2 page 212 6

onIans TSCA rnf~i-oIflW
Page 5 of 23 pages Report 97 10 42 412 B

VAcute Dermal Irritation/Corrosion

Translation of page 2 of the GLP-Certificate by BioChem.

Test category 1: Tests on physico-chemical

properties and content

determinations

Test category 2: Tests on toxicological

properties

Test category 3: Tests on mutagenic properties

(in vitro, in vivo)

Test category 4: Ecotoicological tests on.

effects on aquatic and

terrestrial organisms

Test category 5: Tests on behaviour in soil, in water

and in the air,

bioaccumulation; metabolism

Test category 6: Tests on residues

T"his corresponds to the categories according to the General Administration

Regulation of the Procedure of the Official Supervision of the Compliance with Good

Laboratory Practice (Chem VwV-GLP) dated October 29, 1990:

"Physico-chemical Properties and Content Determinations"

"Toxicological Properties"

"Ecotoxicological Properties"

"Behaviour in Soil, in Water and in the Air"

"Residues"

Stuttgart February 26, 1996

Bade-WiittemergContains TSCA Confidential Business inf ormation

Department of the Environment
Page 6 of 23 pages

oR eport10 42 412 B

YAueDermal Irritation/Corrosion

Quality Assurance Unit Statement

Title: Final Report

Acute Dermial Irritation /Corrosion

project number- 97 1042 412

protocol number- 97 10 42412 B

Guideline: OECD-Guideline for Testing of Chemicals No. 404 "Acute

Dermal Irritation /Corrosio&"

Study / test system: Determination of the potential of[ Ito provoke skin

irritation /corrosion /albino rabbits ~

Test substance:Q

Batch No.: not stated

Sample designation: 137291 2001

Date of arrival: November 05, 1997

Study Director: Dr. H.-J. Kramer

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Page 7 of 23 pages

Report 97 10 42 412 B

!-~ Acute Dermal Irritation/Corrosion

The following checks were effected:

reports to the Study

Director and to the

date verified procedure Management

November 14, 1997 protocol November 14, 1997

December 09, 1997 audit 1 December 09, 1997

December 16, 1997 audit 2 December 16, 1997

December 19, 1997 final report in English December 19, 1997

The data which were taken as a basis for this report were verified by the Quality Assurance Unit.

The accordance of the study with its protocol, the OECD-Guideline for Testing of Chemicals No.

404 and the Principles of Good Laboratory Practice (GLP) is guaranteed.

The methods, procedures and observations are exactly described in the report. The Quality Assurance Unit ensures that the reported results exactly describe the raw data of the study.

The Study Director and the Management are generally informed of any findings of the Quality

Assurance Unit.

As for this study no departures from the prescribed course of the test were ascertained.

Karlsruhe, December 22, 1997 Dr. H. Merten

(Quality Assurance Manager)

Contains TSCA Confidential Business Information
Page 8 of 23 pages

Report 97 10 42 412 B

(~ )/Acute Dermal Irritation/Corrosion

Contents

page

1 Preface 9

2 Summary 10

3 Guideline 11

4 Protocol 11

5 Test Substance I1I

6 Method "Acute Denmal Irritation /Corrosion" 12

6.1 Animals and laboratory conditions 12

6.2 Composition of diet and water 13

6.3 Acclimatizaion /preparations 16

6.4 Study procedures 17

6.5 Evaluation of the skin reaction 18

7 Findings "Acute Dermal Irritation /Corrosion" 19

8 Archiving 23

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Page 9 of 23 pages

Report 97 10 42 412 B

( '/Acute Dermal Irritation/Corrosion

I Preface

Test system: Acute Dermal Irritation /Corrosion

Laboratory animals: Albino rabbits

Guideline: OECD-Guideline for Testing of Chemicals,

Section 4, 404 "Acute Dermal Irritation / Corrosion", adopted July 17, 1992

GLP: Gesetz zum Schutz vor gefdhrlichen

Stoffen (Cheinikaliengesetz ChemG, § 19 a-d

yam 25. Juli 1994)

Project No.: 971042412

Report No.: 97 1042 412 B

Test substance:

Sponsor:

Monitor

Test Facility: BioChem GmbH

DaimlerstraBe 5 b

D-76 185 Karlsruhe

Germany

Management: Dr. W. Massmann

Dr. M. Mailer

Study Director. Dr. H.-J. Kramer

Quality Assurance Manager: Dr. H. Merten

Contains TSCA Confidential Business Information
Page 10 of 23 pages

Repr97 10 42 412 B ( 'Acute Dermal Irritation/Corrosion

2 Summary

Report: Acute Dermal Irritation /Corrosion of in

compliance with OECD-Guideline 404

Sponsor:

Test Facility: BioChem GmbH

DaimlerstraBe 5 b

D-76 185 Karlsruhe

Germany

Project No.: 971042412

Protocol No.: 97 10 42412 B

Report No.: 97 10 42 412 B

Findings: The test substance was applied in a single dose of 0.5

ml dermally to each of 3 albino rabbits.

Under the conditions of this experiment did

not produce any acute systemic toxicological signs or

mortality.

After evaluation of table I in OECD Guideline 404

(grades of skin lesions - page 18 of this report) in the

following observation period none of the three rabbits

showed a dermal irritation of the exposure area.

During the experiment all rabbits showed a gain in

body weight.

Contains ISCA Confidential Business Information
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Report 97 10 42 412 B

( )IAcute Dermal Irritation/Corrosion

3 Guideline I b

The study procedures were performed in compliance with the OECD-Guideline for

Testing of Chemicals 404 "Acute Dermal Irritation / Corrosion", adopted July 17, 1992.

4. Protocol

Project No.: 971042412

Protocol No.: 97 10 42 412 B

Date of Protocol: November 14, 1997

Ratification: November 20, 1997

Monitor. 

'

5. Test Substance

Name:

Characterization of

the test substance: alcohol alkoxylate

Main ingredients:()

Date of arrival: Yovember 05, 1997

Sample designation: 137291 2001

Purity: not stated

Physical state: liquid

Colour colorless

Homogeneity: homogeneous

Stability: not stated

Expiry date: not stated

Storage conditions: not stated, storage class 10 (VCI)

In compliance with OECD-Guideline 404, a reference substance was not tested.

Contains TSCA Confidential Business Information
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/Acute Dermal Irritation/Corrosion

6 Method "Acute Dermal Irritation/Corrosion" in Compliance with

OECD-Guideline 404

6.1 Animals and Laboratory Conditions

Species: Young albino rabbits

Identification numbers: 149, 150, 151 (all male)

Breeding: Dr. Zimmermann

Schweizer Hof

D-73453 Untergrdningen/Aalen

Veterinary preliminaryGemn

examination: Without morbid signs

Diet: Raiffeisen-Ringfu~tter, Kaninchen Trockenfutter 52/P,

Raiffeisen Kraftfutterwerk Kehl, ad libitum.

Water- Community tap water from Karlsruhe; free access for

the animals by daily changing of the watering-bottles.

Husbandry: The rabbits housed individually in wire grating cages

(area 2450 c&, height 42 cm) without bedding.

Temperature: 20 - 23 0 C

Relative humidity: 30-70 %

Light-dark-rhythm: 12: 12

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Page 13 of 23 pages

Report 97 10 42 412 B L Acute Dermal Irritation/Corrosion

6.2 Composition of Diet and Water

Composition of diet "Raiffeisen Ringfutter Kaninchen".

Manufacturer. Raiffeisen Kraftfutterwerke Sold GmbH, Werk Kehl

Manufacturer specification of lot No. 50891/039

INGREDIENTS:

crude protein 17 %

crude fat 3 %

crude fibres 17.5 %

crude ash - 8.5 %

calcium I %

phosphorus 0.6 %

sodium 0.2 %

ADDED VITAMINS PER KG DIET

vitamin A 12.000 I.E.

vitamin D3 1.000 I.E.

vitamnin E 50 mg

fiavophospholipol 4 mg

BHT

Analysis performed on lot No. 50891/039 by Dr. Nedvidek /BioChem GmbH

in December 1994/January 1995.

INGREDIENTS:

moisture method: 115*C/ 5h 9.0%

ash method: 550 'C /16 h 9.5%

crude protein Kjeldahl method according

to BioChem. SOP L 10 15.3 %

crude fat Weibull Stoldt method in analogy

to §35 LMBGL 01.00-20 3.1 %

Contains TSCA Confidential Business Information
Page 14 of 23 pages

Report 97 10 42 412 B ( Acute Dermal Irritation/Corrosion

CONTAMINANTS:

oc-HCH < 0.01 mg/kg

1-HCH < 0.01 mg/kg

y-HCH < 0.01 mg/kg

8-HCH < 0.01 mg/kg

pentachlorobenzene < 0.01 mg/kg

hexachlorobenzene < 0.01 mg/kg

heptachlor < 0.01 mg/kg

aldrin < 0.01 mg/kg

bromophos < 0.01 mg/kg

heptachiorepoxide A + B < 0.01 mg/kg

cc-endosulfan < 0.01 mg/kg

dieidrin < 0.01 mg/kg

o,p-DDT < 0.01 mg/kg

p,p'-DDT < 0.01 mg/kg

o,p-DDE < 0.01 mg/kg

p,p'-DDE < 0.01 mg/kg

o,p-DDD < 0.01 mg/kg

p,p'-DDD < 0.01 mg/kg

endrin < 0.01 mg/kg

f3-endosulfan < 0.01 mg/kg

methoxychlor < 0.01 mg/kg

mirex < 0.01 mg/kg

propazine < 0.01 mg/kg

atrazine < 0.01 mg/kg

atrazine-desethyl < 0.01 mg/kg

terbuthylazine < 0.01 mg/kg

prometlyn < 0.01 mg/kg

ametryn. < 0.01 mg/kg

simetryn < 0.01 mg/kg

terbutryn < 0.01 mg/kg

malathion < 0.01 mg/kg

parathion-ethyl < 0.01 mg/kg

cyanazine < 0.01 mg,/kg

simazine < 0.01 mg/kg

prometon < 0.01 mg/kg

quintozene < 0.01 mg/kg

Contains TSCA Confl~ntiAic'oe 1mm-nn 7
Page 15 of 23 pages (eport 97 10 42 412 B

)Acute Dermal Irritation/Corrosion

COMPOSrrON OF DRINKING WATER:

according to the latest available information of Stadtwerke Karlsruhe (1995):

average of determinations

aluminium < 0.03 mg/i

ammonium < 0.05 mg/I

antimony < 0.01 mg/i

arsenic < 0.001 mg/I

barium 0.09 mg/I

boron 0.01 mg/I

cadmium < 0.0001 mg/I

calcium 119 mg/I

chromium < 0.001 mg/I

copper < 0.01 mg/i

iron < 0.01 mg/I

lead < 0.001 mg/I

magnesium 10.6 mg/i

manganese < 0.01 mg/i

mercury < 0.00004 mg/I

nickel < 0.003 mg/i

potassium 1.7 mg/I

selenium < 0.01 mg/i

silicon 5.7 mg/i

silver < 0.0005 mg/i

sodium 11.3 mg/I

zinc < 0.02 mg/i

chlorides 25 mg/i

cyanides < 0.01 mg/I

fluorides 0.08 mg/i

nitrites < 0.01 mg/I

nitrates 6.0 mg/I

phosphates < 0.02 mg/I

sulfates 71 mg/l

Contains TSCA Confidential Business Information
Page 16 of 23 pages

Report 97 10 42 412 B

( Acute Dermal Irritation/Corrosion Ps.

dissolved organic carbon 1.4 mg/i

pesticides not verificable

polychlorinated biphenyls not verificable

polycyclic aromatic hydrocarbons < 0.00005 mg/i

trihalogenmethane < 0.0001 mg/i

halogenated hydrocarbons < 0.01 mg/I

organic chlorinated compounds < 0.0005 mg/I

surfactants < 0.01 mg/I

6.3 Acclimatization / Preparations

The animals were acclimated to the test conditions for 5 days prior to the administration.

On the fifth day the dorsal area of the trunk was shaved in anarea of 3 x5cm'.

Contains TSCA Confidential Business Information
Page 17 of 23 pages

Report 97 10 42 412 B

( _ Acute Dermal Ir-ritat on/Corrosion

6.4 Study Procedures

Acclimatization: December 04, 1997 - December 08, 1997

Administration: December 09, 1997

The test substance was applied undiluted dermally in

a single dose of 0.5 mll animal onto the dorso-lumbar

,region of 3 albino rabbits. The region treated was

protected with a patch (width 6 cm) (Werovil, No. 15

00 20, WERO-MEDICAL, D-65232 Taunusstein). It

was held in place for a period of 4 hours by an occlusive bandage made from a dressing (width 8 cm,

with selvages, coated completely with cellophane)

(No. 09 00 22, WERO-MEDICAL). The bandage

was fixed with 1.25 cm broad strips of Adhesive

Tape ST (WERO-MEDICAL).

After removal of the bandage at the end of the exposure period the residual test substance was removed

by gently washing with water.

Observation: December 09, 1997 - December 18, 1997

The study was finished on day 9 after the application

because no toxicological symptoms could be noted

Judgement: Mortality, toxicological symptoms, behaviour, body

weight, skin reactions.

Contains TSCA Confidential Business Information
Page 18 of 23 pages

Report 97 10 42 412 B

t Acute Dermal Irritation/Corrosion

6.5 Evaluation of the Skin Reaction (according to table 1 in OECD-Guideline

Table I - 1: Erythema and eschar formation

Value

No erythema 0

Very slight erythema

(barely perceptible) I

Well-defined erythema 2

Moderate to severe

erythema 3

Severe erythema (beet

redness) to eschar

formation preventing

grading of erythema 4

maximum possible 4

Table 1- 2: Oedema formation

Value

No oedemna 0

Very slight oedema,

(barely perceptible) I

Slight oedema (edges

of area well defined

by definite raising) 2

Moderate oedema.

(raised approximately

1 mm) 3

Severe oedema

(raised more than 1 mm

and extending beyond

area of exposure) 4

maximum possible 4

Contains TSCA Confidential Business Information
Page 19 of 23 pages

Report 97 10 42 412 B

L Acute Dermal Irritation/Corrosion

7 Findings "Acute Dermal Irritation/Corrosion"

Mortality: After the dermal administration of 0.5 nml ( )per

animal for a period of 4 hours none of the 3 rabbits

died.

Toxicological symptoms: During the observation period of 9 days none of the

animals showed any systemic toxicological symptoms.

Behaviour: The reaction of the rabbits treated was quiet and

watchful. The behavioural pattern was not different to

that of other rabbits which were not in the test.

Body weight: During the experiment the body weight of all six

rabbits had an increase in weight.

Contains TSCA Confidential Business Information
Page 20 of 23 pages

Report 97 10 42 412 B

C IAcute Dermal Irritation/Corrosion

Table 11: Body weight (statements in g):

3 rabbits, male

Identification numbers: 149 - 151

______________ Rabbit No. ________

phase of test day 149 150 151

acclimatization -5 2772 2580 2770

-4 2795 2646 2879

-3 2879 2669 2914

-2

_________ -l 2875 2670 2978

administration 0 2848 2733 3042

observation 1 2946 2783 3166

2 2998 2819 3142

3 3110 2851 3286

4 3116 2940 3357

5-

6 3219 2932 3356

7 3214 3016 3386

8 3221 3028 3469

________ 9 3307 3080 3478

Contains TSCA Confidential Business Information
Page 21 of 23 pages

Report 97 10 42 412 B

( _/ Acute Dermal Irritation/Corrosion

Skin reaction: After removing the occlusive bandage after a period 

Is®

of 4

hours, the exposure area was washed and cleaned with water.

In the following observation period all three rabbits showed

no dermal irritation of the exposure area.

The findings of the animals 149, 150, and 151 are specified

in table M - V.

Table VI shows the mean value of all assessments.

Table Ill: Acute Dermal Irritation/Corrosion.

Evaluation of the sin reaction.

Rabbit 149, male

Time after administration Test substance Control

_____________ Erytema Oedema Erytema Oedema

Ibhrafter bandage removal 0 0 0 0

dayl1 0 0 0 0

day 2 0 0 0 0

day 3 0 0 0 0

day 4 0 0 0 0

day 5 -

day 6 0 0 0 0

day 7 0 0 0 0

day 8 0 0 0 0

day 9 0 0 0 0

Skin reaction control: Table III, IV, V and VI

The shaved area close to the exposure area was used as the

control.

The control area did not show any pathological skin reaction.

According to table I of OECD-Guideline 404, the value was

always classified as 0.

Contains TSCA Confidential Business Infor=6O

efo
Page 22 of 23 pages

Report 97 10 42 412 B

JAcute Dermal Irritation/Corrosiona

Table IV: Acute Dermal Irritation/Corrosion.

Evaluation of the skin reaction.

Rabbit 150, male

Time after administration Test substance Control

___________ Erytema, Oedemna Erytema Oedema

Ilhr after bandage removal 0 0 0 0

dayl1 0 0 0 0

day 2 0 0 0 0

day 3 0 0 0 0

day 4 0 0 0 0

day 5 -

day 6 0 0 0 0

day 7 0 0 0 0

day 8 0 0 0 0

day 9 0 0 0 0

Table V: Acute Dermal Irritation/Corrosion.

Evaluation of the skin reaction.

Rabbit 151, male

Time after administration Test substance Control

______Eryth_____ema Oedema Erytema Oedema

Ilhr after bandage removal 0 0 0 0

dayl1 0 0 0 0

day 2 0 0 0 0

day 3 0 0 0 0

day 4 0 0 0 0

day 5 -

day 6 0 0 0 0

day 7 0 0 0 0

day 8 0 0 0 0

day 9 0 0 0 0

Contains ISCA Confiential Business Information
-- - -L_ _ _ _ _ __ __ _ _ _ _ _ _

Page 23 of 23 pages

Reprt97 10 42 412 B

('Acute Dermal Irritation/Corrosion Is,

Table VI: Acute Dermal Irritation/Corrosion.

Evaluation of the skin reaction.

Mean values of sin irritation scores of all three animals

Time after administration _ Test substance Control

_____________ Erytema Oedema E ema Qedemna

Ibhrafter bandage removal 0 0 0 0

dayl1 0 0 0 0

day 2 0 0 0 0

day 3 0 0 0 0

day 4 0 0 0 0

day 5 -

day 6 0 0 0 0

day 7 0 0 0 0

day 8 0 0 0 0

day 9 0 0 0 0

9 Archiving

BioChemn GmbH, D-76185 Karlsruhe, Germany, will archive raw data, the protocol, a

copy of the report and the test article archive sample for the stipulated period of time

according to GLP regulations.

Contains TSCA Confidential Business Information
-~ attachment 10

Daimlerstrafle 5 b -D-76185 Karisrhe. Bioh- Tel. 07.21/9737-0 -Fax 0721/9737-222Bi he

E-mail: biochem~biochem.de Labor fOr bialagische und Internet: htpiMwmbiochem.de chemische Analytik GmbH

Report 97 10 42 412 A

Acute Oral Toxicity

Project No.: 971042412

Report No.: 97 1 At2 412 A

Test substance: 1

Batch No: not stated

BioChem sample designation: 137291 1 001

Date of arrival: November 05, 1997

Sponsor

Monitor.)

Test facility: BioChem GmbH

Daixnlerstr. 5 b

D-76 185 Karlsruhe

Germany

Management: Dr. W. Massmaxm

Dr. M. Miller

Study Director. Dr. H.-J. Kramer

Quality Assurance Manager Dr. H. Merten

Karlsruhe, January 07, 1998

BioChem GmbH

Page 1 of 22 pages

Report 97 10 42 412 ACotisTCCofdnalBies frm

(j '/Acute Oral Toxicity CVtiSTC ofdnilBsns nom:
Page reort 2 of 22 pages 97 10 42 412 A

~/Acute Oral Toxicity

Statement of GLP compliance

We declare that all tests are made in accordance with the protocol and with the regulations of the

OECD-Principles of the "Good Laboratory Practice" (GLP).

GLP: "Good Laboratory Practice" Regulations of the European Communities, EC, enacted in Germany in the "Chemikaliengesetz" (Chemicals Act), 2nd Amendment, dated July 25, 1994,

BGBI 1, pp. 1703, 1994 and "AndVO" dated May 14, 1997, BGBl. 1, pp. 1060, 1997.

The documented data correspond to the measured values.

Management

... . 6...........................

Dr. Manfred Mfiller Date

Study Director

Dr. H-ans-Joachim Kramer Date

Quality Assurance Manager

Dr. Hans Merten Date

Contains TSCA Confidential Business Information
Page 3 of 22 pages -. Reoort97l1042412 A ( 'fAcute Oral Toxicity

Umweltmin isterium

Baden-Wfirttemberg

GLP-Beschem*Iigung

Bescheinigung Certificate

Hermit wird bestktgt, daB die It is hereby certified that the test

Prilfungseinrichtung ficlity

in 76185 Krisruhe in 76185 Karlsruhe

der BioChem, Labor fr biologiche und of BioChezn, Labor far biologiche und

chemische Analytik GmbH, chemische Analytik GmbH,

Daimlerstr. 5b . Daimlerstr. 5b

am 19J20.06.95, 27.09.95 und 26.01.96 'on 19J20. June 95, 27. September 95

and 26.IJanuary 96

von der ffir die Oberwachung mistandigen was inspected by the competent

Beharde Ober die Einhaltung der Glrund- authority regarding compliance with the

sA~e der Guten Laborpraxis inspiziert .Principles of Good Laboratory Practice.

worden ist.

Es wird hiermit bestltigt, daB folgende It is hereby certified that studi~s in this test

Pnifingen in dieser Prafeinxichtung nach facility are conducted in compliance with the

den Grundsitzen der Guten Laborpraxis Principles of Good Laboratory Practice.

durchgeflhrt werden.

BADEN-

-Stuttgart, den RMERG

Seite 1/2 page 1/2 4-

Contains TSCA Confidential Business information
'Page 4 of 22 pages

Report 97 10 42 412 A ( Acute Oral Toxicity

Seite 2/2 mir GLP-Bescheinigung

Prilfungskategorie 1 Pilfungen auf physikalischchemische Eigenschaften umd

Gehaltsbestimmangen

Priifungskategorie 2 Prifungen iuf toxikologische

Eigenschaften

Priifungskategorie 3 Piflfungen auf mutagene

Eigenschaften (Cm vitro, in vivo)

Priifungskategorie 4 Umweittoxikologische Priffun-en

zu Auswikngen auf aquaibche

and terrestrische Organismen.

Priifungskategorie 5 Priifungen zam Verhalten im

Boden, im Wasser und in der

Luft;.Bioakkumulatdou;

Metabolismus

Priifungskategorie 6 Prifungen auf Rfickstinde

* Dies entsprichf den Kategorien nach der Ailgemeinen Verwaltungsvorscbrift mmn

Verfabren der behdrdlichen tOberachung der Ezinhaltung der Guten Laborpaxs

(ChemVwV-43LP) vom 29. Qlctober 1990:

"Physkalsch-chemische Elgenschaften and

Gehaltsbestimmungen"

"Toxikologische Eigenschaften"

"Okotoxikologische Eigenschaften"

"Verhalten im Boden, im Wasser und in der Luft"

"Rickstfinde"

Stuttgart, den 26.02.1996

MODEN.

Dr. lbreht ontans SCA Confidenltial Business 1nf or-m--'

Seite 2/2 4- ~page 212
Page 5 of 22 pages Revort 97 10 42 412 A

/ Acute Oral Toxicitya Is,

Translation of page 2 of the GLP-Certificate by BioChem.

Test category 1: Tests on physico-chemical

properties and content

determinations

Test category 2: Tests on toxicological

properties

Test category 3: Tests on mutagenic properties

(in vitro, in vivo)

Test category 4: Ecotoxicological tests on*

effects on aquatic and

terrestrial organisms

Test category 5: Tests on behaviour in soil, in water

and in the air,

bioaccumulation; metabolism

Test category 6: Tests on residues

This corresponds to the categories according to the General Administration

Regulation of the Procedure of the Official Supervision of the Compliance with Good

Laboratory Practice (Chem VwV-GLP) dated October 29, 1990:

"Physico-chernical Properties and Content Determinations"

"Toxicological Properties"

"Ecotoxicological Properties"

"Behaviour in Soil, in Water and in the Air"

"Residues"

Stutgar, bruryF 6, 996Contains TSCA Confid,.ntcl~ Business lnform t,:c

Baden-Wiirttemberg

Department of the Environment
Page 6 of 22 pages

Report 97 10 42 412 A

-( '/Acute Oral Toxic ity Is,

Quality Assurance Unit Statement

Title: Final Report

Acute Oral Toxicity

E -7'-

project numbe 971042412

protocol numbe 97 10 42 412 A

Guideline: OECD-Guideline for Testing of Chemicals No. 401 "Acute

Oral Toxicity", adopted February 24, 1987

Study Itest system: Judgement of the acute oral toxicity/Albino rats (SpragueDawley)

Test substance:

Batch No.: not stated

Sample designation: 137291 1 001

Date of arrival: November 05, 1997

Study Director: Dr. H.-J. Kramer

contains ISCA Confidenl Buiness information
Page 7 of 22 pages

Report 97 10 42 412 A

( Acute Oral Toxic ity

The following checks were effected:

reports to the Study

Director and to the

date verified procedure Management

November 14, 1997 protocol November 14, 1997

December 16, 1997 audit 1 December 16, 1997

December 20, 1997 audit 2 December 20, 1997

January 02, 1998 final report in English January 02, 1998

The data which were taken as a basis for this report were verified by the Quality Assurance Unit.

The accordance of the study with its protocol, the OECD-Guideline for Testing of Chemicals No.

401 and the Principles of Good Laboratory Practice (GLP) is guaranteed.

The methods, procedures and observations are exactly described in the report. The Quality Assurance Unit ensures that the reported results exactly describe the raw data of the study.

The Study Director and the Management are generally informed of any findings of the Quality Assurance Unit.

As for this study no departures from the prescribed course of the test were ascertained.

Karlsruhe, January 07, 1998 Dr. H. Merten

(Quality As-ac Manager)

C~nafSTSCA Conf-def,1 Business nomOf
Page 8 of 22 pages

Report 97 10 42 412 A

Acute Oral Toxicity

Contents

Page

1. Preface 9

2. Summary 10

3. Guideline 11

4. Protocol 11

5. Test substance 11

6. Method "Acute Oral Toxicity" 12

6.1 Animals and laboratory conditions 12

6.2 Composition of diet and water 13

6.3 Acclimatization / Preparations 16

6.4 Study procedures 16

7. Findings "Acute Oral Toxicity" 17

7.1 Mortality/LD 50 17

7.2 Body weight 18

7.3 Toxicological symptoms 20

7.4 Pathological-anatomical findings (macroscopic) 22

8. Archiving 22

Contains ISCA Confidential Business InIorrnatiOfl
Page 9 of 22 pages

Report 7 10 42 412 A

(VAcute Oral Toxicity

1. Preface

Study: Acute Oral Toxicity

Laboratory animals: Albino rats, Sprague-Dawley breed

Guideline: OECD-Guideline for Testing of Chemicals,

Section 4, 401 "Acute Oral Toxicity", adopted

February 24, 1987

GLP: Gesetz zumn Schutz vor gefahrlichen Stoffen (Chemnikaliengesetz ChemG, § 19 a-d vomn

25. Juli 1994) [Chemicals Act of Germany]

Project No.: 971042412

Report No.: 97 1042 412 A

Test substance:

Sponsor:

Monitor.

Test Facility: BioChem GmbH

DaimlerstraBe 5 b

D-76185 Karlsruhe

Germany

Management: Dr. W. Massmann

Dr. M. Miller

Study Director- Dr. H.-J. Kramer

Quality Assurance Manager Dr. H. Merten

Contains TSCA Confidefl MIM Business .
Page 10 of 22 pages

Report 97 10 42 412 A

Acute Oral Toxicity

2. Summary

Report: Acute Oral Toxicity of( in compliance with OECDGuideline 401

Sponsor:

Test Facility: BioChemn GmbH

DaimlerstraBe 5 b

D-76 185 Karlsruhe

Germany

Project No.: 971042412

Protocol No.: 97 10 42 412 A

Report No.: 97 1042 412 A

Findings: The test substance was administered orally in a single dose

of 2000 mg per kg weight to one group of 5 male and 5 female Sprague-Dawley rats. Under the conditions of this experiment ( )caused no mortality. The experiment was

finished as limit test (LD50 (oral) more than 2000 mng per kg

weight).

Acute toxicological symptoms attributed to the exposure to

)could not be observed in all rats on the day of administration and during the observation period. All rats had

an increase in body weight during the 15-day observation

period.

The post-mortemn findings after euthanasia did not show any

macroscopic organ changes.

Contains TSCA Confidential Business Information
Page I11 of 22 pages

Renort7 10 42412 A

S I/ Acute Oral Toxicity0

3. Guideline

The study procedures were performed in compliance with the OECD-Guideline for Testing of Chemicals 401 "Acute Oral Toxicity", adopted February 24, 1987.

4. Protocol

Project No.: 971042412

Protocol No.: 97 10 42 412 A

Date of Protocol: November 14, 1997

Ratification: November 20, 1997

Monitor:(

5. Test Substance

Name:

Characterization of

the test substance: alcohol alkoxylate

Main ingredients:Q

Date of arrival: November 05, 1997

Sample designation: 137291 1 001

Purity: not stated

Physical state: liquid

Colour: colourless

Homogeneity: homogeneous

Stability: not stated

Expiry date: not stated

Storage conditions: not stated, storage class 10 (VC1)

In compliance with the OECD-Guideline 401, a reference substance was not tested.

Contains TSCA Confidential Business Information

qJ3
Page 12 of 22 pages

cenort 97 10 42 412 A

Acute Oral Toxicitya

6. Method "Acute Oral Toxicity"

6.1 Animals and Laboratory Conditions

Species: Albino rat

Breed: Sprague-Dawley

Breeding: Harlan Winkelmann GmbH

Gartenstra~e 27

D-33178 Borchen

Quantity: 1 group of 10 rats (5 males and 5 females)

Identification numbers: 156 - 160 (female)

161 - 165 (male)

Veterinary preliminary examination: without morbid signs; females are nulliparous and non-pregnant

Diet: Haltungsdilt "ALMA 0801 H 1003"

twice 8 g daily

Water~ Free access by daily changing of the watering bottles

Husbandry: The rats housed individually in Makrolon

cages (area 800 cm2, height 17 cm)

Temperature: 19 -23 *C

Relative humidity: 30-70 %

Light-dark-rhythm: 12: 12

Contains ISCA Confidential Business Info rm @111
Page 13 of 22 pages R Mort ?7 10 42 412 A

Acute Oral ToxicityPl

6.2 Composition of Diet and Water

Composition of diet "ALMA 0801 H 1003":

Manufacturer- ALMA-Futter Friedrich Botzenhardt KG

D-87437 Kempten/Allgiu

Manufacturer specification of lot No. 6984

INGREDIENTS:

crude protein 20.00 %

lysine, 1.25 %

methionine, 0.45 %

crude fat 4.00%

crude fibres 5.00 %

crude ash 7.00%

calcium 1.00 %

phosphorus 0.80%

sodium 0.50 %

magnesium 0.25 %

ADDED VITAMINS PER KG DIET:

vitamin A 15.000 I.E.

vitamin D3 500 I.E.

vitamilnE 150 mg

vitamin C 100 mg

Analysis performed on lot No. 6984 by Dr. Nedvidek /BioChem GmbH in March 1994

INGREDIENTS:

moisture method: 105 *C/17 h 12.9 %

ash method. 550 'C/ 6 h 7.1%

crude protein Kjeldahl method according

to BioChemn SOP L 10 24.9 %

crude fat Weibull method in analogy

-to § 35 LMBG L 01.00-20 4.7 %

Contains TSCA Confidential Business Information

96-
Page 14 of 22 pages

Report ?710 42 412 A

j -Y Acute Oral Toxicity,

CONTAMINANTS:

lindane 0.012 mg/kg

a-HCH < 0.01 mg/kg

f3-HCH < 0.01 mg/kg

y-HCH < 0.01 mg/kg

heptachlor < 0.01 mg/kg

aldrin < 0.01 mg/kg

bromophos < 0.01 mg/kg

heptachlorepoxide < 0.01 mg/kg

ac-endosulfan < 0.01 mg/kg

dieldrin < 0.01 mg/kg

p,p'-DDT < 0.01 mg/kg

p,p'-DDE < 0.01 mg/kg

p,p'-DDD <0.01 mg/kg

endrin < 0.01 mg/kg

f3-endosulfan < 0.01 mg/kg

methoxychlor < 0.01 mg/kg

mirex < 0.01 mg/kg

propazine < 0.01 mg/kg

atrazine <0.01 mg/kg

terbuthylazine < 0.01 mg/kg

prometryn < 0.01 mg/kg

ametryn < 0.01 mg/kg

simetryn < 0.01 mg/kg

terbutryn < 0.01 mg/kg

malathion < 0.01 mg/kg

parathion < 0.01 mg/kg

cyanazine < 0.01 mg/kg

Contains TSCA Confidential Business Information

q6
Page 15 of 22 pages

1eor97 10 42 412 A

/ Acute Oral Toxicity

COMPOSITON OF DRINKING WATER:

according to the latest available information of Stadtwerke Karlsruhe (1995):

average of determinations

alum'num < 0.03 mg/I

ammonium < 0.05 mg/I

antimony < 0.01 mg/i

arsenic <0.001 mg/i

barium 0.09 mg/i

boron 0.01 mg/i

cadmium < 0.0001 mg/i

calcium 119 mg/i

chromium < 0.001 mng/I

copper < 0.01 mg/i

iron < 0.01 mg/I

lead < 0.001 mg/i

magnesium 10.6 mg/i

manganese < 0.01 mg/i

mercury < 0.00004 mg/I

nickel < 0.003 mg/I

potassium 1.7 mg/I

selenium < 0.01 mg/I

silicon 5.7 mg/I

silver < 0.0005 mg/i

sodium 11.3 mg/I

zinc < 0.02 mg/i

chlorides 25 mg/i

cyanides < 0.01 mg/i

fluorides 0.08 mg/i

nitrites < 0.01 mg/I

nitrates 6.0 mg/i

phosphates < 0.02 mg/i

sulfates 71 mg/I

dissolved organic carbon 1.4 mg/I

pesticides not verificable

polychiorinated biphenyls not verificable

polycyclic aromatic hydrocarbons < 0.00005 mg/i

trihalogenmethane < 0.0001 mg/l

halogenated hydrocarbons < 0.01 mg/l

organic chlorinated compounds < 0.0005 mg/i

surfactants < 0.01 mg/i

Contains TSCA Confidential Business Information
Page 16 of 22 pages

9Ceor,7 10 42 412 A

/Acute Oral Toxicity

6.3 Acclimatization / Preparations

After randomization the animals were acclimated to the test conditions for 5 days prior to

the administration.

6.4 Study Procedures

Acclimatization: Group (2000 mg/kg):

December I1I - December 15, 1997 Animal No.: 156 - 165

Food was withheld overnight before the day of administration.

Administration: December 16, 1997 (1.30 p.m. - 2.02 p.m.)

Anixnal No.: 156- *165

Calculated amounts of( I according to the dosage of 2 *000

mg/kg were administered with the aid of a stomach tube to the

rats. Food was withheld for fuirther 4 hours after the administration.

Observation: December 16 - December 31, 1997 Animal No.: 156 - 165

Judgement: Mortality, LD50, body weight, behaviour, toxicological symnptoms and at the end of the observation period pathological findings.

Pathology: December 31, 1997 Animal No.: 156 - 165

Contains TSCA Confidential Business Information
Page 17 of 22 pages

Repor 97 10 42 412 A

/Acute Oral Toxicity

7. Findings "Acute Oral Toxicity"

7.1 Mortality / LDSO

Table 1: Mortality

The test group consists of 5 male and 5 female Sprague-Dawley rats

sex male female

dosage (mg/ kg) 2000 2000

mortality 0 0

number of rats 0 out of 5 0out of 5

%0 0

Table 2: LD50 (Sprague-Dawley rat) - Test substance:

[LD50 (mg/kg) 95 % confidence interval I__________

lower limit upper limit

Sprague-Dawley; male > 2000-ISprague-Dawley female > 2000

After the oral administration of 2000 mg( iper kg body weight none of the rats died

during the observation period of 15 days.

The LD50 of "p1 the acute oral toxicity study in rats of both sexes was estimated to

be greater than 2000 mg/kg. Therefore, the study was finished as limit test.

Contains TSCA Confidential Business Information
Page 18 of 22 pages

Report R7 10 42 412 A C 'Acute Oral Toxicity--

7.2 Body Weight

Table 3: Body weight (statements in g)

5 Sprague-Dawley rats, female

Identification numbers: 156 - 160

phase of test day 156 157 158 159 160

acclimatization -5 208 210 209 210 210

-4 212 213 211 210 214

-3 205 206 206 205 209

-2 -

_______ -1 204 200 202 204 -208

administration 0 195 192 199 194 199

1 205 204 210 186 209

_______ 2 212 208 223 197 214

_______ 3 219 214 223 202 215

4 220 216 226 207 221

5 ---

6 233 224 235 218 228

_______ 7 232 223 237 220 230

_______ 9 241 234 246 229 237

_______ 10 - - - -

______ 11 239 230 242 227 237

_______ 12 - - - -

_______ 13 240 234 250 234 247

_______ 14 242 235 250 238 245

pathology 115 1 245 1 240 251 240 1 246

Contains ISCA Confidential Business Inf ormatio
Page 19 of 22 pages

Revort 7 10 42 412 A

-Acute Oral ToxicityTable 4: Body weight (statements in g)

5 Sprague-Dawley rats, male

Identification numbers: 161 - 165

phase of test day 161 162 163 164 165

acclimatization -5 216 218 212 221 211

______-4 224' 215 213 227 224

-3 223 219 228 223 226

_______ ______ -2- --

-l 215 214 221 219 - 215

administration 0- 206 202 214 211 208

1 209 224 223 220 208

2 214 228 235 227 223

3 226 245 243 242 232

_______ 4 232 252 243 250 239

_______ 6 254 262 263 260 250

7 252 265 264 260 259

8- --

9 264 272 269 267 261

10 - - - -

11 278 286 290 276 278

_______ 12 - - - -

_______ 13 289 304 307 304 295

_______ 14 299 307 312 304 294

pathology 1 15 302, 308 1 315 307 302

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Page 20 of 22 pages

Rep~ort,97 10 42 412 A

c Acute Oral Toxicity.-

7.3 Toxicological Symptoms

The observed behavioural pattern and the toxicological symptoms of rats of the different

groups are shown in table 5 - 6, page 20 -2 1.

The criteria of judgement were changes of hair, skin, nose and mouth with mucous membrane, eye, anal region, excrements, urine (macroscopic), respiration, circulation, nervous

system and behaviour.

Table 5: Toxicological symptoms, rat No. 156 - 160, female

symptom day rat No.

(maximum grade)_ ___ _

156 157 158 159 160

0. (5 min) 0 0 0 1 0

0. (12Omin) 0 0 0 0 0

1 . 0 0 0 0 0

2. 0 0 0 0 0

3. 0 0 0 0 0

4. 0 0 0 0 0

5.-----

6. 0 0 0 0 0

apathy (3) 7. 0 0 0 0 0

8.-----

9. 0 0 0 0 0

10. - - - - -

11. 0 0 0 0 0

12. - - - - -

13. 0 0 0 0 0

14 0 0 0 0 0

1 _ _ _ _ __ _ _ _ 115. 0 1 0 1 0 1 0 1 0 _

0 sign not observed 0. further observations during day of administration

I slight symptom 1.- 14. days of observation

2 moderate symptom

3 severe symptom

+ death

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Report P7 10 42 412 A

.-I Acute Oral Toxicity~-

Table 6: Toxicological symptoms, rat No. 161 - 165, male

symptom day rat No.

(maximum grade)____

161 162 163 164 165

0. (5 min) 0 0 0 0 0

0. (12O min) 0 0 0 0 0

1. 0 0 0 0 0

2. 0 0 0 0 0

3. 0 0 0 0 0

4. 0 0 0 0 0

5.- ---

6. 0 0 0 0 0

no symptoms observed 7. 0 0 0 0 0

8. - -

9. 0 0 0 0 0

10. - - - - -

11. 0 0 0 0 0

12. - - - - -

13. 0 0 0 0 0

14 0 0 0 0 0

._________15. 0 0 0 0 0

0 sign not observed 0. fuirther observations during day of administration

I slight symptom 1.- 14. days of observation

2 moderate symptom

3 severe symptom

+ death

Contains TSCA Confidential Business InforM~tion
Page 22 of 22 pages

Report 97 10 42 412 A

I/Acute Oral Toxicity

7.4 Pathological - anatomical Findings (Macroscopic)

No animal died during the observation period. The narcotization and euthanasia of all

animals were performed in an atmosphere of approximately 80 % per cent CO2 at the end

of the observation period.

Table 7: Macroscopical pathological findings, test group

macroscopical individual finding rat No. sex organ chaniges

156 female ___________

157' female -

158 female ___________

159 female ___________

160 female ___________

no pathological findings noted _ _ ____________

161 male ___________

162 male ___________

163 male ___________

164 male ___________

___________________ 165 male __________

8. Archiving

BioChem GmbH. D- 76185 Karlsruhe, Germany, will archive raw data, the protocol, a

copy of the report and the test article archive sample for the stipulated period of time according to GLP regulations.

Contains TSCA Confidential Business Information
auacnmenm 1 -

Oaimlerstrafe 5 b -0-76185 Kartsrhe, i~ e

Tel. 

E-mail: 

07 21/97 

biochem~biochern.de 

37-0 -Fox 07 21/97 37-222I s

Labor fOr biologische und Internet: http./Awwbtochem.de ch &emische =ntlk GmbH

Determination of Ready Biodegradability of

"Closed Bottle Test" According to OECD Guideline 301 D

Study No.: 975040420

Sponsor:

Test substance:(

Lot No.: not stated

BioChem sample designation: 1372915S001

Date of arrival: November 05,1997

BioChern GmbH

Karlsruhe, January 23, 1998 H W/HW

Page I of 26 pages

Report 97 50 40 420

S Ready Biodegradability

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Report ?7 50 40 420

( 1' Ready Biodegradability

Specification of the GLP-Regulations as follows:

We declare that all tests are made in accordance with the regulations of the OECD-principles of

Good Laboratory Practice (GLP).

GLP: "Good Laboratory Practice" Regulation of the European Communities, EC, enacted in

Germany in the "Chernikaliengesetz" (Chemicals Act) 2nd Amendment, dated July 25, 1994, BGBI

1pp. 1703, 1994 and ,,AndVO", dated May 14, 1997, BGBI. 1, pp. 1060, 1997.

The documented data correspond to the measured values.

Study Director

____ ___ ___ aL'" 2

H. Werle ate

Management

"7 2 3L tI'

Dr. Massmann Date

Quality Assurance Manager

Dr. Merten Date

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Page 3 of 26 pages

Reprto Inhibition Test

U~mweltminis terium

Baden-Wuirttemberg

GLP-Besclieinigung

Bescheinigung Certificate

Hierinit wird bestatigt, daB die It is hereby certified that the test

PrOflingeinrichting ficility

in 76185SKarisruhe in 76185SKarlsruhe

der BioChem, Labor fi~r biologische und of BioChern, Labor far biologische umd

chemische Analytik GmbH, chemiache Analytic GmbH,

Daiznlerstr. 5b . Daimlerstr. Sb

am 19J20.06.95, 27.09.95 und 26.0 1.96 on 19120. June 95, 27. September 95

and 26. January 96

von der far die 13berwachung zustAridigen was inspected by the competent

Behorde a±ber die Einhaltung der GJrand- authority regarding compliance with the

s~te der Guten Laborpraxis inspiziert -Principles of Good Laboratory Practice.

worden it

Es wird hierxnit bestatigt;daB folgende It is hereby certified that studies in Thfis test

Priftingen in dieser Prffinrichtung; nach facility are conducted in compliance with the

den Grundsa±zen der Guten Laborprawds Principles of Good Laboratory Practice.

durcbgef~hrt werden.

den BADEN,4

Stut0gaM den r"ERG 0

Seite 1/2 page 1/2 ;

nins TSCA Confidential Business Information
* Page 4 of 26 pages

R eport 97 50 40 420

/Ready Biodegradability--

*Seite 2/2 zur GLP-Bescheimigung

Priifungskategorie 1 Priifungen auf physikaischchemische Eigenschzften und

Gehltsbestimmungen

Prifungskategorie 2 Prilfungen auf toikologische

Eigenschaften

Prilfungskategorie 3 Prfifungen auf mutagene

Eigenschaften (in vitro, in vivo)

Priifungskategorie 4 Umwelttozikologische Prifiunen

zu Auswirknngen anf aquatisdie

und terrestrische Orgaismen

Priifungskategorie 5 Priifungen zum Verhalten in

Boden, im Wasser und in der

Luft;.Bioakkumulation;.

Metabolismus

Priifungskategorie 6 Prifungen auf Riickstinde

Dies entspricht dea Kategoxien nach der Ailgemeinen Verwaltungsvorschriftm

* Verfibren der behdrdlichen O0berwachung der Binhaltung der Guxten Laborpra-ds

* (ChemVwV-GLP) vom 29. Oktober 1990:

"Physikalisch-chemische Eigenschaften and

Gehaltsbestimmungen"

"Toxikologische Eigenschaften"

"Okotoxikologische Eigenschaften"

"Verhalten im Boden, iM Wasser und in der Luft"

"Rfickstinde"

Stuttgart, den 26.02.1996

et4.4< WTJRTTEMBERG ContainsBu~es

Dr. Albrecht '$'SCCofdnalBsesIno

Seite 2/2 page 212
Page 5 of 26 pages

R epot 97 50 40 420

SIRespiration Inhibition Test

Translation of page 2 of the GLP-Certificate by BioChem,

Test category 1:TeTs on physico-chernical

properties and content

determinations

Test category 2: Tests on toxicological

properties

Test category 3: Tests on inutagenic properties

(in vitro, in vivo)

Test category 4: Ecotoxicological tests on*

effects on aquatic and

terrestrial organisms

Test category 5: Tests on behaviour in soiL in water

and in the air-,

bioaccumulation; metabolism

Test category 6: Tests on residues

This corresponds to the categories according to the General Administration

Regulation of the Procedure of the Official Supervision of the Compliance with Good

Laboratory Practice (Chem VwV-GLP) dated October 29, 1990:

"Physico-chemical Properties and Content Determinations"

"Toxicological Properties"

"Ecotoxicological1 Properties"

"Behaviour in Soil, in Water and in the Air"

"Residues"

Stuttgart, February 26, 1996 Contains TSCA Confidential Business Inforui .I

Bade n-Wii rtte mbe rg

Department of the Environment
Page 6 of 26 pages

Report 97 50 40420 ( ~ /Ready Biodegradability_

Quality Assurance Unit Statement

Final Report

Title: Determination of Ready Biodegradability of[ I.-Closed

Bottle Test" according to OECD Guideline ko. 301'D

study number. 97 50 40 420

Guideline: OECD Guideline -for Testing of Chemicals No. 301 D,

"Ready Biodegradability: Closed Bottle Test", Adopted July

17, 1992.

Study /test system: ready biodegradability of( oxygen meter

Test substance:

Lot No.: not stated

BioChem sample designation: 137291 5 001

Date of arrival: November 05, 1997

Study Director: H. Werle

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Report 97 50 40 420

Y Respiration Inhibition Test

The following checks were effected:

reports to the Study Director

date verified procedure and to the Management

November 27, 1997 protocol November 27, 1997

December 09, 1997 audit 1 December 09, 1997

December 23, 1997 audit 2 December 23, 1997

January 21, 1998 finalireport January 22,1998

The data which were taken as a basis for this report were verified by the Quality Assurance Unit.

The accordance of the study with the protocol, the OECD Guideline for Testing of Chemicals No.

301 D, "Ready Biodegradability: Closed Bottle Test", Adopted July 17, 1992 and the Principles of

Good Laboratory Practice (GLP) is guaranteed.

The methods, procedures and observations are exactly described in the report. The Quality

Assurance Unit ensures that the reported results fully comply with the raw data of the study.

The Study Director and the Management are generally informed of any findings of the Quality

Assurance Unit.

As for this study no deviations from the prescribed course of the test were ascertained.

Karlsruhe, January 23, 1998 Dr. Merten

(Quality Assurance Manager)

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Page 8 of 26 pages

Report97 50 40 420

( / Ready Biodegradability

CONTENTS

I Study Identification

2 Test / Guideline

3 Summary

4 Test Material

5 Test Procedure

5.1 Principle

5.2 Equipment and Reagents

5.2.1 Equipment

5.2.2 Reagents

5.3 Performance of the Test

5.3.1 Test Controls

5.3.2 Test Solutions

5.4 TestProcedure

5.5 Test Criteria

6 Results

6.1 Toxicity Evaluation - Inhibition Test

6.2 ( Determination of ThOD/COD

6.3 Reference Materials, Determination of ThOD

6.4 Percentage of Biological Degradation (Calculations)

6.5 Conclusion

6.5.1 Biochemical Degradation of Reference Substances

6.5.2 Toxicity Test

6.5.3 Biochemical Degradation of

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Page 9 of 26 pages

Report 97 50 40 420 (1 VRespiration Inhibition Test

STUDY IDENTIFCATION

Test: Determination of Ready Biodegradability

Guideline: The study procedures described in this report

are based on the following guideline:

OECD Guideline for Testing of Chemicals

No. 301 D, "Ready Biodegradability: Closed

Bottle Test", Adopted July 17, 1992.

Sponsor:-

Monitor:()

Test facility: BioChem, GmbH

Daimlerstr. 5 b

D-76 185 Karlsruhe

Germany

Management: Dr. Massmann,

Dr. Mailer

Quality Assurance Manager: Dr. Merten

Study Director: Mr. H. Werle

Project staff: Mr. Hufliagel

Study number: 97 5040420

Date of protocol: November 28, 1997

Test substance:

Homogeneity: homogeneous

Expiration date/stability: not stated

Packing: 30 ml in brown glass container

Contains TSCA Confidential Business Information

fl/3
Page 10 of 26 pages

R eport 97 50 40 420

/Roady Biodegradability Is, Storage conditions: dark storage at room temperature

Safety precautions: wear gloves and protective goggles, avoid

inhalation and skin contact

Experimental start date: November 26, 1997 (determination of COD)

Experimental termination date: December 30, 1997

Literature: OECD Guideline for Testing of Chemicals

No. 301 D, "Ready Biodegradability: Closed

Bottle Test", Adopted July 17, 1992,

and

German standard methods for the examination

of water, waste water and sludge: "Deutsche

Einheitsverfahren zur Wasser-, Abwasser- und

Schlanimuntersuchung", VCH Verlagsgesellschaft; Norm H 43-1. Bestimmung des

Cheniischen Sauerstoflbedarfs (CSB)-Kurzzeitverfabren,

and

EC Directive 92/69/EEC of 31 July 1992

adapting to technical progress for the

seventeenth time Council Directive

67/548/EEC on approximation of laws,

regulations and administrative provisions

relating to the classification, packaging and

labelling of dangerous substances, Annex Part

C, Methods for Determination of Ecotoxicity;

Method C.4-E, "Closed Bottle Test", Official

Journal of the European Communities No. L

383 A/1 79, dated December 29, 1992.

Archiving: BioChemn GmbH will archive the following

data for 15 years:

protocol, raw data, report, audits (QAU).

Test substance reference samples will be

archived for 12 years.

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Page I1I of 26 pages

Report 97 50 40 420

( /Respiration Inhibition Test

2 TEST / GUIDELIN

Determination of Ready Biodegradability

OECD Guideline for Testing of Chemicals No. 301 D, "Ready Biodegradability: Closed

Bottle Test", Adopted July 17, 1992.

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Page 12 of 26 pages

Report 97 50 40420

Ready Biodegradability -

3 SUMMARY

Determination of Ready Biodegradability of( according to

OECD Guideline No. 301 D

Study objectives

The objective of this study is to determine the Ready Biode raability of(1

The determination of the Ready Biodegradability of ( )~as performed in

accordance with the OECD Guideline for Testing of Chemicals Po. 301 D, "Ready

Biodegradability: Closed Bottle Test", Adopted July 17, 1992.

Conclusion

Validity of the test

Both reference samples, potassium hydrogen phthalate and sodium acetate, reached a

degradation level of more than 60 % within 10 days alter the substance degradation had

reached a 10 % threshold degradation level. So the performed test on biodegradability of [ ]according to OECD-Guideline No. 301 D is valid.

Measured temperatures and 02 consumption of blank solutions are within the range

demanded by the guideline.

Toxicity Test

Since the presence of )h}ad no influence on the sodium acetate degradation by the

inoculum, lacking test substance degradation is not due to any toxicity of[ to

inoculumn bacteria.

Biochemical Degradation ofL

The observed degradation ofC was below 10 % in the course of the test.

On account of this observation the test sample( can not be classified as ready

biodegradable according to the "Closed Bottle Test" (OECI) Guideline No. 301 D).

Contains TSCA Confidential Business Informaticv
Page 13 of 26 pages

Report 97 50 40 420 L )/Respiration Inhibition Test

4 TEST MATERIAL

Name: N

Active ingredient: alcohol alkoxylate

Ingredients:()

Colour: colourless, clear

Homogeneity: homogeneous

Purity (declaration): not stated

Expiry date/stability: not stated

BioChem sample designation: 137291 5 001

Date of arrival at BioChem: November 05, 1997

5 TEST PROCEDURE

5.1 PRINCIPLE

Aqueous solutions of the test substance in different concentrations, including inorganic

nutrient salts, are inoculated with the effluent from a municipal sewage plant. The

solutions are incubated in the dark at 22 0C ± 2 *C for 28 days in closed bottles. The

oxygen content is measured on several days and the biochemical oxygen demand (BOD) is

calculated. The oxygen content of two readily degradable reference substances without

test substance is determined to check the validity of the test.

Also a control with inoculuni, but without any test material as well as a control with pure

mineral nutrient solution are run in parallel for the determination of oxygen blanks.

Furthermore, a toxicity control is included in the test design by analyzing test substance,

sodium acetate (reference chemical) and inoculumn jointly.

A microbiological inhibition test is furthermore performed to exclude inhibitory effects in

the closed bottle test.

The theoretical oxygen demand (ThOD) of the test substance is calculated via the

molecular formula. Moreover, especially with substances of unknown composition or

limited purity the chemical oxygen demand (COD) is analytically determined. The COD is

the maximum possible oxygen consumption during degradation.

Contains ISCA Confidential Business Informatiem
Page 14 of 26 pages

Report 97 50 40 420 C / Ready Biodegradability

The biological degradability of the test material is stated as a percentage of the BOD/COD

or BOD/ThOD ratio.

Test materials giving a measured BOD greater than 60 % of the ThOD or COD within

28 days should be regarded as readily biodegradable. This level must be reached within 10

days of biodegradation exceeding a 10 % biodegradation threshold.

5.2 EQUIPMENT AND REAGENTS

5.2.1 Equipment

Standard laboratory glassware and tools

glass bottles: 250 ml volume, glass stoppered

Pipettes: Eppendorf

Analytical balances: Mettler AT 261 Delta Range

Sartorius Analytic A200S

pH-meter: knick calimatic

02-meter: oximeter 196 inclusive thermoelement, WTW

Determination of COD: Radiometer Titralab VIT 90 Video Titrator and

ABU93 Triburette

5.2.2 Reagents

Water Millipore Milfi-Q

Trypton Soya Broth (Caso bouillon): caseine peptone-soja flour peptone bouillon, Merck,

order No. 5459

Contains TSCA Confidential Business Informal2-I
Page 15 of 26 pages

Report 97 50 40 420

( / Respiration Inhibition Test 

Nl

Mineral nutrient

(All of the following reagents were p.a. grade)

Stock solutions for mineral nutrient:

(a): 8.50 g KH2PO4, potassium dihydrogen phosphate

23.61 g K2BPO4 x 3 H,O, dipotassium hydrogen

phosphate trihydrate

33.40 g Na2HPO4 x 2 H,0, disodium hydrogen

phosphate dihydrate

0.50 g NH4CI, ammonium chloride

dissolved in 11 H2 0

(b): 36.40 g CaC12 x 2 H20, calcium chloride dihydrate,

dissolved in I1I H2 0

(c): 22.50 g MgSO4 x 7 H20, magnesium sulfate heptahydrate, dissolved in 11I H20

(d): 0.25 g FeC13 x 6 H2 0 iron (M1) chloride hexahydrate,

dissolved in 11I H2 0

Preparation of mineral nutrient: 32 ml of each of the solutions (a) - (d) were added to

32 1 purified water. The mineral medium was aerated

for 45 minutes. The pH was determined to about 7. 1.

Reference material: Sodium acetate, anhydrous, Fluka, lot No. 298125

1090, order No. 71180

Potassium hydrogen phthalate, Fluka, p.a., lot No.

2 87995/1 992, order No. 60359)

Inoculum: Sewage plant effluent from municipal sewage plant KarlsruhelNeureut

Date of sampling: December 0 1, 1997

The sewage was filtered through glass wool and was

stored aerobe overnight at +4 *C until start of the

closed bottle test on December 02, 1997.

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Page 16 of 26 pages

Report 97 50 40 420

)/'j Ready Biodegradability&

5.3 PERFORMANCE OF THE TEST

5.3.1 Test Controls

Inhibition test:

A test for inhibitory effects on the development of the bacteria was performed by cultivation

of the activated sludge in a culture medium (Trypton Soya Broth) in presence of the test

substance in its highest concentration made use of in the Closed Bottle Test (24 h; 20 - 25

0

C).

The development of the bacteria was compared visually with a control without test material.

5.3.2 Test Solutions

If the test substance has an unknown stoichiometric composition like ( ).the COD of

this solution was determined (see section 6. 1).

N Test solutions of

As the test substance is not a nitrogen-containing compound, no correction for nitrification

was made

An aliquot of a f~ solution was mixed with water containing mineral salts and

bacteria from a municipal sewage plant (inoculum). The aliquot was fixed to ensure a

remaining concentration of at least 2 mg 02 /1 H2 0 in the test solution after complete

biotic degradation at the end of the test.

For that, 34.5 mg ( )was weighed into a 250 mal volumetric flask. The sample was

dissolved with purified water and the flask was made up to volume with this solvent.

Content: 138 mgfl (= stock solution)

Four different test concentrations were prepared by dilution of aliquots of this stock

solution with mineral medium in separate volumetric; flasks:

test conctr t on stock soln. final volume

[M.\ /I m][l

0.69 15 3000

1.38 30 3000

2.76 60 3000

14.14 90 3000

Each flask was fortified with three drops of inoculum and the solutions were shaken.

Thereafter the test procedure (see section 5.4) was performed immediately.

Contains ISCA Confidential Business Information
Page 17 of 26 pages

Report 97 50 40 420

L Respiration Inhibition Test I

0 Test solutions of reference material

Sodium acetate:

252.2 mg sodium acetate was weighed into a I I volumetric flask which was filled up to

volume with purified water.

60 ml of this stock solution were pipetted into a 3 1 volumetric flask. The sample was

diluted with mineral nutrient and the flask was made up to volume with this solvent.

concentration: 5.044 mg/i.

Potassium hydrogen phthalate

170.0 mg potassium hydrogen phthalate was weighed into a I1I volumetric flask which was

filled up to volume with purified water.

60 ml of this stock solution were pipetted into a 3 1 volumetric flask. The sample was

diluted with mineral nutrient and the flask was made up to volume with this solvent.

concentration: 3.400 mg/i.

Each of both flasks was fortified with three drops of inoculum and the solutions were

shaken. Thereafter the test procedure (see section 5.4) was performed immediately.

5.4 TEST PROCEDURE

Test Sample'( Reference Material

The test solutions of , 4nd the reference solutions (see section 5.3.2) were each

filled bubble-free into ten 250 ml glass bottles up to their maximum capacity. The bottles

were closed airtight with glass stoppers. Subsequently the bottles were stored for 28 days

in darkness at a temperature of 22 ± 2 *C. On day 0, 7, 14, 21 and 28 two bottles per

sample were opened and the oxygen content was measured using an 02 electrode. The

contents of each bottle were poured off after oxygen measurement with exception of the

bottles of day 0. These bottles were refilled with their corresponding test solution and they

were stored in darkness as the others as reserve. The BOD of LA 409 was determined via

the decrease of oxygen in the bottles.

Additional Samples (Test Control)

Degradation test:

The biological degradability of sodium acetate and potassium hydrogen phthalate in the

closed bottle test was determined (see above). The degradability had to reach at least a

level of 60 per cent, i.e. sodium acetate and potassium hydrogen phthalate had to exhibit a

BOD which was greater than 60 per cent of their ThOD within 14 days.

Contains TSCA Confidential Business Information
Page 18 of 26 pages

Report 97 50 40 420 _

( '/Ready Biodegradability

Toxicity test:

To remove any trace of doubt about the test substance toxicity to bacteria,( )nits

highest tested concentration was fortified with sodium acetate stock solution:

90 ml of (- )stock solution and 60 ml sodium acetate stock solution were pipetted

into a 3 1 volumetric flask. The sample was diluted with mineral nutrient and the flask

was made up to volume with this solvent.

concentration: L4.14 mg/I,

sodium acetate: 5.044 mgfl

The flask was fortified with three drops of inoculum and the solution was shaken.

Thereafter the test procedure (see above) was performed immediately.

Any indication o( 1toxicity would have resulted in a decreased sodium acetate

BOD level.

Blank control:

For test control two additional samples were prepared: Pure mineral nutrient and mineral

nutrient fortified with 3 drops inoculumn per 3 1 (test performance see above).

5.5 TEST CRITERIA

Test substances reaching a pass level of 60 % removal of ThOD within 28 days are so

called "ready biodegradable".

Pass-level-criterion:

60% degradation must be reached within at least 10 days after exceeding the 10%

degradation level.

* Validity of study:

1. Oxygen depletion in the inoculum blank must not exceed 1.5 mg dissolved 02/1 after 28

days.

2. The residual concentration of oxygen in the test bottles must not fall below 0.5 mg 02/I

at any time.

3. Difference of extremes of replicate values of the removal of the test substance at the

plateau or at the end of test :5 20 %

4. Percentage degradation of reference compound reaching pass level by day 14.

Contains TSCA Confidential Business Information
Page 19 of 26 pages

Report 97 50 40 420

I/1 Respiration Inhibition Test

6 RESULTS

6.1 Toxicity Evaluation - Inhibition Test

Test concentration: 0. 84 mg( )and 200 ml caso-bouillon; added volume of

activated sludge: 50 jgd per flask (double determination)

sample evaluation of bacteria development

after 24 h incubation time at 20 - 25*C

control

test sample, 4.2 mg

-no growth, + = low, -H- = middle, +- high growth

There are no inhibitory effects on the development of the bacteria by the test material in its

highest concentration or test blank used in the Closed Bottle Test.

Moreover, a respiration inhibition test carried out with activated sludge accor~ing to OECD

Guideline No. 209 revealed no inhibitory effect to bacteria attributed to ( ',This

outcome was recorded in detail in BioChemn Report 97 50 40 806, dated January 20, 1998.

6.2 ( ,Determination of ThOD/COD

Theoretical Oxygen Demand (ThOD)

Due to the unknown exact stoichiometric composition of( )the theoretical oxygen

demand (ThOD) was not used as a basis for degradation determination. Therefore, the

chemical oxygen demand was determined experimentally.

Chemical Oxygen Demand (COD)

German'Standard Methods of Water Analysis ("Deutsche Einheitsverfahren zur

Wasseruntersuchung"), Method H 43-1 was used for COD determination.

52.9 mg ) was weighed into a 250 ml volumetric flask. The sample was dissolved

with purified water and the flask was made up to volume with this solvent.

Concentration: 211.6 mg/l.

Used device: Radiometer Titralab VIT 90 Video Titrator and ABU93 Triburette

Contains TSCA Confidential Business Information A
Page 20 of 26 pages

Report 97 50 40 420 ( /Ready Biodegradability

A fourfold determination of COD of )was performed after equilibrium of l

titrimetric system:

sample No. COD [Mg 02/11 COD [mg 02 / Mg1

1 469.1

2 476.3

3 471.5

4 472.0

mean 472.2 2.23

calculation: COD [Mg 02/1] )conc. (mg/I] = COD [mg O2Img]

The chemical oxygen demand of( )was determined to 2.23 g 02/g test substance.

6.3 Reference Materials, Determination of ThOD

Sodium acetate, Determination of theoretical oxygen demand (ThiOD)

Calculated according to OECD Guideline for Testing of Chemicals 301 D,

Annex IV

Substance: sodium acetate, anhydrous

Formula: C2H3NaO2

element mol/formula molecular mass

g/mol g/mol substance

carbon, C 2 12.011 24.022

hydrogen. H 3 1.0079 3.024

sodium, Na 1 22.9898 22.990

oxygen,O0 2 15.9994 31.999

molecular weight (g/mol) 82.04

ThOD (mg O,/g substance) 780

The theoretical oxygen demand (ThOD) of sodium acetate was calculated to 780 Mg 0 2/g

or 0.78 g 0 2/g sodium acetate.

Contains TSCA Confidential Business Information
Page 21 of 26 pages

Report 97 50 40 420

)l~ Respiration Inhibition Test

Potassium hydrogen phthalate, Determination of theoretical oxygen dema~s

Calculated according to OECD Guideline for Testing of Chemicals 301 D,

Annex IV

Substance: potassium hydrogen phthalate, anhydrous

Formula: CsH5KO4

element mol/formula molecular mass

g/mol g/mol substance

carbon, C 8 12.011 96.088

hydrogen, H 5 1.0079 5.0395

potassiumn,K 1 39.0983 39.0983 -

oxygen, 0 4 15.9994 63.9976

molecular weight (g/mol) 204.22

ThOD (mg 0,/g substance) 1175

The theoretical oxygen demand (ThOD) of potassium hydrogen phthalate was calculated

to 1175 mg 02/g or 1. 175 g 02/g potassium hydrogen phthalate.

6.4 Percentage of Biological Degradation (Calculations)

* 02 decrease (mg oxygen/I) after x days:

oxygen consumption [mg 02/I] =(m,~, - nj- (mbo - mb.)

M0= 02 value in mg/l at day 0 (test substance)

Ml,= 02 value in mg/I at day x (test substance)

MbO =02 value in mg/I at day 0 (mean of blanks with inoculum)

=b 02 value in mg/i at day x (mean of blanks with inoculum)

M Biological oxygen demand (ROD) in mg oxygen / g substance

BOD [mgO2/g substance] = (oxygen consumption / c(test substance)) .1000

- oxygen consumption in mg oxygen / I

- c(test substance) = concentration of test substance in mg/I

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Page 22 of 26 pages

Report 97 50 40 420

( '/Ready Biodegradability

N Calculation of the percentage of biological degradation

BOD in mg/g - 100

% biological degradation= ________

COD* in mg/g

*or ThOD, respectively

Contains TSCA Confid,.ninI Business Information
Page 23 of 26 pages

Reprto Inhibition TestIs

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Report 97 50 40 420 ( Ready Biodegradability

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Report 97 50 40 420

'~iRespiration inhibition Test

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Page 26 of 26 pages

Report 97 5040 420

)/ Ready Biodegradability

6.5 Conclusion

6.5.1 Biochemical Degradation of the Reference Substances

Sodium acetate

Within 7 days after start of the test the biodegradability of sodium acetate had reached a

plateau of about 70 % degradation. The degradation already exceeded the required

minimum of 60 % at this time. A final degradation of about 81 % was determined after

28 days.

Potassium hydrogen phthalate

Within 7 days after start of the test the biodegradability of potassium hydrogen phthalate

had reached a plateau of about 70 % degradation. The degradation already exceeded the

required minimum of 60 % at this time. A final degradation of about 81 % was

determined after 28 days.

Summary:

Both reference samples, potassium hydrogen phthalate and sodium acetate, reached a

degradation level of more than 60 % within 10 days after the substance degradation had

reached a 10 % threshold degradation level. So the performed test on biodegradability of

f-J according to OECD-Guideline No. 301 D is valid.

'"Measuied temperatures and 02 consumption of blank solutions are within the range

demanded by the guideline.

6.5.2 Toxicity Test

Since the presence of( > had no influence on the sodium acetate degradation by the

inoculum, lacking test substance degradation is not due to any toxicity of( -0

inoculumn bacteria.

6.5.3 Biochemical Degradation of(

The observed degradation of(_ )was below 10 % in the course of the test.

Since this observation the test sample (- - )can not be classified as ready

biodegradable according to the "Closed Bottle Test" (OECD Guideline No. 301 D).

Contains TSCA Confidential Business Information
attachment 12

Daimlerstralfe 5 b -D-76185 Kadruhe i he Tel. 07.21/97 37-0O- Fax 07 21/97 37-222Bi he E-mail: biochrO biochem.de Labor fOr biologische und Internet: http-/wvwbiohem.de chemish MAioyik GmbH

Activated Sludge, Respiration Inhibition Test

According to OECD Guideline No. 209

of(

Study No.: 975040806

Sponsor:

Test substance: )

Lot No.: not stated

BioChem sample designation: 137291 7001

Date of arrival: November 05,1997

BioChemn GmbH

Karlsruhe, January 20, 1998 HWIHW

Page I of 23 pages

Report 97 50 40 806 C ~ IRespiration Inhbition Test

Contains TSCA Confidential Business Information
Page 2 of 23 pages

Report 97 50 40 806

( Respiration Inhibition TestIs

Specification of the GLP-Regulations as follows:

We declare that all tests are made in accordance with the regulations of the OECD-principles of

Good Laboratory Practice (GLP).

GLP: "Good Laboratory Practice" Regulation of the European Communities, EC, enacted in

Germany in the "Chemikaiiengesetz" (Chemicals Act) 2nd Amendment, dated July 25, 1994,

BGBI1I, pp. 1703, 1994 and ,,ndVO", dated May 14, 1997, BOBI. 1, pp. 1060, 1997.

The documented data correspond to the measured values.

Study Director

______________ uaw f o /S0

H. Werle Date

Management

Dr. Mfiller Dae 1

Quality Assurance Manager

Dr. Merten Date ,zc7

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Page 3 of 23 pages

Report 97 50 40 806

S Respiration Inhibition Test I

Umweltm=nisterium

Baden-Wfirttemberg

GLP-Beschehuigunig

Bescheinigung Certificate

lflerinit wird bestitigt, daB die It is hereby certified that the test

Profungseinrichtung: flility

in 7618SKrisruhe in 76185Karsruhe

der BioChem, Labor ffir biologische und of BioChem, Labor fdr biologische und

cheisheAnalytik GmbH, - chemiscite Analytik G~mbH,

Dainlerstr. 5b . Daimerstr. 5b

am 19J20.06.95, 27.09.95 und 26.0 1.96 on 19J20. Tune 95, 27. September 95

and 26. January 96

von der fir die Oberwachung nist~ndigen was inspected by the competent

Beharde Qber die Einhaltuing der Giund- authority regarding compliance with the

satze der Guten Laborprawds inspiziert Principles of Good Laboratory Practice.

worden ist

-Es wird hiermit bestAtigt, daB3 folgende It is hereby certified that studies in this test

Pra1fungen in dieser Prafeinuichtung nach facility are conducted in compliance with the

den Grunds~tzen der Guten Laborpraxis Principles of Good Laboratory Practice.

durchgefihfrt werden.

Contains ISCA Confidential B snttss Inlormation Sttgtdn 6 RfER

Seite 1/2pae12j

page11/2
*Page 4-of23 Pages

Report 97 50 40 806 ( )/Respiration Inhibition Test

*Seite 2/2 mur GLP-Bescheinigung r

Priifungskategorie 1 Pxiifungen auf physikaichchemische Eigenschateu urid

Gehaltsbestimmungen

Priifungskategorie 2 Prilfungen auf toikologische

Eigenschaften

Prifungskategorie 3 Prg~fungen auf mutagene

Eigenschnften (in vitro, in vivo)

Priifungskateg-orie 4 Umwelttoxikologische Prilfungen

zu Auswikngen auf aquatische

und terresfrische Organismen

Prtifungskategorie 5 Priifungen zam Verhalten im

Boden, im Wasser und in der

Luft;'Bioakkumulation;.

Metabolismus

Priifungskategorie 6 Priifungen auf Riickstiinde

Dies entspricht den Kategorien nach der Ailgemeinen Ve waltumgsvorschrift 71M

Ver~bren der behard~ichen Oberwachtng der Einhl~tung der Guten Labo rads

* (ChemVwV-GLP) voam 29. Oktober 1990:

IlPhysik- 1 ich-chmische Eigenschaften und

Gehaltsbestimmungen"

"Toxikologische Eigenschaften"

'

t

6kotoklogische Eigenschaften"

"Verhalten im Boden, im Wasser und in der Luft"

"Rickstlinde"

Stuttgart, den 26.02.1996

Dr. Albrecht Cnan SACniefB u~ns nomf

Seite 2/2 page 212
Page 5 of 23 pages

Report 97 5040 806

/Respiration Inhibition Test

Translation of page 2 of the GLP-Certificate by BioChem.

Test category 1: Tests on physico-chemicat

properties and content

determinations

Test category 2: Tests on toicological

properties

Test category 3: Tests on mutagenic properties

(in vitro, in vivo)

Test category 4: Ecotoxidcological tests on*

effects on aquatic and

terrestrial organisms

Test category 5: Tests on behaviour in soil, in water

and in the air;

bioaccumulation; metabolism

Test category 6: Tests on residues

This corresponds to the categories according to the General Administration

Regulation of the Procedure of the Official Supervision of the Compliance with Good

Laboratory Practice (Chem VwV-GLP) dated October 29, 1990:

"Physico-chernical Properties and Content Determinations"

1?oxicological Properties"

"Ecotoxicological Properties"

"Behaviour in Soil, in Water and in the Air"

"Residues"

Stuttgart, February 26, 1996 Contains TSCA Confidential Business InformallOl

Baden-Wilrtemberg

Department of the Environment134
Page 6 of 23 pages

Report 97 50 40 806 ( /Respiration Inhibition Test

Quality Assurance Unit Statement

Final Report

Title: Respiration Inhibition Test of( )according to OECD

Guideline No. 209

study number: 97 50 40 806

Guideline: OECD-Guideline for Testing of Chemicals No. 209,

"Activated Sludge, Respiration Inhibition Test", adopted

April 04, 1984.

Study / test system: respiration rate of activated sludge in presence of LA 409/

oxygen meter

Test substance:

Lot No.: not stated

BioChemn sample designation: 137291 7 001

Date of arrival: November 05, 1997

Study Director. H. Werle

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Page 7 of 23 pages

Report 97 5040 806 ( Respiration Inhibition Test

The following checks were effected:

dateveriied roceurereports to the Study Director

dateveriied roceureand to the Management

December 16, 1997 protocol December 16, 1997

December 23, 1997 audit December 23, 1997

January 19, 1998 final report January 19,1998

The data which were taken as a basis for this report were verified by the Quality Assurance Unit.

The accordance of the study with the protocol, the OECD-Guideline for Testing of Chemicals No.

209, "Activated Sludge, Respiration Inhibition Test", adopted April 04, 1984 and the Principles of

Good Laboratory Practice (GLP) is guaranteed.

The methods, procedures and observations are exactly described in the report. The Quality

Assurance Unit ensures that the reported results fully comply with the raw data of the study.

The Study Director and the Management are generally informed of any findings of the Quality

Assurance Unit.

As for this study no deviations from the prescribed course of the test were ascertained.

Karlsruhe, January 20, 1998 Dr. Merten

(Quality Assurance Manager)

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Report 97 50 40 806 ( )/ Respiration Inhibition Test I

CONTENTS

I Study Identification

2 Test / Guideline

3 Summary

4 Test Material

5 Test Procedure

5.1 Principle

5.2 Equipment and Reagents

5.2.1 Equipment

5.2.2 Reagents

5.3 Performance of the Test

5.3.1 Reference Material

5.3.2 Solutions of the Test Substance

5.3.3 Solutions of the Reference Substance

5.3.4 Test Concentrations

5.3.5 Synthetic Sewage Feed

5.3.6 Activated Sludge

5.3.7 Test Conditions

5.3.8 Test Procedure

5.3.9 Test Validity

6 Results

6.1 Activated Sludge - Determination of Dry Matter

6.2 Observations / Treatment of Results

6.3 Table of Results

6.4 Graphic Evaluation of EC50 of the Reference Substance

6.5 Exemplary Recorder Plots

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Report 97 50 40 806

Il/ Respiration Inhibition Test 

r

1 STUDY IDENTIFICATION

Test: Activated Sludge, Respiration Inhibition Test

Guideline: The study procedures described in this report

are based on the following guideline:

OECD-Guideline for Testing of Chemicals

No. 209, "Activated Sludge, Respiration

Inhibition Test", adopted April 04, 1984.

Sponsor:

Monitor: r )

Test facility: BioChem GmbH

Dainilerstr. 5 b

D-76185 Karlsruhe

Germany

Management: Dr. Massmann,

Dr. MUler

Quality Assurance Manager: Dr. Merten

Study Director: Mr. H. Werle

Project staff- Mr. Hufniagel

Study number: 97 50-40806

Date of protocol: December 17, 1997

Test substance:(

Homogeneity: homogeneous

Expiration date/stability: not stated

Packing: 30 ml in brown glass container

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Page 10 of 23 pages

Report 97 50 40 806

( )IRespiration Inhibition Test

Storage conditions: dark storage at room temperature I

Safety precautions: wear gloves and protective goggles, avoid

inhalation and skin contact

Experimental start date: December 15, 1997 (weight of synthetic

sewage feed ingredients)

Experimental termination date: December 23, 1997 (performance of

respiration inhibition test)

Literature: OECD-Guideline for Testing of Chemicals No.

209, 'Activated Sludge, Respiration Inhibition

Test", adopted April 04, 1984,

EC Directive 88/302/EEC of November 18,

1987 adapting to technical progress for the

ninth time Council Directive 67/548/EEC on

the approximation of laws, regulations and

administrative provisions relating to the

classification, packaging and labelling of

dangerous substances, Annex Part C, Methods

for Determination of Ecotoxicity; Official

Journal of the European Communities No. L

133 All, dated May 30, 1988 and corrected in

L 136 A/20.

Archiving: BioChemn GmbH will archive the following

data for 15 years:

protocol, raw data, report, audits (QAU).

Test substance reference samples will be

archived for 12 years.

2 TEST / GUIDELINE

Activated Sludge, Respiration Inhibition Test.

OECD-Guideline for Testing of Chemicals No. 209, "Activated Sludge, Respiration

Inhibition Test", adopted April 04, 1984.

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Page I1I of 23 pages

Report 97 50 40 806 ( /Respiration Inhibition Test

3 SUMMARYIs

Activated Sludge, Respiration Inhibition Test of )according to

OECD-Guideline No. 209

Study objectives

The objective of this study is to determine the potential respiration inhibition oft

The determination of the respiration inhibition of( )vas performed in accordance

with the OECD-Guideline for Testing of Chemicals No. 209, "Activated Sludge,

Respiration Inhibition Test", adopted April 04, 1984.

Conclusion

Results of the Test Control:

The test controls document the correct progress of the respiration inhibition test.

The respiration rates of the two controls are within 15 per cent of each other.

The EC50 (3 h) value of the reference substance is in the accepted range of 5 to 30 mg/I

(19.4 mg/l 3,5-dichlorophenol determined from the course of a linear regression curve).

Results of the Test:

The study on respiration inhibition of five concentrations of the test substance shows no

inhibitory effect. The tested concentrations had a respiration rate higher than the control

samples. Following this findings, no EC50 value can be deduced from this result.

In summary, it was found that )lot No. not stated, is not a

respiration inhibitory agent according to the OECD Guideline No. 209.

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Report 97 50 40 806

~)/ Respiration Inhibition Test

4 TEST MATERIALIs

Name:(

Active ingredient: alcohol alkoxylate

Ingredients:()

Colour: colourless, clear

Homogeneity: homogeneous

Purity (declaration): not stated

Expiry date/stability: not stated

BioChemn sample designation: 137291 7001

Date of arrival at BioChem: November 05, 1997

5 TEST PROCEDURE

5.1 PRI"cIPLE

The respiration rate of an activated sludge fed with a standard amount of synthetic

sewage feed is measured after a contact time of 30 minutes or 3 hours, or both. The

respiration rate of the same activated sludge in the presence of various concentrations of

the test substance under otherwise identical conditions is also measured. The inhibitory

effect of the test substance at a particular concentr-ation is expressed as a percentage of the

mean respiration rates of two controls. An EC50 value is calculated from determination at

differrent concentrations.

Defnitions

The respiration rate is the oxygen consumption of aerobic sludge or waste-water microorganisms expressed generally as Mg 02 per litre per hour.

EC50 in this study is the concentration of the test substance at which the respiration rate is

50 per cent of that shown by the control under conditions described in this report.

Contains TSCA Confidential Business Information
P age 13 of 23 pages

Renort 97 50 40 806 L~ Respiration Inhibition Test

Applicability I b

Chemicals which are non-volatile and are soluble in water are assessed by this method.

For test substances with limited solubility in the test media, it may not be possible to

determine the EC50.

5.2 Equipment and Reagents

5.2.1 Equipment

Standard Laboratory Equipment and Glassware

Analytical balance: Mettler AT 261 Delta Range

Sartorius Analytic A200S

Centrifuige: SIGMA 3-1

Air supply: Behromat membrane pump 06002, airflow 7-9

1/min, distributed on 12 outlets resulting in

airflows of 0.58 - 0.75 I/min per vessel

Air flow meter: Rota M40 2 - 16 1/min

pH-meter: Knick Calimatic (deviation from protocol)

Oxygen meter: WTW OXI 537 with electrode EO 96 connected

to a Lavmann, Mini 220N xy-plotter

Measuring apparatus: BOD flask

Vessels: PE bottles (deviation from protocol)

Autoclave: Webeco GmbH

5.2.2 Reagents

Sodium hydroxide: Merck, p. a., Lot No. K22393704 551

Sulfuric acid: 0.5 M, Merck, p.a.

Sodium hydroxide: I mol/l, Merck, p.a., Lot No. 70234 255

3,5-dichlorophenol 97 %: Aldrich No. D 7,060-0, Lot No. 00410-055

Peptone: Merck, Lot No. V 778513 618

Meat extract: Fluka, Lot No. 354902/1 12897

Urea: Fluka, p. a., Lot No. 283942 389

Sodium chloride, NaCi: Merck 1.06404 p.a., Lot No. K22393704 551

Calcium chloride, CaCI2 - 2 H20: Merck, p.a., Lot No. 129TA129682

Magnesium sulfate, MgSO4 - 7 H20: Merck, p.a., Lot No. A918686630

Dipotassium hydrogen phosphate: Merck, p.a., Lot No. A913699633

K,2HP0 4 - 3 H20:

Potable water: tap water Karlsruhe

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Page 14 of 23 pages

Report V7 50 40 806 f VRespiration Inhibition Test

5.3 PERFORMANCE OF THE TEST

5-3.1 Reference Material

3,5-dichiorophenol as a known inhibitor of respiration was used as reference compound

and tested for EC50 in order to check that the sensitivity of the used sludge was not

abnormal.

5.3.2 Solutions of the Test Substance

Solutions of the test substance was freshly prepared at the start of the study using astock

solution. A stock solution concentration of 0. 150 g/l was prepared. Due to the low

solubility of the test substance a higher concentration (e.g. 0.5 g/l as cited in the

guideline) was not appicable:

37.5 mg(- ?lwas filled into a 250 ml volumetric flask. The sample was dissolved

with purified water and the flask was made up to volume with this solvent.

concentration: 150 mg/l(

5.3.3 Solutions of the Reference Substance

0.5000 g 3,5-dichiorophenol was weighed into a 100 ml beaker. The sample was

dissolved by addition of 10 ml I M NaOH. After addition of approximately 20 ml

purified water, 0.5 M sulfuric acid was added under stirring until the point of incipient

precipitation (about 8.5 - 9 ml of 0.5 M sulfuric acid was required) was reached.

Thereafter the solution was quantitatively transferred into a 11I volumetric flask and the

flask was made to volume with purified water. The pH of the resulting solution was

determined to 7.80.

5.3.4 Test Concentrations

Five concentrations of the test substance, spaced by a constant factor not exceeding 3.2,

were used for the test:

From the stock solution with a content of 150 mg/l five concentrations were made (factor

2): 100, 50, 25, 12.5 and 6.25 ml of the stock solution were diluted each to test solutions

of 500 ml volume corresponding to concentrations of 30, 15, 7.5, 3.75 and 1.88 mg/l in

the test solutions.

Three concentrations of the reference substance were used: 10, 25 and 50 mg/I (made

from 10, 25 and 50 ml of a stock solution with the concentration of 0.5 g/l).

Contains TSCA Confidential Business Information /4
Page 15 of 23 pages

Report 97 50 40 806 ( 'Respiration Inhibition Test

5.3.5 Synthetic Sewage Feed

A synthetic sewage feed was prepared by dissolving the following amounts of substances

in I litre of water:

16.00 g peptone

11.00 g meat extract

3.00 g urea

0.70 g sodium chloride, NaCI

0.40 g calcium chloride, CaCI2 - 2 H20

0.20 g magnesium sulfate, MgSO4 -7 H20

2.80 g dipotassium hydrogen phosphate, K2BPO4 - 3 H20*

C... This sewage feed was prepared on December 19, 1997, filled into 100 ml PE-bottles and

was autoclaved. The tightened bottles were held at + 4*C in the dark until start of the

respiration inhibition test (December 23, 1997). The pH value of the sewage feed was

determined to 8.0.

5-3.6 Activated Sludge

A fresh sample of activated sludge (suspended in approximately 8 1 water) -of the

municipal sewage plant Neureut/Karlsruhe taken on December 22, 1997 was treated as

mentioned in the cited guideline:

The sludge was precipitated. The sludge was washed with potable water, the supernatant

liquid was sucked off using a tube connected with a water jet pump (deviation from

protocol). This procedure was repeated three times. A small amount of the sludge was

filled into a glass dish in duplicate, weighed and dried overnight at 105 IC in order to

obtain the dry matter. From this result the amount of wet sludge was calculated which had

to be suspended in water in order to obtain an activated sludge with a mixed liquor

suspended solids level of 4 g/l. This level gave a concentration of 1.6 g/I in the test

medium following the procedure listed below.

The so prepared sludge was kept aerated. Due to the fact that it could not be used on the

day of sampling, 50 ml of synthetic sewage feed was added to each litre of the activated

sludge prepared as described above; this mixture was then aerated overnight at 20 ± 2*C.

*Deviation from protocol: K2HPO4 *- 3 H20 instead of K2HPO, was used.

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Page 16 of 23 pages

Report 97 50 40 806 ( )/Respiration Inhibition Test

5.3.7 Test Conditions

Ies

Duration/contact time: 3 hours, during which aeration took place

Vessels: PE bottles

Water: Potable water Karlsruhe

Air supply: Bebromat membrane pump 06002

pH meter. Knick calimatic

Measuring apparatus: BOD flask

Oxygen meter: WTW OXI 537 connected to an xy-plotter

Nutrient solution: Synthetic sewage feed (see above)

Test substance: Freshly prepared test solution (5 different concentrations)

Reference substance: 3,5-dichlorophenol (3 different concentrations)

Controls: Inoculated samples without test substance (2 samples)

Temperature: 20.3 - 22.6 0C

5.3.8 Test Procedure

10 PE-bottles (in deviation from protocol) were used.

Control

At time "0" 16 ml of the synthetic sewage feed was made up to 300 ml with water. 200

ml of microbial inoculum was added and the total mixture (500 ml) was poured into the

first bottle (first control). Aeration was started.

Test substance / reference substance

At time "15 min" the above was repeated except that 100 ml of the test substance

(reference substance, respectively) stock solution was added to the 16 ml of sythetic

sewage feed before adding water to 300 ml and microbial inoculumn to make a volume of

500 ml. This mixture was then poured into the second vessel and aerated as above.

This procedure was repeated at 15-minute intervals with different volumes of the test

substance stock solution and the reference substance stock solution to give a series of

vessels containing 5 different concentrations of test substance and three for the reference

substance. Finally, a second control was prepared.

After three hours the pH of the solutions was measured. Then contents of the first vessel

were poured into the BOD flask and the oxygen consumption was measured over a period

of 10 minutes. This determination was repeated on the contents of each vessel at 15-

minute intervals, in such a way that the contact time in each vessel was three hours.

5.3.9 TEST VALIDITY

The test results are considered to be valid if

the two control respiration rates are within 15 per cent of each

other,

the EC50 (3 hours) of 3,5-dichlorophenol is in the accepted

range 5 to 30 mg/I.

Contains TSCA Confidential Business Information
Page 17 of 23 pages

R eport 97 50 40 806

)/Respiration Inhibition Test I

6 RESULTS

6.1 Activated Sludge - Determination of Thy Matter

Initial weights before drying

Test No. glass dish / glass dish + sample

tare sample weight

114.2411 g 24.7516 g 10.5105 g

2 13.8688 g 22.1165 g 8.2477g

Final weights after drying overnight at 105 0C

Test No. glass dish / glass dish + sample weight dry matter

___ ___ ___ tare sample _ _ _ _ _ _ _ _ _ _ _ _

1 14.2411 g 14.4244 g 0.1833 g 1.74%

2 13.8688 g 14.0138 g 0.1450 g 1.76%

The dry matter (mean value) of the activated sludge was determined to about 1.8 %

From this result the amount of wet sludge was calculated which had to suspended in

water in order to obtain an activated sludge with a mixed liquor suspended solids level of

4 g/l:

555.69 g activated sludge was diluted to a volume of 2.5 1 with water.

Contains TSCA Confidential Business Information

A/7
Page 18 of 23 pages

Report 97 50 40 806

( /Respiration Inhibition Test Is

6.2 Observations / Treatment of Results

Oxygen Consumption was measured and recorded after an aeration time of 3 hours

(contact time) by means of an oxygen meter connected with an xy recorder.

The respiration rate was calculated from the recorder trace as mg 02/1 and h between

approximately 6.5 mg 02/1 and 2.5 mg 02/, or over a 6 minute period (in deviation from

protocol) when the respiration rate was low. The respiration rate was measured over the

linear portion of the respiration curve.

The respiration rate was expressed as a percentage of the mean of the two control

respiration rates:

2Rs

1 -_______ *100 =percent inhibition

Rc1 + Rc2

where

Rs oxygen-consumption rate at tested concentration of test substance

Rc, oxygen-consumption rate, Control I

RC2 oxygen-consumption rate, Control 2

OECD No. 209: The respiration rates of the two control samples must be within 15 per

cent of each other. The EC50 value (3 h) of the reference substance

must be in the accepted range. Otherwise the test is invalid and has to

be repeated.

The per cent inhibition was calculated at each test concentration as above. The per cent

inhibition was plotted against the logarithm of concentration and the EC50 was derived.

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Page 19 of 23 pages

Report 97 50 40 806

( IRespiration Inhibition Test

6.3 Table of Results I

Tabellel

C J- Respiration inhibition Test (OECD 209)

stock soin. stock samn.

sample content volume sample conc. temperature start time

control 1I - - 20,3 10:45 a.m.

reference 1 500 10 10 20.8 11:00 a.m.

reference 2 500 25 25 21,3 11:15 a.m.

rerence 3 500 50 50 21.5 11:30 a.m.

150IS 6,25 1,88 22,1 11:45 a.m.

/2 150 12,5 3,75 22,3 12:00 a.m.

/3 150 25 7,5 22,2 0:15 P.M.

/4 150 50 15 22,4 0:30 p.m.

150 IS 100 30 22,6 0:45 p.m.

n o1 2 1 - I - 122,3 1:00 p.m.I

pH determinations: - synthetic sewage feed: 8.0

- reference sample / stock. soln.: 7.8

I stock soln.: 7.5 - aerated sewage: 7.75

measurement of oxygen content

evaluation

sample time duration from to difference consumption inhibition

____ f~m min] 1mjA [m/1 [m [mgl(irh)] A1% . control 1 1:45 p.m. 6 5.1 1,6 3,5 35 -

reference 1 2:00 p.m. 6 5.8 3,4 2,4 24 34.2

reference 2 2:15 p.m. 6 7,0 5,4 1,6 16 56,2

r~mfernce,3 2:30 p.m. 6 7,3 6,3 1 10 72,6

1 l 2:45 p.m. 6 5,5 1,0 4,5 45 -23.3

1 /2 3:00 p.m. 6 5,8 1,7 4,1 41 -12,3

1 /3 3:15 p.m. 6 5,1 1,1 4 40 -9,6

( 4 3:30 p.m. 6 5,6 1,3 4,3 43 -17.8

conol 2 4:00 p.m. 1 6 1 5,5 1,7j 3.8 38 1 -

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Report 97 50 40 806 ( )/ Respiration Inhibition Test

6.4 Graphical Evaluation of EC,, of the Reference Substance I

Tabelle2

Reference sample 3,5-dichiorophenol

Calculation of EC5O

10~coflc.. inhibition N

.,000 34,2xcofiin: 5,6

13858,2 constnt: 0,72

1.699 72,6 correl. coeff.: 0,999992

3,5 dichlorophenol

70

s.~ .....

C30-

0.000 0,500 1,000 1,500 2,000

log (com-.)

Determination of ECSO at 50% inhibition via graphical evaluation:

inhibition [%] = (Iog(conc.) *54.956).- 20.72

Iog(conc.) = (inhibition [%] + 20.72) / 54.956

log(conc.) = (50 + 20.72) / 54.956

Iog(conc.) = 1.287

concentration (EC5O) = 19,36 mg/I

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Report 97 50 40 806 ( \/Respiration Inhibition Test

6.5S Exemplary Recorder Plots

Control I

10.4 1111IIT1

I Ii~ I

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Report 97 50 40 806 t )/Respiration Inhibition Test

Reference 2 25 mg/I 3,5-dichiorophenol

I N II It I I

11 11 1111%-4 H il I I I I 1 1 1 !1

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Report ~7 50 40 806

~Respiration Inhibition Test

/3 7.5 mg/I LA 409

44 ' I! KI

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attachment 13

BioChem

Am Wieseneck 7 -D-04451 Cunnersdorf I s gr Tel. 03 42 91/416-0 -Fax 03 42 91/416 22Lao E-mail: biochem@ btiochemn.de rbilgshun Internet: http:/Avww.biochem.dechmseAaltkGb Laborch furbilgih undH

Final report

97 10 48078

Acute Toxicity Fish Test

Zebra rish - Brachydanio rerio

according to OECD Guideline 203 (1992)

BioChemn project No.: 97 1048078

Date of order: 30.10.97

Test substance:(

Manufacturing date: 30.06.97

Sponsor:

Monitor:

TESTIN FAciLrry: Bi oChem, agrar Am Wieseneck 7

D-0445 1 Cunnersdorf

Testing Facility Management: Dr. 1. Winkler

Study Director: Dr. R. Kleiner

Quality Assurance Manager: A- Kistner

ANALYTICAL LABORATORY: BioChemn GmbH Karlsruhe

Daimlerstrale 5b

D-76 175 Karlsruhe

Responsible scientist: Dr. Walter

Quality Assurance Manager: Dr. Merten

Study completion date: 12.03.98

Final report 97 10 48 078

Acute Toxicity Fish Test(

Original 2 of 2

Page I of 18 Contains ISCA Confidential Business Information
Final report 97 10 48078

Acute Toxicitiy Fish Test

Original 2 of 2 i

Page 2 of 18

.. ISACHSISCHES STAATSMILNTSTERRJMI FUR UMWELT

.IND LANPESENTWICKLUNG

G LF-aescheinigung

Elescheinlgung Crtificate

Hiemfnit wird best~ligt. dali die Prfifungseinrichtung(en) It is h~ereby certified that the test facflity(ies)

04451- Cunnersdorf.,V 04451 CunnersdCrE

~ in AM illne c 7 . iseneq*c< ar 7

de................................ . .....................................

27./28.02.1996 und ain .2 1 0 5 1996 -Februai Y.27-23,1990' and '..ay 21,139;

tO.2tum) (date)

von der fOr die Ciberwachung :uslAndigen Sehfrde Ober was (were) inspected by the competent authority regarale Einhaltung der Geundsdtze der Guten Laborpraxis ding compliance with U'.e Priicacles ci Good Laboratory

inspiziert warden ist (sind). Practice.

Es wird hierrnit bestatigt. dali folgende Pri~fungen in It is hereby certified that szudies in this test facility are

dieser Prlifeinnichtung nach den Grundsdtzen der Guten conducted in compliance with the Principles or Good

Laborpraxis durchge(Ohrt werden. . Laboratory Practice.

PrUfkatagoria 4 Umwelttoxikalogiscna PrUfungan zu

Auswirkungen auf aquatische und4

tercastrische Crganismner

?riifkategorie 5 Pri~fung3n zu.n Van't!- :.T, v

;assar und i.n z.az LuEt;

?rjE a gorie '6 P r,'fungan aur -C.s tan=

Crasden, den 13.06.1995

R I(LatrSA H E

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Acute Toxicitiy Fish Test(~

Origina 2 of 2S

Page 3 of 18

GLP Compliance Statement

Project No.: 97 1048 078

Title: Acute Toxicity Fish Test

Zebra fish - Brachydanlo rerio

according to OECD Guideline 203 (1992)

I hereby declare that this study was performed according to the study plan by the procedure

described in compliance with the "Principles of Good Laboratory Practice (GLP)"

("Chemikafiengesetz, Juli 1994, Anhang 1").

The documented data correspond to the experimentally determined values.

There were no circumstances that may have adversely affected the quality or integrity of the data.

Study Director: D..R .Ken ...........

Quality Assurance Statement

Procedures of this study were periodically inspected by the Quality Assurance Manager and the

results reported to the Study Director and to the Testing Facility Management.

Inspections/ Dates of inspections/ Dates of report

audits audits

Study plan 20.10.97 20.10.97

Experiments 05.109.12.97 12.12.97

Final report 09.03.98 12.03.98

I herewith attest the authenticity and the correct and accurate performance of this study according

to the "Principles of Good Laboratory Practice (GLP)" ("Chemikaliengesetz, Juli 1994,

Anhang 1").

Quality Assurance Manager: ............. ........ A. Kiistner Date

Testing Facility Management: >. L:& ................. .-..... 4.......

Dr. J. Winkler Date

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Acute Toxicitiy Fish Test

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Page 4 of 18

Table of contents

Page

Title page I

GLP Certificate 2

GLP Compliance Statement 3

Quality Assurance Statement 3

Table of contents 4

Summary 5

1. General information 6

1. 1. Objective 6

1.2. Project staff 6

1.3. Time schedule 6

1.4. Test guidelines 6

1.5. Amendments to the study plan 6

1.6. Archiving 6

2. Performance of the test 7

2.1. Test substance 7

2.2. Test system 8

2.3. Experimental procedure 9

2.3.1. Preparation of the test solutions 9

2.3.2. Course of the trial (chronological schedule) 10

2.3.3. Assessment of the test 11

2.4. Calculation I1I

3. Results 12

3.1. Cumulative mortality of each concentration 12

3.2. Abnormal responses of the fish ., 13

3.3. Results of the analytical determination of( ) 13

in the test solutions

3.4. Oxygen concentrations, pH values and temperatures 14

3.5. Length and weight of fish 16

4. Conditions for the validity of the test 17

5. References 17

Appendix: Analysis of the tap water 18

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Acute Toxicitiy Fish TM(t

Origina 2 of 2

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Summary

Acute toxicity fish test J

Test substance:

Manufacturing date: 30,06.97

Chemical name:

Solution stability: not stated

Fish species: zebra fish - Brachydanio rerlo

Test procedure: statically

Analytical method: H1igh Performance Liquid Chromatography

The method and the results of the analytical determination of the test concentrations are described

in the related report "Determination of Test Sample Concentration in Combination with the Acute

Toxicity Fish Test of( " dated March 04, 1998 (as separate annex to this report).

Results

Nominal concentrations of( ),nominal / measured, all in mg/I)

48 h after application 96 ii after application

NOEC 19.8 / 13.7 29.6 / 20.0

LC0 29.6 / 21.2 29.6 / 20.0

LC50 36.3 / 27.4 36.3 / 26.8

LCo 44.4 / 35.4 44.4 I36.0

The LC,0 values are determined according to the statement of the EC Directive 92/69/EEC, C. 1:

"When two consecutive concentrations ... give only 0 and 100 % mortality, these two values are

sufficient to indicate the range within which the LCso falls."

Only moderate degradation was observed at relevant concentrations.

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Acute Toxicitiy Fish Test,

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1. General information

1.1. Objective

The objective of the study was to determine the acute toxicity of the test substance ) characterized by the median lethal concentration at 96 hours after application (96 hours L 5o).

The examination of the toxic effects was performed with the zebra fish (Brachydano rerio) as a

representative test organism for aquatic vertebrates according to OECD Guideline for Testing of

Chemidcals 203.

1.2. Project staff

Study Director: Dr. agr. Ralf Kleiner

Personnel: Martina Mittag (laboratory assistant)

Responsible scientist for analysis: Dr. Walter

Mr. Werle was the responsible technical assistant for the analytical examidnations

(Labor Karlsruhe).

1.3. Time schedule

Study initiation date: 14.11.97

Signature of the study plan by the sponsor: 20.11.97

Experimental start date: 05.12.97

Experimental termination date: 13.12.97

Study completion date: 12.03.98

1.4. Test guidelines

- OECD Guideline for Testing of Chemicals 203 (1992) "Fish, Acute Toxicity Test" - Standard operative procedures (SOPs) of the testing facility

1.5. Amendments to the study plan

The study was performed in compliance with the study plan.

1.6. Archiving

Study plan and final report as well as the raw data are kept in the GLP archives of BioChemn agrar

in compliance with the GLP principles.

A retention sample of the test substance is stored for 5 years.

Destruction of any data or retention sample of the test substance requires the sponsor's consent.

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Acute Taxicitly Fish Test,(

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2. Performance of the test

2.1. Test substance

Code No.:()

Manufacturing date: 30.06.97

Expiry date: no information

Active ingredient:(

Density (20 *C): 0.96 g/cm3

Appearance: colourless clear liquid

Solubility in water: <~ 0.5 g/l (25 -C)

Storage conditions: room temperature, container keep closed,

class of storage (VCI): 10

Stability/homogeneity: not stated

Classification: designation according to EC-Classification/GefStoffV:

"30' - Irritant; R 36/3 8; S 24/25, 36/37/3 9

Further details: "Sicherheitsdatenblatt" (91/1S5/EWG) of 03.11.97

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Acute Toxicitiy Fish Test,.( Original 2 of 2

Page 8 of 18

2.2. Test system

Test organism: zebra fish - Brachydanio renio

Origin of the animals used in the test: purchase from the fish farmning Erfiirter Neon GbR

Total length of the test fish: 2.5 - 3.7 cm

Holding conditions

The test fish were in good health and were acclimatized for 15 days, of which the last 7 days they

were kept in water of the same quality used in the test.

Light-dark-cycle: 16 h fight : 8 h dark, daily

Temperature: 21 - 25 0C

Oxygen concentration: at least 80 % of air saturation value

Feeding: daily (Tetra Mmi Hauptfutter)

until 24 hours before the test was started

Tap water of Cunnersdorf of the following quality was used during the holding time and the first

period of acclimatization (see appendix):

acid capacity K. 4.3: 1.26 mmol/l

sum of calcium and magnesium ions: 77 mg/l calcium Ca2 ': 63.1 mg/I

magnesium Mg'*: 13.9 mg/I

sodium Nae: 25.2 mg/I

potassium K~: 6.8 mg/i

pH: 7.3

During the acclimatization period the fish were not treated. Not any fish died.

Test conditions

Test vessel (pool): 10-l-aquarium (40 cm x 25 cm x 25 cm)

Test volume/vessel: 101

Number of fish/vessel

(= concentration or control): 10

Loading: maximum loading of 1.0 g fish/i

Test procedure: static test

Use of vehicles: none

Climatic conditions

Temperature: 22.5 - 23.2 0C

Oxygen concentration: 8.5 - 9. 1 mg/I (i. e. 93 -100 % air saturation value)

Feeding: none

Light-dark-cycle: 16 h light : 8 h dark, daily

Test duration: 96 hours

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Page 9 of 18

Test water

Conductivity of deionized water: 0.4 - 1.5 pS/cm

Dilution water used in the test period was reconstituted water:

CaCI2 2 rnnol/l

MgSO 4 0.5 mmol/l

NaHCO 3 0.75 minol/I

KCI 0.075 mmol/l

The sum of calcium and magnesium ions in the dilution water was 2.5 mmol/l. The proportion of

Ca : Mg ions was 4 : 1 and of Na : K ions 10 : 1. The acid capacity K. 4.3 of the solution was

0.8 mmol

Derived from the CaCl2 addition, the hardness of water was equal to 2 mmol/l CaCO3, which

corresponds to a hardness of 200 mg/l CaCO3 (valid: 10 - 250 mg/l).

pH value of test water: light basic (8.5 - 9. 1)

2.3. Experimental procedure

2.3.1. Preparation of the test solution

The solubility of the active ingredient ( )in potable water is <0.5 g/l (as stated by the

sponsor).

Each pool contained 10 1 dilution water. The control pool (pool No. 1) contained dilution water

only.

In a range finding test the concentration range of 10 to 100 mg/l was determined for the maintest. The fac~or for dilution used was 1.5.

3 gL ( vas, transferred into a 3000 ml volumetric flask and filled up to volume with dilution

water This solution was stirred overnight for the purpose of saturation.

Stock solution A: 1000 mg/I

The preparation of the test solutions is summarized in the following tables:

Preparation of the stock solutions

solution amount total amount stock solution

amount name of dilution water of solution name concentration

21 A + 1 31 B 667 mg/I

21 B + 1 3 1 C 444 mg/i

21 C + 1 31 D 296 mg/I

21 D + 1 31 E 198 mg/I

21 E + 1 31 F 132 mgfi

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Page 10 of 18

Filling of the test vessels (pools)

pool concentration of (amount of stock solution

No. (mg/I) dilution water amount name

1 0 (control) 10 1

2 100 91 + I11 A

3 66.7 91 + I1I B

4 44.4 91 + I11 C

5 29.6 91 + I I D

6 19.8 91 + I11 E

7 13.2 91 + I1I F

The saturated solution and the dilution water as well were prepared the day before (in the

afternoon). They were stirred overnight in the same cabinet and at the same temperatures as the

fish test was performed.

The pools were permanently aerated by an aeration pump (type SP 402 SLR).

Not dissolved )~was not discernible in the pools.

2.3.2. Course of the trial (chronological schedule)

The chronological schedule is valid for each performed test.

7 days before the start of the test

- Transferring of the fish in fresh dilution water

24 hours before the start of the test

- Determination of physico-chemnical measured values of the prepared dilution water and

adaptation to the methodical requirements (pH, oxygen concentration, temperature)

- Preparing of the stock solutions of the test substance

At the beginning of the test

- Control of pHL oxygen concentration and temperature of the dilution water

- Preparing of the test vessels and filling of the dilution water into the test vessels

- Preparing of the required test concentrations by addition of the corresponding volumes of the

stock solutions into the test vessels (= application)

- Sampling of the test solution of each test concentration for analysis (main-test)

- Transferring of the test animals into the test solutions

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Final report 97 10 48078

Acute Toxicitiy Fish Test,(

Origi"a 2 of 2

Page 11 of 18

3, 6, 24, 48, 72 and 96 hours after application

- Determination of the number of immobile fish

0, 24, 48, 72 and 96 hours after application

- Measurements of pH, oxcygen concentration and temperature

- Sampling of the test solution of each test variant for analysis (main-test, only 48 and 96 hours)

- Waste disposal

2.3.3. Assessment of the test

Time:

- 3, 6, 24, 48, 72 and 96 hours after application

Parameters:

at the beginning of the test: pH*)

oxygen concentration~

temperature

substance concentration

*3, 6, 24, 72 and 96 hours after application: number of immobile fish

in each test vessel

*24, 48, 72 and 96 hours after application: pH*)

oxygen concentration

temperature

substance concentration~

=determination of each test concentration

2.4. Calculation

The LDso values (48 and 96 h) were derived from calculation (par example: 41 29.6 * 44.4) being

considered the geometric means of LCo and LC too concentrations.

The LC5o value (3 h) was graphically obtained resulting from the plot on probit paper.

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Acute Toxicitiy Fish Test(

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Page 12 of 18

3. Results

3.1. Cumulative mortality at each concentration after 3, 6, 24, 48, 72 and 96 hours

Number. of dead fish

LA 409 3 h 6 h 24 h 48 h 72 h 96 h

noinal concentr.___________

control 0 0 0 0 0 0

100.0 mg/I 10 10 10 10 10 10

66.7 mg/I 10 10 10 10 10 10

44.4 mg/I 9 10 10 10 10 10

29.6 mg/I 0 0 0 0 0 0

19.8mrg/I 0 0 0 0 0 0

1 13.2 mg/i 0 1 0 1 0 1 0 1 0 1 0

The NOEC (no observed effect concentration) after 96 hours was 20 mg<l )(measured

concentration).

e hghest tested concentration causing no mortality after 96 hours (96 h LCo) was 20 mg/I

Ymeauredconcentration).

The lowest tested concentration causing 100 % mortality after 96 hours (96 h LC100) was 36 mg/i (- measured concentration).

The 3 h median lethal concentration of (3 h Ljie.the concentration which proved

lethal to 50 % of fish at the time of 3 h wL~ 35o) mg drie rphicaly).

The median lethal concentration of f ),i.e. the concentration which oroved lethal to 50 % of

fish was 27.4 mg/l~ at the timie o 48 hours and 26.8 mgA( ) t 96 hours (derived

arithmetically).

The fish were inspected at least daily. Fish were considered to be dead if touching the caudal

peduncle produced no reaction. Dead fish were removed when observed.

The control fish showed neither mortality nor any visible sign of abnormality during the test time

of 96 hours. Incidents in the course of the test which might have influenced the results could not

be observed during the test period.

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Final report 97 1048O078

Acute Toxicitiy Fish Tes/

Original 2 of 2

Page 13 of 18 

a

3.2. Abnormal responses of the fish

observation time concentration of( N)visible signs of abnormalities

/ of surviving test fish

3 h 66.7 and 10mg/I 10 fish perished

44.4 mg/I 1 fish lies upside down on the bottom of

the pool. wvith respiration movements

6 h 29.6 mg/I 2 fish shows intensive respiration

activity and uncontrolled movements

24 h 29.6 mg/I 2 fish shows intensive respiration

activity and uncontrolled movements

48 h 29.6 mg/i I fish shows intensive respiration

activity and uncontrolled movements

At any other time of inspection no visible signs of abnormalities of surviving test fish were

observed.

3.3. Results of the analytical determination of( ')n the test solution

Concentration of( the test water

Date of measurement: 18. - 21.02.98

nominal mean of measured concentrations (mg/I)

concentration at the time of

(Mt)0h 48hI 96 h

control <0.5 <0.5 <0.5

100.0 100.4 96.4 93.7

66.7 65.6 61.8 61.2

44. 41.4 35.4 36.0

29.6 25.6 21.2 20.0

19.8 16.9 13.7 13.1

13.2 10.4 8.8 8.5

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Acute Toxicidy Fish Test,(

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Page 14 of 18

Degradation of

nominal( recovery in % of recovery after 96 h in %

concentrtion the nominal ( bae)ntemaue

control -

100.0 100.4 93.7 93.3

66.7 98.4 91.8 93.3

44.4 93.2 81.1 87.0

29.6 86.5 67.6 78.1

19.8 85.4 66.2 77.5

13.2 78.8 64.1 81.3

For precise statements see the related analytical report.

3.4. Oxygen concentrations, pH values and temperatures

The dissolved oxygen concentrations of the control and of the test solutions were checked daily at

24 hours intervals.

Dissolved oxygen concentration of the test water (mg/i1):

concentrations (nominal) 0 h 24 ht 48 ht 72 ht 96 h

control 9.1 8.9 8.6 8.6 8.6

100.0 mg/I 9.1 8.9 8.6 8.6 8.6

66.7 mg/i 9.0 8.9 8.6 8.6 8.6

44.4 mg/I 9.0 8.9 8.6 8.5 8.5

29.6 mg/I 9.0 9.0 9.0 8.9 8.9

19.8 mg/I 9.0 9.0 9.0 8.9 8.9

13.2 mg/I 9.0 9.0 9.0 8.9 8.9

volumetric instrument: 02-meter OXI 96

WTW Wissenschaftlich-Technische Werkstatten GmibH

D-82362 Wejiheim

/97
Final report 97 10 48 078

Acute Toxicitly Fish Test

Original 2of2 1

Page 15 of 181 Is

The pH values of the control and of the test solutions were checked daily at 24 hours intervals.

The pH values of the test solutions were not adjusted.

pH value O h 24 h 48 h 72 h 96 h

control 7.79 7.79 7.76 7.76 7.77

100.0 mg/I 7.79 7.79 7.76 7.76 7.76

66.7 mg/i 7.79 7.79 7.76 7.77 7.76

44.4 mng/i 7.79 7.80 7.78 7.76 7.76

29.6 mg/i 7.79 7.86 7.78 7.77 7.76

19.8 mg/I 7.78 7.89 7.83 7.86 7.86

13.2 mng/I 7.89 7.90 7.86 7.86 7.86

volumnetric instrument: pH-meter pH 95

WTW Wissenschaftfich-Technische Werkstitten GmnbH

D-82362 Weilheirn

During the test period the following water temperatures were measured:

O h: 22.5 0 C

24 h: 22.7 OC

48 h: 23.0 0 C

72 h: 23.2 0C

96 h: 23.0 0C

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Acute Toxicitiy Fish Test,(

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Page 16 of 18

3.5. Length and weight of the fish

Length of fish

total length of 10 fish (cm)

control -nominal concentration (mg/I)

100 66.7 44.4 29.6 19.6 13.2

2.5 3.1 3.3 3.4 3.1 3.5 3.1

2.9 3.4 3.4 3.5 3.3 3.5 3.0

2.8 3.0 3.4 3.5 3.1 3.4 3.1

3.0 3.4 3.6 3.2 3.2 3.1 3.1

353.3 3.5 3.5 3.1 3.0 3.2

3.0 3.6 3.1 3.4 3.5 2.8 3.3

3.1 2.7 3.4 3.1 3.4 3.0 3.2

3.4 3.4 3.4 3.4 2.7 3.0 3.2

3.2 3.2 3.5 3.3 3.1 3.5 3.0

3.0 3.7 3.1 2.8 3.2 3.1 3.4

average 3.04 3.28 3.37 3.31 3.17 3.19 3.16

Weight offish

total weight of 10 fish (mg)

control (-nominal concentration (mg/I)

100 66.7 44.4 29.6 19.6 13.2

134 245 281 307 248 471 248

166 289 308 293 321 407 228

164 174 299 312 310 426 277

200 298 392 231 298 303 290

301 254 345 340 247 256 276

212 422 229 235 313 178 318

257 177 283 244 346 253 306

301 228 304 269 157 240 326

234 246 267 288 279 453 212

175 385 252 195 283 276 349

average 214.4 271.8 296.0 271.4 280.2 326.3 283.0

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Page 17 of 18

4. Conditions for the validity of the test

Constant conditions had been maintained throughout the test.

There was no mortality in the control.

The dissolved oxygen concentration had been at least 60 per cent of air saturation value.

In all test solutions the measured pH-fluctuation was below 1 pH-unit during 96 hours.

There was analytical evidence that the( )concentration had been satisfactorily maintained

during the test period.

5. References

OECD Guideline for Testing of Chemicals, 203 "Fish, Acute Toxicity Test".

OECD July 17, 1992.

EC Directive 92/69/EEC, Annex, Method C. 1., Official Journal of the European Communities

No. L 383 A, December 29, 1992.

DIN 38412, Teil 15: Testverfahren mit Wasserorganismen (Gruppe L).

Bestimmung der Wvirkung von Wasserinhaltsstoffen auf Fische.

Deutsche Norm, Deutsches Institut tur Normung e.V. Berlin, Juni 1982.

Letale Wirkcung beimn ZebrabIrbling Brachydanfo renao (LCO, LC5O, LC 100; 48-96 Stunden). Verfahrensvorschlag der ad hoc-Arbeitsgruppe des Tjmweltbundesamtes zur Entwicklung 6kotoxikologischer Testverfahren in aqjuatischen Systemen, Mai 1984.

LrrcHFMEL, J.T. and WILCOXoN, F.:

A simpled method of evaluating dose effect experiments.

J. Pharm. Exper. Ther. 9-6 99-113, 1949.

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Orignal 2 of 2

Page 18 of 18

Appendix: Analysis of the tap water

Kocturas Waaksw Leipzg CjmbL

Wauta hc PnmfxdlIc

Triakwusgfabm

Duizbsazamogym 1996

Emahmvicie. %W Tbiuft

cinhch nub T%".a

AntaI dr OWC bei cli. KaiIn, in% 0

" d. RWt Koc2 b .C in% V.0

7.9U d. RWC Xalos~a'jhI & 36C m %

PH-wen 6.9.0 12

p11-Wot &. CaCVWdmzq &.

Deb apX .0.2 .0.3

L*US;~da P$'cm 2000 37 S

1I.-bmng T~rl L5 0.3

Fkbua Eam 0.3 C0.2

OXierakagbI m4e/ 5 1.76

QUvdm~.cheijmwt 3 <

Aauiomauma 0.5 < 0.05

.Ninit mgW 0.1 -0.0 1

?'1iti M&O s 13.1

L-Alcdd Mal 250 35

m4an LS 0.79

Sufit -ngl 24m .4

Phoiphae SOIt 6.7 00

Natrtw MVSO5 23.2

Kalium =91v 12 6.:

K~um z' 00 $3.1

I"aali Sw s .13.9

fr.gumCO2 ~ 69

gm.Kjsm Mql 0.2 0.05

U.aaga owl 0.011 O.C0I

Aluuu 0.2 0402

Sbc LIM 10

Dau e1 0.05

w,'1 I z 0.2

g&eaL.-CWI XcwIwwma& RIC 0.01 <0.0 1

C)=d U1 140 <10

Oberflicmnaktive Subdaza-vocib SLO& C401 0.2 < 0.01

~L140 .. 4

1LUI 0.71

Nlmkti 940 50 C.1

Quedrealbcr ;L84 I1 .a.)1

SONGs ;iL 10

AnoM410 <1

Kuizp Mal 3 1 0.05

Zink Mal1 5 0.15

Swumm Tnhmioqawchac )W 10 V? I

TanmJddmJica A9l 3 < 0.5

'wLrm PIv 0.3 < 0.02

SUOI f""Yarb )wl .s < 0.03 Contafns&odb Cofienia Inr.05~ 1 pestCAd <uins Contains Confidential TSCA Business In~rat= l~rM P e0. .00

gq~~~~rm/1 - /-vv-xr 
LC O Inventory Expert Service

A division of the American Chemical Society Phone: 800-631 -1884, 614-447-3600 (ext. 2308)

Fax: 614-447-3747

E-mail: answers@cas.org

INVENTORY EXPERT SERVICE REPORT

IES-ORDER NUMBER: 33902

REGISTRY NUMBER:(

CA INDEX NAME

Please print the above CA Index Name on the appropriate page of your PMN.

V If this box is checked, CAS has made correction(s) marked in red to ___ your IES order. Please make the same correction(s) to your PMN before

submitting it to the EPA.

Contains TSCA Confidential Business Information

2540 Olentangy River Road - P.O. Box 3343 Columbus, Ohio 43210-0334
PreManufacture Notice and

Notice of Commencement Forms

EPA Form 7710-25 (Rev. 5-95)

EPA Form 7710-56 (Rev. -96)

Installation and Use Instructions

MS Word 6. Ofor Windows

U.S. EPA Approved December 12, 1996

by Anthony Cheatham

TSCA Data Administration Section

Lw

r "401riil Responsible Cares

~,A Public Commitment

Page 173
6h

GOLDSCHMIDT CHEMICAL CORPORATION

March 10, 1999

Environmental Protection Agency

HQ Accounting Operations Board

Attention: TSIPMN

P.O. Box 360277 M

Pittsburgh, PA 15251

Gentlelnen:

Please find enclosed our check for $20,000.00 for submission of our Premanufacture

Notices (reference TS-199901; TS-199902; TS-199903; TS-199904; TS-199905;

TS-199906; TS-199907; TS-199908) sent to the Environmental Protection Agency,

Washington, D.C.

Sincerely,

Kelly C. Tiller

Safety & Environmental Technician

Safety & Environmental Compliance

1004 1 6R~0CECK DATE

10044'~~- 002577 03/10/1999

t, ,~.-.___ ~ ___ __ ____t

K. CHECKAMOUNT

S2<0,000-0

O1~OE ~' rIY 5s'rA>Th -. EPA

tf-f,;A 0ONI OPERATQNS- 8ARD

-P0 3'63027'M -',

111100 2577116 nOS0L.OL865t: 0U80?o?~" /

Vp 1'14 f4 17(