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Submission Information

Submission ID : 25349398 Submission Name : 1H-Benzotriazole, 4or 5-methylAR Number : 

Chemical Information

CAS Number : 29385-43-1 Ninth Collective Index Name : 1H-Benzotriazole, 4(or 5)-methylSubmitter Information

Submitter's Name : EPA

Address : 

City : Washington

State : DC

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Additional Technical Point of Contact Information

Technical Point of Contact : 

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Sponsor Information

Sponsor's Name : EPA

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Technical Point of Contact Information

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Title : 

Phone Number : 

Fax Number : 

E-Mail Address : 

 Acute Toxicity to Aquatic Vertebrates

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Vertebrates

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

Referred to as 5-Methyl-1H-Benzotriazole or 4-Methyl-1H-Benzotriazole (MeBT);

Provided by Cincinnati Specialties (Cincinnati Ohio)

Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Measured

Method - Acute Toxicity To Aquatic Vertebrates

Year Study

Performed:

Method/Guideline

Followed:

Other

Other

Method/Guideline:

Methods for measuring the acute toxicity of effluents and receiving waters to

freshwater and marine microorganisms

Deviations from

Method/Guideline:

Species: Pimephales promelas

GLP: No Data

Analytical

Monitoring:

Yes

Test Type: Semi-Static

Test Vessel:

Water Media Type:

Test

Concentrations:

Both

Nominal and

Measured

Concentrations:

nominal: 150.8 and 188.9 mg/L; measured high concentrations of MeBT were between 95.5%

and 99.5% of nominal concentrations at a confidence interval of 95%.
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Exposure Period: 96 Hours

Vehicle Used:

Vehicle Name:

Vehicle Amount

and Units:

Alkalinity:

Dissolved Oxygen:

pH Value:

Test Temperature

and Units:

Value/Lower Range: 20

Photo

(Light/Dark):

16/8

Salinity:

TOC:

Water Hardness:

Method/Guideline

Test Conditions

Remarks:

Fathead minnows were 7 days old at test initiation.

A dilution water control was included.

The toxicity of MeBT to fathead minnows was assessed in moderately hard water using

acute tests according to U.S. Environmental Protection Agency procedures. Mortality

was the endpoint. The nominal (predilution) test concentrations of MeBT were 150.8

and 188.9 mg/L. Test chambers were 500-mL glass beakers. All tests were conducted

at 20 C with a light:dark cycle of 16:8 h. Fathead minnows were fed 0.2 mL of brine

shrimp nauplii (Artemia sp.) at 48 h and were renewed two hours after feeding with

freshly prepared dilutions. LC50 values were calculated using either the graphical,

probit, Spearman-Karber, or trimmed Spearman-Karber methods according to U.S.

Environmental Protection Agency established criteria.

Limit Test:

Test Results - Acute Toxicity To Aquatic Vertebrates

NOEC Exposure

Duration:

NOEC:

LOEC Exposure

Duration:

LOEC:

NOELR Exposure

Duration:

NOELR:

LOELR Exposure

Duration:

LOELR:
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LC/EC Mean Value : Exposure

Duration

Exposure

Units

LC/EC % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Effect

Observed

Basis for

Concentration

96 Hours LC 50 % = 38 65 mg/L Mortality

Results Remarks: Measured high concentrations of MeBT were between 95.5 and 99.5% of nominal

concentrations at a confidence interval of 95%. Lower test concentrations were

calculated as a proportion of the 100% measured concentration. The LC50s were

calculated based on measured 100% concentration.

Reliability/Data Quality - Acute Toxicity To Aquatic Vertebrates

Reliability: 2 - Valid With Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Vertebrates

Reference: Cornell, J.S. et al. 2000. Comparative measures of the toxicity of component

chemicals in aircraft deicing fluid. Environmental Chemistry, 19(6): 1465-1472.

 Acute Toxicity to Aquatic Vertebrates

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Vertebrates

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

Referred to as Tolyltriazole-Cobratec TT-100, Sample 1090. Supplied as tan granules.

Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Measured

Method - Acute Toxicity To Aquatic Vertebrates

Year Study

Performed:

1974

Method/Guideline

Followed:

Other

Other

Method/Guideline:

Fish - Pesticide Acute Toxicity Test guideline; January 20, 1971.
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Deviations from

Method/Guideline:

Species: Lepomis macrochirus

GLP: No Data

Analytical

Monitoring:

Yes

Test Type:

Test Vessel:

Water Media Type:

Test

Concentrations:

Nominal

Nominal and

Measured

Concentrations:

3.16, 3.55, 3.98, 4.47 ppm

Exposure Period: 96 Hours

Vehicle Used: Yes

Vehicle Name: Acetone

Vehicle Amount

and Units:

Alkalinity:

Dissolved Oxygen:

pH Value:

Test Temperature

and Units:

Photo

(Light/Dark):

Salinity:

TOC:

Water Hardness:

Method/Guideline

Test Conditions

Remarks:

Average length of fish was 42 mm; average weight of fish was 0.8 gm.

Fish were held in a plastic wading pool prior to bioassay. The water temperature of

the pool was 72 ±2 F. Fish were held for 10 days before bioassay.

Test Medium -

Container: 5 gallon glass water bucket lined with a clear polyethylene bag which was

discarded and replaced after each use.

Water Quantity: 16 liters per test vessel.

Water Quality: Standard reconstituted distilled water.

Water Temperature: 72 ±2 F.

Test Chemicals - 

Candidate: Tolyltriazole-Cobratec TT-100, Sample 1090, (technical grade chemical)

supplied by Sherwin-Williams Chemicals.
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Solvent, acetone (0.3 mL/liter of test water)

Reference: p, p'DDT, 74%

Solvent, acetone (0.3 mL/liter of test water)

Determinations of Lethal Concentrations - 

Screening tests using 2 fish per dose level were conducted to locate the general level

of toxicity of Tolyltriazole-Cobratec TT-100, Sample 1090, and 74% p, p'DDT to the

particular lot of fish.

From the results of the screening tests, the following concentrations were chosen as

the doses to use in the bioassay:

Cobratec TT-100 (ppm): 3.16, 3.55, 3.98, 4.47

74% p, p'DDT (ppm): 0.016, 0.013, 0.010, 0.008

The test lasted for 96 hours with mortality readings taken every 24 hours.

Twenty fish were exposed to each of the above concentrations. The fish were loaded

ten to a test vessel (0.5 gm fish/liter of water). Twenty fish were untreated and

served as controls.

Reaction of fish: 

1. Controls - there were no deaths and no signs of intoxication.

2. Tolyltriazole - Cobratec TT-100, Sample 1090 - the major symptom of toxicity was

loss of equilibrium prior to death.

3. 74%, p'DDT - death was usually preceded by loss of equilibrium.

Litchfield and Wilcoxon's method was used to compute the confidence limits for the

LC1, LC50, and LC99 (at 24, 48 and 96 hours).

Limit Test:

Test Results - Acute Toxicity To Aquatic Vertebrates

NOEC Exposure

Duration:

NOEC:

LOEC Exposure

Duration:

LOEC:

NOELR Exposure

Duration:

NOELR:

LOELR Exposure

Duration:

LOELR:

LC/EC Mean Value : Exposure

Duration

Exposure

Units

LC/EC % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Effect

Observed

Basis for

Concentration

96 Hours LC 50 % = 3.47 ppm Mortality

48 Hours LC 50 % = 3.55 ppm Mortality

24 Hours LC 50 % = 4.02 ppm Mortality
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Results Remarks: The positive control responded appropriately. It is concluded that Cobratec TT-100 is

less toxic to the bluegill sunfish than is 74% p, p'DDT.

Reliability/Data Quality - Acute Toxicity To Aquatic Vertebrates

Reliability: 2 - Valid With Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Vertebrates

Reference: TSCATS submission (OTS0521121, Doc ID 86-890000613, received 11/06/74): Huntingdon

Research Center, Sherwin Williams Company. 1974. Initial submission: The Acute

Toxicity of Tolyltriazole-Cobratec TT-100 Sample 1090 to the Bluegill Sunfish (Lepomis

macrochirus) with Attachments and Cover Letter Dated 061289.

 Acute Toxicity to Aquatic Vertebrates

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Vertebrates

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

off-white particles

Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Nominal

Method - Acute Toxicity To Aquatic Vertebrates

Year Study

Performed:

1992

Method/Guideline

Followed:

OECD 203

Other

Method/Guideline:

Deviations from

Method/Guideline:

Species: Pimephales promelas

GLP: Yes

Analytical

Monitoring:

No Data

Test Type: Semi-Static

Test Vessel:
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Water Media Type: Freshwater

Test

Concentrations:

Nominal

Nominal and

Measured

Concentrations:

13, 22, 36, 60 and 100 mg/L

Exposure Period: 96 Hours

Vehicle Used:

Vehicle Name:

Vehicle Amount

and Units:

Alkalinity: 184-188 mg/L CaCO3

Dissolved Oxygen: 8.4 to 9.2 mg/L 

pH Value: Value/Lower Range: = 7.8 Upper Range: 8.5

Test Temperature

and Units:

Value/Lower Range: 21 Upper Range: 22.9 °C

Photo

(Light/Dark):

Salinity:

TOC:

Water Hardness: 144 mg/L

Method/Guideline

Test Conditions

Remarks:

Fathead minnows were exposed to a geometric series of five test concentrations and a

negative (well water) control under static-renewal test conditions for 96 hours. Two

replicate test chambers were maintained in each treatment and control group, with 10

fish in each test chamber. Nominal test concentrations were selected in consultation

with the sponsor, and were based upon results of range finding tests. Nominal

concentrations selected were 13, 22, 36, 60 and 100 mg/L.

Immediately prior to initiating the test, fish were impartially removed from holding

tanks in groups of two and distributed among the test chambers until each contained 10

fish. At 48 hours, new test solutions were prepared and fish were transferred to the

new solutions. Observations of mortality and other clinical signs of toxicity were

made at approximately 24, 48, 72, and 96 hours after test initiation. Cumulative

percent mortality values in each treatment group were used to calculate LC50 values

and 95% confidence limits at 24, 48, 72 and 96 hours. The no mortality concentration

was determined by visually examining the mortality data.

Limit Test: Yes

Test Results - Acute Toxicity To Aquatic Vertebrates

NOEC Exposure

Duration:

NOEC:

LOEC Exposure

Duration:

LOEC:

NOELR Exposure
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Duration:

NOELR:

LOELR Exposure

Duration:

LOELR:

LC/EC Mean Value : Exposure

Duration

Exposure

Units

LC/EC % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Effect

Observed

Basis for

Concentration

96 Hours LC 50 % = 43 mg/L Mortality

96 Hours LC 0 % = 22 mg/L Mortality

Results Remarks: Water temperatures taken daily with a hand-held thermometer ranged from 21.0 to 22.9

C. Dissolved oxygen and pH readings ranged from 8.4 to 9.2 mg/L and 7.8 to 8.5,

respectively. Dissolved oxygen concentrations exceeded 60% of saturation throughout

the test. Conductivity, alkalinity, and hardness measured at test initiation (0

hours) in the Negative Control Replicate A were 295 umhos/cm, 188 mg/L as CaCO3, and

144 mg/L as CaCO3, respectively. At test termination (96 hours) they were 325

umhos/cm, 184 mg/L as CaCO3, and 144 mg/L as CaCO3, respectively, in the Negative

Control Replicate A.

After the test substance was added to the vortex, a clear colorless solution as formed

in the test chambers. White particulate matter was noted on the bottom of one tank

(Replicate A) of the 36 mg/L treatment group, approximately 24 hours after test

initiation. The particulate matter remained until test termination.

Fish in the negative control, with the exception of one fish which appeared lethargic

at 48 hours, appeared healthy and normal throughout the test. The 13 and 22 mg/L

groups appeared normal and healthy throughout the test with no signs of disease or

stress. Overt signs of toxicity observed at the 36 mg/L concentration, beginning

approximately 24 hours after test initiation, included lethargy, loss of equilibrium,

and erratic swimming. At the 36 mg/L concentration, 15% mortality was observed at 24

hours with no further mortality occurring by 96 hours. At the 60 and 100 mg/L

concentrations, 100% mortality was observed at 24 hours. 

Reliability/Data Quality - Acute Toxicity To Aquatic Vertebrates

Reliability: 1 - Valid Without Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Vertebrates

Reference: TSCATS submission (OTS0557287, Doc ID 86940000877S, received 06/02/94): Wildlife

International Ltd, CBI. 1993. Initial submission: A 96-hour Static Renewal Toxicity

Test with the Fathead Minnow (Pimephales promelas) of Tolyltriazole - 1HBenzotriazole, Methyl with Cover Letter Dated 04/29/94 (SANITIZED).
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 Acute Toxicity to Aquatic Vertebrates

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Vertebrates

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

Referred to as Tolyltriazole-Cobratec TT-100, Sample 1090. Supplied as tan granules.

Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Measured

Method - Acute Toxicity To Aquatic Vertebrates

Year Study

Performed:

1975

Method/Guideline

Followed:

Other

Other

Method/Guideline:

Fish - Pesticide Acute Toxicity Test guideline; January 20, 1971

Deviations from

Method/Guideline:

Species: Lepomis macrochirus

GLP: No Data

Analytical

Monitoring:

Yes

Test Type:

Test Vessel:

Water Media Type: Freshwater

Test

Concentrations:

Nominal

Nominal and

Measured

Concentrations:

25.1, 28.2, 31.6, 35.5, 39.8 and 44.7 ppm

Exposure Period: 96 Hours

Vehicle Used: Yes

Vehicle Name: Acetone

Vehicle Amount

and Units:

Alkalinity:

Dissolved Oxygen:
High Production Volume Information System (HPVIS) | High Production Volume (HPV) Challenge | OPPT | US EPA

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pH Value:

Test Temperature

and Units:

Photo

(Light/Dark):

Salinity:

TOC:

Water Hardness:

Method/Guideline

Test Conditions

Remarks:

Average length of fish was 38 mm; average weight of fish was 0.9 gm.

Fish were held in a plastic wading pool prior to bioassay. The water temperature of

the pool was 72 ±2 F. Fish were held for 10 days before bioassay.

Test Medium -

Container: 5 gallon glass water bucket

Water Quantity: 18 liters per test vessel.

Water Quality: Standard reconstituted distilled water.

Water Temperature: 72 ±2 F.

Test Chemicals - 

Candidate: Tolyltriazole-Cobratec TT-100, Sample 1090, (technical grade chemical)

supplied by Sherwin-Williams Chemicals.

Solvent, acetone < 0.3 mL/liter of test water

Determinations of Lethal Concentrations - 

Screening tests using 2 fish per dose level were conducted to locate the general level

of toxicity of Tolyltriazole-Cobratec TT-100, Sample 1090 to the particular lot of

fish.

From the results of the screening tests, the following concentrations were chosen as

the doses to use in the bioassay:

Cobratec TT-100 (ppm): 25.1, 28.2, 31.6, 35.5, 39.8 and 44.7 ppm

The test lasted for 96 hours with mortality readings taken every 24 hours.

Twenty fish were exposed to each of the above concentrations. The fish were loaded

ten to a test vessel (0.5 gm fish/liter of water). Twenty fish were untreated and

served as controls.

Reaction of fish: 

1. Controls - there were no deaths and no signs of intoxication.

2. Tolyltriazole - Cobratec TT-100, Sample 1090 - the major symptom of toxicity was

loss of equilibrium, sporadic swimming, and lying on their side prior to death

Litchfield and Wilcoxon's method was used to compute the confidence limits at 96

hours.

Limit Test:

Test Results - Acute Toxicity To Aquatic Vertebrates

NOEC Exposure

Duration:

NOEC:

LOEC Exposure

Duration:

LOEC:
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NOELR Exposure

Duration:

NOELR:

LOELR Exposure

Duration:

LOELR:

LC/EC Mean Value : Exposure

Duration

Exposure

Units

LC/EC % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Effect

Observed

Basis for

Concentration

96 Hours LC 50 % = 31 ppm Mortality

Results Remarks: Cobratec TT-100 No. Dead/No. Dosed

35.5 20/20

31.6 10/20

28.2 2/20

25.1 1/20

Reliability/Data Quality - Acute Toxicity To Aquatic Vertebrates

Reliability: 2 - Valid With Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Vertebrates

Reference: TSCATS submission (OTS0521122, Doc ID 86-890000614, received 09/29/75): Huntingdon

Research Center, Sherwin Williams Company. 1975. Initial submission: The Acute

Toxicity of Tolyltriazole-Cobratec TT-100 Sample 1090 to the Bluegill Sunfish (Lepomis

macrochirus) with Attachments and Cover Letter Dated 061289.

 Acute Toxicity to Aquatic Vertebrates

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Vertebrates

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

Referred to as Tolyltriazole-Cobratec TT-100, Sample 1090. Supplied as tan granules.
High Production Volume Information System (HPVIS) | High Production Volume (HPV) Challenge | OPPT | US EPA

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Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Measured

Method - Acute Toxicity To Aquatic Vertebrates

Year Study

Performed:

Method/Guideline

Followed:

Other

Other

Method/Guideline:

Fish - Pesticide Acute Toxicity Test guideline; January 20, 1971

Deviations from

Method/Guideline:

Species: Salmo sp.

GLP: No Data

Analytical

Monitoring:

Yes

Test Type:

Test Vessel:

Water Media Type: Freshwater

Test

Concentrations:

Nominal

Nominal and

Measured

Concentrations:

25.1, 28.2, 31.6, 35.5, 39.8 and 44.7 ppm

Exposure Period: 96 Hours

Vehicle Used: Yes

Vehicle Name: Acetone

Vehicle Amount

and Units:

Alkalinity:

Dissolved Oxygen:

pH Value:

Test Temperature

and Units:

Photo

(Light/Dark):

Salinity:

TOC:

Water Hardness:

Method/Guideline Average length of fish was 43 mm; average weight of fish was 0.35 gm.
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Test Conditions

Remarks: Fish were held in an indoor tank prior to bioassay. The water temperature of the

pool was 52 ±2 F. Fish were held for 10 days before bioassay.

Test Medium -

Container: 5 gallon glass water bucket

Water Quantity: 14 liters per test vessel.

Water Quality: Standard reconstituted distilled water.

Water Temperature: 52 ±2 F.

Test Chemicals - 

Candidate: Tolyltriazole-Cobratec TT-100, Sample 1090, (technical grade chemical)

supplied by Sherwin-Williams Chemicals.

Solvent, acetone (0.3 mL/liter of test water)

Reference: p, p'DDT, 74%

Solvent, acetone (0/.3 mL/liter of test water)

Determinations of Lethal Concentrations - 

Screening tests using 5 fish per dose level were conducted to locate the general level

of toxicity of Tolyltriazole-Cobratec TT-100, Sample 1090, and 74% p, p'DDT to the

particular lot of fish.

From the results of the screening tests, the following concentrations were chosen as

the doses to use in the bioassay:

Cobratec TT-100 (ppm): 10.01, 14.10, 20.00, 28.20

74% p, p'DDT (ppm): 0.0200, 0.0251, 0.0316, 0.0308

The test lasted for 96 hours with mortality readings taken every 24 hours.

Twenty fish were exposed to each of the above concentrations. The fish were loaded

ten to a test vessel (0.5 gm fish/liter of water). Twenty fish were untreated and

served as controls.

Reaction of fish: 

1. Controls - there were no deaths and no signs of intoxication.

2. Tolyltriazole - Cobratec TT-100, Sample 1090 - the major symptom of toxicity was

loss of equilibrium prior to death.

3. 74% p, p'DDT - death was usually preceded by loss of equilibrium.

Litchfield and Wilcoxon's method was used to compute the confidence limits at 96

hours.

Limit Test:

Test Results - Acute Toxicity To Aquatic Vertebrates

NOEC Exposure

Duration:

NOEC:

LOEC Exposure

Duration:

LOEC:

NOELR Exposure

Duration:

NOELR:

LOELR Exposure

Duration:

LOELR:
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LC/EC Mean Value : Exposure

Duration

Exposure

Units

LC/EC % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Effect

Observed

Basis for

Concentration

96 Hours LC 50 % = 21.4 ppm Mortality

Results Remarks:

Reliability/Data Quality - Acute Toxicity To Aquatic Vertebrates

Reliability: 2 - Valid With Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Vertebrates

Reference: TSCATS submission (OTS0521123, Doc ID 86-890000615, received 06/13/89): Huntingdon

Research Center, Sherwin Williams Company. 1974. Initial submission: The Acute

Toxicity of Tolyltriazole-Cobratec TT-100 Sample 1090 to the Rainbow Trout (Salmo

gairdneri) with Attachments and Cover Letter Dated 061289.

 Acute Toxicity to Aquatic Vertebrates

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Vertebrates

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

50% solution in water of the sodium salt of tolyltriazole. Referred to as Cobratec

TT 50S. Described as dark amber, water-soluble liquid.

Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Nominal

Method - Acute Toxicity To Aquatic Vertebrates

Year Study

Performed:

1979

Method/Guideline

Followed:

Other

Other

Method/Guideline:

Committee on Methods for Toxicity Tests with Aquatic Organisms (1975)
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Deviations from

Method/Guideline:

Species: Lepomis macrochirus

GLP: No Data

Analytical

Monitoring:

No

Test Type: Static

Test Vessel:

Water Media Type: Freshwater

Test

Concentrations:

Nominal

Nominal and

Measured

Concentrations:

~ 0, 16.0, 28.0, 50.0, 90.0 and 160.0 mg sodium tolyltriazole/L

(0, 32.0, 56.0, 100.0, 180.0 and 320.0 mg Cobratec TT 50S / L)

Exposure Period: 96 Hours

Vehicle Used:

Vehicle Name:

Vehicle Amount

and Units:

Alkalinity: 30 mg/L

Dissolved Oxygen: 5.2 to 9.3 mg/L

pH Value: Value/Lower Range: = 7.18 Upper Range: 9.4

Test Temperature

and Units:

Photo

(Light/Dark):

Salinity:

TOC:

Water Hardness: 42 mg/L

Method/Guideline

Test Conditions

Remarks:

Dilution water for the test was obtained from a well on the Tarrytown site, treated

with a Continental Model 3020 Reverse Osmosis Water System, and deionized; then,

following procedures of Marking and Dawson (1973), it was reconstituted to a pH of

7.46, total hardness of 42 mg/L as CaCO3, total alkalinity of 30 mg/L as CaCO3 and

specific conductance of 155 μmhos/cm. Hardness and alkalinity were determined

according to standard analytical procedures (American Public Healthy Association,

1976), pH with an ORION pH Meter, conductivity with a YSI Conductivity Bridge and

dissolved oxygen with a YSI Oxygen Meter.

Test practices followed these recommended by the Committee on Methods for Toxicity

Tests with Aquatic Organisms (1975), except that replicate concentrations were not

used. Samples of Cobratec TT 50S were weighed to a precision of 0.1 mg and added

directly to the test containers. Test vessels were 19.6 liter, chemically clean

glass jars, each containing 15 liters of soft reconstituted dilution water and

immersed in a constant temperature recirculating water bath. 

Testing was begun by adding measured quantities of test material to the water in
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five test vessels, and mixing thoroughly, to yield nominal concentrations of 32.0,

56.0, 100.0, 180.0 and 320.0 mg/L. A sixth vessel, as control, contained 100%

dilution water. Ten fish were introduced at random into each of the test and

control vessels; these were approximately 4 months old with a mean (10 organisms)

length of 37 mm and mean weight of 0.50 grams. Biological loading was 0.33 g/L.

At the beginning of the test, and every 24 hours thereafter, dissolved oxygen and

pH of the control and each test concentration, and temperature of the water bath,

were determined. Mortalities among the test fish, and any observable abnormal

behavioral responses, were noted and recorded every 24 hours. The no effect level

was determined, by observation, at 96 hours.

The concentration of test material lethal to 50% of the test population, and 95%

confidence limits, were determined for the 72- and 96-hour exposure periods by the

Spearman-Karber Estimator. LC50s could not be calculated for the 24 and 48 hour

exposure periods due to the lack of sufficient mortalities. LC50 calculations are

based on nominal concentrations of the test material in soft reconstituted water.

Limit Test:

Test Results - Acute Toxicity To Aquatic Vertebrates

NOEC Exposure

Duration:

NOEC:

LOEC Exposure

Duration:

LOEC:

NOELR Exposure

Duration:

NOELR:

LOELR Exposure

Duration:

LOELR:

LC/EC Mean Value : Exposure

Duration

Exposure

Units

LC/EC % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Effect

Observed

Basis for

Concentration

Results Remarks: The 96-hour LC50 was ~ 95.6 mg sodium tolyltriazole/L (191.2 mg Cobratec TT 50S /

L). The 96-hour NOEC was ~ 28.0 mg sodium tolyltriazole/L (56.0 mg Cobratec TT 50S

/L).

pH ranged from 7.18 to 9.4. Dissolved oxygen ranged from 5.2 to 9.3 mg/L.

Reliability/Data Quality - Acute Toxicity To Aquatic Vertebrates

Reliability: 1 - Valid Without Restrictions

Reliability

Remarks:
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Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Vertebrates

Reference: TSCATS submission (OTS0521126, Doc ID 86-890000618, received 06/13/89): Union

Carbide Corp, Sherwin Williams Company. 1979. Initial submission: The Acute

Toxicity of Cobratec TT 50S to the Bluegill Sunfish (Lepomis macrochirus) Rafinesque

with Cover Letter Dated 061289.

 Acute Toxicity to Aquatic Vertebrates

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Vertebrates

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

50% solution in water of the sodium salt of tolyltriazole. Referred to as Cobratec

TT 50S. Described as dark amber, water soluble liquid.

Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Nominal

Method - Acute Toxicity To Aquatic Vertebrates

Year Study

Performed:

1979

Method/Guideline

Followed:

Other

Other

Method/Guideline:

Committee on Methods for Toxicity Tests with Aquatic Organisms (1975)

Deviations from

Method/Guideline:

Species: Salmo sp.

GLP: No Data

Analytical

Monitoring:

No

Test Type: Static

Test Vessel:

Water Media Type: Freshwater

Test

Concentrations:

Nominal

Nominal and

Measured

Concentrations:

~ 0, 2.8, 5.0, 9.0, 16.0 and 28.0 mg sodium tolyltriazole / L

(0, 5.6, 10.0, 18.0, 32.0 and 56.0 mg Cobratec TT 50S/L)

Exposure Period: 96 Hours
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Vehicle Used:

Vehicle Name:

Vehicle Amount

and Units:

Alkalinity: 28 mg/L

Dissolved Oxygen: 6.2 to 9.4 mg/L

pH Value: Value/Lower Range: = 7.03 Upper Range: 8.42

Test Temperature

and Units:

Photo

(Light/Dark):

Salinity:

TOC:

Water Hardness: 42 mg/L

Method/Guideline

Test Conditions

Remarks:

Dilution water for the test was obtained from a well on the Tarrytown site, treated

with a Continental Model 3020 Reverse Osmosis Water System, and deionized; then,

following procedures of Marking and Dawson (1973), it was reconstituted to a pH of

7.47, total hardness of 42 mg/L as CaCO3, total alkalinity of 28 mg/L as CaCO3 and

specific conductance of 180 umhos/cm. Hardness and alkalinity were determined

according to standard analytical procedures (American Public Healthy Association,

1976), pH with an ORION pH Meter, conductivity with a YSI Conductivity Bridge and

dissolved oxygen with a YSI Oxygen Meter.

Test practices followed these recommended by the Committee on Methods for Toxicity

Tests with Aquatic Organisms (1975), except that replicate concentrations were not

used. A fresh working stock solution of the test material in deionized water was

prepared by weight to a precision of 0.1 mg. Five concentrations were tested, with

a control. Test vessels were 19.6 liter, chemically clean glass jars, each

containing 15 liters of soft reconstituted dilution water and immersed in a

constant temperature recirculating water bath.

Testing was begun by adding measured volumes of stock solution to the water in five

test vessels, and mixing thoroughly, to yield nominal test material concentrations

of 5.6, 10.0, 18.0, 32.0 and 56.0 mg/L. A sixth vessel, as control, contained 100%

dilution water. Ten fish were introduced at random into each of the test and

control vessels; these were approximately 3 months old with a mean (10 organisms)

length of 54 mm and mean weight of 1.27 grams. Biological loading was 0.85 g/L.

At the beginning of the test, and every 24 hours thereafter, dissolved oxygen and

pH of the control and each test concentration, and temperature of the water bath,

were determined. Mortalities among the test fish, and any observable abnormal

behavioral responses were noted and recorded every 24 hours. The no effect level

was determined, by observation, at 96 hours. 

The concentration of test material lethal to 50% of the test population was

determined for the 24, 48, 72 and 96 hour exposure periods by the Spearman-Karber

Estimator. Confidence limits for the 48 hour exposure period were also

calculated. LC50 calculations are based on nominal concentrations of the test

material in soft reconstituted water.

Limit Test:

Test Results - Acute Toxicity To Aquatic Vertebrates

NOEC Exposure

Duration:

NOEC:
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LOEC Exposure

Duration:

LOEC:

NOELR Exposure

Duration:

NOELR:

LOELR Exposure

Duration:

LOELR:

LC/EC Mean Value : Exposure

Duration

Exposure

Units

LC/EC % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Effect

Observed

Basis for

Concentration

Results Remarks: The 96-hour LC50 was ~ 11.9 mg sodium tolyltriazole/L (23.7 mg Cobratec TT 50S

/L). The 96-NOEC was ~ 5.0 mg sodium tolyltriazole/L (10.0 mg Cobratec TT 50S /L).

pH ranged from 7.03 to 8.42. Dissolved oxygen ranged from 6.2 to 9.4 mg/L.

Reliability/Data Quality - Acute Toxicity To Aquatic Vertebrates

Reliability: 1 - Valid Without Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Vertebrates

Reference: TSCATS submission (OTS0521128, Doc ID 86-890000620, received 06/13/89): Union

Carbide Corp, Sherwin Williams Company. 1979. Initial submission: The Acute

Toxicity of Cobratec TT50S to the Rainbow Trout (Salmo gairdneri) Richardson with

Cover Letter Dated 061289.

 Acute Toxicity to Aquatic Vertebrates

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Vertebrates

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Referred to as Tolyltriazole-Cobratec TT-100, Sample 1090. Supplied as tan granules.
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Comments:

Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Nominal

Method - Acute Toxicity To Aquatic Vertebrates

Year Study

Performed:

1974

Method/Guideline

Followed:

Other

Other

Method/Guideline:

Fish - Pesticide Acute Toxicity Test guideline. January 20, 1971

Deviations from

Method/Guideline:

Species: Pimephales promelas

GLP: No Data

Analytical

Monitoring:

Yes

Test Type:

Test Vessel:

Water Media Type:

Test

Concentrations:

Nominal

Nominal and

Measured

Concentrations:

17.80, 22.40, 28.20, 35.50 ppm

Exposure Period: 96 Hours

Vehicle Used: Yes

Vehicle Name: Acetone

Vehicle Amount

and Units:

Alkalinity:

Dissolved Oxygen:

pH Value:

Test Temperature

and Units:

Photo

(Light/Dark):

Salinity:

TOC:

Water Hardness:
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Method/Guideline

Test Conditions

Remarks:

Average length of fish was 65mm; average weight of fish was 1.2 gm.

Fish were held in a plastic wading pool prior to bioassay. The water temperature of

the pool was 68 ±2 deg F. Fish were held for 10 days before bioassay.

Test Medium -

Container: 5 gallon glass water bucket lined with a clear polyethylene bag which was

discarded and replaced after each use.

Water Quantity: 12 liters per test vessel.

Water Quality: Standard reconstituted distilled water.

Water Temperature: 68 ±2 deg F.

Test Chemicals - 

Candidate: Tolyltriazole-Cobratec TT-100, Sample 1090, (technical grade chemical)

supplied by Sherwin-Williams Chemicals.

Solvent, acetone (0.3 mL/liter of test water)

Reference: p, p'DDT, 74%

Solvent, acetone (0.3 mL/liter of test water)

Determinations of Lethal Concentrations - 

Screening tests using 2 fish per dose level were conducted to locate the general level

of toxicity of Tolyltriazole-Cobratec TT-100, Sample 1090, and 74% p, p'DDT to the

particular lot of fish.

From the results of the screening tests, the following concentrations were chosen as

the doses to use in the bioassay:

Cobratec TT-100 (ppm): 17.80, 22.40, 28.20, 35.50

74% p, p'DDT (ppm): 0.0563, 0.0709, 0.0892, 0.1120, 0.1410

The test lasted for 96 hours with mortality readings taken every 24 hours.

Twenty fish were exposed to each of the above concentrations. The fish were loaded

five to a test vessel (0.5 gm fish/liter of water). Twenty fish were untreated and

served as controls.

Reaction of fish: 

1. Controls - there were no deaths and no signs of intoxication.

2. Tolyltriazole - Cobratec TT-100, Sample 1090 - the major symptom of toxicity was

loss of equilibrium prior to death.

3. 74%, p'DDT - death was usually preceded by loss of equilibrium.

Litchfield and Wilcoxon's method was used to compute the confidence limits for the

LC1, LC50, and LC99 (at 24, 48 and 96 hours).

Limit Test:

Test Results - Acute Toxicity To Aquatic Vertebrates

NOEC Exposure

Duration:

NOEC:

LOEC Exposure

Duration:

LOEC:

NOELR Exposure

Duration:

NOELR:

LOELR Exposure

Duration:

LOELR:
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LC/EC Mean Value : Exposure

Duration

Exposure

Units

LC/EC % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Effect

Observed

Basis for

Concentration

96 Hours LC 50 % = 25.5 ppm Mortality

48 Hours LC 50 % = 29.8 ppm Mortality

24 Hours LC 50 % = 36.9 ppm Mortality

Results Remarks: The positive control responded appropriately. It is concluded that Cobratec TT-100 is

less toxic to the fathead minnow than is 74% p, p'DDT.

Reliability/Data Quality - Acute Toxicity To Aquatic Vertebrates

Reliability: 2 - Valid With Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Vertebrates

Reference: TSCATS submission (OTS0521120, Doc ID 86-890000612; received 11/06/74): Huntingdon

Research Center, Sherwin Williams Company. 1974. Initial submission: The Acute

Toxicity of Tolyltriazole-Cobratec TT-100 Sample 1090 to the Fathead Minnow

(Pimephales promelas) with Attachments and Cover Letter dated 061289.

 Acute Toxicity to Aquatic Invertebrates

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Invertebrates

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

5-Methyl-1H-Benzotriazole or 4-Methyl-1H-Benzotriazole (MeBT), CAS No. 29385-43-1

Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Measured

Method - Acute Toxicity To Aquatic Invertebrates

Year Study

Performed:

Method/Guideline

Followed:

Other

Other

Method/Guideline:

Methods for measuring the acute toxicity of effluents and receiving waters to

freshwater and marine microorganisms
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Deviations from

Method/Guideline:

Species/In Vitro

System:

Ceriodaphnia dubia

GLP: No Data

Analytical

Monitoring:

No Data

Test Type:

Test Vessel:

Water Media Type:

Test

Concentrations:

Both

Nominal and

Measured

Concentrations:

nominal: 152.1 and 183.8 mg/L; measured: Measured high concentrations of MeBT were

between 95.5 and 99.5% of nominal concentrations at a confidence interval of 95%.

Exposure Period: 48 Hours

Vehicle Used:

Vehicle Name:

Vehicle Amount

and Units:

Alkalinity:

Dissolved Oxygen:

pH Value:

Test Temperature

and Units:

Value/Lower Range: 20 °C

Photo

(Light/Dark):

16/8

Salinity:

TOC:

Water Hardness:

Method/Guideline

Test Conditions

Remarks:

All C. dubia used in the tests were <24 (±8) h old at test initiation. The toxicity

of MeBT to C. dubia was assessed in moderately hard water using acute tests according

to U.S. Environmental Protection Agency procedures. Mortality was the endpoint. The

nominal (predilution) test concentrations of MeBT were 152.1 and 183.8 mg/L. A

dilution water control was also included. Test chambers were 30-mL plastic cups. All

tests were conducted at 20 C with a light:dark cycle of 16:8 h. Testing was

conducted under static conditions for a 48 hour duration. The LC50 values were

calculated using either the graphical, probit, Spearman-Karber, or trimmed SpearmanKarber methods according to U.S. Environmental Protection Agency-established

criteria.

Limit Test:

Test Results - Acute Toxicity To Aquatic Invertebrates
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NOEC Exposure

Duration:

NOEC:

LOEC Exposure

Duration:

LOEC:

NOELR Exposure

Duration:

NOELR:

LOELR Exposure

Duration:

LOELR:

LC/EC Mean Value : Exposure

Duration

Exposure

Units

LC/EC % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Effect

Observed

Basis for

Concentration

48 Days EC 50 % = 102 108 mg/L

Results Remarks: Measured high concentrations of MeBT were between 95.5 and 99.5% of nominal

concentrations at a confidence interval of 95%. Lower test concentrations were

calculated as a proportion of the 100% measured concentration. The LC50s were

calculated based on measured 100% concentration.

Reliability/Data Quality - Acute Toxicity To Aquatic Invertebrates

Reliability: 2 - Valid With Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Invertebrates

Reference: Cornell, J.S. et al. 2000. Comparative measures of the toxicity of component

chemicals in aircraft deicing fluid. Environmental Chemistry, 19(6): 1465-1472.

 Acute Toxicity to Aquatic Invertebrates

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Invertebrates

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methyl-
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Test Substance

Purity/Composition

and Other Test

Substance

Comments:

Referred to as RM 27513 (EnviroSystems Sample Number 2694E). Delivered on November 15,

1990. RM 27513 was supplied by 3M Company, 935 Bush Avenue, St. Paul, Minnesota. 

Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Nominal

Method - Acute Toxicity To Aquatic Invertebrates

Year Study

Performed:

1991

Method/Guideline

Followed:

Other

Other

Method/Guideline:

based on procedures of the OECD (1984)

Deviations from

Method/Guideline:

Species/In Vitro

System:

Daphnia magna

GLP: Yes

Analytical

Monitoring:

No

Test Type: Static

Test Vessel:

Water Media Type: Freshwater

Test

Concentrations:

Nominal

Nominal and

Measured

Concentrations:

0 mg/L (control), 15 mg/L, 25 mg/L, 40 mg/L, 60 mg/L, and 100 mg/L

Exposure Period: 48 Hours

Vehicle Used: No

Vehicle Name:

Vehicle Amount

and Units:

Alkalinity:

Dissolved Oxygen: 8.3 to 9.7 mg/L

pH Value: Value/Lower Range: = 7.7 Upper Range: 8.4

Test Temperature

and Units:

Value/Lower Range: 19 Upper Range: 20.7 °C

Photo

(Light/Dark):

16/8

Salinity:
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TOC:

Water Hardness:

Method/Guideline

Test Conditions

Remarks:

Dilution water - Water used for acclimation of test organisms and for all toxicity

testing was collected from wells at EnviroSystems in Hampton, New Hampshire. Water was

stored in 500-gallon polyethylene tanks where it was aerated. 

Test organism - Juvenile daphnids employed as test organisms were from a single source

and were identified using an appropriate taxonomic key. Daphnids used in the test were

produced from an in-house culture and were less than 24 hours old at the start of the

test. Control daphnids were weighed at the conclusion of the toxicity test. Prior to

testing, daphnids were maintained in 100% dilution water under static conditions in 4

liter glass jars. During acclimation daphnids were not treated for disease and they were

free of apparent sickness, injuries, and abnormalities at the beginning of the

test. During the acclimation period 24 hours prior to the test initiation the

temperature was 19.0 - 19.8 C. Daphnids were fed yeast, trout chow, and/or the

freshwater alga Selenastrum capricornutum once or twice daily before the test.

Toxicity testing - A screening test with the test substance was conducted during November

27 to 29, 1990. Nominal concentrations of test substance were: 0.1, 1.0, 10, 100, and

1000 mg/L. After 48 hours of exposure there was 0% survival at 1000 mg/L, 75% survival

at 100 mg/L, and 100% survival at all other tested concentrations.

The definitive toxicity test was performed during December 12 to 14, 1990, according to

EnviroSystems Test Protocol 90169-3 (Static, Acute Toxicity Test With RM 27513 And The

Daphnid, Daphnia magna), which was signed by the Study Director on November 16, 1990. It

is based on procedures of the OECD (1984).

The test was conducted at a target temperature of 20 ± 1 C with five concentrations of

test substance and a dilution water control. A 1000 mg/L stock solution was prepared for

combining 1.0 g of test substance with dilution water and adjusting the final

concentrations to 1000 ml without the use of a solvent. Nominal concentrations of the

test material were: 0 mg/L (control), 15 mg/L, 25 mg/L, 40 mg/L, 60 mg/L, and 100 mg/L RM

27513.

Twenty daphnids were randomly and equally distributed among four replicates of each

treatment. The test was performed in 250 ml glass beakers that contained 200 ml of test

solution (water depth was approximately 6 cm). Test vessels were randomly arranged in an

incubator during the 48 hour test (a random numbers table was used to select the location

of each vessel). A 16 hour light and 8 hour dark photoperiod was automatically

maintained with cool-white fluorescent lights that provided a light intensity of 30 uEs1m-2. Aeration was not required to maintain dissolved oxygen concentrations above

acceptable levels. Daphnids were not fed during the test.

The number of surviving organisms and the occurrence of sublethal effects and

immobilization (inability to swim within 15 seconds after gentle agitation) or other

sublethal effects (loss of equilibrium, erratic swimming, loss of reflex, excitability,

discoloration, or change in behavior) were determined visually and recorded initially

after 24 and 48 hours. Dead test organisms were removed when first observed. Dissolved

oxygen, pH, conductivity and temperature were measured and recorded daily in each test

chamber that contained live animals.

Results of the toxicity test were interpreted by standard statistical techniques (Stephan,

1983). Nominal concentrations of test substance the number of daphnids killed or

immobilized were used to calculate the EC50 value.

Limit Test:

Test Results - Acute Toxicity To Aquatic Invertebrates

NOEC Exposure

Duration:

NOEC:

LOEC Exposure

Duration:

LOEC:

NOELR Exposure
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Duration:

NOELR:

LOELR Exposure

Duration:

LOELR:

LC/EC Mean Value : Exposure

Duration

Exposure

Units

LC/EC % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Effect

Observed

Basis for

Concentration

48 Hours EC 50 % = 19 mg/L Immobilization Nominal

24 Minutes EC 50 % = 34 mg/L Immobilization Nominal

Results Remarks: Test vessels containing RM 27513 appeared clear initially and remained free of insoluble

material throughout the test. Ninety-five percent survival occurred in the control

exposure. Control daphnids had an average wet weight (blotted dry) of 0.001 g at the end

of the test. Loading rate during the toxicity test was approximately 0.02 g/L. Exposure

of daphnids to the reference toxicant sodium dodecyl sulfate resulted in a 48 hour median

lethal concentration (LC50) of 13.8 mg/L.

pH ranged from 7.7 to 8.4 throughout testing. Conductivity ranged from 12000 to 13000

umho/cm. Temperature ranged from 19.0 to 20.7 C. Dissolved oxygen ranged from 8.3 to

9.7 mg/L.

Reliability/Data Quality - Acute Toxicity To Aquatic Invertebrates

Reliability: 2 - Valid With Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Invertebrates

Reference: TSCATS submission (OTS0533466, Doc ID 86-920000103S, received 10/31/91): Resource

Analysts, Inc., 3M Company. 1991. Initial submission: Environmental Effects of

Tolyltriazole with Attachments and Cover Letter 102891 (SANITIZED). pp. 75-91.

 Acute Toxicity to Aquatic Plants

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity To Aquatic Plants

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Referred to as RM 27513 (EnviroSystems Sample Number 2694E). Delivered on

November 15, 1990. RM 27513 (white granules) was supplied by 3M Company, 935 Bush
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Substance

Comments:

Avenue, St. Paul, Minnesota. 

Sponsored

Chemical Result

Type:

Measured

Test Substance

Result Type:

Nominal

Method - Acute Toxicity To Aquatic Plants

Year Study

Performed:

1991

Method/Guideline

Followed:

Other

Other

Method/Guideline:

Based on procedures of the OECD (1984)

Deviations from

Method/Guideline:

Species: Pseudokirchnerella subcapitata

GLP: No Data

Analytical

Monitoring:

No

Test Type: Static

Test Vessel: Closed

Water Media Type: Freshwater

Test

Concentrations:

Nominal

Nominal and

Measured

Concentrations:

0 mg/L (control), 62.5 mg/L, 125 mg/L, 250 mg/L, 500 mg/L, and 1000 mg/L

Exposure Period: 96 Hours

Vehicle Used:

Vehicle Name:

Vehicle Amount

and Units:

Alkalinity:

Dissolved Oxygen:

pH Value: Value/Lower Range: = 6.7 Upper Range: 9.4

Test Temperature

and Units:

Value/Lower Range: 23.1 Upper Range: 23.5 °C

Photo

(Light/Dark):

24/0

Salinity:

TOC:
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Water Hardness:

Method/Guideline

Test Conditions

Remarks:

Dilution water - Water used for acclimation of test organisms and for all toxicity

testing was sterile enriched media (U.S. EPA, 1978) with a pH of approximately

8.0. 

Test organism - Algae used for the test was from a culture originally procured

from the Culture Collection of Algae at the University of Texas at Austin and

delivered to Envirosystems on April 12, 1990. The identity of the culture was

verified using an appropriate taxonomic key. The culture was transferred to

sterile enriched media identical to media used for this test and maintained at

test conditions.

Toxicity testing - The definitive toxicity test was performed during April 30 to

May 4, 1991. It is based on procedures of the OECD (1984). The test was

conducted at a target temperature of 22 ± 2 C with five concentrations of test

substance and a dilution water control. A 1,000 mg/L stock solution was prepared

in dilution water. The stock solution was added directly to dilution water

contained in the test vessels without the use of a solvent. Nominal

concentrations of the test material were: 0 mg/L (control), 62.5 mg/L, 125 mg/L,

250 mg/L, 500 mg/L, and 1000 mg/L. 

Algae was randomly and equally distributed among three replicates of each

treatment at the rate of 10,000 cells/mL. The test was performed in 250 mL glass

Erlenmeyer flasks that contained 100 mL of test solution. The flasks were capped

with inverted glass beakers. Test vessels were randomly arranged on the rotary

shaker in an incubatory during the 96 hour test (a random numbers table was used

to select the location of each vessel). A 24 hour light and 0 hour dark

photoperiod was maintained with cool-white fluorescent lights that provided a

light intensity of 35 uEs-1m-2.

The number of algal cells/mL in each test vessel was determined visually every 24

hours by means of direct microscopic counts with a hemacytometer. The pH (Beckman

model pHI 12 meter) was determined in each test flask at the beginning and end of

the test, and temperature (ASTM mercury thermometer) was measured and recorded

daily in the incubator

Statistical methods - Results of the toxicity test were interpreted by standard

statistical techniques (Stephan, 1983). The binomial or moving average method was

used by the author to calculate EC50 values using nominal concentrations of test

substance. The NOEC was calculated using Dunnett's test, which includes a

parametric one-way analysis of variance (ANOVA). The average specific growth rate

was calculated as the natural log of the number of cells per ml at 96 hours minus

the natural log of the number of cells per ml at 0 hours divided by the exposure

period. The percent change from control is calculated by subtracting the sample

average specific growth rate from the control average specific growth rate,

dividing that value by the average specific growth rate in the control and

multiplying that value by 100. The percent change from control was used to

compute EC50 and NOEC values.

Limit Test:

Test Results - Acute Toxicity To Aquatic Plants

NOEC Exposure

Duration:

96 Hours

NOEC: = 250 mg/L Nominal

LOEC Exposure

Duration:

LOEC:

NOELR Exposure

Duration:

NOELR:

LOELR Exposure
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Duration:

LOELR:

Effect : Exposure

Duration

Exposure

Units

Type % Value

Description

Mean

Value

or

Lower

Mean

Value

Upper

Mean

Value

Units Basis

for

Effect

Basis for

Concentration

72 Hours EC 50 % < 62.5 mg/L Growth

Rate

96 Hours EC 50 % = 87 mg/L Growth

Rate

Results Remarks: Temperature varied from 23.1 to 23.5 C throughout testing. pH varied from 6.7 to

9.4 throughout testing.

Reliability/Data Quality - Acute Toxicity To Aquatic Plants

Reliability: 2 - Valid With Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity To Aquatic Plants

Reference: TSCATS submission (OTS0533466, Doc ID 86-920000103S, received 10/31/91): Resource

Analysts, Inc., 3M Company. 1991. Initial submission: Environmental Effects of

Tolyltriazole with Attachments and Cover Letter 102891 (SANITIZED). pp. 30-44

 Other

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

5-Methyl-1H-Benzotriazole or 4-Methyl-1H-Benzotriazole

(MeBT), CAS No. 29385-43-1

Other End Point Name: Microtox assay

Other End Point

Description:

Reference: Cornell, J.S. et al. 2000. Comparative measures of the

toxicity of component chemicals in aircraft deicing

fluid. Environmental Chemistry, 19(6): 1465-1472.

Description: Microtox studies measured the luminescence of the marine

bacterium Vibrio fischeri (formerly known as Photobacterium

phosphoreum) according to the basic method described by

Beckman Instruments. The nominal (predilution) test

concentration was 47.8 mg/L. Two replicates were
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tested. Five- and 15-min EC50s were calculated using methods

described by AZUR Environmental.

Results: 15-min EC50 = 7 mg/L

Measured high concentrations of MeBT were between 95.5 and

99.5% of nominal concentrations at a confidence interval of

95%. Lower test concentrations were calculated as a

proportion of the 100% measured concentration. The LC50s

were calculated based on measured 100% concentration.

 Acute Toxicity

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

Referred to as Cobratec TT 100 Sample No. 5459

Sponsored Chemical Result

Type:

Measured

Method - Acute Toxicity

Route of Administration: Oral

Type of Exposure: No Data

Species: Rat

Mammalian Strain: No Data

Gender: Both M/F

Number of Animals per

Dose:

10

Dose: 502, 632, 795 and 1001 mg/kg

Year Study Performed:

Method/Guideline

Followed:

No Data

GLP: No Data

Method/Guideline and

Test Condition Remarks:

Cobratec TT 100 Sample No. 5459 was administered orally in

graduated doses to 4 groups of 10 rats. Five males and five

females in each group. They were observed for 14 days. 

Test Results - Acute Toxicity

Concentration (LC/LD) : LC/LD % Value

Description

Value/Lower

Concentration

Upper

Concentration

Units

LD 50 % = 675 mg/kgbw
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Number of Deaths ( Male

):

Number of Deaths ( Female

):

Number of Deaths ( Total

):

23

Results Remarks: LD50 = 675 mg/kg (95% confidence limits were 535-851 mg/kg)

Mortalities were as follows:

502 mg/kg - 3 deaths on day 0

632 mg/kg - 1 death on day 0; 2 deaths on day 1

795 mg/kg - 6 deaths on day 0; 1 death on day 1

1001 mg/kg - 10 deaths on day 0

Conclusion: None stated

Reliability/Data Quality - Acute Toxicity

Reliability: 2 - Valid With Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity

Reference: TSCATS submission (Doc ID 86-890000263, received 06/05/89):

Huntingdon Research Center, Union Carbide

Corp. 1989. Initial submission: Health and Safety Studies

for 19 Chemicals with Cover Letter Date 060289. pp. 32-34.

 Acute Toxicity

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

Referred to as Cobratec RTT 100, Sample No. 3000. The

compound was a fine powder, off-white in color. The melting

point was reported to be 76  ̈¬C, with rapid decomposition

occurring at temperatures ¡Ã 300  ̈¬C.

Sponsored Chemical Result

Type:

Measured

Method - Acute Toxicity

Route of Administration: Inhalation
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Type of Exposure: Aerosol

Species: Rat

Mammalian Strain: Sprague-Dawley

Gender: Both M/F

Number of Animals per

Dose:

20

Concentration: 1.73 mg/L

Year Study Performed:

Method/Guideline

Followed:

Other

GLP: No Data

Method/Guideline and

Test Condition Remarks:

: One group of ten male and ten female Sprague-Dawley rats

(Charles River) was exposed for one hour to a concentration

of 1.73 +/- 0.67 mg/L of an aerosol of tolyltriazole. Vapor

phase compound was also monitored. Following the exposure,

all animals were maintained for an additional fourteen days;

with food and water ad libitum. Body weights were taken

prior to exposure and at seven and fourteen days postexposure. All animals were observed for signs of toxicity

during the exposure and daily during the post-exposure

observation period. All animals that died or were sacrificed

after fourteen days were necropsied. At this time, all

organs were observed for macroscopic abnormalities. The

following tissues were stored in 10% neutral buffered

formalin in the event that histopathology might be required:

lungs and trachea, heart, liver, kidneys, spleen, stomach,

intestines, gonads, whole head, abnormal tissue.

Animals were allowed 7 days to accommodate to the laboratory

conditions before being assigned to the study.

Aerosol generation system: Approximately one hundred grams of

tolyltriazole is placed in the round bottom three necked

flask and melted. The temperature of the flask and delivery

tube is maintained at 200  ̈¬C by the heating mantle, air

heater and heating tape. A controlled air flow of ten L/min

is provided to the vapor generating flask. The tolyltriazole

vapor-laden air is taken from the top of the generating

flask, diluted and cooled by admixture of a controlled flow

of clean dry air. This system provided a very fine white

aerosol. Chamber concentration was controlled by varying the

air flow rate through the generator.

Exposure concentrations were determined by sampling the

chamber atmosphere. Actual exposure concentrations were

determined by drawing (at 0.82 L/min) a known volume of

chamber air through a pre-weighted glass fiber filter in a

closed face filter holder. The sample time was three

minutes. The filter was then re-weighed and the chamber

concentration calculated.

Vapor phase compound was determined by sampling the chamber

atmosphere. The sampling rate was 0.82 L/min for six

minutes. The sample was drawn through two 25 mL fritted gas

bubblers in series, each containing 10 mL of 0.5N sodium

hydroxide. After sample collection the contents of each gas

bubbler was transferred to a 25 mL volumetric flask, brought
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to volume with additional 0.5N sodium hydroxide and mixed

well. The absorbance was determined at 274 nm using a UV

spectrophotometer and matched 1 cm. silica cells. The

tolyltriazole content was obtained by reference to a standard

curve.

Particle size analysis was also conducted during the

exposure. An Anderson cascade impactor was used for this

determination. Pre-weighted glass fiber filters, specially

cut to fit the impactor, were used for collecting the aerosol

on each stage. The filter discs were re-weighed immediately

after sampling. A particle size distribution graph was then

constructed from the weight of material collected on each

stage. The aerosol was found to have an Equivalent

Aerodynamic Diameter (EAD) of 0.35 µm with a geometric

standard deviation of 2.62. The sample flow rate was 1 CFM

for ten minutes.

Test Results - Acute Toxicity

Concentration (LC/LD) : LC/LD % Value

Description

Value/Lower

Concentration

Upper

Concentration

Units

LC 50 % > 1.73 mg/L

Number of Deaths ( Male

):

0

Number of Deaths ( Female

):

0

Number of Deaths ( Total

):

0

Results Remarks: During the exposure, the only pharmacologic or toxicologic

sign observed was a decrease in general activity. By 24

hours post-exposure, all animals were apparently normal and

remained so during the entire 14-day observation period. All

surviving animals appeared to gain weight normally throughout

the 14 day observation period.

Since there were no deaths during the exposure or 14-day

post-exposure observation period, all animals were necropsied

following the 14-day observation period. All organs were

observed for gross abnormalities. Respiratory tract

involvement, characterized by slight to moderate incidence of

hemorrhage or hepatization, was found in all animals. Some

incidence of pale liver and kidneys was also noted in males

and females.

Conclusion: None specified

Reliability/Data Quality - Acute Toxicity

Reliability: 1 - Valid Without Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:
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Reference - Acute Toxicity

Reference: TSCATS submission (OTS0521119, Doc ID 86-890000610, received

06/13/89): Huntingdon Research Center, Sherwin Williams

Co. 1976. Initial submission: Evaluation of the Acute

Inhalation Toxicity of Tolyltriazole (Final Report) with

Attachments and Cover Letter Dated 061289.

 Acute Toxicity

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

Substance was labeled Cobratec TT-100 (Tolyltriazole) Sample

No. 6022. The material was light tan granular in nature.

Sponsored Chemical Result

Type:

Measured

Method - Acute Toxicity

Route of Administration: Dermal

Type of Exposure: Occlusive

Species: Rabbit

Mammalian Strain: No Data

Gender: Female

Number of Animals per

Dose:

10

Dose: 2000 mg/kg

Year Study Performed:

Method/Guideline

Followed:

No Data

GLP: No Data

Method/Guideline and

Test Condition Remarks:

The material was applied to the clipped skin of 5 female

rabbits and to the clipped and abraded skin of 5 female

rabbits. The material was administered and allowed to remain

in contact with the skin for 24 hours. The trunk of each

animal was wrapped with a dam. After 24 hours the dams were

removed. The residue was washed off with water. All rabbits

were observed for mortality and toxic effects on the day of

application and daily thereafter until death occurred or the

14 day observation period ended.

Test Results - Acute Toxicity

Concentration (LC/LD) : LC/LD % Value

Description

Value/Lower

Concentration

Upper

Concentration

Units
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LD 50 % > 2000 mg/kgbw

Number of Deaths ( Male

):

Number of Deaths ( Female

):

0

Number of Deaths ( Total

):

0

Results Remarks: No mortalities were observed. Body weights remained stable

throughout the study (varying from 2.8 to 3.1 kg). Very

slight erythema (grade 1) was observed in three rabbits, each

from the abraded group; these effects subsided after the

third day of observation. No additional signs of skin

irritation were observed.

Conclusion: None stated

Reliability/Data Quality - Acute Toxicity

Reliability: 2 - Valid With Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Acute Toxicity

Reference: TSCATS submission (OTS0516797, Doc ID 86-890000263, received

06/05/89): Huntingdon Research Center, Union Carbide

Corp. 1989. Initial submission: Health and Safety Studies

for 19 Chemicals with Cover Letter Date 060289. pp. 10-15. 

 Acute Toxicity

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

Referred to as Olin 58734. Pinkish grey granular material

Sponsored Chemical Result

Type:

Measured

Method - Acute Toxicity

Route of Administration: Dermal
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Type of Exposure:

Species: Rabbit

Mammalian Strain: No Data

Gender: Both M/F

Number of Animals per

Dose:

4

Dose: 500, 1000, 2000 and 4000 mg/kg-bw

Year Study Performed:

Method/Guideline

Followed:

Other

GLP: No Data

Method/Guideline and

Test Condition Remarks:

Method/guideline followed: Method in Appraisal of the Safety

of Chemicals in Foods, Drugs and (illegible) Association of

Food and Drug Officials of the U.S. 1959

Animals - Four albino rabbits per four levels in weight range

of 2.2 to 3.6 kg depilated over the trunk.

Maintenance conditions - Trunk encased in sleeve with animal

restrained in stock during application. Thereafter individual

housing with food and water ad libitum.

Dosage - 24 hour continuous contact with the skin. Test

substance was administered as a 40% aqueous paste.

Observations - Body weight, appearance, behavior, skin

irritation and mortality over a 14-day period

Test Results - Acute Toxicity

Concentration (LC/LD) : LC/LD % Value

Description

Value/Lower

Concentration

Upper

Concentration

Units

LD 50 % > 4000 mg/kgbw

Number of Deaths ( Male

):

0

Number of Deaths ( Female

):

0

Number of Deaths ( Total

):

0

Results Remarks: No deaths occurred at any level in 14 days. The primary

irritation scores were 1.4, 1.5, 1.1 and 1.4, for the 0.5, 1,

2 and 4 g/kg levels, respectively. No other noteworthy

findings were observed.
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Conclusion: The acute dermal toxicity of the tested substance is greater

than 4 g/kg-bw and this product evokes no irritation.

Reliability/Data Quality - Acute Toxicity

Reliability: 3 - Not Valid

Reliability Remarks: Lack of study details; study is outdated

Key Study Sponsor

Indicator:

Reference - Acute Toxicity

Reference: TSCATS submission (OTS0520643, Doc ID 86-89000060A, received

06/13/89): Huntingdon Research Center, Sherwin Williams

Co. 1968. Initial submission: Summary of Mucuous Membrane

(Eye) Irritations in Rabbits Treated with Olin 58734 with

Attachment and Cover Letter Dated 061289.

 Acute Toxicity

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance - Acute Toxicity

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

Referred to as Olin 58734. Pinkish grey granular material

Sponsored Chemical Result

Type:

Measured

Method - Acute Toxicity

Route of Administration: Oral

Type of Exposure: Gavage

Species: Rat

Mammalian Strain: Other

Other Strain:

Gender: Both M/F

Number of Animals per

Dose:

5

Dose: 500, 1000, 2000, 4000 and 8000 mg/kg-bw

Year Study Performed:

Method/Guideline

Followed:

Other

GLP: No Data
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Method/Guideline and

Test Condition Remarks:

Method/guideline followed: Method in Appraisal of the Safety

of Chemicals in Food, Drugs, and Cosmetics, Association of

Food and Drug Officials of the U.S. 1959

Animals - Adult albino rats, FDRL strain, 5 rats per dose

level

Observations - Body weight, behavior, appearance, mortality,

gross changes at autopsy of rats that died and of

representative survivors sacrificed at 14 days

Test substance was administered intragastrically as a 25%

suspension in 0.5% (illegible).

Groups were comprised of three males and two females.

Test Results - Acute Toxicity

Concentration (LC/LD) : LC/LD % Value

Description

Value/Lower

Concentration

Upper

Concentration

Units

LD 50 % = 1830 mg/kgbw

Number of Deaths ( Male

):

Number of Deaths ( Female

):

Number of Deaths ( Total

):

14

Results Remarks: Deaths occurred in 3 hours to 5 minutes, following CNS

depression, shallow breathing, lacrimation, and

salivation. Liver congestion and pale stomach mucosae were

seen at autopsy.

One mortality was observed at 0.5 g/kg; 0 mortalities at 1.0

g/kg; 3 mortalities at 2.0 g/kg; 5 mortalities at 4.0 g/kg; 5

mortalities at 8.0 g/kg.

Conclusion: The acute oral LD50 for rats is 1.83 g per kg body weight;

confidence limits = 1.20 to 2.81 g per kg body weight.

Reliability/Data Quality - Acute Toxicity

Reliability: 3 - Not Valid

Reliability Remarks: Study is dated; lack information on test substance

Key Study Sponsor

Indicator:

Reference - Acute Toxicity

Reference: TSCATS submission (OTS0520643, Doc ID 86-89000060A, received

06/13/89): Huntingdon Research Center, Sherwin Williams

Co. 1968. Initial submission: Summary of Mucuous Membrane

(Eye) Irritations in Rabbits Treated with Olin 58734 with

Attachment and Cover Letter Dated 061289.
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 Genetic Toxicity in vitro

CAS Number : 29385-43-1

Sponsored Chemical

Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Genetic Toxicity in vitro

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

Referred to as TK 10 637/1

Sponsored Chemical

Result Type:

Measured

Method - Genetic Toxicity in vitro

Type of Study: Bacterial reverse mutation assay

Concentrations: 25, 75, 675, 2025 μg/0.1 mL without activation and 500, 1000,

2000, 

4000, 8000 μg/0.1 mL with activation.

Year Study

Performed:

1982

Method/Guideline

Followed:

Other

GLP:

Postive, Negative, and

Solvent

Control Substance(s):

negative control: acetone; Positive controls: 1) for strain TA98:

daunorubicin-HCl, 5 and 10 μg/0.1 mL phosphate buffer; 2) for

strain TA100: 4-nitroquinoline-N-oxide, 0.125 and 0.25 ug/0.1 mL

phosphate buffer; 3) for strain TA1535: N-methyl-N'-nitro-Nnitrosoguanidine, 3 and 5 μg/0.1 mL phosphate buffer; 4) for

strain TA1537: 9(5)aminoacridine hydrochloride monohydrate, 50 and

100 μg/0.1 mL DMSO; 5) for strain TA1538: 2-nitrofluorene, 5 and

10 μg/0.1 mL DMSO. 

Method/Guideline and

Test Condition

Remarks:

Method/guideline followed: Ames, B.N., F.D. Lee, and W.E. Durston

(1973), An Improved Bacterial Test System for the Detection and

Classification of Mutagens and Carcinogens. Proc. Natl. Acad. Sci.

USA 70, 782-786.

The substance was dissolved in acetone. Acetone alone 

was used for the negative controls. Each Petri dish contained: 1)

approx. 20 mL of minimum 

agar, 2) 0.1 mL of the solution of the test substance or the

vehicle and 0.1 mL of a bacterial 

culture in 2.0 mL of soft agar. The soft agar was composed of:

100 mL of 0.6% agar solution 

with 0.6% NaCl and 10 mL of a solution of l-histidine, 0.5 mM and

+biotin 0.5 mM. In the 

experiments in which the substance was metabolically activated,

0.5 mL of an activation mixture 

was added also. 1 mL activation mixture contained: 0.3 mL S9

fraction of liver from rats 

induced with Aroclor 1254 and 0.7 mL of a solution of cofactors. 

Positive control experiments were carried out simultaneously with
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the following substances: 1) for strain TA98: daunorubicin-HCl, 5

and 10 μg/0.1 mL phosphate buffer; 2) for strain TA100: 4-

nitroquinoline-N-oxide, 0.125 and 0.25 ug/0.1 mL phosphate buffer;

3) for strain TA1535: N-methyl-N'-nitro-N-nitrosoguanidine, 3 and

5 μg/0.1 mL phosphate buffer; 4) for strain TA1537:

9(5)aminoacridine hydrochloride monohydrate, 50 and 100 μg/0.1 mL

DMSO; 5) for strain TA1538: 2-nitrofluorene, 5 and 10 μg/0.1 mL

DMSO. The activation mixture was tested with strain TA1535 and

cyclophosphamide, 250 μg/0.1 mL phosphate buffer and with strain

TA98 and 2-aminoanthracene, 5 μg/0.1 mL DMSO.

In the experiments without and with the addition of microsomal

activation mixture three Petri dishes were prepared per strain and

per group (i.e. per concentration or per control group).

The plates were incubated for about 48 hours and 37 oC in

darkness.

When the colonies bad been counted, the arithmetic mean was

calculated. The test substance is generally considered to be nonmutagenic if the colony count in relation to the negative control

is not doubled at any concentration.

Test Results - Genetic Toxicity in vitro

Details on Cytogenetic

Assay:

Statistics:

Effect : Species Other

Species

Strain Other

Strain

Metabolic

Activation

Genotoxic

Effect

Conclusion

Bacteria S.

typhimurium

TA 98

With and

Without

Positive Positive

With

Metabolic

Activation

Bacteria S.

typhimurium

TA 100

With and

Without

Negative

Bacteria S.

typhimurium

TA 1535

With and

Without

Negative

Bacteria S.

typhimurium

TA 1537

With and

Without

Negative

Bacteria S.

typhimurium

TA 1538

With and

Without

Positive Positive

With

Metabolic

Activation

Results Remarks: In the experiments performed without microsomal activation, none

of the tested concentrations of TK 10 637/1 led to an increase in

the incidence of histidine-prototrophic mutants comparison with

the negative controls.

In the experiments with microsomal activation treatment with TK 10

637/1 led to an increase in the number of back-mutant colonies of

strains TA98 and TA1538 at the concentrations of 2000 ug/0.1 mL

and above.

Conclusion:

Reliability/Data Quality - Genetic Toxicity in vitro

Reliability: 1 - Valid Without Restrictions
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Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Genetic Toxicity in vitro

Reference: TSCATS submission (OTS0519041, Doc ID 86-890000272, received

06/07/89): CIBA GEIGY Corp. 1982. Initial submission:

Salmonella/Mammalian - Microsome Mutagenicity Test with TK 10

637/1 with Cover Letter Dated 060589.

 Genetic Toxicity in vitro

CAS Number : 29385-43-1

Sponsored Chemical

Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Genetic Toxicity in vitro

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

Code Identification - TT-100; Sample Number - X18BT7565 Cobratec

78669A; Description - Tan round crystals

Sponsored Chemical

Result Type:

Measured

Method - Genetic Toxicity in vitro

Type of Study: Bacterial reverse mutation assay

Concentrations: 0, 80 μg, 400 μg, 2 mg, 10 mg

Year Study

Performed:

Method/Guideline

Followed:

No Data

GLP: No Data

Postive, Negative, and

Solvent

Control Substance(s):

Positive control/strain/concentration (μg/plate)

Nonactivation

Sodium azide / TA1535 / 10

Quinacrine / TA1537 / 200

2-nitrofluorene / TA1538, TA98, TA100 / 10

Activation

2 amino anthracene / TA1535, TA1537 / 10

2 amino fluorine / TA1538, TA98, TA100 / 10

Method/Guideline and

Test Condition

Remarks:

Dimethylsulfoxide (DMSO) was used to prepare stock 

dilutions of test materials and positive control

chemicals. Approximately 108 cells from an overnight culture of

each indicator strain (TA1535, TA1537, TA1538, TA98, TA100) were

added to separate test tubes containing 2.0 mL of molten agar

supplemented with a trace of histidine and biotin. For

nonactivation tests, three dose levels of the extract were added

to the 

content of the appropriate tubes and poured over the surface of

selective agar plates. In activation tests, the extract
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concentrations were likewise added to the appropriate tubes with

cells. Then, just prior to pouring, an aliquot of the S-9 mix

containing the rat liver microsomes was added to each of the top

agar tubes which were then mixed and the contents poured over the

surface of a minimal agar plate and allowed to solidify. The

plates were incubated for 48 hours at 37 oC, and scored for the

number of colonies growing on each plate. 

Positive control/strain/concentration (μg/plate)

Nonactivation

Sodium azide / TA1535 / 10

Quinacrine / TA1537 / 200

2-nitrofluorene / TA1538, TA98, TA100 / 10

Activation

2 amino anthracene / TA1535, TA1537 / 10

2 amino fluorine / TA1538, TA98, TA100 / 10

Test Results - Genetic Toxicity in vitro

Details on Cytogenetic

Assay:

Statistics:

Effect : Species Other

Species

Strain Other

Strain

Metabolic

Activation

Genotoxic

Effect

Conclusion

Bacteria S.

typhimurium

TA 98

With and

Without

Equivocal Equivocal

With

Metabolic

Activation

Bacteria S.

typhimurium

TA 100

With and

Without

Negative Negative

Bacteria S.

typhimurium

TA 1535

With and

Without

Negative Negative

Bacteria S.

typhimurium

TA 1537

With and

Without

Negative Negative

Bacteria S.

typhimurium

TA 1538

With and

Without

Equivocal Equivocal

With

Metabolic

Activation

Results Remarks: The initial data obtained by the mutagenic bioassay of compound

TT-100 suggested a possible response with tester strains TA1538

and TA98. Confirmatory assays were carried out two additional

times. The results do not clearly indicate that TT-100 is a

frameshift mutagen. The revertant response elicited at the

highest concentration tested indicated a slight elevation of the

revertant rate but did not produce a response of at least an

excess of two times background. Higher concentrations of the test

compound indicated a toxic response. It appears that the TT-100

may possess some very weak mutagenic activity in this system when

enzymatically activated; however, confirmation of this activity is

negated by the toxicity of the compound in this system.

For the initial assay, in the absence of metabolic activation,

toxicity was observed at 10 mg in all strains tested. In the

presence of metabolic activation, toxicity was observed at 10 mg

in TA1535 and TA1537.

For the confirmation assay, toxicity was observed with metabolic

activation at 10 mg for TA1538 and TA98.
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Positive and negative controls responded appropriately.

Conclusion:

Reliability/Data Quality - Genetic Toxicity in vitro

Reliability: 2 - Valid With Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Genetic Toxicity in vitro

Reference: TSCATS submission (OTS0516797, Doc ID 86-890000263, received

06/05/89): Huntingdon Research Center, Union Carbide

Corp. 1989. Initial submission: Health and Safety Studies for 19

Chemicals with Cover Letter Date 060289. pp. 72-91.

 Genetic Toxicity in vitro

CAS Number : 29385-43-1

Sponsored Chemical

Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Genetic Toxicity in vitro

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

Sponsored Chemical

Result Type:

Measured

Method - Genetic Toxicity in vitro

Type of Study: Mammalian cell gene mutation assay

Concentrations: 12.5 - 750 ìg/mL

Year Study

Performed:

Method/Guideline

Followed:

No Data

GLP: No Data

Postive, Negative,

and Solvent

Control

Substance(s):

For each assay, there were 2-4 solvent controls, a positive control

of ethyl methylsulfonate at 4.7 x 10-6 M (or methyl

methanesulfonate at 10-20 ìg/mL) for the test without metabolic

activation, and a positive control of 3-methylcholanthrene at 1.86

x 10-5 M (or dimethylbenz[a]-anthracene at 0.5-4 ìg/mL) for the

test with metabolic activation.

Method/Guideline

and

Test Condition

Remarks:

L5178Y TK+/- 3.7.C mouse lymphoma cells were 

Originally obtained from Dr. Donald Clive, Burroughs Wellcome Co.

(Research

Triangle Park, NC). The cells were grown in Fischer's medium for

leukemic cells of mice (Gibco, Grand Island, NY, or Quality

Biological, Gaithersburg, MD) supplemented with 10%
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horse serum (Gibco or Hyclone, Logan, UT) and 0.02% pluronic F-68

(BASF Wyandotte Corp., Wyandotte, MI). Cells were screened for the

presence of mycoplasma after cryopreservation. New cultures were

initiated at approximately 3 month intervals from cells stored in

liquid N2.

The toxicity of each chemical was determined both with and without

liver S9 prepared from Aroclor 1254-induced male Sprague-Dawley

rats. S9 mix was prepared according to the procedure of Clive et

al. Cells at a concentration of 6 x 105/mL (6 x 106 cells total)

were exposed for 4 h to a range of concentrations from 0.0005 to

10000 ìg/mL. The cells were then washed,

resuspended in growth medium, and incubated at 37 ±1 °C for 48 h.

The rate of cell growth was determined for each of the treated

cultures and compared to the rate of growth of the solvent

controls. The doses of chemical selected for testing were within

the range yielding approximately 0-90% cytotoxicity. For each

assay, there were 2-4 solvent controls, a positive control of ethyl

methylsulfonate at 4.7 x 10-6 M (or methyl methanesulfonate at 10-

20 ìg/mL) for the test without metabolic activation, and a positive

control of 3-methylcholanthrene at 1.86 x 10-5 M (or

dimethylbenz[a]-anthracene at 0.5-4 ìg/mL) for the test with

metabolic activation.

The mutagenicity assay was performed according to the procedure

described by Clive and Spector. A total of 1.2 x 107 cells in

duplicate cultures were exposed to the test chemical, positive

control, and solvent control for 4 h at 37 ± 1 °C, washed twice

with growth medium, and maintained at 37 ±1 °C for 48 h in logphase growth to allow recovery and mutant expression. Cells in the

cultures were adjusted to 3 x 105/mL at 24 h intervals. They were

then cloned (1 x 106 cells/plate for mutant selection and 200

cells/plate for viable count determinations) in soft agar medium

containing Fischer's medium, 20% horse serum, 2 mM sodium pyruvate,

0.02% pluronic F-68, and 0.23% granulated agar (BBL, Inc.,

Cockeysville, MD). Resistance to trifluorothymidine (TFT) was

determined by adding TFT (final concentration, 3 ìg/mL) to

the cloning medium for mutant selection. The 100x stock solution of

TFT in saline was stored at -70 °C and was thawed immediately

before use. Plates were incubated at 37 ±1 °C in 5% CO2 in air for

10-12 days and then counted with an Artek automated colony counter

(Artek 982, DynaTech) or ProtoCol colony counter (Synbiosis,

Frederick, MD). Only colonies larger than x0.2 mm in diameter were

counted. Mutant frequencies were expressed as mutants per 106

surviving cells. Although there are several different methods for

evaluating mouse lymphoma data, results from this study were

interpreted using a doubling of the mutant frequency

over the concurrent solvent-treated control value as an indication

of a positive effect, together with evidence of a dose-related

increase. Doubling of the mutant frequency was previously reported

as representing a positive effect. Only doses yielding total growth

values of 10% were used in the analysis of induced mutant

frequency. Doses yielding less than 10% total growth were used in

determining dose response.

The size of mutant mouse lymphoma colonies was also determined

using an Artek 982 colony counter/sizer or the ProtoCol colony

counter. An internal discriminator was set to step sequentially to

exclude increasingly larger colonies in approximate increments of

0.1 mm in colony diameter. The size range used was from 0.2 to 1.1

mm.

Test Results - Genetic Toxicity in vitro

Details on

Cytogenetic Assay:

Statistics:

Effect : Species Other

Species

Strain Other

Strain

Metabolic

Activation

Genotoxic

Effect

Conclusion
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Mammalian

Cell Line

Mouse

Lymphoma

L5178Y

Cells

With and

Without

Positive Positive

With

Metabolic

Activation

Results Remarks: In the nonactivated test, the old evaluation was Positive at a

concentration of 750 ìg/mL, and the new evaluation was

inconclusive. In the activated test, the old evaluation was

positive at 25 ìg/mL, and the new evaluation was positive at 50

ìg/mL.

Conclusion: Positive in the presence of metabolic activation. Inconclusive

without metabolic activation.

Reliability/Data Quality - Genetic Toxicity in vitro

Reliability: 2 - Valid With Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Genetic Toxicity in vitro

Reference: Seifried et al. 2006. Compilation of two decades of mutagenicity

test results with the Ames Salmonella typhimurium and L5178Y mouse

lymphoma cell mutation assays. Chem. Res. Toxicol. 19: 627-644.

 Genetic Toxicity in vitro

CAS Number : 29385-43-1

Sponsored Chemical

Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Genetic Toxicity in vitro

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

Code Identification - TT-100

Sample number - X18BT7565 Cobratec 78669A

Description - Tan round crystals

Sponsored Chemical

Result Type:

Measured

Method - Genetic Toxicity in vitro

Type of Study: in vitro mammalian cell transformation assay

Concentrations: 6, 60 and 600 μg

Year Study

Performed:
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Method/Guideline

Followed:

No Data

GLP: No Data

Postive, Negative,

and Solvent

Control

Substance(s):

positive control: dimethylbenzanthracene

Method/Guideline

and

Test Condition

Remarks:

The C3H10T1/2 line is a fibroblastic cell line, derived from 

C3H mouse embryos which are highly sensitive to postconfluence

inhibition of cell division. 

When C3H10T1/2 cells are exposed to a variety of chemical

carcinogens morphologically 

transformed foci develop with a frequency that is dose

dependent. The foci have been shown to 

produce fibrosarcomas when injected into syngeneic mice. Therefore,

the C3H10T1/2 cell line 

can be used to assay for chemical carcinogens.

Two sets of 60-mm culture dishes are seeded with cells. One set of

dishes, 12 x replicates per dose, is seeded with 1000 cells. This

set is used for the transformation assay. The second set was

seeded with 200 cells per dish, 3 x per dose, and is utilized for

cytotoxicity determinations.

Both sets of cells are treated in an identical manner with

solutions of the test material. Following a 24-hour exposure

period, the toxicant is washed from all the plates and the cells

refed with normal growth media. Ten days following the initial

seeding, the plates designated for the cytotoxicity assay are

washed, fixed with methanol, and stained. The clones on all of the

plates are counted and the plating efficiency determined. This

plating efficiency is expressed as the number of colonies formed as

a percentage of the number of cells seeded.

The transformation set are incubated for 6 weeks with weekly medium

changes. During this time, the clones grow to form a confluent

monolayer of cells on the bottom of the plates. At the termination

of the incubation period, all remaining plates are washed, fixed,

stained and scored for the transformed foci:

Type I Foci: A Type I focus is composed of tightly packed cells 

Type II Foci: The cells display massive piling into opaque

multilayers.

Type III Foci: These foci are composed of highly polar, densely

stained cells, which form a criss-crossed multilayer.

Type II and Type III foci are considered to be malignantly

transformed.

The transformation frequency is calculated as the percentage of

cells that give rise to transformed foci corrected for plating

efficiency which is determined by the cytotoxicity assay.

Test Results - Genetic Toxicity in vitro

Details on

Cytogenetic Assay:

Statistics:

Effect : Species Other

Species

Strain Other

Strain

Metabolic

Activation

Genotoxic

Effect

Conclusion

Mammalian

Cell Line

Other -

C3H10T1/2

Mouse

Fibroblast

Not

Applicable

Negative Negative
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Results Remarks: For the 6, 60, 600 μg and control groups, the transformation

frequency was 0.32, 0.15, 0.07 and 0.07, respectively.

None of the sample chemicals at the dose levels tested produced

significant increases in transformation above control cell

population levels. As a positive control the cells did respond in

a dose-dependent manner, to the chemical carcinogen

dimethylbenzanthracene (DMBA).

Conclusion:

Reliability/Data Quality - Genetic Toxicity in vitro

Reliability: 2 - Valid With Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Genetic Toxicity in vitro

Reference: TSCATS submission (OTS0516797, Doc ID 86-890000263, received

06/05/89): Huntingdon Research Center, Union Carbide

Corp. 1989. Initial submission: Health and Safety Studies for 19

Chemicals with Cover Letter Date 060289. P 72-91

 Genetic Toxicity in vitro

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance - Genetic Toxicity in vitro

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

Sponsored Chemical Result

Type:

Measured

Method - Genetic Toxicity in vitro

Type of Study: Bacterial reverse mutation assay

Concentrations: 100 - 10,000 ug/plate

Year Study Performed:

Method/Guideline

Followed:

Other

GLP: No Data
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Postive, Negative, and

Solvent

Control Substance(s):

Method/Guideline and

Test Condition Remarks:

The experimental materials, methods, and procedures as 

explained below are based on those described by Ames et al.

For initial mutagenicity testing, the 

plate incorporation assay was used; some compounds were also

tested using the flavin 

mononucleotide (FMN)-dependent reducing conditions described

by Prival and Mitchell to break 

the diazo bonds of the condensed dyes, forming the individual

constituents.

S9 Preparation. Liver S9 homogenate was prepared from male

Sprague-Dawley rats and Syrian golden hamsters that had been

injected with Aroclor 1254 at 500 mg/kg body weight. The

postmitochondrial (microsomal) enzyme fractions were prepared

as described by Ames et al. (26). The components of the S9

mix were 8 mM MgCl2, 33 mM KCl, 5 mM glucose-6-phosphate, 4

mM NADP, 100 mM sodium phosphate (pH 7.4), and the

appropriate S9 homogenate at a concentration of 0.1 mL/mL of

mix. For each plate receiving microsomal enzymes, 0.5 mL of

S9 mix was added.

Plate Incorporation Methodology. For testing in the absence

of S9 mix, 100 ìL of the tester strain and 50 ìL of the

solvent or test chemical were added to 2.5 mL of molten

selective top agar at 45 ±2 °C. When S9 was used, 0.5 mL of

S9 mix, 50 ìL of tester strain, and 50 ìL of solvent or test

chemical were added to 2.0 mL of molten selective top agar at

45 ±2 °C. After it was vortexed, the mixture was overlaid

onto the surface of 25 mL of minimal bottom agar. After the

overlay had solidified, the plates were incubated for 48 h at

37 ±2 °C. Five doses of test chemical, together with the

appropriate concurrent solvent and positive controls, were

tested in triplicate on each tester strain without metabolic

activation and also with activation by induced rat and

hamster liver S9 preparations.

The doses that were tested in the mutagenicity assay were

selected based on the levels of cytotoxicity observed in a

preliminary dose range-finding study using strain TA100. Ten

dose levels of the chemical, one plate per dose, were tested

in both the presence and the absence of induced hamster S9.

If no toxicity was observed, a total maximum dose of 10 mg of

test chemical per plate was used. The criteria used to

evaluate a test were as follows: for a

test article to be considered positive, it had to induce at

least a doubling (TA98, TA100, and TA1535) in the mean number

of revertants per plate of at least one tester strain. This

increase in

the mean revertants per plate had to be accompanied by a dose

response to increasing concentrations of the test chemical.

If the study showed a dose response with a less than 3-fold

increase on TA1537 or TA1538, the response had to be

confirmed in a repeat experiment.

Preincubation Methodology under Reductive Conditions. For

tests using the FMN-modified assay, strains TA98 and TA100

were used. The bacteria, uninduced hamster liver S9 (30%

v/v),

cofactors (FMN, NADH, glucose-6-phosphate dehydrogenase, and

glucose-6-phosphate), and test chemical were added, mixed,

and incubated at 30 °C for 30 min without shaking. Nitrogen

was blown over the preincubation tube to keep the atmosphere

reduced. At the end of the incubation period, 2 mL of molten

top agar was added to each sample tube and the mixture was

poured on a minimal agar plate containing 0.5% glucose rather

than the 2% glucose specified by Ames et al. The plates were

then incubated at 37 °C for 48 h. The positive control in all

FMN experiments was Congo red. All plates were counted with
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an Artek automated colony counter (Artek 880, DynaTech,

Chantilly, VA) or Minicount colony counter (Imaging Products

International, Inc., Chantilly, VA), which was calibrated

prior to use.

Test Results - Genetic Toxicity in vitro

Details on Cytogenetic

Assay:

Statistics:

Effect : Species Other

Species

Strain Other

Strain

Metabolic

Activation

Genotoxic

Effect

Conclusion

Bacteria With and

Without

Negative Negative

Results Remarks:

Conclusion:

Reliability/Data Quality - Genetic Toxicity in vitro

Reliability: 2 - Valid With Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Genetic Toxicity in vitro

Reference: Seifried et al. 2006. Compilation of two decades of

mutagenicity test results with the Ames Salmonella typhimurium

and L5178Y mouse lymphoma cell mutation assays. Chem. Res.

Toxicol. 19: 627-644.

 Genetic Toxicity in vitro

CAS Number : 29385-43-1

Sponsored Chemical

Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Genetic Toxicity in vitro

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

99+ % pure

Sponsored Chemical Measured
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Result Type:

Method - Genetic Toxicity in vitro

Type of Study: Bacterial reverse mutation assay

Concentrations: 0, 33, 100, 333, 1000, 1666, 3333, 6666 ìg/plate

Year Study

Performed:

1983

Method/Guideline

Followed:

Other

GLP: No Data

Postive, Negative, and

Solvent

Control Substance(s):

vehicle control: dimethylsulfoxide; The positive controls in the

absence of metabolic activation were sodium azide (TA1535 and

TA100), 9-aminoacridine (TA97 and TA1537), and 4-nitro-ophenylenediamine (TA98). The positive control for metabolic

activation with all strains was 2-aminoanthracene.

Method/Guideline and

Test Condition

Remarks:

Preparation of Liver S-9 Fractions - The S-9 (9,000 g

supernatant) fractions of Aroclor 1254-induced, male SpragueDawley rat and male Syrian hamster livers were prepared as

described previously [Haworth et al, 1983]. The S-9 mixes were

prepared immediately prior to use and contained either 10% or 30%

S-9; occasionally, other levels were used. All chemicals were

tested in the absence of metabolic activation and with rat and

hamster S-9 fractions.

Test Protocol - The preincubation assay was performed as described

previously [Haworth et al., 1983], with some differences, as

described below. The test chemical (0.05 mL), Salmonella culture

(0.10 mL), and S-9 mix or buffer (0.50 mL) were incubated at 37

ºC, without shaking, for 20 min. Chemicals known or

suspected to be volatile were incubated in capped tubes. The top

agar was added and the contents of the tubes were mixed and

poured onto the surface of petri dishes containing Vogel-Bonner

medium [Vogel and Bonner, 1956]. The histidine-independent (his+)

colonies arising on these plates were counted following two days

incubation at 37 ºC. Plates were machine counted unless

precipitate was present which interfered with the count, or the

color of the test chemical on the plate reduced the contrast

between the colonies and the background agar. At the discretion

of the investigators, plates with low numbers of colonies were

counted by hand.

Variations in the protocol among the tested chemicals reflect the

evolution of the protocol originally described by Haworth et al.

[1983]. Four protocol variations are evident. 1) Testing in

strains TA97, TA98, TA100 and TA1535, with some additional testing

in strain TA1537; 10% S-9 was used. 2) The first test of a

chemical was without activation and with 10% S-9 in the mix. If

a positive result was obtained the test was repeated. If the

tests were negative they were repeated without S-9 and with 30%

S-9. 3) The order of use of 10% and 30% S-9 was reversed. 4)

Initial testing was in strains TA98 and TA100 without activation

and with 30% rat and hamster S-9s. If a positive result was

obtained in one of these two strains it was repeated and the

other strains were not used. If the tests were negative, the

other strains were used with 30% and 10% S-9. A chemical was not

designated nonmutagenic unless it had been tested in strains TA98,

TA100, TA1535, and TA97 and/or TA1537, without activation and with

10% and 30% rat and hamster S-9. Occasionally, 5% S-9 was also

used in all protocol variations.

All chemicals were tested initially in a toxicity assay to

determine the appropriate dose range for the mutagenicity

assay. The toxicity assay was performed using TA100 or the system

developed by Waleh et al. [1982]. Toxic concentrations were those
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that produced a decrease in the number of his+ colonies, or a

clearing in the density of the background lawn, or both.

Each chemical was tested initially at half-log dose intervals up

to a dose that elicited toxicity, or to a dose immediately below

one which was toxic in the preliminary toxicity test. Subsequent

trials occasionally used narrower dose increments and may not have

included doses in the toxic range. Experiments were repeated at

least one week following the initial trial. A maximum of 0.05 mL

solvent was added to each plate.

Concurrent solvent and positive controls were run with each

trial. The positive controls in the absence of metabolic

activation were sodium azide (TA1535 and TA100), 9-aminoacridine

(TA97 and TA1537), and 4-nitro-o-phenylenediamine (TA98). The

positive control for metabolic activation with all strains was 2-

aminoanthracene. Occasionally, a laboratory used a single solvent

and/or positive control for more than one chemical tested on the

same day. Each laboratory made its own determination regarding

positive control dose levels.

A chemical was judged mutagenic or weakly mutagenic if it produced

a reproducible dose-related response over the solvent control in

replicate trials. A chemical was judged questionable if the

results of the individual trials were not reproducible, if

increases in his+ revertants did not meet the criteria for a

'weakly mutagenic' response, or if only single doses produced

increases in his+ revertants in repeat trials. Chemicals were

judged nonmutagenic if they did not meet the criteria for a

mutagenic or questionable response. 

Test Results - Genetic Toxicity in vitro

Details on Cytogenetic

Assay:

Statistics:

Effect : Species Other

Species

Strain Other

Strain

Metabolic

Activation

Genotoxic

Effect

Conclusion

Bacteria S.

typhimurium

TA 97

With and

Without

Bacteria S.

typhimurium

TA 98

With and

Without

Bacteria S.

typhimurium

TA 100

With and

Without

Bacteria S.

typhimurium

TA 1535

With and

Without

Bacteria S.

typhimurium

TA 1537

With and

Without

Results Remarks: Test substance is weakly positive.

Toxicity was observed at 6666 ìg/plate in TA97 with activation and

in TA1535 with activation.

Conclusion: Weakly positive.

Reliability/Data Quality - Genetic Toxicity in vitro

Reliability: 1 - Valid Without Restrictions

Reliability Remarks:
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Key Study Sponsor

Indicator:

Reference - Genetic Toxicity in vitro

Reference: NTP study 651117. Zeiger, E., Anderson, B., Haworth, S., Lawlor,

T., and Mortelmans, K. Salmonella mutagenicity tests. IV. Results

from the testing of 300 chemicals Environ. Molec. Mutagen. Vol. 11

(Suppl 12) (1988) 1-158

 Genetic Toxicity in vitro

CAS Number : 29385-43-1

Sponsored

Chemical Name :

1H-Benzotriazole, 4(or 5)-methylTest Substance - Genetic Toxicity in vitro

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test

Substance

Comments:

Code Identification - TT-100; Sample number - X18BT7565 Cobratec

78669A; Description - Tan round crystals

Sponsored

Chemical Result

Type:

Measured

Method - Genetic Toxicity in vitro

Type of Study: DNA damage or repair assay, unscheduled DNA synthesis in mammalian cells

in vitro

Concentrations: not legible

Year Study

Performed:

Method/Guideline

Followed:

No Data

GLP: No Data

Postive, Negative,

and Solvent

Control

Substance(s):

Method/Guideline

and

Test Condition

Remarks:

This test system is utilized to determine the potential of compounds to

damage DNA in mammalian cells.

One mL of the cell suspension (2 x 105 cells/mL) is added to four mL of

media in 60-mm tissue culture plates (6 x replicates per dose

level). The cells are allowed to grow to confluency (5-7 days) then

exposed to 10 mM hydroxyurea for 2 hours. The cells are then washed

and exposed to a mixture of the test material and tritiated thymidine

(3H-TdR) for 24 hours. After the exposure the plates are washed 3 X

with phosphate buffered saline (pH 7.2) and lysed with 2 mL of 0.1M

NaOH. After neutralization with 0.2 mL of 1M HCl, 2.2 mL of cold 10

percent TCA is added, and the plates incubated at 4 oC for 1 hour. The

lysate is then filtered through a GF/A filter which is subsequently

washed 3 X with TCA. The filters are dried for 8 hours and counted in

a Nuclear Chicago Mark III liquid scintillation counter.

Dosage levels which have caused DNA damage will exhibit elevated counts

due to the increased update of 3H-TdR are mediated by the DNA repair

mechanism in the cells.
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Test Results - Genetic Toxicity in vitro

Details on

Cytogenetic Assay:

Statistics:

Effect : Species Other

Species

Strain Other

Strain

Metabolic

Activation

Genotoxic

Effect

Conclusion

Mammalian

Cell Line

Mammalian

Cell Line:

Other

- WI-38

Human

Embryonic

Lung

Fibroblasts

Without Negative Negative

Results Remarks: The potential for damaging cellular DNA was monitored in WI-38 human

embryonic lung fibroblasts by measuring the amount of 3H-TdR

incorporated into cells exposed to subtoxic levels of the chemical

samples.

Samples of TT-100 did not elicit significant incorporation above

control levels at any of the dose levels tested.

Conclusion: Samples of TT-100 did not elicit significant incorporation above

control levels at any of the dose levels tested.

Reliability/Data Quality - Genetic Toxicity in vitro

Reliability: 2 - Valid With Restrictions

Reliability

Remarks:

Key Study Sponsor

Indicator:

Reference - Genetic Toxicity in vitro

Reference: TSCATS submission (OTS0516797, Doc ID 86-890000263, received 06/05/89):

Huntingdon Research Center, Union Carbide Corp. 1989. Initial

submission: Health and Safety Studies for 19 Chemicals with Cover Letter

Date 060289. P 72-91

 Skin Irritation

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance - Skin Irritation

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

Referred to as Cobratec TT-100 Sample No. 6022
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Sponsored Chemical Result

Type:

Measured

Method - Skin Irritation

Species: Rabbit

Mammalian Strain: New Zealand White

Type of Coverage: Occlusive

Preparation of Test Site: Shaved

Gender: Female

Number of Animals per

Dose:

12

Amount/Concentration

Applied:

0.5 grams

Year Study Performed:

Method/Guideline

Followed:

No Data

GLP: No Data

Exposure Period: 24 Hours

Total Volume applied and

Units:

Control Group Type:

Vehicle Used:

Vehicle Name:

Vehicle Amount and Units:

Post-Exposure Period: 48 Hours

Grading Scale:

Method/Guideline and

Test Condition Remarks:

Twelve adult female New Zealand albino rabbits were

used. Prior to application the backs were clipped free of

hair. The skin was left intact on six of the rabbits and was

abraded on the other six. One half gram of the sample was

applied to an area of skin approximately one inch square, two

single layers thick, were applied over the samples and

covered with a non-reactive tape. The rabbits were then

wrapped with a binder.

After the 24-hour exposure, the binders and the tape were

removed and the skin reactions evaluated. An evaluation was

also made at 72 hours after application.

The reactions were scored according to the method described

in the Federal Hazardous Substances Act.

Test Results - Skin Irritation

Grade:
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Primary Irritation Index: 0

Lesions:

Erythema:

Edema:

Results Remarks: The primary irritation index was 0.0. There was no skin

irritation.

Interpretation of Results: Not Irritating

Conclusion: Not irritating to rabbit skin.

Reliability/Data Quality - Skin Irritation

Reliability: 2 - Valid With Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Skin Irritation

Reference: TSCATS submission (OTS0516797, Doc ID 86-890000263, received

06/05/89): Huntingdon Research Center, Union Carbide

Corp. 1989. Initial submission: Health and Safety Studies

for 19 Chemicals with Cover Letter Date 060289. pp. 51-57.

 Eye Irritation

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance - Eye Irritation

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

Referred to as Olin 58734. Pinkish grey granular material

Sponsored Chemical Result

Type:

Measured

Method - Eye Irritation

Species: Rabbit

Mammalian Strain: No Data

Gender: Both M/F

Number of Animals per

Dose:

6

Amount/Concentration

Applied:

10 mg
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Year Study Performed:

Method/Guideline

Followed:

Other

GLP: No Data

Exposure Period:

Total Volume applied and

Units:

10 mg

Control Group Type:

Vehicle Used:

Vehicle Name:

Vehicle Amount and Units:

Post-Exposure Period: 72 Hours

Grading Scale:

Tool:

Method/Guideline and

Test Condition Remarks:

Method/guideline followed: Method in Appraisal of the Safety

of Chemicals in Food, Drugs, and Cosmetics, Association of

Food and Drug Officials of the U.S. 1959, p. 46

Dosage - 10 mg of powdered sample in right eye, without

washout.

Maintenance conditions - Animals maintained in stocks for 6

hours after dosage. Thereafter, individual housing with food

and water ad libitum.

Observations - Evaluation of degree of eye irritation at 24,

48, and 72 hours, as described in Regulations

Test Results - Eye Irritation

Overall Grading Score:

Overall Irritation Score:

Primary Irritation Index:

Lesions:

Cornea:

Iris:

Conjunctivae (Chemosis):

Conjunctivae (Redness):

Results Remarks: Cornea - Moderate opacity in small area in 3 on day 1, in 2

on day 2, in 1 on day 3.
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Iris - Irritation in 1 on days 1, 2, and 3.

Conjunctiva - Lid redness slight in 6 on day 1, very slight

in 2 on day 3. Chemosis slight in 4 on day 1. Discharge

slight in 6 on day 1, very slight in 1 on day (illegible).

Interpretation of Results: Slightly Irritating

Conclusion: The test substance is slightly and transiently irritating to

the eyes of rabbits.

Reliability/Data Quality - Eye Irritation

Reliability: 2 - Valid With Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Eye Irritation

Reference: TSCATS submission (OTS0520643, Doc ID 86-89000060A, received

06/13/89): Huntingdon Research Center, Sherwin Williams

Co. 1968. Initial submission: Summary of Mucuous Membrane

(Eye) Irritations in Rabbits Treated with Olin 58734 with

Attachment and Cover Letter Dated 061289.

 Eye Irritation

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance - Eye Irritation

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

Sponsored Chemical Result

Type:

Measured

Method - Eye Irritation

Species: Rabbit

Mammalian Strain: New Zealand White

Gender: Female

Number of Animals per

Dose:

6

Amount/Concentration

Applied:

100 mg

Year Study Performed:

Method/Guideline

Followed:

No Data

GLP: No Data
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Exposure Period:

Total Volume applied and

Units:

100 mg

Control Group Type: Negative

Vehicle Used:

Vehicle Name:

Vehicle Amount and Units:

Post-Exposure Period: 14 Days

Grading Scale:

Tool: Fluorescein

Method/Guideline and

Test Condition Remarks:

Six female albino New Zealand rabbits were used. Both eyes

of each animal were examined with the aid of an

ophthalmoscope before testing. 100 mg of test material was

placed in one eye of each animal by gently pulling the lower

lid away from the eyeball to form a cup into which the test

substance was dropped. The lids were gently held together

for one second and the animal released. The other eye,

remaining untreated, served as a control.

The eyes were examined and the grade of ocular reaction

recorded at 24, 48, 72 hours and 14 days. Reading of

reactions was facilitated by the use of an

ophthalmoscope. After the recording of observations at 24

hours, the eyes were stained with fluorescein and examined

with an ultra-violet lamp to detect corneal damage if

present.

The reactions were scored from: "Illustrated Guide for

Grading Eye Irritation by Hazardous Substances," Section

191.12 of the Federal Hazardous Substances Labeling Act

Regulations. 

Test Results - Eye Irritation

Overall Grading Score:

Overall Irritation Score:

Primary Irritation Index:

Lesions:

Cornea:

Iris:

Conjunctivae (Chemosis):

Conjunctivae (Redness):

Results Remarks: The test substance was found to be irritating to rabbit

eyes. Conjunctival irritation scores ranged from 0 to 4
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throughout the observation period. Corneal irritation scores

ranged from 0 to 3. Iridial irritation scores ranged from 0

to 1.

One rabbit died prior to the 14 day reading. All rabbits

exhibited treatment-related effects on the conjunctiva and

cornea. 4/6 rabbits exhibited treatment-related effects on

the iris. 

No. Observed / No. Exposed

Cornea observations -

2/6 - a diffuse area of corneal opacity

3/6 - discernible translucent areas of opacity, details of

iris slightly obscured

1/6 - no details of iris visible

6/6 - fluorescein examination positive

Iris observations - 4/6 - moderate circumcorneal injection

Conjunctival redness - 

4/6 - diffuse crimson red

2/6 - diffuse beefy red

Chemosis - 

1/6 - swelling with partial eversion of lids

2/6 - swelling with lids about half closed

3/6 - swelling with lids half closed or greater

Interpretation of Results: Irritating

Conclusion: Irritating to rabbit eyes

Reliability/Data Quality - Eye Irritation

Reliability: 2 - Valid With Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Eye Irritation

Reference: TSCATS submission (Doc ID 86-890000263, received 06/05/89):

Huntingdon Research Center, Union Carbide

Corp. 1989. Initial submission: Health and Safety Studies

for 19 Chemicals with Cover Letter Date 060289. P 60-65.

 Skin Sensitization

CAS Number : 29385-43-1

Sponsored Chemical Name

:

1H-Benzotriazole, 4(or 5)-methylTest Substance - Skin Sensitization

Test Substance: (29385-43-1) 1H-Benzotriazole, 4(or 5)-methylTest Substance

Purity/Composition

and Other Test Substance

Comments:

Referred to as TK 10 637/1, Lot 1179/1186, 97.8% pure

Sponsored Chemical Result

Type:

Measured
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Method - Skin Sensitization

Test Type: In Vivo

Study Type: Maurer Optimization Test

Species: Guinea pig

Mammalian Strain: Pirbright-Hartley

Route of Induction: Intradermal

Route of Challenage

Exposure:

Intradermal and Epicutaneous

Gender: Both M/F

Number of Animals per

Dose:

20

Concentration: 0.1% for intradermal application

10% for epidermal application

Year Study Performed: 1982

Method/Guideline

Followed:

Other

GLP: No Data

Exposure Period:

Induction Frequency of

Treatment:

once every second day (except weekends) to a total of 10

Challenge Exposure

Period:

24 Hours

Challenge Frequency of

Treatment:

Total Volume applied and

Units:

Control Group Type: Vehicle

Vehicle Used: Yes

Vehicle Name: Other

Other Vehicle Name: physiological saline

Vehicle Amount and Units:

Positive Control

Substance:

Negative Control

Substance:

phsyiological saline

Post-Exposure Period: 24 Hours

Method/Guideline and

Test Condition Remarks:

The optimization test was used, an intracutaneous

sensitization procedure exceeding the sensitivity of the

method recommended in the "Appraisal of the Safety of
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Chemicals in Foods, Drugs and Cosmetics" (1959), the US

Association of Food and Drug Officials (AFDO).

The test was performed on 10 male and 10 female guinea pigs

of the Pirbright white strain, bred on the premises and

weighing between 221 to 431 grams (~10 weeks old). 

The animals were housed individually in Macrolon cages type

3, assigned to the different groups by means of random

numbers generated by the random number generator incorporated

in the Hewlett-Packard desk computer, identified with

individual ear tags, kept at a constant room temperature of

20 ±1 C, at a relative humidity of 50 ± 10% and on a 14

hours light cycle. The animals received ad libitum standard

guinea pig pellets and water, supplemented with fresh

carrots. The animals were acclimatized for 12 days. 

After the 24-hour exposure, the binders and the tape were

removed and the skin reactions evaluated. An evaluation was

also made at 72 hours after application. The reactions were

scored according to the method described in the Federal

Hazardous Substances Act.

During the induction period the animals received one

injection every second day (except weekends) to a total of 10

intracutaneous injections of a freshly prepared 0.1% solution

of TK 10 637/1 in physiological saline. One control group

was treated with the vehicle alone ("negative control").

On the first day, injections of 0.1 mL were administered into

the shaved skin of the right flank and the back, while on the

following days a single intracutaneous injection was given

into the back. 

During the second and third week of the induction period the

test material was incorporated in a mixture of the normal

vehicle with complete Bacto Adjuvant.

(vehicle:adjuvant = 1:1).

Fourteen days after the last sensitizing injection, a

challenge injection of 0.1 mL of a freshly prepared 0.1%

solution of TK 10 637/1 in physiological saline was

administered into the skin of the left flank. 

Twenty-four hours after each injection during the first week

of the induction period and 24 hours after the challenge

injection the reactions were recorded.

The two largest perpendicular diameters (in mm) and the

increase in the skin-fold thickness (in mm) were measured and

by multiplication of these values a "reaction volume" was

obtained (in μl) for each reading from each animal. The mean

volume plus one standard deviation of the induction reactions

observed in the individual animal in the first week was taken

as representing the skin irritation "threshold" for each

animal. Any challenge reaction greater than this threshold

value in the induction period was graded as an allergic

reaction and the animal termed "positive". The number of

"positive" animals in the test group was compared with the

number of animals in the control group (treated with the

vehicle alone) that showed a non-specific reaction of at

least the same magnitude ("negative control").

Ten days after the intracutaneous challenge injection a

subirritant dose of the test compound (10% TK 10 637/1 in

vaseline) was applied epicutaneously under occlusive dressings

which were left in place for 24 hours. The reactions were

evaluated 24 hours after removing of the bandages according

to Draize scoring scale.
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Test Results - Skin Sensitization

Measurement Period 

and Units:

Percent Sensitized 

Test Substance:

Percent Sensitized 

Positive Control:

Percent Sensitized 

Negative Control:

Sensitization Score:

Results Remarks: Under the experimental conditions employed, no differences

between the test group and the vehicle-treated controls were

seen, after either intradermal or epidermal challenge

application of TK 10 637/1. TK 10 637/1 was found to be

devoid of skin-sensitizing (contact allergenic) potency in

albino guinea-pigs.

Interpretation of Results: Not Sensitizing

Conclusion: Not a sensitizer

Reliability/Data Quality - Skin Sensitization

Reliability: 1 - Valid Without Restrictions

Reliability Remarks:

Key Study Sponsor

Indicator:

Reference - Skin Sensitization

Reference: TSCATS submission (OTS0519042, Doc ID 86-890000273, received

06/07/89): Ciba Geigy Corp. 1982. Initial submission: Report

on Skin Sensitizing Effect in Guinea Pigs of TK 10 637/1 with

Cover Letter Dated 060589.