1,6-Bis(methoxybenzoyloxy)hexane

Toxicity to reproduction

Administrative data

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods with acceptable restrictions

Justification for type of information:

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This report addresses the use of a combination of data for hexane-1,6-diol (CAS No. 629-11-8) and for vanillin (CAS No. 121-33-5) to address the reproductive/developmental toxicity screening data requirement for 1,6-bis(methoxybenzoyloxy)hexane (CAS No. 390359-18-9) in an analogue read-across approach. The basis for this read-across approach is that, upon oral administration, the target substance is expected to hydrolyse prior to absorption into hexane-1,6-diol and p-anisic acid (CAS No. 100-09-4). The reproductive/developmental toxicity of the hexane-1,6-diol metabolite will be assessed using information on hexane-1,6-diol, and the reproductive/developmental toxicity of the p-anisic acid metabolite will be assessed using information on vanillin, to which it is structurally very similar.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
See attached justification for full details
PURITY/IMPURITIES
The target substance is known to be of high purity (typically 96.8 % w/w), and to contain two known impurities. The first known impurity, typically present at 1.72 % w/w, is the mono-ester analogue of the target substance, which is an intermediate hydrolysis product of the target substance, and which is itself expected also to hydrolyse to the same products as the target substance. The second known impurity, typically present at 0.52 % w/w, is one of the final hydrolysis products of the target substance. On this basis, the source substances effectively represent typically >99 % w/w of the target substance. The purities of the samples of source material that were tested are not specifically known, but it is assumed that they would not have been sufficiently impure as to substantially affect the study results. On this basis, the applicability of the data on the source substance to the target substance is not expected to be compromised by the presence of impurities in either substance.

The p-anisic acid metabolite and vanillin source substance are both oxygenated benzene compounds, carrying both carbonyl and methoxy substituent groups, and are identified as possessing 50.2% structural similarity in the ChemIDplus advanced database, which is one of ECHA’s recommended data sources for analogue identification. They differ only in that, in p-anisic acid the carbonyl group is the carboxylic acid, and is positioned para to the methoxy group, whereas in vanillin the carbonyl group is the aldehyde and is positioned meta to the methoxy group and para to a hydroxy group.

3. ANALOGUE APPROACH JUSTIFICATION
See attached justification for full details
The basis for this read-across approach is that the target substance is a bis carboxylic ester that, upon oral administration, is expected to metabolise, hydrolytically and prior to absorption, into hexane-1,6-diol and p-anisic acid.

4. DATA MATRIX
See attached justification for full details

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

read-across source

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Qualifier:

according to guideline

Guideline:

OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)

GLP compliance:

not specified

Limit test:

no

Specific details on test material used for the study:

Hexanediol

Species:

rat

Strain:

Wistar

Sex:

male/female

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on exposure:

Male and female Wistar rats received 0, 100, 400 and 1,000 mg/kg body weight Hexanediol by gavage for 4 (males) to 6 (females) weeks in compliance with OECD test method 421 to screen the effect of hexanediol on reproduction and developmental toxicity. The premating exposure period was at least 14 days and the study was terminated 4 days post partum of the F1 generation pups.

Dose / conc.:

100 mg/kg bw/day (nominal)

Dose / conc.:

400 mg/kg bw/day (nominal)

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

4 (males) to 6 (females)

Clinical signs:

not specified

Mortality:

not specified

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Marginal retarded body weight development in males at 1,000 mg/kg b.w. was the only effects noted in this study.

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Dose descriptor:

NOAEL

Effect level:

400 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

body weight and weight gain

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Remarks on result:

not determinable due to absence of adverse toxic effects

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Reproductive effects observed:

no

Conclusions:

In a valid OECD 421 study no indication of toxic effect on reproductive function or developmental toxicity were observed.

Cross-reference 2

Reason / purpose for cross-reference:

read-across source

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Route of administration:

oral: gavage

Vehicle:

not specified

Details on exposure:

no data

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

no data

Details on mating procedure:

no data

Duration of treatment / exposure:

1 week before mating until 4 days post parturition

Frequency of treatment:

no data

Duration of test:

no data

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

125 mg/kg bw/day (nominal)

Dose / conc.:

250 mg/kg bw/day (nominal)

Dose / conc.:

500 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10 females

Control animals:

yes, concurrent vehicle

Details on study design:

no data

Maternal examinations:

clinical signs, body weight and feed consumption were recorded throughout the study. Mating performance, fertility, duration of gestation/
parturition, maternal behavior, litter sizes, dystocia, number of implantation sites, and gross lesions at necropsy were also examined.

Ovaries and uterine content:

no data

Fetal examinations:

viability, sex, external morphology, and body weight at birth and on day 4 postpartum

Statistics:

no data

Indices:

no data

Historical control data:

no data

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
Toxic signs included death, clinical observations and changes in body weight and feed consumption

Dose descriptor:

NOAEL

Effect level:

250 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

clinical signs

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no data

Dose descriptor:

NOEL

Effect level:

500 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: teratogenicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Conclusions:

Vanillin was considered to have no teratogen effects

Executive summary:

10 female rats per group were exposed one week before mating until 4 days post partum to 0, 125, 250, 500 mg/kg bw/day of Vanillin. Maternal toxicity had been reported with only few details: death, clinical signs and change in body weight and feed consumption. The NOAEL maternal was considered to be 250 mg/kg bw. No effect on pups were reported and the NOEL for teratogenicity was the highest dose tested 500 mg/kg bw.

Cross-reference 3

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

water solubility

Type of information:

experimental study

Adequacy of study:

key study

Study period:

November 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: EU A.6 and OECD 105 test with GLP compliance

Qualifier:

according to guideline

Guideline:

OECD Guideline 105 (Water Solubility)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method A.6 (Water Solubility)

Deviations:

no

GLP compliance:

yes

Type of method:

column elution method

Water solubility:

8.8 µg/L

Temp.:

19.5 °C

pH:

7.5

Details on results:

Nominal concentration was 18.9g/L.
Maximum deviation is 1.4% for all the test runs.

Conclusions:

Interpretation of results (migrated information): insoluble (< 0.1 mg/L)
Water solubility of the substance was meassured at 0.0000088g/L

Executive summary:

Water solubility of the substance was meassured at 0.0000088g/L

Cross-reference 4

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

water solubility

Type of information:

(Q)SAR

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model, but not (completely) falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

1. SOFTWARE: US EPA Toxicity Estimation Software Tool (T.E.S.T.) Version 4.2

2. MODEL: Predicted Water solubility at 25°C for 629-11-8 from Consensus method

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL: CAS Number 629-11-8

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
- Defined endpoint: Water solubility at 25°C mg/L
- Unambiguous algorithm: In the consensus method, the predicted water solubility is simply the average of the predicted toxicities from the other QSAR methodologies, specifically hierarchical clustering and single model, (taking into account the applicability domain of each method). If only a single QSAR methodology can make a prediction, the predicted value is deemed unreliable and not used. This method typically provides the highest prediction accuracy since errant predictions are dampened by the predictions from the other methods. In addition, this method provides the highest prediction coverage because several methods with slightly different applicability domains are used to make a prediction.
- Defined domain of applicability: Chemicals with water solubility less than 1,000,000 mg/L.
- Appropriate measures of goodness-of-fit and robustness and predictivity:
Mean absolute error in -Log10(mol/L): Entire set 0.58
Mean absolute error in -Log10(mol/L): Similarity coefficient (≥ 0.5) 0.27

5. APPLICABILITY DOMAIN
- Descriptor domain:
10 analogues formed the dataset
Predicted water solubility is less than 1,000,000 mg/L
- Similarity with analogues in the training set:
The target substance is similar to the analogues within the training set (similarity coefficient = 0.27).

6. ADEQUACY OF THE RESULT
The prediction provides a discrete water solubility prediction and there are similar analogues within the training set. The prediction indicates that the target substance is soluble in water.

Guideline:

other: US EPA Toxicity Estimation Software Tool (T.E.S.T.) Version 4.2

Principles of method if other than guideline:

Software tool(s) used including version: US EPA Toxicity Estimation Software Tool (T.E.S.T.) Version 4.2
- Model(s) used: Predicted Water solubility at 25°C for 629-11-8 from Consensus method
- Justification of QSAR prediction: see field 'Justification for type of information'

GLP compliance:

no

Type of method:

other: Not specified - QSAR prediciton

Specific details on test material used for the study:

CAS Number: 629-11-8

Water solubility:

35 032 mg/L

Conc. based on:

test mat.

Temp.:

25 °C

Remarks on result:

other: QSAR predicted value

Conclusions:

The model provided a valid water solubility prediction of 35032 mg/L at 25°C

Executive summary:

The US EPA Toxicity Estimation Software Tool (T.E.S.T.) Version 4.2 model predicted a water solubility of 35032 mg/L at 25°C.

The model result is valid and indicates that the substance CAS Number 629-11-8 is solubility in water.

Cross-reference 5

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

water solubility

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Principles of method if other than guideline:

- Principle of test: Determine water solubility
- Short description of test conditions: Unknown, textbook value
- Parameters analysed / observed: water solubility

GLP compliance:

not specified

Type of method:

other: Not specified

Water solubility:

10 000 mg/L

Conc. based on:

test mat.

Remarks on result:

completely miscible

Remarks:

reported as: 1g dissolves in 100mL water

Details on results:

The reference reports 1 gram dissolves in 100 mL water.

Conclusions:

The handbook data reported the substance was solube in water (1 g in 100 mL water).

Cross-reference 6

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

water solubility

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Principles of method if other than guideline:

- Principle of test: Determine water solubility
- Short description of test conditions: Unknown, textbook value
- Parameters analysed / observed: water solubility

GLP compliance:

not specified

Type of method:

other: Not specified

Water solubility:

400 mg/L

Conc. based on:

test mat.

Remarks on result:

completely miscible

Remarks:

reported as: soluble in 2500 parts water

Details on results:

The reference reports soluble in 2500 parts water.

Conclusions:

The handbook data reported the substance was solube in water (soluble in 2500 parts water).

Cross-reference 7

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

partition coefficient

Type of information:

experimental study

Adequacy of study:

key study

Study period:

September 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: According to OECD and EEC test methods and to GLP standards.

Qualifier:

according to guideline

Guideline:

OECD Guideline 117 (Partition Coefficient (n-octanol / water), HPLC Method)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method A.8 (Partition Coefficient)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: Rekker Calculation method for preliminary estimation of Partition coeficient.

Deviations:

no

GLP compliance:

yes

Type of method:

HPLC method

Partition coefficient type:

octanol-water

Analytical method:

high-performance liquid chromatography

Type:

log Pow

Partition coefficient:

5.2

Temp.:

24.5 °C

Remarks on result:

other: pH not specified

Details on results:

The results of the Calculation method and the HPLC method are not in agreement. Since the HPLC method is a more accurate method than the Calculation method, the result of the HPLC method is reported as the partition coefficient (n-octanol/water), Pow of Setafix X 11342.

The Pow value for the major component in Setafix X 11342 is 18000 (log Pow= 5.2) at 24.5 ± 0.5°c.
The Pow values for eleven impurities in Setafix X 11342 are <2, 170, 350, 4800, 6100, 11000, 13000, 28000 , 49000, 82000 and 120000 (log Pow values of <0.3, 2.2, 2.5, 3.7, 3.8, 4.0, 4.1, 4.4, 4.7, 4.9 and 5.1) at 24.5 ± 0.5°C.

Conclusions:

The Pow value for the major component in Setafix X 11342 is 18000 (log Pow= 5.2) at 24.5 ± 0.5 °C.

Executive summary:

The Pow value for the major component in Setafix X 11342 is 18000 (log Pow = 5.2) at 24.5 ± 0.5 °C.

Cross-reference 8

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

partition coefficient

Type of information:

(Q)SAR

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model and falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

1. SOFTWARE
EPI Suite (US EPA) 4.1
2. MODEL (incl. version number)
KOWWIN v1.68
3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL
OCCCCCCO
4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
- Defined endpoint:
Partition coefficient (n-Octanol/Water)
- Unambiguous algorithm:
Results of the two successive multiple regressions (first for atom/fragments and second for correction factors) yield the following general equation for estimating log P of any organic compound: log P = Σ(fini ) + Σ(cjnj ) + 0.229 (num = 2447, r2 = 0.982, std dev = 0.217, mean error = 0.159)
- Defined domain of applicability:
Log P estimates are less accurate for compounds outside the MW range (18.02 - 199.98) of the training set compounds, and/or that have more instances of a given fragment than the maximum for all training set compounds.
- Appropriate measures of goodness-of-fit and robustness and predictivity:
R² = 0.94
- Mechanistic interpretation:
KOWWIN uses a "fragment constant" methodology to predict log P. In a "fragment constant" method, a structure is divided into fragments (atom or larger functional groups) and coefficient values of each fragment or group are summed together to yield the log P estimate. KOWWIN’s methodology is known as an Atom/Fragment Contribution (AFC) method. Coefficients for individual fragments and groups were derived by multiple regression of 2447 reliably measured log P values. KOWWIN’s "reductionist" fragment constant methodology (i.e. derivation via multiple regression) differs from the "constructionist" fragment constant methodology of Hansch and Leo (1979) that is available in the CLOGP Program (Daylight, 1995).
5. APPLICABILITY DOMAIN
- Descriptor domain:
Molecular weight = 118.18
- Structural and mechanistic domains:
Functional groups are within the domain of the model.
-CH2- (aliphatic carbon) 6 fragments (model maximum: 28)
-OH (hydroxy, aliphatic attach) 2 fragments (model maximum: 9)
- Multi-alcohol correction 1 factor (model maximum: 1)
- Similarity with analogues in the training set:
There are close structural analogues in the training set.
6. ADEQUACY OF THE RESULT
The prediction is reliable and log Kow is required for risk assessment and classification and labelling purposes.
This endpoint study record is supporting information reported within the read-across justification.

Guideline:

other: KOWWIN v1.68 (September 2010)

Principles of method if other than guideline:

Software tool(s) used including version: EPI Suite (US EPA) 4.1
- Model(s) used: KOWWIN v1.68 (September 2010)
- Model description: see field 'Justification for type of information'
- Justification of QSAR prediction: see field 'Justification for type of information'

GLP compliance:

no

Type of method:

calculation method (fragments)

Partition coefficient type:

octanol-water

Specific details on test material used for the study:

SMILES: OCCCCCCO

Type:

log Pow

Partition coefficient:

0.765

Remarks on result:

not measured/tested

Conclusions:

The EPI Suite (US EPA) Version 4.1 model KOWWIN v1.68 (September 2010) estimated the Log Kow to be 0.7648.

Cross-reference 9

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

partition coefficient

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Guideline:

other: Unknown, textbook value

Principles of method if other than guideline:

- Principle of test: Determine octanol/water partition coefficient
- Short description of test conditions: Unknown, textbook value
- Parameters analysed / observed: octanol/water partition coefficient

GLP compliance:

not specified

Type of method:

other: Not specified

Partition coefficient type:

octanol-water

Specific details on test material used for the study:

CAS Number: 121-33-5

Type:

log Pow

Partition coefficient:

1.21

Remarks on result:

other: textbook value

Conclusions:

The reference reports a Log Kow of 1.21.

Cross-reference 10

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

partition coefficient

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Guideline:

other: Unknown, textbook value

Principles of method if other than guideline:

- Principle of test: Determine octanol/water partition coefficient
- Short description of test conditions: Unknown, textbook value
- Parameters analysed / observed: octanol/water partition coefficient

GLP compliance:

not specified

Type of method:

other: Not specified

Partition coefficient type:

octanol-water

Specific details on test material used for the study:

CAS Number: 100-09-4

Type:

log Pow

Partition coefficient:

1.96

Remarks on result:

other: textbook value

Conclusions:

The reference reports a Log Kow of 1.96.

Cross-reference 11

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

dissociation constant

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Guideline:

other: textbook reference

Principles of method if other than guideline:

- Principle of test: Determine dissociation constant
- Short description of test conditions: Unknown, textbook value
- Parameters analysed / observed: dissociation constant

GLP compliance:

not specified

Specific details on test material used for the study:

4-Methoxybenzoic acid

Dissociating properties:

yes

pKa:

4.5

Temp.:

25 °C

Remarks on result:

other: textbook reference

Conclusions:

The reference reports the pKa for the substance as 4.50 at 25°C.

Cross-reference 12

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

dissociation constant

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Guideline:

other: OECD QSAR Toolbox reference

Principles of method if other than guideline:

- Principle of test: Determine dissociation constant
- Short description of test conditions: Unknown, textbook value
- Parameters analysed / observed: dissociation constant

GLP compliance:

not specified

Specific details on test material used for the study:

CAS Number 121-33-5

Dissociating properties:

yes

pKa:

7.4

Temp.:

25 °C

Remarks on result:

other: textbook reference

Conclusions:

The reference reports the pKa for the substance as 7.4 at 25°C.

Cross-reference 13

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

06/07/2000

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study according to OECD test method and to GLP standards

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Deviations:

no

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

yes

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: approx 8 weeks old
- Weight at study initiation: +/- 20% of the sex mean (Mean of 190g for females, 303g for males on Day 1)
- Fasting period before study: Overnight, maximum of 20 hours prior to dosing until approx 3-4 hours after administration of the test substance.
- Housing: 3 animals per sex per cage in labelled polycarbonate cages containing purified sawdust as bedding material.
- Diet (e.g. ad libitum): Free access to standard pelletd laboratory animal diet (from Carfil Quality BVBA, Oud-Turnhout, Belgium)
- Water (e.g. ad libitum): Free access to tap water.
- Acclimation period: 5 days before start of treatment

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21Centigarde
- Humidity (%): 50% relative
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12 hours artificial fluorescent light and 12 hours dark per day.

Route of administration:

oral: gavage

Vehicle:

propylene glycol

Remarks:

SG 1.036

Details on oral exposure:

Maximum dose volume applied: 2000mg/kg

Doses:

Single dose on day one. 2000mg/kg (10ml/kg) body weight

No. of animals per sex per dose:

1 group of 3 males and 1 group of 3 females, all with the same 2000mg/kg dose.

Control animals:

no

Details on study design:

- Duration of observation period following administration: 15 days
- Frequency of observations and weighing: Twice daily for mortaility/viability and Body weights on days 1, 8 and 15
- Necropsy of survivors performed: Yes
- Other examinations performed: clinical signs, body weight and macroscopic findings

Statistics:

No statistical analysis was performed.

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Sex:

male

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No Mortality occured

Clinical signs:

other: No clinical signs were noted

Other findings:

Macroscopic findings: No abnormalities were found at macroscopic post mortem examination of the animals

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The oral LD50 value of 1070 GA 051 in Wistar rats was established to exceed 2000mg/kg body weight.
Based on these results and according to EC criteria for classification and labelling requirements for dangerous substances and preparations (Guidelines in commision Directive 93/21/EEC), 1070 GA 051 does not have to be classified and has no obligatory labelling requirement for oral toxicity.

Executive summary:

The oral LD50 value of 1070 GA 051 in Wistar rats was established to exceed 2000mg/kg body weight.

Based on these results and according to EC criteria for classification and labelling requirements for dangerous substances and preparations (Guidelines in commision Directive 93/21/EEC), 1070 GA 051 does not have to be classified and has no obligatory labelling requirement for oral toxicity.

Cross-reference 14

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

GLP compliance:

no

Test type:

standard acute method

Limit test:

no

Species:

rat

Strain:

Osborne-Mendel

Sex:

male/female

Details on test animals or test system and environmental conditions:

Groups of 10 young adult Osborne-Mendel rats evenly divided by sex were fasted for approximately 18 hr prior to treatment.

Animals had access to water at all times, and the food was replaced in cages as soon as animals received their respective doses.

Route of administration:

oral: gavage

Vehicle:

propylene glycol

Remarks:

20%

Details on study design:

All animals were maintained under close observation for recording toxic signs and time of death. Such observation was continued until animals appeared normal and showed weight fain. The usual observation period was 2 weeks.

Statistics:

LD50'S were computed by the method of Litchfield & Wilcoxon.

Sex:

male/female

Dose descriptor:

LD50

Effect level:

1 580 mg/kg bw

Based on:

test mat.

95% CL:

1 390 - 1 810

Clinical signs:

other: Coma soon after treatment Death Time 4 hr - 4 days

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

The acute oral LD50 of vanillin is 1580 mg/kg bw in rats.

Cross-reference 15

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

acute toxicity: oral

Type of information:

(Q)SAR

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model and falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

1. SOFTWARE: US EPA Toxicity Estimation Software Tool (T.E.S.T.) Version 4.2

2. MODEL: Predicted Oral rat LD50 for 121-33-5 from Consensus method

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL: CAS Number 121-33-5

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
- Defined endpoint: Oral rat LD50 mg/kg
- Unambiguous algorithm: It was not possible to develop a single model or a group contribution model that fit the entire training set. The consensus method achieved the best results in terms of both prediction accuracy and prediction coverage. The prediction statistics for this endpoint were not as good as those for the other endpoints. This is not surprising because this endpoint has a higher degree of experimental uncertainty and has been shown to be more difficult to model than other endpoints
- Defined domain of applicability:
Compounds can only contain the following element symbols: C, H, O, N, F, Cl, Br, I, S, P, Si, or As
Compounds must represent a single pure component
- Appropriate measures of goodness-of-fit and robustness and predictivity:
Mean absolute error in -Log10(mol/L): Entire set 0.43
Mean absolute error in -Log10(mol/L): Similarity coefficient (≥ 0.5) 0.25

5. APPLICABILITY DOMAIN
- Descriptor domain:
10 analogues formed the dataset
Compound is a single pure component containing C, H, O
- Similarity with analogues in the training set:
The target substance is similar to the analogues within the training set (similarity coefficient = 0.25).

6. ADEQUACY OF THE RESULT
The prediction provides a discrete acute oral toxicity prediction and there are similar analogues within the training set.

Guideline:

other: QSAR prediction

Principles of method if other than guideline:

- Software tool(s) used including version: US EPA Toxicity Estimation Software Tool (T.E.S.T.) Version 4.2
- Model(s) used: Consensus method
- Model description: see field 'Justification for non-standard information', 'Attached justification' and/or 'Cross-reference'
- Justification of QSAR prediction: see field 'Justification for type of information', 'Attached justification' and/or 'Cross-reference'

GLP compliance:

not specified

Test type:

other: not specified

Specific details on test material used for the study:

CAS Number: 121-33-5

Species:

rat

Strain:

not specified

Sex:

not specified

Route of administration:

oral: unspecified

Vehicle:

not specified

Sex:

not specified

Dose descriptor:

LD50

Effect level:

1 930 mg/kg bw

Based on:

test mat.

Remarks on result:

other: predicted value

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

The predicted value for the oral rat LD50 is 1930 mg/kg bw.

Cross-reference 16

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

acute toxicity: oral

Type of information:

(Q)SAR

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model and falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

1. SOFTWARE: US EPA Toxicity Estimation Software Tool (T.E.S.T.) Version 4.2

2. MODEL: Predicted Oral rat LD50 for 100-09-4 from Consensus method

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL: CAS Number 100-09-4

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
- Defined endpoint: Oral rat LD50 mg/kg
- Unambiguous algorithm: It was not possible to develop a single model or a group contribution model that fit the entire training set. The consensus method achieved the best results in terms of both prediction accuracy and prediction coverage. The prediction statistics for this endpoint were not as good as those for the other endpoints. This is not surprising because this endpoint has a higher degree of experimental uncertainty and has been shown to be more difficult to model than other endpoints
- Defined domain of applicability:
Compounds can only contain the following element symbols: C, H, O, N, F, Cl, Br, I, S, P, Si, or As
Compounds must represent a single pure component
- Appropriate measures of goodness-of-fit and robustness and predictivity:
Mean absolute error in -Log10(mol/L): Entire set 0.43
Mean absolute error in -Log10(mol/L): Similarity coefficient (≥ 0.5) 0.33

5. APPLICABILITY DOMAIN
- Descriptor domain:
10 analogues formed the dataset
Compound is a single pure component containing C, H, O
- Similarity with analogues in the training set:
The target substance is similar to the analogues within the training set (similarity coefficient = 0.33).

6. ADEQUACY OF THE RESULT
The prediction provides a discrete acute oral toxicity prediction and there are similar analogues within the training set.

Guideline:

other: QSAR prediction

Principles of method if other than guideline:

- Software tool(s) used including version:
US EPA Toxicity Estimation Software Tool (T.E.S.T.) Version 4.2
- Model(s) used:
Consensus method
- Model description: see field 'Justification for non-standard information', 'Attached justification' and/or 'Cross-reference'
- Justification of QSAR prediction: see field 'Justification for type of information', 'Attached justification' and/or 'Cross-reference'

GLP compliance:

not specified

Test type:

other: not specified

Specific details on test material used for the study:

CAS Number: 100-09-4

Species:

rat

Strain:

not specified

Sex:

not specified

Route of administration:

oral: unspecified

Vehicle:

not specified

Sex:

not specified

Dose descriptor:

LD50

Effect level:

2 479 mg/kg bw

Based on:

test mat.

Remarks on result:

other: predicted value

Interpretation of results:

Category 5 based on GHS criteria

Conclusions:

The predicted value for the oral rat LD50 is 2479 mg/kg bw.

Cross-reference 17

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Deviations:

yes

Remarks:

missing details on test substance, all doses not reported, number of animals used was not clearly stated, no information on control animals was reported, full results not reported.

GLP compliance:

no

Remarks:

study pre-dates GLP requirements

Test type:

standard acute method

Limit test:

no

Specific details on test material used for the study:

1,6-Hexanediol

Species:

rat

Strain:

Sherman

Sex:

male

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Control animals:

not specified

Sex:

male

Dose descriptor:

LD50

Effect level:

3.73 mL/kg bw

95% CL:

2.68 - 5.21

Interpretation of results:

GHS criteria not met

Conclusions:

The acute oral toxicity is reported as 3.73 ml/kg bw.

Cross-reference 18

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20-02-2004 to 18-03-2004

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The study includes data generated according to generally valid and internationally accepted testing guidelines and performed according to GLP. The study was based on the following guideliens: -EC Directive 96/54/EEC, B.7 Repeated Dose (28 days) Toxicity (oral), 1996 -OECD 407, Repeated Dose 28-day Oral Toxicity Study in Rodents, 1995 -OPPTS 870.3050, Repeated dose 28-day Oral Toxicity Study in Rodents. Office of Prevention, Pesticides and Toxic Substances (7101), EPA 712-C-00-366,200.

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OPPTS 870.3050, Repeated dose 28-day oral toxicity study in rodents.

Deviations:

no

GLP compliance:

yes

Remarks:

22 July 2004 and 26 July 2004

Limit test:

no

Species:

rat

Strain:

other: Wistar Crl:(WI) BR (outbred, SPF-Quality)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: Approximately 6 weeks
- Weight at study initiation: approx. 213 g (Male) and approx. 170 g (Female)
- Housing: Group housing of 5 animals per sex per cage in stainless steel suspended cages with wire mesh floors (55 x 34 x 21.5 cm height). During activity monitoring, animals were individually housed overnight in Macrolon plastic cages (type III, height 15 cm) with sterilised sawduct (woody-Clean type 3/4, Tecnilab-BMI BV, Someren, The Netherlands) provided as bedding.
- Diet (e.g. ad libitum): Free access to standard pelleted laboratory animal diet (from Altromin (code VRF-1, Lage, Germany). Each batch is analysed for nutrients and contaminants are analysed on regular basis. Results of analysis for ingredients and/or contaminants of diet, sawdust were assessed and did not reveal any findings that were considered to have affected study integrity.
- Water (e.g. ad libitum): Free access to tap water. Results of analysis for water were assessed and did not reveal any findings that were considered to have affected study integrity.
- Acclimation period: At least 5 days before start of treatment under laboratory conditions

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.0 ± 3.0 (actual range 19.0 - 25.3)
- Relative Humidity (%): 30 - 70 (actual range: 32 - 58%)
- Air changes (per hr): approx. 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours artificial fluorescent light and 12 hours darkness per day

IN-LIFE DATES: From: 20 February 2004 To: 18 March 2004

Route of administration:

oral: gavage

Vehicle:

propylene glycol

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
Method of formulation: Formulations (w/w) were prepared daily within 4 hours prior to dosing (within approximately 5 hours on day 1) and were homogenised to a visually acceptable level. Adjustment was made for specific gravity of vehicle. The solutions were stored at ambient temperature.
Stability of test substance in vehicle was determined to be at least 4 hours (determined at NOTOX)

VEHICLE
- Vehicle: propylene glycol, specific gravity 1.03628 days
- Justification for use and choice of vehicle (if other than water): Rationale for vehicle was ased on trial formulations performed at NOTOX
- Concentration in vehicle: Analysis of the accuracy of dose preparations revealed values within the range of 96 - 102 % of nominal, which was considered to represent an acceptable level of accurancy for formulations of this type.
- Amount of vehicle (if gavage): 5 ml/kg body weight. Actual dose volumes were calculated weekly according to the latest body weight.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Samples of formulations prepared on day 14 were analysed to check homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations). Stability in vehicle over 4 hours was also determined (highest and lowest concentration). The nalytical method used was based on the results of a separate project for the development and validation of the analytical method (NOTOX project 331008).
Test substance formulations in propylene glycol were noted as stable for at least 4 hours and formed a homogeneous suspension at the concentrations tested. Analysis of the accuracy of dose preparations revealed values within the range of 96 - 102 % of nominal, which was considered to reresent an acceptable level of accurancy for formulations of this type.

Duration of treatment / exposure:

28 days

Frequency of treatment:

Once daily for at least 28 days , 7 days per week, approximately the same time as each day with a maximum of 4 hours difference between the earliest and latest dose. Animals were dosed up to the day prior to necropsy.

Remarks:

Doses / Concentrations:
0 (vehicle)
Basis:
other: Vehicle

Remarks:

Doses / Concentrations:
50 mg/kg/day
Basis:
nominal in diet

Remarks:

Doses / Concentrations:
150 mg/kg/day
Basis:
nominal in diet

Remarks:

Doses / Concentrations:
1000 mg/kg/day
Basis:
nominal in diet

No. of animals per sex per dose:

5 animals per sex per dose

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale:
The dose levels were selected on the basis of a 5-day dose range finding study (NOTOX Project 399061). based on the results of this range finding study, dose levels selected for the main study (28 days toxicicty study) were: 50, 150 and 1000 mg/kg body weight/day

- Rationale for animal assignment:
20 males, 20 females (females were nulliparous and non-pregnant)
Animals were randmly assigned prior to comencement of treatment, by computer-generated random algorithm according to body weight, with all animals within ± 20 % of sex mean.

Positive control:

Not applicable

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Once prior to start of treatment and on a weekly basis thereafter.

MORTALITY/VIABILITY
At least twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: At least once daily, detailed clinical observations were made in all animals. Once prior to start of treatment and on a weekly basis thereafter, this was also performed outside the home cage in a standard arena. The symptoms were graded according to fixed scales and the time of onset, degree and duration were recorded:
Maximum grade 1: grade 0 = absent, grade 1 = present
Maximum grade 3 or 4: grade 1 = slight, grade 2 = moderate, grade 3 = severe, grade 4 = very severe

NEUROBEHAVIOURAL EXAMINATION: Yes
- During week 4 of treatment, the following tests were performed on all animals:
- Hearing ability, Pupillary reflex, Static righting reflex and Grip strength (Score 0 = normal/present, score 1 = abnormal/absent)
- Motor Activity Test (recording period: 12 hours during overnight for individual animals). Motor activity measurements for males commenced on the last day of week 3.

BODY WEIGHT: Yes
- Time schedule for examinations: On days 1, 8, 15, 22 and 28

FOOD CONSUMPTION: Weekly

WATER CONSUMPTION: No
- Subjective appraisal was maintained during the study, but no quantitative investigation introduced as no effect was suspected

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Immediately prior to scheduled post mortem examination, between 7.30 and 10.30 a.m
- Anaesthetic used for blood collection: Yes, under iso-flurane anaesthesia
- Animals fasted: Yes (overnight, with a maximum of 20 hours)
- How many animals: 40
- Parameters examined: Erythrocytes count, Haemoglobin, Haematocrit, Mean corpuscular volume, Mean corpuscular haemoglobin, Mean corpuscular haemoglobin concentration, Platelet count, Red cell distribution width, Differential leucocyte count (neutrophils, lymphocytes, monocytes, eosinophils, basophils).

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Immediately prior to scheduled post mortem examination, between 7.30 and 10.30 a.m
- Anaesthetic used for blood collection: Yes, under iso-flurane anaesthesia
- Animals fasted: Yes (overnight, with a maximum of 20 hours)
- How many animals: 40
- Parameters examined: Alanine aminotransferase, alkaline phosphatase, Aspartate aminotransferase, total Bilirubin, Chloride, total Cholesterol, Creatine, Glucose, Phosphorus, total protein, Protein albumin, Urea, Calcium, Potassium and Sodium.

URINALYSIS: No data

CLOTTING POTENTIAL: Yes.
Parameters examined: Prothrombine, Partial throboplastin time

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

Necropsy:
All animals survivinf to the end of the observation period were deeply anaesthetised using iso-flurane and subsequently exsanguinated. All animals assigned to the study were necropsied and descriptions of all macroscopic abnormalities recorded.

Organs weights:
The following organ weights (and terminal body weight) were recorded from the animals on the scheduled day of necropsy: Adrenal glands, Brain, Liver, Heart, Kidneys, Spleen, Testes, Thymus and Epididymides

HISTOPATHOLOGY: Yes
The following slides were examined by a pathologist:
- all tissues and organs collected at the scheduled sacrifice from all animals of the control and the highest dose group
- all gross lesions of all animals
All abnormalities were described and documented.

Statistics:

The following statistical methods were used to analyse the data:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test (many-to-one test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control groups for each sex. The Student's t-test was applied for motor activity data.
- The Steel-test (many-to-one rank test) was applied when the data could not be assumed to follow a normal distribution.
- The exact Fisher-test was applied to frequency data.

All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance. Group means were calculated for continuous data and medians were calculated for discrete data (scores) in the summary tables. Test statistics were calculated on the basis of exact values for means and pooled variances. Individual values, means and standard deviations may have been rounded off before before printing.. Therefore, two groups may display the same printed means for a given parameter, yet dispaly different test statistics values.

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Weight gains of high dose males were significantly lower than those of male controls on days 8 and 22 (p< 0.05). Body weights and body weight gain of other groups of treated animals remained in the same range as controls over the 4-week study period.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

Food consymption before or after allowance for body weight was similar between treated and control animals.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not examined

Description (incidence and severity):

Subjective appraisal was maintained during the study, but no quantitative investigation introduced as no effect was suspected.

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Description (incidence and severity):

slight chnages observed were considered to be of no toxicological significance.

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

No dose-related trend was apparent and any deviations were considered to be of no toxicological relevance.

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

No toxicologically significant changes were noted

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

No toxicologically significant changes were noted

Gross pathological findings:

no effects observed

Description (incidence and severity):

No toxicologically relevant changes were noted

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

No toxicologically significant changes were noted

Histopathological findings: neoplastic:

not specified

Details on results:

MORTALITY
No mortality occurred during the study.

CLINICAL SIGNS
There were no clinical signs over the 28-day observation period that were considered to be related to treatment.

Incidental findings that were noted included watery discharge from and/or dullness of the eys, alopecia on the neck, broken tail apex, diarrhoea and chromodacryorrhoea. These findings are occasionally noted in rats of this age and strain which are housed and treated under the conditions in this study. At incidence observed, these were considered signs of no toxicological significance. No clinical signs were shown by males at 150 mg/kg/day, females at 1000 mg/kg/day and control females.

NEUROBEHAVIOUR
Increased motor activity recordings were obtained for three high dose males (nos. 17, 18 and 20) and two high dose females treated with Setafix X 11342, when compared to control animals. Motor activity data between other treated and control animals were similar.

BODY WEIGHT AND WEIGHT GAIN
Slightly lower total body weight gains were noted in both genders at the high dose. Weight gains of high dose males were significantly lower than those of male controls on days 8 and 22 (p < 0.05).
Body weights and body weight gain of other groups of treated animals remined in the same range as conctrols over the 4-week study period.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
Food consumption before or after allowance for body weight was similar between treated and control animals.

WATER CONSUMPTION
Subjective appraisal was maintained during the study, but no quantitative investigation introduced as no effect was suspected.

HAEMATOLOGY
A higher prothrombin time (PT) was measured in high dose males (p < 0.05). However, the control mean was slightly low and the mean was within the normal range of values for this type of study. The statistically lower partial thromboplastin time (PTT) in males at 50 mg/kg/day occurred in the absence of a treatment-related distribution. Therefore, this change was consiered to be of no toxicological significance. No further differences were noted in haematology parameters between treated and control rats.

CLINICAL CHEMISTRY
The following deviations in clinical biochemistry parameters distinguished high dose females from their controls:
- Increased alanine aminotransferase activity (ALAT; p < 0.05);
- Increased alkaline phosphatase activity (ALP; non-significant);
- Increased urea levels in female nos. 37 and 40 (p < 0.05);
- Increased glucose level in female no. 37 ( mean non-significant);
- Slightly increased inorganic phospate level (non-significant);
- Lower total protein level (non-significant).

The statistically significant lower total protein and albumin levels among males at 50, 150 and/or 1000 mg/kg/day were considered to be related to the slightly high control levels of these parameters. Also, no dose-related trend was apparent. In one high dose male (no. 19) a very low platelet count was measured. Since other animals of this dose group displayed normal values for this parameter, this was considered to have occurred incidentally. Therefore, these deviations were considered to be of no toxicological relevance.

ORGAN WEIGHTS
No toxicologically significant changes were noted regarding organ weights and organ:body weight ratios.
Slight changes in organ weights of high dose males (e.g. liver weights) were considered to be related to slightly lower terminal body weights. These changes were therefore considered not to be a sign of toxicity.

GROSS PATHOLOGY

- Macroscopic Examination:
Necropsy did not reveal any toxicologically relevant alterations.
Incidental findings among control and/or treated animals included enlargement of mandibular lymp nodes, pelvic dilation of the kidneys, a fractured tip of the tail, diaphragamatic hernia of the left lateral lobe of the liver, fluid in the uterus and reddish discolouration of the thymus. These findings are occasionally seen among rats used in these types of studies. In the absence of a treatment-related distribution they were considered changes of no toxicological significance.
No necropsy findings were noted in females at 50 mg/kg/day and above.

- Microscopic Examination:
There were no microscopic findings recorded which could be attributed to treatment with the test substance.
All microscopic findings were within the range of background pathology encountered in Wistar rats of this age and strain and occurred at similar incidencies and severity in both control and treated rats.

Dose descriptor:

NOAEL

Effect level:

150 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: see 'Remark'

Critical effects observed:

not specified

RESULTS:

Accuracy, homogeneity and stability over 4 hours of formulation of test substance in propylene glycol were demonstrated by analyses.

Findings that distinguished treated from control animals included:

50 mg/kg/day:

No treatment-related findings noted

150 mg/kg/day:

Increased individual motor activity recordings

1000 mg/kg/day:

- Increased individual motor activity recordings

- Slightly low weight gain

- Increased prothrombin time

- Deviations in clinical biochemistry parameters in females included:

- Increased alanine aminotransferase activity;

- Increased alkaline phosphatase activity;

- Increased urea levels in individual females;

- Increased glucose level in one female;

- Slightly increased inorganic phosphate level;

- Lower total protein level.

Conclusions:

Based on the results of this study and in the absence of functional or morphological disturbances supporting the higher motor activity recordings for individual females at 150 mg/kg/day, a No Observed Adverse Effect Level (NOAEL) for Setafix X 11342 of 150 mg/kg/day was established.

Executive summary:

Subacute 28 -day oral toxicity with Setafix X 11342 by daily gavage in the rat.

The nature and purpose of this sub-acute toxicity study was to assess the toxic potential of the test substance when administered to rats by oral gavage for a period of 28 days. A No Observed Adverse Effect Level (NOAEL) was evaluated. The study should provide a part of a rational basis for toxicological risk assessment in man. The oral route was selected as it is a possible route of human exposure during manufacture, handling or use of the test substance.

Based on the results of a 5 -day range finding study, the dose levels for the 28 -day toxicity study were selected to be 0, 50, 150 and 1000 mg/kg/day.

The study was based on the following guidelines:

- EC Directive 96/54/EEC, B.7 Repeated Dose (28 days) Toxicity (oral), 1996.

- OECD 407, Repeated Dose 28 -day Oral Toxicity Study in Rodents, 1995

- OPPTS 870.3050, Repeated Dose 28 -day Oral Toxicity Study in Rodents. Office of Prevention, Pesticides and Toxic Substances (7101), EPA 712 -C-00 -366, 2000.

The test substance was administered daily for 28 days by oral gavage to SPF-bred Winstar rats.

One control group and three treated groups were tested, each consisting of 5 males and 5 females.

The following parameters were evaluated: clinical signs daily; functional observation tests; body weight and food consumption weekly; clinical pathology and mcroscopy at termination; organ weights and histopathology on a selection of tissues.

No in-life observations indicative of toxicity were obtained except for a slightly low weight gain for high dose animals during treatment. This change was, however, not accompanied by concurrent changes in food intake. Also, increased motor activity recordings were obtained for individual high dose males, and females at 150 and 1000 mg/kg/day. However, concurrent and similar changes of the low or high sensors were not always apparent and there was no supportive clinical signs of hyperactivity. A relationship to treatment was considered to be uncertain.

There were no changes at determination of clinical appearance, performance of functional observations, food consumption measurements, macroscopic examination, organ weight determination and microscopic examination that were considered to be an effect of treatment.

Based on the results of this study and in the absence of functional or morphological disturbances supporting the higher motor activity recordings for individual females at 150 mg/kg/day, a No Observed Adverse Effect Level (NOAEL) for Setafix X 11342 of 150 mg/kg/day was established.

Cross-reference 19

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 452 (Chronic Toxicity Studies)

GLP compliance:

no

Remarks:

pre-dates GLP

Limit test:

no

Species:

rat

Sex:

male/female

Route of administration:

oral: feed

Clinical signs:

no effects observed

Gross pathological findings:

no effects observed

Dose descriptor:

NOAEL

Effect level:

>= 250 - <= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Conclusions:

When three groups of 24 rats (twelve of each sex) received 0.5-2% vanillin in the diet for 2 yr [equivalent to about 250-1000 mg/kg bw/day], no adverse clinical effects were evident, and gross and microscopic examination of the major organ tissues did not reveal any abnormalities.

Cross-reference 20

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification.

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male/female

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Duration of treatment / exposure:

Hexanediol by gavage for 28 days in compliance with OECD test method 407 to assess the effect of hexanediol with respect to repeated toxicity and for 28 days (males) / 42 days (females) in compliance with OECD test method 421.

Dose / conc.:

100 mg/kg bw/day (nominal)

Dose / conc.:

400 mg/kg bw/day (nominal)

Dose / conc.:

1 000 mg/kg bw/day (nominal)

Control animals:

yes

Clinical signs:

effects observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Slight changes in body weight and body weight gain at 1000 mg/kg b.w./d were observed always only in one sex: fermales (OECD 407) and males (OECD 421). This effect is assessed as a borderline effect of questionable toxicological relevance because of the following reasons: only one sex (either males or females) is affected; there is no correlation to foods consumption, no changes in hematology, clinical chemistry and histopathology were found.

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Dose descriptor:

NOAEL

Effect level:

400 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

Conclusions:

In valid OECD studies, tested up to the highest recommended dose of 1,000 mg/kg b.w. hexanediol revealed no effects of toxicological relevance beside a borderline effect on body weight either in males (OECD 421) or in females (OECD 407). NOAEL = 400 mg/kg b.w..

Cross-reference 21

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

30 October 2001 - 30 November 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The study includes data generated according to generally valid and internationally accepted testing guidelines and performed according to GLP. The study was based on the following guidelines: -Organisation for Economic Co-operation and Development (OECD), OECD Guidelines for Testing of Chemicals; Guideline no. 471: "Genetic Toxicology: Bacterial Reverse Mutation Test". (Adopted July 21, 1997). - European Economic Community (EEC). Adapting to technical progress for the twenty-sixth time Annex V of the EEC Directive 67/548/EEC, Part B: Methods for the Determination of Toxicity; B.13/14: "Muta1~enicity: "Reverse Mutation Assay using bacteria". EEC Publication Commission Directive {Published June 8, 2000).

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

GLP compliance:

yes

Remarks:

GLP Statement date: 10/12/2001 and 21/12/2001

Type of assay:

bacterial reverse mutation assay

Target gene:

Salmonella typhimurium bacteria and Escherichia coli bacteria

Species / strain / cell type:

other: Salmonella typhimurium TA98, TA100, TA1535, TA1537 and Escherichia coIi WP2uvrA

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

S9-mix

Test concentrations with justification for top dose:

DOSE RANGE FINDING TEST:
3, 10, 33, 100, 333, 1000, 3330 and 5000 μg/plate in the absence and presence of 59-mix.

MUTATION ASSAY:

EXPERIMENT 1: 3, 10, 33, 100, 333 μg/plate in the absence and presence of 59-mix (Strains: TA 1535, TA 1537 and TA98)

EXPERIMENT 2: 3, 10, 33, 100, 333 μg/plate in the absence and presence of 59-mix (Strains: TA 1535, TA 1537, TA98, TA 100 and WP2uvrA)

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Dimethyl sulfoxide (DMSO

- Justification for choice of solvent/vehicle:
Not specified but likely to be solubility of test substance

Untreated negative controls:

yes

Remarks:

Dimethyl sulfoxide

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Remarks:

Strain TA1535

Positive control substance:

sodium azide

Remarks:

Without metabolic activation(-S9-mix)

Untreated negative controls:

yes

Remarks:

Dimethyl sulfoxide

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Remarks:

Strain TA1537

Positive control substance:

9-aminoacridine

Remarks:

Without metabolic activation(-S9-mix)

Untreated negative controls:

yes

Remarks:

Dimethyl sulfoxide

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Remarks:

Strain TA98

Positive control substance:

other: Daunomycine

Remarks:

Without metabolic activation(-S9-mix)

Untreated negative controls:

yes

Remarks:

Dimethyl sulfoxide

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Remarks:

Strain TA100

Positive control substance:

methylmethanesulfonate

Remarks:

Without metabolic activation(-S9-mix)

Untreated negative controls:

yes

Remarks:

Dimethyl sulfoxide

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Remarks:

Strain WP2uvrA

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

Without metabolic activation(-S9-mix)

Untreated negative controls:

yes

Remarks:

Dimethyl sulfoxide

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Remarks:

Strains: TA1537, TA1535, TA98, TA100 and WP2uvrA

Positive control substance:

other: 2-aminoanthracene

Remarks:

With metabolic activation (+S9-mix)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

Preparation of Bacterial cultures:
Samples of frozen stock cultures of bacteria were transferred into enriched nutrient broth (Oxoid no. 2) and incubated in a shaking incubator (37°C, 150 spm), until the cultures reached an optical density of 1.0 ± 0.1 at 700 nm (109 cells/ml}. Freshly grown cultures of each strain were used for a test.

Permeabilization of the Escherichia coli strain:
WP2uvrA bacteria were washed twice in 0.25 the original volume of ice-cold 0.12 M Tris-HCL buffer pH 8.0, then gently resuspended in 0.2 vol. 0.12 M TrisHCL, 0.5 mM EDTA pH 8.0, and shaken for 2.5 min at 37°C. MgCl2 was then added to a final concentration of 10 mM. The cells were centrifuged and resuspended in the original volume of nutrient broth.

Agar Plates:
Agar plates (0 9 cm) contained 25 ml glucose agar medium. Glucose agar medium contained per liter: 18g purified agar (Oxoid, code L2) in Vogel-Bonner Medium E, 20 g glucose.
N.B. The agar plates for the test with the Salmonella typhimurium strains also contained 12.5 μg/plate biotin and 15 μg/plate histidine and the agar plates for the test with the Escherichia coli strain contained 15 μg/plate tryptophan.

Top Agar:
Top agar medium, containing 0.6% (w/v) purified agar and 0.5% (w/v) NaCl, was heated to dissolve the agar. Samples of 3 ml top agar were transferredinto 10 ml glass tubes with metal caps. Top agar tubes were autoclaved for 20 min at 121 ± 1 °C.

Environmental conditions:
All incubations were carried out in the dark at 37 ± 1 °C. The temperature was monitored during the experiment.

Evaluation criteria:

A test substance is consideried negative (not mutagenic) in the test if:

a) The total number of reveirtants in any tester strain at any concentration is not greater than two times the solvent control value, with or without metabolic activation.

b) The negative response should be reproducible in at least one independently repeated experiment.

A test substance is considered positive (mutagenic) in the test if:

a) It induces a number of revertant colonies, dose related, greater than two-times the number of revertants induced by the solvent control in any of the tester strains, either with or without metabolic activation.
However, any mean plate count of less than 20 is considered to be not significant.

b) The positive response should be reproducible in at least one independently repeated experiment.

The preceding criteria were not absolute and other modifying factors might enter into the final evaluation decision.

Statistics:

No formal hypothesis testing was done.

Species / strain:

other: Salmonella typhimurium TA98, TA100, TA1535, TA1537 and Escherichia coIi WP2uvrA

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

DOSE RANGE FINDING TEST

SETAFIX X 11342 was testeid in the tester strains TA100 and WP2uvrA with concentrations of 3, 10, 33, 100, 333, 1000, 3330 and 5000 μg/plate in the absence and presence of 59-mix.

Precipitate:

The test substance precipitated in the top agar at concentrations of 100 μg/plate and upwards. Precipitation of SETAFIX X 11342 on the plates was observed at the start and at the end of the incubation period at concentrations of 333 μg/plate and upwards.

Toxicity:

To determine the toxicity of SETAFIX X 11342, the reduction of the bacterial background lawn, the increase in the size of the microcolonies and the reduction of the revertant colonies were examined.

No reduction of the bacterial background lawn and no biologically relevant decrease in the number of revertants were observed.

MUTATION ASSAY

Based on the results of the close range finding test, SETAFIX X 11342 was tested up to concentrations of 333 μg/plate in the absence and presence of S9-mix in two mutation assays.

The first mutation experiment was performed with the strains TA 1535, TA 1537 and TA98 and the second mutation experiment was performed with the strains TA 1535, TA 1537, TA98, TA 100 and WP2uvrA.

Precipitate:

SETAFIX X 11342 precipitated in the top agar at concentrations of 100 and 333 μg/plate. Precipitation of SETAFIX X 11342 on the plates was observed at the start and at the end of the incubation period in all tester strains at the highest concentration tested.

Toxicity

The bacterial background lawn was not reduced at all concentrations tested and no biologically relevant decrease in the number of revertants was observed.

Number of revertants:

All bacterial strains showed negative responses over the entire dose range, i.e. no dose-related, two-fold, increase in the number of revertants in two independently repeated experiments.

The negative control values were within our laboratory background historical control data ranges. The strain-specific positive control values were within our laboratory background historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly, except for TA1537 in the presence of S9 -mix (second experiment). However, since this value was just without the limit of the range, the validity of the test was considered to be not affected.

Conclusions:

Interpretation of results (migrated information):
negative

Based on the results of this study it is concluded that SETAFIX X 11342 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Executive summary:

 

The objective of this study was to evaluate the test substance for its ability to induce reverse mutations in a gene of histidine-requiring Salmonella typhimurium bacterial strains resulting in histidine-independent strains, and in a gene of tryptophan-requiring Escherichia coli bacterial strain resulting in a tryptophan-independent strain.

 

SETAFIX X 11342 was tested in the Salmonella typhimurium reverse mutation assay with four histidine-requiring strains of Salmonella typhimurium (TA1535, TA1537, TA100 and TA98) and in the Escherichia coli reverse mutation assay with a tryptophan-requiring strain of Escherichia coli WP2uvrA. The test was performed in two independent experiments in the presence and absence of S9-mix (Aroclor-1254 induced rat liver S9-mix).

 

In the dose range finding test, SETAFIX X 11342 was tested up to concentrations of 5000 μg/plate in the absence and presence of S9-mix in the strains TA100 and WP2uvrA. SETAFIX X 11342 precipita1ted on the plates at dose levels of 333 μg/plate and upwards. The bacterial background lawn was not reduced at all concentrations tested and no biologically relevant decrease in the number of revertants was observed.

 

In the first and in the second mutation assay, SETAFIX X 11342 was tested up to concentrations of 333 μg/plate in the absence and presence of S9-mix. SETAFIX X 11342 precipitated on the plates at this dose level. The bacterial background lawn was not reduced at all concentrations tested and no decrease in the number of revertants was observed.

 

The presence of 5 and 10% (v/v) liver microsomal activation did not influence these findings.

 

SETAFIX X 11342 did not induce a dose-related, two-fold increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA both in the absence and presence of S9-metabolic activation. These results were confirmed in an independently repeated experiment.

 

Based on the results of this study it is concluded that SETAFIX X 11342 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Cross-reference 22

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

(Q)SAR

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model and falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

1. SOFTWARE: US EPA Toxicity Estimation Software Tool (T.E.S.T.) Version 4.2

2. MODEL: Predicted Mutagenicity for 100-09-4 from Consensus method

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL: CAS Number 100-09-4

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
- Defined endpoint: bacterial reverse mutation assay
- Unambiguous algorithm: the consensus method achieved the best prediction accuracy (concordance) and prediction coverage. The single model and group contribution methods could not be applied to this endpoint. All of the methods achieved a nice balance of prediction sensitivity and specificity.
- Defined domain of applicability:
Compounds can not be salts, mixtures or ambiguous compounds
- Appropriate measures of goodness-of-fit and robustness and predictivity:
Concordance with similar chemicals: 0.90 (9 out of 10)
Sensitivity with similar chemicals: 0.83 (5 out of 6)
Specificity with similar chemicals: 1.00 (4 out of 4)

5. APPLICABILITY DOMAIN
- Descriptor domain:
10 analogues formed the dataset
- Similarity with analogues in the training set:
The target substance is similar to the analogues within the training set.

6. ADEQUACY OF THE RESULT
The prediction provides a discrete in vitro gene mutation study in bacteria result of negative and there are similar analogues within the training set.

Guideline:

other: QSAR prediction

Principles of method if other than guideline:

- Software tool(s) used including version: US EPA Toxicity Estimation Software Tool (T.E.S.T.) Version 4.2
- Model(s) used: Predicted Mutagenicity for 100-09-4 from Consensus method
- Model description: see field 'Justification for non-standard information', 'Attached justification' and/or 'Cross-reference'
- Justification of QSAR prediction: see field 'Justification for type of information', 'Attached justification' and/or 'Cross-reference'

GLP compliance:

no

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

CAS Number: 100-09-4

Species / strain:

not specified

Metabolic activation:

not specified

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Remarks on result:

mutagenic potential (based on QSAR/QSPR prediction)

Conclusions:

Based on the consensus method, the AMES mutagenicity Negative.

Cross-reference 23

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

Hexandiol

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Test concentrations with justification for top dose:

up to 5,000 μg/plate

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Conclusions:

Hexandiol was not mutagenic in the Ames test (OECD 471) when Salmonella typhimurium strains (TA 98, TA 100, TA 1535 and TA 1537) were exposed up to 5,000 μg/plate with and without metabolic activation.

Cross-reference 24

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

S. typhimurium TA 98

Species / strain / cell type:

S. typhimurium TA 100

Metabolic activation:

with and without

Test concentrations with justification for top dose:

0.05 to 1000 µg/plate

Vehicle / solvent:

DMSO

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Conclusions:

The mutagenesis assays were run at 0.05 to 1000 µg/pl for vanillin. Vanillin did not induceda number of revertants that was over half the number of spontaneous revertants of TA98 or TAlOO at the indicated dose ranges, either with or without S9 mix.

Cross-reference 25

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

21 January 2004 - 26 March 2004

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The study includes data generated according to generally valid and internationally accepted testing guidelines and performed according to GLP. The study was based on the following guidelines: - OECD Guidelines for Testing of Chemicals, Guideline no. 473: In Vitro Mammalian Chromosome Aberration Test (adopted 21st July 1997). - European Economic Community (EEC). Directive 2000/32/EC, Part B: Methods for the Determination of Toxicity; B.10: "Mutagenicity: In Vitro Mammalian Chromosome Aberration Test".

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Deviations:

yes

Remarks:

.The study integrity was not adversely affected by the deviations.

Qualifier:

according to guideline

Guideline:

EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)

Deviations:

yes

Remarks:

.The study integrity was not adversely affected by the deviations.

GLP compliance:

yes

Remarks:

GLP Statement Date: 01-04-2004

Type of assay:

in vitro mammalian chromosome aberration test

Target gene:

Cultured Human Lymphocytes

Species / strain / cell type:

lymphocytes: cultured peripheral human lymphocytes

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

S9-miix

Test concentrations with justification for top dose:

Dose Range Finding Test: 1, 3, 10, 33 and 100 μg/ml (culture medium with and without S9-mix)

First Cytogenetic Assay: 10, 33 and 100 μg/ml (culture medium with and without S9-mix)

Second Cytogenetic Assay: 10, 33 and 100 μg/ml (culture medium with and without S9-mix)

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Dimethyl Sulfoxide (DMSO)

- Justification for choice of solvent/vehicle:
Not specified but likely to be solubility

Untreated negative controls:

yes

Remarks:

Dimethyl sulfoxide

Negative solvent / vehicle controls:

yes

Remarks:

Dimethyl sulfoxide

True negative controls:

not specified

Positive controls:

yes

Remarks:

Hanks' Balanced Salt Solution (HBSS) without calcium and magnesium

Positive control substance:

mitomycin C

Remarks:

Without metabolic activation (-S9-mix)

Untreated negative controls:

yes

Remarks:

Dimethyl sulfoxide

Negative solvent / vehicle controls:

yes

Remarks:

Dimethyl sulfoxide

True negative controls:

not specified

Positive controls:

yes

Remarks:

Solvent for positive controls: Hanks' Balanced Salt Solution (HBSS) without calcium and magnesium

Positive control substance:

cyclophosphamide

Remarks:

With metabolic activation (+S91-mix)

Details on test system and experimental conditions:

TEST SYSTEM
Cultured peripheral human lymphocytes were used as test system. Peripheral human lymphocytes are recommended in international guidelines (e.g. EPA, OECD, EEC).
These stimulated human lymphocytes were used because they are sensitive indicators of clastogenic activity of a broad range of chemicals

Blood was collected from healthy adult, non-smoking male volunteers. The Average Generation Time (AGT) of the cells is presented below:
Dose range finding study: age 29, AGT = 15.8 h (Nov. 2003)
First cytogenetic assay: age 33, AGT = 14.2 h (Nov. 2003)
Second cytogenetic assay: age 38, AGT = 16.8 h (Feb. 2004)

CELL CULTURE:

Blood samples:
Blood samples were taken from healthy adult male volunteers by venapuncture using the Venoject multiple sample blood collecting system with a suitable size sterile vessel containing sodium heparin. Immediately after blood collection lymphocyte cultures were started.

Culture medium:
Culture medium consisted of RPMI 1640 medium (lnvitrogen Corporation, Breda, The Netherlands), supplemented with 20% (v/v) heat-inactivated (56°C; 30 min) foetal calf serum (lnvitrogen Corporation), L-glutamine (2 mM) {Merck), penicillin/streptomycin (50 U/ml and 50 μg/ml respectively) (lnvitrogen Corporation) and 30 U/ml heparin.

Lymphocyte cultures:
Whole blood (0.4 ml) treated with heparin was added to 5 ml or 4.8 ml culture medium {in the absence and presence of S9-mix respectively). Per culture 0.1 ml (9 mg/ml) phytohaemagglutinin (Murex, Dartford, England) was added.

Environmental conditions:
All incubations were carried out in a humid atmosphere (80-100%) containing 5 ± 0.5% C02 in air in the dark at 37 ± 1°C. ThE~ temperature, humidity and C02-percentage were monitored throughout the experiment.

Metabolic activation system:
Rat liver microsomal enzymes were routinely prepared from adult male Wistar rats (6), which were obtained from Charles River (Sulzfeld, Germany).

Evaluation criteria:

Data evaluation:
A test substance was considered positive (clastogenic) in the chromosome aberration test if:
a) it induced a dose-related statistically significant (Chi-square test, P < 0.05) increase in the number of cells with chromosome aberrations.
b) a statistically significant and biologically relevant increase in the frequencies of the number of cells with chromosome aberrations was observed in the absence of a clear dose-response relationship.

A test substance was considered negative (not clastogenic) in the chromosome aberration test if none of the tested concentrations induced a statistically significant (Chi-square test, P < 0.05) increase in the number of cellls with chromosome aberrations.

Acceptability of the assay:
A chromosome aberration test was considered acceptable if it met the following criteria:
a) The number of chromosome aberrations found in the solvent control cultures should reasonably be within the laboratory historical control data range.
b) The positive control substances should produce a statistically significant (Chi-square test, P < 0.05) increase in the number of cells with chromosome aberrations.
c) A homogeneous response between the replicate cultures is observed.

Statistics:

The incidence of aberrant cells (cells with one or more chromosome aberrations, inclusive or exclusive gaps) for each exposure group was compared to that of the solvent control using Chi-square statistics.

If P is small (P< 0.05) the hypothesis that the incidence of cells with chromosome aberrations is the same for both the treated and the solvent control group is rejected and the number of aberrant cells in the test group is considered to be significantly different from the control group at the 95% confidence level.

Species / strain:

lymphocytes: Cultured peripheral human lymphocytes

Metabolic activation:

with and without

Genotoxicity:

not determined

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: other: Cultured peripheral human lymphocytes

Remarks:

Migrated from field 'Test system'.

DOSE RANGE FINDING TEST

At a concentration of 100 μg/ml Setafix X 11342 precipitated in the culture medium. Therefore, a concentration of 100 μg/ml was used as the highest concentration of Setafix X 11342.

In the dose range finding test lt>lood cultures were treated with 1, 3, 10, 33 and 100 μg Setafix X 11342 /ml culture mE~dium with and without S9-mix.

FIRST CYTOGENETIC ASSAY

Based on the results of the dose range finding test the following dose levels were selected for the cytogenetic assay:

Without and with S9-mix: 10, 33 and 100 μg Setafix X 11342/ml culture medium (3 h 1:ixposure time, 24 h fixation time).

All dose levels were selected for scoring of chromosome aberrations. Both in the absence and presence of S9-mix Setafix X 111342 did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations.

SECOND CYTOGENETIC ASSAY

To obtain more information about the possible clastogenicity of Setafix X 11342, a second cytogenetic assay was performed in which human lymphocytes were continuously exposed to Setafix X 11342 in the absencE of S9 mix for 24 or 48 hours. In the· presence of S9-mix, cells were fixed after 48 hours following a 3 hour exposure to Setafix X 11342. The following dose levels were selected for the seicond cytogenetic assay:

Without S9-mix: 10, 33 and 100 μg Setafix X 11342/ml culture medium (24 hand 48 h exposure time, 24 hand 48 h fixation time)

With S9-mix: 10, 33 and 100 μg Setafix X 11342/ml culture medium (3 h exposure time, 48 h fixation time)

All doses were selected for scoring of chromosome aberrations. Both in the absence and presence of S9-mix Setafix X 11342 did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations.

Conclusions:

Interpretation of results (migrated information):
negative

The test is valid and that Setafix X 11342 is not clastogenic in human lymphocytes under the experimental conditions of this study.

Executive summary:

 

The objective of this study was to evaluate Setafix X 11342 for its ability to induce structural chromosome aberrations in cultured human lymphocytes, either in the presence or absence of a metabolic activation system (S9-mix).

 

The effect of Setafix X 11342 on the number of chromosome aberrations in cultured peripheral human lymphocytes in the presence and absence of a metabolic activation system (Aroclor-1254 induced rat liver S9-mix) is summarised. The possible clastogenicity of Setafix X 11342 was tested in two independent experiments.

 

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

- OECD Guidelines for Testing of Chemicals, Guideline no. 473: In Vitro Mammalian Chromosome Aberration Test (adopted 21st July 1997).

- European Economic Community (EEC). Directive 2000/32/EC, Part B: Methods for the Determination of Toxicity; B.10: "Mutagenicity: In Vitro Mammalian Chromosome Aberration Test".

 

Batch PP 03 of Setafix X 11342 was a light yellowish solid with a purity of >95%. The test substance was crushed in a test tube and dissolved in dimethyl sulfoxide at concentrations of 33 mg/ml and lower.

 

In the first cytogenetic assay, Setafix X 11342 was tested up to 100 μg/ml for a 3 h exposure time with a 24 h fixation time in the absence and presence of S9-mix. Setafix X 11342 precipitated in the culture medium at this dose level.

 

In the second cytogenetic assay, Setafix X 11342 was tested up to 100 μg/ml for a 24 hand 48 h continuous exposure time with a 24 h and 48 h fixation time in the absence of S9-mix. In the presence of 1.8% (v/v) S9-fraction Setafix X 11342 was also tested up to 100 μg/ml for a 3 h exposure time with a 48 h fixation time.

 

Positive control chemicals, mitomycin C and cyclophosphamide, both produced a statistically significant increase in the incidence of cells with chromosome aberrations, indicating that the test conditions were adequate and that the metabolic activation system (S9-mix) functioned properly.

 

Setafix X 11342 did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in the absence and in the presence of S9-mix, in two independently repeated experiments.

 

Finally, it is concluded that this test is valid, and that Setafix X 11342 is not clastogenic in human lymphocytes under the experimental conditions of this study.

Cross-reference 26

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

GLP compliance:

not specified

Type of assay:

in vitro mammalian chromosome aberration test

Specific details on test material used for the study:

Hexandiol

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Species / strain:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Conclusions:

Hexandiol was not mutagenic for chromosome aberration according to OECD 473 performed with the same cell line with and without S9 mix.

Cross-reference 27

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / bone marrow chromosome aberration

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study without detailed documentation

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 475 (Mammalian Bone Marrow Chromosome Aberration Test)

GLP compliance:

not specified

Type of assay:

mammalian bone marrow chromosome aberration test

Species:

mouse

Strain:

not specified

Sex:

male

Route of administration:

oral: gavage

Dose / conc.:

500 mg/kg bw/day (nominal)

No. of animals per sex per dose:

3 male mice

Sex:

male

Genotoxicity:

negative

Toxicity:

not specified

Vehicle controls validity:

not specified

Negative controls validity:

not specified

Positive controls validity:

not specified

Conclusions:

A dose of 500 mg/kg given by stomach tube did not induce chromosome damage (micronuclei) in the bone marrow of three male mice.

Cross-reference 28

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Study period:

04 Jul 2017 to 04 Sep 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 490 (In Vitro Mammalian Cell Gene Mutation Tests Using the Thymidine Kinase Gene)

GLP compliance:

yes

Type of assay:

in vitro mammalian cell gene mutation tests using the thymidine kinase gene

Specific details on test material used for the study:

Identification: Weylchem di-ester
Batch: DECG020911
Purity: 94.9%

Target gene:

testing the ability to induce forward mutations at the thymidine kinase (TK) locus in L5178Y mouse lymphoma cells

Species / strain / cell type:

mouse lymphoma L5178Y cells

Details on mammalian cell type (if applicable):

L5178Y/TK+/--3.7.2C mouse lymphoma cells [American Type Culture Collection, (ATCC, Manassas, USA) (2001)]

Basic medium: RPMI 1640 Hepes buffered medium (Dutch modification) (Life Technologies) containing penicillin/streptomycin (50 U/ml and 50 μg/ml, respectively) (Life Technologies), 1 mM sodium pyruvate (Sigma, Zwijndrecht, The Netherlands) and 2 mM L-glutamin (Life Technologies).

Metabolic activation:

with and without

Metabolic activation system:

Rat liver microsomal enzymes (S9 homogenate) were obtained from Trinova Biochem GmbH and was prepared from rats that have been dosed orally with a suspension of phenobarbital (80 mg/kg body weight) and ß-naphthoflavone (100 mg/kg body weight).

Test concentrations with justification for top dose:

First Mutagenicity Test: Based on the results of the dose-range finding test, the following dose-range was selected for the first mutagenicity test: Without and with S9-mix: 1.22, 2.44, 4.88, 9.75, 19.5, 39, 78 and 156 μg/ml exposure medium.

Second Mutagenicity Test: Based on the results of the dose-range finding test and experiment 1, the following dose levels were selected for mutagenicity testing: 1.22, 2.44, 4.88, 9.75, 19.5, 39, 78, 117 and 156 μg/ml exposure medium.

Since the test item was not toxic and difficult to dissolve in aqueous solutions the highest concentration was determined by the solubility in the culture medium.

Vehicle / solvent:

The vehicle for the test item was dimethyl sulfoxide (DMSO, SeccoSolv, Merck Darmstadt, Germany).

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Remarks:

historical control data range

Positive controls:

yes

Positive control substance:

cyclophosphamide

methylmethanesulfonate

Details on test system and experimental conditions:

METHOD OF APPLICATION: in suspension

DURATION
In the first experiment, cell cultures were exposed for 3 hours to the test item in exposure medium in the absence and presence of S9-mix. In the second experiment, cell cultures were exposed to the test item in exposure medium for 24 hours in the absence of S9-mix.

For expression of the mutant phenotype, the remaining cells were cultured for 2 days after the treatment period. During this culture period at least 4000000 cells (where possible) were subcultured every day in order to maintain log phase growth. Two days after the end of the treatment with the test item the cells were plated for determination of the cloning efficiency and the mutation frequency.

SELECTION AGENT
For determination of the cloning efficiency the cell suspensions were diluted and seeded in wells of a 96-well dish. One cell was added per well (2 x 96-well microtiter plates/concentration) in non-selective medium.

For determination of the mutation frequency a total number of 960000 cells per concentration were plated in five 96-well microtiter plates, each well containing 2000 cells in selective medium (trifluorothymidine-selection), with the exception of the positive control groups (methyl methanesulfonate and cyclophosphamide) where a total number of 960000 cells/concentration were plated in ten 96-well microtiter plates, each well containing 1000 cells in selective medium (trifluorothymidine-selection). The microtiter plates for cloning efficiency and mutation frequency were incubated for 11 or 12 days. After the incubation period, the plates for the trifluorothymidine-selection were stained for 2 hours, by adding 0.5 mg/ml 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (Sigma) to each well. The plates for the cloning efficiency and mutation frequency were scored with the naked eye or with the microscope.

DETERMINATION OF THE MUTANT COLONIES:
The colonies were divided into small and large colonies. Mutant cells that have suffered extensive genetic damage have prolonged doubling times and thus form small colonies. Less severely affected mutant cells grow at rates similar to the parental cells and form large colonies. The small colonies can be associated with the induction of chromosomal mutations. The large colonies appear to result from mutants with single gene mutations (substitutions, deletions of base-pairs) affecting the TK gene.

The small colonies are morphologically dense colonies with a sharp contour and with a diameter less than a quarter of a well. The large colonies are morphologically less dense colonies with a hazy contour and with a diameter larger than a quarter of a well. A well containing more than one small colony is classified as one small colony. A well containing more than one large colony is classified as one large colony. A well containing one small and one large colony is classified as one large colony.

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Remarks:

vehicle control served as negative control

Positive controls validity:

valid

Additional information on results:

The mutation frequency found in the solvent control cultures was within the acceptability criteria of this assay and within the 95% control limits of the distribution of the historical negative control database.

Positive control chemicals, methyl methanesulfonate (MMS) and cyclophosphamide, both produced significant increases in the mutation frequency. In addition, the mutation frequency found in the positive control cultures was within the 95% control limits of the distribution of the historical positive control database. Although in the second experiment the response of MMS was just above the upper control limits, these limits are 95% control limits and a slightly higher response is within the expected response ranges. It was therefore concluded that the test conditions were adequate and that the metabolic activation system (S9-mix) functioned properly.

The suspension growth over the two-day expression period for cultures treated with DMSO was between 20 and 26 (3 hour treatment) and 51 and 53 (24 hour treatment).

In the absence of S9-mix, the test item did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in a repeat experiment with modification in the duration of treatment.

In the presence of S9-mix, the test item did not induce a significant increase in the mutation frequency.

Conclusions:

In conclusion, the test item is not mutagenic in the TK mutation test system under the experimental conditions described in this report.

Executive summary:

The objective of this study was to evaluate the mutagenic potential of the test item by testing its ability to induce forward mutations at the thymidine kinase (TK) locus in L5178Y mouse lymphoma cells, either in the absence or presence of a metabolic system (S9-mix). The TK mutational system detects base pair mutations, frame shift mutations and small deletions.

 

The test was performed in the absence of S9-mix with 3 and 24 hour treatment periods and in the presence of S9-mix with a 3 hour treatment period.

 

The study procedures described in this report were based on the most recent OECD guideline.

 

Batch DECG020911 of the test item was a slightly yellow solid. The test item was dissolved in dimethyl sulfoxide.

 

In the first experiment, the test item was tested up to concentrations of 156 μg/ml in the absence and presence of S9-mix. The incubation time was 3 hours. No toxicity was observed at this dose level in the absence and presence of S9-mix. The test item precipitated in the culture medium at the dose levels of 78 and 156 μg/ml.

 

In the second experiment, the test item was tested up to concentrations of 130 μg/ml in the absence of S9-mix. The incubation time was 24 hours. No severe toxicity was observed at this dose level in the absence of S9-mix. The test item precipitated in the culture medium at the dose levels of 115 and 130 μg/ml.

 

The mutation frequency found in the solvent control cultures was within the acceptability criteria of this assay and within the 95% control limits of the distribution of the historical negative control database.

 

Positive control chemicals, methyl methanesulfonate (MMS) and cyclophosphamide, both produced significant increases in the mutation frequency. In addition, the mutation frequency found in the positive control cultures was within the 95% control limits of the distribution of the historical positive control database. Although in second experiment the response of MMS was just above the upper control limits, these limits are 95% control limits and a slightly higher response is within the expected response ranges. It was therefore concluded that the test conditions were adequate and that the metabolic activation system (S9-mix) functioned properly.

 

In the absence of S9-mix, the test item did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent experiment with modification in the duration of treatment.

 

In the presence of S9-mix, the test item did not induce a significant increase in the mutation frequency.

 

In conclusion, the test item is not mutagenic in the mouse lymphoma L5178Y test system under the experimental conditions described in this report.

Cross-reference 29

Reason / purpose for cross-reference:

read-across: supporting information

Reference

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study without detailed documentation

Justification for type of information:

This endpoint study record is supporting information reported within the read-across justification

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

GLP compliance:

not specified

Type of assay:

in vitro mammalian cell micronucleus test

Specific details on test material used for the study:

Hexandiol

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Species / strain:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Conclusions:

Hexandiol was not mutagenic in the in vitro point mutation studied in mammalian cells (Chinese hamster V79 HPRT locus, OECD 476) with no indication of a mutagenic response either in the presence or absence of metabolic activation.

Materials and methods

Test material

Test material information

Constituent 1

Reference substance name:

1,6-Bis(methoxybenzoyloxy)hexane

Cas Number:

390359-18-9

Molecular formula:

C22H26O6

IUPAC Name:

1,6-Bis(methoxybenzoyloxy)hexane

Test material form:

other: Light yellowish solid

Details on test material:

- Name of test material (as cited in study report): Setafix X 11342
- Substance type: Organic
- Physical state: Solid
- Analytical purity: >95%
- Lot/batch No.: PP03
- Expiration date of the lot/batch: 31 December 2004
- Stability under test conditions:
- Storage condition of test material: At room temperature in the dark
- Stability under storage conditions: Stable

Specific details on test material used for the study:

1,6-Bis(methoxybenzoyloxy)hexane value is read-across from supporting 1,6-hexanediol and vanillin

Results and discussion

Results: P0 (first parental generation)

Effect levels (P0)

Dose descriptor:

NOAEL

Effect level:

ca. 316 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

Vanillin

Sex:

female

Basis for effect level:

clinical signs

Remarks on result:

other: based on the experimental data for vanillin (NOAEL = 250 mg/kg bw) and the amount of p-anisic acid metabolite produced during metabolism.

Results: F1 generation

Effect levels (F1)

Dose descriptor:

NOEL

Generation:

F1

Effect level:

ca. 633 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

Vanillin

Basis for effect level:

other: teratogenicity

Remarks on result:

other: based on the experimental data for vanillin (NOEL = 500 mg/kg bw) and the amount of p-anisic acid metabolite produced during metabolism.

Overall reproductive toxicity

Reproductive effects observed:

no

Any other information on results incl. tables

The 1,6-hexanediol metabolite is clearly identical to the first source substance, and the amount produced will be equivalent to 31% w/w of the dose of target substance.

The p-anisic acid metabolite is structurally very similar to the second source substance, and the amount produced will be equivalent to 79% w/w of the dose of target substance.

The sum of the above values exceeds 100% due to the mass added by the incorporation of water of hydrolysis.

As such, based on the experimental data for vanillin (NOEL = 500 mg/kg bw), the NOEL of the target substance for developmental toxicity can be predicted to be approximately 633 mg/kg bw. The toxicity data on the vanillin source substance will represent a very similar or slightly worse case than, and provide a sound basis for a slightly conservative assessment of, the toxicity of the target substance.

Applicant's summary and conclusion

Conclusions:

Vanillin was considered to have no teratogen effects

In a valid OECD 421 study no indication of toxic effect on reproductive function or developmental toxicity were observed with 1,6-hexanediol.

Information on the two source substances will represent a similar or worse case than the target substance.