Diethyl (hydroxymethyl)phosphonate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Deviations:

no

Principles of method if other than guideline:

n/a

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Diethyl (hydroxymethyl)phosphonate

EC Number:

221-391-6

EC Name:

Diethyl (hydroxymethyl)phosphonate

Cas Number:

3084-40-0

Molecular formula:

C5H13O4P

IUPAC Name:

diethyl (hydroxymethyl)phosphonate

Test material form:

other: meduim viscosity liquid

Details on test material:

E06-16 (Lot# 1041-37-6, Sample: T# 207), transparent "medium-viscoosity" liquid.

Specific details on test material used for the study:

Lot number 540-1013-14

In vivo test system

Test animals

Species:

mouse

Strain:

CBA/Ca

Sex:

female

Details on test animals and environmental conditions:

Healthy female CBA/Ca mice (a total of 34 females) from Harlan Netherlands Ltd.
weight range: 17.0 to 22.4 g a
eight to twelve weeks of age
Acclimatisation to experimental environment: at least 6 days prior to the start of the study.
Two or three animals per cage
Temperature and relative humidity: range of 19 to 23oCC and 40 to 70% respectively.
Artificial lighting was controlled to give a cycle of 12 hours continuous light and 12 hours continuous dark per 24 hours.

Study design: in vivo (LLNA)

Vehicle:

dimethylformamide

Concentration:

50% v/v

No. of animals per dose:

5

Details on study design:

The study comprised of three treated groups, each comprising five female mice receiving E06-16 at concentrations of 25, 50% v/v and the test substance as supplied. Similarly constituted groups received a sham dose (naïve control), the vehicle dimethylformamide (DMF) or positive control substance (25% v/v hexyl cinnamic aldehyde).
The mice were treated by daily application of 25 uL of the appropriate concentration or control (vehicle or positive), to the dorsal surface of both ears for three consecutive days. The naïve control were not dosed with test substance or vehicle, just the tip of the pipette passed over the ears on each dosing occasion.
The proliferative response of the lymph node cells (LNC) from the draining auricular lymph nodes was assessed five days following the initial application, by measurement of the incorporation of 3H-methyl Thymidine (3HTdR) by beta-scintillation counting of LNC suspensions. The response was expressed as radioactive disintegrations per minute per lymph node (dpm/node) and as the ratio of 3HTdR incorporation into LNC of test nodes relative to that recorded for control nodes (test/control ratio), termed as Stimulation Index (SI).

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Statistics:

Statistical Analysis of Radioactivity Data (dpm) for the E06-16 Local Lymph Node Assay was performed.
Varous tests were applied as follows: Bartlett’s test, Dunnett's test Shirley's test, Steel's test, Wilcoxon rank sum test and t-test.

Results and discussion

Positive control results:

The SI for the positive control substance hexyl cinnamic aldehyde was 9.2, which demonstrates the validity of this study.There was a significant increase in mean dpm for the HCA treated group in comparison with the vehicle control (p=0.001).

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

>= 0.7

Test group / Remarks:

5 animals

Remarks on result:

other: 25% v/v

Results 2

Key result

Parameter:

SI

Value:

>= 1

Test group / Remarks:

5 animals

Remarks on result:

other: 50% v/v

Results 3

Key result

Parameter:

SI

Value:

>= 2.2

Test group / Remarks:

5 animals

Remarks on result:

other: as supplied

Cellular proliferation data / Observations:

Product: DPM*/node SI
vehicle DMF 237.5 n/a
Naive control 181.6 n/a
25% v/v E06-16 167.6 0.7
50% v/v E06-16 232.7 1.0
E06-16 As supplied 391.8 2.2
HCA 25% v/v 2191.5 9.2

*dpm- Disintegrations per minute (less background count of 43.0 dpm)

Any other information on results incl. tables

There was no evidence of a treatment effect for the E06-16 treated groups when compared with the appropriate control group. This result was consistent with stimulation indices less than 3 for these groups.

Applicant's summary and conclusion

Interpretation of results:

other: Not sensitizing

Conclusions:

E06-16 is not regarded as a potential skin sensitizer

Executive summary:

The study was performed to assess the skin sensitization potential of E06-16 using the local lymph node assay (LLNA).

The study comprised of three treated groups, each comprising five female mice receiving E06-16 at concentrations of 25, 50% v/v and the test substance as supplied. Similarly constituted groups received a sham dose (naïve control), the vehicle dimethylformamide (DMF) or positive control substance (25% v/v hexyl cinnamic aldehyde). The mice were treated by daily application of 25 uL of the appropriate concentration or control (vehicle or positive), to the dorsal surface of both ears for three consecutive days. The naïve control were not dosed with test substance or vehicle, just the tip of the pipette passed over the ears on each dosing occasion.

The proliferative response of the lymph node cells (LNC) from the draining auricular lymph nodes was assessed five days following the initial application, by measurement of the incorporation of 3H-methyl Thymidine (3HTdR) by -scintillation counting of LNC suspensions. The response was expressed as radioactive disintegrations per minute per lymph node (dpm/node) and as the ratio of 3HTdR incorporation into LNC of test nodes relative to that recorded for control nodes (test/control ratio), termed as Stimulation Index (SI). The test substance is regarded as a sensitizer if at least one concentration of the chemical has a SI of three or more.

Results

The SI obtained for 25, 50% v/v and as supplied were 0.7, 1.0 and 2.2 respectively which indicates that E06-16 did not show the potential to induce skin sensitization. The SI for the positive control substance hexyl cinnamic aldehyde was 9.2, which demonstrates the validity of this study.

Conclusion

E06-16 is not regarded as a potential skin sensitizer.