Diammonium sodium hexakis(nitrito-N)rhodate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 August - 18 September 2013

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

Guideline study performed largely to GLP (except that the stability of the test item was determined in a separate analytical study, and no analysis was conducted to determine the homogeneity or concentration of the test item). Minor deviations (humidity outside 30-70% range) not expected to affect study validity.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

on occasions, relative humidity was outside the target range of 30-70%. This was not considered to affect study integrity

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

GLP compliance:

yes (incl. QA statement)

Remarks:

Except that the stability of the test item was determined in a separate analytical study, and no analysis was conducted to determine the homogeneity or concentration of the test item.

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Diammonium sodium hexakis(nitrito-N)rhodate

EC Number:

264-713-0

EC Name:

Diammonium sodium hexakis(nitrito-N)rhodate

Cas Number:

64164-17-6

Molecular formula:

H4N.1/2N6O12Rh.1/2Na

IUPAC Name:

Diammonium sodium hexakis(nitro-N)rhodate

Test material form:

solid - liquid: suspension

Details on test material:

- Name of test material (as cited in study report): diammonium sodium hexakis(nitrito-N)rhodate
- Substance type: no data
- Physical state: no data
- Analytical purity: no data [19.55% given as % for all elements in Certificate of Analysis]
- Impurities (identity and concentrations) (as %): platinum (0.01), ruthenium (0.05), iridium (0.06), silver (0.01), iron (0.07), nickel (0.59), bismuth (0.13), copper (1.51), tellurium (0.25), lead (0.84), chromium (0.07), aluminium (0.09)
- Composition of test material, percentage of components: rhodium at 15.88%
- Isomers composition: no data
- Purity test date: no data
- Lot/batch No.: RHL 633
- Expiration date of the lot/batch: 19 September 2014
- Stability under test conditions: stable in propylene glycol for at least 2 hours
- Storage condition of test material: room temperature in the dark

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/Ca (CBA/CaOlaHsd)

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories Ltd., Oxon, UK
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 15-23 g
- Housing: individually housed in suspended solid floor polypropylene cages furnished with softwood woodflakes
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): target: 19-25 (no data on actual values, but presumably not outside this range)
- Humidity (%): target: 30-70 (no data on actual values, but occasional deviations outside this range were noted, and were not considered to affect study integrity)
- Air changes (per hr): ~15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: no data

Study design: in vivo (LLNA)

Vehicle:

propylene glycol

Concentration:

Range-finding study: 25% (w/w)
Main study: 0, 5, 10 or 25%

No. of animals per dose:

Range-finding study: 1
Main study: 5

Details on study design:

RANGE FINDING TESTS:
- Compound solubility: solubility in various common vehicles assessed to find the vehicle giving the highest concentration suitable for dosing
- Irritation: yes. Range-finding mouse observed twice daily on days 1, 2 and 3, and once daily on days 4, 5 and 6, with erythema scored from 0-4.
- Lymph node proliferation response: not performed

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: local lymph node assay
- Criteria used to consider a positive response: the proliferation response of lymph node cells was expressed as the number of radioactive disintegrations per minute per animal and as the ratio of 3HTdR incorporation into lymph node cells of test nodes relative to that recorded for the control nodes (Stimulation Index). The test item was regarded as a sensitiser if at least one concentration resulted in a threefold or greater increase in 3HTdR incorporation compared to control values. Any test item failing to produce a threefold or greater increase in 3HTdR incorporation was classified as a "non sensitiser."

TREATMENT PREPARATION AND ADMINISTRATION: the test item was freshly prepared as a suspension in propylene glycol. The test item was formulated within two hours of being applied to the test system. 25 µl was applied topically to the ears of mice on days 1, 2 and 3 in both the range-finding and the main study.

Positive control substance(s):

other: phenylacetaldehyde (>90% purity) at 2.5% (v/v) in propylene glycol

Statistics:

Data were processed to give group mean values for disintegrations per minute and standard deviations where appropriate. Individual and group mean disintegrations per minute values were assessed for dose response relationships by analysis of homogeneity of variance followed by one way analysis of variance (ANOVA). In the event of a significant result from the ANOVA, pairwise comparisons were performed between control and treated groups. For homogenous datasets, Dunnett’s Multiple Comparison test was used, and for non homogenous datasets, Dunnett’s T3 Multiple Comparison Method was used.

Results and discussion

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: See Table 1 in "any other information on results incl. tables".

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: See Table 1 in "any other information on results incl. tables".

Any other information on results incl. tables

In the range-finding study, there were no signs of systemic toxicity, visual skin irritation, or ≥25% increased ear thickness (considered a sign of local irritation).

Table 1      Individual Disintegrations per Minute and Stimulation Indices

Treatment Group	Animal Number	dpm/ Animal^a	Mean dpm/Animal (Standard Deviation)	Stimulation Index^b	Result
Vehicle propylene glycol	1-1	1030.91	2038.00 (±812.38)	N/A	N/A
	1-2	2227.51
	1-3	3264.38
	1-4	1801.09
	1-5	1866.09
Test Item 5% w/w in propylene glycol	2-1	1495.69	2421.31 (±646.90)	1.19	Negative
	2-2	1993.61
	2-3	2898.45
	2-4	2752.74
	2-5	2966.04
Test Item 10% w/w in propylene glycol	3-1	1719.93	1949.44 (±493.19)	0.96	Negative
	3-2	1565.86
	3-3	2793.73
	3-4	1963.41
	3-5	1704.28
Test Item 25% w/w in propylene glycol	4-1	1923.37	2781.17 (±763.81)	1.36	Negative
	4-2	3983.18
	4-3	2742.12
	4-4	2399.61
	4-5	2857.56
Positive Control Item 2.5% v/v in propylene glycol	5-1	25068.56	12617.71 (±8237.59)	6.19	Positive
	5-2	16448.63
	5-3	10121.97
	5-4	5236.74
	5-5	6212.65

 

The results of the statistical analysis of the data indicated there was no significant difference between the control group and the test groups.

 

dpm=    Disintegrations per minute

N/A =     Not applicable

a=  Total number of lymph nodes per animal is 2

b=  Stimulation Index of 3.0 or greater indicates a positive result

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Diammonium sodium hexakis(nitrito-N)rhodate was non-sensitising in a mouse LLNA, conducted according to OECD guideline 429.

Executive summary:

The skin sensitising potential of diammonium sodium hexakis(nitrito-N)rhodate was assessed in a murine local lymph node assay (LLNA) conducted in accordance with OECD Test Guideline 429 and to GLP. Following a preliminary test using a concentration of 25% w/w, female CBA/Ca mice (5/group) received 25 µL applications of the test material, in propylene glycol at 0, 5, 10 or 25%, to the skin of both ears for three consecutive days. Two additional groups of 5 females were respectively treated with propylene glycol alone and phenylacetaldehyde, a known sensitiser, at 2.5%.

 

Mice were monitored for mortality and overt signs of toxicity, as well as changes in body weight. Three days after the induction period (on day six), mice were given tail vein injections of radiolabelled thymidine, and were sacrificed after approximately five hours. Lymph nodes were excised and processed for radioactivity.

 

No signs of skin irritation or systemic toxicity were observed in the preliminary test. In the main experiment, SI values were 1.19, 0.96 and 1.36 for animals given respective concentrations of 5%, 10% and 25%. An SI value of over 3 is judged to be indicative of a substance being sensitising. Hence, under the conditions of this assay, diammonium sodium hexakis(nitrito-N)rhodate was considered non-sensitising.

 

Based on the results of this study, diammonium sodium hexakis(nitrito-N)rhodate does not warrant classification for skin sensitisation, according to EU CLP criteria (EC 1272/2008).