Bis(1-n-butyl-3-methylcyclopentadienyl)zirconium dichloride

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Justification for type of information:

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
Zirconium chloride is expected to be one of the key degradation products of alkylated zirconocene dichloride. This metal salt is expected to be the most likely substance with a sensitization potential.

Data source

Reference

Reference Type:

publication

Title:

Evaluation of skin sensitization potential of nickel, chromium, titanium and zirconium salts using guinea-pigs and mice

Author:

Ikarashi, Y., J. Momma, T. Tsuchiya, and A. Nakamura

Year:

1996

Bibliographic source:

Biomaterials 17(21): 2103-2108

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

Deviations:

yes

Remarks:

Assay was performed with the addition of a intradermally injected sensitization phase.

GLP compliance:

not specified

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Zirconium tetrachloride

EC Number:

233-058-2

EC Name:

Zirconium tetrachloride

Cas Number:

10026-11-6

Molecular formula:

Cl4Zr

IUPAC Name:

zirconium tetrachloride

Test material form:

solid

Specific details on test material used for the study:

Zirconium chloride (ZrCl,) was obtained from Wako Pure Chemical Industries, Ltd. (Osaka, Japan).

In vivo test system

Test animals

Species:

mouse

Strain:

Balb/c

Sex:

female

Details on test animals and environmental conditions:

Adult female mice were purchased from Japan SLC Inc. (Shizuoka, Japan).

Study design: in vivo (LLNA)

Vehicle:

dimethyl sulphoxide

Remarks:

Saline was used for as vehicle for intradermal injection, while DMSO was used as vehicle for topical application

Concentration:

0.02, 0.2 or 2% intradermal injection.
5% topically applied.

No. of animals per dose:

3 mice per dose group

Details on study design:

This assay was performed according to the method described previously (Ikarashi et al., 1993). Groups of mice (n = 3) were initially injected intradermally, totaling 50 ul of test chemical-FCA emulsion into two sites of the abdominal skin. Five days after injection, the mice received 25 ul of test chemical in vehicle on both sides of each ear for three consecutive days. Control mice were treated by intradermal injection of vehicle-FCA emulsion into the abdomen and, after 5 days, the mice were exposed to vehicle alone on the ears for three consecutive days. The day after the final application, auricular lymph nodes were excised, pooled for each experimental group and weighed. A single cell suspension of lymph node cell (LNC) was prepared by mechanical disaggregation through sterile 200-mesh gauge, and washed once with Hanks’ balanced salt solution. The LNCs were resuspended in RPMI-1640 culture medium supplemented with 25 mM HEPES, 100 units/ml penicillin, 100 pg/ml streptomycin and 10% fetal calf serum, and total LNC number was determined using an automated cell counter. The increase in LNC number relative to vehicle-treated controls was derived for each experimental group and recorded as a stimulation index of LNC number (SIn). The LNC suspensions (1 x 10^6 cells) were seeded into 96-well culture plates (five wells per group) and cultured with 0.5 uCi [3H] methyl thymidine (3HTdR) for 24 h at 37°C in a humidified atmosphere of 5% CO, in air. Culture was terminated by using a semiautomatic cell harvester, and the 3HTdR incorporation was determined by liquid scintillation counting. The increase in 3HTdR incorporation relative to controls was derived for each experimental group and recorded as a stimulation index of LNC proliferation [SIp). SItotal was also defined as SIn x SIp and it indicates the total lymph node activation induced by test chemicals. A chemical was regarded as positive (a sensitizer) if it showed an SItotal value of 3 or greater.

Ikarashi Y, Tsuchiya T, Nakamura A (1993) A sensitive mouse lymph node assay with two application phases for detection of contact allergens. Arch Toxicol 67: 629-636

Positive control substance(s):

not specified

Results and discussion

Positive control results:

Not applicable

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Remarks:

stimulation index of lymph node cell proliferation

Value:

> 0.8 - <= 0.93

Results 2

Parameter:

SI

Value:

0.8

Test group / Remarks:

0.02% ZrCl4, n=3

Results 3

Parameter:

SI

Value:

0.92

Test group / Remarks:

0.2% ZrCl4, n=3

Results 4

Parameter:

SI

Value:

0.93

Test group / Remarks:

2% ZrCl4, n=3

Cellular proliferation data / Observations:

Two different parameters were reported for all 3 test concentrations. SIn was calcualted as the stimulation index of lymph node cell proliferation and SIp represented the stimulation index of lymph node cell proliferation. Results for SIn and SIp were all below 1.2 fold greater than vehicle control.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

ZrCl4 did not show any sensitization responses.

Executive summary:

Contact sensitization potential of metal salts, including Zirconium chloride, were evaluated using the sensitive mouse lymph node assay. Zirconium chloride was determined to not be a skin sensitizer.