Cobalt(2+) hydrogen citrate

Endpoint summary

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

15 Feb 2012 - 06 Mar 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-Guideline study (The LLNA is not recommended for certain metals)

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted in 2010

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted in 2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Hess. Ministerium fuer Umwelt, Energie, Landwirtschaft und Verbraucherschutz, Wiesbaden, Germany

Type of study:

mouse local lymph node assay (LLNA)

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Additional information on strain: CBA/CaOlaHsd
- Source: Harlan Laboratories B.V., Postbus 6174, 5960 AD Horst, The Nederlands
- Age at study initiation: 8-9 weeks (pre-test: 10-12 weeks)
- Mean weight at study initiation: 19.7 g (pre-test: 21.1 g)
- Housing: 3/cage in Makrolon Type II (pre-test)/Type III (main test) with wire mesh top (Ehret GmbH, Emmendingen, Germany)
- Diet: pelleted standard diet (Harlan Laboratories V.V., Horst, Netherlands); ad libitum
- Water: tap water; ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 32 - 65 (acclimation phase); 45-65 (pre-test and main study)
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

acetone/olive oil (4:1 v/v)

Concentration:

0.5, 1 and 2.5% (w/v)

No. of animals per dose:

4 (pre-test: 2)

Details on study design:

RANGE FINDING TESTS:
- Compound solubility: A solubility experiment was performed according to the recommendations given in the OECD guideline 429. The highest concentration which could be technically used was a 25% suspension in AOO. Grinding of the test substance was necessary.
- Irritation: To determine the highest non-irritant test concentration, that did not induce systemic signs of toxicity, a pre-test was performed in 2 animals. The animals were treated with 10 and 25% of the test substance once daily on 3 consecutive days. Besides recording of body weight, signs of toxicity and grading of erythema, ear thickness was measured before sacrifice on day 6. Excessive local erythema were seeen; therefore, a second pre-test was performed using test item concentrations of 2.5 and 5%. At both concentrations an increase in ear thickness was observed that exceeded the threshold value for excessive local skin irritation. Both animals showed erythema of the ear skin on day 4 and 5 which decreased to score 1 on day 6. At 5%, an increase in ear weight was observed that distinctly exceeded the threshold value for excessive local skin irritation. Thus, in the main study the test item was assayed at 0.5, 1, and 2.5%.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Criteria used to consider a positive response: A test item is regarded as a sensitiser in the LLNA if the following criteria are fulfilled: 1) Exposure to at least one concentration of the test item resulted in an icorporation of ³HTdR at least 3-fold or greater than that recorded in control mice, as indicated by the Stimulation index (SI). 2) The data are compatible with a conventional dose response, although allowance must be made for either local toxicty or immunological suppression.

TREATMENT PREPARATION AND ADMINISTRATION:
- Site: topical application at the dorsum of each ear once daily each on three consecutive days with 3 different concentrations of the test substance in AOO
- Application volume: 25 µL per application
- Control: same treatment with vehicle only
5 days after the first application, the mice were intravenously injected with radio-labelled thymidine (³H-methyl thymidine; ³HTdR) in PBS (250 µL). Approx. 5 hours after injection, the mice were sacrificed and the draining auricular lymph nodes excised and pooled per group. The proliferative capacity of pooled lymph node cells was determined by the incorporation of ³HTdR measured in a beta-scintillation counter. ³HTdR incorporation was expressedas the number of radioactive disintegrations per minute (DPM).

Positive control substance(s):

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

Statistics:

The mean values and standard deviations were calculated.

Positive control results:

The EC3 value of the positive controls (test performed in December 2011) was 14.4%.

Parameter:

SI

Remarks on result:

other: - 0.5% : 3.73 - 1%: 4.18 - 2.5%: 6.07 Since the SI values for all treatment groups were > 3, no EC3 value was determined. A clear dose-response was observed.

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: - Vehicle control: 1886 - 0.5%: 6976 - 1%: 7819 - 2.5%: 11340

Table 1: LLNA – Details on results

Test item concentration (%)	Group	Measurement DPM	Calculation			Result
			DPM-BG a)	No. of lymph nodes	DPM per lymph node b)	S.I.
-	BG I	19	-	-	-	-
-	BG II	20	-	-	-	-
0	1	1886	1867	8	233.3	1.00
0.5	2	6976	6957	8	869.6	3.73
1	3	7819	7800	8	974.9	4.18
2.5	4	11340	11321	8	1415.1	6.07

BG = background (5% trichloroacetic acid)

a) calculated with the mean value of BG I and II

b) DPM/node was determined by dividing the measured values by the number of lymph nodes pooled.

The animals did not show any signs of systemic toxicity during the study. On day 2, the animals of all test groups showed erythema of the ear skin (score = 1) and from day 3 to day 6 the observed erythema was score = 2 in all test groups.

The body weights and body weight gains were within the normal range.

Interpretation of results:

sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: Skin sens 1A, H317
DSD: Xi, R43

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

The skin sensitising potential was evaluated in a Local Lymph Node Assay (LLNA) following OECD 429 (Wieland, 2012). Cobalt citrate was tested in concentrations of 0.5%, 1.0%, and 2.5% in acetone/olive oil (AOO) based on the result of a range-finding test. Hexyl cinnamic aldehyde in (4:1% v/v) AOO was used as positive control. 4 mice per group were topically treated with 25 µL of the appropriate formulations of the test item, the positive control substance and the vehicle using a pipette, on the dorsal surface of each ear. Each animal was dosed once a day on 3 consecutive days. There was no treatment on days 4, 5, and 6. On day 6, the animals were intravenously injected via the tail vein with 250 µL of sterile PBS containing 20 µCi of ³H-methyl-thymidine. After sacrifice, the draining auricular lymph nodes were excised, removed and pooled per group and a single cell suspension of lymph node cells was prepared. The incorporated radioactivity was measured and expressed as the amount of radioactive disintegration per minute (DPM) and transformed in DPN (DPN = DPM/number of pooled lymph nodes). The SI is the ratio of DPN of the treated mice and the negative control. The results for the SI values were 1.0, 3.73, 4.18, and 6.07 for the negative control and for 0.5%, 1.0%, and 2.5% of the test item, respectively. The criteria used to consider a positive response, is a 3-fold increase of the stimulation index (SI) compared to the negative control. Based on the results of the LLNA, cobalt citrate is considered as skin sensitising.

There are no epidemiological data available on skin sensitisation for cobalt hydrogen citrate. However, there are reliable data for soluble cobalt compounds considered suitable for read-across using the analogue approach.

Cobalt hydrogen citrate is a metal-organic compound, which is water soluble and nearly completely dissociates in aqueous solutions. For identifying hazardous properties of cobalt hydrogen citrate concerning human health effects, the existing forms of the target chemical at physiological and very acidic pH conditions (e. g. in the stomach) are relevant for risk assessment. For cobalt hydrogen citrate, it can be assumed that cobalt cations are released under biological conditions that are considered to be toxicologically relevant. Furthermore it is anticipated that the cobalt cation released by the parent compound is the determining factor for toxicological effects, same as for other soluble cobalt compounds. Therefore, data originating from soluble cobalt compounds can be used in the derivation of toxicological endpoints for cobalt hydrogen citrate.For further details, please refer to the read-across justification attached in section 13 of the technical dossier.

In humans, dermal exposure arising from soluble cobalt salts has been also observed to result in skin sensitisation. Contact allergy was reported in 22 of 223 (9.9%) nurses who were tested with a patch test of 1% cobalt(II)chloride (Kiec-Swierczynska and Krecisz, 2000), as well as 16 of 79 (20.3%) of examined dentists (Kiec-Swierczynska and Krecisz, 2002). Numerous human data also show that soluble cobalt salts are skin sensitisers (for example Kanerva et al., 1988; Goh et al., 1986; Alomar et al., 1985).

References:

Kiec-Swierczynska M and Krecisz B, 2002, Allergic contact dermatitis in dentists and dental nurses. Exogenous Dermatology. 1(1): 27 -31

Kanerva L et al., 1988, Occcupational skin disease in Finland, International Archives of Occupational and Environmental Health, 60: 89 -94

Goh et al., 1986, Occupational dermatitis in a prefabrication construction factory. Contact dermatitis, 15: 235 -240

Alomar et al., 1985, Occupational dermatosis from cutting oils. Contact dermatitis, 12:129 -138

Migrated from Short description of key information:
Cobalt citrate
Local Lymph Node Assay (OECD 406): sensitising

Justification for selection of skin sensitisation endpoint:
There is only one study available.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

There are no data available on respiratory sensitisation for cobalt hydrogen citrate. However, there are reliable data for soluble cobalt compounds considered suitable for read-across using the analogue approach.

Cobalt hydrogen citrate is a metal-organic compound, which is water soluble and nearly completely dissociates in aqueous solutions. For identifying hazardous properties of cobalt hydrogen citrate concerning human health effects, the existing forms of the target chemical at physiological and very acidic pH conditions (e. g. in the stomach) are relevant for risk assessment. For cobalt hydrogen citrate, it can be assumed that cobalt cations are released under biological conditions that are considered to be toxicologically relevant. Furthermore it is anticipated that the cobalt cation released by the parent compound is the determining factor for toxicological effects, same as for other soluble cobalt compounds. Therefore, data originating from soluble cobalt compounds can be used in the derivation of toxicological endpoints for cobalt hydrogen citrate. For further details, please refer to the read-across justification attached in section 13 of the technical dossier.

The available case reports revealed that soluble cobalt (II) salts are capable of inducing hypersensitive reactions in the respiratory tract after inhalation exposure. These hypersensitive reactions include e. g. respiratory irritation, bronchial asthma, wheezing and pneumonia.

Shirakawa et al. (1989) reported that inhalation of cobalt(II)chloride aerosols can provoke an asthmatic attack in sensitised individuals. Inhalation exposure to cobalt(II)salts among glass bangle workers resulted in decreases in ventilatory function relative to the control workers (Rastogi et al., 1991). In the cross-sectional study of Nemery et al. (1992), 194 workers (166 men and 28 women) in the diamond polishing industry were exposed to cobalt dusts. A significant increase in the prevalence of eye, nose, and throat irritation and reduced lung function compared to 59 unexposed control workers (46 men and 13 women) were observed. Cobalt exposure groups were defined based on air measurements at the time of the study, and exposure was confirmed by measurement of cobalt in urine. The duration of employment was not discussed. In another cross-sectional study, workers in a cobalt refinery who were exposed to cobalt metal, salts and oxides for up to 39 years at an average concentration of 0.125 mg Co/m3 had increased dyspnoea and wheezing, and decreased lung function compared to unexposed controls (Swennen et al., 1993). Linna et al (2003) also found that asthma symptoms were more prevalent in workers in a cobalt plant who were exposed to cobalt compounds. However, many of the epidemiological studies on inhalation exposure to cobalt have been conducted on workers in the hard metal industry where subjects are co-exposed to elemental cobalt and other substances such as tungsten carbide. It is difficult to assess the effects of cobalt alone from these studies as the toxicity of cobalt metal is increased in the presence of tungsten carbide, and it has been proposed that a mixture of cobalt and tungsten carbide behaves as a unique entity (IARC 2006).

References not cited in the IUCLID:

Roto P, 1980, Asthma, symptoms of chronic bronchitis and ventilatory capacity among cobalt and zinc production workers. Scand J Work Environ Health 6: 1-49

Kusaka Y et al., 1996a, Epidemiological study of hard metal asthma. Occup Environ Med 53: 188-193

Kusaka Y et al., 1996b, Decreased ventilatory function in hard metal workers. Occup Environ Med 53: 194-199

Ruokonen EL et al., 1996, A fatal case of hard-metal disease. Scand J Work Environ Health 22: 62-65

IARC (International Agency for Research on Cancer). 2006. IARC Monographs on the Evaluation of Carcinogenic Risks to Humans. Volume 86. Cobalt in hard metals and cobalt sulphate, gallium arsenide, indium phosphide and vanadium pentoxide.

Migrated from Short description of key information:
Case reports evidenced respiratory sensitisation by soluble cobalt (II) salts in humans. Based on read-across, cobalt hydrogen citrate is also expected to be a respiratory sensitiser.

Justification for selection of respiratory sensitisation endpoint:
No study was selected as hazard assessment was conducted by a weight of evidence of appropriate read-across substances. Available data on soluble cobalt salts were pooled for assessment based on the basic assumption that the cobalt ion is the determining factor for biological activity. Please refer to the endpoint discussion for further details.

Justification for classification or non-classification

The available data on skin sensitisation meet the criteria for classification as Category 1 (H317) according to Regulation (EC) 1272/2008 and as Xi, R43 according to Directive 67/548/EEC.

Based on read-across, the available data on respiratory sensitisation meet the criteria for classification as Category 1 (H334) according to Regulation (EC) 1272/2008 and as Xn, R42 according to Directive 67/548/EEC.