[[2,2',2''-[29H,31H-phthalocyaninetriyltris(methylene)]tris[1H-isoindole-1,3(2H)-dionato]](2-)-N29,N30,N31,N32]copper

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

migrated information: read-across based on grouping of substances (category approach)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Well documented and acceptable study with the following restrictions: A 91 day subchronic dosed feed toxicity study was conducted in mice, similar to OECD Guideline 408 (Subchronic Oral Toxicity - Rodent: 90-day Study), but not a toxicokinetics study (similar to OECD Guideline 417). However, the study additionally included the examination of systemic copper absorption after exposure to the test material, analyzed in livers and kidneys of the animals.

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

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

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Peer reviewed well documented study.

Reason / purpose for cross-reference:

reference to same study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

yes

Remarks:

; no clinical chemistry, hematology, or urinalysis were conducted and no organ weights were taken (as recommended in OECD Guideline 408).

Principles of method if other than guideline:

This subchronic toxicity study, similar to OECD Guideline 408 "Subchronic Oral Toxicity - Rodent: 90-day Study" was a range finding study, conducted to assist in selecting Maximum Tolerated Dosages (MTD) for a 104 week chronic study in a dosed feed subchronic study.
No clinical chemistry, hematology, or urinalysis were conducted and no organ weights were taken, as recommended in OECD Guideline 408.

GLP compliance:

no

Limit test:

no

Specific details on test material used for the study:

- Name of test material (as cited in study report): Pigment Blue 15B
- Analytical purity: The chemical analysis, performed at Midwest Research Institute indicated that the purity was 104.7 % +- 1.1 % (elemental analysis)
- Elemental analysis indicated that the compound contained less than 0.01 % chlorine.
- Structural formula attached as image file (if other than submission substance): see Fig.

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Industries
- Age at study initiation: 8.5 weeks
- Weight at study initiation: males: 16 - 23 g; females: 16 - 19 g
- Housing: polycarbonate cages: groups of 5 mice per cage
- Diet: weighed portions of Purina Lab Chow in meal form, mixed together with weighed portions of the test material (see details at "doses/concentrations")
- Water: ad libitum
- Acclimation period: 15 days

ENVIRONMENTAL CONDITIONS
- Temperature: 21 - 23 °C
- Humidity: 40 - 60 %
- Air changes: at least 15 per hour
- Photoperiod: 12 hrs dark / 12 hrs light

Route of administration:

oral: feed

Vehicle:

other: 12 % water was added to the test material as a dust control agent

Details on oral exposure:

Dose levels of 5.0, 2.5, 1.25, 0.6 and 0.3 % (w/w) were selected for both males and females. The selected doses were prepared by mixing weighed portions of purina Lab Chow in meal form with weighed portions of the test material. 12 % water was added to the test material as a dust control agent prior to mixing with the meal. For each dose level, one weekly lot of 4500 g (+ 12 % water compensation) was prepared.

The actual mixtures were composed of the following ingredients:
- Dose level 5.0 % (w/w): 252 g test material and water + 4275 g meal
- Dose level 2.5 % (w/w): 126 g test material and water + 4387.5 g meal
- Dose level 1.25 % (w/w): 63 g test material and water + 4443.75 g meal
- Dose level 0.6 % (w/w): 30.24 g test material and water + 44735 g meal
- Dose level 0.3 % (w/w): 15.12 g test material and water + 4486.5 g meal

Each diet was mixed in a Patterson-Kelly twin shelled V blender for 15 min.
The doses were mixed one or two days prior to the week of their use in the study, and stored at 23 °C.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

One analysis was perfomed to determine the accuracy of the mixture concentration. Results were within +- 10 % of the desired dose concentration.

Duration of treatment / exposure:

91 days

Frequency of treatment:

daily

Dose / conc.:

0.3 other: %

Remarks:

nominal in diet, approx. 1000 mg/kg bw/day for males and 1100 mg/kg bw/day for females

Dose / conc.:

0.6 other: %

Remarks:

nominal in diet, approx. 2000 mg/kg bw/day for males and 2200 mg/kg bw/day for females

Dose / conc.:

1.25 other: %

Remarks:

nominal in diet, approx. 4000 mg/kg bw/day for males and 4700 mg/kg bw/day for females

Dose / conc.:

2.5 other: %

Remarks:

nominal in diet, approx. 8000 mg/kg bw/day for males and 9400 mg/kg bw/day for females

Dose / conc.:

5 other: %

Remarks:

nominal in diet, approx. 16000 mg/kg bw/day for males and 18700 mg/kg bw/day for females

No. of animals per sex per dose:

10 males and 10 females per dose

Control animals:

yes, plain diet

Details on study design:

- 10 animals were used per sex and dose group.
- Five dose levels of 0.0, 0.3, 0.6, 1.25, 2.5 and 5.0 % in feed were used in this study (approx. 0, 1000, 2000, 4000, 8000 and 16000 mg/kg bw for males [based on 7.3 g/d average food consumption, 0.023 kg average bw] and approx. 0, 1100, 2200, 4700, 9400 and 18700 mg/kg bw for females [based on 7.1 g/d average food consumption, 0.019 kg average bw], respectively).
- The selected doses were prepared by mixing together weighed portions of Purina Lab Chow in meal form with weighed portions of the chemical. 12% water was added to the chemical as a dust control agent prior to mixing with the meal.
- Each dosed group received on 91 consecutive days of dosed feed mixture.

Observations and examinations performed and frequency:

Animals were observed twice each day for clinical signs, with at least 6 hours between observations. All clinical signs (or negative observations) were recorded daily. Additionally, blood sampling was conducted from 10 control mice, 6 treated males and 4 treated females.

Sacrifice and pathology:

- Mice were necropsied on day 92 and 93.
- Gross examination were performed on all animals from all dosage groups.
- Microscopic examinations were performed on following organs from all animals in the control group and the highest dose treatment group: Kidney, liver, lung, (only in control group: heart).

Other examinations:

Copper analyses were completed in the liver and kidney tissues and the formalin preserving those tissues from male mice in the highest dose group (5 % w/w) and control groups. See details and results in endpoint "7.1.1. Basic toxicokinetics" in endpoint study record "Batelle 76-34-106002, mouse".

Clinical signs:

no effects observed

Description (incidence and severity):

During the course of the 91-day study, neither treatment related signs of toxicity, nor abnormal clinical signs were observed. Cannibalism was observed within one cage group of control females and among four different dose levels in male mice, but was not considered to be drug-related or significant.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

There were seven early deaths during this study. Four male mice from four different dose or control groups died during weeks 3, 7, and 10, while 3 control female mice died during the week 3.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

The body weight data ranged from -7 to +11 % differential weight gain in the dosed females, without any dose-relation. Dosed male mouse groups showed a positive weight differential in all groups except the lowest dosage group of 0.3 %. No trends coinciding with dosage levels or food consumption are evident in these data.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

there were no trends in diet consumption among dosed animals compared with controls in either male or female mice. Female control mice consumed 2 to 3 grams more feed than dosed animals. However, diet consumption data in this group were based on seven survivors during the last ten weeks of the study and have a higher value than expected. The dosed female mouse groups had average and expected levels of diet consumption.

Haematological findings:

not examined

Description (incidence and severity):

No hematology was performed.

Clinical biochemistry findings:

not examined

Description (incidence and severity):

No clinical chemistry was performed.

Urinalysis findings:

not examined

Description (incidence and severity):

No urinalysis was performed.

Organ weight findings including organ / body weight ratios:

not examined

Description (incidence and severity):

No organ weights were taken.

Gross pathological findings:

no effects observed

Description (incidence and severity):

No substance related changes were reported on macroscopic and microscopic examination of the animals. At necropsy there were no consistent lesions noted.

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

Microscopically there were no compound related lesions noted.

Details on results:

The authors recommended an dosage level of 5 % and 2.5 % test substance to be used in the chronic study, due to lack of compound-related lesions.

Dose descriptor:

NOAEL

Effect level:

ca. 16 000 mg/kg bw/day (nominal)

Sex:

male

Basis for effect level:

other: No treatment-related adverse effects upt to the higest tested dose.

Dose descriptor:

NOAEL

Effect level:

ca. 18 700 mg/kg bw/day (nominal)

Sex:

female

Basis for effect level:

other: No treatment-related adverse effects upt to the higest tested dose.

Critical effects observed:

no

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1979

Report date:

1979

Reference 2

Reference Type:

study report

Title:

Unnamed

Year:

1980

Report date:

1980

Reference 3

Reference Type:

other: expert judgement

Title:

Phthalocyanine Blue 15 B and Phthalocyanine Green 7

Author:

Mennear JH

Year:

1980

Bibliographic source:

Assessment from Dr. Mennear JH, Expert Toxicologist to Dr. Moore JA, Deputy Director NTP

Report date:

1980

Reference 4

Reference Type:

secondary source

Title:

Final SIDS Initial Assessment Profile on C.I. Pigment Green 7 Copper, [tetradecachloro-29H, 31 H-phthalocyaninato(2-)-N{29},N{30},N{31},N{32}]-

Author:

OECD

Year:

2005

Report date:

2005

Materials and methods

Objective of study:

distribution

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

other: OECD Guideline 408 (Subchronic Oral Toxicity - Rodent: 90-day Study)

Principles of method if other than guideline:

This subchronic toxicity study, similar to OECD Guideline 408, was conducted to assist in selecting Maximum Tolerated Dosages (MTD) for a 104 week chronic study in a dosed feed subchronic study. Additionally the study examined, whether or not systemic absorption of copper occured after exposure to the test material. Copper analyses were conducted in livers and kidneys of the animals.

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

29H,31H-phthalocyaninato(2-)-N29,N30,N31,N32 copper

EC Number:

205-685-1

EC Name:

29H,31H-phthalocyaninato(2-)-N29,N30,N31,N32 copper

Cas Number:

147-14-8

Molecular formula:

C32H16CuN8

IUPAC Name:

[29H,31H-phthalocyaninato(2-)-kappa~2~N~29~,N~31~]copper

Test material form:

solid

Specific details on test material used for the study:

- Name of test material (as cited in study report): Pigment Blue 15B
- Analytical purity: The chemical analysis, performed at Midwest Research Institute indicated that the purity was 104.7 % +- 1.1 % (elemental analysis)
- Elemental analysis indicated that the compound contained less than 0.01 % chlorine.

- Structural formula attached as image file (if other than submission substance): see Fig.

Radiolabelling:

no

Test animals

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Industries
- Age at study initiation: 8.5 weeks
- Weight at study initiation: males: 16 - 23 g; females: 16 - 19 g
- Housing: polycarbonate cages: groups of 5 mice per cage
- Diet: weighed portions of Purina Lab Chow in meal form, mixed together with weighed portions of the test material (see details at "doses/concentrations")
- Water: ad libitum
- Acclimation period: 15 days

ENVIRONMENTAL CONDITIONS
- Temperature: 21 - 23 °C
- Humidity: 40 - 60 %
- Air changes: at least 15 per hour
- Photoperiod: 12 hrs dark / 12 hrs light

Administration / exposure

Route of administration:

oral: feed

Vehicle:

other: 12 % water was added to the test material as a dust control agent

Details on exposure:

Dose levels of 5.0, 2.5, 1.25, 0.6 and 0.3 % (w/w) were selected for both males and females. The selected doses were prepared by mixing weighed portions of purina Lab Chow in meal form with weighed portions of the test material. 12 % water was added to the test material as a dust control agent prior to mixing with the meal. For each dose level, one weekly lot of 4500 g (+ 12 % water compensation) was prepared.

The actual mixtures were composed of the following ingredients:
- Dose level 5.0 % (w/w): 252 g test material and water + 4275 g meal
- Dose level 2.5 % (w/w): 126 g test material and water + 4387.5 g meal
- Dose level 1.25 % (w/w): 63 g test material and water + 4443.75 g meal
- Dose level 0.6 % (w/w): 30.24 g test material and water + 44735 g meal
- Dose level 0.3 % (w/w): 15.12 g test material and water + 4486.5 g meal

Each diet was mixed in a Patterson-Kelly twin shelled V blender for 15 min.
The doses were mixed one or two days prior to the week of their use in the study, and stored at 23 °C.
One analysis was perfomed to determine the accuracy of the mixture concentration.

Duration and frequency of treatment / exposure:

90 days

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0.3 other: %

Remarks:

in diet, approx. 1000 mg/kg bw for males [based on 7.3 g/d average food consumption, 0.023 kg average bw] and approx.1100 mg/kg bw for females [based on 7.1 g/d average food consumption, 0.019 kg average bw], respectively).

Doses / concentrations 2

Dose / conc.:

0.6 other: %

Remarks:

in diet, approx. 2000 mg/kg bw for males [based on 7.3 g/d average food consumption, 0.023 kg average bw] and approx. 2200 mg/kg bw for females [based on 7.1 g/d average food consumption, 0.019 kg average bw], respectively).

Doses / concentrations 3

Dose / conc.:

1.25 other: %

Remarks:

in diet, approx. 4000 mg/kg bw for males [based on 7.3 g/d average food consumption, 0.023 kg average bw] and approx. 4700 mg/kg bw for females [based on 7.1 g/d average food consumption, 0.019 kg average bw], respectively).

Doses / concentrations 4

Dose / conc.:

2.5 other: %

Remarks:

in diet, approx. 8000 mg/kg bw for males [based on 7.3 g/d average food consumption, 0.023 kg average bw] and approx. 9400 mg/kg bw for females [based on 7.1 g/d average food consumption, 0.019 kg average bw], respectively).

Doses / concentrations 5

Dose / conc.:

5 other: %

Remarks:

in diet, approx. 16000 mg/kg bw for males [based on 7.3 g/d average food consumption, 0.023 kg average bw] and approx. 18700 mg/kg bw for females [based on 7.1 g/d average food consumption, 0.019 kg average bw], respectively).

No. of animals per sex per dose / concentration:

10 males and 10 females per dose

Control animals:

yes, plain diet

Details on study design:

- 10 animals were used per sex and dose group.
- Five dose levels of 0.0, 0.3, 0.6, 1.25 and 5.0 % in feed were used in this study (approx. 1000, 2000, 4000, 8000 and 16000 mg/kg bw for males [based on 7.3 g/d average food consumption, 0.023 kg average bw] and approx. 1100, 2200, 4700, 9300 and 18700 mg/kg bw for females [based on 7.1 g/d average food consumption, 0.019 kg average bw], respectively).
- The selected doses were prepared by mixing together weighed portions of Purina Lab Chow in meal form with weighed portions of the chemical. 12% water was added to the chemical as a dust control agent prior to mixing with the meal.
- Each dosed group received on 91 consecutive days of dosed feed mixture.
- Mice were necropsied on day 92 and 93.

Copper analyses were completed in the liver and kidney tissues and the formalin preserving those tissues from male mice in the highest dose group (5 % w/w) and control groups:
- Tissue samples were prepared for analysis by digesting in 10 ml of concentrated nitric acid until most of the organic material was destroyed. Perchloric acid was then added and the solutions were evaporated to strong fumes, additional nitric acid being added as required. The solutions were then fumed to dryness, the residues were dissolved in 5 % nitric acid and the solutions were diluted to 10 ml.
- Formalin samples were filtered through a Millex-GS 0.22 µm filter unit and 5 ml portions of each sample were prepared for analysis by the procedure used to prepare the tissue samples.
- The samples were then subjected to atomic absorption spectrophotometry to determine copper content:
A Perkin-Elmer Model 5000 atomic absorption spectrophotometer was utilized for the work. A series of 10 ml standard solutions, ranging from 0.05 to 2.0 ppm were prepared in 5 % nitric acid by dilution of a certified standard copper stock solution. These solutions were used to calibrate the instrument, which was programmed to print out data as total microgramms of copper per sample. The prepared sample solutions were used in the same manner as the standards. Concentrations of copper in the tissue samples were calculated by dividing the total microgramms found by the weight of the sample. Concentrations of copper in the formalin samples were calculated on a volume basis.

Statistics:

Student´s T-test (alpha = 0.05) was used to compare the highest dose group results with control results.

Results and discussion

Preliminary studies:

No data given.

Toxicokinetic / pharmacokinetic studies

Details on absorption:

No data given.

Details on distribution in tissues:

Slight, but statistically significant increases of copper incorporation were reported in the liver tissues (3.98 ppm +- 1.16 ppm; p < 0.05) and kidney (7.47 ppm +- 2.86 ppm; p < 0.05) tissues of treated male animals of the highest dose group, compared to controls (liver: 3.0 ppm +- 0.34 ppm; kidney: 4.66 ppm +- 0.6 ppm). From all the formalin analyses performed, the authors drawed the conclusion that no detectable levels of copper were leached from the preserved tissue into the formalin bath.
However, an evaluation of the results was conducted by expert judgement and the author draw the following conclusions:
The results of the study do not provide unequivocal evidence for a lack of absorption of the test material. However, the total lack of findings during the 13 week study, coupled with the insolubility of the test material and the minimal changes in tissue copper residues strongly suggests that the test material was not appreciably absorbed.

Details on excretion:

No data given.

Metabolite characterisation studies

Metabolites identified:

not measured

Details on metabolites:

No data given.

Any other information on results incl. tables

Table 1: Copper determinations in tissues and formalin of male mice from the subchronic study, treated with the test material for 91 days

male animal #	ppm copper
	liver	kidney	formalin	remark
highest dose group (5 % w/w)
1	7.0	14.5	--
2	3.2-3.6 *	6.8	--	* results of 2 analyses
3	3.4	7.7	--
4	3.4	4.8	--
5	3.5	6.3	--
6	3.7	7.3	--
7	3.9	6.4	--
8	4.1	8.2	--
9	3.3-3.6 *	5.2	--	* results of 2 analyses
control
1	3.5	4.2	0.1
2	3.1	3.8	0.1
3	3.1	4.6	0.1
4	2.8	4.6	0.1
5	2.9	4.2	0.1
6	3.2	4.7	0.1
7	3.2	5.8	0.1
8	2.9	5.3	0.1
9	2.3	4.7	0.1

Applicant's summary and conclusion

Conclusions:

Interpretation of results: no bioaccumulation potential based on study results