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TI: Sylvon VonDerPool RAD Disposition March 20, 2009 

PM: Zofia Kosim 

 Addendum to P09-142-144 

Background Information

P09-142/143/1/44 (P09-142C), submitted by the Nano-C, Inc.,is 3’HCyclopropa[1,9][5,6]fullerene-C60-1h-3’

-phenyl-, methyl ester. The PMN is expected to be used 

 

Physicochemical Properties

P09-142 P09-143 P09-144 

MW: 910.91 

Physical State: neat solid 

BP: > 400 

VP: < 0.000001 

1031.02 

neat solid 

> 400 

< 0.000001 

1031.02 

neat solid 

> 400 

< 0.000001 

 Additional Chemical Info: P09-143/144 each consists of two enantiomers; however, the specific 

composition of the enantiomers is undefined. (Chemistry Report Attachment # 5 hard copy 

only). 

 

Chemical Structure

 

 

 

O

O

O

O

 P09-142 

 

 
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O

O

O

O

 

 P09-143 

 

 

O

O

O

O

 P09-144 

General Information

The Submitter (Nano-C, Inc.), made no claim that the PMN chemical contains any unique/novel 

properties. 

Health Effects

For particles with size less than 100 nanometers, absorption is expected to be poor via all routes 

based on analogues. For particles with size greater than 100 nanometers, absorption is expected 

to be nil via all routes based on physical/chemical properties. There is concern for lung overload 

if respirable particles are inhaled. For any poorly soluble particles (PSP), there is potential 

concern for adverse lung effects including cancer, lung fibrosis, lung inflammation and induce 

systemic effects including immunotoxicity if inhaled (Human Health Report 1/13/09). 

Carcinogenicity 

P09-142C is a fullerene (C60, buckyball) attached with a 3’H-cyclopropyl-3’-butanoic 

acid, 3’-phenyl-, methyl ester group. They are insoluble particles with diameter ranging from 

about 0.2-160 μm. An ICB document for P09-142-144, posted in the last day or so listed the 

particle size of the PMN at 0.15 – 176 microns (based on three-run analysis). There were no 

particles detected below 100 nanometers and approximately 90% of the particles were larger 

than 1,000 nanometers. 
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 There are no data on the potential human health effects of the P09-142C. These 

aggregates/agglomerates mentioned above have health concerns of particles under the PSP 

category. A number of PSP such as crystalline silica, TiO2 and carbon black have been shown 

to be carcinogenic. Particle surface characteristics are considered to be the key factors in free 

radicals and reactive oxygen species (ROS) formation and in the development of fibrosis and 

cancer by quartz (crystalline silica). By analogy to these carcinogenic mineral particles, there is 

a concern for the PMN for pulmonary fibrosis and cancer if inhaled (Health Hazard Review for 

P09-142C, see attachment # 1 hard copy only). 

 In addition, there is concern that the PMN may disperse/disaggregate in the lung fluids to 

particles within the nanoscale. Hence, there would be a concern for systemic effects of 

nanoparticles beyond the lungs. For instance, Nurkiewcz et al. (2008) found that adverse cardiovascular effects were seen on exposure to ultrafine TIO2 at inhalation doses below those that 

induced pulmonary effects. 

Using energy-filtered transmission electron microscopy and electron tomography, uptake 

of nC60 (60-270 nm in diameter) to the cytoplasm and nucleus of human macrophage cells has 

been reported. In a number of in vitro and in vivo studies utilizing the organic solvent 

tetrahydron furan (THF) to prepare water-soluble suspensions of nC60, cytotoxicity of nC60 to 

human cells or toxicity in the brain of juvenile largemouth bass due to lipid peroxidation and 

generation of reactive oxygen species (ROS) were reported. However, the use of nC60 

suspensions prepared by solvent exchange methods in toxicity studies has been suspected to be 

due to the toxic effects of THF; the toxicity data of nC60 remains a topic of debate within the 

scientific community. In a subchronic inhalation toxicity study, C60 fullerene nanoparticles (2.22 

mg/m3

, 55 nm in diameter) and microparticles (2.35 mg/m3, 0.93 μm in diameter) were exposed 

to rats for 3 hr/day for 10 consecutive days. No gross or microscopic lesions were observed in 

either group of treated rats, and only minimal changes in hematology and serum chemistry, and 

in bronchoalveolar lavage (BAL) fluid parameters were noted. In a second 90-day subchronic 

study, Wistar-Han rats and B6C3F1 mice were exposed by nose-only inhalation to two sizes of 

C60 (0.05 um or 1 um) 3 hours per day 5 days per week for 90 days. Exposure concentrations 

were 0, 0.5, 2.5 mg/m3

 for 0.05 um C60, and 0, 5, 15, and 30 mg/m3 for the 1um C60. There 

were no biologically significant exposure-related changes in these parameters in rats or mice 

exposed to either 0.05 um or 1 um C60 (N.J.Walker et al. 2009). No chronic exposure inhalation 

studies on fullerenes are available (Health Hazard Review for P09-142C see attachment # 1 hard 

copy only). 

Mutagenicity

No mutagenicity data were provided with PMNs 09-142, -143 and -144. Based upon their 

nanotech properties, the PMNs may induce gene mutation in bacterial cells and in mammalian 

cells in culture and in vivo, chromosomal effects in mammalian cells in culture and in vivo, and 

DNA effects in mammalian cells in culture and in vivo. They may not induce gene mutation and 

DNA effects in lower eukaryotes in vivo or cell transformation in mammalian cells in culture. 

Due to uncertainty about their ability to penetrate cells, and to caveats based upon the 

nonstandard methodologies employed to obtain the majority of the positive results so far 

demonstrated in nanotech materials, testing of these materials in genotoxicity assays remains 

problematic, and negative responses would not conclusively rule out the potential mutagenicity 

hazard of nanotubule materials in humans. No testing of the PMNs is recommended at this time 
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(Mutagenicity Hazard Report see Attachment # 2 hard copy only). 

Immunotoxicity

 Based on published scientific data, some nanoparticles appear to have immunotoxicity 

hazard potential. Pulmonary exposure to these PMNs may lead to toxicity to alveolar 

macrophage, impaired clearance of inhaled bacteria and possible systemic immunosuppression 

(Immunotoxicity Hazard Report, see attachment # 3 hard copy only). 

Developmental Toxicity

This report summarizes the potential adverse developmental effects of PMN P-09-

0142/0143/0144. The three PMN substances are carbon-60 fullerene derivatives. No data were 

available on the developmental toxicity of the PMN substances themselves. There are limited 

data (one study, described below) for C60-fullerene, an analogue of the PMN substance; 

however, this study had several deficiencies (see section IV of this report). Due to these 

deficiencies and the use of a route of administration (i.p.) that is not an expected route of human 

exposure, this study does not appear acceptable for assessing the potential developmental 

toxicity of P-09-0142/0143/0144. This study was previously reviewed in 2004 as part of a 

standard review for (Developmental Toxicity Hazard Report see Attachment # 4 hard 

copy only). 

Environmental Effects: Ecotoxicity concerns for the PMN chemicals are mitigated based on no 

releases to water. 

Occupational Exposure

 TABLE 1 (P09-142) 

Scenario # Sites # Workers Inhal (mg/d) Derm (mg/d) 

Manufacturing 0.64 (particle) 3100 (solid) 

Processing 0.0064 “ 3100 “ 

18 (liquid) 

Use negligible 87 “ 

 TABLE 2 (P09-143/144) 

Scenario # Sites # Workers Inhal (mg/d) Derm (mg/d) 

Manufacturing 0.32 (particle) 3100 (solid) 

Processing 0.032 “ 3100 “ 

18 (liquid) 

Use Negligible 87 “ 

Risk Assessment

Rationale: No to minor effects were seen with pure C60 fullerenes in the two subchronic studies. 

However, it is not possible to draw conclusions about these derivitized PMN fullerenes based on 

these subchronic studies with pure C60 fullerenes. Some fraction of the PMN material is in the 

respirable range. Therefore, the concern for any poorly soluble particles (PSP) support a health 

concern for these PMNS. In addition, there is a concern for cardiovascular effects. 
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A comparison of Tables 1 and 2 above indicate a greater inhalation exposure value (0.64 

mg/day) during manufacturing for P09-142. Therefore, since P09-142/143/144 are close 

analogues, the inhalation exposure value of 0.64 mg/day (0.064 mg/m3) with the NCEL of 0.05 

mg/m3 for crystalline silica will be used for the risk assessment calculations. 

Therefore, the worst case worker inhalation exposure of 0.064 mg/m3

 for P09-142C during 

manufacturing when compared to the NCEL of 0.05 mg/m3 for crystalline silica poses a potential 

carcinogenic risk to workers exposed via the inhalation route. 

Testing Recommendations

Inhalation is the preferred method of exposure of the respiratory tract for hazard 

identification and to obtain dose-response data for use in quantitative risk assessment. A 90-

day inhalation study by nose-only administration in male or female rats with up to 3 months 

observation post exposure is recommended for evaluating the toxicity of nanoscale 

substances (CFR 870.3465). There are several additional considerations for using the 90-day 

subchronic protocol at CFR 870.3465: 

i. Evaluation includes markers of damage, oxidant stress, cell proliferation, 

the degree/intensity and duration of pulmonary inflammation, fibrosis, and 

cytotoxic effects in the broncoalveolar lavage fluid (BALF) and 

histopathology of pulmonary and ex-pulmonary organs/tissues 

(cardiovascular, CNS, liver, kidney, etc., ). It is also recommended that 

the potential for cardiovascular toxicity be determined through monitoring 

of the most sensitive blood and/or plasma endpoints indicative of 

cardiovascular effects. 

ii. Data on pulmonary deposition, clearance half-life (biopersistence) and 

translocation of the test material are desirable to aid in interpreting the test 

results. 

iii. Size distributions should be determined prior to, and during, the 

inhalation tests. 

iv. Differences in the physicochemical properties of the as-administered 

material – relative to the as-produced material -- should be provided 

Techniques used to produce the as-administered material should also be 

described. 

v. The proposed protocol for testing should be provided to EPA for review 

and comment prior to the initiation of testing. 

vi. The results of the subchronic inhalation study may support the need to 

conduct a 2-year inhalation study bioassay in rats with the PMN nanoscale 

substance. 

File:TSCA_PMN-90-day_InhalationTesting_12032008Sayre.doc 

References

1. Timothy R Nurkiewicz, Dale W Porter, Ann F Hubbs, Jared L Cumpston, Bean T Chen, 

David G Frazer, and Vincent Castranova, Particle and Fibre Toxicology, 2008. 
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2. N.J.Walker, G.L. Baker, J.A. Dill, D.R. Germolec, K.L. White. J.H. Roycroft, Evaluation 

of the effect of particle size on the toxixity and toxicokinetics of fullerene C60 in rats and 

mice following nose-only inhalation exposure.. Abstract No. 233. 2009 Itinery Planner. 

Baltimore, MD: Society of Toxicvology. 

 

 

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