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Basic toxicokinetics

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basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Referenceopen allclose all

Reference Type:
Report date:
Reference Type:
Preparation of 14C-labeled Multi-walled carbon nanotubes for biodistribution investigations
Georgin D, Czarny B, Botquin M, Mayne-L'Hermite M, Pinault M, Bouchet-Fabre B, Carrière M, Poncy J-L, Chau Q, Maximilien R, Dive V, Taran F
Bibliographic source:
J. AMER. CHEM. SOC., 131, 14658-14659

Materials and methods

Objective of study:
Principles of method if other than guideline:
Organ and blood samples were collected and evaluated for the presence of MWCNTs after the i.v. administration of MWCNT radiolabeled with 14C atoms to allow detection in the body.
GLP compliance:
not specified

Test material

Constituent 1
Chemical structure
Reference substance name:
Tangled Multi-Walled Carbon Nanotubes
EC Number:
Cas Number:
Molecular formula:
Hollow tubular carbon, 1-dimensional nano structures with hexagonal arrangement of carbon atoms
Tangled Multi-Walled Carbon Nanotubes
Constituent 2
Reference substance name:
Test material form:
solid: nanoform
see below

Test animals

Details on test animals or test system and environmental conditions:
- Source: Charles River, France
- Weight at study initiation: 226 and 234g for females and 256 and 266g for males

No data

Administration / exposure

Route of administration:
other: 0.05 wt% Rat Serum Albumin (RSA) in ultra pure water
Details on exposure:
A 2.56 mg/ml stock solution was prepared by prewetting the CNT powder in 0.5 vol % ethanol followed by dispersion in 0.05 wt% Rat Serum Albumin (RSA) in ultra pure water. The suspensions were sonicated for 15/16 minutes on ice.

After preparation dispersed 14Cmultiwall carbon nanotubes (MWCNT) solutions were observed by optical microscopy to control sample homogeneity. According to the observations, depending on the MWCNT, the dispersion solutions were stable from one to several hours and therefore were immediately used after their preparation for IV injection in rats.
Duration and frequency of treatment / exposure:
single (day 1) or repeated (on 5 consecutive days, day 1-5) administrations
Doses / concentrations
Doses / Concentrations:
single dose groups: 9.6 – 10 mg/kg b.w. for male animals, and 10.9 – 11.3 mg/kg b.w. for female.
repeated dose groups: total cumulative dose between 48 – 50 mg/kg b.w after 5 days treatment, for male animals, and 54.5 – 56.5 mg/kg b.w. for female.
No. of animals per sex per dose / concentration:
6 animals per time/group (3 males and 3 females), except for day 14 and day 30 for repeated dosages for which only 3 males were investigated.
Control animals:
other: Vehicle treated animals were investigated as follows; males n=2 at days 2, 6, and 90, and females n=1 at days 2, 6 and 90.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled:
Various organs and blood were collected including liver, lung, spleen, kidney, heart and testes. Organs were immediately removed and immersed in an - 80°C mixture of dry ice and isopentane to prevent MWCNT tissue redistribution.
Blood samples were collected on abdominal aortic vein at days post administration of MWCNT. Quantitative determination of the radioactivity in blood was carried out by 1 µL blood deposition on glass slides and radioactivity counting using the b-imagerTM 2000.
For tissue collection rats were anesthetised with isoflurane 2% and sacrificed by exsanguination (abdominal aorta). Organs were immediately removed and immersed in an -80° C mixture of dry ice and isopentane to prevent MWCNT tissue redistribution. After blocking organs in mounting medium, 50 tissue sections (20 m) were made at -20° C with a slicing microtome (LEICA Microsystems, France), selecting representative organ regions. After lay down on glass slides, these sections were left either in freezer for optical study, or were kept at room temperature for 1 day in the presence of silica gel to ensure complete drying, before radioimaging analysis. The quantitative determination of the radioactivity in dried tissue sections and imaging were carried out using a high performance
autoradiography imager (b-imagerTM 2000, detection threshold of 0.01 cpm/mm2 for 14C Biospace Lab, Paris, France).
For each animal, the percentage (%) of the injected dose for whole organs was calculated by radioactivity integration to the whole organ volume, using the radioactivity level detected in the 50 tissue sections for each organ (integration procedure developed previously in our laboratories). The % of the injected dose for each analysed organ is reported.
- Time and frequency of sampling:
Tissue samples were obtained at various time points after the administration. Samples were collected at day 2 (24 hour after the single administration at day 1), and at day 90 after the single administration. After the five times repeated administrations at days 1-5, samples were collected at day 6 (24 hour after the last of the administrations days 1 to 5), and at day 14, day 30 and day 90. At autopsy at day 2, day 6, day 14, day 30 and day 90 after administration blood was collected.

Design of toxicokinetic study for determination of tissue distribution of NM 402:
Treatment Autopsy
Day 2 Day 6 Day 14 Day 30 Day 90
MN (1x) 3M+3F - - - 3M+3F
MN (5x) - 3M+3F 3M 3M 3M+3F
Vehicle 2M+1F 2M+1F - - 2M+1F

Results and discussion

Main ADME resultsopen allclose all
Single IV dose, total recovery in organs on days 1 and 90, 8 and 4% (male) and 24 and 6 % (female). In liver, 7 and 3% and 23 and 5%.
5-day IV dosing, total recovery in organs on days 6 and 90, 5 and 4% (male) and 11 and 4 % (female). In liver, 4 and 3% and 10 and 3%.

Any other information on results incl. tables

In blood obtained at 24 hours after the administration of the 14C-MWCNT very low to almost no radioactivity was detected whatever the protocol of MWCNT administration in either male or female rats. Table 6-2 shows the 14C tissue distribution after a single IV MWCNT administration, and Table 6-3 shows the distribution after five repeated IV administrations.

After a single IV dose for male rats for NM-402 only 8% of the injected dose was observed in organs at day 1, with the liver being the main target organ (7%). In females the recovery was 24% after a single dose (Table 6-2). At day 90 a 50% reduction of the level of NM-402 was observed when compared to day 1 in most organs, while for the lung a similar % of injected dose was observed both on day 1 after a single administration and at day 90 after repeated administration. After the repeated IV dosing for male rats compared to the data of day 6, almost no elimination of NM-402 was observed at day 90 after administration (Table 6-3). However, when comparing the elimination after repeated exposures and single exposure, NM-402 elimination appeared much more reduced. After a single exposure a 50% elimination was observed while almost no elimination was observed after repeated IV doses. A particular feature with this NM-402 was the observation that spleen was more loaded than lung at any time point.

Applicant's summary and conclusion