Inhalation Exposure to Carbon Nanotubes (CNT) and Carbon Nanofibers (CNF): Methodology and Dosimetry

Figures & data

TABLE 1. Methods of Dosing Rodent Lower Respiratory Tract with Particles

Method	Anesthesia	Depos. all Reg. Resp. Tr.	Evenness of Deposition	Physiol. Mode	Dose Rate	Use for Toxicity Ranking	Use for Quant. Risk Assessm.	NP Pre-treatment to avoid Agglomer	Repeat/Long-term Exposures	Animal Stress at daily Exposure	Special Expertise/Equipment Required	Costs
Bolus Type
Intranasal instillation A	yes	yes	good to poor	no	high	yes	no	yes	limited	high	no	low
Intratrach instillations A	yes	No	good to poor	no	high	yes	no	yes	limited	high	no	low
Orophar. aspiration B	yes	no	good to poor	no	high	yes	no	yes	limited	high	no	low
Inhalation C
whole body	no	yes	excell.	yes	low	yes	yes	no D	yes	low	yes	high
nose only	no	yes	excell.	yes	low	yes	yes	no D	yes	med	yes	high

A - synchronize with inspiration; B - caveat: significant inflammation in rats; C - requires larger amounts of material: D- most aerosolization methods result in significant, yet realistic, agglomeration of NPs.

TABLE 2. Physicochemical Characteristics which influence Toxicity of CNTs and CNFs

1. Method of Generation

2. Shape (length, width, morphology)

3. Agglomeration/aggregation

4. Surface Properties (area, charge, defects, coating, reactivity)

5. Impurities

6. Density

FIGURE 1 a. MWCNT aerosols at a research facility.

Aerosols collected at a research facility during manufacture of MWCNT revealing a multiplicity of structures. (Han et al. 2008). Shown are samples collected in the facility after opening the furnace, demonstrating many non-fibrous structures in addition to some single fibers but mostly agglomerates of different sizes. Initially high concentrations could be significantly reduced following installation of proper enclosure and ventilation systems.

FIGURE 1 b. MWCNT aerosols collected at a research facility.

Aerosols collected at a research laboratory generating MWCNT by chemical vapor deposition (A: Tsai et al. 2009) and several commercial and research facilities manufacturing MWCNT (B, C, D: Methner et al. 2010). In the research facility, mostly agglomerates were seen and only some single filaments with numerous attached clusters of nanoparticles were found that consisted of carbon and included Fe from the catalyst (A). The aerosol in a commercial facility was described as “various forms like agglomerates, clusters, bundles, or nests rather than individual fibers or spherical particles” (B, C, D). These were collected under different working tasks (e.g., sawing, dumping). An example of the appearance of the few individual fibers and spherical particles is shown in (C). Total carbon concentrations measured on filters according to NIOSH NMAM Method 5040 (Issue 2, 1998) were equivalent to 1.8 mg/m³. In general, samples from both facilities indicate that long and individual fibers were hardly seen, in contrast to clusters/tangles of different sizes. The MWCNT aerosol generated experimentally for use in animal inhalation studies seems to be much better dispersed.

FIGURE 2. CNF aerosols collected on impactor stages during thermal treatment.

Transmission electron microscopy images of airborne CNF size selected by impactors located in a thermal treatment processing area for CNF. Impactor Stage A: 2.5–10 μm; Impactor Stage B: 1.0–2.5 μm; Impactor Stage C: 0.5–1.0 μm; Impactor Stage D: 0.25–0.5. Lacey carbon-coated Ni and SiO-coated Ni grids were used in impactors. (From: Birch et al. 2011).

FIGURE 3. Dosimetric Extrapolation of Inhaled Particles from Rats to Humans.

Concept of dosimetric extrapolation of effects of inhaled materials in experimental animals to humans. A Human Equivalent Concentration (HEC) is derived based on study in rats (scenario 2 described in text) by using the MPPD model for rats to estimate the inhaled (delivered) and daily deposited dose in rats resulting from the specific experimental exposure concentrations. Knowing the material rat-specific clearance rates or retention halftime (T1/2) in the lung, the retained dose that has accumulated over the duration in the lung can be calculated, but may also be determined by analytical methods at the end of exposure. Using then human-specific clearance rates—if available—and breathing scenarios as inputs to the MPPD model for humans the HEC can be estimated for either the MMAD and GSD of the experimental aerosol, or for an aerosol size distribution that is known to be present for the human exposure scenario. The HEC is equivalent to the rat exposure in terms of resulting in the same deposited dose—or retained dose—in both species. Through using the MPPD model human and rat inhalability and respirability differences are taken into consideration. Deposited and retained dose can be expressed either per unit of epithelial surface in the respiratory tract—tracheobronchial or alveolar—, or per unit mass of the lung. In addition, the material based dose-metric can be mass (as indicated in the scheme), or surface area or number of CNT/CNF, if respective information is available. [From: (Oberdörster 1989)]

FIGURE 4. Exposure-Response and Dose-Response.

Exposure-Response and Dose-Response relationships of five 3-month inhalation studies in rats with MWCNT, CNF and CB. The carbon black (CB) study is added as a well studied low toxicity particle for comparison.Percent increase in lung lavage neutrophils (PMN) and retained lung dose were not reported for all of the five subchronic rat inhalation studies. Thus, to approximate retained lung burden of the MWCNT study by Ma-Hock et al. and for the CNF study by DeLorme et al., deposited and retained doses were calculated with the MPPD model, Version 2.90.3, using MMAD and GSD data and the packing density of the materials provided by the authors and a normal rat-specific pulmonary retention halftime (T1/2) of 70 days as model inputs. This T1/2 is appropriate for the rats exposed to the lower concentrations; for the high concentrations and resulting high lung burdens, a prolongation of T1/2 has to be expected (for example, as determined by Pauluhn et al., 2010, Table 3). Therefore, the lung burdens calculated for the high dose groups of these two studies are likely underestimated. The x-axes have been truncated to better separate the individual MWCNT/CNF data, although as a consequence the high exposure/date points could not be displayed.A: Lung weight and lung lavage neutrophil exposure-responses. B: Lung weight dose-responses based on retained lung burden expressed as mass, surface area and volume of the retained material.

- MWCNT (Pauluhn, 2010);

- MWCNT (Ma-Hock et al., 2009a);

- CNF (DeLorme et al., 2012);

- CB (Elder et al., 2005);

- MWCNT (Kasai et al., 2015).

FIGURE 5. MWCNT aerosol size distribution in rodent inhalation study.

A: Subchronic inhalation of different types of cigarettes and air-exposed controls in nose-only tubes.B: Chronic inhalation of different types of cigarette smoke and diesel engine exhaust and air-exposed controls in nose-only tubes.Note: Effect of restraint stress on bodyweight in sham (clean air) exposed controls, highlighted in red.

FIGURE 6. MWCNT aerosols collected in rodent inhalation study.

Aerosolized MWCNT collected on filter (A, B) and carbon-coated 200 mesh N: grids (C, D). The MWCNT were aerosolized using an acoustic aerosol generation system (McKinney, Chen, and Frazer 2009) for dry powder generation. Large variations of the airborne structures were observed, ranging from a few single tubes to differently-sized structures of agglomerated CNT to larger tangles resulting in an airborne size distribution as shown in Figure 5. The aerodynamic, settling and diffusional behavior cannot be inferred from these pictures; the large tangle in section D may not be inhalable by rodents and is likely not respirable, but can be estimated from their aerodynamic size distribution. Such experimentally generated MWCNT aerosol for rodent exposures needs to be compared to atmospheres found at workplaces. (See Figures 1, 2).

FIGURE 7. MWCNT aerosols collected in rodent inhalation study.

Aerosolized MWCNT collected on filter (A, B) and carbon-coated 200 mesh N: grids (C, D). The MWCNT were aerosolized using an acoustic aerosol generation system (McKinney et al., 2009) for dry powder generation. Large variations of the airborne structures were observed, ranging from a few single tubes to differently-sized structures of agglomerated CNT to larger tangles resulting in an airborne size distribution as shown in Figure 8. The aerodynamic, settling and diffusional behavior cannot be inferred from these pictures; the large tangle in section D may not be inhalable by rodents and is likely not respirable, but the aerodynamic behavior can be estimated from their aerodynamic size distribution. Such experimentally generated MWCNT aerosol for rodent exposures needs to be compared to atmospheres found at workplaces. (See Figures 1, 2).

TABLE 3. 90-Day Rat Inhalation Studies with MWCNT and CNF, Exposure-Dose-Response Comparison

	Ma-Hock et al. (2009a)		Kasai et al. (2015)
Material	MWCNT (Nanacyl NC7000)	Pauluhn (2010) MWCNT (Baytubes)	(Hodogaya Chemical Co., LTD; MWNT-7)	DeLorme et al. (2012) CNF (VGCF-H)
Characterization
Length/diameter, nm	100–10,000 /5–15 nm	200–1000/10 nm (micronization)	2000–5700/40–90nm	1000–14000/40–350 nm
Agglomeration	Yes	Yes (coiled structures)	Yes	Yes
Density, g/cm^3	Packing: 0.043	Bulk: 0.11 – 0.33	Bulk; 0.007(production plant)	Bulk: 0.077
Impurities	9.6% Al; <0.2% Co	–0.5% Co	0.2 – 0.4%(not specified)	0.003% Fe
BET surf area, m^2/g	250 – 300	255	24–28 m^2/g	13.8
Animals
Rat strain	Wistar; males/females	Wistar; males/females	F344, males/females	Sprague-Dawley; males/females
Body weight (age)	B.W. not reported (8–9 weeks)	–228 g (m); 180 g (f); (–10 weeks)	B.W. not reported (6 wks)	B.W. not reported(>5 weeks)
Exposure
Exposure mode	Nose-only	Nose-only	Whole body	Nose-only
Generator	Brush feed generator + cyclone	Wright-Dust Feeder	Cyclon-sieve generator	Accurate Bin Feeder
Duration	90 days: 6 hr/d; 5d/wk	90 days: 6 hr/d; 5d/wk	90 days: 6 hr/d; 5d/wk	90 days: 6 hr/d; 5d/wk
Conctr, mg/m^3	0; 0.1; 0.5; 2.5	0, 0.1; 0.4; 1.5; 6	0; 0.2; 1; 5	0.54; 2.5; 25
MMAD μm (GSD)	2.0(2.8); 1.5(2.1); 0.7(3.6) (Berner L.P. Cascade Impactor)	3.05(1.98); 2.74(2.11); 3.42(2.14) (Marple Cascade Impactor)	1.5 (2.8); 1.5 (2.8); 1.5 (2.6) (Micro orifice Uniform Depos. Impact.)	1.9(3.1); 3.2(2.1); 3.3 (2.0) (Sierra cyclone/cyclone impactor)
Size distrib.nm(SMPS)	58 ± 1.8 – 64 ± 1.7	75; 320; 908; 1763	Not done	Not done
Post-expos. recovery	No	6 mos. (males only)	No	3 mos. (males, females; 0.25 mg/m^3)
Dosimetry/Biokinetics
Lung burden (90 days, males)	47; 243; 1170 μg (deposited)^a	–5, 25; 100; 420 μg (retained)^b (also post-exposure)	9.69;63.60;360.90 μg (retained)^c	Not determined
Extrapulmonary Translocation	Mediastinal lymphnodes (inside granulomatous nodules forming acrophages)	Hilar lymphnodes; other organs not reported	Mediastinal lymphnodes; parietal pleura (diaphragm, capillaries of muscle), spleem	Tracheo-bronchial lymphnodes; rarely in brain, heart, liver, kidneys, spleen, intestinal tract, mediastinal lymphnodes (no adverse histopath.)
Retention halftime (T1/2, days)	Not done	151; 350; 318; 375^b	Not done	Not done
Response
Body wt. change	No significant difference	No significant difference	No significant difference	15% lower in lowest conc. group; no sign. difference in other groups
Lung weight (90 days)	+ 1%; + 23%; + 81% (males)	+0; + 12; +27; +61% (males)	+5;+17;+30% (males)	–2; + 8; +22; (males)
Lymphnode wt.	Increased size (high conc)	Increased wt (1.5 and 6 mg/m^3)	Not reported	not reported
BAL–PMN (90 days)	Not reported	–0.5; 3.8; 13; 19%	0.6;13.5;45.7%	1.4; 2.7; 11.0%
BAL-PMN post-exposure (3 mos)[6 mos] high dose only	Not done	(22%) [19%]	Not done	(13%) [not done]
Neutrophilic, histiocytic inflam.	Yes	Yes	Yes	Yes
Granulomat. inflam.	All concentrations	6 mg/m3	All concentrations	Not reported
Interstitial fibrosis	Fibrotic changes	At 1.5 and 6 mg/m^3 groups	At 1 and 5 mg/m^3 groups	Interstitial thickening, no fibrosis
Alv. lipoproteinosis	Yes, 0.5; 2.5	No	No	No
Hypercellularity	Not reported	Yes (0.4 and higher)	Goblet cells in nasal cavity, all conc.	Yes (2.5 and higher conc.)
Lymphnode histology	Granul. inflam, all conctr.	Increased cellularity, all conctr. (signif. at 0.4 and higher)	Not reported	Not reported
Visceral pleura	Not reported	Thickening, collagen at 1.5 and 6 mg/m^3	Inflamm. Infiltration (5 mg/m^3)	Not reported
Blood neutrophilia	Yes (highest conc.)	Not reported	Yes, females only, all conc.	Not reported
Systemic Toxicity	Not detected	Not detected	Yes (increased mRNA of inflammatory cytokines in spleen, all conc.)	Not detected
Evaluation
NOAEL	No	100 μg/m^3	No	540 μg/^m3
LOAEL	100 μg/m3	400 μg/^m3	200 μg/m^3	2500 μg/m^3

^a estimated by authors, no clearance considered; ^b based on retained Co in lung; ^c extrapolated from amount determined in left lung and using a left to right lung weight ratio of 1:2.

short-legendFIGURE 8.

TABLE 4. Surface Areas of Respiratory Tract Regions at FRC

	Rat		Human
	cm^2	% of total	cm^2	% of total
Nasal	18.5	0.75	210	0.03
Trach-bronch	24	1.00	4149	0.65
Alveolar	2422	98.25	634620	99.32

Keyhani et al., 1997; Kimbell et al., 1997; Miller et al., 2011.

(Keyhani, Scherer, and Mozell 1997; Kimbell et al. 1997; Miller et al. 2011).

TABLE 5. Acute to Chronic Inhalation Exposures of Rodents to CNT/CNF for Toxicity Testing. (rat as preferred species; 4–6 hrs/day, 5 d/wk; whole-body)

Acute/Subacute (14–28 days)	Subchronic (90 days)	Chronic (2 years)
• to obtain hazard ID and ranking (pos/neg control?) • may be preceded by i.t. instillation or 1 day inhalation with range of doses to estimate inhaled concentration with MPPD model • assure and test rat respirable aerosol with range of concentr. • if available use workplace or consumer exposure data to inform aerosol generation • to determine concentration for 90-day; (range-finding) • to collect biokinetic data for portal of entry, and possibly identification of secondary target organs, incl. pleura, and fetus • to provide guidance for dose levels for mechanistic in vitro testing, incl. secondary organs • post-exposure observation period desirable (˜2 months)	• to derive NOAEL • use minimum 3 conc, including known or expected human exposure levels; both sexes optional • if no effect at 50mg/m^3 rodent respirable aerosol, then no need to do chronic study* • to identify hazard: total resp.tract; pleura, cardiovascular, CNS, bone marrow • identify target organs • to select conc, for chronic study • detailed biokinetics: retention, clearance, organ accumulation, • predicting long-term effects? • to perform risk assessment by extrapolation to human via dosimetric extrapolation • post-exposure observation period for longer-term effects to assess progression-regression (˜3 months)	• to determine long latency effects (cancer); life shortening; extrapulmonary target organs • 3 conc; based on 90-day or range-finding study results; include human exposure level; high dose: MTD; low dose: no significant effect • to assess total respiratory tract, pleura and systemic effects, nose to alveoli; cardiovascular, CNS, bone marrow, others (reproductive?) • to determine detailed biokinetics: resp.tract retention, clearance, organ accumulation • to perform extrapolation to human for risk assessment • post-exposure observation period up to a total study duration of 30 months (if survival of ≥20%)

*This suggestion is based on the fact that such high concentrations will never be reached in a repeat exposure scenario, and it would be a waste of animals (ethical) and resources ($), (Bernstein et al. 2005) to design a chronic workplace study. It should even be considered to move this suggestion to the acute/subacute category and not follow up with a subchronic study if an appropriate postexposure (˜60 d) observation period is included. Even shorter-term (5 d) inhalation studies with CNT/CNF less than 10 mg/m³ or a single 6-hr. inhalation at 30 mg/m³ have induced significant effects (Ma-Hock et al. 2009; Ryman-Rasmussen et al. 2009a; b; Stapleton et al. 2012; Shvedova et al. 2008). Thus, a cutoff at 50 mg/m³ in a subchronic study is very conservative.

TABLE 6. Percent Deposition of Inhaled Particles of Unit Density (ρ = 1 g/cm³) in Extrathoracic, Tracheobronchial, and Alveolar Regions of Rodents

		Mouse				Rat
MMAD	(GSD)	ET	TB	Alv	Total Lung	ET	TB	Alv	Total Lung
4 μm	(1)	51.75	1.34	0.79	2.13	70.3	0.14	0.13	0.27
4 μm	(2)	42.94	1.71	1.56	3.27	61.4	0.69	1.28	1.97
4 μm	(3)	35.86	1.70	1.83	3.53	49.79	1.14	2.50	3.64
3 μm	(1)	55.34	2.93	2.26	5.19	75.92	0.55	0.75	1.3
3 μm	(2)	45.74	1.83	1.81	3.64	59.45	1.15	2.38	3.53
3 μm	(3)	37.28	1.60	1.90	3.50	43.58	1.17	3.02	4.19
2 μm	(1)	53.26	3.49	3.37	6.86	70.45	2.06	3.80	5.86
2 μm	(2)	46.25	2.54	2.55	5.09	50.11	1.24	3.10	4.34
2 μm	(3)	39.16	1.85	2.23	4.08	39.14	0.99	3.23	4.22
1 μm	(1)	39.16	2.77	3.43	6.20	21.87	2.00	5.68	7.68
1 μm	(2)	38.41	3.02	3.78	6.80	32.98	1.89	5.53	7.42
1 μm	(3)	36.35	3.48	4.37	7.85	38.14	1.62	5.38	7.00

MPPD Model 2.90.3

+ Inhalability Adjustment

Rat: Asymmetrical Multiple Path

Mouse: Balb-C

Default settings (nasal breathing; FRC; tidal volume; breathing frequency; no pause)

FIGURE 9. MWCNT aerosol size distribution in rodent inhalation study.

MWCNT aerosol size distribution as determined by a nano-MOUDI through weighing of deposits on the individual stages. The aerosol was generated with an acoustic aerosol generation system (Fig. 6) and collected with a 13-stage nano-MOUDI (NanoMoudi-II^TM Model 125A, MSP Corp., Shoreview, MN) for 10 minutes at a flow rate of 10 l/min. Lower stages (10–13) were removed because deposits could not be reliably weighed; they contained less than 2% of the total mass. Larger structures with aerodynamic sizes >5 μm contribute much of the total mass concentration of the aerosol, but not to the deposition in the lower respiratory tract of rodents. Thus, describing a CNT/CNF aerosol exposure only by its total airborne mass concentration can be highly misleading when used as comparison to a human exposure concentration. Concepts of dosimetric rodent to human extrapolation discussed in this document have to be taken into consideration. Placing a cyclone separator in line before the aerosol enters the inhalation chamber may eliminate the non-inhalable tangles. On the other hand, if such larger structures are still inhalable by humans and if they are present at the workplace, it could be considered to use intratracheal inhalation (Oberdörster et al., 1997) in rats to circumvent the upper respiratory tract. However, intratracheal inhalation requires anesthesia of rats and is not recommended for day-to- day repeat exposures.

FIGURE B-1. CNT/CNF thoracic fractions for humans and rats.

CNT/CNF Thoracic fraction for different sizes for humans and rats at minute ventilations of 15 LPM and 7.4 mL/s, respectively.

TABLE 7. Instruments and techniques for characterizing nano-aerosol exposure (modified from ISO TR12885)

Metric	Devices	Remarks	CNT Monitoring *(M) and Quantitation (Q)
Mass directly	Membrane filter sampling	Requires size selective inlet cutoff device to collect only structures <100 nm. However, most aerosols generated from nanomaterials contain larger agglomerates and aggregates so a size limitation may not be meaningful. Gravimetric and chemical analyses can be used. Major problem: separation of background.	Yes(M + Q) (average/integration over longer time)
	Size selective static sampler	The only devices offering a cut point around 100 nm are cascade impactors (Berne-type low pressure impactors, or Microorifice impactors). Allows gravimetric and chemical analysis of samples on stages below 100 nm	Yes (M + Q) (average/integration over longer time)
	TEOM*	Sensitive real-time monitors such as the Tapered Element Oscillating Microbalance (TEOM) might be useable to measure nanoaerosol mass concentration on-line, with a suitable size selective inlet.	Yes (M + Q)
Mass by calculation	ELPI™	Real lime size selective (aerodynamic diameter) detection of active surface-area concentration giving aerosol size distribution. Mass concentration of aerosols can be calculated, only if particle charge and density are assumed or known. Sue selected samples might be further analyzed off-line (as above).	Yes (M)
			No(Q)
	DMAS	Real-time size-selective (mobility diameter) detection of number concentration, giving aerosol size distribution. Mass concentration of aerosols can be calculated, only if particle shape and density are known or assumed.	Yes (M)
			No (Q)
Number directly	CPC	CPCs provide real-time number concentration measurements between their particle diameter detection limits. Without a nanoparticle preseparator, they are not specific to the nanometre size range. P-Trak has diffusion screen to limit top size to 1 μm.	Yes (M)
			No(Q)
Number directly	DMAS	Real-time size-selective (mobility diameter) detection of number concentration, giving number-based size distribution.	Yes (M)
			No (Q)
	Electron Microscopy	Offline analysis of electron microscope samples can provide information on size-specific aerosol number concentration	Yes (M + Q) and morphology
Mass by calculation	ELPI™	Real-time size-selective (aerodynamic diameter) detection of active surface-area concentration, giving aerosol size distribution, Data might be interpreted in terms of number concentration. Size-selected samples might be further analyzed off-line.	Yes (M)
			No (Q)
	Diffusion Charger	Real-time measurement of aerosol active surface area. Active surface area does not scale directly with geometric surface area for particles larger than 100 nm. Note that not all commercially available diffusion chargers have a response that scales with particle active surface area for particles smaller than 100 nm. Diffusion chargers are only specific to nanoparticles if used with an appropriate inlet preseparator.	No (M+Q)
Surface-area directly	ELPI™	Real-time size-selective (aerodynamic diameter) detection of active surface-area concentration. Active surface area does not scale directly with geometric surface area for particles larger than 100 nm.	No (M + Q)
	Electron Microscopy	Off-line analysis of electron microscope samples can provide information on particle surface area with respect to size. TEM analysis provides direct information on the projected area of collected particles, which might be related to geometric area for some particle shapes.	Yes (M + Q)

TABLE 8. Recommendation for exposure characterization of sampled and airborne CNT/CNF (modified from Oberdörster et al., 2005)

	Human exposure		Rodent Exposure
Characterization Type	Aerosol	Supplied material	Aerosol	Filter samples
Size distribution (primary particles)	E	D	E	D
Agglomeration/aggregation state	E	D	E	D*
Density	D	E	D	N
Shape	E	E	E	E
Surface area	O	E	D	D*
Concentration, mass and number	E	−	E	E
Composition	E	E	E	E
Surface chemistry	O	E	D	D
Surface contamination	D	E	D	E
Surface charge – suspension/solution	O	O	D	E
Surface charge – powder	D	O	D	E
Crystal structure	D	E	D	E
Particle physicochemical structure	D	E	D	E
Solubility	N	E	N	E
Porosity	N	E	N	N
Method of production	E	E	E	E
Preparation process	E	E	E	E
Heterogeneity	D	E	E	E
Prior storage of material	E	E	E	E

E: These characterizations arc considered to be essential and to be available for each characterization at least once.

D: These characterizations are considered to provide valuable information, but are not recommended as essential due to constraints associated with complexity, cost and method availability.

O: These characterizations are considered to provide valuable but non-essential information.

N: These characterizations are not considered to be essential for all studies.

*The CNT/CNF property is assumed to be the same as in the supplied material and therefore no need to repeat measurement. If the CNT/CNF material has been altered from that supplied (e.g., micronization), then the new properties as used in the rodent assay have to be determined.

FIGURE A-1. CNT/CNF inhalability for humans and rats.

CNT/CNF inhalability for humans via oral and nasal, and rats nasal breathings. Equivalent impaction diameter in the figure includes shape and orientation effects. d_ei is found from Equation (1) for cylindrical shapes. Additional size measurements are needed to develop more realistic models of d_ei for CNT/CNF which, while being elongated, generally are not cylindrical.

FIGURE B-2. Human to rat ratios for equal thoracic fractions.

Ratio of human equivalent concentration to rat exposure concentration based on equal thoracic fractions for minute ventilations of 15 LPM and 7.4 mL/s in humans and rats respectively.

FIGURE B-3. CNT/CNF respiratory fractions for humans at different aspect ratios.

CNT/CNF deposition fraction in the human lung via nasal breathing for different aspect ratios. A minute ventilation of 7.5 LMP is used in the calculations.

FIGURE B-4. HEC/CR ratios for CNT/CNF (100 nm—2 μm diameter) at aspect ratio of 10.

Ratio of HEC/C_R for different diameter CNT/CNF and aspect ratio of 10. Minute ventilations for humans and rats were 7.5 LPM and 0.241 LPM respectively.

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