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Physical & Chemical properties

Nanomaterial agglomeration / aggregation

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Administrative data

Endpoint:
nanomaterial agglomeration/aggregation
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference
Reference Type:
publication
Title:
Preparation and characterization of stable dispersions of carbon black and nanodiamond in culture medium for in vitro toxicity assessment
Author:
Kato et al.
Year:
2011
Bibliographic source:
Carbon 49 (2011) 3989-3997

Materials and methods

Test guideline
Qualifier:
no guideline available
Principles of method if other than guideline:
The study investigated the stability of carbon black suspensions (pre-mixed) in NaCl and synthetic cell culture medium (DMEM-FBS: Dulbecco’s modified Eagle’s medium supplemented with 10% heat-inactivated fetal bovine serum) via dynamic light scattering (DLS). The effect of BSA on the aggregation and stability of carbon black was determined alongside with measurements of zeta-potentials and calculations of the theoretical strength of electrostatic interactions.
GLP compliance:
no
Details on methods and data evaluation:
Preparation of carbon black dispersions
Commercial carbon black aqueous dispersions (19.2 wt %) were mixed with an aqueous solution of bovine serum albumin (BSA). Pre-mixed carbon black – BSA suspensions were then shaken and immediately added to the DMEM-FBS solution (Dulbecco’s modified Eagle’s medium supplemented with 10% heat-inactivated fetal bovine serum). Carbon black dispersions in aqueous NaCl solution (6.5 mg/L) were also prepared at the same concentration as the DMEM-FBS dispersions.

Stable carbon black dispersion in aqueous dispersion
As-received carbon black aqueous solution (AB001, 19.2 wt%, carboxylate stabilized carbon black) was diluted with ultrapure water using a Milli-Q system (0.1 µm filter).

Stable carbon black dispersion in DMEM-FBS
As-received carbon black aqueous solution (AB001, 19.2 wt%, carboxylate stabilized carbon black) was pre-mixed in 600 µL BSA solution at concentrations of 33 or 66 mg/L. 900 µL of the dispersions were added to 5100µL DMEM-FBS and gently stirred.

Stable carbon black dispersion in NaCl aqueous solution
300 mL of carbon black aqueous solution (AB001, 19.2 wt%, carboxylate stabilized carbon black) were pre-mixed in 600 µL BSA solution at concentrations of 33 or 66 mg/L. 900 µL of the dispersions were added to 5100µL NaCl solution (6.5 mg/mL) and gently stirred.

Test material

Constituent 1
Chemical structure
Reference substance name:
Carbon black
EC Number:
215-609-9
EC Name:
Carbon black
Cas Number:
1333-86-4
Molecular formula:
C
IUPAC Name:
Carbon black
Test material form:
solid: nanoform
Details on test material:
Carbon black aqueous solution (AB001) obtained from Tokai Carbon Co., Ltd. (Japan)

Results and discussion

Agglomerate / aggregate diameter
Key result
Mean diameter:
>= 1 μm
Medium:
6.5 mg/mL NaCl and DMEM-FBS
Remarks on result:
other: Suspensions unstable, mean hydrodynamic diameter >1 µm
Remarks:
DLS measurement

Any other information on results incl. tables

Stability of carbon black suspensions

The stability of carbon black suspensions was investigated in suspensions pre-mixed in BSA solution and those not pre-mixed in BSA solution. In addition, agglomeration and sedimentation processes of carbon black in the respective media were monitored for two different carbon black nanoparticle concentrations. Nanoparticle hydrodynamic diameters were monitored for one week in water, 3.3 mg/mL BSA solution, 6.5 mg/mL NaCl solution (with/without premixed BSA solution) or DMEM-FBS (with/without premixed BSA solution).

Stable suspensions could not be obtained in solutions without previous mixing in BSA solution. However, with pre-mixing, at nanoparticle concentrations of 0.1 mg/L and 1 mg/L, average hydrodynamic diameters ranged from 179 to 188 and 171 to 187 nm, respectively, in BSA, NaCl and DMEM-FBS solution. With average hydrodynamic diameters of 161 and 158 nm, carbon black nanoparticles showed similar aggregation in water.

In sum, average hydrodynamic diameters of particles in the 0.1 mg/L and 1 mg/L carbon black suspensions ranged from approx. 150 to 190 nm in presence of BSA under all conditions investigated with no obvious changes in particle diameters over the course of seven days and no indication of gravitational settling (DLS measurements).

However, the stability of carbon black nanoparticles was highly dependent on the presence of BSA. Without addition of BSA, 0.1 and 1.0 mg/mL suspensions of carbon black were unstable in NaCl (6.5 mg/mL) solution and DMEM-FBS (Dulbecco’s modified Eagle’s medium supplemented with 10% heat-inactivated fetal bovine serum).

Zeta-potentials, BSA adsorption

In addition, changes in surface potentials including treatments after pre-mixing in aqueous BSA solutions were determined (Table 1). Zeta-potentials in BSA shifted to approximately -14 to -15 mV, whereas the zeta-potential in pure water amounted to approx. -70 to -60 mV, indicating BSA adsorbtion. The adsorbed amount of BSA was further estimated using AFFFF (field-flow fractionation, molecular cutoff 10 kDa), yielding an observed weigth ratio between BSA and and carbon black particles of approximately 1:2, i.e. 0.05 and 0.56 mg/mL BSA bound to carbon black suspensions at concentrations of 0.11 and 1.09 mg/mL, respectively.

Table 1: Zeta-potentials of 0.1 mg/mL carbon black dispersions

condition

Pre-mixed in BSA

zeta-potential (first day)

SD

zeta potential (7 days)

SD

C1

water

-

-67

4.6

-70.9

3.5

C2

3.3 mg/mL BSA

-

-16.2

1.4

-15.4

0.9

C3

6.5 mg/mL NaCl

yes

-15.8

2.5

-15.2

2.4

C5

DMEM-FBS

yes

-14.2

0.7

-14.7

3

Applicant's summary and conclusion

Conclusions:
The study highlights the role of BSA on the stability of carbon black nanoparticle suspensions (AB001, 19.2 wt%, carboxylate stabilized carbon black). Whereas no stable suspensions could be obtained without addition of BSA in 6.5 mg/mL NaCl and DMEM-FBS (Dulbecco’s modified Eagle’s medium supplemented with 10% heat-inactivated fetal bovine serum) after 7 days, stable suspensions (0.1 mg/L and 1 mg/L carbon black) were obtained in presence of BSA with average hydrodynamic diameters of particles in suspension ranging from approx. 150 to 190 nm.