Biomedical Engineering Reference
In-Depth Information
generated by passing the sprayed biomaterials through the electrohydrodynamic atomization instru-
ment, and the pDNA was exposed to shear damage only once. Therefore, electrospraying is a more
appropriate technique for the aerosolization of delicate macromolecules such as a naked DNA.
A schematic diagram of the setup for aerosolizing pDNA is shown in Figure 11.50. Electrosprayed
pDNA was generated using a 3 mm cylinder of stainless steel with a central bore of 1 mm and a
polyacetyl cap with a 500 μm spraying aperture. The application of
5 kV to the nozzle using a
high-voltage power supply generated cone-jet spraying. The solution consisting of 80% ethanol,
20% water, 1 mM ethanoic acid, and pDNA was pumped to the nozzle at different liquid fl ow rates
using a syringe pump. The aerosolized pDNA was collected by bubbling the aerosol through a
Dreschel bottle containing 30 mL of chloroform. The plasmid DNA was separated by centrifuga-
tion of the chloroform and extraction of the aqueous supernatant.
A formulation of 80% ethanol and 20% water was chosen for stable electrospraying in the work
because the aqueous pDNA solution was diffi cult to be aerosolized in a stable mode due to high
surface tension and the relatively high conductivity of pDNA formulated. The stability of electro-
spraying can be increased by the addition of 1 mM ethanoic acid. A stable electrosprayed aerosol
containing up to 1 mg/mL of pDNA was generated by using this formulation. Figure 11.51a shows
the size of the electrosprayed droplets generated using various concentrations of plasmid pCIKLux.
The electrosprayed aerosols containing 1 mg/mL pDNA were stable at fl ow rates up to 1.4 μL/s.
Figure 11.51b shows that at a fl ow rate of 0.4 μL/s, the pDNA aerosol had a volume median diameter
(VMD) of 3.45 μm, and a GSD of 0.61. Over 95% of droplets had a smaller diameter than 5 μm.
The experiment revealed that very little degradation of the aerosolized pDNA was observed when
compared with nonaerosolized pDNA. Aerosolization studies
in vivo
showed detectable levels of
pDNA deposition and measurable luciferase reporter gene expression in the lungs of exposed mice.
This study demonstrates that the respirable aerosols of naked pDNA can be generated without plas-
mid degradation through electrospraying technique, which has been proved to be an appropriate
technique for the aerosolization of delicate gene transfer agents.
Gomez et al. produced insulin protein nanoparticles through aerosolizing a solution by elec-
trospraying [110]. The technique was suffi ciently good enough to provide no fragmentation of the
macromolecules and was gentle enough to aerosolize insulin protein. The investigation demon-
strated the feasibility of producing monodispersed and biologically active insulin particles using
electrospraying. Figure 11.52 is a schematic illustration of the apparatus, which consisted of a grav-
ity-controlled feed-line to adjust the fl ow rates, a disposable syringe, a needle with a tapered end,
and high-voltage power supply. The needle was perpendicularly positioned to the ground electrode
and was kept approximately 3 cm apart from the collecting electrode. A voltage of 5 kV was applied
between the two electrodes to establish a necessary electric fi eld to form a stable conical spraying.
−
Liquid
supply
24 V
supply
EHD
nozzle
High-voltage
generator
V
Discharging
needles
Aerosol
FIGURE 11.50
A schematic diagram of electrospraying apparatus for aerosolization of DNA. (Reprinted
from Davies, L.A. et al.,
Pharmaceut. Res.
, 22, 1294, 2005. © Springer Science and Business Media. With
permission.)
