Biomedical Engineering Reference
In-Depth Information
particle with a diameters 0.21 um (product № 1-200, batch 1312.1; 0.91 um (product № 1-
900; Batch № 20-326-9.3);3.0 um (product № S37223; Batch № 2272,1);5.0 um (Batch №
2434R 1*; product № S37227, Lot 51993A); 3.0 um CML/sulfonate latex contains a
negatively charged SO
2
O
−
groups grafted to the surface of the polymer particle (Batch № 636,
product № 23-3000); 3.0 um aliphatic amino latex contains positively charged NH
3
+
groups
grafted to the surface of the polymer particle (Lot 466786, Batch № 271-4MD-1,1*) were
produced by Invitrogen, USA.
Preparation of the protein/polysaccharide and protein-polysaccharide-particles
mixtures.
Most experiments were performed in the much diluted phosphate buffer (ionic
strength,
I
= 0.002). To prepare molecularly dispersed solutions of SC, SA,
or
DSS with the
required concentrations, phosphate buffer (Na
2
HPO
4
/NaH
2
PO
4
, pH 7.0,
I
=0.002) was gradually
added to the weighed amount of biopolymer sample at 298 K, and stirred, first for 1 h at this
temperature and then for 1 h at 318 K.
The solutions of SC, SA, and DSS were then cooled to
296 K and stirred again for 1 h.
The required pH value (7.0) was adjusted by addition of 0.1-
0.5M NaOH or HCl. The resulting solutions were centrifuged at 60,000
g
for 1 h at 296K, to
remove insoluble particles. Concentrations of the solutions are determined by drying at 373K
up to constant weight. The semidiluted W-SC-SA, W-SC-SA-DSS, and W-SC-DSS systems
with required compositions were prepared by mixing solutions of each biopolymer at 296 K.
The suspensions of the particles were initially diluted by double bidistilled deionized
water up to 2 wt% of particles. Two phase protein-polysaccharide systems containing 0.5
wt% particles were prepared by slow and careful mixing of the solution containing 2 wt%
particles with two phase W-SC-SA-DSS system or single phase W-SC-DSS system in such
way to obtain the final system containing 0.5 wt% particles.
All experiments with W-SC-SA-
DSS system were performed at the DSS/SC weight ratio, q equal to 0.14.
After mixing for 1
h, two phase systems were kept at room temperature for 3 h. to separate the phases.
Bright light microscopy
is used to visualized particles distributed in coexisting phases,
using an Olympus BX51W1 fixed stage microscope equipped with a high resolution CCD-
camera, (1000[1000 pixels, C-8800-21, Hamamatsu).
Determination of the phase diagram
. The effect of the presence of particles on the
isothermal phase diagrams demixed SC-SA systems
was investigated
using a methodology
described elsewhere[19]. The weight DSS/SC ratio in the system, (q) was kept
at
0.14. The
threshold point was determined from the plot as the point where the line with the slope −1 is
tangent to the binodal. The critical point of the system was defined as the point where the
binodal intersects the rectilinear diameter, which is the line joining the centre of the tie lines.
Symmetry coefficient K
s
was determined as a ratio of concentrations SC and SA at the critical
point.
Rheological measurements
were performed
using a Physica Rheometer, type CSL2 500
A/G H/R, with a cone-plate geometry CP50-1/Ti ~diameter 5 cm, angle 0,993°, Anton Paar.
The temperature was controlled at 23 °C by using a Peltier element. For each sample, flow
curves were measured at increasing shear rate ~from 0.1 to 150 s
-1
. The ramp mode was
logarithmic and the time between two measurements was 30 s. Frequency sweeps ~0.1-200
rad/s were carried out as well for a strain of 3.0%, which was in the linear response regime.
During the rheological measurements, all samples were covered with paraffin oil to avoid
drying.
Environment scanning electron microscopy (ESEM).
Micro structural investigation was
performed with the environment scanning electron microscop
e
Philips XL30 ESEM FEG.
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