Biomedical Engineering Reference
In-Depth Information
different BSA/gelatin weight ratio (q) using CARY 300 Bio UV-visible spectrometer. The
samples were heated up to temperatures 40
o
C, 50
o
C,52
o
C, 54
o
C, 56
o
C and 58
o
C before
measurements.
2.6. High-Sensitivity DSC
Thermal denaturation of BSA in aqueous solution in the absence and in the presence of
dextran was monitored with a highly sensitive differential scanning calorimeter, model
DASM-4, Puschino, Russia.
Thermograms were obtained between 20 and 90
o
C, at a scan rate of 60
o
C h
-1
. Previous
experiments performed at a scan rate of 20
o
C h-1 showed the same curves, indicating
insignificant contributions from kinetic effects. For all the measurements, the pH values was
5.4. Some test experiments performed at different concentrations of BSA (0.075 mol dm
-3
-
0.149 mol dm
-3
) produced essentially the same curves, ruling out significant contributions
from concentration effects. All results are averages of three independent measurements.
Degassing during the calorimetric experiments was prevented by additional constant
pressure of 1.7 atm over the liquids in the cells. At first, the water was placed in the sample
and in the reference compartments. A DSC curve corresponding to a water vs water run was
used as the instrumental baseline for BSA solution.
3.
R
ESULTS
3.1. Hydrodynamic Radiuses and Zeta Potentials of BSA, Gelatin
and their Mixtures
We focus our attention to the interaction between BSA and gelatin in aqueous solutions at
pH 5.4 within the region of pair interaction. The scattering intensity as a function of size for
0.25 wt% solutions of gelatin and BSA is shown in Figure 1. Z-Average diameter of BSA at
given conditions is about 9 nm that is close to that (8.39 nm) reported on the web site of the
California NanoSystems Institute (CNSI). About 90 % of all particles of BSA has a average
diameter 7.4 nm (Figure 1). The sample of gelatin is strongly polydisperse but the main peak (
50 %) has the average diameter 280 nm. Gelatin sample also contain small parts of the low
molecular weight fractions with the average diameter 7.6 nm (27 %) and 18 nm (23%).
The scattering intensity as a function of BSA/gelatin weight ratio (q) at pH 5.4 is
presented in Figure 2. The concentration of gelatin in mixture was kept 0.25 wt%.
Since the main peak of the gelatin sample at 140 nm place far enough from the main BSA
peak we shall consider mainly possible changes of the main gelatin peak in the presence of
BSA. As can be seen from Figure 1 the presence of small amount of BSA in the gelatin
solution (at q =0.2 and 0.4) do not affect on the intensity of the main gelatin peak. However at
a higher q the intensity of this peak first decrease considerably (q=1.0) and then completely
disappear at q=2.0. At such compositions the new big peak appear corresponding complex
particles with the Z average diameter 28 nm. At a higher content of BSA in the mixture (at
q=8) the shoulder is appear which correspond to “free” BSA molecules. In order understand
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