Biomedical Engineering Reference
In-Depth Information
However, there is another peculiarity: at pH 7 one sees a semi-equivalence
point, indicating that the molecule resists being charged beyond 28 units. To
study the details of this process, the pH titration was investigated 11 with 31 P
NMR. Figure 10.3 shows the results. The 31 P NMR spectrum shows four lines,
two of those with double intensity, corresponding to mirror symmetry in the
molecule (see Figure 10.3). The titration of the individual phosphate groups
shows that every group experiences a semi-equivalence point during the
titration. Hence the extended titration range and equivalence is not due to
differences in pK a values of the individual phosphate groups; the NMR data
suggest a molecule-wide origin of the titration anomalies. The figure shows
that the behavior of the individual groups is even more esoteric than the
overall
deprotonation—especially
the
blue
signal
(corresponding
to
two
intensities) which is not titrating at all between pH 7.5 and 9.5.
The asymmetry of the titration reflects the asymmetry of the interactions
between phosphate groups. Each group has two close neighbours, two groups
that are at intermediate distance and one group that is the farthest away. The
data can be qualitatively interpreted in those terms: deprotonation of IP6toa
state in which two groups that are the farthest away can be achieved with
relatively little electrostatic repulsion. This leads to the semi-equivalence point
Figure 10.3
The pH dependence of the 31 P NMR spectrum of IP6. The blue and
green trajectories belong to signals with double intensity, the red and
brown lines to signals with single intensity. Adapted from ref. 11.
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