The titration of an amino acid
These data represent the pH titration curve of an amino acid. A plot of pH against
volume of base added produces the classic S-shaped curve expected. The values of
pK1, pK2 and the Ionisation Potential (pI) can be obtained from the graph. A more
accuarate esitmate of pI can be achieved using a derivative plot -- that is a plot of
rate of change of pH against volume (see below).
pH
Volume / ml
1.00 2.34 pH change
1.50 2.42 12.00
2.00 2.47
2.50 2.57
10.00
3.00 2.66 pK2
3.50 2.78
4.00 2.90 8.00
4.50 3.03
5.00 3.25
pH
6.00
5.50 3.63 pI
6.00 4.32
7.50 8.53 4.00
8.00 9.27
8.50 9.37 pK1
2.00
9.00 9.50
9.50 9.60
0.00
10.00 9.70 1 3 5 7 9 11 13 15 17 19
10.50 9.76
Volume NaOH / ml
11.00 9.84
11.50 9.97
Using a derivative plot
Here the idea is to graph the rate of change (i.e. first derivative) of pH against volume of
base added. We start with the same data as before:
pH
Volume / ml Rate of change of pH (1) Click in cell D39, and type an
1.00 2.34 equals sign.
1.50 2.42 0.08 (2) Click in cell C39.
2.00 2.47 0.05 (3) Type a minus sign.
2.50 2.57 0.10 (4) Click in cell C38.
3.00 2.66 0.09 (5) The formula should read "=C39-
3.50 2.78 0.12 C38"
4.00 2.90 0.12 (6) Click on the green tick to confirm.
4.50 3.03 0.13 (7) The value for C39-C38 is
calculated and inserted in cell D39.
5.00 3.25 0.22
5.50 3.63 0.38 You can manually enter the remainder
6.00 4.32 0.69 of the formulae, but it is much easier to
7.50 8.53 4.21 use Excel's Fill function:
8.00 9.27 0.74
(1) Click in cell D39.
8.50 9.37 0.10 (1) Click in cell D39.
9.00 9.50 0.13 (2) Move your cursor to the bottom
9.50 9.60 0.10 right cormer of the cell where it will
10.00 9.70 0.10 change to a cross.
10.50 9.76 0.06 (3) Hold down the left mouse button
11.00 9.84 0.08 and pull down to automatically fill the
11.50 9.97 0.13 remaining cells.
You now need to plot the derivative. If you have followed the instructions above you will
have three columns of data. But you only want to plot the first (volume) and last (rate of
change of pH). To plot these data:
(1) Select the data in the first column in the usual way, then, while it is still selected press
and hold down the Ctrl key.
(2) Now select the data in the second column.
(3) Call the chart wizard and edit your graph.
Derivative plot
4.5
Rate of change of pH
4
3.5
3
2.5
2
1.5
1
0.5
0
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
Volume base added / ml
e remainder
uch easier to