UV-Vis absorption reaction curves

Isosbestic point

An isosbestic point is a wavelength at which two chromophores possess the same extinction coefficient. The absorbance of a sample containing only these two species, at this specific wavelength, remains constant as long as their total concentration stays the same.

Simulate UV-Vis reaction curve series containing two spectral components in a|e

This covers the theoretical production of isosbestic reaction curves (i.e. a series of absorption spectra) from two spectral components in different ratios (i.e. two component absorption spectra). This operation was originally implemented in a|e in order to test the SVD analysis routine but the feature may also be useful for other purposes as well.

In this example, the FAM and TAMRA absorption spectra were imported into a|e using the reference spectra download function.

1. Activate the Calculator toggle button in the toolbar. This will activate the Isosbestic plot panel.

The 'Number of spectra' edit box defines how many spectra, in different component ratios, should be produced. The 'Multiplication factor' can be used to multiply all produced spectra by a constant for scaling.

2. Press the Go button in the Isosbestic plot panel. If no spectra have been selected using the data cursor tool, or if the data cursor selection is not exactly two spectra, a selection dialog box will be opened.

3. In the dialog box, select the two component spectra. Press down the Shift or Alt key for multiselection. Press OK.

In this example seven new spectra are produced and plotted in the graph window, i.e. the two component spectra plus five new spectra being the two component spectra added in different ratios:

'New1' = a*'Component1' + (1-a)*'Component2'


where a is a constant in between 0 and 1.

Note that two spectra can also be specified using the data cursor tool, in which case the selection dialog box step is omitted:

First select two spectra.

Then press the Go button in the Isosbestic plot panel.