The DOSY sequence includes a delay (for diffusion) and a pulse (the gradient pulse). It is a necessity to know their values, otherwise the raw data can only be processed as an array of 1-D spectra. The exponential decay (due to diffusion) of the signals can be expressed in the general form:
signal(z) = max(signal) exp(-zD)
where D is the diffusion constant and z a function of the diffusion delay and the gradient pulse.
A script does the job of extracting the values from the original files and calculating
the list of z values.
iNMR itself only reads the calculated values of z from a text file called
“zeta”, already written by the script.
Examples of scripts can be found on the web site.
You need to run the script only once (to create the file “zeta”), with the DOSY experiment
already open and into the foreground.
You are free to edit this file or generate it with your own program or script, using the language of your choice.
Note that the quantity of zD is dimensionless. You are free and responsible to choose any unit for z.
The diffusion coefficients will be expressed in the inverse unit. iNMR will utterly ignore those units.
A DOSY spectrum must first be processed like an arrayed spectrum (for example, like an experiment of inversion recovery). In other words, you process it as a 2-D spectrum, but applying the FT along the rows only. Then it is necessary to correct the phase accurately. A baseline correction might be useful or deleterious. Try with and without and judge by yourself. iNMR assumes that the chemical shifts are constant during the experiment. If this is not the case, apply binning along the rows only: the spectrum will be less resolved, but the peaks will not wander from row to row.
It is important that iNMR recognizes a DOSY experiment as an arrayd experiment. What this means in detail is that iNMR should understand that one of the indirect dimensions (normally f-1) is not suitable for FT processing. This recognition is normally automatic and happens the first time you open the spectrum with iNMR. The program then stores the information by means of a trick, namely by defining a dummy atomic species, whose symbol is a dot. If iNMR fails to recognize a DOSY experiment as an arrayed experiment, open the dialog Edit > Metadata and modify the atomic symbol manually, substituting a dot for the H (for example). Avoid using the “fake FT” option: it solves the problem only partially, because iNMR can't tell which dimension to apply the DOSY processing along.
The command “Process > DOSY > Mono-Exponential” can effectively process spectra where each column contains a single signal, or where all the signals at any given chemical shift belong to a single component of a mixture. In this case it is enough to fit the experiment with a simple mono-exponential decay. Here is the meaning of the parameters:
If you select again the command Process > DOSY > Mono-Exponential and then close the dialog with the button “Cancel” you'll return to the frequency-domain spectrum before the DOSY-like transformation.
The diffusion scale (vertical scale) is either logarithmic (to be more correct, it corresponds to the negative of the logarithm)
or linear. In the latter case, all the diffusion coefficients are multiplied by 1e10.
They are more readable in this way.
An handy method to calculate the value of the diffusion coefficient is to put an horizontal mark on the peak of interest
(or up to 5 such marks on different peaks) and choose the command Process > DOSY > Diffusion Coefficients...
A dialog appears.
The calculated coefficents will be reported on the right, in their true dimension.
You can store them into the document, by clicking the button
at the bottom.
Once you have created a list of diffusion coefficients with the command above, you can extract the single components with the command “Extract Components”. You still get the same kind of DOSY plot but, this time, if a column contains more than one signal, you will get two different peaks. The plot generated by this command contains much less rows than the mono-exponential DOSY, actually you can't specify the number of rows. There are so few rows that it is necessary to use a stacked plot to display the result.
The dialog shows a few options, at the top, that are identical to the ones seen for the mono-exponential case. The completely new parameter is called “Ignore components under ...%”. The percentage is calculated relative to the highest component of the column. If you discover that the separation is not good, increase this value and repeat the processing. The risk is that a true minor component can disappear. If you already know, however, that the signals of the components of the mixture fall at different frequencies, than it's OK to ignore even components under 99%.
If the list of the diffusion coefficients contains more than 2 values, you can select a subset. This strategy can be useful when only a portion of the spectrum is processed.