Tuesday 4 May 2010

Help on: Peak Purity by PDA


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“I want to know why we are not able to determine Peak purity by use of a UV detector instead of Photodiode Array detecion (PDA)?”

“The HPLC detectors, variable wavelength UV detectors and photodiode array detectors, both work on the principle that molecules of interest are detected by the absorption of UV light. However the design of the instruments differs.

In a variable wavelength detector the instrument can be set up to monitor a particular wavelength and thus a chromatogram is obtained which corresponds to the absorption of UV light at that wavelength at each time point during the injection. Some UV detectors are capable of acquiring multiple wavelengths and so chromatograms can be obtained for several wavelengths.
A photodiode array detector collects information on all the wavelengths within a range specified by the operator, thus obtaining a UV spectrum for each time point during the injection. An individual wavelength may be extracted so that a chromatogram can be displayed for a particular wavelength of interest if required.
To measure peak purity it is necessary to be able to inspect the UV spectrum at a number of time points across the peak of interest. In the example shown in Figure 1, spectra are extracted at retention times of 8.11, 8.21 and 8.31. Since the UV spectra are not comparable it can be concluded that the peak is not pure. Only the photodiode array detector is able to provide this information.
Figure 1
A word of caution, although peak purity by UV can be very useful it cannot conclusively prove that a peak is pure. Molecules which have very similar structures may produce identical UV spectra and thus cannot be differentiated using this technique.”

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