Chromatography Forum

how do we determine the best working wavelength for analysis of drug n drug preparations by HPLC using UV detector.

^ max of drug in diluent

or

^ max of drug in working mobile phase

or both.

regards


Adam

Dear Friend,

It is always better to determine the UV spectrum of the analyte in the mobile phase to determine the optimum for detection. This is because those are the conditions in which your compound will be detected, and also the UV bands and maxima can be different in the diluent. If you have a PDA detector, the lambda max position can be easily determined.

I can also tell you that it is always better to use wavelength maxima rather than one on either side of the bands. However, in many cases more than one compound needs to be analyzed and therefore it is not always feasible to find a maximum for all the signals of interest.

Good Luck

Benjamin

We usually run first on a DAD detector, scanning. Typically, like Benjamin does, we will use lambda max if it is higher than 250 nm, but for some we choose a different wavelength to avoid potential interferences from the matrix. Obviously, low wavelengths like 210 and 220 have more sensitivity but less specificity, so we usually go higher.