Anyone has experience with more than one drug detection at a time in HPLC? I am having problem to detect 3 drugs individually, even though all of the 3 drugs appear at a time in a single run.
To get different peak separation simultaneously of the three different drugs you should consider below points first:
1. Nature of the molecules (Acidic, Basic or Neutral), their pKa value (very important!!!), and solubility
2. HPLC column properties according to the molecule
3. Mobile Phase (Buffer: Organic) composition types: Isocratic or gradient
4. Wavelength of the molecules (nm) and detector response
Please note, if your molecules are too different from each other, then it would be very difficult to separate them through the similar HPLC method.
If you have no problems in detecting your compounds separately with the same column and using the same gradient, then it is possible that they interact among themselves to generate adducts with different retention times. Check also for possible differences in the pH of the samples.
Hi, you can detect 3 drugs at the same time in single injection of your samples. Gradient method is best for this detection, for this purpose you should know the chemical nature of your three drugs. Based on this, you can create a gradient program.
This is basis of method development in HPLC, however, before any specific answer can be given the three drugs in question need to be supplied. Once this is known the column can be selected based on the type of chromatography best suit to the molecules, (some compromise will be necessary). With a column selected a "rough gradient" of 20% ACN:H20 to 80% ACN :H20 can be run and a method can be developed from the results seen (or not seen as the case maybe). Based on the information you have provided this is best anybody can do to help you, we all need more information to help you further
It seems the 3 drugs are eluting together under the HPLC conditions you are employing. First check the retention time(s) of each drug (standard) , then run a mixture consisting of 3- drugs. This should give you and idea how to proceed next. You can start changing other parameters such as column, solvent mixture and delivery (gradients vs normal). One more factor you need to consider is the chemical structure(s) of the 3-drugs.
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