Information on HPLC method development required
Posted: Sun Mar 18, 2007 8:49 am
I have only little idea regarding hplc method development can any body explain me in detail.Considering the following aspects- if the structure of the molecule is known then how to proceed?
1. Which mobile phase I have to select?
2. What buffer I have to use?
3. Which column I have to use?
4. Which wavelength I have to use?
5. In what concentration I have to inject the sample(do I have to do
linearity study)?
6. If HPLC is not having PDA how to fix the wavelength should we
use UV-VIS spectrophotometer?If yes in which solvent I have to
dissolve the compund and find its lamda maximum?
7. If there is a variety of columns to select which one I will
choose?
8. During method development if peak tailing is observed how to
improve the peak shape ? can this be improved by changing the
buffer concentration or changing the buffer?What is the
role of buffer for the improvement in peak shape?
9. Do I have to look for pKa values if so how? what is the
influence ? explain in detail
10.Why buffers are used in hplc mobile phase?
1. Which mobile phase I have to select?
2. What buffer I have to use?
3. Which column I have to use?
4. Which wavelength I have to use?
5. In what concentration I have to inject the sample(do I have to do
linearity study)?
6. If HPLC is not having PDA how to fix the wavelength should we
use UV-VIS spectrophotometer?If yes in which solvent I have to
dissolve the compund and find its lamda maximum?
7. If there is a variety of columns to select which one I will
choose?
8. During method development if peak tailing is observed how to
improve the peak shape ? can this be improved by changing the
buffer concentration or changing the buffer?What is the
role of buffer for the improvement in peak shape?
9. Do I have to look for pKa values if so how? what is the
influence ? explain in detail
10.Why buffers are used in hplc mobile phase?