This course is taught at the intermediate level and is intended to serve chemists working in the pharmaceuticals and biotechnology industries, though the topics covered are of equal benefit to chemists working in other fields. The audience should consist of persons who routinely work with HPLC and/or other chromatographic techniques, and whose understanding of chromatography is at the intermediate level.
Topics include all stages of method development: sample handling, selection of the separation technique, and column and mobile phase selection. Special attention will be given to the most popular modes of liquid chromatographic separation, LC-MS, CE and CEC as well as on specific techniques such as separation of peptides and proteins, and of ionic and ionizable components (pharmaceuticals). Participants are also encouraged to bring their own separations problems for discussion.
The course is appropriate for individuals with prior
HPLC experience, though beginners are most welcome. The course was developed specifically for those working in
the pharmaceuticals industry – research chemists, laboratory technicians,
biochemists, and biologists searching for practical solutions to separations
broader approach to developing HPLC analysis strategies
Sample handling and pretreatment
Principles of sample preparation in view of the ensuing analytical
Matrix effects; extraction modes; diluent selection
Reverse phase HPLC column selection and pharmaceutical method development
column parameters, system suitability, surface chemistry,
eluent type and composition effects
HPLC techniques for the analysis of ionized or ionizable analytes
General approach to selection of a chromatographic
technique. Principles and
applications of ion suppression
HPLC, ion-exchange HPLC, ion chromatography and ion pair HPLC.
Specific modes of detection for carbohydrates, etc.
Chromatographic separations of proteins and peptides
Special considerations in protein and peptide
separations. Selection of an
appropriate HPLC mode, optimization of analysis parameters, and recovery of
analyte. Specific modes of
detection for proteins and peptides.
The effect of eluent pH on reverse phase chromatographic behavior of
tuning of the HPLC separation with pH adjustment. Buffer concentration effect.
Influence of counteranions on the retention of basic pharmaceutical components
Principles of LC-MS analysis
types of LC-MS interfaces, operation principles. Strategy of HPLC method
development suitable for LC-MS, sensitivity enhancement.
Emerging techniques in
liquid phase separations – CE and CEC.