Practical HPLC Method Development Course - 3 days

Course duration:

Location:

Laserchrom - Rochester

Course content:

HPLC in terms of :

• Selectivity
• Relative Retention Time
• Efficiency
• Resolution
• Asymmetry
• Solvophobic Theory

Requirements of an HPLC method

Objectives of an HPLC method

• Resolution
• Speed
• Accuracy/Precision
• Robustness
• Sensitivity

Establishing a starting point for method development

Running an initial gradient - What can be learned from the result

Choice of Column

Choice of solvent

%B Optimisation

Temperature Optimisation

Choice of buffer

Optimising buffer concentration

Choice of Isocratic or gradient elution

Effect of changing gradient profile and run time

Setting up integration conditions

Setting up an appropriate calibration

• Area %
• Internal Standards
• External Standards
• Calibration Range
• Limit of Detection (LOD)
• Limit of Quantitation (LOQ)

Analysis of errors and how to minimise them

Changing detector to increase sensitivity

Establishing confidence in peak identification

Sample preparation

IQ/OQ/PQ

Standard operating procedures

Method Validation

Risk assessment

Case Studies

Course assessment

The practical part of the course is run on an instrument in the training room. During the course we run a sample mixture using different column, we run a 0-100% gradient to establish approximate k' conditions, and then optimise the eluent composition for methanol, acetonitrile and THF using the Snyder triangle. We use diode array detection to monitor changes in peak elution order, and Drylab method development software to optimise for temperature, pH, buffer concentration and %B.

Course Dates in 2010