A robust HPLC method is built on science, not serendipity.
EXPERIENCED CHROMATOGRAPHERS
PREREQUISITES
ADVANCED LEVEL
- Designed for: experienced chromatographers with good working knowledge of HPLC
- Completion of “Understanding HPLC” and “Troubleshooting” is advantageous
- Method development experience recommended
- Solid grounding in chromatography and chemistry
- Practical HPLC user experience
- Familiarity with system suitability parameters
Learning outcomes
- Set clear method development objectives aligned with sample and regulatory needs
- Optimize sample preparation, column chemistry, and mobile phase for selectivity and resolution
- Apply Quality by Design (QbD) principles to build robustness into methods
- Master gradient elution, column selection (HSM model), and method optimisation strategies
- Understand pre‑validation, system characterisation, and documentation for method transfer
ADVANCED HPLC METHOD DEVELOPMENT
LIVE ONLINE INTERACTIVE COURSE · process‑driven & systematic
For the experienced chromatographer – a comprehensive, process‑driven approach to developing optimal and robust HPLC methods. Move beyond trial and error.
📌 COURSE OBJECTIVES
- Understand key principles of HPLC method development
- Apply Quality by Design (QbD) principles for robust analytical methods
- Optimize separation parameters for improved resolution, selectivity, and efficiency
- Ensure method reproducibility, sensitivity, and robustness
⚙️ QUALITY BY DESIGN (QbD) IN HPLC
What is QbD?
- Importance of systematic method development
- Defining Critical Quality Attributes (CQAs) and Critical Method Parameters (CMPs)
Building robustness
- Identifying and controlling variables affecting method performance
- Strategies for reducing variability and enhancing reproducibility
Key factors influencing chromatographic resolution
- Retention factor (k’)
- Efficiency (N)
- Selectivity (α)
🧪 SAMPLE PREPARATION STRATEGIES
- Optimizing sample solubility: effect of solvents, pH, temperature
- Choosing the right diluent: matching polarity, avoiding solvent interactions with mobile phase
💧 MOBILE PHASE OPTIMIZATION
- Effect of sample polarity on separation
- pH control and buffer selection – impact on ionizable compounds, pH stability
- Optimizing flow rates for resolution and throughput
🧬 COLUMN SELECTION AND OPTIMIZATION
- Physical dimensions: length, internal diameter, efficiency
- Particle size and type: conventional HPLC vs. UHPLC
- Column temperature effects on retention and peak shape
- Chemically modified silica‑based phases: reversed‑phase ligands (C18, C8, phenyl, etc.)
- Hydrophobic Subtraction Model (HSM) classification for column selection
📈 GRADIENT ELUTION & METHOD OPTIMIZATION
- Isocratic vs. gradient elution: when to use each approach
- Optimizing retention factor (k’) for improved separation and peak shape
- Effect of gradient parameters: slope, dwell volume, mobile phase composition
🔧 BUILDING A PRACTICAL HPLC METHOD
- Comprehensive sample preparation techniques
- Optimizing retention and selectivity (α): screening column chemistries and mobile phase conditions
- Chromatographic optimisation strategies – resolution, peak symmetry
- Assessing linearity, range, and sensitivity
- Pre‑validation considerations: system suitability, robustness, repeatability
🗣️ Q&A AND CLOSING DISCUSSION
- Summary of key concepts
- Open discussion on challenges faced in HPLC method development
- Resources for further learning
💻 ONLINE PRACTICAL COMPONENT
- Interactive decision exercises: column screening, gradient simulations, robustness testing scenarios
- Case‑based optimisation workshops