Chapter 23

Pharmaceutical Analysis

return to the Course index
previous | next

High Performance (Pressure) Liquid Chromatography (HPLC)

The General Approach

• Collect blood sample
• Separate plasma (partial clean-up)
• Extraction into organic solvent (polarity/pH)
• Injection onto column
• Separation on column
• Quantitation with detector
• Record and analyse results

HPLC Instrumentation

• Solvent Reservoir
• Pump
  • High pressure - 1000 to 5000 psi
• Injector
  • Low pressure - stop flow
  • High pressure value
• Column
  • Normal Phase - organic (water-free) mobile phase
    • Silica gel - non-aqueous
    • Adsorption
  • Reverse phase (C8, C18) - aqueous mobile phase
    • Partitioning
  • Ion-exchange - aqueous mobile phase
  • Molecular sieve - aqueous mobile phase
    • Size
• Detector
  • Specific
    • Absorbance
    • Fluorescence
    • Mass Spectrometry
    • Electrochemical
  • Non-specific
    • Refractive index
    • Radioactivity
    • Conductivity
• Recorder

Fig 23.5.1 Diagram illustrating a Typical HPLC System
Redrawn from: Pieper and Rutledge, Laboratory Techniques for Pharmacists, Upjohn 1989, page 27, figure 6

Figure 23.5.2 A HPLC system in the lab

Figure 23.5.3 Example Chromatogram

Figure 23.5.4 Example Standard Curve - Peak Height Ratio versus Concentration

return to the Course index

Material on this website should be used for Educational or Self-Study Purposes Only

Copyright © 2001 - 2025 David W. A. Bourne (david@boomer.org)

Catch the Sig A game to aid in interpreting Prescription Sig instructions See how many Sigs you can catch before you run out of lives