Post column derivatisation analyses review. Is post-column derivatisation incompatible with modern HPLC columns?

Area of Science:

• Analytical Chemistry
• Chromatography

Background:

• Post-column derivatization (PCD) coupled with High-Performance Liquid Chromatography (HPLC) and Ultra-High-Performance Liquid Chromatography (UHPLC) is a key analytical technique.
• A significant drawback of PCD is the increased post-column dead volume from reaction coils, leading to peak broadening and reduced separation efficiency.

Purpose of the Study:

• To review the compatibility of existing PCD methods with modern UHPLC systems.
• To analyze system parameters in published PCD methods, focusing on column dimensions, stationary phase, particle size, and reaction loop geometry.

Main Methods:

• A systematic review of 87 PCD methods published between 2009 and 2014.
• Analysis of column dimensions (length, internal diameter, particle size) and reaction loop volumes.
• Experimental evaluation of dead volume effects on column performance (resolution, separation efficiency).

Main Results:

• Most reviewed PCD methods utilized conventional HPLC columns (e.g., 250 × 4.6 mm i.d. with 5 μm particles) rather than UHPLC-compatible columns.
• The majority of methods employed reaction loops exceeding 500 μL.
• Experimental data demonstrated a severe loss of performance with reaction loops larger than 100 μL.

Conclusions:

• Practitioners of PCD methods predominantly use column formats incompatible with UHPLC.
• The dead volume introduced by typical PCD reaction coils significantly compromises the efficiency gains offered by modern HPLC and UHPLC technologies.
• There is a fundamental incompatibility between conventional PCD setups and the requirements of high-efficiency UHPLC separations.