Pickering Laboratories Pinnacle PCX - AMINO ACIDS

6.2
Section 6
ApplicAtions
Pinnacle Operators Manual
Pickering Laboratories Inc.
AMINO ACIDS
6.2 Introduction
6.2-1 Background
6.2-2 Basic Sample Preparation
6.2-3 Reagent Preparation
6.2-4 Analytical and Post-column Conditions
6.2-5 Procedure
6.2-6 Sample Chromatograms and Gradient Programs
6.2-18 Precautions
6.2-21 Notes
Introduction
High performance liquid chromatography (HPLC) with post-column derivatization is a technique for rendering
analytes more detectable than they would otherwise be in their native forms. Post-column derivatization gives
improved sensitivity or better selectivity (reduction of interference) leading to lower detection limits. The
Pickering Laboratories Pinnacle PCX was developed to facilitate the determination of amino acids in
using sodium ion-exchange or in native samples using lithium ion-exchange columns. There are two options
for post-column detection of amino acids. The first is the use of Pickering’s patented TRIONE
®
ninhydrin
reagent, which will react with both primary and secondary amino acids. The second is the use of
o-phthalaldehyde (OPA), a fluorescent reagent that gives greater sensitivity but will detect only primary amino
acids.
A complete post-column analysis system for amino acids consists of the following components:
• HPLC ternary or greater gradient pump
• Manual injector or autosampler equipped with high pH compatible Tefzel
®
or PEEK
™
seals
• Pickering Laboratories ion-exchange columns
• Pickering Pinnacle PCX post-column derivatization instrument
• Eluants, reagents, and standards
• Visible or fluorescence detector
• Chart recorder, integrator, or data system
Ion-exchange chromatography followed by post-column derivatization has been the method of choice for amino
acid analysis since S. Moore, D.H. Spackman and W.H. Stein published it in 1958—work which merited a
Nobel prize.