Flow Injection Tutorial

Amino acids are routinely assayed in foodstuffs, proteins, and peptides,

and monitored as essential nutrients in fermentors and bioreactors. The

most common method used for this purpose is reversed-phase liquid

chromatography. The advantage of RP-LC is its well-established

wide usage, which assures that laboratory personnel are familiar with

its operation. The chief downside of RP-LC for amino acid assay is the

necessity for pre-column derivatization, required to render amino

acids retentive on reversed-phase columns, and to allow their detection

by means of absorbance or fluorescence.

There exist instrument configurations that allow for precolumn

derivatization of amino acids in an automated fashion. Typically, an

autosampler is used to aspirate aliquots of the

sample and the derivatization agent from separate

vials and to inject the derivatized sample into the

RP-LC instrument.

SI Chromatography offers a far simpler solution

using an instrument that comprises only two

moving parts: a piston pump and a multiposition valve.

Monitoring of amino acids in cell culture medium

by SI chromatography is based on an assay

protocol that comprises the following operations:

1. recovery of amino acids from the bacterial culture

2. precolumn derivatization of amino acids by OPA

3. separation of amino acids

4. detection by fluorescence measurement