Separation of Select Organic Phosphorus Compounds
Reverse phase ion pairing chromatography (RP-ICP) was used in this
study for the separation and detection of Inositol Hexakisphosphate (lHP),
Adenosine 5' Triphosphate (ATP), Adenosine 5' Diphosphate (ADP), and
Adenosine 5' Monophosphate (AMP). The RP-1PC system uses elements of
both reverse phase and ion exchange chromatography. The reverse phase
chromatography system is composed of a non-polar stationary phase, usually
carbon chains of varying length, with a polar mobile phase. Unlike regular
reverse phase chromatography, the RP-ICP system has a mobile phase
containing an ion-pairing reagent, typically a charged molecule with a
hydrophobic portion that will 'pair up' with ions in solution. The ion pairs are
retained by the hydrophobic stationary phase. Ion pairs do not exist in the
aqueous solution, but in the nonpolar stationary phase. Because the stationary
phase retains ion pairs, the reverse-phase column effectively functions as an
ion exchange column. This methodology offers several advantages. First,
ionic compounds can be separated without having to use an ion exchange
column, which eliminates complicated sample preparation. Second, reverse
phase chromatography is relatively easy to use and the columns are durable
and inexpensive. This allows for the analysis of large sample sets with
minimal preparation time and without costly sample preparation techniques.