HPLC Method for Fructose 1,6 Bisphosphate, Phosphate, Fructose 6-Phosphate on Newcrom B by SIELC Technologies

High Performance Liquid Chromatography (HPLC) Method for Analysis of Fructose 1,6 Bisphosphate, Phosphate, Fructose 6-Phosphate.

Fructose 1,6-biphosphate, alternatively known as Harden-Young ester, is the main form glucose and fructose take when they enter cells. It is a key component of the glycolysis metabolic pathway and is the product of the phosphorylation of fructose 6-phosphate. It has the chemical formula C₆H₁₄O₁₂P₂.
Using a Newcrom B mixed-mode column and a mobile phase consisting of water and acetonitrile with an ammonium formate (AmFm) buffer, Fructose 1,6-bisphosphate can be retained, separated (from fructose 6-phosphate), and measured. Fructose 1,6 bisphosphate can be detected using charged aerosol detection (CAD) with high resolution.

Sugar phosphates occur in biological systems. Sugar phosphates are parts of two metabolic pathways: glycolisis and pentose phosphate pathways. One of the purposes of sugar phosphates is to store and transfer energy in cells. Because of the presence of sugar fragment and phosphate moiety these molecules are very polar in nature. Mixed-mode chromatography was applied for separation of glucose and fructose phosphates and diphosphates. Compounds are separated based on anion-exchange properties with diphosphates retaining longer than mono phosphates. Monitoring is done by ELSD.