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A high performance liquid chromatographic method for the separation of hexose monophosphates and UDP glucose from plant extracts and its use to determine specific activities in radiotracer experiments
Author(s) -
Viola R.,
Davies H. V.,
Sommerville L. B.
Publication year - 1994
Publication title -
phytochemical analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.574
H-Index - 72
eISSN - 1099-1565
pISSN - 0958-0344
DOI - 10.1002/pca.2800050104
Subject(s) - chemistry , chromatography , hexose , high performance liquid chromatography , fructose , enzyme , chromatographic separation , phosphate , biochemistry
Anion exchange high performance liquid chromatography (HPLC) was used to separate UDP glucose (UDPGlc) and hexose monophosphates (HMPs) in crude ethanol extracts of potato tubers. Detection and quantification of UDPGlc and [U‐ 14 C]UDPGlc was achieved with UV (A 254 ) and radioactivity monitors connected in series. The 14 C content in glucose 6‐phosphate (Glc‐6‐P), fructose 6‐phosphate (Fru‐6‐P) and glucose 1‐phosphate (Glc‐1‐P) was obtained following their enzymatic conversion to gluconate 6‐P and subsequent separation by HPLC. Individual HMPs were quantified by enzyme‐coupled reaction systems using a microplate reader equipped with a 340 nm filter. This approach allowed the specific activities of UDPGlc, Glc‐6‐P, Fru‐6‐P and Glc‐1‐P to be quantified in radiolabelled potato extracts within 2 h.