Enzymatic post-column cleavage and electrochemical detection of glycosides separated by high-performance liquid chromatography

Crude extract of Helix pomatia , commercially available as β-glucuronidase, was immobilized on porous glass and packed in a column (50 × 3 mm I.D.). Similarly, β-glucuronidase from bovine liver, immobilized on agarose beads, was used as a post-column reactor in the high-performance liquid chromatography of phenolic glycosides. Electrochemical oxidation by glassy carbon electrodes was used for the detection of the phenolic products formed by the enzymatic reaction and proved to be useful for the identification and sensitive detection of phenolic glycosides. Enzymatic activities were in the range 0.1–1 I.U. The detection limits of various phenolic glycosides were 3–23 pmol. Peak broadening and the linearity of the system were evaluated. The reactor containing immobilized Helix pomatia crude extract was also shown to posses enzymatic activity towards cyanogenic glycosides.