Abstract:

Silica gel was used as the carrier to prepare α-amylase affinity chromatography column. The column was then
used for evaluating the α-amylase inhibitory activity of five glycoside compounds (neomangiferin, mangiferin, timosaponin AⅡ,
timosaponin BⅡ, and timosaponin BⅢ) from Rhizoma anemarrhenae by linear scan cyclic voltammetry. The effect
of immobilization conditions on the immobilized enzyme and the stability of the affinity chromatography column were
investigated. Frontal affinity chromatography was applied to explore the binding affinity of these five glycoside compounds
to α-amylase. Finally, the α-amylase inhibitory activity of samples was assayed. Results showed that the α-amylase
binding affinity of the glycosides followed the decreasing order: neomangiferin (Kd = 0.230 5 μmol/L) > mangiferin
(Kd = 0.299 5 μmol/L) > timosaponin BⅡ (Kd = 0.312 4 μmol/L) > timosaponin BⅢ (Kd = 0.316 2 μmol/L) > timosaponin
AⅡ (Kd = 0.453 3 μmol/L). The same ranking was observed for α-amylase inhibitory activity. Therefore, frontal affinity
chromatography can be used to quantitatively detect the α-amylase inhibitory activity of five glycoside compounds and the
α-amylase inhibitory activity follows the same order as the binding affinity.

Key words: frontal affinity chromatography, linear scan cyclic voltammetry, α-amylase immobilization, α-amylase inhibitory activity, Rhizoma anemarrhenae