Inhibition of amylases present in ruminal particle‐associated micro‐organisms

The effect of different substances potentially inhibitory of ruminal amylase activity in sheep was assessed using biochemical and electrophoretic assays. Most amylase activity was detected in the particle‐associated fraction (70%) of the ruminal contents (which was selected for the assays) in comparison with the bacterial (21%) and extracellular (9%) fractions. Salts of divalent ions such as Sn²⁺, Hg²⁺, Cu²⁺ and Zn²⁺ produced 90, 82, 65 and 44% inhibition of amylase activity respectively when assayed at a relative concentration of 5 × 10^?3 mol l^?1. Organic acids such as tannic, formic, ascorbic and benzoic acid produced 79, 48, 43 and 37% inhibition respectively, whereas chelators such as EDTA, EGTA and 1,10‐phenanthroline produced an inhibition ranging from 32 to 37%. Substrate SDS‐PAGE zymograms allowed the identification of different amylase‐active bands in ruminal extracts, showing a wide range of relative molecular masses (from 36 to more than 100 kDa). Such zymograms also confirmed the effect of some inhibitors. The reversibility of the inhibitory effect of some of the assayed substances was assessed. ZnSO₄ was the most persistent inhibitory substance even at low concentrations and, owing to its low toxicity, appears to be an adequate substance to reduce the high in vitro ruminal degradation of starch. Implications for the process of enzymatic digestion of starch are discussed. © 2002 Society of Chemical Industry