Enzymatic inhibition and thermal inactivation in the hydrolysis of chestnut purée with an amylases mixture

Hydrolysis of starch present in 225 g L−1 chestnut purée was performed at 70 °C in a sole step with a thermostable α-amylase (Termamyl 120L, type S) and glucoamylase (AMG 300L) mixture. Applying an optimal ratio for both enzymes at a high enzyme concentration (60 enzymatic units g−1 raw chestnut, corresponding to 13.5 enzymatic units mL−1 chestnut purée) total polysaccharide conversion to glucose was obtained in 15 min. Complete hydrolysis was not possible when operating at a 10-fold lower enzyme concentration after 48 h incubation. Glucoamylase product inhibition and thermal deactivation appeared as the main reasons for the incomplete reaction of the enzymes mixture at the lower concentration assayed. An empirical model was applied to describe substrate and competitive and non competitive product inhibition and operational kinetic parameters were calculated for the enzymes mixture in chestnut purée. While α-amylase was not affected by the thermal treatment in the operational conditions, glucoamylase showed a strong thermal deactivation that did not follow first order kinetics but fitted to a parallel biexponential model, which points towards the presence in AMG 300L of two glucoamylase forms with different thermostabilities. An empirical model was developed to describe thermal deactivation of the enzymes mixture in chestnut purée considering the presence of different glucoamylase forms and a sinergistic effect between them and the α-amylase.