Assessment of the enzymatic hydrolysis profile of cellulosic substrates based on reducing sugar release

Abstract

The activity profile of a 1:0.30 mixture of Celluclast 1.5L FG and Novozym 188 (Novozymes) was investigated using Whatman #1 filter paper (W1FP) as a single substrate for hydrolysis. The procedure was based on the ability of the enzymes to release total (RS(Tot)), insoluble (RS(Insol)) and soluble (RS(Sol)) reducing sugars from W1FP. RS(Insol) was used to estimate endoglucanase (EnG) activity whereas exoglucanases (ExG) were assessed by measuring RSSol in the presence of δ-gluconolactone. Finally, the β-glucosidase (βG) activity was derived from the difference between RS(Sol) measurements in the presence and absence of δ-gluconolactone. When this analytical procedure was applied to W1FP using 9.64 mg mL(-1) of the enzyme mixture, the relative contributions of EnG, ExG and βG to the total cellulase activity were 63.28%, 12.02% and 24.70%, respectively. Also, this ratio changed with changes in the enzyme loading, giving a new insight into the synergy that exists among the enzymes.

Keywords: 3,5-dinitrosalicylic acid; ARS; ARS able to release soluble reducing sugars; ARS due to endo-β-1,4-glucanases; ARS due to exo-β-1,4-glucanases; ARS due to β-1,4-glucosidases; ARS(EnG); ARS(ExG); ARS(Sol); ARS(Tot); ARS(βG); CMC; Cellulases; DNS; EnG; Enzyme activity; ExG; Filter paper; RS; RS(Insol); RS(Sol); RS(Tot); RSI(Sol); RSβ(Sol); Reducing sugars; Synergism; activity able to release reducing sugars; carboxymethyl cellulose; endo-β-1,4-glucanase; exo-β-1,4-glucanase; insoluble reducing sugars; reducing sugars; reducing sugars released by β-1,4-glucosidase activity; soluble reducing sugars; soluble reducing sugars obtained under β-1,4-glucosidase inhibition by δ-gluconolactone; total ARS; total reducing sugars; β-1,4-glucosidase; βG.