Cellulase Inhibition by Pretreatment-Released Biomass Degradation Products

The influence of pretreatment chemistry and the severity of dilute acid pretreatment on the subsequent enzymatic hydrolysis of soluble oligomer-derived and insoluble crystalline cellulose using a commercial enzyme preparation (Novozymes) was investigated. A dilute acid pretreatment was performed on three feedstocks: corn stover, pine and mixed hardwood, at 0.7 % H₂SO₄ (w/v) and 1% solid concentration with a 100 mL Accelerated Solvent Extraction (ASE, Dionex Corp.). The pretreatment conditions (residence time, temperature) with constant acid concentration were varied over a range of six reaction severities. Different pretreatment chemistries were studied on pine and included dilute acid, liquid hot water, buffered hot water, aqueous ammonia and lime, all carried out at the same severity.

Hydrolysis kinetics were analyzed using a Yellow Springs Instruments (YSI Corp.) glucose analyzer. Enzyme kinetics were measured at different dilutions of biomass hydrolysate, with all three sources of biomass causing inhibition of the enzymes to different degrees. Alkaline pretreatments were observed to cause less enzyme inhibition than acidic pretreatments. It was noted that the presence of soluble oligomers in the hydrolysate resulted in apparent diminution of the inhibitory effect, resulting from an increased release of oligomer-derived glucose at higher concentrations of hydrolysate. Correction for this glucose contribution from the oligomeric sugars rendered the inhibitory effect of the hydrolysate more apparent.

Analytical interrogation of the hydrolysates quantified 40 different compounds present in the hydrolysates. Comparison of analytical to hydrolysis results indicate that enzyme inhibition can be correlated to the presence of select degradation compounds present in the hydrolysate.