Unravelling the effect of pretreatment severity on the balance of cellulose accessibility and substrate composition on enzymatic digestibility of steam-pretreated softwood

Pretreatment is essential for effective enzymatic digestion of lignocellulosic biomass. Steam pretreatments increase the digestibility by increasing the accessibility of the carbohydrates to the enzymes. However, they can also cause yield loss and lowered digestibility via increased non-productive binding of enzymes to lignin. The relative importance of these effects is not well defined, especially for softwoods which require more severe pretreatments than other types of biomass. Pinus radiata wood was steam pretreated at 180 °C and 215 °C to Combined Severity Factors of −3.31 or −2.61 and the digestibilities of the washed insoluble fractions examined before and after ball-milling to a common accessibility, as determined by Simons' stain measurements. Pretreatments at 215 °C for 2 min with citric and sulfuric acid catalysts were also investigated. Results showed that the digestibility of the pretreated substrates increased with pretreatment severity, rising from ∼5% with no pretreatment to ∼20% after the most severe pretreatment. However, when the substrates were ball-milled to a common accessibility, the digestibility decreased on increasing pretreatment severity. At a common accessibility and low enzyme dose the digestibility dropped six-fold from ∼30% for the original wood to ∼5% for the most severely pretreated substrate. This showed that while increasing pretreatment severity does lead to greater enzyme inhibition, this was being overridden by increases in the accessibility.