Author: Rafał Łukasik

One of the most challenging objectives of the FRONTSH1P project is the valorisation of marginal streams and by-products in innovative circular biorefineries. Sugars are one of the most important streams, that can be valorised in multiple bio-products. Therefore, the implementation of any biochemical-based biorefinery centred on the so-called “sugar platform” requires the prior hydrolysis of its constituent polysaccharides, i.e. of cellulose and hemicellulose fractions, in order to obtain the respective monomers (mostly glucose and xylose). This hydrolysis can be promoted by chemical, thermal or biological methods.

Enzymatic hydrolysis is advantageous due to its greater intrinsic specificity, with the consequent absence of formation of degradation products (possibly obtained from sugars and lignin) and increased potential yield, reduced energy consumption due to the moderate reaction conditions and its non-corrosive nature (involving less investment in equipment and maintenance costs) and non-polluting. However, most sources of lignocellulosic materials are structurally rigid and compact and therefore their polysaccharides are not easily accessible to enzymes. This recalcitrance of lignocellulosic biomass is mainly due to two factors: the low accessibility of microcrystalline cellulose fibres, which prevents the efficient action of cellulases, and the presence of lignin (mainly) and hemicellulose on the surface of the cellulose, which prevents access effect of cellulases to the substrate.

Thus, lignocellulosic materials, and in particular residual forest and agricultural biomass, unlike saccharine and starch-based (1st generation) raw materials, need to be subjected to mechanical comminution and pre-treatment before applying an enzymatic process, which poses an added challenge in the development of the technology to be implemented. The pre-treatment aims to release the cellulose and hemicellulose fractions from the lignin to reduce the cellulose crystallinity and increase the porosity of the material. Given the high recalcitrance of lignocellulosic biomass, this initial stage of the process is considered one of the key points. It is the most complex step from a technical point of view, and with the highest cost of the conversion technology of lignocellulosic biomass into fermentable sugars. The literature reports show that pre-treatment may constitute more than 40% of the total cost of cellulosic bioethanol production, and as such, it limits the implementation of new advanced commercial biorefineries.

Since this pre-treatment step is common to all value chains for the conversion of biomass into sugars and is of critical importance, advanced biorefineries that use lignocellulose as a raw material can be differentiated based on the different technologies applied as pre-treatment. Pre-treatment technologies are grouped by type of process, having been applied – at the laboratory, pilot and demonstration scales – biological, physical, chemical and physico-chemical processes. However, pre-treatment of the biomass requires a correct integration with the subsequent enzymatic hydrolysis step. There are currently very robust cellulases on the market, acting very efficiently even when applied at a low dose, as is the case of Cellic® CTec3. In combination with enzymes produced by the fungus talaromyces amestiolkiae, β-glycosidases can be enriched to provide 2G monomeric sugars (namely glucose and pentoses, such as xylose and arabinose).

To achieve efficient process intensification coupled with the liquefaction and subsequent enzymatic hydrolysis, the fermentation of the obtained concentrated sugar stream to lipids must be done with a high yield. The fermentation of sugars to lipids can be performed using recombinant cell factories, e.g., the oleaginous bacterium rhodococcus opacus and the oleaginous yeast yarrowia lipolityca. This way the main products are mainly C16-18 triglycerides. Either oleaginous bacteria or yeasts, able to naturally metabolize glucose, can be engineered to assimilate at the same time the pentoses (i.e., xylose and arabinose) present in the hydrolysates. Thus, this can turn the production of free fatty acids from the agriculture wastes to be the carbon, energy and cost-efficient.