Microbial Fermentation

Several microorganisms including fungi (e.g. Aspergillus, Candida shehatae, Fusarium sp., Kluyveromyces sp., Neurospora sp„ Phanerochaete sp., Penicillium sp., Pichia kudriavzevii, Saccharomyces cerevisiae, Schizophyllum sp., Sclerotium sp., Trichoderma sp., etc.) and bacteria (e.g. Acetovibrio sp.. Bacillus sp., Clostridium thermocellum, Erwinia sp., Escherichia coli, Klebsiella oxytoca, Ruminococcus sp., Zymomonas mobilis, etc.) accomplish fermentation of biomass hydrolysates to produce ethanol (Nanda et al. 2014b). S. cerevisiae is a model microorganism for ethanol fermentation because of its high efficiency, stability, a faster rate of sugar conversion and high solvent (alcohol) tolerance. Moreover, it is also considered as GRAS (generally regarded as safe). S. cerevisiae is also a potential producer of zymase, an enzyme complex that manifests the biocatalysis of sugar fermentation into ethanol and CO, (Lin and Tanaka 2006).

Although S. cerevisiae is efficient in fermenting hexose sugars (glucose) and starch, it lacks the natural ability to ferment pentose sugars (i.e. xylose and arabinose) from hemicelluloses. However, a significant development in synthetic biology and genetic engineering has made it possible to express metabolically engineered pathways for D-xylose and L-arabinose metabolism in 5. cerevisiae (Nijland and Driessen 2020). Moreover, a few fungi such as Candida parapsilosis, Candida shehatae and Pichia stipitis have demonstrated the xylose metabolism with the aid of xylose reductase and xylitol dehydrogenase (Nanda et al. 2014b). Xylose reductase transforms xylose to xyli- tol, whereas xylitol dehydrogenase further converts xylitol to xylulose. Xylulose can be metabolized through the pentose-phosphate pathway.

Pretreatment and enzymatic saccharification generate simple monomeric sugars, w'hich serve as the source of carbon and energy for various microorganisms. Fermentation typically operates in the temperature range of 30-36°C, whereas the process of enzymatic hydrolysis demands a temperature range of 45-50°C. The two most widely used fermentation techniques are solid-state fermentation and submerged fermentation. Solid-state fermentation technology employs the microbial grow'th on moist solid substrates for producing high value-added products. This technology is an expanding approach for the commercial production of many enzymes. This technology has grabbed enormous attention since it is better than submerged fermentation, which involved fermentation in a liquid-based (i.e. hydrolysate-rich) media.

Another iteration of fermentation, i.e. separate hydrolysis and fermentation (SHF) is easy to manage the enzymatic hydrolysis and fermentation processes. However, SHF encounters a limitation of accumulation of glucose and cellobiose during enzymatic hydrolysis, which may cause feedback inhibition for the hydrolytic enzymes. To resolve such an issue, the addition of p-glucosidase becomes necessary, thereby making the process costly. The necessity of [3-glucosidase can be curbed in simultaneous saccharification and fermentation (SSF) process due to restricted chances of feedback inhibition, which can make SSF cost-effective than SHF (Elumalia and Thangavelu 2010; Kont et al. 2013). Furthermore, SSF is a faster process with a higher yield of ethanol in contrast to SHF. The presence of ethanol in the fermentation medium creates an environment where the chances of contamination and spoilage of the sugar-rich hydrolysate are less (Sasikumar and Viruthagiri

2010).

Conclusions

The production of second-generation biofuels such as bioethanol from lignocellulosic biomass plays a pivotal function in the sustainability of biorefineries. As the building blocks of these renewable feedstocks, recovery of the fermentable sugars is imperative for a highly efficient and profitable bioethanol production process. Genetically modified microorganisms and their engineered metabolisms can help utilize a wide variety of monomeric sugars extracted from the pretreatment and enzymatic hydrolysis of lignocellulosic biomass. Furthermore, optimization of the fermentation process and minimizing the chances of contamination and undesired byproduct formation can lead to improve bioethanol yields and lower the overall process expenditures. Pretreatment and enzymatic hydrolysis are crucial steps for the recovery of fermentable sugars from heterogeneous lignocellulosic biomass. Novel pretreatment processes can be adopted that can allow enzymes to act upon biomass simultaneously, thereby maximizing the extraction of monomeric sugars.

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