Development of Fermentation Process for Production of Bio-based Energy Carriers from Spirodela polyrhiza

Abstract

Energy is the driving force for socioeconomic development. Currently, world’s energy needs are fulfilled by non-renewable energy sources like fossil fuels which are depleting readily and come with the price of environmental pollution. This has led to the quest for alternative energy sources like renewable biofuels. Bioethanol and biogas are two of the promising biofuels that can help in minimizing dependence on crude oil and natural gas, respectively. However, currently bioethanol is produced from food crops that is raising food versus fuel feud. Spirodela polyrhiza is one of the novel feedstock that can be used to produce bioethanol at large scale without causing any food and fuel competition. Spirodela polyrhiza is a hydrophyte with high growth rate, ability to accumulate high starch content in its biomass and has high biomass yield which makes it an excellent alternative feedstock for the production of bioethanol and biogas. In addition to high starch accumulation in its biomass, Spirodela polyrhiza has also the ability to to phytoremediate wastewater like agricultural runoff. The excessive use of agrochemicals to fulfill the high demand of agricultural products has been imposing numerous negative impacts on the environment for decades worldwide. Untreated wastewater with high chemical load and unutilized nutrients of fertilizers and pesticides released from agricultural farms results in contamination and intoxication of clean water bodies. Utilization of Spirodela polyrhiza for the treatment of agriculture wastewaters is an efficient and cost-effective method due to its excellent nutrient uptake abilities. Production of valuable Spirodela polyrhiza biomass during phytoremediation of agricultural runoff also serves as an alternative feedstock for the production of biofuel (bioethanol) which is another precedence of using this method. This research work comprised of four phases. In phase 1 Spirodela polyrhiza from a local pond was collected and its growth was established in Hoagland growth media. The plant was then given nutrient starvation stress to enhance its starch content by 78%. The high starch containing plant biomass was acid pretreated and 99.3% starch to glucose conversion was achieved. In order to ferment plant sugars, yeast strain Saccharomyces cerevisiae QG1 MK788210 was indigenously isolated. In order to achieve high ethanol yield the isolated yeast strain was statistically optimized using Plackett-Burman and Central Composite Design. The fermentation of plant sugar by Saccharomyces cerevisiae QG1 MK788210 resulted in 100% theoretical ethanol yield thus successfully achieving complete conversion of Spirodela polyrhiza starch to bioethanol. The study conducted demonstrated effective optimization of indigenously isolated yeast viii strain Saccharomyces cerevisiae QG1 MK788210 to deliver high ethanol yield from Spirodela polyrhiza. Application of nutrient stress also indicated Spirodela’s excellent absorptive capacity of various nutrients from water which indicates that it can also be used as biophytoremediator for nutrient polluted wastewaters like agricultural runoff. In phase 2 of this research work, the potential of Spirodela polyrhiza for taking nitrogen and phosphates from the agricultural runoff was determined by growing it in the agricultural runoff under lab conditions. Up to 70% and 78% removal of nitrogen and phosphates respectively was achieved during the treatment of agricultural runoff by Spirodela polyrhiza. The starch content of Spirodela polyrhiza was elevated by 58% during 10-12 days of nutrient starvation at the end of the agricultural runoff treatment process. The biomass obtained was pretreated with dilute acid that liberated 96.4% glucose from the starch-rich biomass. In the last step, glucose was used as a substrate for the fermentation process carried out by Saccharomyces cerevisiae QG1 MK788210 yeast strain which yielded 99% volumetric conversion of glucose into bioethanol. In the phase 3 of this research work, we evaluated the potential of Spirodela polyrhiza for production of both bioethanol and biogas in a sequential process and analyzed the ability to generate energy using this process. Spirodela was given nutrient stress to enhance its starch content by 80%. The plant biomass with high starch was dilute acid pretreated and starch to glucose conversion rate of 99.6% was achieved. The plant derived glucose was fermented with indigenously isolated and optimized yeast strain and 99.8% theoretical ethanol yield was obtained. The fermentation vinasse was anaerobically digested and 0.88 NL/g VS biogas yield was obtained. Further output energy analysis of processes producing bioethanol, biogas and both bioethanol and biogas sequentially from Spirodela feedstock was carried out which showed that highest amount of energy i.e., 18.5 MJ/g VS is produced in the process where high starch containing Spirodela plant biomass was used for the production of biomethane only. The results obtained showed that high starch Spirodela biomass is best used in terms of energy when it is anaerobically digested for production of biogas only. However, the type of fuel produced depends upon the need for which fuel production is desired. By now, we have established that high starch Spirodela biomass can effectively be used for bioethanol production. In phase 4 of this research work the cell walls of duckweeds were studied and the results depicted that the duckweed cell walls have high proportion of three fermentable sugars that are galactose, glucose and xylose. The starch content of two duckweed specie, Spirodela polyrhiza and Lemna minor was enhanced by 80% and 72% ix by applying nutrient stress, respectively. The cell wall analysis of high starch duckweeds was carried out and three fermentable sugars i.e., galactose, xylose and glucose were identified. The cell walls of these high starch duckweed biomass were treated with sequential hydrothermal and enzymatic saccharification and ionic liquid cholinium phosphate. The sequential hydrothermal pretreatment and enzymatic saccharification in case of Lemna and IL pretreatment in case of Spirodela were effective in releasing high amount of xylose and glucose from cell wall which, when fermented can effectively enhance the overall bioethanol yield from high starch duckweed biomass rather than utilizing duckweed starch alone